Locus of Heat Resistance (LHR) in Meat-Borne \u3ci\u3eEscherichia coli\u3c/i\u3e: Screening and Genetic Characterization

Microbial resistance to processing treatments poses a food safety concern, as treatment tolerant pathogens can emerge. Occasional foodborne outbreaks caused by pathogenic Escherichia coli have led to human and economic losses. Therefore, this study screened for the extreme heat resistance (XHR) phenotype as well as one known genetic marker, the locus of heat resistance (LHR), in 4,123 E. coli isolates from diverse meat animals at different processing stages. The prevalences of XHR and LHR among the meat-borne E. coli were found to be 10.3% and 11.4%, respectively, with 19% agreement between the two. Finished meat products showed the highest LHR prevalence (24.3%) compared to other processing stages (0 to 0.6%). None of the LHR1 E. coli in this study would be considered pathogens based on screening for virulence genes. Four high-quality genomes were generated by whole-genome sequencing of representative LHR+ isolates. Nine horizontally acquired LHRs were identified and characterized, four plasmid-borne and five chromosomal. Nine newly identified LHRs belong to ClpK1 LHR or ClpK2 LHR variants sharing 61 to 68% nucleotide sequence identity, while one LHR appears to be a hybrid. Our observations suggest positive correlation between the number of LHR regions present in isolates and the extent of heat resistance. The isolate exhibiting the highest degree of heat resistance possessed four LHRs belonging to three different variant groups. Maintenance of as many as four LHRs in a single genome emphasizes the benefits of the LHR in bacterial physiology and stress response. IMPORTANCE Currently, a “multiple-hurdle” approach based on a combination of different antimicrobial interventions, including heat, is being utilized during meat processing to control the burden of spoilage and pathogenic bacteria. Our recent study (M. Guragain, G. E. Smith, D. A. King, and J. M. Bosilevac, J Food Prot 83:1438–1443, 2020, https://doi.org/10.4315/JFP-20-103) suggests that U.S. beef cattle harbor Escherichia coli that possess the locus of heat resistance (LHR). LHR seemingly contributes to the global stress tolerance in bacteria and hence poses a food safety concern. Therefore, it is important to understand the distribution of the LHRs among meat-borne bacteria identified at different stages of different meat processing systems. Complete genome sequencing and comparative analysis of selected heat-resistant bacteria provide a clearer understanding of stress and heat resistance mechanisms. Further, sequencing data may offer a platform to gain further insights into the genetic background that provides optimal bacterial tolerance against heat and other processing treatments

Determining the Impact of Probicon L28 and BioPlus 2B on Finishing Pig Growth Performance and Carcass Characteristics

These data represent the growth performance of pigs enrolled in a study to determine the impact of two direct fed microbial products on Salmonella and Escherichia coli prevalence pre- and post-harvest. A total of 650 finishing pigs in two groups were randomly assigned to pen via a completely randomized design, and pens were assigned to one of three treatments: 1) a control treatment with pigs fed a standard corn-soybean meal finishing diet (with no added probiotic); 2) the control diets with Probicon L28 (NexGen Innovations, LLC, Lubbock, TX) supplemented through water lines using a water medicator system at a target concentration of 1.0 × 106 CFU/head/day; and 3) the control diet with added BioPlus 2B (5.0 × 108 CFU/lb of feed; ~3.0 × 109 CFU/ head/day; CHR Hansen, Inc, Milwaukee, WI). No evidence of difference (P \u3e 0.10) between treatments was observed for overall ADG, ADFI, or F/G or any of the carcass traits. However, there was a tendency for a treatment effect for loin depth (P = 0.070). Pigs fed the BioPlus 2B treatment had numerically greater loin depth compared to other treatments, but there were no significant pairwise differences between treatments (P \u3e 0.05). The results of this study suggested that probiotics used in this study and supplied through the water or feed had no impact on growth or carcass characteristics of finishing pigs

Exploring the Use of Probicon L28 and BIOPLUS 2B as Direct-Fed Microbials to Reduce Salmonella and Shiga Toxin-Producing Escherichia coli in Market Pigs

