umukoro

what is computer

2007-12-26T06:15:00.000-08:00

Definitions of Computer
An electronic, digital device that stores and processes information
An electronic device that has the ability to store, retrieve, and process data, and can be programmed with instructions that it remembers. The physical parts that make up a computer (the central processing unit, input, output, and memory) are called hardware. ...
The term computer can refer to just a (1) a boxy device which contains a hard disk drive (HDD), memory, a CPU (central processing unit) and other circuitry, or it can refer to (2) the boxy device plus peripheral devices, such as a display, keyboard and mouse. ...
The ability to access a computer or a network from a remote site
a programmable machine that inputs, processes and outputs dat
A "computer" is defined for Current Population Surveys as a personal or home workstation having a typewriter-like keyboard connected to a laptop computer, mini-computer, or mainframe comp
Originally, the term referred to a person who did mathematical calculations using a mechanical calculation device or an analogue computer. Today, the term 'computer' refers to the electronic machine that is able to store, process and manipulate data. ...

This document includes a brief overview of basic computing concepts. It is the first in a series of three self-instruction Internet tutorials produced by the Bedford Public Library.You can scroll through the document or jump to any of the topics listed below. You can also print the entire document by printing this page.
What is a Computer?Software and Hardware Hardware Components Hardware Accessories Operating System Software Software Applications Reference

What is a Computer?
Computers are not very intelligent devices, but they handle instructions flawlessly and fast. They must follow explicit directions from both the user and computer programmer. Computers are really nothing more than a very powerful calculator with some great accessories. Applications like word processing and games are just a very complex math problem.

Software and Hardware
If you use a player piano as an analogy, the piano can be thought of as the hardware and the roll of music as the software.The software a series of very simple computer instructions carefully organized to complete complex tasks. These instructions are written in programming languages (like BASIC, PASCAL, C...) to help simplify the development of applications.The hardware is what sits on your desk and executes the software instructions. The player piano is useless unless the roll of music has been written correctly.

Hardware Components
Input Devices -- "How to tell it what to do"
- A keyboard and mouse are the standard way to interact with the computer. Other devices include joysticks and game pads used primarly for games.
Output Devices -- "How it shows you what it is doing"
- The monitor (the screen) is how the computer sends information back to you, whether it be surfing the web or writing a memo. A printer is also an output device.
Storage Devices -- "How it saves data and programs"
- Hard disk drives are an internal, higher capacity drive which also stores the operating system which runs when you power on the computer.- "Floppy" disk drives allow you to save work on small disks and take the data with you.

Memory -- "How the processor stores and uses immediate data"
- When you use a program, the computer loads a portion of the program from the hard drive to the much faster memory (RAM). When you "save" your work or quit the program, the data gets written back to the hard drive.
Microprocessors -- "The brain of the computer"
- PCs primarily use microprocessors (sometimes called the chip) manufactured by Intel. The older Intel versions include the 386, 486 and now the Pentium line.
- Macintoshes use PowerPC processors by Motorola.
- Megahertz (MHz) is the internal processor speed in which computer instructions are performed. The MHz speed does not always indicate the power of the microprocessor. Newer processors can execute more instructions at the same or slower MHz. For example, an Intel 486 @100MHz is less powerful than a Pentium @75 MHz (but the MHz is "faster").

Hardware Accessories
Modems
- Modems allow you to communicate with other computers using a phone line. Modem speeds are in bits per second (14.4, 28.8 and 56 thousand bits per second are standard).
CD-ROM Drives
- A CD-ROM drive is a high capacity storage device which lets you read data from the disk, but not write data back. The speed of the drive (how fast the CD platter spins) is measured in multiples from the first generation drives. New drives are up to 24X (or 24 times the first drives), but while the CD spins faster, it is not really 24 times faster in actual output.
Printers
- There are different types of printers (laser, ink jet, dot matrix) with differing quality of output. They are measured in dpi (dots per inch) and ppm (pages per minute), the higher the better.
Scanners
- Scanners "digitize" printed material (like photos and graphics) and save it to a graphic file format (like .GIF or .JPG) for display on the computer.

Operating System Software
Operating system software provides a "user interface" for users to manage files, start programs, customize computer settings, and other tasks. The operating system also provides the fundamental core computer functionality for programmers.Intel based PCs use Microsoft Windows version 3.1 (older) or Windows 95 as the operating system. Macintoshes use the Macintosh operating system.

Software Applications
Application software uses the operating system software and provides the real functionality of a computer. Applications include:
- Word Processing (MS Word, WordPerfect, Ami...)- Spreadsheets (Lotus 123, MS Excel...)- Database (DBase, Fox Pro, Oracle...)- Presentation (MS PowerPoint, Persuasion...)- Internet Browsers (Netscape Navigator, MS Internet Explorer)- Games

Computer viruses are small software programs that are designed to spread from one computer to another and to interfere with computer operation.
A virus might corrupt or delete data on your computer, use your e-mail program to spread itself to other computers, or even erase everything on your hard disk.
Viruses are most easily spread by attachments in e-mail messages or instant messaging messages. That is why it is essential that you never open e-mail attachments unless you know who it's from and you are expecting it.
Viruses can be disguised as attachments of funny images, greeting cards, or audio and video files.
Viruses also spread through downloads on the Internet. They can be hidden in illicit software or other files or programs you might download.
To help avoid viruses, it's essential that you keep your computer current with the latest updates and antivirus tools, stay informed about recent threats, and that you follow a few basic rules when you surf the Internet, download files, and open attachments.
Once a virus is on your computer, its type or the method it used to get there is not as important as removing it and preventing further infection.

Umukoro david

Slow cookers

2007-12-11T02:30:00.000-08:00

6-Quart Meal Maker Slow Cooker by Hamilton Beach
//

Slow cookers are one of the most appreciated kitchen appliances. For a ready-made dinner after work, all the home chef needs to do is put the ingredients in the crock the night before and refrigerate, then in the morning before leaving for work, place the crock in the cooker base and turn the knob on. Succulent roasts or stews are done by dinner time. Decorated with garden-fresh vegetables, this six-quart oval-shaped model holds a 6-pound beef roast, 6-pound chicken, two 3-pound rib packs, or approximately 5 quarts of stew or soup. Wraparound heat ensures food cooks from the sides, not just the bottom. The temperature knob features two cook settings, low and high, and a keep warm setting for serving after food is thoroughly cooked. An included latch keeps the lid secured for transporting. The crock removes from the base for use in a microwave or oven, and both the crock and lid are dishwasher-safe.
Price
$46.99

Availability
Available to Ship
Quantity

Breast cancer

2007-12-11T01:38:00.000-08:00

Breast cancer is a growth of abnormal cells within the breast. Breast cancer is not a single disease but rather a group of diseases that can develop in the ducts (which carry milk to the nipple), the lobules (milk producing areas) or other parts of the breast.
After non-melanoma skin cancer, breast cancer is the most common form of cancer in women. For 2007, the American Cancer Society (ACS) estimates that more than 178,000 new cases of breast cancer will be diagnosed, adding to the 2 million women who have been diagnosed and treated previously for this disease. In addition, the ACS estimates that nearly 40,500 women are expected to die from breast cancer in 2007, making it the second leading cause of cancer death among women (surpassed only by lung cancer).
Although rare, men can also develop breast cancer. In the United States, about 2,000 men are expected to develop breast cancer in 2007, and about 450 men will die from the disease, according to the ACS. However, breast cancer primarily affects women.
Common ways to screen for breast cancer include monthly breast self-examination, clinical breast examination performed by a physician and screening mammography. However, to confirm a diagnosis of breast cancer, a biopsy is performed in which all or part of the mass is removed and then analyzed by a pathologist who views the specimen under a microscope.
Based on the findings by the pathologist, if the tumor is cancerous (malignant) then additional tests, such as an MRI, ultrasound or PET scan may be ordered. All these tests provide the physician with additional information as to the location and stage (extent of spread) of the cancer and other information about the tumor and how it may respond to treatment.
There are several options for breast surgery ranging from the breast-sparing lumpectomy to the now rarely used Halsted radical mastectomy, which removes the breast, the chest muscles and nearby lymph nodes.
In addition to surgery, a patient may receive additional treatment based on the type of breast cancer, its size and other important factors. The size of the tumor, the possible spread of the cancer to the lymph nodes and any possible spread to other organs or tissues helps the oncologist (cancer physician) determine the stage of the cancer. This information, in turn, gives the physician the necessary input to select the best treatment options for the patient. These options can include a wide variety of combinations of treatment, including surgery, radiation therapy, chemotherapy, hormone therapy or biological therapy.
A major study released in 2006 indicated that the number of new cases of breast cancer dropped significantly in the reporting period of 2002 to 2003. The ACS has reported a leveling off of new cases from 2001 to 2003, after 20 years of increases.
Overall survival rates for breast cancer are quite high when the disease is detected and treated in its earliest stages. When breast cancer progresses undetected to later stages, the survival rates drop. The 5-year survival rate for cancer that is localized (not spread to the lymph nodes or areas outside of the breast) is 98 percent. If the cancer has spread to nearby tissues, the 5-year survival rate is 83 percent.
According the ACS, large gains in survival rates for several cancers, including breast cancer, have been recorded over the past two decades. The overall survival rate for all stages of cancer at 5 years is 89 percent and at 10 years is 80 percent.

Strep throat infection

2007-12-11T01:21:00.000-08:00

Strep throat is an infection caused by group A streptococci bacteria. These bacteria can cause inflammation and infection in a variety of areas throughout the body, including the respiratory system, skin and vital organs.
A strep throat infection can be transmitted by kissing, coughing, sneezing or touching contaminated surfaces. Symptoms usually appear within five days of exposure to the bacteria and include a sore throat, fever, swollen neck glands and tonsils. Symptoms tend to begin suddenly and are usually more severe than those experienced with viral infections. Coughing and a runny nose are not signs of strep throat.
Most cases of strep throat occur in children between the ages of 5 and 15 years. Strep throat rarely occurs in infants and toddlers. Children who spend time in crowded environments (e.g., daycare facilities, school) or who live in northern areas of the United States face an increased risk of developing strep throat.
Parents who suspect their child has strep throat should consult their child’s pediatrician. A physical examination and medical history may be performed, although the only way to identify the presence of strep throat is with a throat culture and/or rapid strep test (test that is similar to a throat culture, except with faster results).
Children with strep throat are treated with antibiotics. This kills the bacteria that causes infection, limits the spread of infection to others, and prevents serious medical conditions that may occur if an untreated infection spreads throughout the body. Children remain contagious for at least a day after beginning treatment. While children are recuperating from infection, parents can help make recovery as comfortable as possible.
It may be impossible to completely avoid people carrying the bacteria that cause strep throat. Some people may carry the bacteria but never develop an infection themselves and will not display any symptoms. Perhaps the best way to prevent infection is to practice good hygiene (e.g., handwashing with soap and water). Replacing a child’s toothbrush after infections is a good way to prevent the recurrence of infection.

Prevention (scarlet fever)

2007-12-11T01:09:00.000-08:00

Prevention methods for scarlet fever
The bacteria that cause scarlet fever can be passed through direct contact and through droplets in mucus and nasal fluids. Close contact with an infected person or items they have used can spread the disease. Good hygiene may prevent transmission. Regular handwashing and carefully separating and washing utensils and dishes used by an infected person can help prevent transmission. Avoid touching hands to the eyes, nose or mouth.
Infected patients should be kept out of work, school or daycare until they have been treated with antibiotics for at least 24 hours. Schools or daycare centers should be notified if a child contracts scarlet fever so other children and parents can attempt to prevent further transmission. If a child frequently gets a sore throat or strep throat, a physician may recommend tonsillectomy (removal of the tonsils) to prevent future strep infections

Diagnosis and treatment for scarlet fever

2007-12-11T00:51:00.000-08:00

Diagnosis and treatment for scarlet fever
The rash that accompanies scarlet fever is unique and may be readily identified by a physician. The physician will also compile a medical history, including any potential exposure to infection, and perform a physical examination. Although scarlet fever is not usually difficult to diagnose, milder cases (in which throat examination does not conclusively show strep infection) may resemble other conditions, such as rubella.
Identification of the Streptococci bacteria is also important. A physician or nurse may use a swab to take a sample of material from the back of the throat. A rapid strep test of this sample may identify strep throat in about five to 10 minutes. A throat culture takes two to three days to identify strep throat as the source of the scarlet fever.
Physicians treat scarlet fever with antibiotics. Patients should remain away from school, work or other public areas until they have taken antibiotics for at least 24 hours. The antibiotic prescription should be used until it is finished (usually 10 days), even if the symptoms have subsided. Other symptoms of scarlet fever may also be treated to make the patient more comfortable. Foods and beverages to soothe a sore throat include soup and ice cream. Patients should drink plenty of liquids. A pain reliever such as acetaminophen may be given to children with fever. Children with fever should never take aspirin because of the potential for developing Reye syndrome.

Scarlet fever

2007-12-11T00:10:00.000-08:00

Scarlet fever is a bacterial infection that results in a characteristic red skin rash. It is caused by the same bacteria that cause strep throat and frequently occurs with strep throat.
Streptococci bacteria can invade the body and cause throat inflammation and strep throat. Some of these cases also develop into scarlet fever, which may also include a rash, fever and enlarged tongue. The symptoms of scarlet fever develop quickly, only a day or two after exposure to the bacteria.
Scarlet fever is passed through fluids of the nose and mouth, often by coughing, sneezing or having direct contact with an infected person. Scarlet fever can occur at any age, but is most common among children, who spread it readily in schools and daycare centers.
Physicians can diagnose scarlet fever by identifying the characteristic rash. They also may take a swab from the throat to perform a throat culture or rapid strep test to confirm the presence of streptococcal bacteria. Scarlet fever is easily treated with antibiotics and other measures to alleviate symptoms of the rash or fever. Prevention methods revolve around good hygiene and keeping patients from transmitting the disease for several days after they begin antibiotic therapy. The effect of scarlet fever on public health has changed dramatically in the past 100 years. Before antibiotics were available, scarlet fever was a common cause of serious childhood illness and death, mostly occurring when its complications affected other body systems. Quarantining patients in their homes was the only way to prevent transmission and minimize epidemics. With the antibiotics available today, scarlet fever can be readily cured within a few weeks and rarely causes serious complications. Most patients who have scarlet fever develop immunity to it and do not contract it again.About scarlet fever
Scarlet fever is an upper respiratory disease characterized by a red skin rash and high fever. The rash is caused by an allergic reaction to toxins produced by streptococcal A bacteria, the same bacteria that cause strep throat.
Streptococcal bacteria normally can live on healthy skin and may cause no problems. When the bacteria get inside the body, they can cause infections such as strep throat or the skin and tissue infections of impetigo or cellulitis. Untreated, strep throat can develop into scarlet fever. In rare cases, other streptococcal infections may cause scarlet fever, but most cases originate with strep throat.
The rash associated with scarlet fever can affect most parts of the body, except the area around the mouth. It may itch or feel like a sunburn and after several days begins to peel.
Although it can occur at any age, scarlet fever usually affects children, though some children may be more susceptible than others. The bacteria can be transmitted through mouth and nasal fluids or droplets, either directly by coughing or sneezing or sometimes from objects an infected person has touched. The disease is commonly spread in areas of close contact such as schools and daycare centers. Scarlet fever occurs more often in colder months when children spend more time indoors and in close contact.
The disease is easily treatable with antibiotics. However, left untreated, scarlet fever can infect other body systems and create other serious inflammations. Rheumatic fever can inflame connective tissue such as heart valves and damage them permanently. Glomerulonephritis is a kidney inflammation that may eventually lead to kidney failure. The advent of antibiotics has changed the course of scarlet fever. Before the middle of the 20th century, scarlet fever was a serious and often deadly childhood illness. Patients were quarantined in their homes for weeks and often their belongings were burned. People who later developed rheumatic fever spent months convalescing. Current antibiotic treatment and attention to hygiene has eliminated the need for quarantine. Most people who contract scarlet fever develop immunity to further infection with the same strain of the bacteria.

MOMS SKIN RASH CURE

2007-12-11T00:06:00.000-08:00

WARNING:
"The New Eczema/Dermatitis Skin Rash Healing Secret That Skin Doctors Don't Want You To Know About ...
... Guaranteed To Heal Your 'Worst Case' Skin Problems In 3 Days Or You Don't Pay A Cent"
You've been to doctors and dermatologists who prescribedyou steroids, creams, antibiotics and drugs that did nothing ...
You're sick and tired of itching and scratching yourself all the time and you're embarrassed when people stare at your skin in public ...
You're about to discover a revolutionary new skin cream solution that'salready helped over 99,824 skin rash sufferers from 52 different countries ...

My Risk-Free Guarantee - Your Skin Rash Symptoms Healed In Days Or You Don't Pay A Cent!
I fully guarantee your satisfaction 100%. If after using Freederm HC your skin is not totally healed (or dramatically improved) within 3 days then simply mail the jar back to us (even if its empty) and we'll refund your money in full. No questions asked. You've got nothing to lose except your painful eczema.

Freederm HC has been proven to heal many different skin conditions, such as:
>> Dermatomyositis>> Psoriasis>> Pityriasis Rosea
>> Lichen Planus>> Keratosis Pilaris>> Diaper Rash
>> Lichen Sclerosis>> Dyshidrosis>> Seborrheic Dermatitis
Keep reading to discover how you can "test drive" this new skin product risk-FREE ...

Do you suffer from eczema? Then you must read this now ...
"Oh My God!
I Pulled Back The Cover Of His Stroller And Screamed For Dear Life ..."
Recently, we received this alarming letter ...
"My child just turned 5 months. But at 1 month he broke out with this really bad rash. So I took him to see the doctor. I picked up the medication prescribed by the doctor and it didn't do a thing but make my child scream when I tried to put it on him.
I tried all types of lotions from Vaseline, gold bond, eucerin and more but to my surprise nothing helped. I have nearly spent over $80.00 plus just trying to find something to help my child.
Late one night after my many attempts to calm him down and put him to bed, I came across your web site. What really got to me was the testimonials and the before and after pictures. I could not believe my eyes. I was telling my self this is what my son has.
Well when my husband got home from work and I told him about the web site and showed him. He was amazed then skeptical like I was. But I said hell if I wasted over $80.00 why not go for it, after all its for my sons sake. So I ordered it with the guarantee of the 100% money back. It got to me in 3 days. I was amazed at the fast shipping.
Then we gave my child the bath with the soap and put on the cream and moisturizer. And we went out for the day. Around lunch time we sat at the mall in the food court and he woke up. I pulled back the cover of his stroller and screamed for dear life.
I was so shocked at what I saw that I yelled "OH MY GOD". My husband came running over and almost dropped his food. He asked me what was wrong and when I looked back at our son in his stroller he looked also. He looked like he saw a ghost.
Our sons face no longer had the scabs, and it wasn't oozing and it wasn't swollen and red and he looked happy. The eczema was light pinkish color and his face was smooth and he was smiling with no pain or worries.
If you ever asked for some type of miracle I would tell you this is the cream for you. I have enclosed some before pictures and after pictures. So a total of only 2 days with the treatment and look at the results. Once again THANK YOU SO MUCH FROM THE BOTTOM OF OUR HEARTS.
Baby Mahalo Just 2 Days After Using Freederm HC!

Before
After
- Aleena (and baby Mahalo) from Honolulu Hawaii
Dear Eczema Sufferer,
If you're like most eczema sufferers, then ...
You're sick of itching and scratching all the time! You're frustrated because you can't stop scratching the skin on your hands, face and other effected body parts.
Your doctor or dermatologist didn't help you. You've been to see a skin specialist and they gave you a prescription for some expensive drugs or antibiotics (like steroids, triamcinolone, Elidel, Protopic) but had no effect on your eczema!
You've tried everything but nothing works. You're family and friends have told you to try "proven" products like creams and lotions, but nothing works!
You're embarrassed to go out in public. You wear clothes to cover your scars, rashes, cracked and peeling skin. You're afraid of people who stare at you, make nasty comments, don't want to shake your hand and treat you like a leaper!
You can't get a good nights sleep! You wake up throughout the night with an unbearable urge to itch and scratch yourself. You're regularly tired and fatigued because you haven't been sleeping well.
Bottom line is this: If you suffer from eczema, tried every steroid, drug, cream the doctors and dermatologists have prescribed you but nothing works - and want a solution that heals your eczema in just days - then stay here and read this letter, because I've got some great news for you! But first ...
How I Stumbled Upon The "Miracle" That's Now Healed 99,824 Eczema Sufferers ...
Hi, my name is Michelle Grover. My son pictured to the right was effected by eczema.

