Development of antibiotic resistance in foodborne pathogens, Salmonella spp. and Campylobacter, is a public health concern. Public demand to reduce the use of sub-therapeutic antibiotic growth promoters (AGP) in poultry feeding has resulted in greater adoption of antibiotic-free poultry production systems. There is a need to understand the effects of AGP removal from poultry feed on gut microbiota and its impact on prevalence of foodborne pathogens. The effect of antibiotic withdrawal from poultry feed on gut microbial community, host performance and immunity, and prevalence of Salmonella and Campylobacter was evaluated. Birds were raised on three phase diets (starter [d0-22], grower [d23-35] and finisher [d36-42]) with and without bacitracin dimethyl salicyclate (BMD). At early growth stage, bird performance was improved (P ≤ 0.05) with BMD treatment, whereas performance was better (P ≤ 0.05) in control group (no BMD in the feed) at the time of commercial processing. Acetate and butyrate production was affected (P ≤ 0.05) by age, whereas propionate production was affected (P ≤ 0.05) by both the treatment and age. The bacterial communities in the cecum were more diverse (P ≤ 0.001) and rich compared to the ileal communities, and they shifted in parallel to one another as the chicks matured. Differences in diversity and species richness were not observed (P > 0.05) between the BMD-fed and control groups. Comparing all ages, treatments and diets, the composition of cecal and ileal bacterial communities was different (P ≤ 0.001). Inclusion of BMD in the feed did not affect the bacterial phyla. However, predictable shift in the ileal and cecal bacterial population at lower taxonomic level was observed in control vs BMD-fed group. Cytokines gene expression (IL-10, IL-4, IFN-γ, beta-defensin, and TLR-4) was affected (P≤ 0.05) in the BMD-fed group at early stages of growth. The prevalence of foodborne pathogens, Campylobacter spp. and Salmonella spp. showed higher abundance in the ilea of BMD-fed chicks compared to control group. Overall, this study provided insight of the impact of AGP supplementation in the feed on gut microbial modulations, bird performance, host immunity and pathogen prevalence. This information can assist in designing alternative strategies to replace antibiotics in modern poultry production and for food safety.

Background: Despite a strong association between Salmonella isolation and slaughter hygiene, as measured by the Enterobacteriaceae levels on pre-chill carcass surfaces, a high variation in this association was observed between sampling days within the same slaughterhouse. It was hypothesised that in a scenario of high exposure on the farm, batches with a high prevalence of carrier pigs shedding a high number of Salmonella may enhance the risk of contamination on some slaughter days. Thus, the aim of this study was to assess the profile of Salmonella carried in the intestinal contents of slaughter pigs.Materials, Methods & Results: Ten pig batches slaughtered in a slaughterhouse were investigated for the presence of Salmonella. From each pig, the following samples were taken: i. blood collected at bleeding; ii. sponges rubbed on the carcass surface after bleeding and before chilling; iii. fragment of the ileocecal region of the intestine. Serum samples were subjected to a ELISA-Typhimurium test. Sponges were investigated for the presence of Salmonella and total aerobic mesophilic (TAM) and Enterobacteriaceae (EC) bacterial counts. Salmonella was enumerated in the intestinal contents. Selected Salmonella strains were subjected to an antimicrobial resistance disk diffusion test, macro-restriction with Xba-I (PFGE) and whole genome sequencing (WGS). From the 50 sampled pigs, 96% were positive in the ELISA-Typhimurium test and 64% were Salmonella-positive in the intestinal contents. The amount of Salmonella in the intestinal content samples was highly variable, and the mean log of fitted distributions of Salmonella in the batch ranged from -2.97 to 2.25 cfu.g-1. The slaughter process achieved a logarithmic reduction, ranging from 0.64 to 2.35 log cfu.cm-2 for TAM and from 0.55 to 2.57 log cfu.cm-2 for EC. Salmonella was isolated from 16% of the carcasses after bleeding; this frequency decreased to 8% at the pre-chill step. All positive pre-chill carcasses originated from pigs carrying Salmonella in the intestinal content and from batches with a high number of carrier pigs. Salmonella Typhimurium and its monophasic variant were the most frequent in the intestinal contents and carcasses. Resistance was detected against ampicillin (42.5%), tetracycline (42.5%), sulfonamide (40%), gentamicin (25%) and ciprofloxacin (12.5%). Regarding colistin, 85% of the tested strains were classified as non-susceptible. The monophasic variant S. Typhimurium strains subjected to PFGE and WGS presented different profiles; several antimicrobial resistance genes were identified and all belonged to ST-19.Discussion: In this study, almost all sampled pigs entering the slaughter line had been exposed to Salmonella on the farm and a high number were carrying Salmonella in their guts. While the three batches with Salmonella-positive carcasses at the pre-chill step presented TAM media that was not significantly different from the other batches, there was a higher number of positive pigs carrying Salmonella in their intestinal contents. Moreover, the batch with the highest number of positive carcasses also presented the highest Salmonella mean count in their intestinal contents. The profile of Salmonella carried in the intestinal content of slaughter pigs proved to be highly variable in terms of the frequency, number of bacteria, serovars, antimicrobial resistance, and genotypes. Results indicate that the day-to-day variability in the prevalence and number of Salmonella in the intestinal contents of slaughter batches is likely to influence the frequency of contaminated pre-chill carcasses. Salmonella Typhimurium isolated from the intestinal contents of slaughter pigs may belong to genotypes involved in human disease and may carry several antimicrobial resistance genes. These aspects should be taken into account when planning Salmonella control in swine.