Pigs are hosts for Salmonella and Shiga toxin-producing Escherichia coli (STEC) and these pathogens can commonly be isolated from the pig farm environment. Pigs can carry pathogens to the abattoir and contaminate pork products, posing a risk to public health. Identifying an intervention that effectively reduces pathogens in commercial pigs before harvest is imperative. Due to the need for effective pre-harvest interventions in the pig industry, the objective of this study was to investigate BIOPLUS 2B (Bacillus licheniformis and Bacillus subtilis) and Probicon L28 (Lactobacillus salivarius L28) as pre-harvest interventions to reduce Salmonella and STEC in commercial growing-finishing pigs. Two groups of pigs (group 1, N = 294; group 2, N = 356, initial body weight = 106.6 lb) were fed a standard corn-soybean meal (SBM) finishing diet according to the following treatments: Probicon L28 supplementation through water lines at 1.0 × 106 CFU/head/day (Probicon); BIOPLUS 2B supplemented at 3.0 × 109 CFU/head/day (BIOPLUS 2B); and a control with no added probiotic (Control). With each group of pigs, 12 pens were used per treatment (N = 24 total), for a total of 36 pens per group (N = 72 pens total). Each group was sampled upon arrival/baseline, midway through the grow-finish phase/6 weeks post-placement, and prior to loadout/13 weeks post-placement to collect fecal samples (4 pigs/pen), boot covers (2/pen), and ropes (1/pen). Market pigs were followed to the abattoir and superficial inguinal lymph nodes (SILNs) were collected. All samples were analyzed for STEC (stx, eae genes, and O157:H7, and O26, O111, O121 O45, O103, and O145 serogroups) and Salmonella using the BAX System (real-time polymerase chain reaction). Overall, Salmonella and O111 prevalence was very low for all sample types, and Escherichia coli O157:H7 was not detected in any samples throughout the study. When compared to the control, there was no evidence (P \u3e 0.05) that BIOPLUS 2B and Probicon L28 impacted the prevalence of STEC (stx and eae genes) or serogroups O26, O121, O45, O103, and O145 in feces, boot covers, ropes, and SILNs of market pigs

EFSA BIOHAZ Panel (EFSA Panel on Biologicial Hazards), 2013. Scientific Opinion on the public health hazards to be covered by inspection of meat (solipeds)

A risk ranking process identified Trichinella spp. as the most relevant biological hazard in the context of meat inspection of domestic solipeds. Without a full and reliable soliped traceability system, it is considered that either testing all slaughtered solipeds for Trichinella spp., or inactivation meat treatments (heat or irradiation) should be used to maintain the current level of safety. With regard to general aspects of current meat inspection practices, the use of manual techniques during current post-mortem soliped meat inspection may increase microbial cross-contamination, and is considered to have a detrimental effect on the microbiological status of soliped carcass meat. Therefore, the use of visual-only inspection is suggested for “non-suspect” solipeds. For chemical hazards, phenylbutazone and cadmium were ranked as being of high potential concern. Monitoring programmes for chemical hazards should be more flexible and based on the risk of occurrence, taking into account Food Chain Information (FCI), covering the specific on-farm environmental conditions and individual animal treatments, and the ranking of chemical substances, which should be regularly updated and include new hazards. Sampling, testing and intervention protocols for chemical hazards should be better integrated and should focus particularly on cadmium, phenylbutazone and priority “essential substances” approved for treatment of equine animals. Implementation and enforcement of a more robust and reliable identification system throughout the European Union is needed to improve traceability of domestic solipeds. Meat inspection is recognised as a valuable tool for surveillance and monitoring of animal health and welfare conditions. If visual only post-mortem inspection is implemented for routine slaughter, a reduction in the detection of strangles and mild cases of rhodococcosis would occur. However, this was considered unlikely to affect the overall surveillance of both diseases. Improvement of FCI and traceability were considered as not having a negative effect on animal health and welfare surveillance

Scientific Opinion on the public health hazards to be covered by inspection of meat from farmed game

