High-pressure synthesis of acentric sodium pyrocarbonate, Na2[C2O5].

The inorganic pyrocarbonate salt Na2[C2O5] crystallizes in the acentric, monoclinic space group P21 with Z = 2. It contains monovalent alkali metal cations together with isolated pyrocarbonate anions. The [C2O5]2--groups consist of two planar [CO3]2--groups which are slightly tilted with respect to each other around the bridging oxygen atom. Na2[C2O5] was synthesized in a laser-heated diamond anvil cell at 20(2) GPa by heating a mixture of Na2[CO3] + CO2 to ≈800(200) K. Its crystal structure was obtained by single crystal synchrotron X-ray diffraction and confirmed by density functional theory-based calculations in combination with Raman spectroscopy. Second harmonic generation measurements verified the acentric space group symmetry. The crystal structure is characterized by alternating layers of Na+-cations and [C2O5]2--complex anions.

Genomic analysis of Shiga toxin-producing Escherichia coli O157:H7 from cattle and pork-production related environments.

Three E. coli O157:H7 outbreaks have been attributed to contaminated pork in Alberta, Canada, recently. This study investigates the phylogenetic relatedness of E. coli O157:H7 from pigs, cattle, and pork-production environments for source attribution. Limited strain diversity was observed using five conventional subtyping methods, with most or all strains being in one subgroup. Whole-genome single nucleotide polymorphism analysis confirmed the recent ancestry of the isolates from all three sources. Most environmental isolates clustered closer with pig isolates than cattle isolates. Also, a direct link was observed between 2018-outbreak environmental isolates and isolates collected from a pig farm in 2018. The majority of pig isolates harbor only one Shiga toxin gene, stx2a, while 70% (35/50) of the cattle isolates have both stx1a and stx2a. The results show some E. coli O157:H7 strains could establish persistence on pig farms and as such, pigs can be a significant source of the organism.

Genome Sequences of 104 Escherichia coli O157:H7 Isolates from Pigs, Cattle, and Pork Production Environments in Alberta, Canada.

Genome sequences of Escherichia coli O157:H7 originating from pigs are limited in the public databases. We sequenced 104 E. coli O157:H7 isolates from pig and cattle feces and pork production environments in Alberta, Canada. The information will aid studies investigating sources of E. coli O157:H7 contaminating pork and the associated environments.

Escherichia coli: Physiological Clues Which Turn On the Synthesis of Antimicrobial Molecules.

Zoonotic pathogens, like Shiga toxin-producing Escherichia coli (STEC) are a food safety and health risk. To battle the increasing emergence of virulent microbes, novel mitigation strategies are needed. One strategy being considered to combat pathogens is antimicrobial compounds produced by microbes, coined microcins. However, effectors for microcin production are poorly understood, particularly in the context of complex physiological responses along the gastro-intestinal tract (GIT). Previously, we identified an E. coli competitor capable of producing a strong diffusible antimicrobial with microcin-associated characteristics. Our objective was to examine how molecule production of this competitor is affected by physiological properties associated with the GIT, namely the effects of carbon source, bile salt concentration and growth phase. Using previously described liquid- and agar-based assays determined that carbon sources do not affect antimicrobial production of E. coli O103F. However, bile salt concentrations affected production significantly, suggesting that E. coli O103F uses cues along the GIT to modulate the expression of antimicrobial production. Furthermore, E. coli O103F produces the molecule during the exponential phase, contrary to most microcins identified to date. The results underscored the importance of experimental design to identify producers of antimicrobials. To detect antimicrobials, conventional microbiological methods can be a starting point, but not the gold standard.

Whole-Genome Draft Assemblies of Difficult-to-Classify Escherichia coli O157 and Non-O157 Isolates from Feces of Canadian Feedlot Cattle.

