Escherichia coli (Ecoli)


Escherichia coli is a bacterium that is commonly found in the lower intestine of warm-blooded animals. Most E. coli strains are harmless, but some, such as serotype O157:H7, can cause serious food poisoning in humans, and are occasionally responsible for costly product recalls. The harmless strains are part of the normal flora of the gut, and can benefit their hosts by producing vitamin K2, or by preventing the establishment of pathogenic bacteria within the intestine.

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E. coli are not always confined to the intestine, and their ability to survive for brief periods outside the body makes them an ideal indicator organism to test environmental samples for fecal contamination. The bacteria can also be grown easily and its genetics are comparatively simple and easily-manipulated, making it one of the best-studied prokaryotic model organisms, and an important species in biotechnology. E. coli was discovered by German pediatrician and bacteriologist Theodor Escherich in 1885, and is now classified as part of the Enterobacteriaceae family of gamma-proteobacteria.



Strains

A strain of E. coli is a sub-group within the species that has unique characteristics that distinguish it from other E. coli strains. These differences are often detectable only on the molecular level; however, they may result in changes to the physiology or lifecycle of the bacterium. For example, a strain may gain pathogenic capacity, the ability to use a unique carbon source, the ability to inhabit a particular ecological niche or the ability to resist antimicrobial agents. Different strains of E. coli are often host-specific, making it possible to determine the source of fecal contamination in environmental samples.Depending on which E. coli strains are present in a water sample, for example, assumptions can be made about whether the contamination originated from a human, other mammal or bird source.


New strains of E. coli evolve through the natural biological process of mutation, and some strains develop traits that can be harmful to a host animal. Although virulent strains typically cause no more than a bout of diarrhea in healthy adult humans, particularly virulent strains, such as O157:H7 or O111:B4, can cause serious illness or death in the elderly, the very young or the immunocompromised.

Biology and biochemistry

E. coli is Gram-negative, facultative anaerobic and non-sporulating. It can live on a wide variety of substrates. E. coli uses mixed-acid fermentation in anaerobic conditions, producing lactate, succinate, ethanol, acetate and carbon dioxide. Since many pathways in mixed-acid fermentation produce hydrogen gas, these pathways require the levels of hydrogen to be low; E. coli can only use these processes when hydrogen-consuming organisms such as methanogens or sulfate-reducing bacteria are present.

Optimal growth of E. coli occurs at 37°C, but some laboratory strains can multiply at temperatures of up to 49°C. Growth can be driven by aerobic or anaerobic respiration, using a large variety of redox pairs, including the oxidation of pyruvic acid, formic acid, hydrogen and amino acids, and the reduction of substrates such as oxygen, nitrate, dimethyl sulfoxide and trimethylamine N-oxide.

Strains that possess flagella can swim and are motile, but other strains lack flagella. The flagella of E. coli have a peritrichous arrangement.

E. coli and related bacteria possess the ability to transfer DNA via bacterial conjugation, transduction or transformation, which allows genetic material to spread horizontally through an existing population. It is believed that this process led to the spread of shiga toxin from Shigella to E. coli.

Normal Role
E. coli normally colonizes an infant's gastrointestinal tract within 40 hours of birth, arriving with food or water or with the individuals handling the child. In the bowel, it adheres to the mucus of the large intestine. Wild-type E. coli has no growth factor requirements; it can synthesize all the components of its cell from glucose. It is the primary facultative organism of the human gastrointestinal tract. As long as these bacteria do not acquire genetic elements encoding for virulence factors, they remain benign commensals.

Role in disease
Virulent strains of E. coli can cause gastroenteritis, urinary tract infections, and neonatal meningitis. In rarer cases, virulent strains are also responsible for peritonitis, mastitis, septicemia and Gram-negative pneumonia. Recently it is thought that E. coli and certain other foodborne illnesses can sometimes trigger serious health problems months or years after patients survived that initial bout.Food poisoning can be long-term problem.