Salmonella spp. in farmed wild boar and Toxoplasma gondii in farmed deer and farmed wild boar were ranked as a high priority for meat inspection. Trichinella spp. in wild boar was ranked as low priority due to current controls, which should be continued. For chemical hazards, all substances were ranked as medium or lower potential concern. More effective control of biological hazards could be achieved using an integrated farm to chilled carcass approach, including improved food chain information (FCI) and risk-based controls. Further studies are required on Salmonella spp. in farmed wild boar and T. gondii in farmed wild boar and farmed deer. If new information confirms a high risk to public health from meat from these species, setting targets at carcass level should be considered. Palpation and incision should be omitted, as it will not detect biological hazards considered to be a high priority for meat inspection while increasing the potential spread and cross-contamination of the carcasses with Salmonella. Palpation and/or incision may be applied where abnormalities have been detected but away from the slaughter line. However the elimination of routine palpation and incision would be detrimental for detecting tuberculosis. As farmed deer and farmed wild boar can act as tuberculosis reservoirs, any reduction in the detection, due to changes in the post-mortem inspection procedures, will have consequences for the overall surveillance of tuberculosis. Monitoring programmes for chemical hazards should be more flexible and based on the risk of occurrence, taking into account FCI, which should be expanded to reflect the specific environmental conditions of the farms where the animals are reared, and the ranking of chemical substances, which should be regularly updated and include new hazards. Control programmes across the food chain, national residue control programmes, feed control and monitoring of environmental contaminants should be better integrated

EFSA BIOHAZ Panel (EFSA Panel on Biological Hazards), 2013. Scientific Opinion on the public health hazards to be covered by inspection of meat (bovine animals).

A risk ranking process identified Salmonella spp. and pathogenic verocytotoxin-producing Escherichia coli (VTEC) as current high-priority biological hazards for meat inspection of bovine animals. As these hazards are not detected by traditional meat inspection, a meat safety assurance system for the farm-to-chilled carcass continuum using a risk-based approach was proposed. Key elements of the system are risk-categorisation of slaughter animals for high-priority biological hazards based on improved food chain information, as well as risk-categorisation of slaughterhouses according to their capability to control those hazards. Omission of palpation and incision during post-mortem inspection for animals subjected to routine slaughter may decrease spreading and cross-contamination with the high-priority biological hazards. For chemical hazards, dioxins and dioxin-like polychlorinated biphenyls were ranked as being of high potential concern; all other substances were ranked as of medium or lower concern. Monitoring programmes for chemical hazards should be more flexible and based on the risk of occurrence, taking into account the completeness and quality of the food chain information supplied and the ranking of chemical substances, which should be regularly updated to include new hazards. Control programmes across the food chain, national residue control programmes, feed control and monitoring of environmental contaminants should be better integrated. Meat inspection is a valuable tool for surveillance and monitoring of animal health and welfare conditions. Omission of palpation and incision would reduce detection effectiveness for bovine tuberculosis and would have a negative impact on the overall surveillance system especially in officially tuberculosis free countries. The detection effectiveness for bovine cysticercosis, already low with the current meat inspection system, would result in a further decrease, if palpation and incision are removed. Extended use of food chain information could compensate for some, but not all, the information on animal health and welfare lost if only visual post-mortem inspection is applied

Enumeration of \u3ci\u3eSalmonella\u3c/i\u3e from Poultry Carcass Rinses \u3ci\u3evia\u3c/i\u3e Direct Plating Methods

Aim: To evaluate direct plating methods for the estimation of Salmonella load in poultry carcass rinses. Methods and Results: Two direct plating tools, the spiral plate count method (SPCM) and the hydrophobic grid membrane filtration (HGMF) method, were adapted to support quantification of Salmonella during poultry processing. Test samples consisted of 180 broiler carcasses from a commercial abattoir, 60 from each of three points in the processing line [pre-inside–outside bird wash (pre- IOBW), prechill and postchill]. The SPCM was used to estimate Salmonella load in pre-IOBW rinses, while HGMF was used to estimate Salmonella levels in prechill and postchill rinses. Carcass rinses were also evaluated for Salmonella prevalence by enrichment methods. Mean prevalences of Salmonella were 95%, 100% and 41·7%, and the geometric mean loads were 3·7 × 101, 5·6 × 100 and 5·0 × 10-2 CFU ml-1 for pre-IOBW, prechill and postchill rinses, respectively. Conclusions: The methods described are useful for estimating the concentration of viable and typical Salmonella in poultry carcass rinses. Significance and Impact of the Study: Direct plating enumeration methods can facilitate the monitoring of Salmonella load on poultry carcasses throughout the production process, and the evaluation of new processing intervention strategies