After 18 months of watching him suffer and trying every eczema treatment available, this product completely healed him in just 3 days.
To me, calling this cream a "miracle" isn't an exaggeration. The worst pain I have ever felt was not being able to do anything to relieve my son's pain and discomfort of terrible eczema for the first 18 months of his life. The frustration nearly drove me insane. As a Mother, I'd have given anything in the world to have helped him.
In desperation, I tried everything. All the "off the shelf" products, everything that the numerous doctors at different clinics I took him to prescribed, but none of them could stop his painful, itching symptoms.
Every time I'd see those ads in magazines that say "New Breakthrough Product for Eczema" I'd get my hopes up and run out and buy it, only to find out it didn't work. But then ...
One Day An Amazing Thing Happened
On one of my many trips to the local clinic, a different pediatrician was on duty, and he recommended that I pay a visit to a pharmacist in my home town. I remember thinking, "why would a doctor send me to a pharmacist without prescribing anything?"
... But I went to see the pharmacist, and he gave me a jar of cream to try. He said it was a cream that he had created for his own eczema and it worked so well, he'd started mixing it for a few of his customers. I was hopeful but skeptical as I walked out the door.
That Night "The Miracle" Started ...
Right after giving my son his evening bath and applying the cream, I tucked him into bed and kissed him goodnight. The next morning when I awoke, I was a bit surprised to realize that I had slept all night!
My son didn't wake me up with his crying and scratching from the agony of his itchiness. I immediately took his pajamas off and I yelled to my husband in shock ...
"Honey, You Have To Come And See This!"
His skin was already showing dramatic healing and I couldn't wait to see what the next application would do. I placed it all over his body and I kept checking every couple of hours.
It was the first day in a long time that I took the cotton socks off of his hands. He walked around touching things as if it was the first time he had touched anything.
After about 72 hours, he was like a new person. Not only was his skin soft and smooth but he was also happy and enjoying himself for the first time.I called the pharmacist the next week overjoyed, and thanked him over and over until I could feel him blushing over the phone. And that's when I realized ...
I Had To Share 'Eczema Miracle' With The World
I asked the pharmacist if he could make the product available to the general public. He wasn't interested in going that route, so I had to go forward on my own.
I contacted an FDA licensed manufacturer and worked with their on staff chemist in order to come up with a more up to date version of the cream. I decided that day that nothing would stop me from getting this product to you, and after a year of hard work, I have finally made that a reality.
Since then, almost 100,000 people have benefited from this remarkable eczema solution. In fact ...
Here's Just A Few Of The 92,874 Eczema Sufferers Freederm HC Has Already Helped ...
(Guaranteed To Work No Matter How Old You Are Or What Country You're From)
Success Stories:
"Your Product Got Rid Of The Eczema That I've Had For Over 10 years"

Before
After
"Hi, I wanted to be your first African American Testimonial. When I arrived at your site, I noticed that there were not any. Being in college and suffering with eczema is not a fun thing. I was always trying to cover up the areas on my body that had eczema. Your product got rid of the eczema that I have had for over 10 years. Here are the before and after pictures. I hope this helps you get more African American customers as I know it will work for them as it has for me. Thank you so much for putting this product on the market. I feel like a new person.
- Anisha (Raleigh, North Carolina)
"After One And Half Days It Was Actually Healing!"

Before
After
"We are thankful that our son did not suffer for months and months (although his eczema was severe) before discovering your cream. We tried various creams, lotions, and ointments, but none seemed to really help and we did not want to use straight hydrocortisone on his delicate skin.
When the Freederm cream arrived, we eagerly read the instructions and applied the cream on his entire body. His cheeks were oozing, cracked, and red. His forehead, ears and neck suffered the same and the rest of his body was full of eczema to a lesser degree, but nonetheless itchy and irritated (as seen in the photo).
After one and half days, he was actually healing! My heart was relieved. I must note that the flare ups come and go and Freederm surely keeps it to a minimum. As time has passed however, I have seen the quality of his skin only improve. Also, using Freederm cream has become less frequent. I am thankful for this product and have already recommended to several people."
- Daisy Farrales
Eczema Sufferer Of 31 Years Says ..."Such Improvement In Just 3 Days -I Couldn't Believe My Eyes ..."

Before
After
I have had problems for the past 31 years... After looking at your pictures of different people on your site with the same condition as I have I thought that I would try your product. What a great product. After just three days there was so much improvement in my hands I couldn't believe my eyes. I will never be without FreeDerm again.
Thank You so very much for such a great product.
- Colleen (Whitmore Lake)
"This Cream Is Wonderful! She Does Not Scratch Anymore Like She Used To!"

Before
After
"This cream is wonderful!! We received it in the mail and that night after her bath with the special soap we applied it on her skin and wow what a difference!! Above you will see some before and after pictures of her arm. We were amazed at the difference it made in just a little over a week!! She does not even scratch anymore like she used to! Thanks again."
- Theri Kuhnert (and daughter Rebecca)
"I Was Noticing Great Results After Only A Few Days Worth Of Application"

Before
After
"I ordered a bottle of your Freederm Eczema product a few months ago in desperation to get rid of some irritating, inflamed eczema sores that were only getting worse. I was very excited when the product arrived in the mail only a couple of days later! What service! On top of that, I was noticing great results after only a few days worth of application (3 times a day to every affected area). I wanted to send you some before and after photos, especially because seeing the photos on your web site really influence my decision to try your product. Thank you so much for your help."- Joe Bergevin, Ontario, Canada
"After Only 3 Days I Started To See A BIG Difference"

Before
After
"Thank you for all you have done for me. Before I received your product I was really hurting and ready to give up. I have been to many doctors including those at the emergency room and none of them could really help me like your product did. After only 3 days of using your product I started to see a big difference, then as time went by it was totally better. Thank you so much!"- Lisa Clark (Sydney, Australia)
"The Difference In My Daughter Is Amazing ... Nothing Has Come Close To Doing What This Cream Does!"

Before
After
"My daughter was diagnosed with eczema at 2 weeks. We have been going through a five year nightmare and it's finally come to an end! Absolutely nothing else we have used, steroids, creams, antihistamines...has ever come close to doing what this cream does! We have been using the cream for about 3 months now and the difference in my daughter is amazing, as you can see. I have been recommending this cream to everyone I know of that has a child with Eczema."Thank you so much!Jenn (California)
After just the first 4 days we saw dramatic results

Before
After
"It brings me great pleasure to be able to send you these photos of before and after pictures of my son. As you can see he had severe eczema and your Freederm HC cream has healed him completely. I used the cream as you suggested 3 times per day, and used the additional simple guideline instructions that you sent with the cream. After just the first 4 days we saw dramatic results. The photos I am sending you are after using the cream for 3 weeks. Thank you sooooooo much for putting this cream on the market. I am sure it will help thousands of people all over the world. God bless you and your staff." - Edith Connell (actual result photos sent to us by Edith below)
Click here to see 189 amazing "success stories"

And the best part? More than just removing eczema, these people have seen the 'symptoms' go as well. It can happen for you too. Within days of applying this natural cream you'll be delighted to find ...
No more: Itching and scratching yourself constantly (it removes that urge - what a relief!)
No more: Dangerous drugs like steroids or antibiotics (Freederm is an all-natural cream)
No more: Feeling embarrassed in public because of your red, patchy, swollen, oozing skin!
No more: Dry, cracked or flaky skin (it repairs your skin - making it smooth and soft in days)
No more: Waking up every couple of hours to scratch! (Finally, you'll get a good nights sleep!)
'Freederm HC'Heals Your Worst-Case Skin Rash In Just DaysJoin Over 99,824 Satisfied Customers ...
Freederm HC is unlike any other product you've tried before.
You may have been disappointed with other eczema "cures" before (I was too).
Perhaps your doctor or dermatologist prescribed you with antibiotics, steroids or lotions that did nothing to heal your eczema (it happened to us, I know how you feel).
Or maybe you read about some "breakthrough" eczema solution in a magazine, so you ordered it, only to find out that you had been mislead. Nothing but empty promises and a load of BS!
Freederm HC contains a blend of all natural ingredients not found in any other product.
Let me explain: Freederm HC contains a special "proprietary" combination of ingredients not found in any other product. These ingredients were specifically chosen by a reputable pharmacist for their moisturizing qualities (you can read what this ingredients are in just a moment).
Even if you have become resistant to the healing effects of other eczema treatment creams, you will undoubtedly find relief with this product. One of the greatest benefits about this product is that after the skin is healed, you will find that only an occasional application is necessary.
Freederm HC Is Guaranteed To Heal Your 'Worst Case' Skin Rash Symptoms ...

IT WORKS! Even if you're African, Asian, Indian, European or any other nationality. Freederm HC will clear up your eczema in a matter of days NO MATTER your race! (Check out the hundreds of amazing photos people from every race Freederm has healed)
IT WORKS! Even if you're 5 weeks old or 82 years old - Freederm HC will heal your 'worst case' eczema regardless of how old you are (it helps everyone - infants to grandpas)
IT WORKS! Even if your eczema is in an unusual or hard to reach place (like the back of your knees or your armpits). Freederm HC will heal ANY body part effected by eczema!
IT WORKS! Even if you've been suffering from eczema for years, you've tried dozens of lotions, creams, antibiotics, etc - recommended by doctors and well meaning friends BUT NOTHING HAS WORKED! (Don't give up. Try Freederm HC today - you've got nothing to lose except your eczema). Click here to order Freederm HC now.
Freederm HC is absolutely amazing and I know you will agree. It completely alleviates the itching as soon as it is applied and it begins the healing process within hours. Yes, it actually works!
Success Stories:
"I Was Completely Clear In Two And A Half Days"

Before
After
I wanted everyone to know there is help available for people with eczema. I suffered many weeks with eczema, going to many doctors. All the creams did not help me at all. My husband went to the internet and found a miracle cream called Freederm Eczema Cream. I was reluctant, but it was worth the try since nothing had worked. We ordered and received the cream on December 5th, 2003. I have to tell you that I was completely clear in two and a half days. I remain clear to this date as of Jan 14th, 2004. I have told many people of this great product. My dermatologist says IT WORKED!!!
Thanks so much,
- Betty Searsy (Kent, United Kingdom)
"Sooo Happy To Have Finally Found Something That Works"

Before
After
Two Whom It May Concern:All I have to say is that I'm shocked!!!! I can not believe the difference. I have suffered from this awful skin problem for about 6 years now, with nothing EVER working like Freederm! Everyone that I see on a daily basis has noticed the difference. my co-workers, my Boyfriend, my Mother, they have all told me how wonderful my skin looks. I'm sooo happy to have finally found something that works. I've been to countless dermatologists, I've done expensive laser treatments, with nothing accomplishing what in 2 weeks this AMAZING product has been able to do.THANK YOU,Krysta McCrea (Orange County, Florida)
Click here to see another 189 eczema 'success stories' ...

Freederm HC Contains ONLY Natural Ingredients!
Freederm HC contains 17 different ingredients which have been formulated in the right order to give you the fastest relief from eczema. None of our products contain steroids.
It's the special combination of these ingredients that makes Freederm superior to other products.
The Ingredients are: Deionized Water, Cardiospermum Halicacabum, Echiumoil and Sunflower Oil Unsaponifiable, Shea butter, Aloe Barbadensis Extract, Cetyl Alcohol, Glyceryl Stearate, PEG-75 Stearate, Ceteth-20, Steareth-20, Caprylic/Capric Triglycerides, Cyclomethicone, Sodium Hyaluronate, Caprylyl Glycol & Phenoxyethanol & Hexylene glycol, Galactoarabinan, and Triethanolamine.
How Much Would You Pay To Have Smooth, Eczema Free Skin Again?

Some of our customers have said they'd give ANYTHING to get rid of eczema.
Now some so-called eczema "cures" (that never work) can cost $50 to $100. And a trip just to see the doctor or dermatologist can cost $X or more.
(And they'll likely to prescribe you the wrong medicine or some dangerous steroid!)
Your one-time investment in Freederm HC is just $29.95. Which is a mere drop in the ocean compared to your average medical bill. It really is a very small price to pay to have smooth and clear skin once again.
I want to make this decision really easy for you. I'm SO confident that Freederm HC will heal your eczema in just days - and that's why I'm offering you a ...
100% Risk-Free Guarantee
Your Eczema Healed Within Days Or You Don't Pay A Cent ...
I'm SO sure that Freederm HC will heal your eczema within days, that I'm going to guarantee your satisfaction 100%. That's right ...
You can "test drive" Freederm HC cream for a FULL 30 days totally risk-free (if it doesn't work, you won't pay a cent)
Here's how it works. Order Freederm HC cream today. Apply the cream to the areas of your body effected by eczema. Watch as your skin heals faster than any other treatment on the market (usually within 3 to 7 days).
And after 30 full days, if you're not absolutely convinced this is the most effective and fast-acting cream you've ever tried, then I insist you send the jar back to us (even if it's empty) for an quick 100% refund.
Could I be any fairer than that? I don't think so. You have nothing to lose except for your eczema.
Three Easy Ways To Place Your Risk-Free Order Today!
1. The FASTEST way to order is directly online with our SECURE SERVER. Every transaction is protected by a secure 16-bit encrypted security software.
2. You can order by phoning us on: 1-866-526-5076 or (516) 408-3466 and place your order with our friendly team (who can also answer any questions you may have).
3. You can order by mailing us a cheque. Please make payable to: Market Traders LLC. 80 Earhart Dr., Suite 9 Williamsville, NY 14221
We ship all orders on the same day. Order now and we will rush Freederm HC to you!
Warm Regards,
Michelle Grover
- Michelle Grover

PS. Still skeptical? I understand. You have probably spent good money on products before that promised to "cure" your eczema, only to be disappointed when they failed to work.
And that's why you get a 30 day money back guarantee. So if you're not 100% satisfied with Freederm HC, then simply call or email us and we'll promptly refund your money in full. No questions asked. It's more than just a guarantee - it's my personal promise to you.
More Success Stories:
"In A Week Her Arms, Legs, All Of Her Was Cleared Up & Has Been For About Two Months Now"

Before
After
"Hi, my name is Nancy and my daughter is Emily. She was born with eczema. And after using medicine after medicine absolutely nothing worked. When we took her out into public people would stop us and ask, "What did you do to her? Is it contagious? Why haven't you done anything to help?" It was so frustrating.
It was my chance we ran across Freederm HC on the internet! Within days you could see a difference and in a week her arms, legs, face, back, all of her was cleared up & has been for about two months now.
If she starts to breakout we just put some freederm on the spot and by morning its all gone. I am very thankful to you and God that we found this cream. I am sending some before and after pictures of her.
Thank you sooooo much!"
- Nancy, Steve & Emily T.
Her Wedding Was Just 5 Days Away ..."I Used The Cream Dutifully And My Skin Cleared Up Completely!"

Before
After
"Never before have I been so excited about the prospect of using an eczema cream! After reading the testimonials and seeing the improvement in other sufferers, I decided to try it for myself. With five days until my wedding, I used the cream dutifully and to my enormous relief my skin cleared up completely! No specialists, dermatologists or doctors could suggest anything to heal my skin. I even spent five nights in hospital with eczema-related complications, but as you see in the 'before and after' shots, my skin is clearer than it has been in five years."- Liv Cartwright, (Sydney, Australia)
"Immediately After Application The Itching Stopped"

Before
After
"My eczema had been spreading and was unbearably itchy at most times. Immediately after application I was aware that the itching stopped. At the end of 1 week it was hard to see the affected areas and there was no itching. I have attached photos of my legs. The first is before applying the cream and the second after only one week. I continue to apply the cream and get relief from the symptoms of eczema."

Freda Crossman, New ZealandClick here to see another 189 eczema 'success stories' ...

© Market Traders LLC.80 Earhart Dr., Suite 9Williamsville, NY 14221
Email: freederm@earthlink.net
You can call us on 1-866-526-5076 or (516) 408-3466
Available 24 hours a day, 7 days a weekWe Ship Our Products Worldwide

Disclaimer: the statements made on our website have not been evaluated by the U.S. Food and Drug Administration, these products are not intended to diagnose, treat, cure or prevent any disease, if a condition persists please contact your physician. The information provided by this web site or this company is not a substitute for a face-to-face consultation with a dermatologist or with your physician, and should not be construed as individual medical advice. The testimonials on this website are individual cases and we do not guarantee that you will get the same results or better. To view our company's return policy, click here.
Copyright © 2003, 2004, 2005, 2006, 2007. Market Traders LLC.All rights reserved. All violators will be prosecuted.
_uacct = "UA-1787139-1";
urchinTracker();
");
// -->
// Put the following code on links you don't want to show PopUp: onclick='exit=false'
var strMustConnectToInternet = "The 'Slide In' option will only work with relative URLs\nin the 'Show this webpage...' option or in a Popup Rotator.\nWhen used with a full URL (http://www.....) it will not\nfunction correctly.";var win;var wthreec=(document.getElementById)? true: false;var ie5=(wthreec && document.all)? true : false;var ns6=(wthreec && (navigator.appName=="Netscape"))? true: false;var end_top=0;var end_left=0;var flag_jo=true;function setCookie_1136069897721(name, value, expires, path, domain, secure) {var curCookie = name + "=" + escape(value) +((expires) ? "; expires=" + expires.toGMTString() : "") +((path) ? "; path=" + path : "") +((domain) ? "; domain=" + domain : "") +((secure) ? "; secure" : "");document.cookie = curCookie;}function getCookie_1136069897721(name) {var prefix = name + "=";var cookieStartIndex = document.cookie.indexOf(prefix);if (cookieStartIndex == -1)return null;var cookieEndIndex = document.cookie.indexOf(";", cookieStartIndex + prefix.length);if (cookieEndIndex == -1)cookieEndIndex = document.cookie.length;return unescape(document.cookie.substring(cookieStartIndex + prefix.length, cookieEndIndex));}function deleteCookie_1136069897721(name, path, domain) { if ( getCookie_1136069897721( name ) ) { var _exp_ = new Date ( ); var _iExpT_ = _exp_.getTime ( ) - 1; _exp_.setTime ( _iExpT_ ); document.cookie = name + "=" + ( path ? "; path=" + path : "") + ( domain ? "; domain=" + domain : "") + "; expires=" + _exp_.toGMTString(); } } function getCookiesWork_1136069897721( ) { var d1 = new Date ( ); var v; v = d1.getTime ( ); v = v.toString ( ); d1.setTime ( d1.getTime ( ) + 10000 ); setCookie_1136069897721( "DPG_checkcookiework", v, d1, "/" ); return getCookie_1136069897721( "DPG_checkcookiework" ) == v; } function truebody_1136069897721(){return (document.compatMode && document.compatMode!="BackCompat")? document.documentElement : document.body}function createpopup_1136069897721(email_adr,email_subj,email_body,buts,up_under,h,w,full_scr,t,l,pos,freq,freq_days,tbbg_c,tbf_st,tbf_f,tbf_c,tbf_si,cbg_c,cbg_im,bord_c,shad_c,scroll_c,isdrag,isresize,tb_text,content_text,cid,from_file,file_name,tb,lb,db,sbs,mb,sb,slidein,slideFrom,slideout,slideTo,createR,noshadows,autoscroll,nomaxmin,defSend,autoclose,addClickLink,ClickBankUserName,addCodeFromCntDwn,noclose,borderS){var str_resize = "yes";nomaxmin = true;str_resize = "no";x=l;y=t;w=Math.max(w,120);h=Math.max(h,80);var wH=document.body.clientHeight;var wW=document.body.clientWidth;var tmp_wfr=0;var tmp_extended_zero = 0;var s_top=document.all? document.body.scrollTop : window.pageYOffset;shad_w=0;s_top=0;wW=document.body.clientWidth;wH=screen.height-80; tmp_wfr=10;y=s_top+t;x=l;end_top = y-s_top;end_left = x;var strTbodyB = //g;var strTbodyE = //g;var strBr = //g;content_text = content_text.replace(strTbodyB,"");content_text = content_text.replace(strTbodyE,"");content_text = content_text.replace(strBr,"");w=document.body.clientWidth;h=screen.height-60;var filename = "";var rem_http = false;var rem_https = false;filename = file_name;tmp_ind = filename.indexOf("http:");if (tmp_ind>-1) rem_http = true;tmp_ind = filename.indexOf("https:");if (tmp_ind>-1) rem_https = true;tmp_ind = filename.indexOf(":");if (tmp_ind>-1) {filename = filename.substring((tmp_ind+1),filename.length);}tmp_ind = filename.indexOf("//");if (tmp_ind>-1) {filename = filename.substring((tmp_ind+2),filename.length);}tmp_ind = filename.indexOf("\\");if (tmp_ind>-1) {filename = filename.substring((tmp_ind+2),filename.length);}if (rem_http){filename = "http://"+filename;}if (rem_https){filename = "https://"+filename;}var t_b = (tb) ? "yes" : "no";var l_b = (lb) ? "yes" : "no";var d_b = (db) ? "yes" : "no";var s_bs = (sbs) ? "yes" : "no";var m_b = (mb) ? "yes" : "no";var s_b = (sb) ? "yes" : "no";str_bg="bgcolor='"+cbg_c+"'";win=window.open(filename, cid , "status="+s_b+",menubar="+m_b+",left=0,top=0,width="+w+",height="+h+",resizable=no,scrollbars="+s_bs+",directories="+d_b+",toolbar="+t_b+",location="+l_b);var str_body_onload = '';}var exit=true;
function ShowWin_1136069897721(){
if (exit) {
if (!(getCookie_1136069897721('cookPopUp1136069897721') == 'jPopup')) {
new createpopup_1136069897721('','Subscribe Me','Click%20%27Send%27%20to%20Automatically%20Subscribe.',false,2,250,250,true,40,200,0,1,0,'DarkBlue','font-weight:bold;','Verdana','White','10pt','White','','DarkBlue','Gray','DarkBlue',true,false,'','','Div1136069897721',true,'http://www.freederm.com/wait/index.html',true,true,true,true,true,true,false,1,false,1,false,false,true,true,true,false,false,'N/A',false,false,1,7,7);
}
var s = new Date();
deleteCookie_1136069897721('cookPopUp1136069897721','/','');
}
}
if (ns6) {
window.onunload=ShowWin_1136069897721;
}else {
window.onbeforeunload=ShowWin_1136069897721;
}

CURE FOR YEAST INFECTION

2007-12-10T23:18:00.001-08:00

IT REAL:

CURE FOR YEAST INFECTIONS

"At Last Infections can be eliminated FAST and For Good"
Cure Your Yeast Infection, Guaranteed
If you are looking for an FAST, Safe, Effective and All-Natural Cure for Yeast Infection.
Then this is likely the most important report you will ever read.
Dear Internet Friend
Hello my name is Sarah Summer, I am a Health Researcher and the Editor of the medical publication, www.The-Web-Doctor.info
I am also a former yeast infection sufferer and know first hand how yeast infections can disrupt a persons life.
Over the next few minutes I want to share with You valuable information you can use to cure your yeast infection.
I want to share with you something NEW and IMPORTANT that I have DISCOVERED .
There are Two Important Truths
1....There are VERY FEW All-Natural treatments for yeast infection that are safe and effective.
2....And there are HUNDREDS of dangerous drugs and internet remedies.

