The Use of Multiplex Real-Time PCR for the Simultaneous Detection of Foodborne Bacterial Pathogens.

Foodborne pathogens continue to be a major health concern worldwide. Culture-dependent methodologies are still considered the gold standard to perform pathogen detection and quantification. These methods present several drawbacks, such as being time-consuming and labor intensive. The implementation of real-time PCR has allowed to overcome these limitations, and even reduce the cost associated with the analyses, due to the possibility of simultaneously and accurately detecting several pathogens in one single assay, with results comparable to those obtained by classical approaches. In this chapter, a protocol for the simultaneous detection of two of the most important foodborne pathogens, Salmonella spp. and Listeria monocytogenes, is described.

Application of MinION Long-Read Sequencer for Semi-targeted Detection of Foodborne Pathogens.

Foodborne pathogens remain a serious health issue in developed and developing countries. Safeness of food products has been assured for years with culture-based microbiological methods; however, these present several limitations such as turnaround time and extensive hands-on work, which have been typically address taking advantage of DNA-based methods such as real-time PCR (qPCR). These, and other similar techniques, are targeted assays, meaning that they are directed for the specific detection of one specific microbe. Even though reliable, this approach suffers from an important limitation that unless specific assays are design for every single pathogen potentially present, foods may be considered erroneously safe. To address this problem, next-generation sequencing (NGS) can be used as this is a nontargeted method; thus it has the capacity to detect every potential threat present. In this chapter, a protocol for the simultaneous detection and preliminary serotyping of Salmonella enterica serovar Enteritidis, Salmonella enterica serovar Typhimurium, Listeria monocytogenes, and Escherichia coli O157:H7 is described.

Frequency, serotyping, antibiogram, and seasonality of Salmonella isolated from red meat markets.

Bacterial pathogenic strains are as adaptable as Salmonella strains and cause diverse intestinal and extraintestinal diseases in humans and other mammals worldwide. Red meat and its products are important hosts for many zoonotic diseases. This work was designed to investigate the frequency, serotypes, and antimicrobial resistance profile of isolated Salmonella spp. in red meat (cattle, sheep, and goats) sold in Dhamar Governorate, Yemen. A total of 250 red meat samples were collected from the retail seller market between July and December 2022. All samples were transported immediately to the laboratory, subcultured on selective enrichment agar, and identified by serotyping and antimicrobial susceptibility tests via disk diffusion methods. The results indicated 26 positive samples of Salmonella out of the 250 samples (10.4%). Notably, isolates belong to ten various serotypes: S. Typhimurium 19.2%, S. Anatum 15.4%, S. Newport 11.5%, S. Enteritidis 11.5%, S. Muenchen 11.5%, S. Infantis 7.7%, S. Montevideo 7.7%, S. Dublin 7.7%, S. Senftenberg 3.9%, and S. Arizona 3.9%. The antibiotic resistance profile revealed that 57.5%, 53.9%, and 53.9% of isolates are resistant to erythromycin, tetracycline, and norfloxacin, respectively. This resistance among Salmonella spp. suggests a significant threat to health, which will in turn require an active safety measure and response. On the other hand, the seasonal variations "August and July" were found to be associated with an increased frequency of Salmonella isolation.

The PARK2_e01(-697) Polymorphism does not Associate with Susceptibil-ity to Typhoid in Punjabi Population, Pakistan: A Case Control Study.

BACKGROUND: SNP based association studies have revolutionized the field of biomed-icines. Enteric fever is a systemic disease with etiologic agents Salmonella enterica serovar typhi and paratyphi. It is a serious health issue worldwide and presents wide variations in incidence, rates, and severity. Previous investigations have revealed that genetic variations may lead to sus-ceptibility to typhoid fever. A current study was performed to investigate the potential association of PARK2_e01(-697) polymorphism with the susceptibility to typhoid in the Punjabi population. METHODS: For this case-control study, blood samples obtained from typhoid patients with positive Typhidot or blood culture test (n=72) and healthy controls (n=73) were processed for DNA ex-traction. The polymorphism PARK2_e01(-697) analysis was carried out by using PCR and RFLP. RESULTS: No allelic association was found between PARK2_e01(-697) and susceptibility to ty-phoid fever in the understudy population. CONCLUSION: This case control study is the demonstration of the non-association of PARK2_e01(-697) with typhoid in the Pakistani population. Future research, using a larger population size, will help to elucidate the role of PARK2_e01(-697) polymorphism in typhoid pathogenesis.

