Cattle hides are a main source of enterohemorrhagic Escherichia coli (EHEC) contamination of beef carcasses. The objectives of this study were to (1) determine the prevalence of "top 6" non-O157 plus O157:H7 EHEC (EHEC-7) on feedlot cattle hides and their matched preintervention carcasses; (2) assess the agreement among detection methods for these matrices; and (3) conduct a molecular risk assessment of EHEC-7 isolates. Samples from 576 feedlot cattle were obtained at a commercial harvest facility and tested for EHEC-7 by a culture-based method and the polymerase chain reaction/mass spectrometry-based NeoSEEK(™) STEC Detection and Identification test (NS). Prevalence data were analyzed with generalized linear mixed models. The cumulative prevalence of EHEC-7 in hide samples as detected by NS was 80.7%, with a distribution of 49.9%, O145; 37.1%, O45; 12.5%, O103; 11.0%, O157; 2.2%, O111; 2.0%, O121; and 0.2%, O26. In contrast, the cumulative prevalence of EHEC-7 in hide samples by culture was 1.2%, with a distribution of 0.6%, O157; 0.4%, O26; 0.2%, O145; and 0%, O45, O103, O111, and O121. The cumulative prevalence of EHEC-7 on matched preintervention carcasses as detected by NS was 6.0%, with a distribution of 2.8%, O157; 1.6%, O145; 1.2%, O103; 1.1%, O45; 0.2%, O26; and 0.0%, O111 and O121. Although the culture-based method detected fewer positive hide samples than NS, it detected EHEC in five hide samples that tested negative for the respective organism by NS. McNemar's chi-square tests indicated significant (p<0.05) disagreement between methods. All EHEC-7 isolates recovered from hides were seropathotype A or B, with compatible virulence gene content. This study indicates that "top 6" and O157:H7 EHEC are present on hides, and to a lesser extent, preintervention carcasses of feedlot cattle at harvest. However, continued improvement in non-O157 detection methods is needed for accurate estimation of prevalence, given the discordant results across protocols.

Ubc9 was identified as a cellular protein that interacts with the Gag protein of Mason-Pfizer monkey virus.We show here that Ubc9 also interacts with the human immunodeficiency virus type 1 (HIV-1) Gag protein and that their interaction is important for virus replication. Gag was found to colocalize with Ubc9 predominantly at perinuclear puncta. While cells in which Ubc9 expression was suppressed with RNA interference produced normal numbers of virions, these particles were 8-to 10-fold less infectious than those produced in the presence of Ubc9. The nature of this defect was assayed for dependence on Ubc9 during viral assembly, trafficking, and Env incorporation. The Gag-mediated assembly of virus particles and protease-mediated processing of Gag and Gag-Pol were unchanged in the absence of Ubc9. However, the stability of the cellassociated Env glycoprotein was decreased and Env incorporation into released virions was altered. Interestingly, overexpression of the Ubc9 trans-dominant-negative mutant C93A, which is a defective E2-SUMO-1 conjugase, suggests that this activity may not be required for interaction with Gag, virion assembly, or infectivity. This finding demonstrates that Ubc9 plays an important role in the production of infectious HIV-1 virions.

BackgroundSince 1982, specific serotypes of Shiga toxin-producing Escherichia coli (STEC) have been recognized as significant foodborne pathogens acquired from contaminated beef and, more recently, other food products. Cattle are the major reservoir hosts of these organisms, and while there have been advancements in food safety practices and industry standards, STEC still remains prevalent within beef cattle operations with cattle hides implicated as major sources of carcass contamination. To investigate whether the composition of hide-specific microbial communities are associated with STEC prevalence, 16S ribosomal RNA (rRNA) bacterial community profiles were obtained from hide and fecal samples collected from a large commercial feedlot over a 3-month period. These community data were examined amidst an extensive collection of prevalence data on a subgroup of STEC that cause illness in humans, referred to as enterohemorrhagic E. coli (EHEC). Fecal 16S rRNA gene OTUs (operational taxonomic units) were subtracted from the OTUs found within each hide 16S rRNA amplicon library to identify hide-specific bacterial populations.ResultsComparative analysis of alpha diversity revealed a significant correlation between low bacterial diversity and samples positive for the presence of E. coli O157:H7 and/or the non-O157 groups: O26, O111, O103, O121, O45, and O145. This trend occurred regardless of diversity metric or fecal OTU presence. The number of EHEC serogroups present in the samples had a compounding effect on the inverse relationship between pathogen presence and bacterial diversity. Beta diversity data showed differences in bacterial community composition between samples containing O157 and non-O157 populations, with certain OTUs demonstrating significant changes in relative abundance.ConclusionsThe cumulative prevalence of the targeted EHEC serogroups was correlated with low bacterial community diversity on pre-harvest cattle hides. Understanding the relationship between indigenous hide bacterial communities and populations may provide strategies to limit EHEC in cattle and provide biomarkers for EHEC risk assessment.Electronic supplementary materialThe online version of this article (doi:10.1186/s40168-016-0155-4) contains supplementary material, which is available to authorized users.

The primary objective of this study was to determine the prevalence and level of enterohemorrhagic Escherichia coli (EHEC) O26, O45, O103, O111, O121, and O145 (collectively EHEC-6) plus EHEC O157 in fecal, hide, and preintervention carcass surface samples from culled dairy cows. Matched samples (n =300) were collected from 100 cows at harvest and tested by a culture-based method and two molecular methods: NeoSEEK STEC (NS) and Atlas STEC EG2 Combo. Both the culture and NS methods can be used to discriminate among the seven EHEC types (EHEC-7), from which the cumulative prevalence was inferred, whereas the Atlas method can discriminate only between EHEC O157 and non-O157 EHEC, without discrimination of the serogroup. The EHEC-7 prevalence in feces, hides, and carcass surfaces was 6.5, 15.6, and 1.0%, respectively, with the culture method and 25.9, 64.9, and 7.0%, respectively, with the NS method. With the Atlas method, the prevalence of non-O157 EHEC was 29.1, 38.3, and 28.0% and that of EHEC O157 was 29.1, 57.0, and 3.0% for feces, hides, and carcasses, respectively. Only two samples (a hide sample and a fecal sample) originating from different cows contained quantifiable EHEC. In both samples, the isolates were identified as EHEC O157, with 4.7 CFU/1,000 cm2 in the hide sample and 3.9 log CFU/g in the fecal sample. Moderate agreement was found between culture and NS results for detection of EHEC O26 (κ =0.58, P &lt; 0.001), EHEC O121 (κ = 0.50, P &lt; 0.001), and EHEC O157 (κ = 0.40, P &lt; 0.001). No significant agreement was observed between NS and Atlas results or between culture and Atlas results. Detection of an EHEC serogroup in fecal samples was significantly associated with detection of the same EHEC serogroup in hide samples for EHEC O26 (P =0.001), EHEC O111 (P =0.002), EHEC O121 (P &lt; 0.001), and EHEC-6 (P = 0.029) based on NS detection and for EHEC O121 (P &lt; 0.001) based on detection by culture. This study provides evidence that non-O157 EHEC are ubiquitous on hides of culled dairy cattle and that feces are an important source of non-O157 EHEC hide contamination.