The aim of this study was to investigate the presence of shiga toxin-producing Escherichia coli (STEC) and atypical enteropathogenic Escherichia coli (aEPEC) in frozen chicken carcasses sold at stores in southern Brazil. Typical E. coli colonies were enumerated in 246 chicken carcasses, and the presence of stx1, stx2, eae genes was investigated in their rinse liquid and in E. coli strains isolated from those carcasses.Strains of E. coli were also investigated for the presence of bfp gene. A median of 0.6 cfu.g -1 (ranging from <0.1 to 242.7 cfu.g -1 ) of typical E. coli colonies was found in the carcasses. Shiga toxin-encoding genes (stx1 and stx2) were not detected, indicating that the chicken carcasses were negative for STEC. The intimin protein gene (eae) was detected in E.coli isolated from 4.88% of the carcasses; all tested strains were negative for the bfp gene and were classified as aEPEC. Twenty-two aEPEC strains were tested for resistance to ten antimicrobials and subjected to macrorestriction (PFGE). All the tested aEPEC strains were fully susceptible to cephalosporins, ciprofloxacin and colistin. Resistance to sulfonamide (65%), ampicillin (55%), tetracycline (50%) and gentamicin (45%) were the most frequent. The PFGE profile demonstrated a low level of similarity among the resistant strains, indicating that they were epidemiologically unrelated. The results indicate that aEPEC strains can contaminate chicken meat, and their association with strains implicated in human diarrhea needs to be further investigated.

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The increasing antimicrobial resistance observed worldwide in bacteria isolated from human and animals is a matter of extreme concern and has led to the monitoring of antimicrobial resistance in pathogenic and commensal bacteria. The aim of this study was to evaluate the antimicrobial resistance profile of Escherichia coli isolated from pig carcasses and to assess the occurrence of relevant resistance genes. A total of 319 E. coli isolates were tested for antimicrobial susceptibility against different antimicrobial agents. Moreover, the presence of extended-spectrum β-lactamase (ESBL) and inducible ampC-β-lactamase producers was investigated. Eighteen multi-resistant strains were chosen for resistance gene detection and PFGE characterization. The study showed that resistance to antimicrobials is widespread in E. coli isolated from pig carcasses, since 86.2% of the strains were resistant to at least one antimicrobial and 71.5% displayed multi-resistance profiles. No ampC-producing isolates were detected and only one ESBL-producing E. coli was identified. Genes strA (n=15), floR (n=14), aac(3)IVa (n=13), tetB (n=13), sul2 (n=12), tetA (n=11), aph(3)Ia (n=8) and sul3 (n=5) were detected by PCR. PFGE analysis of these multi-resistant E. coli strains showed less than 80% similarity among them. We conclude that antimicrobial multi-resistant E. coli strains are common on pig carcasses and present highly diverse genotypes and resistance phenotypes and genotypes.