Virulence properties
Enteric E. coli (EC) are classified on the basis of serological characteristics and virulence properties. Virotypes include:
  • Enterotoxigenic E. coli (ETEC) – causative agent of diarrhea (without fever) in humans, pigs, sheep, goats, cattle, dogs, and horses. ETEC uses fimbrial adhesins (projections from the bacterial cell surface) to bind enterocyte cells in the small intestine. ETEC can produce two proteinaceous enterotoxins: the larger of the two proteins, LT enterotoxin, is similar to cholera toxin in structure and function, while the smaller protein, ST enterotoxin causes cGMP accumulation in the target cells and a subsequent secretion of fluid and electrolytes into the intestinal lumen. ETEC strains are non-invasive, and they do not leave the intestinal lumen.
  • Enteropathogenic E. coli (EPEC) – causative agent of diarrhea in humans, rabbits, dogs, cats and horses. Like ETEC, EPEC also causes diarrhea, but the molecular mechanisms of colonization and etiology are different. EPEC lack fimbriae, ST and LT toxins, but they utilize an adhesin known as intimin to bind host intestinal cells. This virotype has an array of virulence factors that are similar to those found in Shigella, and may posses a shiga toxin. Adherence to the intestinal mucosa causes a rearrangement of actin in the host cell, causing significant deformation. EPEC cells are moderately-invasive (i.e. they enter host cells) and elicit an inflammatory response. Changes in intestinal cell ultrastructure due to "attachment and effacement" is likely the prime cause of diarrhea in those afflicted with EPEC.
  • Enteroinvasive E. coli (EIEC) – found only in humans. EIEC infection causes a syndrome that is identical to Shigellosis, with profuse diarrhea and high fever. EIEC are highly invasive, and they utilize adhesin proteins to bind to and enter intestinal cells. They produce no toxins, but severely damage the intestinal wall through mechanical cell destruction.
  • Enterohemorrhagic E. coli (EHEC) – found in humans, cattle, and goats. The sole member of this virotype is strain O157:H7, which causes bloody diarrhea and no fever. EHEC can cause hemolytic uremic syndrome and sudden kidney failure. It uses bacterial fimbriae for attachment, is moderately-invasive and possesses a phage-encoded Shiga toxin that can elicit an intense inflammatory response.
  • Enteroaggregative E. coli (EAggEC) – found only in humans. So named because they have fimbriae which aggregate tissue culture cells, EAggEC bind to the intestinal mucosa to cause watery diarrhea without fever. EAggEC are non-invasive. They produce a hemolysin and an ST enterotoxin similar to that of ETEC.

Gastrointestinal infection
Certain strains of E. coli, such as O157:H7, O121 and O104:H21, produce toxins. Food poisoning caused by E. coli are usually associated with eating unwashed vegetables and meat contaminated post-slaughter. O157:H7 is further notorious for causing serious and even life-threatening complications like Hemolytic Uremic Syndrome (HUS). This particular strain is linked to the 2006 United States E. coli outbreak of fresh spinach. Severity of the illness varies considerably; it can be fatal, particularly to young children, the elderly or the immunocompromised, but is more often mild. E. coli can harbor both heat-stable and heat-labile enterotoxins. The latter, termed LT, contains one 'A' subunit and five 'B' subunits arranged into one holotoxin, and is highly similar in structure and function to Cholera toxins. The B subunits assist in adherence and entry of the toxin into host intestinal cells, while the A subunit is cleaved and prevents cells from absorbing water, causing diarrhea. LT is secreted by the Type 2 secretion pathway.

If E. coli bacteria escape the intestinal tract through a perforation (for example from an ulcer, a ruptured appendix, or a surgical error) and enter the abdomen, they usually cause peritonitis that can be fatal without prompt treatment. However, E. coli are extremely sensitive to such antibiotics as streptomycin or gentamycin, so treatment with antibiotics is usually effective. This could change since, as noted below, E. coli quickly acquires drug resistance.

Intestinal mucosa-associated E. coli are observed in increased numbers in the inflammatory bowel diseases, Crohn's disease and ulcerative colitis.Invasive strains of E. coli exist in high numbers in the inflamed tissue, and the number of bacteria in the inflamed regions correlates to the severity of the bowel inflammation.

Epidemiology of gastrointestinal infection


Transmission of pathogenic E. coli often occurs via fecal-oral transmission.Common routes of transmission include: unhygienic food preparation, farm contamination due to manure fertilization, irrigation of crops with contaminated greywater or raw sewage, feral pigs on cropland, or direct consumption of sewage-contaminated water. Dairy and beef cattle are primary reservoirs of E. coli O157:H7, and they can carry it asymptomatically and shed it in their feces. Food products associated with E. coli outbreaks include raw ground beef, raw seed sprouts or spinach, raw milk, unpasteurized juice, and foods contaminated by infected food workers via fecal-oral route.