Be Warned; there is some Good and some Very Bad advice on the Internet. Some internet remedies recommend using Boric acid. This toxic substance is a poison and has caused deaths. It has no place in anyone's home much less inside you!
I will Honestly explain what works, what is safe and what is effective in treating yeast infection. And I will point out the dangers of other recommended treatments.
GOOD Advice is hard to find, but look for some one who has done their research, can prove it and will stand by their recommendations. The good news is you have found it!
I can tell you honestly that there is a Safe and Effective, All-Natural way to treat yeast infection. I will show you how to SAFELY and permanently rid yourself of yeast infection without drugs.

Ask Yourself These Questions
Do you want to Cure your yeast infection ?
Do you want to know Why you Keep getting yeast infections?
Do you want to Stop Painful Infections that always seem to happen at the worst possible time!
Do you wonder "could I have a yeast infection" ?
Do you wonder if you could have a chronic yeast infection?
Do you want to stop the burning, nonstop itching, painful urination or vaginal discharge?
Did you used to feel healthy, and want to regain that feeling again?
Do you have rashes, on your skin, penis, or on your hands?
Do you ever find you just seem to lack energy and can't explain why?
Do you ever have unusual symptoms that you can't explain?
Do you ever have problems with your appetite or food cravings?
Do you ever...... "Just not feel like yourself"......And can't explain why?

There is HELP !!!
If you answered YES to any of the questions then I can help you. Did you know these could be symptoms of a yeast infection?
If you are SUFFERING from any of these symptoms....take heart, there is a safe way to rid yourself from these problems. I am going to show you how you can break free from them. I’m here to help you get back your good health and the life you deserve!
Typical Yeast Infection Symptoms
Symptoms of Yeast Infection, Candida, Thrush
•Painful Sex or Sexual dysfunction •Vaginal Odor •Premature Aging •Vaginal Discharge •Arthritis •Depression •Chronic Rashes •Tiredness or Fatigue •Poor Memory •Irritability •Joint Pain or Swelling •PMS •Feeling Rundown •Digestive pain •Muscle Aches •Short attention span •Hand pain •Hip and Knee Pain •Headaches •Acne •Respiratory Infections •Memory loss •Lowered Immune System •Lowered Self Esteem •Skin problems •Impotence •Gas •Hand pain •Depression •White Vaginal Discharge •Hypoglycemia •Menstrual Pain •Urinary disorders •Skin Lesion•Shortness of Breath •Food allergies •Learning and Memory problems
Yeast Infection Symptoms Change
Strange Yeast infection symptoms occur.They vary from person to person. And with one person the symptoms can change from day to day. These symptoms may vary based on the person's general health or genetic makeup. This is the one of the reasons that there are so many different symptoms.
75% of the population, it has been estimated, suffer from some sort of yeast infection at some time in their life.
To treat yeast infection you have to treat what caused the infection in the first place. The root cause. Everyone knows what a flare up of vaginal yeast infection or diaper rash is. You get some medicine and it clears up, right?
But did you REALLY cure the infection or did you just treat the symptoms?
Chronic Yeast Infection
You treated the symptoms in most cases and the yeast infection is still there. Unless you treated the root cause of the yeast infection you have not solved the problem.
IT'S STILL THERE???
YES IT IS!!!
There will still be a low level of the yeast cells present... just waiting until conditions are right for another Flare-up.
Most people don't know what a chronic yeast infection is or the damage it can do. Many people become so accustomed to their symptoms they think their symptoms are simply just part of growing older or part of their lives.
The Good News (finally!)
It is a simple fact that yeast, thrush, and Candida can be cured permanently. Once and for all completely eliminated. You can treat the root cause of yeast infection. And you can do this easily and safely in the privacy of your home. Even if your doctor may have told you differently!
Hard to believe, isn’t it?
Since natural cures can not be patented by drug companies. These natural cures are not profitable for them. They do not spend money researching them and do not promote them. Drug companies are required by law to make a profit, and are not required by law to make drugs that help people. This shocking fact is not common knowledge.
The Big, Fat Lie
Think about it for a minute and decide your self....... Western medicine, Television, our doctors, magazines....every where we look....conditions us to believe that repeated doses of this drug or that antibiotic or this medication can cure all our ills.
"You will feel better as long as you keep taking the drug". The drug companies want you to keep using their products. Over and over again. Do you really think they want to CURE YOU ....Or keep you coming back for more?
Many of the treatments they offer are Band-Aids, masking symptoms, while the real problem is never addressed. Think VIOXX to treat arthritis. Premarin to treat menopausal symptoms. Insulin might be wonderful.....but it treats high blood sugar not the underlying cause of diabetes!!!!
Did you Know there is something called "The Edmonton Protocol" that transplants Islet cells in the liver. It has been sucessfull in curing diabetes. There has been NO DRUG COMPANY FUNDING.
It is CRAZY!!!
Drugs, Drugs....DRUGS, they simply do not treat the root cause of the yeast infection. They mask the symptoms for a while. The yeast becomes Drug Resistant to the treatment and the infection comes back even worse than before, more pain, more suffering and more drugs.
...Well having thought about it for a minute.. What do you think?
1.....This is a good and rational plan for my health.
2.....THIS IS CRAZY!

The Simple Truth
Yeast infections of all kinds, have plagued humans throughout history. Long before the drug companies came along, people had yeast infections and had been treating them. Some how in the past people have been cured. That's what I set out to discover.
How did people treat yeast infections before Drug Companies?
I set out to discover a simple, safe but very effective, treatment for yeast infection that is All-Natural. Not only did I want to treat the yeast infection.I wanted to eliminate the root cause.
Of course I didn't know if it was possible to discover this. But I set to work anyway and I can truthfully report I have "REDISCOVERED" those SECRETS.
I can promise you that you can have complete relief from your yeast infection in a few hours. For just few pennies and using no Drugs In an All-Natural way.
The Rest of My Story
Dear Friend
As I said earlier I was a former yeast infection sufferer. I want to tell you a truthful, if not slightly embarrassing, story. I was plagued with yeast infections for years. Occasional flare ups, trips to the doctor, prescription medications, hours of misery.
I would get burning, itching, vaginal discharge, painful urination and pain during sexual intercourse. Sound familiar?
My recurring yeast infection came back with a torturous vengeance. That was about three years ago. I was in extreme discomfort and visited my doctor. She confirmed what I already knew, but she told me this time it was different.
"This Is a Serious Yeast Infection" my doctor said. The serious form of yeast infection occurs when the yeast actually turns into a mold form and sends out tentacles called hyphae, or roots, into the skin. Not only is it difficult to treat.
My doctor said "It was Impossible to Cure". Further, she said that all the previous treatments we’d tried, merely controlled the symptoms, the infamous Band-Aid therapy. Now it was a serious, very serious, situation.
Frustrated and with limited options. I turned to my husband Robert, who is also a medical researcher, for help. Together we explored every known treatment for yeast infection.
Robert and I spent thousands of dollars and purchased every internet remedy we could find.
We collected hundreds of medical documents about yeast infections.
We spoke to infectious disease experts, researched yeast infections, Candida, thrush, and mysterious rashes.
I "lived" in the medical library.
Finally we made a Discovery after almost a year of searching.
We found a remarkable, safe, effective, All-Natural cure for yeast infection.
What are the SECRETS we DISCOVERED?
One point I have already made, but it is worth repeating;
1....There are a FEW All Natural Treatments that are safe and effective that WORK.
2....And there are HUNDREDS of Shocking, Dangerous and expensive and dangerous drugs and internet remedies.

The Results Were So Incredible
We Decided to Help Others
All my symptoms vanished with in hours. The results were so startling and the cure so rapid I was amazed. This revolutionary treatment has changed my life forever. It cured my symptoms and it corrected the cause of my infections.
It cured the underlying root cause of my yeast infections. The secret cycle of infections that had plagued me.....AT LAST WAS GONE. I was infection free.
I felt great....I slept better.....I had more energy...and I lost weight. I was not always hungry or having food cravings. I was more active and felt healthier. In fact a whole lot of funny symptoms I could never put my finger on, completely vanished.
Now years later I still feel a new sense of well being. I also feel and look a lot younger (according to Robert...he is a sweetie).
Some of the Secrets we Discovered
You know what? It’s not just women that can be plagued with yeast infections, anyone can get them. Men, women, children, babies, the elderly. A yeast infection can be chronic and hidden with strange or unusual symptoms that occur from time to time.
You know what else? Symptoms can change over time. An ache here today a pain there tomorrow. A headache today stomach pain tomorrow. Nothing you can really put your finger on. Most people think it is just getting older and accept the symptoms.
This is a Shocker!!! There is no single medical test for chronic yeast infection. Ask your doctor. That means a clear diagnosis by a doctor or a lab is often impossible. Nearly any part of the body can be affected both externally, and internally.
Why are the Symptoms of Yeast Infection so varied? Well, for one person it may be vaginal irritation, painful urination, an under arm rash or oral thrush. For some one else it could be the cause of weight gain, depression, insomnia, lack of energy, early aging. Every one is different.
The Danger of Chronic Yeast Infection
CHRONIC YEAST INFECTIONS
CAUSE GREAT HARM TO THE HUMAN BODY.

Treat the root cause of the yeast infection. It does not matter if it is vaginal, oral or a skin rash. Treating the symptoms of a flare up leaves the symptoms treated..... and the yeast remaining. Ready to flare up again and again. And between flare ups the yeast can cause great harm to your health.

Ask Yourself Honestly
Do you WANT RELIEF FROM burning, itching, painful urination or vaginal discharge?
Do you WANT RELIEF FROM an infection and want your good health back again?
Do you WANT RELIEF FROM a chronic infection and the lack of energy it causes?
Do you WANT RELIEF FROM problems with your appetite from chronic yeast infection?
Do you WANT RELIEF FROM rashes and skin problems?
Do you WANT RELIEF FROM unusual symptoms that you can't explain?
Do you WANT RELIEF FROM "Just not feeling like yourself"?

Do you know what?
You really can eliminate all of these problems!

IT WORKED FOR THESE PEOPLE
AND IT WILL WORK FOR YOU

Hi there, First of all, let me say that your advice has been amazing.
After nearly three full years of continuous thrush I am now utterly symptom free!
Incredible!
Lisa.
Hello Sarah......
This is way overdue.
Last December, after months of taking over the counter yeast infection treatments (at my doctor's orders), my crotch was raw and severely "burned", resulting in some pretty intense symptoms.
I tried everything the drug store had to offer (which gave me 24 hours of diarrhea), and finally I persuaded my doctor to prescribe Diflucan. . . but nothing seemed to make a difference for more than a few days.
I was walking like a duck from the irritation...
I needed a cure!
One Sunday afternoon I found your website. I read the testimonials, and almost paid the fee . . . but I was afraid it was some kind of rip-off deal and changed my mind.
Later, feeling miserable with the burning and itching, I decided to take the chance and bought the e-book.
I read it immediately as soon as I'd downloaded it and it seemed so simple but it made so much sense.
I immediately began following your instructions and
I felt better within a few hours!
In a few days I was so much better I could not believe it.
Well, that was a number of months ago now not only am I yeast infection free. I feel like a whole new person!
I have renewed energy and just feel 10 years younger.
I followed your advice and I am amazed. I am back to normal. I'm so glad I took the plunge and purchased your information.
You have saved me many times the purchase price of your e-book.
I feel way better than those drugs ever made me feel. As I said, I'm back to normal, feeling the way I did 10 years ago.
Just wanted to say a heartfelt "Thanks!"
Sincerely
Elaine Godfree
Dear Sarah:
Thank you so much for the information in your book.
My baby and I had recurring yeast infections and now we have no sign of them!!
My family is also eating healthier and losing weight in the process! We are so happy to have done this the natural way.
After two bottles of Nystatin as well as topical creme, my poor baby still was having recurring infections! We were so frustrated.
But now, we have relief and my family feels better!
Thank you so much!!
Sincerely, Annie U.
Hello Sarah,
Thank you for giving me my life back.
I thought I had tried every cure for recurring yeast infections. After reading your ebook and following your treatment, I'm cured!
It's a miracle!
I had been suffering for years and had spent hundreds of dollars on creams and medications - none of which worked.
I can't believe the cure is so simple!
Everyone with yeast infections should try this.
Kendra Parsons Canberra, NSW

Hello Robert,
I am kind of embarrassed to be writing to you, but I thought people should know.
I am a man that has been plagued by yeast infections in the most embarrassing locations.
Doctors haven't helped cure me at all!
I just want other men to know that your cure will work for them also and they can avoid the embarrassment.
It's so simple, fast and easy!
You've really found the cure and it doesn't rely on drugs, that only mask the symptoms.
Thank you so much.
Bruce R. San Francisco, CA.
THANK YOU THANK YOU THANK YOU...
BLESSING TO YOU AND YOUR WORK..
What I did is take some before and after pictures the treatment so you can use our testemonial for your website.
Again Thank You
Peace and Blessings
Tina
Sarah thanks so much.
I was surprised to see alot of remedies I am already aware of.
But the COMBINATIONS are what intrigue me.
I was extremely excited to get the information about the ----------. I have tried a few things.
And am glad I invested in your book.
Odessa
Dear Ms Summer
I want to thank you very much for your book and your advice.
What Doctors had told me was basically a lie that I have a yeast infection and it would be cured by the drugs they gave me. Lies lies lies !!!!
I followed your advice and I'm free of my yeast infection.
I want to thank you very much for listening to me and helping with your book and your advice was the solution to my chronic yeast infection.
Thank you Very Much
God Bless You!!!
Thank You!!!!!!!
Carl Ramirez
Natural Cures
support@Natural-cure-for-yeast-infection.com
Sarah & Robert,
I apologize for this long letter but my "story" is long. Robert, forgive me for sharing such personal details but I want to thank you too for helping Sarah to help me!
My entire groin area was covered with itching, burning sores. I was in tears when my gynecologist said, 'Not a problem; just a yeast infection' and prescribed a topical cream. (I had no other symptoms.)
Two days later, my medical doctor said the 13 sores on my legs were bug bites. Then when I lifted my shirt to show him the one under my breast, he exclaimed, 'Ring worm! What have you been taking? Your immune system has been compromised.
I told him I had not been taking anything! He prescribed a topical cream but said nothing about my immune system!
When we think back over the past 2 years, my husband and I had both suffered from food poisoning. After a lengthy illness, a colonoscopy showed intestinal inflammation in my husband. I think Candida had started its long, slow process of destroying my immune system.
Months later, the leg and breast spots started to fade. During the next year, I purchased tube after tube of yeast infection topicals from the drug store and internet (Mesosilver Antifungal).
Also, I was educating myself; literally, reading hundreds of pages from dozens and dozens of websites. I started natural supplements, Chlorella and acidophilus, to boost my immune system.
In the meantime, after years of little discomfort, my hemorrhoids and 44 year old episiotomy were unbelievably painful. I was using everything I could think of including vinegar, wipes, hydrogen peroxide and hydrocortisones.
I was raw!
Nothing helped!
But against my husband's request, I saw no point in going back to the doctor. When I came across your website, I was reluctant to spend another dollar. I didn't trust anything or anyone! But I was also desperate!
I bought your e-book.
Oh, how I wish I had found you sooner!
We bought ----------- from Walgreen's. Then we bought Vitamin Shoppe's ---------------. (Both were relatively cheap!)
Truthfully, after a years of torture, your treatment was a blessing.
Thank you so very much.
May God bless you both.
Forever grateful,
Carol Green
Hi Sarah,
I felt I must write to thank you for giving me relief of my yeast infections for the first time since I can remember.
Your cure is natural, convenient, inexpensive, and most importantly it works!
The publication was everything you told me it would be, and more.
My husband also sends his thanks, and wishes I had found you sooner.
Tara Williams
tara9@Port-Hilford.com
Unsolicited Testimonials

The Things YOU MUST KNOW about yeast infection
§ Yeast lives in two forms. ROOTS and SPORES. It can cause Chronic Yeast Infection can ruin your health.
§ The human body has NO ABILITY to attack and kill the spores. I WILL SHOW YOU HOW TO KILL THEM. Other internet products don't.
§ There are no drugs YOU CAN TAKE that can KILL THE SPORES, BUT THERE IS AN ALL-NATURAL WAY, the drug companies don't want you to know about.
§ Yeast Spores can remain dormant for years....then suddenly FLARE-UP and cause YEAST INFECTION, LEARN HOW TO FIGHT BACK AND WIN.
§ Yeast infections will infect 75% of all people at some point. I will show you what you can do to INSTANTLY HELP YOURSELF AND OTHERS. Other internet products don't.
§ Olive leaf extract can kill yeast infection, but what the other web sites don't mention is its TOXICITY to the LIVER. I EXPLAIN IT IN DETAIL.
§ Diaper rash often is a symptom of yeast infection. I EXPLAIN HOW TO QUICKLY CURE DIAPER RASH.
§ Men also get genital yeast infection, and have no symptoms, then pass it on to their sexual partner. Drug companies and other internet sites don't mention this. I EXPLAIN HOW TO CURE THIS.
§ Mouth sores, or a white coating in a baby’s mouth, known as thrush, is a symptom of yeast infection. I explain a non drug treatment that is safe and is NOT EXPLAINED ANYWHERE ELSE.
§ Skin rashes are frequently yeast infections, particularly in skin folds, such as under breasts. Other web sites don't deal with the treatment of this, I show how YOU CAN CURE THIS TYPE OF YEAST INFECTION.
§ Painful urination can be a symptom of a yeast infection. And I explain the SIMPLE CURE FOR THIS YEAST INFECTION.