Evaluation of Salmonella biofilm attachment and hydrophobicity characteristics on food contact surfaces.

Salmonella forms biofilms, and persist on food contact surfaces. Once a biofilm is formed cleaning and sanitation protocols may be inadequate for effective removal. This study evaluated attachment characteristics, surface properties, and structure of Salmonella biofilms on food contact surfaces commonly used in the tree-fruit industry. Multi-strain Salmonella biofilms were grown in a Centers for Disease Control and Prevention (CDC) biofilm reactor at 22 ± 2 °C and sampling was conducted at 2, 24 and 96-h. After each incubation period, coupons weregently rinsed and the remaining cells enumerated. Biofilms were analyzed with Laser Scanning Confocal Microscopy (LSCM). Hydrophobicity was evaluated by measuring the contact angles of reference liquids method using a drop tensiometer instrument. Material type and biofilm age significantly influenced attachment and biofilm hydrophobicity (P < 0.05). The strength of attachment, across all time points, was highest on nylon followed by wood and high-density polyethylene. The highest contact angle measurements were observed after 96-h of biofilm formation for each material. All the results and observations from this study contribute to a better understanding of the attachment and hydrophobicity characteristics of Salmonella and might help producers make informed decisions when selecting containers for harvesting and storing in order to minimize biofilm formation and potential for cross-contamination.

Monophasic variant of Salmonella Typhimurium 4,[5],12:i:- outbreak: an investigation by the Competent Authority.

Foodborne diseases represent a significant public health issue, regarding both collective health and the economy, with implications for healthcare costs and agribusiness. This paper shows the description and results of the investigation of a Salmonella enterica subsp. enterica foodborne outbreak that occurred in the Marche Region (Italy) in 2022, which was linked to the consumption of a roasted pork product (porchetta). As part of the outbreak investigations discussed in this article, molecular analysis to evaluate the genomic correlation between clinical, food, and environmental origin strains was carried out. All Salmonella strains of different origins were serotyped as monophasic variant of Salmonella Typhimurium with an antibiotic-resistance pattern and an allelic profile such as to define the "cluster strain" allowing the correlation between clinical and food/environmental strains, definitively confirmed by whole genome sequencing analysis. Following the laboratory evidence, corrective measures at the porchetta processing plant and at the retail stores involved were carried out by the Local Animal Based Food Hygiene Service. The results of this study show that effective intervention is only possible if efficient data exchange, standardized procedures, and staff training are guaranteed. The latter aspect also concerns the food business operator, who must take appropriate measures to minimize the risk.

Incidence, antimicrobial susceptibility & out of pocket expenditure of severe enteric fever in Chandigarh, north India.

Background & objectives Burden estimates of enteric fever are required to make policy decisions on introducing typhoid vaccine in India. Incidence, antimicrobial susceptibility, and out-of-pocket expenditure (OOPE) of enteric fever are estimated in Chandigarh, India. Methods A hybrid (facility and community-based) surveillance system was set up at a secondary care hospital to enrol patients above six months of age, hospitalized with fever, from a defined catchment population from May 2018 to March 2020. Blood samples were collected and cultured using an automated system (BD BACTECTM blood culture system). The Salmonella Typhi and S. Paratyphi isolates were characterized for antimicrobial susceptibility. OOPE was recorded after 14 and 28 days of discharge. Results Blood samples were collected from 97 per cent of the 1650 study participants enrolled. The incidence of enteric fever was 226.8 per 1,00,000 person-years (PY), severe typhoid fever 156.9 per 1,00,000 PY, and severe paratyphoid fever 69.9 per 1,00,000 PY. Salmonella was highly susceptible to ampicillin, azithromycin, and ceftriaxone (99.25%) and least susceptible to ciprofloxacin (11.3%). The OOPE due to hospitalization of individuals infected with S. Paratyphi [INR 8696.6 (USD 116)] was significantly higher than the individuals infected with S. Typhi [INR 7309 (USD 97.5), P=0.01], and among cases who were hospitalized for more than seven days [INR 12,251 (USD 163.3)] as compared with those with a stay of 3-7 days [INR 8038.2 (USD 107.2)] or less than three days [INR 5327.8 (USD 71), P<0.001]. Interpretation & conclusions There was a high incidence of enteric fever, high OOPE, and resistance to ciprofloxacin.