According to the U.S. Food and Drug Administration, the fecal-oral cycle of transmission can be disrupted by cooking food properly, preventing cross-contamination, instituting barriers such as gloves for food workers, instituting health care policies so food industry employees seek treatment when they are ill, pasteurization of juice or dairy and proper hand washing requirements.

Shiga toxin-producing E. coli (STEC), specifically serotype O157:H7, have also been transmitted by flies, as well as direct contact with farm animals, petting zoo animals,and airborne particles found in animal-rearing environments.

Diagnosis


Enteropathogenic E. coli (EPEC) and Enterotoxigenic E. coli (ETEC) - UTI or GIT infections in infants are caused by EPEC which presents as watery diarrhea, meaning that PMN's will not be observed in the stool neither with methylene blue nor Gram stain. First off, G - ve, rods, with no particular arrangement are seen in Gram stain. Then, either MacConkey agar or EMB agar (or both) are inoculated with the stool. On MacConkey agar, deep red colonies are produced as the organism is lactose positive, and this utilization will cause the medium's pH to drop leading to darkening of the medium. Growth on EMB agar would show black colonies with greenish-black metallic sheen. This is diagnosic of E. coli. The organism is lysine positive, and grows on TSI slant with a (A/A/g+/H2S-) profile. Also, IMViC test is ++-- for E. coli; as it's indol positive (red ring) and methyl red positive (bright red), but VP negative (no change-colorless) and citrate negative (no change-green color). Serology is done using the SSS-Coagglutination test.

Enterohaemorrhagic E. coli O157:H7 - isolated either from urine or, more commonly, stool. There exist three protocols for diagnosis:

(a) Diagnosis is carried out as is the case with EPEC and ETEC.

(b) Gram stain:- G - ve, rods, with no particular arrangement. Sorbitol-MacConkey agar is a modified MacConkey agar which has sorbitol instead of lactose. EHEC would produce colorless colonies as it can't utilize sorbitol. TSI slant and IMViC are then performed. Serology detects O157:H7 antigens.

(c) Two bottles of verocells are used. One is inoculated with equal portions of a stool extract and antitoxin, while the other has only a stool extract. If the first bottle shows neutralization (no cytopathic effects) while the second doesn't, then the test is considered positive. Otherwise, it's not. This is based on the fact that the toxin produced by this strain is neutralized in the presence of its specific antibody, meaning that it won't be able to exert its effects on cells.

Role in biotechnology
Because of its long history of laboratory culture and ease of manipulation, E. coli also plays an important role in modern biological engineering and industrial microbiology.The work of Stanley Norman Cohen and Herbert Boyer in E. coli, using plasmids and restriction enzymes to create recombinant DNA, became a foundation of biotechnology.

Considered a very versatile host for the production of heterologous proteins,researchers can introduce genes into the microbes using plasmids, allowing for the mass production of proteins in industrial fermentation processes. Genetic systems have also been developed which allow the production of recombinant proteins using E. coli. One of the first useful applications of recombinant DNA technology was the manipulation of E. coli to produce human insulin. Modified E. coli have been used in vaccine development, bioremediation, and production of immobilised enzymes. E. coli cannot, however, be used to produce some of the more large, complex proteins which contain multiple disulfide bonds and, in particular, unpaired thiols, or proteins that also require post-translational modification for activity.

Model organism
E. coli is frequently used as a model organism in microbiology studies. Cultivated strains (e.g. E. coli K12) are well-adapted to the laboratory environment, and, unlike wild type strains, have lost their ability to thrive in the intestine. Many lab strains lose their ability to form biofilms.These features protect wild type strains from antibodies and other chemical attacks, but require a large expenditure of energy and material resources.

In 1946, Joshua Lederberg and Edward Tatum first described the phenomenon known as bacterial conjugation using E. coli as a model bacterium, and it remains the primary model to study conjugation. E. coli was an integral part of the first experiments to understand phage genetics, and early researchers, such as Seymour Benzer, used E. coli and phage T4 to understand the topography of gene structure. Prior to Benzer's research, it was not known whether the gene was a linear structure, or if it had a branching pattern.

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