Forty-eight Escherichia coli strains were chosen due to variable detection of stx or serogroup by PCR. Although all strains were initially determined to be Shiga toxin-producing Escherichia coli (STEC), their genomes revealed 11 isolates carrying stx 1a, stx 1b, stx 2a, and/or stx 2b Assembled genome sizes varied between 4,667,418 and 5,556,121 bp, with N 50 values between 79,648 and 294,166 bp and G+C contents between 50.3% and 51.4%.

Properties of an Antimicrobial Molecule Produced by an Escherichia coli Champion.

Over recent decades, the number and frequency of severe pathogen infections have been increasing. Pathogen mitigation strategies in human medicine or in livestock operations are vital to combat emerging arsenals of bacterial virulence and defense mechanisms. Since the emergence of antimicrobial resistance, the competitive nature of bacteria has been considered for the potential treatment or mitigation of pathogens. Previously, we identified a strong E. coli competitor with probiotic properties producing a diffusible antimicrobial molecule(s) that inhibited the growth of Shiga toxin-producing E. coli (STEC). Our current objective was to isolate and examine the properties of this antimicrobial molecule(s). Molecules were isolated by filter sterilization after 12 h incubation, and bacterial inhibition was compared to relevant controls. Isolated antimicrobial molecule(s) and controls were subjected to temperature, pH, or protease digestion treatments. Changes in inhibition properties were evaluated by comparing the incremental cell growth in the presence of treated and untreated antimicrobial molecule(s). No treatment affected the antimicrobial molecule(s) properties of STEC inhibition, suggesting that at least one molecule produced is an efficacious microcin. The molecule persistence to physiochemical and enzymatic treatments could open a wide window to technical industry-scale applications.

Bacteriocin Occurrence and Activity in Escherichia coli Isolated from Bovines and Wastewater.

The increasing prevalence of antimicrobial resistant (AMR) E. coli and related Enterobacteriaceae is a serious problem necessitating new mitigation strategies and antimicrobial agents. Bacteriocins, functionally diverse toxins produced by most microbes, have long been studied for their antimicrobial potential. Bacteriocins have once again received attention for their role as probiotic traits that could mitigate pathogen burden and AMR bacteria in livestock. Here, bacteriocins were identified by activity screening and whole-genome sequencing of bacteriocin-producers capable of inhibiting bovine and wastewater E. coli isolates enriched for resistance to cephalosporins. Producers were tested for activity against shiga toxin-producing E. coli (STEC), AMR E. coli, and related enteric pathogens. Multiple bacteriocins were found in 14 out of 90 E. coli isolates tested. Based on alignment within BACTIBASE, colicins M, B, R, Ia, Ib, S4, E1, E2, and microcins V, J25, and H47, encoded by identical, variant, or truncated genes were identified. Although some bacteriocin-producers exhibited activity against AMR and STEC E. coli in agar-based assays, most did not. Despite this idiosyncrasy, liquid co-cultures of all bacteriocinogenic isolates with luciferase-expressing generic (K12) or STEC E. coli (EDL933) resulted in inhibited growth or reduced viability. These abundant toxins may have real potential as next-generation control strategies in livestock production systems but separating the bacteriocin from its immunity gene may be necessary for such a strategy to be effective.

Comparison of heating block and water bath methods to determine heat resistance in Shiga-toxin producing Escherichia coli with and without the locus of heat resistance.

This study found variability in the time required for tubes of media in heating block wells to reach 60 °C, resulting in significant effects on heat resistance measurements. To determine the extent that methodology changed heat resistance measurements, we compared the heat resistance of Shiga-toxin producing Escherichia coli (STEC) strains with and without the locus of heat resistance (LHR) using both heating block and water bath methods. A total of 34 strains of STEC were used along with a generic E. coli which has been identified as heat-resistant and used as a positive control. The E. coli strains were incubated in a water bath and a heating block set at 60 °C to determine come up time to 60 °C (T0) and for 6 additional minutes (T6) to calculate the D60 value. After incubation, the colony forming units (CFU) were enumerated and mean log CFU/mL from biological replicates was calculated. To compare reductions from T0 to T6, standard deviations among replicates within heating method and correlation of the D60 values generated across methods were determined using Mixed model and Correlation analyses. Our findings indicate that the method chosen to evaluate heat resistance of E. coli can dramatically influence results as there was not a significant correlation between D60 values for the same isolate determined by water bath and heating block methods. The water bath method generates more reliable and consistent heat resistance data and should be used in future evaluations of heat resistance in E. coli. Moreover, PCR screening for the LHR would only be moderately useful for predicting phenotypic heat-resistance of E. coli. Considering water bath data only, LHR-positive STEC isolates were either moderately heat-resistant (1 to 5 log reduction) or heat-sensitive (> 5 log reduction). As LHR-negative STEC were also moderately heat-resistant, prediction of phenotypic heat resistance from genotype requires further refinement.

