In December 1998, the WHO convened an international meeting in Japan to examine the epidemiology and progress towards vaccine development for enterotoxigenic Escherichia coli (ETEC) and enterohemorrhagic E. coli (EHEC).

Vaccines against E. coli O157:H7— enterohemorrhagic E. coli.

In humans:

Success claimed as vaccine beats E. coli

Washington, AP

A vaccine against E. coli, the deadly food poisoning bacteria that forced the recall last year of millions of kilograms of US beef, has been tested successfully on a small group of volunteers, researchers said on Monday.

To make this effective vaccine, the scientists chemically linked a polysaccharide (sugar chain) from the capsule of the bacterium to the genetically inactivated toxin from another bacterium, Pseudomonas aeruginosa

Scientists at the US National Institute of Child Health and Human Development and the Carolinas Medical Center in Charlotte, North Carolina, report that a preliminary study using 87 volunteers showed that the vaccine causes an immune reaction that could protect against infection by E. coli O157. "This is still very early in the research," said Dr. Dwayne Alexander, director of the Institute of Child Health and Human Development, one of the US National Institutes of Health. "This is the first human study of this proposed vaccine."

He said the important finding is that the vaccine produced a level of antibody in the volunteers that could kill E. coli O157 in the test tube. "We don't know yet if it will kill the bacteria in the body," he said.

Dr. Alexander said the next step is tests to determine if the vaccine will prevent E. coli infection in cattle, which are thought to be the most common source of the infection.

E. coli O157 is a deadly new strain of bacteria that can contaminate beef, fruit juice and other foods, causing severe symptoms of food poisoning, including bloody diarrhea and damaged kidneys. People can also become infected by swimming in lakes or rivers contaminated with the organism.

Children are most seriously affected by the infection. An outbreak in Japan last year infected more than 10,000 people in just two months.

E. coli is spread most frequently from cattle manure that can get into meat during butchering or onto fruits or vegetables in the field. Water runoff from pastures where there are infected cattle can contaminate rivers and lakes.

Researchers believe that some genes of a dangerous bacterium, called Shigella, were transferred into E. coli during a Shigella epidemic in Central America in the 1970s. This transformed one strain of a usually harmless germ into a pathogen that does not respond well to antibiotics and can cause severe food poisoning.

Experimental Vaccine Against E. coli O157 Proven Safe and Effective In Preliminary Trials

UBC research breakthrough holds promise for E. coli vaccine

Bacterial Ghosts as an oral vaccine

Another kind of experimental vaccine that is currently being developed is an Oral Vaccine.

Dead bacteria that contain empty cell envelopes are used in many diseases to produce antibody protection. The cell envelopes contain antigenic epitopes that can be recognized by the immune system and can lead to the development of an immune reaction. Those envelopes are called "bacterial ghosts". They have the same surface components of live cells and are capable of inducing strong immune responses, but they lack the potentially harmful genetic material.

 Use of "bacterial ghosts" can lead to the induction of immune memory and a secondary immune response.

The existing ways of "killing" bacteria usually involve the use of high temperature or chemical-processing treatments.

These methods are problematic in all types of bacteria and especially problematic in E. coli because they lead to denaturation and harm the structure of the antigenic epitopes that are on the cell envelopes.

For this reason, scientists are looking for ways to produce bacterial ghosts without damaging the epitope structure.

Pathogenomic research has led to such production by degrading the E. coli DNA using a plasmid that has the nuclease gene in combination with the lysis gene pML1.

This plasmid is activated by a temperature shift from 28 to 42o Celcius. This activation causes lysis of the E. coli bacterial DNA without damaging the envelope structure.

To test the protection afforded by this vaccine against exposure to E. coli, it was given to mice via oral vaccination following the EHEC ghost and challenge scheme.

Mice were divided into four groups, A to D, with different schemes of immunization. All mice were deprived of food 24 h before oral immunization. Mice from group A were immunized once on day 0. Group B mice were immunized twice, once on day 0 and once on day 28. For all immunizations, the oral dose was 1 mg of freeze-dried EHEC ghost strain in 30 (ml) of phosphate-buffered saline (PBS), applied through a soft polyethylene catheter.

Group C mice, were immunized with 30 (ml) of PBS once on day 0. Mice from group D were immunized with 30 (ml)  of PBS twice, once on day 0 and once on day 28. On day 55, all mice were orally challenged with 108 CFU of heterologous EHEC.

The survival rate of the mice was checked 21 days after the challenge.

The results are presented in chart1:

For specific antibody determinations, blood was taken from the orbital sinuses

of the surviving mice in groups A, B, C, and D at various time points.

The average results are presented in chart 2:

Vaccine may protect against E coli

A hundred cows awaited judging in a fairground barn. Groomed and pampered, they were the pride of their farms. One harbored a deadly germ. But nobody knew that. Not until later, when medical sleuths figured out what killed a 3-year-old girl and a 79-year-old man, and made more than 1,000 other fair-goers terribly sick. The malignant microbe at the Washington County Fair was Escherichia coli 0157:H7.

The bacterium does not make cows sick and does not cause clinical symptoms in cattle, but in humans it causes serious disease that can lead to kidney damage.

Until recently, it wasn’t even clear how widespread the organism was in cattle. Last year, scientists estimated that about 1 to 3 percent of cattle were infected with E. coli 0157:H7. Now, USDA researchers say that the numbers appear to be much higher. "In every herd we’ve tested, there have been at least some animals positive for 0157."

Cattle are the primary source of E. coli infection for humans. This infection can occur in two ways:

The first occurs when people eat undercooked infected meat.