Short communication: A multispecies bacterial-based direct-fed microbial alleviates Salmonella invasion and supports in vitro epithelial integrity.

Managing bacterial infections is of great importance in livestock production, particularly those caused by Salmonella enterica serovars Typhimurium or Dublin, which can impact both animal health and performance, as well as human food safety. Direct-fed microbials (DFM) can support gastrointestinal function and alleviate the potential negative effects of bacterial infections. In the present study, the capacity of a multispecies bacterial-based DFM containing Ligilactobacillus (formerly Lactobacillus) animalis 506, Propionibacterium freudenreichii 507, Bacillus licheniformis 809, and B. subtilis 597 to reduce S. Typhimurium ATCC14028 invasion was investigated using a co-incubation model with the HT29-MTX-E12 cell line (Exp. 1). Next, a possible antagonistic effect of the DFM against S. Dublin ATCC 41286 was evaluated using an in vitro agar well diffusion method following a co-incubation of 48 h (Exp. 2). At last, a series of experiments were performed to evaluate how different doses (6.25 × 106, 2.50 × 107, or 1.00 × 108 CFU/well) of the DFM would support the integrity of intestinal epithelial cells challenged or not with S. Typhimurium ATCC14028 or hydrogen peroxide under a transepithelial electrical resistance (TEER) assay with Caco-2 cells (Exp. 3 and 4). In Exp. 1, BDP significantly (P < 0.001) reduced by 90.8% the invasion of S. Typhimurium into HT29-MTX-E12 cells, whereas viability of the potentially harmful bacteria was reduced by 21.0% (P < 0.0001). In Exp. 2, the antagonistic properties of BDP towards S. Dublin were confirmed by the detection of a clear inhibition zone (size = 8.6 mm). Lastly, without challenge, the lowest dose of the DFM (6.25 × 106 CFU) provided the greatest support on the cells (treatment × hour; P < 0.0001). However, when the cells were challenged with S. Typhimurium, all doses alleviated the loss of integrity caused by the pathogen (treatment × hour; P < 0.0001). In cells challenged with hydrogen peroxide, the greater dose (1.00 × 108 CFU) supported the cells for a longer period of time (treatment × hour; P < 0.0001). These in vitro findings set the stage for exploring the potential benefits of using a novel DFM as a promising tool and strategy to mitigate Salmonella enterica infections in ruminants and improve animal health, food safety, and public health. Further in vivo confirmation needs to be developed to validate these preliminary in vitro results.

Phenotypic variation in the lipopolysaccharide O-antigen of Salmonella Paratyphi A and implications for vaccine development.

Enteric fever remains a major public health problem in South and Southeast Asia. The recent roll-out of the typhoid conjugate vaccine protecting against S. Typhi exhibits great promise for disease reduction in high burden areas. However, some endemic regions remain vulnerable to S. Paratyphi A due to a lack of licensed vaccines and inadequate WASH. Several developmental S. Paratyphi A vaccines exploit O-antigen as the target antigen. It has been hypothesised that O-antigen is under selective and environmental pressure, with mutations in O-antigen biosynthesis genes being reported, but their phenotypic effects are unknown. Here, we aimed to evaluate O-antigen variation in S. Paratyphi A originating from Nepal, and the potential effect of this variation on antibody binding. O-antigen variation was determined by measuring LPS laddering shift following electrophoresis; this analysis was complemented with genomic characterisation of the O-antigen region. We found structural O-antigen variation in <10 % of S. Paratyphi A organisms, but a direct underlying genetic cause could not be identified. High-content imaging was performed to determine antibody binding by commercial O2 monoclonal (mAb) and polyclonal antibodies, as well as polyclonal sera from convalescent patients naturally infected with S. Paratyphi A. Commercial mAbs detected only a fraction of an apparently "clonal" bacterial population, suggesting phase variation and nonuniform O-antigen composition. Notably, and despite visible subpopulation clusters, O-antigen structural changes did not appear to affect the binding ability of polyclonal human antibody considerably, which led to no obvious differences in the functionality of antibodies targeting organisms with different O-antigen conformations. Although these results need to be confirmed in organisms from alternative endemic areas, they are encouraging the use of O-antigen as the target antigen in S. Paratyphi A vaccines.