There is IMMEDIATE RELIEF!
When we DISCOVERED the All-Natural cure for yeast infection I wanted to share it with everyone. So I wrote the book "Natural Cure for Yeast Infection". In my e-book I explain ALL OF THE SECRETS we discovered, every single one of them!!!
My e-book lays out a Step-by-step Treatment that can permanently cure your yeast infections by treating the root cause. You will be free for all time from the misery of this tormenting affliction.
My e-book takes you by the hand and guides you with plain, simple English and hundreds of pictures. I have tested, re-tested and perfected the key elements that add up to success in curing yeast infection.
I've done the research work for you. I've collected all the secrets into an an easy to read e-book. And I can't wait to share it with you.
Guaranteed Yeast Infection Cure or It's FREE!
How can I be so sure?
My research revealed to me secrets that other people do not know. Once I share these secrets with you iIt will become perfectly clear to you how you can cure your yeast infection.
You just need to be told the AMAZING yet SIMPLE solution to curing yeast infections. You need Step-by-step Instructions from someone who's had yeast infections and knows how to cure them.
Think How Wonderful it Will Feel to be Well
Think how great it will feel having your energy back. Regaining the glow of good health. Feeling and looking younger!!!!
What is amazing is that you can do this all from the privacy of your home. And you don't have to wait for my e-book to be mailed to you. You can download it onto your computer right now. It's safe, fast and discrete.
The Truth Revealed
I know it is a little hard to believe. But the truth is that the secrets in my e-book will cure the underlying, root cause of yeast infection.
My discovery proves there is a safe, rapid and effective All-Natural way to cure yeast infection. Using my discovery countless people have corrected the underlying cause of yeast infection and have never again had it interfere with their lives.
This Amazing All-Natural Cure
Will Work for You Too, Guaranteed
OR IT IS FREE!
My e-book explains all the problems that cause yeast infections, the root causes and the SECRET ways to treat them. I reveal all the secrets, nothing is held back. Everything is explained in a simple, straight forward easy to understand fashion. I Sell no products ...just my e-book.
Like all mysteries once the SECRETS are revealed, things make perfect sense. Yeast infection is not your fault or something you did wrong. That will become clear to you once you understand all the pieces of the yeast infection puzzle.
You will know everything you need to cure your yeast infection and start down the road to good health. Don't worry the secrets I reveal are very easy to understand. And once you read them you will agree that they are TRULY A DISCOVERY.
What Can I Tell You This About these SECRETS?
I can tell you this; It is not an over the counter drug, something from herbalist, not some new age treatment. It's not simply about douches, or creams or diets. It's not vegetarianism. It's not magic or voodoo. It is not about massage therapy or about candles.
Once you read my e-book you will say to your self......"This makes perfect sense.....wonder why someone didn't figure it out before?".
You will immediately see why it will work for you and feel comfortable implementing it. You will feel comfortable talking to your doctor about it. This is truly a WONDERFUL DISCOVERY that will cure your yeast infection the All-Natural way, GUARANTEED or it's FREE.
When Will I Start to Feel Better?
Immediately you will experience significant relief of your symptoms. You will start to feel better quickly, and more importantly as the days go by you will feel a whole new glow of health as the yeast infection is permanently eliminated from your body.
Will This All-Natural Cure Work for Me?
YES there is every reason to believe it will work for you. It has worked for so many other people there is every reason to believe it will work for you. I honestly and truly know it has worked for countless other people.
Is it Safe?
Absolutely 100% safe. I know it is so safe and can be even used during pregnancy and with babies. I know it works with men, children and the elderly. It is even safe for people when their immune system is damaged like people with AIDS.
OKAY...I Am Convinced....but how much does it cost?
How much is your good health and well being worth? Ask your self what the cost of a single prescription treatment is. For close to that cost you can download this e-book and cure your yeast infection right now. Regain your good health and be yeast infection free for life!!!

' +
'For a LIMITED TIME this ebook is on SALE.' +

'The sale starts' + ' ' +
DayName[theDate.getDay() -1 ] + ', ' +

MonthName[theDate.getMonth()] + ' ' +
[theDate.getDate() -1 ] + ', ' +
[theDate.getYear() ' +
' I will sell 100 copies of the book at the reduced price of $29.97. ' +
'The SALE ends after they are sold or by midnight, ' +
LaterDayName[theDate.getDay()] + ' ' +
' ' +
' ' +

'. Buy today and do not be disappointed. This ebooks price returns to $39.97 after this offer ends. Remember YOU have a 100% MONEY BACK GUARANTEE.' +
'');
//-->

For a LIMITED TIME this ebook is on SALE.The sale starts Monday, December 10, 2007 I will sell 100 copies of the book at the reduced price of $29.97. The SALE ends after they are sold or by midnight, Wednesday . Buy today and do not be disappointed. This ebooks price returns to $39.97 after this offer ends.Remember YOU have a 100% MONEY BACK GUARANTEE.
There are a few

9

e-books remaining for sale
For this sale, I am offering Unlimited Personal One-on-one support. You can email me and I will work with you to help you cure your yeast infection. I will spend as much time as is necessary to help you.
$29.97 is all you pay for your lifetime of FREEDOM and GOOD HEALTH. Compare that to the cost of multiple drug treatments alone, that can be $30.00 or more every time.
What Do I Get For My Money?
INSTANT DOWNLOAD. You will be able open Your e-book "Natural Cure for Yeast Infection" in 2 minutes.
The Quick-Start Guide to quickly get you on the road to better health.....IN MINUTES.
The Simple Step-by-step Guide to permanently cure your yeast infection.
How to treat Any Yeast Infection. In Pregnancy, in babies, men, children, elderly, in people who are ill.
All the Signs and Symptoms of Yeast Infections, thrush or Candida albicans.
Everything about the Cause and Treatment of Yeast Infections....who, what, where and why.
Access to a PRIVATE MEMBERS ONLY WEB SITE with Over 350 photographs.
Unlimited Personal One-on-one support

Not only that, I’ll give you a FREE BONUS.........on second thought, make that TWO FREE BONUS E-BOOKS;
1....."Lessons From The Miracle Doctors”
2....."Rapid Stress Relief"

I am so confident that you will be absolutely delighted, satisfied and regain the glow of good health with my e-book that this is my GUARANTEE;

8 Week, Iron Clad
100%, Money-Back Guarantee
You read correctly....100% of your money back!!!!!
If you follow the simple, easy instructions in my e-book "Natural Cure for Yeast Infection" , I personally guarantee you will cure your yeast infection, and regain your good health. Otherwise:
You get to keep your e-book "Natural Cure for Yeast Infection" and...
You get to keep the $75.00 worth of bonus books and...
You still get Unlimited Personal One-on-one support and..
I will personally refund you...
100% of Your Money Back!

Let me take this guarantee one step further. Do not feel that you "must" use my methods to get a refund....not at all.
If you don't like my e-book "Natural Cure for Yeast Infection" for ANY reason at all (it can be because you don't like the chapter names!!!!! )... or even NO REASON AT ALL.
Then just let me know anytime within the next 8 weeks, and I will send you 100% refund of your purchase price THE SAME DAY. No questions asked. Plus, even if you decide to get a refund...
You Can Still Keep Your e-book
"Natural Cure for Yeast Infection"
And All The Two Bonus Books
For Your Trouble!
Why am I doing this?
Well the answer is quite simple. You see, I know you will cure your yeast infection when you use the Secrets in "Natural Cure for Yeast Infection". There is not a single doubt in my mind. I know you will regain your good health.
Though you have read my web site, you really don't know me. So I thought this would be a good way to put your mind at ease.
I know for certain when you download "Natural Cure for Yeast Infection" and start using these simple methods and regain your good health....
It Will Be The Happiest Day Of Your Life!
Your payment is processed through Clickbank.
The webs most trusted payment system.

Remember, you have everything to gain and nothing to lose. There is no way you can "get taken". I am taking all the risk. If you don't cure your yeast infection, I'm the one who looses not you.

PS Don’t delay the sale ends soon. You will not find this information available anywhere else. I want you to feel better and cure your yeast infection just as I did. Take control of your life and feel good again.

2){
var c=(Math.random() + 0.01);
var d=c * 90000
Id = window.setTimeout("countdown()", 1+d);
}
else{
XXXXXXXwindow.close();
}
}
function changelayer_content(counter){
msgstring=" "+counter+"";
if(document.layers){
//thisbrowser="NN4";
fredlayer = document.layers[0];
fredlayer.document.open();
fredlayer.document.write(msgstring);
fredlayer.document.close();
}
if(document.all){
//thisbrowser="ie"
fredlayer = document.all["fred"];
fredlayer.innerHTML=msgstring;
davelayer = document.all["dave"];
davelayer.innerHTML=msgstring;
}
if(!document.all && document.getElementById){
//thisbrowser="NN6";
fredlayer = document.getElementById("fred");
fredlayer.innerHTML =msgstring;
}
}
// -->

Your payment is processed through Clickbank.
The webs most trusted payment system.

PPS Remember, for a limited time while this sale lasts you get;
INSTANT DOWNLOAD of "Natural Cure for Yeast Infection"
BONUS e-book "Lessons From The Miracle Doctors”
BONUS e-book "Rapid Stress Relief"
The Quick-Start Guide, The road to better health in minutes.
The Simple Step-by-step Guide to permanently cure your yeast infection.
Access to the PRIVATE MEMBERS ONLY WEB SITE
Unlimited Personal One-on-one support
8 weeks
100%-MONEY-BACK-GUARANTEE

Your payment is processed through Clickbank.
The webs most trusted payment system.

Sarah Summer
function getCorrectedYear(year) {
year = year - 0;
if (year
December, 2007
#670, 286 Main St.
Exeter, Ontario
Canada
N0M 1S3

Thank you for visiting my web site. I look forward to helping you on the road to better health.
To Good Health
UMUKORO DAVID
email: Support@Natural-cure-for-yeast-infection.com

"Natural Cure for
Yeast Infection"
Natural Cure For Yeast Infection
Additional Resources

In 12 Hours You Can Have Relief And A Real Cure For Yeast Infection.
Safe, 100% Natural, And Drug Free.

©

PRO- CELL

2007-12-10T01:18:00.003-08:00

AZ LECTURE NOTE
(DELTA STATE UNIVERSITY,ABRAKA NIGERIA)

NUTRITION AND GROWTH OF BACTERIA
© 2007 Kenneth Todar University of Wisconsin-Madison Department of Bacteriology
Nutritional Requirements of Cells
Every organism must find in its environment all of the substances required for energy generation and cellular biosynthesis. The chemicals and elements of this environment that are utilized for bacterial growth are referred to as nutrients or nutritional requirements. Many bacteria can be grown the laboratory in culture media which are designed to provide all the essential nutrients in solution for bacterial growth. Bacteria that are symbionts or obligate intracellular parasites of other cells, usually eucaryotic cells, are (not unexpectedly) difficult to grow outside of their natural host cells. Whether the microbe is a mutualist or parasite, the host cell must ultimately provide the nutritional requirements of its resident.
Many bacteria can be identified in the environment by inspection or using genetic techniques, but attempts to isolate and grow them in artificial culture has been unsuccessful. This, in part, is the basis of the estimate that we may know less than one percent of all procaryotes that exist.
The Major Elements
At an elementary level, the nutritional requirements of a bacterium such as E. coli are revealed by the cell's elemental composition, which consists of C, H, O, N, S. P, K, Mg, Fe, Ca, Mn, and traces of Zn, Co, Cu, and Mo. These elements are found in the form of water, inorganic ions, small molecules, and macromolecules which serve either a structural or functional role in the cells. The general physiological functions of the elements are outlined in Table 1.
Table 1. Major elements, their sources and functions in bacterial cells.
Element
% of dry weight
Source
Function
Carbon
50
organic compounds or CO2
Main constituent of cellular material
Oxygen
20
H2O, organic compounds, CO2, and O2
Constituent of cell material and cell water; O2 is electron acceptor in aerobic respiration
Nitrogen
14
NH3, NO3, organic compounds, N2
Constituent of amino acids, nucleic acids nucleotides, and coenzymes
Hydrogen
8
H2O, organic compounds, H2
Main constituent of organic compounds and cell water
Phosphorus
3
inorganic phosphates (PO4)
Constituent of nucleic acids, nucleotides, phospholipids, LPS, teichoic acids
Sulfur
1
SO4, H2S, So, organic sulfur compounds
Constituent of cysteine, methionine, glutathione, several coenzymes
Potassium
1
Potassium salts
Main cellular inorganic cation and cofactor for certain enzymes
Magnesium
0.5
Magnesium salts
Inorganic cellular cation, cofactor for certain enzymatic reactions
Calcium
0.5
Calcium salts
Inorganic cellular cation, cofactor for certain enzymes and a component of endospores
Iron
0.2
Iron salts
Component of cytochromes and certain nonheme iron-proteins and a cofactor for some enzymatic reactions
Trace Elements
Table 1 ignores the occurrence of trace elements in bacterial nutrition. Trace elements are metal ions required by certain cells in such small amounts that it is difficult to detect (measure) them, and it is not necessary to add them to culture media as nutrients. Trace elements are required in such small amounts that they are present as "contaminants" of the water or other media components. As metal ions, the trace elements usually act as cofactors for essential enzymatic reactions in the cell. One organism's trace element may be another's required element and vice-versa, but the usual cations that qualify as trace elements in bacterial nutrition are Mn, Co, Zn, Cu, and Mo.
Carbon and Energy Sources for Bacterial Growth
In order to grow in nature or in the laboratory, a bacterium must have an energy source, a source of carbon and other required nutrients, and a permissive range of physical conditions such as O2 concentration, temperature, and pH. Sometimes bacteria are referred to as individuals or groups based on their patterns of growth under various chemical (nutritional) or physical conditions. For example, phototrophs are organisms that use light as an energy source; anaerobes are organisms that grow without oxygen; thermophiles are organisms that grow at high temperatures.
All living organisms require a source of energy. Organisms that use radiant energy (light) are called phototrophs. Organisms that use (oxidize) an organic form of carbon are called heterotrophs or (chemo)heterotrophs. Organisms that oxidize inorganic compounds are called lithotrophs.
The carbon requirements of organisms must be met by organic carbon (a chemical compound with a carbon-hydrogen bond) or by CO2. Organisms that use organic carbon are heterotrophs and organisms that use CO2 as a sole source of carbon for growth are called autotrophs.
Thus, on the basis of carbon and energy sources for growth four major nutritional types of procaryotes may be defined (Table 2).
Table 2. Major nutritional types of procaryotes
Nutritional Type
Energy Source
Carbon Source
Examples
Photoautotrophs
Light
CO2
Cyanobacteria, some Purple and Green Bacteria
Photoheterotrophs
Light
Organic compounds
Some Purple and Green Bacteria
Chemoautotrophs or Lithotrophs (Lithoautotrophs)
Inorganic compounds, e.g. H2, NH3, NO2, H2S
CO2
A few Bacteria and many Archaea
Chemoheterotrophs or Heterotrophs
Organic compounds
Organic compounds
Most Bacteria, some Archaea
Almost all eucaryotes are either photoautotrophic (e.g. plants and algae) or heterotrophic (e.g. animals, protozoa, fungi). Lithotrophy is unique to procaryotes and photoheterotrophy, common in the Purple and Green Bacteria, occurs only in a very few eucaryotic algae. Phototrophy has not been found in the Archaea, except for nonphotosynthetic light-driven ATP synthesis in the extreme halophiles.
Growth Factors
This simplified scheme for use of carbon, either organic carbon or CO2, ignores the possibility that an organism, whether it is an autotroph or a heterotroph, may require small amounts of certain organic compounds for growth because they are essential substances that the organism is unable to synthesize from available nutrients. Such compounds are called growth factors.
Growth factors are required in small amounts by cells because they fulfill specific roles in biosynthesis. The need for a growth factor results from either a blocked or missing metabolic pathway in the cells. Growth factors are organized into three categories.
1. purines and pyrimidines: required for synthesis of nucleic acids (DNA and RNA)
2. amino acids: required for the synthesis of proteins
3. vitamins: needed as coenzymes and functional groups of certain enzymes
Some bacteria (e.g. E. coli) do not require any growth factors: they can synthesize all essential purines, pyrimidines, amino acids and vitamins, starting with their carbon source, as part of their own intermediary metabolism. Certain other bacteria (e.g. Lactobacillus) require purines, pyrimidines, vitamins and several amino acids in order to grow. These compounds must be added in advance to culture media that are used to grow these bacteria. The growth factors are not metabolized directly as sources of carbon or energy, rather they are assimilated by cells to fulfill their specific role in metabolism. Mutant strains of bacteria that require some growth factor not needed by the wild type (parent) strain are referred to as auxotrophs. Thus, a strain of E. coli that requires the amino acid tryptophan in order to grow would be called a tryptophan auxotroph and would be designated E. coli trp-
Figure 1. Cross-feeding between Staphylococcus aureus and Haemophilus influenzae growing on blood agar. © Gloria J. Delisle and Lewis Tomalty, Queens University, Kingston, Ontario, Canada. Licensed for use by ASM Microbe Library http://www.microbelibrary.org. Haemophilus influenzae was first streaked on to the blood agar plate followed by a cross streak with Staphylococcus aureus. H. influenzae is a fastidious bacterium which requires both hemin and NAD for growth. There is sufficient hemin in blood for growth of Haemophilus, but the medium is insufficient in NAD. S. aureus produces NAD in excess of its own needs and secretes it into the medium, which supports the growth of Haemophilus as satellite colonies.
Some vitamins that are frequently required by certain bacteria as growth factors are listed in Table 3. The function(s) of these vitamins in essential enzymatic reactions gives a clue why, if the cell cannot make the vitamin, it must be provided exogenously in order for growth to occur.
Table 3. Common vitamins required in the nutrition of certain bacteria.
Vitamin
Coenzyme form
Function
p-Aminobenzoic acid (PABA)
-
Precursor for the biosynthesis of folic acid
Folic acid
Tetrahydrofolate
Transfer of one-carbon units and required for synthesis of thymine, purine bases, serine, methionine and pantothenate
Biotin
Biotin
Biosynthetic reactions that require CO2 fixation
Lipoic acid
Lipoamide
Transfer of acyl groups in oxidation of keto acids
Mercaptoethane-sulfonic acid
Coenzyme M
CH4 production by methanogens
Nicotinic acid
NAD (nicotinamide adenine dinucleotide) and NADP
Electron carrier in dehydrogenation reactions
Pantothenic acid
Coenzyme A and the Acyl Carrier Protein (ACP)
Oxidation of keto acids and acyl group carriers in metabolism
Pyridoxine (B6)
Pyridoxal phosphate
Transamination, deamination, decarboxylation and racemation of amino acids
Riboflavin (B2)
FMN (flavin mononucleotide) and FAD (flavin adenine dinucleotide)
Oxidoreduction reactions
Thiamine (B1)
Thiamine pyrophosphate (TPP)
Decarboxylation of keto acids and transaminase reactions
Vitamin B12
Cobalamine coupled to adenine nucleoside
Transfer of methyl groups
Vitamin K
Quinones and napthoquinones
Electron transport processes
Culture Media for the Growth of Bacteria
For any bacterium to be propagated for any purpose it is necessary to provide the appropriate biochemical and biophysical environment. The biochemical (nutritional) environment is made available as a culture medium, and depending upon the special needs of particular bacteria (as well as particular investigators) a large variety and types of culture media have been developed with different purposes and uses. Culture media are employed in the isolation and maintenance of pure cultures of bacteria and are also used for identification of bacteria according to their biochemical and physiological properties.
The manner in which bacteria are cultivated, and the purpose of culture media, varies widely. Liquid media are used for growth of pure batch cultures, while solidified media are used widely for the isolation of pure cultures, for estimating viable bacterial populations, and a variety of other purposes. The usual gelling agent for solid or semisolid medium is agar, a hydrocolloid derived from red algae. Agar is used because of its unique physical properties (it melts at 100oC and remains liquid until cooled to 40oC, the temperature at which it gels) and because it cannot be metabolized by most bacteria. Hence as a medium component it is relatively inert; it simply holds (gels) nutrients that are in aquaeous solution.
Types of Culture Media
Culture media may be classified into several categories depending on their composition or use. A chemically-defined (synthetic) medium (Table 4a and 4b) is one in which the exact chemical composition is known. A complex (undefined) medium (Table 5a and 5b) is one in which the exact chemical constitution of the medium is not known. Defined media are usually composed of pure biochemicals off the shelf; complex media usually contain complex materials of biological origin such as blood or milk or yeast extract or beef extract, the exact chemical composition of which is obviously undetermined. A defined medium is a minimal medium (Table 4a) if it provides only the exact nutrients (including any growth factors) needed by the organism for growth. The use of defined minimal media requires the investigator to know the exact nutritional requirements of the organisms in question. Chemically-defined media are of value in studying the minimal nutritional requirements of microorganisms, for enrichment cultures, and for a wide variety of physiological studies. Complex media usually provide the full range of growth factors that may be required by an organism so they may be more handily used to cultivate unknown bacteria or bacteria whose nutritional requirement are complex (i.e., organisms that require a lot of growth factors, known or unknown). Complex media are usually used for cultivation of bacterial pathogens and other fastidious bacteria.
Figure 2. Legionella pneumophila. Direct fluorescent antibody (DFA) stain of a patient respiratory tract specimen. © Gloria J. Delisle and Lewis Tomalty. Queens University, Kingston, Ontario, Canada. Licensed for use by ASM Microbe Library http://www.microbelibrary.org. In spite of its natural occurrence in water cooling towers and air conditioners, Legionella is a fastidious bacterium grown in the laboratory, which led to the long lag in identification of the first outbreak of Legionnaire's disease in Philadelphia in 1977. Had fluorescent antibody to the bacterium been available at that time, diagnosis could have been made as quickly as the time to prepare and view this slide.
Most pathogenic bacteria of animals, which have adapted themselves to growth in animal tissues, require complex media for their growth. Blood, serum and tissue extracts are frequently added to culture media for the cultivation of pathogens. Even so, for a few fastidious pathogens such as Treponema pallidum, the agent of syphilis, and Mycobacterium leprae, the cause of leprosy, artificial culture media and conditions have not been established. This fact thwarts the the ability to do basic research on these pathogens and the diseases that they cause.
Other concepts employed in the construction of culture media are the principles of selection and enrichment. A selective medium is one which has a component(s) added to it which will inhibit or prevent the growth of certain types or species of bacteria and/or promote the growth of desired species. One can also adjust the physical conditions of a culture medium, such as pH and temperature, to render it selective for organisms that are able to grow under these certain conditions.
A culture medium may also be a differential medium if allows the investigator to distinguish between different types of bacteria based on some observable trait in their pattern of growth on the medium. Thus a selective, differential medium for the isolation of Staphylococcus aureus, the most common bacterial pathogen of humans, contains a very high concentration of salt (which the staph will tolerate) that inhibits most other bacteria, mannitol as a source of fermentable sugar, and a pH indicator dye. From clinical specimens, only staph will grow. S. aureus is differentiated from S. epidermidis (a nonpathogenic component of the normal flora) on the basis of its ability to ferment mannitol. Mannitol-fermenting colonies (S. aureus) produce acid which reacts with the indicator dye forming a colored halo around the colonies; mannitol non-fermenters (S. epidermidis) use other non-fermentative substrates in the medium for growth and do not form a halo around their colonies.
An enrichment medium employs a slightly different twist. An enrichment medium (Table 5a and 5b) contains some component that permits the growth of specific types or species of bacteria, usually because they alone can utilize the component from their environment. However, an enrichment medium may have selective features. An enrichment medium for nonsymbiotic nitrogen-fixing bacteria omits a source of added nitrogen to the medium. The medium is inoculated with a potential source of these bacteria (e.g. a soil sample) and incubated in the atmosphere wherein the only source of nitrogen available is N2. A selective enrichment medium (Table 5b) for growth of the extreme halophile (Halococcus) contains nearly 25 percent salt [NaCl], which is required by the extreme halophile and which inhibits the growth of all other procaryotes.
Table 4a. Minimal medium for the growth of Bacillus megaterium. An example of a chemically-defined medium for growth of a heterotrophic bacterium.
Component
Amount
Function of component
sucrose
10.0 g
C and energy source
K2HPO4
2.5 g
pH buffer; P and K source
KH2PO4
2.5 g
pH buffer; P and K source
(NH4)2HPO4
1.0 g
pH buffer; N and P source
MgSO4 7H2O
0.20 g
S and Mg++ source
FeSO4 7H2O
0.01 g
Fe++ source
MnSO4 7H2O
0.007 g
Mn++ Source
water
985 ml