The second, when people drink water that has been contaminated by infected cattle feces.

Eliminating E. coli O157 from cattle could go a long way towards the prevention of disease outbreaks in humans.

USDA scientists working on oral E. coli vaccine

A vaccine developed by researchers at the USDA  significantly decreases infection of cattle by the toxic strain of E. coli that contaminated drinking water in Walkerton, Ontario.

The key ingredient of this oral vaccine for cattle is intimin, a protein on the outer membrane of the O157:H7 strain. The bacteria need intimin to attach to intestinal tissue.

(Studies found that rather than making use of a host protein, the bacterium fires a soluble bacterial protein into the host cell membrane. The protein is then modified in the host cell membrane to form a perfect landing site for intimin, a bacterial surface molecule that binds with the host cell surface.)

This study was assisted by an earlier study that showed that calves injected with purified bacterial intimin would develop antibodies against it. This confirmed previous studies in mice that showed that intimin-specific responses reduced the adherence of E. coli O157:H7 bacteria to both cultured tissue cells and intestinal cells in the intact animal.

Previous works also revealed that intimin-fighting antibodies interfere with E. coli O157:H7 colonization and lessen intestinal damage in newborn pigs. Earlier studies found that pregnant pigs vaccinated against bacterial intimin developed antibodies against it in their sera and colostrum. Also, newborn piglets experimentally challenged with a Shiga-toxin-negative E. coli O157:H7 strain, and who ingested colostrum from intimin-vaccinated pigs, had fewer of the inoculated bacteria in their intestines than did piglets nursed by nonvaccinated pigs.

 Another possible way of fighing E.coli infection is by changing the cows' diet:

Hay! What a way to fight E. coli.

Vaccine for 'Montezuma's revenge'/'Delhi belly'—enterotoxigenic Escherichia coli

 Montezuma's revenge is a disease caused by ETEC following ingestion of contaminated food or water. It is characterized by profuse watery diarrhea lasting for several days: 50% of the travelers to Africa, Southeast Asia and the Indian subcontinent, Central and South America and the Caribbean, get it.

The disease is also widespread among children under 5 years of age in the developing world, and probably accounts for approximately 200 million diarrhea episodes and 380,000 deaths annually.

More about: Enterotoxigenic Escherichia coli (ETEC)

Several approaches have been pursued to develop specific ETEC vaccines, including the use of:

For diarrhea to result, ETEC must colonize the small intestine. ETEC attach to specific receptors on the surface of enterocytes in the intestinal lumen by virtue of their hair-like fimbriae also termed pili; or fibrils, which are host-specific proteinaceous hair-like structures on the bacterial cell surface. There are more than 20 types of fimbria antigens, called E. coli surface antigens (CSAs) or colonization factor antigens (CFAs).

Several adhesive CFAs have been localized in the fimbriae.

The most frequent and best characterized of the CFAs are CFA/I and CFA/II.

 ETEC may also produce a heat-stable toxin (ST) that is of low molecular weight (4000) and is resistant to boiling for 30 min. This low-molecular-weight protein is excreted into the medium during the growth of ST-elaborating strains. Unlike LT, ST is poorly antigenic and for this reason, the identification of ST-producing strains has relied on an assay involving the intragastric injection of infant mice.

ST causes an increase in cyclic GMP [O3] in the host cell cytoplasm, leading to the same effects as an increase in cAMP. STa is known to act by binding to a guanylate cyclase that is located on the apical membranes of host cells, thereby activating the enzyme. This leads to the secretion of fluid and electrolytes, resulting in diarrhea.

The infective dose of ETEC for adults has been estimated to be at least 108 cells; but the young, the elderly and the infirm may be susceptible to lower levels.

If ETEC is detected, levels of toxins should also be enumerated to assess the potential hazard of the contaminated food. Both LT and ST genes have been sequenced and PCR and gene probe assays developed.

Edible transgenic plants that express the cholera toxin B subunit (CTB)

Immunization with inactivated oral B subunit/whole-cell (BS-WC) cholera vaccine against the LT toxin offers short-term (3 months) but significant (>67%) protection against travelers' diarrhea caused by LT-related ETEC. Since it expresses the cholera toxin B (CTB) subunit, the live attenuated oral cholera vaccine strain CVD 103-HgR may induce similar protection.

Why are those specific antigens—LT, ST, and colonization factors chosen for making vaccines?

The new Microscience vaccine, spi-VEC [O4] , is targeted to fight infection by ETEC. The vaccine is actually made from Salmonella bacteria, rendered harmless by the removal of several important genes.

The Salmonella bacteria then act as a vehicle to deliver this directly to cells of the immune system, resulting in efficient antigen presentation and strong immune responses.

Key genes from ETEC are added to fool the body into generating antibodies against it. Each dose of vaccine is expected to offer about six months' protection.

What genes can be added to the Salmonella bacteria to produce antibodies against E. coli?

This vaccine is designed to protect people against ETEC, to protect vacationers against the dreaded 'Delhi belly'.

The vaccine, which is taken as a drink, has undergone promising initial trials. Researchers in London gave increasing doses of the spi-VEC vaccine to 36 volunteers to test for safety and success in creating an immune response.

After a single dose, 50% of the volunteers had high levels of immune response against a known protective antigen of ETEC, rising to 70% after two doses.

Dr. David Lewis, clinical investigator with the company developing the vaccine, Microscience, said: "The results of this first study with spi-VEC are exciting and these are the best results achieved to date in humans using this type of oral delivery system."

http://www.vaccine.ac.uk/SkyNews.htm

What are the advantages of giving vaccines orally?

Eat up your vaccines:

Eat up your vaccines