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Cattle are the main reservoir of Enterohemorrhagic Escherichia coli (EHEC), with O157:H7 the distinctive serotype. EHEC is the main causative agent of a severe systemic disease, Hemolytic Uremic Syndrome (HUS). Argentina has the highest pediatric HUS incidence worldwide with 12-14 cases per 100,000 children. Herein, we assessed the genomes of EHEC O157:H7 isolates recovered from cattle in the humid Pampas of Argentina. According to phylogenetic studies, EHEC O157 can be divided into clades. Clade 8 strains that were classified as hypervirulent. Most of the strains of this clade have a Shiga toxin stx2a-stx2c genotype. To better understand the molecular bases related to virulence, pathogenicity and evolution of EHEC O157:H7, we performed a comparative genomic analysis of these isolates through whole genome sequencing. The isolates classified as clade 8 (four strains) and clade 6 (four strains) contained 13 to 16 lambdoid prophages per genome, and the observed variability of prophages was analysed. An inter strain comparison show that while some prophages are highly related and can be grouped into families, other are unique. Prophages encoding for stx2a were highly diverse, while those encoding for stx2c were conserved. A cluster of genes exclusively found in clade 8 contained 13 genes that mostly encoded for DNA binding proteins. In the studied strains, polymorphisms in Q antiterminator, the Q-stx2A intergenic region and the O and P γ alleles of prophage replication proteins are associated with different levels of Stx2a production. As expected, all strains had the pO157 plasmid that was highly conserved, although one strain displayed a transposon interruption in the protease EspP gene. This genomic analysis may contribute to the understanding of the genetic basis of the hypervirulence of EHEC O157:H7 strains circulating in Argentine cattle. This work aligns with other studies of O157 strain variation in other populations that shows key differences in Stx2a-encoding prophages., Competing Interests: The authors have declared that no competing interests exist.

While the differential association of Escherichia coli O157 genotypes with animal and human hosts has recently been well documented, little is known about their distribution between countries and how this might affect regional disease rates. Here, we used a 48-plex single nucleotide polymorphism (SNP) assay to segregate 148 E. coli O157 isolates from Australia, Argentina, and the United States into 11 SNP lineages. We also investigated the relationship between SNP lineages, Shiga toxin (Stx) gene profiles, and total Stx production. E. coli O157 isolates clearly segregated into SNP lineages that were differentially associated with each country. Of the 11 SNP lineages, seven were detected among isolates from a single country, two were detected among isolates from all three countries, and another two were detected only among U.S. and Argentinean isolates. A number of Australian (30%) and Argentinean (14%) isolates were associated with novel, previously undescribed SNP lineages that were unique to each country. Isolates within SNP lineages that were strongly associated with the carriage of stx2a produced comparatively more Stx on average than did those lacking the stx2a subtype. Furthermore, the proportion of isolates in stx2a-associated SNP lineages was significantly higher in Argentina and the United States than Australia (P < 0.05). This study provides evidence for the geographic divergence of E. coli O157 and for a prominent role of stx2a in total Stx production. These results also highlight the need for more comprehensive studies of the global distribution of E. coli O157 lineages and the impacts of regionally predominant E. coli O157 lineages on the prevalence and severity of disease., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)

More than 400 serotypes of Shiga toxin-producing Escherichia coli (STEC) have been implicated in outbreaks and sporadic human diseases. In recent years STEC strains belonging to serogroup O178 have been commonly isolated from cattle and food of bovine origin in South America and Europe. In order to explore the significance of these STEC strains as potential human pathogens, 74 German and Argentinean E. coli O178 strains from animals, food and humans were characterized phenotypically and investigated for their serotypes, stx-genotypes and 43 virulence-associated markers by a real-time PCR-microarray. The majority (n = 66) of the O178 strains belonged to serotype O178:H19. The remaining strains divided into O178:H7 (n = 6), O178:H10 (n = 1), and O178:H16 (n = 1). STEC O178:H19 strains were mainly isolated from cattle and food of bovine origin, but one strain was from a patient with hemolytic uremic syndrome (HUS). Genotyping of the STEC O178:H19 strains by pulsed-field gel electrophoresis revealed two major clusters of genetically highly related strains which differ in their stx-genotypes and non-Stx putative virulence traits, including adhesins, toxins, and serine-proteases. Cluster A-strains including the HUS-strain (n = 35) carried genes associated with severe disease in humans (stx2a, stx2d, ehxA, saa, subAB1, lpfAO113 , terE combined with stx1a, espP, iha). Cluster B-strains (n = 26) showed a limited repertoire of virulence genes (stx2c, pagC, lpfAO113 , espP, iha). Among O178:H7 strains isolated from deer meat and patients with uncomplicated disease a new STEC variant was detected that is associated with the genotype stx1c/stx2b/ehxA/subAB2/espI/[terE]/espP/iha. None of the STEC O178 strains was positive for locus of enterocyte effacement (LEE)- and nle-genes. Results indicate that STEC O178:H19 strains belong to the growing group of LEE-negative STEC that should be considered with respect to their potential to cause diseases in humans.