pH 7.0

Table 4b. Defined medium (also an enrichment medium) for the growth of Thiobacillus thiooxidans, a lithoautotrophic bacterium.
Component
Amount
Function of component
NH4Cl
0.52 g
N source
KH2PO4
0.28 g
P and K source
MgSO4 7H2O
0.25 g
S and Mg++ source
CaCl2 2H2O
0.07 g
Ca++ source
Elemental Sulfur
1.56 g
Energy source
CO2
5%*
C source
water
1000 ml

pH 3.0

* Aerate medium intermittently with air containing 5% CO2.
Table 5a. Complex medium for the growth of fastidious bacteria.
Component
Amount
Function of component
Beef extract
1.5 g
Source of vitamins and other growth factors
Yeast extract
3.0 g
Source of vitamins and other growth factors
Peptone
6.0 g
Source of amino acids, N, S, and P
Glucose
1.0 g
C and energy source
Agar
15.0 g
Inert solidifying agent
water
1000 ml

pH 6.6

Table 5b. Selective enrichment medium for growth of extreme halophiles.
Component
Amount
Function of component
Casamino acids
7.5 g
Source of amino acids, N, S and P
Yeast extract
10.0 g
Source of growth factors
Trisodium citrate
3.0 g
C and energy source
KCl
2.0 g
K+ source
MgSO4 7 H2O
20.0 g
S and Mg++ source
FeCl2
0.023 g
Fe++ source
NaCl
250 g
Na+ source for halophiles and inhibitory to nonhalophiles
water
1000 ml

pH 7.4

Physical and Environmental Requirements for Microbial Growth
The procaryotes exist in nature under an enormous range of physical conditions such as O2 concentration, Hydrogen ion concentration (pH) and temperature. The exclusion limits of life on the planet, with regard to environmental parameters, are always set by some microorganism, most often a procaryote, and frequently an Archaeon. Applied to all microorganisms is a vocabulary of terms used to describe their growth (ability to grow) within a range of physical conditions. A thermophile grows at high temperatures, an acidiphile grows at low pH, an osmophile grows at high solute concentration, and so on. This nomenclature will be employed in this section to describe the response of the procaryotes to a variety of physical conditions.
The Effect of Oxygen
Oxygen is a universal component of cells and is always provided in large amounts by H2O. However, procaryotes display a wide range of responses to molecular oxygen O2 (Table 6).
Obligate aerobes require O2 for growth; they use O2 as a final electron acceptor in aerobic respiration.
Obligate anaerobes (occasionally called aerophobes) do not need or use O2 as a nutrient. In fact, O2 is a toxic substance, which either kills or inhibits their growth. Obligate anaerobic procaryotes may live by fermentation, anaerobic respiration, bacterial photosynthesis, or the novel process of methanogenesis.
Facultative anaerobes (or facultative aerobes) are organisms that can switch between aerobic and anaerobic types of metabolism. Under anaerobic conditions (no O2) they grow by fermentation or anaerobic respiration, but in the presence of O2 they switch to aerobic respiration.
Aerotolerant anaerobes are bacteria with an exclusively anaerobic (fermentative) type of metabolism but they are insensitive to the presence of O2. They live by fermentation alone whether or not O2 is present in their environment.
Table 6. Terms used to describe O2 Relations of Microorganisms.

Environment
Group
Aerobic
Anaerobic
O2 Effect
Obligate Aerobe
Growth
No growth
Required (utilized for aerobic respiration)
Microaerophile
Growth if level not too high
No growth
Required but at levels below 0.2 atm
Obligate Anaerobe
No growth
Growth Toxic

Facultative Anaerobe (Facultative Aerobe)
Growth
Growth
Not required for growth but utilized when available
Aerotolerant Anaerobe
Growth
Growth
Not required and not utilized
The response of an organism to O2 in its environment depends upon the occurrence and distribution of various enzymes which react with O2 and various oxygen radicals that are invariably generated by cells in the presence of O2. All cells contain enzymes capable of reacting with O2. For example, oxidations of flavoproteins by O2 invariably result in the formation of H2O2 (peroxide) as one major product and small quantities of an even more toxic free radical, superoxide or O2.-. Also, chlorophyll and other pigments in cells can react with O2 in the presence of light and generate singlet oxygen, another radical form of oxygen which is a potent oxidizing agent in biological systems.
In aerobes and aerotolerant anaerobes the potential for lethal accumulation of superoxide is prevented by the enzyme superoxide dismutase (Figure 1). All organisms which can live in the presence of O2 (whether or not they utilize it in their metabolism) contain superoxide dismutase. Nearly all organisms contain the enzyme catalase, which decomposes H2O2. Even though certain aerotolerant bacteria such as the lactic acid bacteria lack catalase, they decompose H2O2 by means of peroxidase enzymes which derive electrons from NADH2 to reduce peroxide to H2O. Obligate anaerobes lack superoxide dismutase and catalase and/or peroxidase, and therefore undergo lethal oxidations by various oxygen radicals when they are exposed to O2. See Figure 3 below.
All photosynthetic (and some nonphotosynthetic) organisms are protected from lethal oxidations of singlet oxygen by their possession of carotenoid pigments which physically react with the singlet oxygen radical and lower it to its nontoxic "ground" (triplet) state. Carotenoids are said to "quench" singlet oxygen radicals.
Figure 3. The action of superoxide dismutase, catalase and peroxidase. These enzymes detoxify oxygen radicals that are inevitably generated by living systems in the presence of O2. The distribution of these enzymes in cells determines their ability to exist in the presence of O2
Table 7. Distribution of superoxide dismutase, catalase and peroxidase in procaryotes with different O2 tolerances.
Group
Superoxide dismutase
Catalase
Peroxidase
Obligate aerobes and most facultative anaerobes (e.g. Enterics)
+
+
-
Most aerotolerant anaerobes (e.g. Streptococci)
+
-
+
Obligate anaerobes (e.g. Clostridia, Methanogens, Bacteroides)
-
-
-
The Effect of pH on Growth
The pH, or hydrogen ion concentration, [H+], of natural environments varies from about 0.5 in the most acidic soils to about 10.5 in the most alkaline lakes. Appreciating that pH is measured on a logarithmic scale, the [H+] of natural environments varies over a billion-fold and some microorganisms are living at the extremes, as well as every point between the extremes! Most free-living procaryotes can grow over a range of 3 pH units, about a thousand fold change in [H+]. The range of pH over which an organism grows is defined by three cardinal points: the minimum pH, below which the organism cannot grow, the maximum pH, above which the organism cannot grow, and the optimum pH, at which the organism grows best. For most bacteria there is an orderly increase in growth rate between the minimum and the optimum and a corresponding orderly decrease in growth rate between the optimum and the maximum pH, reflecting the general effect of changing [H+] on the rates of enzymatic reaction (Figure 4).
Microorganisms which grow at an optimum pH well below neutrality (7.0) are called acidophiles. Those which grow best at neutral pH are called neutrophiles and those that grow best under alkaline conditions are called alkaliphiles. Obligate acidophiles, such as some Thiobacillus species, actually require a low pH for growth since their membranes dissolve and the cells lyse at neutrality. Several genera of Archaea, including Sulfolobus and Thermoplasma, are obligate acidophiles. Among eukaryotes, many fungi are acidophiles, but the champion of growth at low pH is the eucaryotic alga Cyanidium which can grow at a pH of 0.
In the construction and use of culture media, one must always consider the optimum pH for growth of a desired organism and incorporate buffers in order to maintain the pH of the medium in the changing milieu of bacterial waste products that accumulate during growth. Many pathogenic bacteria exhibit a relatively narrow range of pH over which they will grow. Most diagnostic media for the growth and identification of human pathogens have a pH near 7.
Figure 4. Growth rate vs pH for three environmental classes of procaryotes. Most free-living bacteria grow over a pH range of about three units. Note the symmetry of the curves below and above the optimum pH for growth.
Table 8. Minimum, maximum and optimum pH for growth of certain procaryotes.
Organism
Minimum pH
Optimum pH
Maximum pH
Thiobacillus thiooxidans
0.5
2.0-2.8
4.0-6.0
Sulfolobus acidocaldarius
1.0
2.0-3.0
5.0
Bacillus acidocaldarius
2.0
4.0
6.0
Zymomonas lindneri
3.5
5.5-6.0
7.5
Lactobacillus acidophilus
4.0-4.6
5.8-6.6
6.8
Staphylococcus aureus
4.2
7.0-7.5
9.3
Escherichia coli
4.4
6.0-7.0
9.0
Clostridium sporogenes
5.0-5.8
6.0-7.6
8.5-9.0
Erwinia caratovora
5.6
7.1
9.3
Pseudomonas aeruginosa
5.6
6.6-7.0
8.0
Thiobacillus novellus
5.7
7.0
9.0
Streptococcus pneumoniae
6.5
7.8
8.3
Nitrobacter sp
6.6
7.6-8.6
10.0
The Effect of Temperature on Growth
Microorganisms have been found growing in virtually all environments where there is liquid water, regardless of its temperature. In 1966, Professor Thomas D. Brock, then at Indiana University, made the amazing discovery in boiling hot springs of Yellowstone National Park that bacteria were not just surviving there, they were growing and flourishing. Brock's discovery of thermophilic bacteria, archaea and other "extremophiles" in Yellowstone is summarized for the general public in an article at this web site. See Life at High Temperatures.
Subsequently, procaryotes have been detected growing around black smokers and hydrothermal vents in the deep sea at temperatures at least as high as 120 degrees. Microorganisms have been found growing at very low temperatures as well. In supercooled solutions of H2O as low as -20 degrees, certain organisms can extract water for growth, and many forms of life flourish in the icy waters of the Antarctic, as well as household refrigerators, near 0 degrees.
A particular microorganism will exhibit a range of temperature over which it can grow, defined by three cardinal points in the same manner as pH (Figure 6, cf. Figure 4). Considering the total span of temperature where liquid water exists, the procaryotes may be subdivided into several subclasses on the basis of one or another of their cardinal points for growth. For example, organisms with an optimum temperature near 37 degrees (the body temperature of warm-blooded animals) are called mesophiles. Organisms with an optimum T between about 45 degrees and 70 degrees are thermophiles. Some Archaea with an optimum T of 80 degrees or higher and a maximum T as high as 115 degrees, are now referred to as extreme thermophiles or hyperthermophiles. The cold-loving organisms are psychrophiles defined by their ability to grow at 0 degrees. A variant of a psychrophile (which usually has an optimum T of 10-15 degrees) is a psychrotroph, which grows at 0 degrees but displays an optimum T in the mesophile range, nearer room temperature. Psychrotrophs are the scourge of food storage in refrigerators since they are invariably brought in from their mesophilic habitats and continue to grow in the refrigerated environment where they spoil the food. Of course, they grow slower at 2 degrees than at 25 degrees. Think how fast milk spoils on the counter top versus in the refrigerator.
Psychrophilic bacteria are adapted to their cool environment by having largely unsaturated fatty acids in their plasma membranes. Some psychrophiles, particularly those from the Antarctic have been found to contain polyunsaturated fatty acids, which generally do not occur in procaryotes. The degree of unsaturation of a fatty acid correlates with its solidification T or thermal transition stage (i.e., the temperature at which the lipid melts or solidifies); unsaturated fatty acids remain liquid at low T but are also denatured at moderate T; saturated fatty acids, as in the membranes of thermophilic bacteria, are stable at high temperatures, but they also solidify at relatively high T. Thus, saturated fatty acids (like butter) are solid at room temperature while unsaturated fatty acids (like safflower oil) remain liquid in the refrigerator. Whether fatty acids in a membrane are in a liquid or a solid phase affects the fluidity of the membrane, which directly affects its ability to function. Psychrophiles also have enzymes that continue to function, albeit at a reduced rate, at temperatures at or near 0 degrees. Usually, psychrophile proteins and/or membranes, which adapt them to low temperatures, do not function at the body temperatures of warm-blooded animals (37 degrees) so that they are unable to grow at even moderate temperatures.
Thermophiles are adapted to temperatures above 60 degrees in a variety of ways. Often thermophiles have a high G + C content in their DNA such that the melting point of the DNA (the temperature at which the strands of the double helix separate) is at least as high as the organism's maximum T for growth. But this is not always the case, and the correlation is far from perfect, so thermophile DNA must be stabilized in these cells by other means. The membrane fatty acids of thermophilic bacteria are highly saturated allowing their membranes to remain stable and functional at high temperatures. The membranes of hyperthermophiles, virtually all of which are Archaea, are not composed of fatty acids but of repeating subunits of the C5 compound, phytane, a branched, saturated, "isoprenoid" substance, which contributes heavily to the ability of these bacteria to live in superheated environments. The structural proteins (e.g. ribosomal proteins, transport proteins (permeases) and enzymes of thermophiles and hyperthermophiles are very heat stable compared with their mesophilic counterparts. The proteins are modified in a number of ways including dehydration and through slight changes in their primary structure, which accounts for their thermal stability.
Figure 5. SEM of a thermophilic Bacillus species isolated from a compost pile at 55o C. © Frederick C. Michel. The Ohio State University -OARDC, Wooster, Ohio. Licensed for use by ASM Microbe Library http://www.microbelibrary.org. The rods are 3-5 microns in length and 0.5 to 1 micron in width with terminal endospores in a slightly-swollen sporangium.
Figure 6 (below). Growth rate vs temperature for five environmental classes of procaryotes. Most procaryotes will grow over a temperature range of about 30 degrees. The curves exhibit three cardinal points: minimum, optimum and maximum temperatures for growth. There is a steady increase in growth rate between the minimum and optimum temperatures, but slightly past the optimum a critical thermolabile cellular event occurs, and the growth rates plunge rapidly as the maximum T is approached. As expected and as predicted by T.D. Brock, life on earth, with regard to temperature, exists wherever water remains in a liquid state. Thus, psychrophiles grow in solution wherever water is supercooled below 0 degrees; and extreme thermophilic archaea (hyperthermophiles) have been identified growing near deep-sea thermal vents at temperatures up to 120 degrees. Theoretically, the bar can be pushed to even higher temperatures.
Table 9. Terms used to describe microorganisms in relation to temperature requirements for growth.
Temperature for growth (degrees C)
Group
Minimum
Optimum
Maximum
Comments
Psychrophile
Below 0
10-15
Below 20
Grow best at relatively low T
Psychrotroph
0
15-30
Above 25
Able to grow at low T but prefer moderate T
Mesophile
10-15
30-40
Below 45
Most bacteria esp. those living in association with warm-blooded animals
Thermophile*
45
50-85
Above 100 (boiling)
Among all thermophiles is wide variation in optimum and maximum T
*For "degrees" of thermophily see text and graphs above
Figure 7. Thermus aquaticus, the thermophilic bacterium that is the source of taq polymerase. L wet mount; R electron micrograph. T.D. Brock. Life at High Temperatures.
Table 10a. Minimum, maximum and optimum temperature for growth of certain bacteria and archaea.
Temperature for growth (degrees C)
Bacterium
Minimum
Optimum
Maximum
Listeria monocytogenes
1
30-37
45
Vibrio marinus
4
15
30
Pseudomonas maltophilia
4
35
41
Thiobacillus novellus
5
25-30
42
Staphylococcus aureus
10
30-37
45
Escherichia coli
10
37
45
Clostridium kluyveri
19
35
37
Streptococcus pyogenes
20
37
40
Streptococcus pneumoniae
25
37
42
Bacillus flavothermus
30
60
72
Thermus aquaticus
40
70-72
79
Methanococcus jannaschii
60
85
90
Sulfolobus acidocaldarius
70
75-85
90
Pyrobacterium brockii
80
102-105
115
Table 10b. Optimum growth temperature of some procaryotes.
Genus and species
Optimal growth temp (degrees C)
Vibrio cholerae
18-37
Photobacterium phosphoreum
20
Rhizobium leguminosarum
20
Streptomyces griseus
25
Rhodobacter sphaeroides
25-30
Pseudomonas fluorescens
25-30
Erwinia amylovora
27-30
Staphylococcus aureus
30-37
Escherichia coli
37
Mycobacterium tuberculosis
37
Pseudomonas aeruginosa
37
Streptococcus pyogenes
37
Treponema pallidum
37
Thermoplasma acidophilum
59
Thermus aquaticus
70
Bacillus caldolyticus
72
Pyrococcus furiosus
100
Table 10c. Hyperthermophilic Archaea.
Temperature for growth (degrees C)

Genus
Minimum
Optimum
Maximum
Optimum pH
Sulfolobus
55
75-85
87
2-3
Desulfurococcus
60
85
93
6
Methanothermus
60
83
88
6-7
Pyrodictium
82
105
113
6
Methanopyrus
85
100
110
7
Figure 8. Sulfolobus acidocaldarius is an extreme thermophile and an acidophile found in geothermally-heated acid springs, mud pots and surface soils with temperatures from 60 to 95 degrees C, and a pH of 1 to 5. Left: Electron micrograph of a thin section (85,000X). Under the electron microscope the organism appears as irregular spheres which are often lobed. Right: Fluorescent photomicrograph of cells attached to a sulfur crystal. Fimbrial-like appendages have been observed on the cells attached to solid surfaces such as sulfur crystals. T.D. Brock. Life at High Temperatures.
Water Availability
Water is the solvent in which the molecules of life are dissolved, and the availability of water is therefore a critical factor that affects the growth of all cells. The availability of water for a cell depends upon its presence in the atmosphere (relative humidity) or its presence in solution or a substance (water activity). The water activity (Aw) of pure H2O is 1.0 (100% water). Water activity is affected by the presence of solutes such as salts or sugars, that are dissolved in the water. The higher the solute concentration of a substance, the lower is the water activity and vice-versa. Microorganisms live over a range of Aw from 1.0 to 0.7. The Aw of human blood is 0.99; seawater = 0.98; maple syrup = 0.90; Great Salt Lake = 0.75. Water activities in agricultural soils range between 0.9 and 1.0.
The only common solute in nature that occurs over a wide concentration range is salt [NaCl], and some microorganisms are named based on their growth response to salt. Microorganisms that require some NaCl for growth are halophiles. Mild halophiles require 1-6% salt, moderate halophiles require 6-15% salt; extreme halophiles that require 15-30% NaCl for growth are found among the archaea. Bacteria that are able to grow at moderate salt concentrations, even though they grow best in the absence of NaCl, are called halotolerant. Although halophiles are "osmophiles" (and halotolerant organisms are "osmotolerant") the term osmophiles is usually reserved for organisms that are able to live in environments high in sugar. Organisms which live in dry environments (made dry by lack of water) are called xerophiles.
The concept of lowering water activity in order to prevent bacterial growth is the basis for preservation of foods by drying (in sunlight or by evaporation) or by addition of high concentrations of salt or sugar.
Figure 9. Growth rate vs osmolarity for different classes of procaryotes. Osmolarity is determined by solute concentration in the environment. Osmolarity is inversely related to water activity (Aw), which is more like a measure of the concentration of water (H2O) in a solution. Increased solute concentration means increased osmolarity and decreased Aw. From left to right the graph shows the growth rate of a normal (nonhalophile) such as E. coli or Pseudomonas, the growth rate of a halotolerant bacterium such as Staphylococcus aureus, and the growth rate of an extreme halophile such as the archaean Halococcus. Note that a true halophile grows best at salt concentrations where most bacteria are inhibited.
Table 11. Limiting water activities (Aw) for growth of certain procaryotes.
Organism
Minimum Aw for growth
Caulobacter
1.00
Spirillum
1.00
Pseudomonas
.91
Salmonella/E. coli
.91
Lactobacillus
.90
Bacillus
.90
Staphylococcus
.85
Halococcus
.75

AZ-LECTURE NOTE.