Ambient alkaline hydrolysis and anaerobic digestion as a mortality management strategy for whole poultry carcasses.

Livestock mortality management is a critical factor for ensuring biosecurity, minimizing environmental impact, and maintaining public trust in livestock production agriculture. The number of technologies currently used for livestock mortality management is small, including composting, burial, incineration, landfilling, and rendering. Each technology has advantages and disadvantages which make their suitability situational. In this study, ambient alkaline hydrolysis (AAH) using 2, 4, or 8 M potassium hydroxide at ambient temperature and pressure was explored as a disposal method for whole broiler chicken carcasses. Alkaline hydrolysate (pH > 14) resulting from the process was neutralized by mixing with acidic corn silage, and then utilized as a substrate for anaerobic digestion in bench top continuously stirred tank reactors. All AAH treatments solubilized broiler carcasses within 20 days. Corn silage neutralized 2 M hydrolysate using a 2:1 (w/w) mixing ratio, while 4 M hydrolysate required a 4:1 mixing ratio. Anaerobic digestion of neutralized hydrolysate reduced volatile solids by >96% for all treatments. Highest methane yields were observed from the 2 M hydrolysate (607.2 ±â€¯47.9 g mL-1 VS), while biogas production from the 8 M hydrolysate was totally inhibited over a total of 42 days. Ambient alkaline hydrolysis followed by silage neutralization and anaerobic digestion provides a feasible, straightforward technology to manage routine and emergency animal mortalities.

Competition among Escherichia coli Strains for Space and Resources.

Shiga toxin-producing Escherichia coli (STEC) are a subgroup of E. coli causing human diseases. Methods to control STEC in livestock and humans are limited. These and other emerging pathogens are a global concern and novel mitigation strategies are required. Habitats populated by bacteria are subjected to competition pressures due to limited space and resources but they use various strategies to compete in natural environments. Our objective was to evaluate non-pathogenic E. coli strains isolated from cattle feces for their ability to out-compete STEC. Competitive fitness of non-pathogenic E. coli against STEC were assessed in competitions using liquid, agar, and nutrient limiting assays. Winners were determined by enumeration using O-serogroup specific quantitative PCR or a semi-quantitative grading. Initial liquid competitions identified two strong non-pathogenic competitors (O103F and O26E) capable of eliminating various STEC including O157 and O111. The strain O103F was dominant across permeable physical barriers for all tested E. coli and STEC strains indicating the diffusion of antimicrobial molecules. In direct contact and even with temporal disadvantages, O103F out-competed STEC O157E. The results suggest that O103F or the diffusible molecule(s) it produces have a potential to be used as an alternative STEC mitigation strategy, either in medicine or the food industry.

Variability in Characterizing Escherichia coli from Cattle Feces: A Cautionary Tale.