Shiga toxin-producing Escherichia coli (STEC) is an important foodborne pathogen, and ruminants are recognized as the main natural reservoir. The purposes of this study were to detect E. coli O157 in bovine feces and surface water in a beef cattle farm of Gualeguaychú, Argentina; to characterize the isolates; and to establish the clonal relatedness by pulsed-field gel electrophoresis. Between September 2005 and November 2006, 288 samples of bovine feces and 79 samples of water troughs were studied. E. coli O157 was detected by immunomagnetic separation and polymerase chain reaction as screening techniques. The rfb(O157) gene was detected in 3.8% of the 288 fecal samples and in 17.7% of the 79 water samples. The stx gene was detected in all rfb(O157)-positive fecal samples and in 5.1% of water samples. Eleven E. coli O157 strains isolated from bovine fecal samples and eight from water samples were characterized. The most frequent stx genotype identified was stx(1) and stx(2c(vh-a)). Twelve (63.2%) strains harbored fliC(H7), eae, and ehxA genes. Using pulsed-field gel electrophoresis with the enzyme XbaI, a total of eight patterns with at least 72.1% similarity were identified among the 19 strains. The patterns of 15 strains were grouped into four clusters: two of them included only bovine strains and the other two only aquatic strains. No genetic correlation was established between the bovine and water STEC strains detected. The prevalence of STEC O157:H7 established in the herd studied was higher than that previously reported for Argentine grazed cattle.

Aims: To study the seasonal variation of Shiga toxin-encoding genes (stx) and to investigate the presence of Shiga toxin-producing Escherichia coli (STEC) O157 in cattle belonging to five dairy farms from Argentina., Methods and Results: Rectal swab samples were collected from 360 dairy cows in each season and 115 and 137 calves in autumn and in spring, respectively. The stx were investigated by multiplex PCR and it was used as the indicator for STEC. Samples positives for stx were tested by PCR for eae-gamma1 of E. coli O157 and then subjected to IMS (immunomagnetic separation). In positive animals significant differences in the prevalence of stx between warm and cold seasons were detected. In warm seasons, stx1 + stx2 increased and stx1 decreased, independently of the animal category. The prevalence of STEC O157 in cows and calves were 0.2% and 0.8%, respectively., Conclusions: This work provides new data about the occurrence of stx and STEC O157 in dairy herds from Argentina and suggests a relationship between the type of stx and season of year., Significance and Impact of Study: The detection of STEC O157 and the seasonality of stx and its types provide an opportunity to improve control strategies designed to prevent contamination of food products and transmission animal-person.

Shiga toxin-producing Escherichia coli (STEC), responsible for the hemolytic uremic syndrome, is an endemic pathogen in Argentina. We studied the prevalence of STEC in fecal samples from cats and dogs of Buenos Aires city and suburbs. Cultures were used for screening stx1/stx2 and rfbO157 by multiplex PCR. All E. coli-positive colonies for these genes were further characterized for the eae gene and for serotypes. In dogs, 17 (3.7%), 19 (4.2%) and 34 (7.5%) of samples were positive for stx2, stx1 and rfb, respectively. In cats, six (4.0%) of the samples were positive for stx2, three (2.0%) for stx1 and four (2.7%) for rfbO157. In 18 (4.0%) of the dog samples, a bacteriological diagnosis was obtained by isolation. The percentage of positive isolates corresponding to the rfbO157 and to the stx2 genotypes were 2.9% and 1.1%, respectively. In four of the cat samples, the bacteriological diagnosis for stx2 (2.6% prevalence of STEC) was confirmed. Although these data suggest that the high infection index of STEC in children in Argentina does not seem to be due mainly to the role of cats and dogs, there are some strains with virulence genes in common for humans and their domestic animals.

Between July 1999 and December 2000, the prevalence of Shiga toxin-producing Escherichia coli (STEC) was established in 200 Argentine healthy young beef steers (14-16 months old) grown under local production systems with a feed grain period of 3-4 months, and the STEC strains isolated were examined in regard to their phenotypic and genotypic characteristics. Stool samples (n = 70) and rectal swabs (n = 130) were taken at the slaughterhouse level. By polymerase chain reaction (PCR), Shiga toxin (stx) gene sequences were detected in 69% of the samples. Eighty-six STEC strains were isolated from 39% of the animals. Serogroups identified, in order of frequency, were: O8 (16 strains), O113 (14), O103 (5), O91 (4), O171 (3), O174 (3), O25 (2), O112 (2), O145 (2), O2, O11, O104, O121, O128, O143, O146, O157. The most frequent serotype isolated was O8:H19 (12.9%). A total of 17 serotypes, including E. coli O157:H7 found in one animal (0.5%), have been previously associated with hemolytic uremic syndrome (HUS), bloody and non-bloody diarrhea in different countries, including Argentina. The prevalent genotype isolated was stx2 (51 of 86, 59.3%). Subtyping of stx2 variants showed the prevalence of stx2vh-b (25.6%) and stx2vh-a types (24.4%), and revealed the presence of an atypical stx2-v. Only 7.0% of STEC strains carried eae, and 33.7% harbored EHEC-hlyA gene. The full virulent genotype (stx/eae/EHEC-hlyA) was found to be present in 4 of the 86 (4.7%) STEC strains isolated. This research indicates that young steers from the main beef-producing area of Argentina are an important reservoir of STEC strains; however, its importance as agents of human diseases in our country has still to be established.