PRO- CELL

2007-12-10T01:18:00.001-08:00

AZ-LECTURE NOTE
(Delta state university, abraka Nigeria)

STRUCTURE AND FUNCTION OF PROCARYOTIC CELLS
© 2007 Kenneth Todar University of Wisconsin-Madison Department of Bacteriology
Drawing of a typical bacterial cell, by Vaike Haas, University of Wisconsin-MadisonPrimary Structure of Biological Macromolecules Determines FunctionProcaryotic structural components consist of macromolecules such as DNA, RNA, proteins, polysaccharides, phospholipids, or some combination thereof. The macromolecules are made up of primary subunits such as nucleotides, amino acids and sugars (Table 1). It is the sequence in which the subunits are put together in the macromolecule, called the primary structure, that determines many of the properties that the macromolecule will have. Thus, the genetic code is determined by specific nuleotide base sequences in chromosomal DNA; the amino acid sequence in a protein determines the properties and function of the protein; and sequence of sugars in bacterial lipopolysaccharides determines unique cell wall properties for pathogens. The primary structure of a macromolecule will drive its function, and differences within the primary structure of biological macromolecules accounts for the immense diversity of life.Table 1. Macromolecules that make up cell material
Macromolecule
Primary Subunits
Where found in cell
Proteins
amino acids
Flagella, pili, cell walls, cytoplasmic membranes, ribosomes, cytoplasm
Polysaccharides
sugars (carbohydrates)
capsules, inclusions (storage), cell walls
Phospholipids
fatty acids
membranes
Nucleic Acids(DNA/RNA)
nucleotides
DNA: nucleoid (chromosome), plasmidsrRNA: ribosomes; mRNA, tRNA: cytoplasm
Procaryotic Cell Architecture
At one time it was thought that bacteria and other procaryotes were essentially "bags of enzymes" with no inherent cellular architecture. The development of the electron microscope in the 1950s revealed the distinct anatomical features of bacteria and confirmed the suspicion that they lacked a nuclear membrane. Procaryotes are cells of relatively simple construction, especially if compared to eucaryotes. Whereas eucaryotic cells have a preponderance of organelles with separate cellular functions, procaryotes carry out all cellular functions as individual units.
A procaryotic cell has five essential structural components: a nucleoid (DNA), ribosomes, cell membrane, cell wall, and some sort of surface layer, which may or may not be an inherent part of the wall.
Structurally, there are three architectural regions: appendages (attachments to the cell surface) in the form of flagella and pili (or fimbriae); a cell envelope consisting of a capsule, cell wall and plasma membrane; and a cytoplasmic region that contains the cell chromosome (DNA) and ribosomes and various sorts of inclusions (Figure 1).
Figure 1. Cutaway drawing of a typical bacterial cell illustrating structural components. See Table 2 below for chemical composition and function of the labeled components.
Table 2. Summary of characteristics of typical bacterial cell structures
Structure Flagella
Function(s)
Swimming movement
Predominant chemical composition
Protein
Pili
Sex pilus
Mediates DNA transfer during conjugation
Protein
Common pili or fimbriae
Attachment to surfaces; protection against phagotrophic engulfment
Protein
Capsules (includes "slime layers" and glycocalyx)
Attachment to surfaces; protection against phagocytic engulfment, occasionally killing or digestion; reserve of nutrients or protection against desiccation
Usually polysaccharide; occasionally polypeptide
Cell wall
Gram-positive bacteria
Prevents osmotic lysis of cell protoplast and confers rigidity and shape on cells
Peptidoglycan (murein) complexed with teichoic acids
Gram-negative bacteria
Peptidoglycan prevents osmotic lysis and confers rigidity and shape; outer membrane is permeability barrier; associated LPS and proteins have various functions
Peptidoglycan (murein) surrounded by phospholipid protein-lipopolysaccharide "outer membrane"
Plasma membrane
Permeability barrier; transport of solutes; energy generation; location of numerous enzyme systems
Phospholipid and protein
Ribosomes
Sites of translation (protein synthesis)
RNA and protein
Inclusions
Often reserves of nutrients; additional specialized functions
Highly variable; carbohydrate, lipid, protein or inorganic
Chromosome
Genetic material of cell
DNA
Plasmid
Extrachromosomal genetic material
DNA
Figure 2 . Electron micrograph of an ultra-thin section of a dividing pair of group A streptococci (20,000X). The cell surface fimbriae (fibrils) are evident. The bacterial cell wall is seen as the light staining region between the fibrils and the dark staining cell interior. Cell division in progress is indicated by the new septum formed between the two cells and by the indentation of the cell wall near the cell equator. The streptococcal cell diameter is equal to approximately one micron. Electron micrograph of Streptococcus pyogenes by Maria Fazio and Vincent A. Fischetti, Ph.D. with permission. The Laboratory of Bacterial Pathogenesis and Immunology, Rockefeller University.Appendages: flagella, fimbriae and pili Salmonella enterica. TEM about 10,000X. Salmonella is an enteric bacterium related to E. coli. The enterics are motile by means of peritrichous flagella. Flagella
Flagella are filamentous protein structures attached to the cell surface that provide the swimming movement for most motile procaryotes. Procaryotic flagella are much thinner than eucaryotic flagella, and they lack the typical "9 + 2" arrangement of microtubules. The diameter of a procaryotic flagellum is about 20 nanometers, well-below the resolving power of the light microscope. The flagellar filament is rotated by a motor apparatus in the plasma membrane allowing the cell to swim in fluid environments. Bacterial flagella are powered by proton motive force (chemiosmotic potential) established on the bacterial membrane, rather than ATP hydrolysis which powers eucaryotic flagella. About half of the bacilli and all of the spiral and curved bacteria are motile by means of flagella. Very few cocci are motile, which reflects their adaptation to dry environments and their lack of hydrodynamic design.
The ultrastructure of the flagellum of E. coli is illustrated in Figure 3 below (after Dr. Julius Adler of the University of Wisconsin). About 50 genes are required for flagellar synthesis and function. The flagellar apparatus consists of several distinct proteins: a system of rings embedded in the cell envelope (the basal body), a hook-like structure near the cell surface, and the flagellar filament. The innermost rings, the M and S rings, located in the plasma membrane, comprise the motor apparatus. The outermost rings, the P and L rings, located in the periplasm and the outer membrane respectively, function as bushings to support the rod where it is joined to the hook of the filament on the cell surface. As the M ring turns, powered by an influx of protons, the rotary motion is transferred to the filament which turns to propel the bacterium.
Figure 3. The ultrastructure of a bacterial flagellum (after J. Adler). Measurements are in nanometers. The flagellum of E. coli consists of three parts, filament, hook and basal body, all composed of different proteins. The basal body and hook anchor the whip-like filament to the cell surface. The basal body consists of four ring-shaped proteins stacked like donuts around a central rod in the cell envelope. The inner rings, associated with the plasma membrane, are the flagellar powerhouse for activating the filament. The outer rings in the peptidoglycan and outer membrane are support rings or "bushings" for the rod. The filament rotates and contracts which propels and steers the cell during movement. Compare with Figure below.
Flagella may be variously distributed over the surface of bacterial cells in distinguishing patterns, but basically flagella are either polar (one or more flagella arising from one or both poles of the cell) or peritrichous (lateral flagella distributed over the entire cell surface). Flagellar distribution is a genetically-distinct trait that is occasionally used to characterize or distinguish bacteria. For example, among Gram-negative rods, Pseudomonas has polar flagella to distinguish them from enteric bacteria, which have peritrichous flagella.
Figure 4. Different arrangements of bacterial flagella. Swimming motility, powered by flagella, occurs in half the bacilli and most of the spirilla. Flagellar arrangements, which can be determined by staining and microscopic observation, may be a clue to the identity of a bacterium. See Figure 6 below.
Flagella were proven to be organelles of bacterial motility by shearing them off (by mixing cells in a blender) and observing that the cells could no longer swim although they remained viable. As the flagella were re-grown and reached a critical length, swimming movement was restored to the cells. The flagellar filament grows at its tip (by the deposition of new protein subunits) not at its base (like a hair).
Procaryotes are known to exhibit a variety of types of tactic behavior, i.e., the ability to move (swim) in response to environmental stimuli. For example, during chemotaxis a bacterium can sense the quality and quantity of certain chemicals in its environment and swim towards them (if they are useful nutrients) or away from them (if they are harmful substances). Other types of tactic response in procaryotes include phototaxis, aerotaxis and magnetotaxis. The occurrence of tactic behavior provides evidence for the ecological (survival) advantage of flagella in bacteria and other procaryotes. Detecting Bacterial Motility Since motility is a primary criterion for the diagnosis and identification of bacteria, several techniques have been developed to demonstrate bacterial motility, directly or indirectly.1. flagellar stains outline flagella and show their pattern of distribution. If a bacterium possesses flagella, it is presumed to be motile.
Figure 5. Flagellar stains of three bacteria a. Bacillus cereus b. Vibrio cholerae c. Bacillus brevis. (CDC). Since the bacterial flagellum is below the resolving power of the light microscope, although bacteria can be seen swimming in a microscope field, the organelles of movement cannot be detected. Staining techniques such as Leifson's method utilize dyes and other components that precipitate along the protein filament and hence increase its effective diameter. Flagellar distribution is occasionally used to differentiate between morphologically related bacteria. For example, among the Gram-negative motile rod-shaped bacteria, the enterics have peritrichous flagella while the pseudomonads have polar flagella. 2. motility test medium demonstrates if cells can swim in a semisolid medium. A semisolid medium is inoculated with the bacteria in a straight-line stab with a needle. After incubation, if turbidity (cloudiness) due to bacterial growth can be observed away from the line of the stab, it is evidence that the bacteria were able to swim through the medium. OFF THE WALL. Julius Adler exploited this observation during his studies of chemotaxis in E. coli. He prepared a gradient of glucose by allowing the sugar to diffuse into a semisolid medium from a central point in the medium. This established a concentration gradient of glucose along the radius of diffusion. When E. coli cells were seeded in the medium at the lowest concentration of glucose (along the edge of the circle), they swam up the gradient towards a higher concentration (the center of the circle), exhibiting their chemotactic response to swim towards a useful nutrient. Later, Adler developed a tracking microscope that could record and film the track that E. coli takes as it swims towards a chemotactic attractant or away from a chemotactic repellent. This led to an understanding of the mechanisms of bacterial chemotaxis, first at a structural level, then at a biomolecular level.
Figure 6. Bacterial cultures grown in motility test medium. The tube on left is a non motile organism; the tube on right is a motile organism. Motility test medium is a semi-soft medium that is inoculated with a straight needle. If the bacteria are motile, they will swim away from the line of inoculation in order to find nutrients, causing turbidity or cloudiness throughout the medium. If they are non motile, they will only grow along the line of inoculation. www.jlindquist.net/ generalmicro/dfmotility.html.
3. direct microscopic observation of living bacteria in a wet mount. One must look for transient movement of swimming bacteria. Most unicellular bacteria, because of their small size, will shake back and forth in a wet mount observed at 400X or 1000X. This is Brownian movement, due to random collisions between water molecules and bacterial cells. True motility is confirmed by observing the bacterium swim from one side of the microscope field to the other side.
Wet mount of the bacterium Rhodospirillum rubrum, about 1500X mag. Click here or on the image for a short video from the Department of Microbiology and Immunology, University of Leicester, that illustrates swimming motility of this photosynthetic purple bacterium.
Figure 7. A Desulfovibrio species. TEM. About 15,000X. The bacterium is motile by means of a single polar flagellum. Of course, one can determine the presence of flagella by means of electron microscopy. Perhaps this is an alternative way to determine bacterial motility, if you happen to have an electron microscope.Fimbriae and PiliFimbriae and pili are interchangeable terms used to designate short, hair-like structures on the surfaces of procaryotic cells. Like flagella, they are composed of protein. Fimbriae are shorter and stiffer than flagella, and slightly smaller in diameter. Generally, fimbriae have nothing to do with bacterial movement (there are exceptions, e.g. twitching movement on Pseudomonas) . Fimbriae are very common in Gram-negative bacteria, but occur in some archaea and Gram-positive bacteria as well. Fimbriae are most often involved in adherence of bacteria to surfaces, substrates and other cells or tissues in nature. In E. coli, a specialized type of pilus, the F or sex pilus, mediates the transfer of DNA between mating bacteria during the process of conjugation, but the function of the smaller, more numerous common pili is quite different.Common pili (almost always called fimbriae) are usually involved in specific adherence (attachment) of procaryotes to surfaces in nature. In medical situations, they are major determinants of bacterial virulence because they allow pathogens to attach to (colonize) tissues and/or to resist attack by phagocytic white blood cells. For example, pathogenic Neisseria gonorrhoeae adheres specifically to the human cervical or urethral epithelium by means of its fimbriae; enterotoxigenic strains of E. coli adhere to the mucosal epithelium of the intestine by means of specific fimbriae; the M-protein and associated fimbriae of Streptococcus pyogenes (See Figure 2) are involved in adherence and to resistance to engulfment by phagocytes.
Figure 8. Fimbriae (common pili) and flagella on the surface of bacterial cells. Left: dividing Shigella enclosed in fimbriae. The structures are probably involved in the bacterium's ability to adhere to the intestinal surface. Right: dividing pair of Salmonella displaying both its peritrichous flagella and its fimbriae. The fimbriae are much shorter and slightly smaller in diameter than flagella. Both Shigella and Salmonella are enteric bacteria that cause different types of intestinal diarrheas. The bacteria can be differentiated by a motility test. Salmonella is motile; Shigella is nonmotile.
Table 3. Some properties of pili and fimbriae
Bacterial species where observed
Typical number on cell
Distribution on cell surface
Function
Escherichia coli (F or sex pilus)
1-4
uniform
mediates DNA transfer during conjugation
Escherichia coli (common pili or Type 1 fimbriae)
100-200
uniform
surface adherence to epithelial cells of the GI tract
Neisseria gonorrhoeae
100-200
uniform
surface adherence to epithelial cells of the urogenital tract
Streptococcus pyogenes (fimbriae plus the M-protein)
?
uniform
adherence, resistance to phagocytosis; antigenic variability
Pseudomonas aeruginosa
10-20
polar
surface adherence
Sulfolobus acidocaldarius (an archaic)
?
?
attachment to sulfur particles
The Cell Envelope: capsules, cell walls and cell membranesThe cell envelope is a descriptive term for the several layers of material that envelope or enclose the protoplasm of the cell. The cell protoplasm (cytoplasm) is surrounded by the plasma membrane, a cell wall and a capsule. The cell wall itself is a layered structure in Gram-negative bacteria. All cells have a membrane, which is the essential and definitive characteristic of a "cell". Almost all procaryotes have a cell wall to prevent damage to the underlying protoplast. Outside the cell wall, foremost as a surface structure, may be a polysaccharide capsule or glycocalyx.
Figure 9. Profiles of the cell envelope the Gram-positive and Gram-negative bacteria. The Gram-positive wall is a uniformly thick layer external to the plasma membrane. It is composed mainly of peptidoglycan (murein). The Gram-negative wall appears thin and multilayered. It consists of a relatively thin peptidoglycan sheet between the plasma membrane and a phospholipid-lipopolysaccharide outer membrane. The space between the inner (plasma) and outer membranes (wherein the peptidoglycan resides) is called the periplasm.
Capsules
Most procaryotes contain some sort of a polysaccharide layer outside of the cell wall polymer. In a general sense, this layer is called a capsule. A true capsule is a discrete detectable layer of polysaccharides deposited outside the cell wall. A less discrete structure or matrix which embeds the cells is a called a slime layer or a biofilm. A type of capsule found in bacteria called a glycocalyx is a thin layer of tangled polysaccharide fibers which occurs on surface of cells growing in nature (as opposed to the laboratory). Some microbiologists refer to all capsules as glycocalyx and do not differentiate microcapsules.
Figure 10. Bacterial capsules outlined by India ink viewed by light microscopy. This is a true capsule, a discrete layer of polysaccharide surrounding the cells. Sometimes bacterial cells are embedded more randomly in a polysaccharide matrix called a slime layer or biofilm. Polysaccharide films that may inevitably be present on the surfaces of bacterial cells, but which cannot be detected visually, are called glycocalyx.
Figure 11. Negative stain of Streptococcus pyogenes viewed by transmission electron microscopy (28,000X). The halo around the chain of cells is the hyaluronic acid capsule that surrounds the exterior of the bacteria. The septa between dividing pairs of cells may also be seen. Electron micrograph of Streptococcus pyogenes by Maria Fazio and Vincent A. Fischetti, Ph.D. with permission. The Laboratory of Bacterial Pathogenesis and Immunology, Rockefeller University.
Capsules are generally composed of polysaccharide; rarely they contain amino sugars or peptides (Table 4).
Table 4. Chemical composition of some bacterial capsules
Bacterium
Capsule composition
Structural subunits
Gram-positive Bacteria

Bacillus anthracis
polypeptide (polyglutamic acid)
D-glutamic acid
Bacillus megaterium
polypeptide and polysaccharide
D-glutamic acid, amino sugars, sugars
Streptococcus mutans
polysaccharide
(dextran) glucose
Streptococcus pneumoniae
polysaccharides
sugars, amino sugars, uronic acids
Streptococcus pyogenes
polysaccharide (hyaluronic acid)
N-acetyl-glucosamine and glucuronic acid
Gram-negative Bacteria