Shiga toxin-producing Escherichia coli (STEC) are diverse bacteria, with seven serogroups (O26, O45, O103, O111, O121, O145, O157; "Top 7") of interest due to their predominance in human disease. Confirmation of STEC relies on a combination of culturing, immunological and molecular assays, but no single gold standard for identification exists. In this study, we compared analysis of STEC between three independent laboratories (LAB) using different methodologies. In LAB A, colonies of Top 7 were picked after serogroup-specific immunomagnetic separation of feces from western-Canadian slaughter cattle. A fraction of each colony was tested by PCR (stx1, stx2, eae, O group), and Top 7 isolates were saved as glycerol stocks (n = 689). In LAB B, a subsample of isolates (n = 171) were evaluated for stx1 and stx2 using different primer sets. For this, approximately half of the PCR were performed using original DNA template provided by LAB A and half using DNA extracted from sub-cultured isolates. All Top 7 isolates were sub-cultured by LAB A and shipped to LAB C for traditional serotyping (TS) to determine O and H groups, with PCR-confirmation of virulence genes using a third set of primers. By TS, 76% of O groups (525/689) matched PCR-determined O groups. Lowest proportions (p < 0.05) of O group matches between PCR and TS (62.6% and 69.8%) occurred for O26 and O45 serogroups, respectively. PCR-detection of stx differed most between LAB A and LAB C. Excluding isolates where O groups by PCR and TS did not match, detection of stx1 was most consistent (p < 0.01) for O111 and O157:H7/NM. In contrast, for O45 and O103, stx1 was detected in >65% of isolates by LAB A and <5% by LAB C. Stx2 was only detected by LAB C in isolates of serogroups O121, O145, and O157:H7/NM. LAB B also detected stx2 in O26 and O157:H12/H29, while LAB A detected stx2 in all serogroups. Excluding O111 and O157:H7/NM, marked changes in stx detection were observed between initial isolation and sub-cultures of the same isolate. While multiple explanations exist for discordant O-typing between PCR and TS and for differences in stx detection across labs, these data suggest that assays for STEC classification may require re-evaluation and/or standardization.

Quantitative surveillance of shiga toxins 1 and 2, Escherichia coli O178 and O157 in feces of western-Canadian slaughter cattle enumerated by droplet digital PCR with a focus on seasonality and slaughterhouse location.

Often Escherichia coli are harmless and/or beneficial bacteria inhabiting the gastrointestinal tract of livestock and humans. However, Shiga toxin-producing E. coli (STEC) have been linked to human disease. Cattle are the primary reservoir for STEC and STEC "super-shedders" are considered to be a major contributor in animal to animal transmission. Among STEC, O157:H7 is the most recognized serotype, but in recent years, non-O157 STEC have been increasingly linked to human disease. In Argentina and Germany, O178 is considered an emerging pathogen. Our objective was to compare populations of E. coli O178, O157, shiga toxin 1 and 2 in western Canadian cattle feces from a sampling pool of ~80,000 beef cattle collected at two slaughterhouses. Conventional PCR was utilized to screen 1,773 samples for presence/absence of E. coli O178. A subset of samples (n = 168) was enumerated using droplet digital PCR (ddPCR) and proportions of O178, O157 and shiga toxins 1 & 2 specific-fragments were calculated as a proportion of generic E. coli (GEC) specific-fragments. Distribution of stx1 and stx2 was determined by comparing stx1, stx2 and O157 enumerations. Conventional PCR detected the presence of O178 in 873 of 1,773 samples and ddPCR found the average proportion of O178, O157, stx1 and stx2 in the samples 2.8%, 0.6%, 1.4% and 0.5%, respectively. Quantification of stx1 and stx2 revealed more virulence genes than could be exclusively attributed to O157. Our results confirmed the presence of E. coli O178 in western Canadian cattle and ddPCR revealed O178 as a greater proportion of GEC than was O157. Our results suggests: I) O178 may be an emerging subgroup in Canada and II) monitoring virulence genes may be a more relevant target for food-safety STEC surveillance compared to current serogroup screening.

Air-Dried Brown Seaweed, Ascophyllum nodosum, Alters the Rumen Microbiome in a Manner That Changes Rumen Fermentation Profiles and Lowers the Prevalence of Foodborne Pathogens.