Evaluation of a postbiotic on Salmonella enterica prevalence, serotype diversity, and antimicrobial resistance in the subiliac lymph nodes of cull dairy cattle.

This study aimed to determine whether the farm-level use of a Saccharomyces cerevisiae-based postbiotic was associated with Salmonella prevalence and concentration, serotype diversity, and antimicrobial resistance in the subiliac lymph nodes (LN) of cull dairy cows. In collaboration with two commercial processing plants in the Southwestern (SW) and Northeastern (NE) regions of the U.S., cull dairy cattle lots processed in the same week from dairy farms that fed or not the postbiotic were sampled. Up to 20 LN were collected from dairy cattle from each supplier farm at least once every season. Samples were analyzed for Salmonella by culture and quantitative PCR methods, and isolates were subjected to serotype identification and antimicrobial susceptibility testing. Although a numerically lower prevalence was observed, the pre-harvest administration of the postbiotic was not significantly associated with Salmonella prevalence in cull dairy cattle. However, Salmonella prevalence significantly varied by region; the SW region showed a higher prevalence than the NE region. Whereas dominant Salmonella serotypes included Montevideo, Mbandaka, Muenster, Cerro, Meleagridis, and Anatum, the probability of isolating a dominant serotype did not significantly vary by feed additive status (FAS) or region, but varied by season. Up to 34 isolates (out of 391) exhibited resistance to each antimicrobial, with the highest number of isolates exhibiting resistance to streptomycin and ciprofloxacin, however, this did not significantly vary by FAS, season, or region. The precise reasons for the lack of effectiveness of the postbiotic in reducing Salmonella burden are unknown, however, varied administration durations of the postbiotic due to culling at different lactation cycle stages or during dry periods may have hindered its impact, especially if dairy cows were culled early. Other factors may include dietary components, farm management practices, and external environmental influences.

Geographical Variation of Antimicrobial Resistance of Salmonella in Japanese Chicken.

Chicken is a potent source of Salmonella infection in humans. Occasionally, patients with severe Salmonella enteritis require antimicrobial therapy. Antimicrobials are used to prevent and treat bacterial infections in broiler and breeder farms. Herein, we investigated the prevalence and antimicrobial resistance of Salmonella in 337 vacuum-packed chicken breast products manufactured in Japan between June and December 2021. Salmonella was isolated from 287 samples (85.2%). Among the products from Eastern Japan, the lowest Salmonella prevalence was observed in those processed in September (65.6%), which was significantly (p < 0.05) lower than that in November or December. Among the products from Western Japan, the lowest Salmonella prevalence was observed in those processed in August (61.9%), which was significantly (p < 0.05) lower than that in June, November, and December. The most frequent serovar was Salmonella Schwarzengrund (223 isolates), followed by S. Infantis (53 isolates), S. Manhattan (9 isolates), and S. Enteritidis (1 isolate). High rates of antimicrobial resistance were observed for streptomycin (64.5%), kanamycin (50.2%), tetracycline (65.2%), nalidixic acid (11.5%), and trimethoprim (35.9%). Resistance rates against these five antimicrobials in S. Schwarzengrund isolates were markedly higher in the isolates from Western Japan than in those from Eastern Japan. All 287 Salmonella isolates were susceptible to ciprofloxacin which belongs to fluoroquinolones and cefotaxime which belongs to third-generation cephalosporins. Salmonella prevalence in chicken products in Japan was found to be extremely high; therefore, chicken meat should be thoroughly heated before consumption. In Japan, fluoroquinolones and third-generation cephalosporins are recommended as the first- and second-choice antimicrobials for patients with severe Salmonella enteritis, respectively. The results of this study show that administering fluoroquinolones or third-generation cephalosporins is an effective option for patients with Salmonella enteritis caused by consuming chicken meat, and efficient strategies for Salmonella management on broiler farms and chicken-processing plants need to be developed.