Acetobacter xylinum
polysaccharide
(cellulose) glucose
Escherichia coli
polysaccharide (colonic acid)
glucose, galactose, fucose glucuronic acid
Pseudomonas aeruginosa
polysaccharide
mannuronic acid
Azotobacter vinelandii
polysaccharide
glucuronic acid
Agrobacterium tumefaciens
polysaccharide
(glucan) glucose
Capsules have several functions and often have multiple functions in a particular organism. Like fimbriae, capsules, slime layers, and glycocalyx often mediate adherence of cells to surfaces. Capsules also protect bacterial cells from engulfment by predatory protozoa or white blood cells (phagocytes), or from attack by antimicrobial agents of plant or animal origin. Capsules in certain soil bacteria protect cells from perennial effects of drying or desiccation. Capsular materials (e.g. dextrans) may be overproduced when bacteria are fed sugars to become reserves of carbohydrate for subsequent metabolism.
Figure 12. Colonies of Bacillus anthracis. The slimy or mucoid appearance of a bacterial colony is usually evidence of capsule production. In the case of B. anthracis, the capsule is composed of poly-D-glutamate. The capsule is an essential determinant of virulence to the bacterium. In the early stages of colonization and infection the capsule protects the bacteria from assaults by the immune and phagocytic systems.
Some bacteria produce slime materials to adhere and float themselves as colonial masses in their environments. Other bacteria produce slime materials to attach themselves to a surface or substrate. Bacteria may attach to surface, produce slime, divide and produce microcolonies within the slime layer, and construct a biofilm, which becomes an enriched and protected environment for themselves and other bacteria.
A classic example of biofilm construction in nature is the formation of dental plaque mediated by the oral bacterium, Streptococcus mutans. The bacteria adhere specifically to the pellicle of the tooth by means of a protein on the cell surface. The bacteria grow and synthesize a dextran capsule which binds them to the enamel and forms a biofilm some 300-500 cells in thickness. The bacteria are able to cleave sucrose (provided by the animal diet) into glucose plus fructose. The fructose is fermented as an energy source for bacterial growth. The glucose is polymerized into an extracellular dextran polymer that cements the bacteria to tooth enamel and becomes the matrix of dental plaque. The dextran slime can be depolymerized to glucose for use as a carbon source, resulting in production of lactic acid within the biofilm (plaque) that decalcifies the enamel and leads to dental caries or bacterial infection of the tooth.
Figure 13. (Left) Dental plaque revealed by a harmless red dye. http://www.medicdirect.co.uk/DentalHealth (Right) Human dental plaque. Transmission electron micrograph by Marilee Sellers, Northern Arizona University. http://www4.nau.edu/electron/TEM_img.htmAnother important characteristic of capsules may be their ability to block some step in the phagocytic process and thereby prevent bacterial cells from being engulfed or destroyed by phagocytes. For example, the primary determinant of virulence of the pathogen Streptococcus pneumoniae is its polysaccharide capsule, which prevents ingestion of pneumococci by alveolar macrophages. Bacillus anthracis survives phagocytic engulfment because the lysosomal enzymes of the phagocyte cannot initiate an attack on the poly-D-glutamate capsule of the bacterium. Bacteria such as Pseudomonas aeruginosa, that construct a biofilm made of extracellular slime when colonizing tissues, are also resistant to phagocytes, which cannot penetrate the biofilm.Cell Wall The cell walls of bacteria deserve special attention for several reasons: 1. They are an essential structure for viability, as described above. 2. They are composed of unique components found nowhere else in nature. 3. They are one of the most important sites for attack by antibiotics. 4. They provide ligands for adherence and receptor sites for drugs or viruses. 5. They cause symptoms of disease in animals. 6. They provide for immunological distinction and immunological variation among strains of bacteria.
Most procaryotes have a rigid cell wall. The cell wall is an essential structure that protects the cell protoplast from mechanical damage and from osmotic rupture or lysis. Procaryotes usually live in relatively dilute environments such that the accumulation of solutes inside the procaryotic cell cytoplasm greatly exceeds the total solute concentration in the outside environment. Thus, the osmotic pressure against the inside of the plasma membrane may be the equivalent of 10-25 atm. Since the membrane is a delicate, plastic structure, it must be restrained by an outside wall made of porous, rigid material that has high tensile strength. Such a material is murein, the ubiquitous component of bacterial cell walls.
Murein is a unique type of peptidoglycan, a polymer of disaccharides (glycan) cross-linked by short chains of amino acids (peptide). Many types of peptidoglycan exist. All Bacterial peptidoglycans contain N-acetylmuramic acid, which is the definitive component of murein. The cell walls of Archaea may be composed of protein, polysaccharides, or peptidoglycan-like molecules, but never do they contain murein. This feature distinguishes the Bacteria from the Archaea.
In the Gram-positive Bacteria (those that retain the purple crystal violet dye when subjected to the Gram-staining procedure), the cell wall consists of several layers of peptidoglycan. Running perpendicular to the peptidoglycan sheets is a group of molecules called teichoic acids which are unique to the Gram-positive cell wall (Figure 14). Figure 14. Structure of the Gram-positive bacterial cell wall. The wall is relatively thick and consists of many layers of peptidoglycan interspersed with teichoic acids that run perpendicular to the peptidoglycan sheets.In the Gram-negative Bacteria (which do not retain the crystal violet), the cell wall is composed of a single layer of peptidoglycan surrounded by a membranous structure called the outer membrane. The outer membrane of Gram-negative bacteria invariably contains a unique component, lipopolysaccharide (LPS or endotoxin), which is toxic to animals. In Gram-negative bacteria the outer membrane is usually thought of as part of the cell wall (Figure 15).
Figure 15. Structure of the Gram-negative cell wall. The wall is relatively thin and contains much less peptidoglycan than the Gram-positive wall. Also, teichoic acids are absent. However, the Gram negative cell wall consists of an outer membrane that is outside of the peptidoglycan layer. The outer membrane is attached to the peptidoglycan sheet by a unique group of lipoprotein molecules.In the Gram-positive Bacteria, the cell wall is thick (15-80 nanometers), consisting of several layers of peptidoglycan. In the Gram-negative Bacteria the cell wall is relatively thin (10 nanometers) and is composed of a single layer of peptidoglycan surrounded by an outer membrane. Peptidoglycan structure and arrangement in E. coli is representative of all Enterobacteriaceae, as well as many other Gram-negative bacteria. The glycan backbone is made up of alternating molecules of N-acetylglucosamine (G) and N-acetylmuramic acid (M) connected by a beta 1,4-glycoside bond. The 3-carbon of N-acetylmuramic acid (M) is substituted with a lactyl ether group derived from pyruvate. The lactyl ether connects the glycan backbone to a peptide side chain that contains L-alanine, (L-ala), D-glutamate (D-glu), Diaminopimelic acid (DAP), and D-alanine (D-ala). MurNAc is unique to bacterial cell walls, as is D-glu, DAP and D-ala. The muramic acid subunit of E. coli is shown in Figure 16 below.
Figure 16. The structure of the muramic acid subunit of the peptidoglycan of Escherichia coli. This is the type of murein found in most Gram-negative bacteria. The glycan backbone is a repeat polymer of two amino sugars, N-acetylglucosamine (G) and N-acetylmuramic acid (M). Attached to the N-acetylmuramic acid is a tetrapeptide consisting of L-ala-D-glu-DAP-D-ala. b. Abbreviated structure of the muramic acid subunit. c. Nearby tetrapeptide side chains may be linked to one another by an interpeptide bond between DAP on one chain and D-ala on the other. d. The polymeric form of the molecule.
Strands of murein are assembled in the periplasm from about 10 muramic acid subunits. Then the strands are connected to form a continuous glycan molecule that encompasses the cell. Wherever their proximity allows it, the tetrapeptide chains that project from the glycan backbone can be cross-linked by an interpeptide bond between a free amino group on DAP and a free carboxy group on a nearby D-ala. The assembly of peptidoglycan on the outside of the plasma membrane is mediated by a group of periplasmic enzymes, which are transglycosylases, transpeptidases and carboxypeptidases. The mechanism of action of penicillin and related beta-lactam antibiotics is to block transpeptidase and carboxypeptidase enzymes during their assembly of the murein cell wall. Hence, the beta lactam antibiotics are said to "block cell wall synthesis" in the bacteria.
The glycan backbone of the peptidoglycan molecule can be cleaved by an enzyme called lysozyme that is present in animal serum, tissues and secretions, and in the phagocytic lysosome. The function of lysozyme is to lyse bacterial cells as a constitutive defense against bacterial pathogens. Some Gram-positive bacteria are very sensitive to lysozyme and the enzyme is quite active at low concentrations. Lachrymal secretions (tears) can be diluted 1:40,000 and retain the ability to lyse certain bacterial cells. Gram-negative bacteria are less vulnerable to attack by lysozyme because their peptidoglycan is shielded by the outer membrane. The exact site of lysozymal cleavage is the beta 1,4 bond between N-acetylmuramic acid (M) and N-acetylglucosamine (G) , such that the muramic acid subunit shown in Figure 16(a) is the result of the action of lysozyme on bacterial peptidoglycan.
In Gram-positive bacteria there are numerous different peptide arrangements among peptidoglycans. The best studied is the murein of Staphylococcus aureus shown in Figure 17 below. In place of DAP (in E. coli) is the diamino acid, L-lysine (L-lys), and in place of the interpeptide bond (in Gram-negatives) is an interpeptide bridge of amino acids that connects a free amino group on lysine to a free carboxy group on D-ala of a nearby tetrapeptide side chain. This arrangement apparently allows for more frequent cross-bonding between nearby tetrapeptide side chains. In S. aureus, the interpeptide bridge is a peptide consisting of 5 glycine molecules (called a pentaglycine bridge). Assembly of the interpeptide bridge in Gram-positive murein is inhibited by the beta lactam antibiotics in the same manner as the interpeptide bond in Gram-negative murein. Gram-positive bacteria are more sensitive to penicillin than Gram-negative bacteria because the peptidoglycan is not protected by an outer membrane and it is a more abundant molecule. In Gram-positive bacteria, peptidoglycans may vary in the amino acid in place of DAP or L-lys in position 3 of the tetrapeptide, and in the exact composition of the interpeptide bridge. At least eight different types of peptidoglycan exist in Gram-positive bacteria.
Figure 17. Schematic diagram of the peptidoglycan sheet of Staphylococcus aureus. G = N-acetyl-glucosamine; M = N-acetyl-muramic acid; L-ala = L-alanine; D-ala = D-alanine; D-glu = D-glutamic acid; L-lys = L-lysine. This is one type of murein found in Gram-positive bacteria. Compared to the E. coli peptidoglycan (Figure 7) there is L-lys in place of DAP (diaminopimelic acid) in the tetrapeptide. The free amino group of L-lys is substituted with a glycine pentapeptide (gly-gly-gly-gly-gly-) which then becomes an interpeptide bridge forming a link with a carboxy group from D-ala in an adjacent tetrapeptide side chain. Gram-positive peptidoglycans differ from species to species, mainly in regards to the amino acids in the third position of the tetrapeptide side chain and in the amino acid composition of the interpeptide bridge.
Gram-negative bacteria may contain a single monomolecular layer of murein in their cell walls while Gram-positive bacteria are thought to have several layers or "wraps" of peptidoglycan. Closely associated with the layers of peptidoglycan in Gram-positive bacteria are a group of molecules called teichoic acids. Teichoic acids are linear polymers of polyglycerol or polyribitol substituted with phosphates and a few amino acids and sugars. The teichoic acid polymers are occasionally anchored to the plasma membrane (called lipoteichoic acid, LTA) apparently directed outward at right angles to the layers of peptidoglycan. The functions of teichoic acid are not known. They are essential to viability of Gram-positive bacteria in the wild. One idea is that they provide a channel of regularly-oriented negative charges for threading positively charged substances through the complicated peptidoglycan network. Another theory is that teichoic acids are in some way involved in the regulation and assembly of muramic acid subunits on the outside of the plasma membrane. There are instances, particularly in the streptococci, wherein teichoic acids have been implicated in the adherence of the bacteria to tissue surfaces.
The Outer Membrane of Gram-negative Bacteria
Of special interest as a component of the Gram-negative cell wall is the outer membrane, a discrete bilayered structure on the outside of the peptidoglycan sheet (see Figure 18 below). For the bacterium, the outer membrane is first and foremost a permeability barrier, but primarily due to its lipopolysaccharide content, it possesses many interesting and important characteristics of Gram-negative bacteria. The outer membrane is a lipid bilayer intercalated with proteins, superficially resembling the plasma membrane. The inner face of the outer membrane is composed of phospholipids similar to the phosphoglycerides that compose the plasma membrane. The outer face of the outer membrane may contain some phospholipid, but mainly it is formed by a different type of amphiphilic molecule which is composed of lipopolysaccharide (LPS). Outer membrane proteins usually traverse the membrane and in one case, anchor the outer membrane to the underlying peptidoglycan sheet.
Figure 18. Schematic illustration of the outer membrane, cell wall and plasma membrane of a Gram-negative bacterium. Note the structure and arrangement of molecules that constitute the outer membrane.
The LPS molecule that constitutes the outer face of the outer membrane is composed of a hydrophobic region, called Lipid A, that is attached to a hydrophilic linear polysaccharide region, consisting of the core polysaccharide and the O-specific polysaccharide.
Figure 19. Structure of LPS
The Lipid A head of the molecule inserts into the interior of the membrane, and the polysaccharide tail of the molecule faces the aqueous environment. Where the tail of the molecule inserts into the head there is an accumulation of negative charges such that a magnesium cation is chelated between adjacent LPS molecules. This provides the lateral stability for the outer membrane, and explains why treatment of Gram-negative bacteria with a powerful chelating agent, such as EDTA, causes dispersion of LPS molecules.
Bacterial lipopolysaccharides are toxic to animals. When injected in small amounts LPS or endotoxin activates macrophages to produce pyrogens, activates the complement cascade causing inflammation, and activates blood factors resulting in intravascular coagulation and hemorrhage. Endotoxins may play a role in infection by any Gram-negative bacterium. The toxic component of endotoxin (LPS) is Lipid A. The O-specific polysaccharide may provide ligands for bacterial attachment and confer some resistance to phagocytosis. Variation in the exact sugar content of the O polysaccharide (also referred to as the O antigen) accounts for multiple antigenic types (serotypes) among Gram-negative bacterial pathogens. Therefore. even though Lipid A is the toxic component in LPS, the polysaccharides nonetheless contribute to virulence of Gram-negative bacteria.
The proteins in the outer membrane of Escherichia coli are well characterized (see Table 5). About 400,00 copies of the Braun lipoprotein are covalently attached to the peptidoglycan sheet at one end and inserted into the hydrophobic interior of the membrane at the opposite end. A group of trimeric proteins called porins form pores of a fixed diameter through the lipid bilayer of the membrane. The omp C and omp F porins of E. coli are designed to allow passage of hydrophilic molecules up to mw of about 750 daltons. Larger molecules or harmful hydrophobic compounds (such as bile salts in the intestinal tract) are excluded from entry. Porins are designed in Gram-negative bacteria to allow passage of useful molecules (nutrients) through the barrier of the outer membrane, but to exclude passage harmful substances from the environment. The ubiquitous omp A protein in the outer membrane of E. coli has a porin like structure, and may function in uptake of specific ions, but it is also a receptor for the F pilus and an attachment site for bacterial viruses.
Table 5. Functions of the outer membrane components of Escherichia coli.
Component
Function
Lipopolysaccharide (LPS)
Permeability barrier
Mg++ bridges
Stabilizes LPS and is essential for its permeability characteristics
Braun lipoprotein
Anchors the outer membrane to peptidoglycan (murein) sheet
Omp C and Omp F porins
proteins that form pores or channels through outer membrane for passage of hydrophilic molecules
Omp A protein
provides receptor for some viruses and bacteriocins; stabilizes mating cells during conjugation
S-layers
S-layer proteins form the outermost cell envelope component of a broad spectrum of bacteria and archaea. S-layers are composed of a single protein or glycoprotein species (Mw 40-200 kDa) and exhibit either oblique, square or hexagonal lattice symmetry with unit cell dimensions in the range of 3 to 30 nm. S-layers are generally 5 to 10 nm thick and show pores of identical size (diameter, 2 - 8 nm) and morphology.Crystalline bacterial cell surface layer (S-layer) proteins have been optimized during billions of years of biological evolution as constituent elements of one of the simplest self-assembly systems in nature. Isolated S-layer proteins have the intrinsic property to recrystallize into two-dimensional arrays on a broad spectrum of surfaces including silicon, metals and polymers, and to interfaces such as planar lipid films and liposomes. The well defined arrangement of functional groups on S-layer lattices allows the binding of molecules and particles in defined regular arrays. S-layers also represent templates for the formation of inorganic nanocrystal superlattices composed of CdS, Au, Ni, Pt, or Pd.The self-assembly of S-layers illustrates a basic building principle in nature for generating large arrays of biomolecules with well-defined geometrical and physicochemical surface properties.
Many Gram-negative and Gram-positive bacteria, as well a many archaea possess a regularly structured layer called an S-layer attached to the outermost portion of their cell wall. It is composed of protein or glycoprotein and in electron micrographs, has a pattern resembling a tiled surface. Transmission electron micrograph of a freeze-etched, metal shadowed preparation of a bacterial cell with an S-layer with hexagonal lattice symmetry. Bar = 100nm.http://www.foresight.org/conference/MNT7/Papers/Pum/index.htmlS-layers have been associated with a number of possible functions. The S-layer may protect bacteria from harmful enzymes or changes in pH. It may contribute to virulence by protecting the bacterium against complement attack and phagocytosis. It is thought to protect E. coli from attack by the predatory bacterium, Bdellovibrio.
The S-layer can function as an adhesin, enabling the bacterium to adhere to host cell membranes and environmental surfaces in order to colonize. Many of the cell-associated protein adhesins used by pathogens are components of the S-layer.
A correlation between Gram stain reaction and cell wall properties of bacteria is summarized in Table 6. The Gram stain procedure contains a "destaining" step wherein the cells are washed with an acetone-alcohol mixture. The lipid content of the Gram-negative wall probably affects the outcome of this step so that Gram-positive cells retain a primary stain while Gram-negative cells are destained.
Table 6. Correlation of Grams stain with other properties of Bacteria.
Property
Gram-positive
Gram-negative
Thickness of wall
thick (20-80 nm)
thin (10 nm)
Number of layers
1
2
Peptidoglycan (murein) content
>50%
10-20%
Teichoic acids in wall
present
absent
Lipid and lipoprotein content
0-3%
58%
Protein content
0
9%
Lipopolysaccharide content
0
13%
Sensitivity to Penicillin G
yes
no (1)
Sensitivity to lysozyme
yes
no (2)
(1) A few Gram-negative bacteria are sensitive to natural penicillins. Many Gram-negative bacteria are sensitive to some type of penicillin, especially semisynthetic penicillins. Gram-negative bacteria, including E. coli, can be made sensitive to natural penicillin by procedures that disrupt the permeability characteristics of the outer membrane. (2) Gram-negative bacteria are sensitive to lysozyme if pretreated by some procedure that removes the outer membrane and exposes the peptidoglycan directly to the enzyme.
Cell Wall-less Forms A few bacteria are able to live or exist without a cell wall. The mycoplasmas are a group of bacteria that lack a cell wall. Mycoplasmas have sterol-like molecules incorporated into their membranes and they are usually inhabitants of osmotically-protected environments. Mycoplasma pneumoniae is the cause of primary atypical bacterial pneumonia, known in the vernacular as "walking pneumonia". For obvious reasons, penicillin is ineffective in treatment of this type of pneumonia. Sometimes, under the pressure of antibiotic therapy, pathogenic bacteria can revert to cell wall-less forms (called spheroplasts or protoplasts) and persist or survive in osmotically-protected tissues. When the antibiotic is withdrawn from therapy the organisms may regrow their cell walls and reinfect unprotected tissues.
The Plasma Membrane
The plasma membrane, also called the cytoplasmic membrane, is the most dynamic structure of a procaryotic cell. Its main function is a s a selective permeability barrier that regulates the passage of substances into and out of the cell. The plasma membrane is the definitive structure of a cell since it sequesters the molecules of life in a unit, separating it from the environment. The bacterial membrane allows passage of water and uncharged molecules up to mw of about 100 daltons, but does not allow passage of larger molecules or any charged substances except by means special membrane transport processes and transport systems. Bacterial membranes are composed of 40 percent phospholipid and 60 percent protein. The phospholipids are amphoteric molecules with a polar hydrophilic glycerol "head" attached via an ester bond to two nonpolar hydrophobic fatty acid tails, which naturally form a bilayer in aqueous environments. Dispersed within the bilayer are various structural and enzymatic proteins which carry out most membrane functions. At one time, it was thought that the proteins were neatly organized along the inner and outer faces of the membrane and that this accounted for the double track appearance of the membrane in electron micrographs. However, it is now known that while some membrane proteins are located and function on one side or another of the membrane, most proteins are partly inserted into the membrane, or possibly even traverse the membrane as channels from the outside to the inside. It is possible that proteins can move laterally along a surface of the membrane, but it is thermodynamically unlikely that proteins can be rotated within a membrane, which discounts early theories of how transport systems might work. The arrangement of proteins and lipids to form a membrane is called the fluid mosaic model, and is illustrated in Figure 20.