The use of Tasco (air-dried Ascophyllum nodosum) as a feed supplement for ruminants has been reported to affect rumen fermentation and reduce Escherichia coli O157:H7 shedding in feces, but the mode of action behind this phenomenon is unclear. In this study, the effects of four Tasco levels (0, 1, 3, and 5%) on rumen microbiota and rumen/fecal E. coli O serogroups in rams were investigated. Rumen total bacteria and archaea were linearly reduced (P < 0.001) and protozoa were linearly increased (P < 0.001) by increasing levels of Tasco. The relative abundances of seven bacterial species and one protozoal species differed among Tasco levels. With Tasco, 14 predicted metabolic pathways were enriched while only 3 were suppressed. A lower ruminal butyrate concentration is possibly associated with enrichment of the "butanoate metabolism" pathway in Tasco-fed rams. The ruminal total E. coli population was linearly reduced (P < 0.001) by Tasco. Supplementation with Tasco only completely eliminated O121 in the rumen and feces, and higher levels of Tasco (3 and 5%) reduced fecal shedding of serogroups O45, O103, and O111 even though these serogroups were present in the rumen. Our results suggest that Tasco effectively reduced pathogenic E. coli but had only minimal impacts on rumen fermentation in rams. IMPORTANCE Maintaining product safety and reducing the carbon footprint of production are two sustainability goals of the livestock industry. The objective of this study was to study the impact of Tasco, a product derived from the brown macroalga Ascophyllum nodosum, on the rumen microbiome and its function. The inclusion of Tasco altered both rumen and fecal microbiota levels without affecting rumen fermentation. Tasco reduced fecal Escherichia coli populations and specifically reduced the prevalence of Shiga toxin-producing E. coli O45, O103, O111, and O121 in feces. The findings of this study highlight the application of Tasco as a potential feed additive to reduce pathogen shedding in rams without interfering with ruminal metabolism.

Dissipation of antimicrobial resistance genes in compost originating from cattle manure after direct oral administration or post-excretion fortification of antimicrobials.

Dissipation of antimicrobial resistance genes (ARG) during composting of cattle manure generated through fortification versus administration of antimicrobials in feed was compared. Manure was collected from cattle fed diets containing (kg-1) dry matter (DM): (1) 44 mg chlortetracycline (CTC), (2) a mixture of 44 mg each of chlortetracycline and sulfamethazine (CTCSMZ), (3) 11 mg tylosin (TYL) or (4) Control, no antimicrobials. Manures were composted for 30 d with a single mixing after 16 d to generate the second heating cycle. Quantitative PCR (qPCR) was used to measure 16S rDNA and tetracycline (tet), erythromycin (erm) and sulfamethazine (sul) genes. Temperature peaks ranged from 48 to 68°C across treatments in the first composting cycle, but except for the control, did not exceed 55°C in the second cycle. Copy numbers of 16S rDNA decreased (P < 0.05) during composting, but were not altered by antimcrobials. Except tet(L), all ARG decreased by 0.1-1.6 log10 g DM-1 in the first cycle, but some genes (tet[B], tet[L], erm[F], erm[X]) increased (P < 0.05) by 1.0-3.1 log10 g DM-1 in the second. During composting, levels of tet(M) and tet(W) in CTC, erm(A), erm(B) and erm(X) in TYL, and sul(1) in CTCSMZ remained higher (P < 0.05) in fed than fortified treatments. The dissipation of ARG during composting of manure fortified with antimicrobials differs from manure generated by cattle that are administered antimicrobials in feed, and does not always align with the dissipation of antimicrobial residues.

Effect of severe weather events on the shedding of Shiga toxigenic Escherichia coli in slaughter cattle and phenotype of serogroup O157 isolates.