Innovative approaches to controlling Salmonella in the meat industry.

Salmonella, a Gram-negative, rod-shaped bacterium from the Enterobacteriaceae family, is a significant cause of illnesses in humans and animals. It resides in the digestive tracts of livestock, poultry, and other warm-blooded animals and can contaminate various environments and foods through fecal matter. Salmonella enterica, the main species that affects humans, is widespread in cattle, pigs, and poultry. Despite efforts to control pathogens in meat systems, over 1.4 million human salmonellosis cases occur annually in the U.S., with serotypes S. enteritidis and S. typhimurium being predominant. Advances in meat processing have targeted pathogen reduction at multiple stages, but more innovative approaches are needed for substantial public health impact. This paper discusses current and future strategies to minimize Salmonella in the food supply. It emphasizes pre- and post-harvest Salmonella prevalence by biomapping it through the whole processing chain, focusing on beef and pork interventions such as probiotics. These interventions have shown promise in reducing pathogen loads in cattle manure and lymph nodes. Techniques such as microbiome, whole genome sequencing (WGS), and electron microscopy (EM) provide detailed insights into Salmonella's genetic and bacterial structural-morphological characteristics, aiding in the development of targeted interventions. Integrating rapid detection, biomapping, and enviromapping enhances pathogen tracking in meat production, reducing Salmonella prevalence and improving risk assessment and food safety. The advanced, current, and innovative techniques allow for timely identification, detailed spatial and quantitative data, and more effective interventions. This leads to safer food products and reduces foodborne illnesses.

Emergence and Characterization of the High-Level Tigecycline Resistance Gene tet(X4) in Salmonella enterica Serovar Rissen from Food in China.

The plasmid-mediated tet(X4) gene has exhibited a high-level resistance to tigecycline (TGC), which has raised concerns globally regarding antibiotic resistance. Although the widespread tet(X4) has been found widely in Escherichia coli, it is scarcely found in other Enterobacteriaceae. This study aimed to characterize a ST469 Salmonella enterica serovar Rissen (S. Rissen) isolate harboring tet(X4) from pork, which was identified and characterized via antimicrobial susceptibility testing, conjugation assays, plasmid curing testing, whole-genome sequencing, and bioinformatic analysis. Ten ST469 S. Rissen isolates of 223 Salmonella spp. isolates were isolated from food samples in China during 2021-2023. One of 10 S. Rissen isolates, SM2301, carrying tet(X4) conferred high-level resistance to TGC (minimum inhibitory concentration > 8 µg/mL). The tet(X4) could be conjugated into different recipients, including E. coli, S. enteritidis, and K. pneumoniae isolates. Plasmid curing confirmed that tet(X4) was plasmid-mediated. Genetic analysis revealed that the tet(X4) in the SM2301 isolate was located in the IncFIA(HI1)-IncHI1A-IncHI1B(R27) hybrid plasmid, and the structure of tet(X4) was abh-tet(X4)-ISCR2. To the best of our knowledge, this is the first report of a tet(X4)-positive food-derived S. Rissen isolate. The extending bacterial species of tet(X4)-bearing plasmids suggested the increasing transmission risk of the mobile TGC resistance gene tet(X4) beyond E. coli. This study highlights the emerging and evolution risk of novel resistance genes across various bacterial species. Therefore, further surveillance is warranted to monitor the prevalence of tet(X4) in Salmonella spp. and other bacterial species.

SalmoFree® Phage Additive Proves Its Safety for Laying Hens.

