Non-sorbitol-fermenting, ␤-glucuronidase-negative Escherichia coli O157:H7 strains are regarded as a clone complex, and populations from different geographical locations are believed to share a recent common ancestor. Despite their relatedness, high-resolution genotyping methods can detect significant genome variation among different populations. Phylogenetic analysis of high-resolution genotyping data from these strains has shown that subpopulations from geographically unlinked continents can be divided into two primary phylogenetic lineages, termed lineage I and lineage II, and limited studies of the distribution of these lineages suggest there could be differences in their propensity to cause disease in humans or to be transmitted to humans. Because the genotyping methods necessary to discriminate the two lineages are tedious and subjective, these methods are not particularly suited for studying the large sets of strains that are required to systematically evaluate the ecology and transmission characteristics of these lineages. To overcome this limitation, we have developed a lineage-specific polymorphism assay (LSPA) that can readily distinguish between the lineage I and lineage II subpopulations. In the studies reported here, we describe the development of a six-marker test (LSPA-6) and its validation in a side-by-side comparison with octamer-based genome scanning. Analysis of over 1,400 O157:H7 strains with the LSPA-6 demonstrated that five genotypes comprise over 91% of the strains, suggesting that these subpopulations may be widespread.

Cattle are an important reservoir of Escherichia coli O157:H7 leading to contamination of food and water, and subsequent human disease. This pathogen colonizes its hosts by producing several proteins such as Tir and EspA that are secreted by a type III secretion system. These proteins play a role in colonization of the intestine, suggesting that they might be useful targets for the development of a vaccine to reduce levels of this organism in cattle. Vaccination of cattle with proteins secreted by E. coli O157:H7 significantly reduced the numbers of bacteria shed in feces, the numbers of animals that shed, and the duration of shedding in an experimental challenge model. Vaccination of cattle also significantly (P=0.04) reduced the prevalence of E. coli O157:H7 in a clinical trial conducted in a typical feedlot setting. This strategy suggests it is possible to vaccinate cattle to decrease the level of E. coli O157:H7 shedding for the purpose of reducing the risk of human disease.

Enterotoxigenic Escherichia coli (ETEC) strains that produce multiple enterotoxins are important causes of severe dehydrating diarrhea in human beings and animals, but the relative importance of these enterotoxins in the pathogenesis is poorly understood. Gnotobiotic piglets were used to study the importance of heat-labile enterotoxin (LT) in infection with an ETEC strain that produces multiple enterotoxins. LT ؊ (⌬eltAB) and complemented mutants of an F4؉ LT ؉ STb ؉ EAST1 ؉ ETEC strain were constructed, and the virulence of these strains was compared in gnotobiotic piglets expressing receptors for F4 ؉ fimbria. Sixty percent of the piglets inoculated with the LT ؊ mutant developed severe dehydrating diarrhea and septicemia compared to 100% of those inoculated with the nalidixic acid-resistant (Nal r ) parent and 100% of those inoculated with the complemented mutant strain. Compared to piglets inoculated with the Nal r parent, the mean rate of weight loss (percent per hour) of those inoculated with the LT ؊ mutant was 67% lower (P < 0.05) and that of those inoculated with the complemented strain was 36% higher (P < 0.001). Similarly, piglets inoculated with the LT ؊ mutant had significant reductions in the mean bacterial colony count (CFU per gram) in the ileum; bacterial colonization scores (square millimeters) in the jejunum and ileum; and clinical pathology parameters of dehydration, electrolyte imbalance, and metabolic acidosis (P < 0.05). These results indicate the significance of LT to the development of severe dehydrating diarrhea and postdiarrheal septicemia in ETEC infections of swine and demonstrate that EAST1, LT, and STb may be concurrently expressed by porcine ETEC strains.