High-event periods (HEPs) occur sporadically when beef carcasses and meat have episodes of acute contamination with Shiga toxin-producing Escherichia coli (STEC). In this study, severe weather events were investigated as catalysts for HEPs based on PCR and isolate prevalence of seven E. coli serogroups in slaughter cattle feces. Winter ambient temperatures with daily means 10.5oC warmer or 12.3°C colder than seasonal norms (-10.4°C) most altered STEC shedding. Fecal samples yielded increased proportions (P < 0.05) of O26 and O157 isolates during winter warm periods, and reduced (P < 0.05) O45 isolates during cold periods compared to samplings during seasonal norms. Based on changing PCR prevalence and isolates collected, O157 was the serogroup most responsive to severe weather events. Consequently, O157 isolates (n = 219) were evaluated for heat resistance, biofilm-forming potential and virulence gene subtypes. Two isolates had heat-resistant phenotypes with thermal death time at 60°C (D60) > 10 min and one also had strong biofilm-forming potential. However, this isolate lacked eae and stx genes. Severe weather can influence STEC shedding, particularly of O157, and could possibly trigger HEPs. However, our data suggest that it is unlikely for isolates to carry virulence genes and possess phenotypes capable of evading post-harvest microbiological interventions.

Subtyping Escherichia coli Virulence Genes Isolated from Feces of Beef Cattle and Clinical Cases in Alberta.

Clinical outcomes of Shiga toxin (stx)-producing Escherichia coli infection are largely determined by virulence gene subtypes. This study used a polymerase chain reaction (PCR)-pyrosequencing assay to analyze single-nucleotide polymorphisms for subtyping three major virulence genes (stx1, stx2, eae) of pathogenic E. coli (O157, O26, O111, and O103) isolated from cattle over a 2-year interval (n = 465) and human clinical cases (n = 42) in western Canada. Most bovine isolates were PCR positive for at least one target virulence gene (367/465), whereas 100% of human isolates harbored eae in combination with at least one stx gene. Four Shiga toxin (1a, 2a, 2c, and 2e) and four eae (λ/γ1-eae, ɛ-eae, θ/γ2-eae, and ß-eae) subtypes were identified in over 25 distinct virulence genotypes. Among cattle isolates, every serogroup, but O103, presented a dominant genotype (O157: stx1a+stx2a+λ/γ1-eae, O26: ß-eae alone, and O111: stx1a+θ/γ2-eae). Similar patterns were found in human isolates, although it was not possible to establish a clear genotypic association between the two sources. Many O157 and non-O157 cattle isolates lacked stx genes; the absence was greater in non-O157 (75/258) and O157:non-H7 (19/40) than in O157:H7 strains (1/164). In addition, there was a greater diversity of virulence genotypes of E. coli isolated from cattle than those of human diseases, which could be due to sample characteristics (e.g., source and clinical condition). However, the majority of cattle strains had virulence profiles identical to those of clinical cases. Consequently, determining the presence of certain stx (stx1a and stx2a) and eae (λ/γ1-eae) subtypes known to cause human disease would be a valuable tool for risk assessment and prediction of disease outcome along the farm-to-fork continuum.

Monitoring Seven Potentially Pathogenic Escherichia coli Serogroups in a Closed Herd of Beef Cattle from Weaning to Finishing Phases.

The goal of this study was to monitor Shiga toxin-producing Escherichia coli (STEC) serogroups and virulence genes in cattle (n = 30) originating from a closed herd. Fecal samples were collected (1) at weaning, (2) upon arrival to a feedlot, (3) after 30 days on feed (DOF), and (4) after 135 DOF. DNA was extracted from feces for detection of virulence and serogroup genes by polymerase chain reaction (PCR) and immunomagnetic separation and pulsed-field gel electrophoresis (PFGE) were performed to collect and subtype STEC isolates. The prevalence of each serogroup measured by PCR from weaning to 135 DOF was 23.3-80.0% for O26, 33.3-46.7% for O45, 70.0-73.3% for O103, 36.7-86.7% for O111, 56.7-6.7% for O121, 26.7-66.7% for O145, and 66.7-90.0% for O157. Total fecal samples positive for virulence genes were 87.5% for ehxA, 85.8% for stx1, 60.0% for stx2, 52.5% for eae, and 44.2% for the autoagglutinating adhesion gene, saa. The prevalence of each serogroup and virulence gene tended to increase by 135 DOF, with the exception of O121, stx2, and saa. The frequency of detection of some virulence genes was largely affected over time, most notably with saa and stx2 decreasing, and eae increasing when cattle were transitioned to concentrate-based diets. PFGE analysis of O157 and O103 fecal isolates revealed dominant pulsotypes, but the presence of identical O103 isolates, which differed in virulence profiles. Overall, this study showed that fecal shedding of E. coli serogroups and virulence-associated genes are highly variable over time as cattle move from ranch to feedlot. To mitigate STEC, it is important to understand the factors affecting both prevalence of individual serogroups and the presence of virulence factors.