Background: The avian pathogen Salmonella Gallinarum causes avian typhosis in laying hens, leading to high mortality rates among adult birds, which poses a significant problem in the poultry industry. Various products, such as vaccines, antibiotics, probiotics, and disinfectants, are commonly used to prevent and control the disease on farms. An alternative to these products is the use of bacteriophages, which may effectively prevent the colonization of S. Gallinarum. Materials and Methods: This study evaluated the safety of SalmoFree®, a bacteriophage cocktail, administered to 276 laying hens from the first week of age until the 28th week. The hens were divided into two groups: a control group (138 birds) and a treatment group (138 birds). Over the 28-week period, eight doses of SalmoFree® (∼1010 UFP per bird) were administered via drinking water in a controlled environment. Results: The results indicate that the consumption of SalmoFree® has no adverse effects on bird health or zootechnical parameters. Additionally, there is a trend toward improving weight homogeneity (up to 19%), feed conversion (up to 68%), and egg weight (up to 2.7%). The detection of phages by PCR in cloacal swabs suggests that they persist in birds for 2 to 8 weeks post-ingestion. Furthermore, phages were detected in organs and eggshells, indicating that they provide protection beyond the gut. Conclusion: The study demonstrates that SalmoFree® is safe for use in laying hens and may offer additional benefits, such as improved zootechnical parameters and extended protection against S. Gallinarum through the persistence of bacteriophages in the birds.

Epigenetic regulation in epithelial cells and innate lymphocyte responses to S. Typhi infection: insights into IFN-γ production and intestinal immunity.

Infection by Salmonella enterica serovar Typhi (S. Typhi), the cause of enteric fevers, is low in high-income countries but persistent in low- and middle-income countries, resulting in 65,400-187,700 deaths yearly. Drug resistance, including in the United States, exacerbates this issue. Evidence indicates that innate lymphocytes (INLs), such as natural killer (NK) cells, and unconventional T lymphocytes (e.g., Mucosal-associated invariant T (MAIT) cells and T-cell receptor gamma delta (TCR-γδ) cells) can impact the intestinal epithelial barrier, the primary site of exposure to S. Typhi. Moreover, INL production of IFN-γ is central in controlling S. Typhi infection. However, the impact of epithelial cells (EC) on the secretion of IFN-γ by INLs and the relationship between these events and epigenetic changes remains unknown. Epigenetic modifications in host cells are fundamental for their differentiation and function, including IFN-γ production. Herein, using a human organoid-derived polarized intestinal epithelial cell monolayer, we investigated the role of H3K4me3 and H3K27me3 epigenetic marks in intestinal immunity, focusing on the function of EC, NK, MAIT, and TCR-γδ cells in response to S. Typhi. This study builds on our previous findings that MAIT subsets exhibiting specific IFN-γ pattern signatures were associated with protection against typhoid fever and that S. Typhi infection regulates changes in chromatin marks that depend on individual cell subsets. Here, we show that cultures exposed to S. Typhi without EC exhibit a significant increase in NK and MAIT cells, and, to a lesser extent, TCR-γδ cells, expressing IFN-γ and H3K4me3 but not H3K27me3 marks, contrasting with cultures where EC is present. The influence of EC on INL H3K4me3 marks might be indirectly mediated through the modulation of IL-18 secretion via the Histone Deacetylase 6 gene during S. Typhi infection.

Time-Resolved Multiomics Illustrates Host and Gut Microbe Interactions during Salmonella Infection.

Salmonella infection, also known as Salmonellosis, is one of the most common food-borne illnesses. Salmonella infection can trigger host defensive functions, including an inflammatory response. The provoked-host inflammatory response has a significant impact on the bacterial population in the gut. In addition, Salmonella competes with other gut microorganisms for survival and growth within the host. Compositional and functional alterations in gut bacteria occur because of the host immunological response and competition between Salmonella and the gut microbiome. Host variation and the inherent complexity of the gut microbial community make understanding commensal and pathogen interactions particularly difficult during a Salmonella infection. Here, we present metabolomics and lipidomics analyses along with the 16S rRNA sequence analysis, revealing a comprehensive view of the metabolic interactions between the host and gut microbiota during Salmonella infection in a CBA/J mouse model. We found that different metabolic pathways were altered over the four investigated time points of Salmonella infection (days -2, +2, +6, and +13). Furthermore, metatranscriptomics analysis integrated with metabolomics and lipidomics analysis facilitated an understanding of the heterogeneous response of mice, depending on the degree of dysbiosis.

Molecular Compositions, Mutagenicity, and Mutation Spectra of Atmospheric Oxidation Products of Alkenes and Dienes Initiated by NOx + UV or Ozone: A Structure-Activity Analysis.