Ozone application has been suggested as an additional measure to the slaughter animals under hygiene programs. In this study, we determined the efficacy of gaseous ozone applied to pig carcasses during chilling (16 h at 2–5°C). Forty carcasses were allocated to each treatment: control, without ozone application (T1) and 5 ppm gaseous ozone application (T2), divided in two 4-h periods. The carcasses were sampled before and after chilling. The average counts of total aerobic mesophilic (TAM) bacteria before chilling were not different (p = 0.55) between T1 and T2. In turn, after chilling, the ozone-treated carcasses had significantly reduced about 0.4 colony-forming units (CFU)/cm2 of TAM counts (p < 0.001) than the control carcasses. No significant reduction was observed in the number of carcasses positive for Listeria sp. and Escherichia coli after gaseous ozone treatment; while a tendency (p = 0.08) of lower number of Salmonella positive carcasses in T2 was observed. Common macrorestriction (pulsed-field gel electrophoresis) patterns of S. enterica were observed in the carcasses before and after chilling. Pork samples from treated and untreated carcasses with ozone showed no lipid oxidation or altered color and pH. The results indicate that the gaseous ozone in the tested protocol is effective in reducing TAM populations, but not effective in decreasing the number of carcasses positive for E. coli and Listeria sp. Regarding Salmonella, the tendency of positive carcasses reduction may encourage further studies by testing other protocols of gaseous ozone application inside the chilling chamber.

Pork salami is an embedded, cured and ripened product commonly consumed in Brazil, and the presence of Salmonella enterica has already been reported in this product. During its preparation, the microbiological safety depends on the meat quality, addition of ingredients with antimicrobial activity, hygiene during processing, pH and water activity (Aw) reduction during maturation. In Brazil, the maturation protocol has not been determined in food regulation; therefore, the objectives of this study were (a) to identify the fermentation and drying phases during salami maturation; (b) to test the survival of S. enterica during salami processing; and (c) to compare xylose lysine deoxycholate (XLD) and thin agar layer (TAL) agar for recovering Salmonella. The salami samples were prepared with a cocktail of S. enterica strains, fermented at 30°C and dried at 20°C with controlled relative humidity (RH). Periodic sampling for S. enterica quantification and Aw and pH analyses were performed during maturation, and curves were constructed. Fermentation occurred during the first 66 hours, and the pH decreased while the population of S. enterica increased over the first 21 hours. The drying step was able to reduce the bacterial population by approximately 5 log CFU after 875 hours, reaching an Aw of less than 0.78. However, elimination of S. enterica was not achieved. For Salmonella recovery, TAL agar was more efficient than XLD agar.

Salami is a ready-to-eat (RTE) product frequently purchased at street fairs in Porto Alegre. Salmonella enterica, Listeria monocytogenes, and coagulase-positive Staphylococcus (CPS) are important causes of foodborne disease and can be transmitted through the consumption of RTE foods. The aim of this study was to evaluate the presence of these pathogens in salami sold at street fairs. Ninety salami samples from three commercial brands available at street fairs were analyzed by routine bacteriological methods for Salmonella spp. and Listeria spp., as well as enumeration of CPS. In addition, two samples from each commercial brand were analyzed for water activity (aw). Samples of brand A showed aw values (0.938 and 0.942) above those set by the legislation, while brand B (0.849 and 0.860) and brand C (0.826 and 0.854) were compliant. Microbiological analyses showed that 67.7% were negative to all investigated bacteria. Salmonella Typhimurium was isolated from 4.4% (4/90) of salami samples, all from commercial brand A. Listeria monocytogenes was detected in 3.3% (3/90) of samples, from commercial brands B and C. Moreover, 7.7% (7/90) of samples contained CPS populations non-compliant with legislation. Although the great majority of salami sold at street fairs of Porto Alegre was compliant with standards, S. enterica, L. monocytogenes, and CPS ≥ 5 × 103 cfu.g-1 could be found in this RTE product. Therefore, control measures in the processing industry and consumer’s education about foodborne illness prevention should be maintained.