The present study investigated the prevalence of zoonotic STEC in small farms within rural communities, located in the agricultural Culiacan Valley along the coastal northwestern part of Mexico. The Culiacan Valley is considered to be one of the most important agricultural regions in Mexico 
. The farms, examined in this study, were located in rural communities where livestock was often raised without an efficient management of animal wastes 
. Thus, a lack of good agricultural practices could potentially contribute to the dispersal and transmission of pathogens throughout the environment. An understanding of the primary animal reservoirs in rural communities within an agricultural region would be imperative for the development of control measures of any risk factors that could lead to human infections with zoonotic STEC.
Only a limited number of studies have previously documented the prevalence of STEC in animal reservoirs in various locations in Mexico. The contamination of animal carcasses with O157:H7 and non-O157 STEC was detected in slaughter plants sampled in northeast and western Mexico 
. The recovered STEC isolates from these slaughter plants were found positive for the virulence factors intimin and hemolysin. A separate study confirmed the presence of E. coli
O157:H7 in animal feces collected from dairy and beef cattle farms and from swine-farrowing facilities throughout Central Mexico 
. Whether the recovered O157 isolates from the farms in Central Mexico harbor genes associated with severe human illness is still to be determined. Thus, additional studies are needed to identify the prevalence and phylogenetic relatedness of O157 and non-O157 STEC in other relevant agricultural regions of Mexico.
Given that cattle and other ruminants are considered relevant reservoirs of zoonotic STEC 
, the present study is the first report that has examined the prevalence of O157 and non-O157 STEC from feces of domestic animals that were raised on the sampled rural farms in the agricultural Culiacan Valley. The results from this study indicated that O157 STEC were detected in 5.4% (13/240) and non-O157 STEC were detected in 7.1% (17/240) of the animal fecal samples. The observed STEC prevalence in farm animals in Mexico’s Culiacan Valley appears to be lower when compared to some countries. Moreover, a published report that comprehensively examined the worldwide prevalence of O157 and non-O157 in livestock feces reported significant wide ranges 
. In particular, the prevalence of O157 ranged from 0.2% to 27.8% and of non-O157 ranged from 2.1% to 70.1% 
. These observations have also indicated that multiple factors, including sampling locations, hosts, periods of study and methods, may contribute to the dynamic nature observed among studies of pathogen prevalence in animals 
Due to the complexity of detecting and enumerating toxigenic E. coli
in naturally-contaminated samples 
, an enrichment method and plating on selective media was employed to increase the efficiency for isolating both O157 and non-O157 STEC. The use of the IMS method facilitated significantly higher recovery of isolates when compared to direct plating. The number of fecal samples that were positive for presumptive STEC using the IMS method was 2.1-times higher than those subjected to direct plating on selective media. These findings are in agreement with other studies that have reported an increased effectiveness in STEC detection in animal feces by using the IMS method 
. Furthermore, the increased recovery of STEC isolates that was observed during the second sampling period can be attributed to performing the enrichment step without antibiotics, a procedure proposed to increase the number of STEC cells before exposure to the selective conditions in the isolation media 
The O157:H7 serotype was identified in 40% (26/65) of the E. coli
isolates that were recovered from the domestic animals sampled in this study. Most O157 isolates were recovered from cattle feces and sheep feces. Interestingly, two O157:H7 isolates were recovered from chicken feces, considered to be a rare animal source for this serotype 
. Genotyping studies revealed that most O157:H7 isolates grouped into two PFGE 2-enzyme profiles with 98.6% homology, thus, the use of MLVA as a genotyping tool facilitated further discrimination of isolates that were indistinguishable by PFGE. Further analysis of the allelic polymorphisms of the 11 VNTR loci of the O157 MLVA system indicated that most alleles had low or intermediate diversity in the O157:H7 isolates from the Culiacan Valley. Vhec1 was the only VNTR locus that was identified to have high allelic diversity. This finding is in agreement with previous reports that documented a particularly high polymorphism at Vhec1 
. The limited allelic diversity in conjunction with the observation that all O157:H7 isolates had the same virulence profile (positive for stx2, eae, ehxA
) suggests that the O157:H7 isolates from this region in Mexico are highly related. It also appears that a small number of genetic types may predominate and persist in different animal reservoirs on the rural farms. In particular, the O157:H7 sheep isolate RM8753 was recovered by direct plating, and six months later, genotypically-identical isolates by MLVA and PFGE were recovered by the IMS method from the same sampling site (site C), but from both sheep and cattle. This finding may suggest persistence of the O157:H7 isolates in the Culiacan Valley and a potential transmission between species.
The non-O157 serotypes O8:H19, O75:H8, O111:H8 and O146:H21 were identified in 35.4% (23/65) of the STEC isolates that were recovered in the Culiacan Valley. These relevant serotypes have been previously implicated with disease outbreaks or with severe symptoms in humans 
. The non-O157 isolates were predominantly recovered from sheep and were identified to harbor either one or both stx
genes. With the exception of five isolates, all other non-O157 isolates (86.5%, 32/37) were identified to be positive for ehxA.
However, only 10.8% (4/37) of the non-O157 isolates harbored eae
gene. Similar to what was observed in the analysis of O157:H7 isolates, a diverse set of non-O157 isolates was identified at sampling site C, located in a valley between two large urban cities (Culiacan and Navolato) and proximal to the mountain ranges along the east.
Among the non-O157 serotypes that were identified in this study, serotype O75:H8 was the most variable serotype by both PFGE and MLVA. The O75:H8 isolates, representing 18.5% (12/65) of the isolates recovered from the Culiacan Valley, were a common sheep serotype. Another sheep serotype was O146:H21. The predominant isolation of O75:H8 and O146:H21 isolates from sheep was in agreement with previous reports that documented the same animal source for these non-O157 serotypes in several countries such as Australia, Brazil, New Zealand, and Spain 
. Moreover, to our knowledge, the present study is the first report of sheep feces as an animal source of STEC O111:H8, a serotype previously identified to be recovered exclusively from cattle 
Genotypic analysis by PFGE and MLVA revealed that cattle isolates with serotype O8:H19 from the Culiacan Valley were closely related. By contrast, a high genotypic diversity was identified in O8:H19 isolates from beef products in Argentina 
. In the present study, the low genomic diversity that was observed in the O8:H19 isolates may indicate that a prevalent genotype may be circulating in this geographical region in Mexico. Additional surveys would be needed to determine whether other STEC serotypes may also represent stable and predominant genotypes in this agricultural region in Mexico. Future work will investigate other sources of O157 and non-O157 STEC by examining wildlife and environmental samples (plants, soil, sediment, and irrigation water) to obtain a better understanding of pathogen diversity, persistence, and movement. Further assessment of additional molecular risk factors associated with STEC pathogenesis would also provide relevant information regarding the virulence potential of toxigenic E. coli
recovered from the Culiacan Valley in Mexico.