Although heat-stable (ST) and heat-labile (LT) enterotoxins produced by enterotoxigenic Escherichia coli (ETEC) have been documented as important factors associated with diarrheal diseases, investigations assessing the contributions of individual enterotoxins to the pathogenesis of E. coli infection have been limited. To address the individual roles of enterotoxins in the diarrheal؉ astA STb ؉ strain did not, although diarrhea developed in several piglets. The changes in the blood packed-cell volume and plasma total protein of gnotobiotic piglets inoculated with the LT-positive strains were significantly greater than those of pigs inoculated with the K88 astA/pBR322 strain (P ‫؍‬ 0.012, P ‫؍‬ 0.002). Immunochemistry image analysis also suggested that LT enhanced bacterial colonization in a gnotobiotic piglet model. This investigation suggested that LT is a major contributor to the virulence of K88؉ ETEC and that isogenic constructs are a useful tool for studying the pathogenesis of ETEC infection.Escherichia coli strains that colonize the small intestines, invade intestinal epithelial cells, and/or produce one or more toxins are important causes of diarrheal disease in both farm animals and humans. The virulence of enterotoxigenic E. coli (ETEC) is believed to be associated with the production of fimbrial adhesins and enterotoxins (1,19,35,36,51,54). Fimbrial adhesins mediate the attachment of bacteria to the surface of host epithelium cells and allow bacterial colonization. Fimbriae produced by different ETEC strains are quite diverse (21). In swine, ETEC strains that produce K88 (F4) or F18 are the most common currently associated with diarrheal diseases (19). These fimbriae apparently bind to glycoconjugates in the porcine enterocyte brush borders, and the absence of the respective glycoconjugate renders the animal resistant to bacterial colonization and consequent diarrheal diseases (14,15,20,48,49).Enterotoxins, including heat-stable enterotoxins (STa and STb) and heat-labile enterotoxin (LT) (23,25,39,45), have been found to disrupt intestinal fluid homeostasis and to cause hypersecretion of fluid and electrolytes through activation of adenylate cyclase (by LT) or guanylate cyclase (by STa) in small intestinal mucosal cells (26, 34). There are two major serogroups of LT found among E. coli strains: LT-I and LT-II. LT-I is associated with diarrheal diseases of both humans and animals, while LT-II is typically associated with diarrheal disease in animals. STs are small and monomeric molecules and may be associated with either human or animal disease (45, 55). STa and STb are the two classes of STs first recognized and differ from each other in both structure and enzyme activity (11,12). STa is produced by ETEC and other bacteria, while STb is found only associated with ETEC. A third ST, enteroaggregative E. coli (EAEC) EAST1, has been more recently identified. It is a plasmid-mediated enterotoxin, of low molecular weight and is frequently but not exclusively associated with EAEC isolated from children with p...

This study was designed to describe the percentage of cattle shedding Escherichia coli O157:H7 in Midwestern U.S. feedlots and to discover relationships between the point prevalence of cattle shedding the organism and the characteristics of those cattle or the conditions of their pens. Cattle from 29 pens of five Midwestern feedlots were each sampled once between June and September 1999. Feces were collected from the rectum of each animal in each pen. Concurrently, samples of water were collected from the water tank, and partially consumed feed was collected from the feedbunk of each pen. Characteristics of the cattle and conditions of each pen that might have affected the prevalence of cattle shedding E. coli O157:H7 were recorded. These factors included the number of cattle; the number of days on feed; and the average body weight, class, and sex of the cattle. In addition, the temperature and pH of the tank water were determined, and the cleanliness of the tank water and the condition of the pen floor were subjectively assessed. The samples of feces, feed, and water were tested for the presence of E. coli O157:H7. E. coli O157:H7 was isolated from the feces of 719 of 3,162 cattle tested (23%), including at least one animal from each of the 29 pens. The percentage of cattle in a pen shedding E. coli O157:H7 did not differ between feedyards, but it did vary widely within feedyards. A higher prevalence of cattle shed E. coli O157:H7 from muddy pen conditions than cattle from pens in normal condition. The results of this study suggest that E. coli O157:H7 should be considered common to groups of feedlot cattle housed together in pens and that the condition of the pen floor may influence the prevalence of cattle shedding the organism.