Influence of Season and Feedlot Location on Prevalence and Virulence Factors of Seven Serogroups of Escherichia coli in Feces of Western-Canadian Slaughter Cattle.

Pooled feces collected over two years from 1749 transport trailers hauling western-Canadian slaughter cattle were analysed by PCR for detection of Escherichia coli serogroups O26, O45, O103, O111, O121, O145, and O157. Sequential immunomagnetic separation was then used to collect bacterial isolates (n = 1035) from feces positive for target serogroups. Isolated bacteria were tested by PCR to confirm serogroup and the presence of eae, ehxA, stx1, and stx2 virulence genes. Based on PCR screening, serogroup prevalence in feces ranged from 7.0% (O145) to 94.4% (O103) with at least 3 serogroups present in 79.5% of samples. Origin of cattle affected serogroup PCR prevalence and O157 was most prevalent in feces from south-west Alberta (P < 0.001). All serogroups demonstrated seasonal variations in PCR prevalence, with O26, O45, O103, O121, and O157 least prevalent (P < 0.001) in cooler winter months, while uncommon serogroups O111 and O145 increased in prevalence during winter (P < 0.001). However, isolates collected during winter were predominantly from serogroups O103 and O45. No seasonal variation was noted in proportion of isolates which were Shiga toxin containing E. coli (STEC; P = 0.18) or positive for Shiga toxin and eae (enterohemorrhagic E. coli; EHEC; P = 0.29). Isolates of serogroups O111, O145, and O157 were more frequently EHEC than were others, although 37.6-54.3% of isolates from other serogroups were also EHEC. Shiga-toxin genes present also varied by geographic origin of cattle (P < 0.05) in all serogroups except O157. As cattle within feedlots are sourced from multiple regions, locational differences in serogroup prevalence and virulence genes imply existence of selection pressures for E. coli and their virulence in western-Canadian cattle. Factors which reduce carriage or expression of virulence genes, particularly in non-O157 serogroups, should be investigated.

Inactivation of Bacillus anthracis Spores during Laboratory-Scale Composting of Feedlot Cattle Manure.

Anthrax outbreaks in livestock have social, economic and health implications, altering farmer's livelihoods, impacting trade and posing a zoonotic risk. Our study investigated the survival of Bacillus thuringiensis and B. anthracis spores sporulated at 15, 20, or 37°C, over 33 days of composting. Spores (∼7.5 log10 CFU g(-1)) were mixed with manure and composted in laboratory scale composters. After 15 days, the compost was mixed and returned to the composter for a second cycle. Temperatures peaked at 71°C on day 2 and remained ≥55°C for an average of 7 days in the first cycle, but did not exceed 55°C in the second. For B. thuringiensis, spores generated at 15 and 21°C exhibited reduced (P < 0.05) viability of 2.7 and 2.6 log10 CFU g(-1) respectively, as compared to a 0.6 log10 CFU g(-1) reduction for those generated at 37°C. For B. anthracis, sporulation temperature did not impact spore survival as there was a 2.5, 2.2, and 2.8 log10 CFU g(-1) reduction after composting for spores generated at 15, 21, and 37°C, respectively. For both species, spore viability declined more rapidly (P < 0.05) in the first as compared to the second composting cycle. Our findings suggest that the duration of thermophilic exposure (≥55°C) is the main factor influencing survival of B. anthracis spores in compost. As sporulation temperature did not influence survival of B. anthracis, composting may lower the viability of spores associated with carcasses infected with B. anthracis over a range of sporulation temperatures.