Figure 20. Fluid mosaic model of a biological membrane. In aqueous environments membrane phospholipids arrange themselves in such a way that they spontaneously form a fluid bilayer. Membrane proteins, which may be structural or functional, may be permanently or transiently associated with one side or the other of the membrane, or even be permanently built into the bilayer, while other proteins span the bilayer and may form transport channels through the membrane.
The membranes of Bacteria are structurally similar to the cell membranes of eucaryotes, except that bacterial membranes consist of saturated or monounsaturated fatty acids (rarely, polyunsaturated fatty acids) and do not normally contain sterols. The membranes of Archaea form bilayers functionally equivalent to bacterial membranes, but archaeal lipids are saturated, branched, repeating isoprenoid subunits that attach to glycerol via an ether linkage as opposed to the ester linkage found in glycerides of eukaryotic and bacterial membrane lipids (Figure 21). The structure of archaeal membranes is thought to be an adaptation to their existence and survival in extreme environments. Figure 21. Generalized structure of a membrane lipids. (top). A phospholipid in the membrane of the bacterium Escherichia coli. The R1 and R2 positions on glycerol are substituted with saturated or monounsaturated fatty acids, with ester linkages to the glyceride. The R3 position is substituted with phosphatidylethanolamine, the most common substituent in this position in Bacteria. (bottom). An Archaeal membrane lipid. In contrast to bacterial phospholipids, which are glycerol esters of fatty acids, the lipids in membranes of Archaea are diethers of glycerol and long-chain, branched, saturated hydrocarbons called isoprenoids or which are made up of repeating C5 subunits. One of the major isoprenoids is the C20 molecule phytanol. The R3 position of glycerol may or may not be substituted. The structure of archaeal membrane lipids is thought to be an adaptation to extreme environments such as hot and acidic conditions where Archaea prevail in nature.
Functions of the Cytoplasmic Membrane
Since procaryotes lack any intracellular organelles for processes such as respiration or photosynthesis or secretion, the plasma membrane subsumes these processes for the cell and consequently has a variety of functions in energy generation, and biosynthesis. For example, the electron transport system that couples aerobic respiration and ATP synthesis is found in the procaryotic membrane. The photosynthetic chromophores that harvest light energy for conversion into chemical energy are located in the membrane. Hence, the plasma membrane is the site of oxidative phosphorylation and photophosphorylation in procaryotes, analogous to the functions of mitochondria and chloroplasts in eukaryotic cells. Besides transport proteins that selectively mediate the passage of substances into and out of the cell, procaryotic membranes may contain sensing proteins that measure concentrations of molecules in the environment or binding proteins that translocate signals to genetic and metabolic machinery in the cytoplasm. Membranes also contain enzymes involved in many metabolic processes such as cell wall synthesis, septum formation, membrane synthesis, DNA replication, CO2 fixation and ammonia oxidation. The predominant functions of procaryotic membranes are listed in Table 7 and discussed below.
Table 7. Functions of the procaryotic plasma membrane
1. Osmotic or permeability barrier
2. Location of transport systems for specific solutes (nutrients and ions)
3. Energy generating functions, involving respiratory and photosynthetic electron transport systems, establishment of proton motive force, and transmembranous, ATP-synthesizing ATPase
4. Synthesis of membrane lipids (including lipopolysaccharide in Gram-negative cells)
5. Synthesis of murein (cell wall peptidoglycan)
6. Assembly and secretion of extracytoplasmic proteins
7. Coordination of DNA replication and segregation with septum formation and cell division
8. Chemotaxis (both motility per se and sensing functions)
9. Location of specialized enzyme system
Permeability Barrier
The cell membrane is the most dynamic structure in the cell. Its main function is as a permeability barrier that regulates the passage of substances into and out of the cell. The plasma membrane is the definitive structure of a cell since it sequesters the molecules of life in the cytoplasm, separating it from the outside environment. The bacterial membrane freely allows passage of water and a few small uncharged molecules (less than molecular weight of 100 daltons), but it does not allow passage of larger molecules or any charged substances except when monitored by proteins in the membrane called transport systems.
Transport of Solutes
The proteins that mediate the passage of solutes through membranes are referred to variously as transport systems, carrier proteins, porters, and permeases. Transport systems operate by one of three transport processes as described below in Figure 22. In a uniport process, a solute passes through the membrane unidirectionally. In symport processes (also called cotransport) two solutes must be transported in the same direction at the same time; in antiport processes ( also called exchange diffusion), one solute is transported in one direction simultaneously as a second solute is transported in the opposite direction.
Types of Transport Systems
Bacteria have a variety of types of transport systems which can be used alternatively in various environmental situations. The elaborate development of transport processes and transport systems in procaryotes probably reflects their need to concentrate substances inside the cytoplasm against the concentration (gradient) of the environment. Concentration of solutes in the cytoplasm requires the operation of an active transport system, of which there are two types in bacteria: ion driven transport systems (IDT) and binding-protein dependent transport systems (BPDT). The definitive feature of an active transport system is the accumulation of the solute in the cytoplasm at concentrations far in excess of the environment. According to the laws of physical chemistry, this type of process requires energy. There are four types of carrier-mediated transport systems in procaryotes. The carrier is a protein (or group of proteins) that functions in the passage of a small molecule from one side of a membrane to the other side. A transport system may be a single transmembranous protein that forms a channel that admits passage of a specific solute, or it may be a coordinated system of proteins that binds and sequentially passes a small molecule through the membrane. Transport systems have the property of specificity for the solute transported. Some transport systems transport a single solute with the same specificity and kinetics as an enzyme. Some transport systems will transport (structurally) related molecules, although at reduced efficiency compared to their primary substrate. Most transport systems transport specific sugars, amino acids, anions or cations that are of nutritional value to the bacterium.
Facilitated diffusion systems (FD) are the least common type of transport system in bacteria. Actually, the glycerol uniporter in E. coli is the only well known facilitated diffusion system. FD involves the passage of a specific solute through a carrier that forms a channel in the membrane. The solute can move in either direction through the membrane to the point of of equilibrium on both sides of the membrane. Although the system is carrier-mediated and specific, no energy is expended in the transport process. For this reason the glycerol molecule cannot be accumulated against the concentration gradient.
Ion driven transport systems (IDT) and Binding-protein dependent transport systems (BPDT) are active transport systems that are used for transport of most solutes by bacterial cells. IDT is used for accumulation of many ions and amino acids; BPDT is frequently used for sugars and amino acids. IDT is a symport or antiport process that uses a hydrogen ion (H+) i.e., proton motive force (pmf), or some other cation, i.e., chemiosmotic potential, to drive the transport process. IDT systems such as the lactose permease of E. coli utilize the consumption of a hydrogen ion during the transport of lactose. Thus the energy expended during active transport of lactose is in the form of pmf. The lactose permease is a single transmembranous polypeptide that spans the membrane seven times forming a channel that specifically admits lactose.
Binding-protein dependent transport systems (BPDT), such as the histadine transport system in E. coli, are composed of four proteins. Two proteins form a membrane channel that allows passage of the histadine. A third protein resides in the periplasmic space where it is able to bind the amino acid and pass it to a forth protein which admits the amino acid into the membrane channel. Driving the solute through the channel involves the expenditure of energy, which is provided by the hydrolysis of ATP.
Group translocation systems (GT), more commonly known as the phosphotransferase system (PTS) in E. coli, are used primarily for the transport of sugars. Like binding protein-dependent transport systems, they are composed of several distinct components. However, GT systems specific for one sugar may share some of their components with other group transport systems. In E. coli, glucose may be transported by a group translocation process that involves the phosphotransferase system. The actual carrier in the membrane is a protein channel fairly specific for glucose. Glucose specifically enters the channel from the outside, but in order to exit into the cytoplasm, it must first be phosphorylated by the phosphotransferase system. The PTS derives energy from the metabolic intermediate phosphoenol pyruvate (PEP). PEP is hydrolyzed to pyruvate and glucose is phosphorylated to form glucose-phosphate during the process. Thus, by the expenditure of a single molecule of high energy phosphate, glucose is transported and changed to glucose-phosphate.
Table 8. Distinguishing characteristics of bacterial transport systemsPD = passive diffusion FD = facilitated diffusion IDT = ion-driven transport BPDT = binding protein dependent transport GT = group translocation
Property
PD
FD
IDT
BPDT
GT
carrier mediated
-
+
+
+
+
conc. against gradient
-
-
+
+
NA
specificity
-
+
+
+
+
energy expended
-
-
pmf
ATP
PEP
solute modified during transport
-
-
-
-
+
Generation of EnergyUnlike eucaryotes, bacteria don't have intracellular organelles for energy producing processes such as respiration or photosynthesis. Instead, the cytoplasmic membrane carries out these functions. The membrane is the location of electron transport systems (ETS) used to produce energy during photosynthesis and respiration, and it is the location of an enzyme called ATP synthetase (ATPase) which is used to synthesize ATP.When the electron transport system operates, it establishes a pH gradient across of the membrane due to an accumulation of protons (H+) outside and hydroxyl ion (OH-) inside. Thus the outside is acidic and the inside is alkaline. Operation of the ETS also establishes a charge on the membrane called proton motive force (pmf). The outer face of the membrane becomes charged positive while inner face is charged negative, so the membrane has a positive side and a negative side, like a battery. The pmf can be used to do various types of work including the rotation of the flagellum, or active transport as described above. The pmf can also be used to make ATP by the membrane ATPase enzyme which consumes protons when it synthesizes ATP from ADP and phosphate. The connection between electron transport, establishment of pmf, and ATP synthesis during respiration is known as oxidative phosphorylation; during photosynthesis, it is called photophorylation.Figure 24 below illustrates the membrane of E. coli. The topographical features of the membrane from top to bottom are 1. lactose transport system; 2. the flagellar motor coupled to the hook and filament; 3. Na+ transport (export) system; 4. Ca++ transport (export) system; 5. electron transport system; 6. ATPase enzyme; 7. proline transport system. The operation ot the electron transport system during respiration produces the H+ charge on the membrane (pmf). The pmf ( H+) is used by the transport systems to move molecules from one side of the membrane to the other; by the flagellar motor ring to rotate the flagellar filament; and by the ATPase enzyme to synthesize ATP.
The plasma membrane of procaryotes may invaginate into the cytoplasm or form stacks or vesicles attached to the inner membrane surface. These structures are sometimes referred to as mesosomes. Such internal membrane systems may be analogous to the cristae of mitochondria or the thylakoids of chloroplasts which increase the surface area of membranes to which enzymes are bound for specific enzymatic functions. The photosynthetic apparatus (light harvesting pigments and ATPase) of photosynthetic procaryotes is contained in these types of membranous structures. Mesosomes may also represent specialized membrane regions involved in DNA replication and segregation, cell wall synthesis, or increased enzymatic activity. Membrane foldings and vesicles sometimes appear in electron micrographs of procaryotic cells as artifacts of preparative techniques. These membranous structures, of course, are not mesosomes, but their existence does not prove that mesosomes are not present in procaryotes, and there are several examples of procaryotic membrane topology and appearance that are suggestive of mesosomes.
There are a few antibiotics (e.g. polymyxin), hydrophobic agents (e.g. bile salts), and proteins (e.g. complement) that can damage bacterial membranes.
The PeriplasmBetween the inner (plasma) and outer membranes of Gram-negative bacteria and spirochetes is a space called the periplasm or periplasmic space (See Figures 9 and 18). Actually, the peptidoglycan sheet resides within the periplasm. The periplasm is a very active compartment of the cell, containing enzymes for assembly of cell wall and membrane components, various degradative or detoxifying enzymes, secretion systems, sensing proteins for chemotaxis and signal transduction, and binding proteins for solutes taken up by BPDT transport systems. Components of the periplasm are needed in this region of the cell and are bounded or "trapped" by the two membranes of the cell. In the case of spirochetes, their flagella (called endoflagella or periplasmic flagella) rotate within the periplasm and impart the flexing and screw-like rotation characteristic of spirochete motility.
Table 9. Representative periplasmic proteins in E. coli.
Binding proteins For amino acids (e.g. histadine, arginine) For sugars (e.g. glucose, maltose) For vitamins (e.g. thiamine, vitamin B12) For ions (e.g. phosphate, sulfate)
Biosynthetic enzymes For murein assembly (e.g. transglycosylases, carboxypeptidases, transpeptidases) For fimbrial subunit secretion and assembly (e.g. chaperonins)
Degradative enzymes phosphatases proteases
Detoxifying enzymes Beta-lactamases (e.g. penicillinase) Aminoglycoside-phosphorylating enzymes
The Cytoplasm
The cytoplasm of bacterial cells consists consists of an aqueous solution of three groups of molecules: macromolecules such as proteins (enzymes), mRNA and tRNA; small molecules that are energy sources, precursors of macromolecules, metabolites or vitamins; and various inorganic ions and cofactors (see Tables 9, 10, 11). The primary structural components found in the cytoplasm are the nucleoid and ribosomes, and possibly some type of inclusion. The cytoplasm of procaryotes is more gel-like than that of eucaryotes and the processes of cytoplasmic streaming, which are evident in eucaryotes, do not occur.
Table 9. Molecular composition of E. coli under conditions of balanced growth. Percentage of dry weight refers to all structural and cytoplasmic components.
Molecule
Percentage of dry weight
Protein
Total RNA
DNA
Phospholipid
Lipopolysaccharide
Murein
Glycogen
Small molecules: precursors, metabolites, vitamins, etc.
Inorganic ions
Total dry weight
55
20.5
3.1
9.1
3.4
2.5
2.5
2.9
1.0
100.0
Table 10. Small molecules present in the cytoplasm of a growing bacterial cell.
Molecule
Approximate number of kinds
Amino acids, their precursors and derivatives
Nucleotides, their precursors and derivatives
Fatty acids and their precursors
Sugars, carbohydrates and their precursors or derivatives
quinones, porphyrins, vitamins, coenzymes and prosthetic groups and their precursors
120
100
50
250
300
Table 11. Inorganic ions present in the cytoplasm of a growing bacterial cell.
Ion
Function
K+
Maintenance of ionic strength; cofactor for certain enzymes
NH4+
Principal form of inorganic N for assimilation
Ca++
Cofactor for certain enzymes
Fe++
Present in cytochromes and other metalloenzymes
Mg++
Cofactor for many enzymes; stabilization of outer membrane of Gram-negative bacteria
Mn++
Present in certain metalloenzymes
Co++
Trace element constituent of vitamin B12 and its coenzyme derivatives and found in certain metalloenzymes
Cu++
Trace element present in certain metalloenzymes
Mo++
Trace element present in certain metalloenzymes
Ni++
Trace element present in certain metalloenzymes
Zn++
Trace element present in certain metalloenzymes
SO4--
Principal form of inorganic S for assimilation
PO4---
Principal form of P for assimilation and a participant in many metabolic reactions
The bacterial chromosome (nucleoid) is typically one large circular molecule of DNA, more or less free in the cytoplasm, although coiled and supercoiled and anchored by proteins. Procaryotes sometimes possess smaller extrachromosomal pieces of DNA called plasmids. The total DNA content of a procaryote is referred to as the cell genome. The cell chromosome is the genetic control center of the cell which determines all the properties and functions of the bacterium. During cell growth and division, the procaryotic chromosome is replicated in a semiconservative fashion to make an exact copy of the molecule for distribution to progeny cells. However, the eucaryotic processes of meiosis and mitosis are absent in procaryotes. Replication and segregation of procaryotic DNA is coordinated by the membrane and various proteins in the cytoplasm. Figure 25. When a bacterium such as E. coli is "gently lysed" the chromosomal DNA leaks out of the cell as a continuous molecule that is many times longer than the length of the cell.The distinct granular appearance of procaryotic cytoplasm is due to the presence and distribution of ribosomes. Ribosomes are composed of proteins and RNA. The ribosomes of procaryotes are smaller than cytoplasmic ribosomes of eucaryotes. Procaryotic ribosomes are 70S in size, being composed of 30S and 50S subunits. The 80S ribosomes of eucaryotes are made up of 40S and 60S subunits. Ribosomes are involved in the process of translation (protein synthesis), but some details of their activities differ in eucaryotes, bacteria and archaea. The 70S ribosomes that occur in eucaryotic mitochondria and chloroplasts contain ssrRNA closely related to bacterial ribosomal RNA. his is taken as a major line of evidence that these organelles are descended from procaryotes. Figure 26. The bacterial chromosome or nucleoid is the nonstaining region in the interior of the cell cytoplasm. The granular structures distributed throughout the cytoplasm are cell ribosomes.
Inclusions
Often contained in the cytoplasm of procaryotic cells is one or another of some type of inclusion granule. Inclusions are distinct granules that may occupy a substantial part of the cytoplasm. Inclusion granules are usually reserve materials of some sort. For example, carbon and energy reserves may be stored as glycogen (a polymer of glucose) or as polybetahydroxybutyric acid (a type of fat) granules. Polyphosphate inclusions are reserves of PO4 and possibly energy; elemental sulfur (sulfur globules) are stored by some phototrophic and some lithotrophic procaryotes as reserves of energy or electrons. Some inclusion bodies are actually membranous vesicles or intrusions into the cytoplasm which contain photosynthetic pigments or enzymes.
Table 12. Some inclusions in bacterial cells.
Cytoplasmic inclusions
Where found
Composition
Function
glycogen
many bacteria e.g. E. coli
polyglucose
reserve carbon and energy source
polybetahydroxybutyric acid (PHB)
many bacteria e.g. Pseudomonas
polymerized hydroxy butyrate
reserve carbon and energy source
polyphosphate (volutin granules)
many bacteria e.g. Corynebacterium
linear or cyclical polymers of PO4
reserve phosphate; possibly a reserve of high energy phosphate
sulfur globules
phototrophic purple and green sulfur bacteria and lithotrophic colorless sulfur bacteria
elemental sulfur
reserve of electrons (reducing source) in phototrophs; reserve energy source in lithotrophs
gas vesicles
aquatic bacteria especially cyanobacteria
protein hulls or shells inflated with gases
buoyancy (floatation) in the vertical water column
parasporal crystals
endospore-forming bacilli (genus Bacillus)
protein
unknown but toxic to certain insects
magnetosomes
certain aquatic bacteria
magnetite (iron oxide) Fe3O4
orienting and migrating along geo- magnetic field lines
carboxysomes
many autotrophic bacteria
enzymes for autotrophic CO2 fixation
site of CO2 fixation
phycobilisomes
cyanobacteria
phycobiliproteins
light-harvesting pigments
chlorosomes
Green bacteria
lipid and protein and bacteriochlorophyll
light-harvesting pigments and antennae
Figure 27. A variety of bacterial inclusions. a. PHB granules; b. a parasporal BT crystal in the sporangium of Bacillus thuringiensis; c. carboxysomes in Anabaena viriabilis, showing their polyhedral shape; d. sulfur globules in the cytoplasm of Beggiatoa
Endospores
A bacterial structure sometimes observed as an inclusion is actually a type of dormant cell called an endospore. Endospores are formed by a few groups of Bacteria as intracellular structures, but ultimately they are released as free endospores. Biologically, endospores are a fascinating type of cell. Endospores exhibit no signs of life, being described as cryptobiotic. They are highly resistant to environmental stresses such as high temperature (some endospores can be boiled for hours and retain their viability), irradiation, strong acids, disinfectants, etc. They are probably the most durable cell produced in nature. Although cryptobiotic, they retain viability indefinitely such that under appropriate environmental conditions, they germinate back into vegetative cells. Endospores are formed by vegetative cells in response to environmental signals that indicate a limiting factor for vegetative growth, such as exhaustion of an essential nutrient. They germinate and become vegetative cells when the environmental stress is relieved. Hence, endospore-formation is a mechanism of survival rather than a mechanism of reproduction.

Property
Vegetative cells
Endospores
Surface coats
Typical Gram-positive murein cell wall polymer
Thick spore coat, cortex, and peptidoglycan core wall
Microscopic appearance
Nonrefractile
Refractile
Calcium dipicolinic acid
Absent
Present in core
Cytoplasmic water activity
High
Very low
Enzymatic activity
Present
Absent
Macromolecular synthesis
Present
Absent
Heat resistance
Low
High
Resistance to chemicals and acids
Low
High
Radiation resistance
Low
High
Sensitivity to lysozyme
Sensitive
Resistant
Sensitivity to dyes and staining
Sensitive
Resistant

By: Mr. Umukoro David azubuike.Delta state university,abraka,Nigeria
Contact address: umukoro71223@gmail.com.

2007-12-10T01:18:00.000-08:00