Photooxidation products resulting from volatile organic compounds (VOCs) reacting with sunlight are important contributors to gas-phase air pollution. We characterized the product-weighted mutagenic potencies (rev m3 mgC-1 h-1) in Salmonella TA100 of atmospheres resulting from the hydroxyl radical (OH)-initiated photochemical oxidation of 11 C4 or C5 alkenes or dienes in the presence of nitric oxide (NO) and from the ozonolysis of four VOCs without NO (isoprene; 1,3-pentadiene; 1,4-pentadiene; and 1,3-butadiene). Irradiated atmospheres from precursors with a single C═C bond (3-methyl-1-butene, 2-methyl-1-butene, cis/trans-2-pentene, 2-methyl-2-butene, 1-butene, and 1-pentene) had low potencies (<5), whereas linear dienes with terminal C═C bonds had high potencies (50-65). Dienes with a branched structure (isoprene) or internal C═C bonds (1,3-pentadiene) had intermediate potencies (15-20). No VOCs were mutagenic without photochemical oxidation. VOCs+O3 in the dark produced less mutagenic atmospheres than photochemistry in the presence of NO. Atmospheres induced primarily C to T and C to A mutations, the main base substitutions in nonsmoker lung cancer. Atmospheres from the photooxidation of isoprene and 1,3-pentadiene also induced GG to TT, the signature mutation of peroxyacetyl nitrate. Five molecular compositions identified by Chemical Ionization Mass Spectrometry (CIMS), most containing nitrogen, correlated (r = 0.76-0.85) with the mutagenic potencies of irradiated atmospheres; most had a likely nitrate functional group. Assessment of the mutagenicity of emitted VOCs should consider VOC photooxidation products, especially dienes with terminal C═C bonds, as these products likely contribute to overall health effects from ambient air pollution.

First report on whole genome sequencing and comparative genomics of Salmonella enterica serovar Abortusequi isolated from Donkey in India.

Salmonella enterica subspecies enterica serovar Abortusequi (S. Abortusequi) is a leading cause of abortion in equines that hinders the rapid growth of equine industry. S. Abortusequi infection in equids has re-emerged over last ten years. In the present study, S. Abortusequi was isolated and characterized from donkeys during an abortion storm in the southern peninsular region of India. Further, whole genome sequencing and phylogenomic analysis revealed that the present isolate was clustered among S. Abortusequi clade. The core genome MLST (cgMLST) analysis based on hierarchical clustering and single nucleotide polymorphism (SNP) core-genome dendrogram of the present isolate against 10 S. Abortusequi isolates revealed that the present isolate established a distinct clade compared to all previously reported isolates. A comparison of cgMLST and SNP analyses revealed the same clustering concordance between isolates. In addition, comparative genomics and phylogenetic analysis was carried out with six S. Abortusequi serovars showed a higher number of core genes than accessory genes. Further, comparative analysis of phenotype and genotype antimicrobial resistance revealed a concordance of 32% and discordance of 68% respectively.

Application of hot water and cold air to reduce bacterial contamination on broiler carcasses.

Two physical treatments (heat via water bath and cold air) with various temperatures (20/70/75/80°C and - 80/-90°C) and exposure times (20, 30, 40 s) were carried out to identify a decontaminating effect on zoonotic pathogens on broiler carcasses. Subsequently, carcasses were analyzed for thermotolerant Campylobacter (C.), Salmonella, Escherichia (E.). coli and total colony count (TCC). Moreover, for the hot water treatment, qPCR with viable/dead differentiation (v-qPCR) was applied to detect viable but non-culturable cells (VBNC) of Campylobacter, referred to as intact but putatively infectious units (IPIU). Hot water immersion was tested on carcasses inoculated with C. jejuni and Salmonella, while cold air treatment was evaluated for naturally contaminated carcasses of broiler flocks colonized with Campylobacter. For hot water treatment, the statistically significant reducing effect was about 1 log10 CFU/ml for both Salmonella and Campylobacter for 70-80°C and 20/30 s treatments. The effect of heat treatment for Campylobacter was smaller when samples were analyzed with v-qPCR with reductions of 0.5-0.8 log10 IPIU/ml in mean. Cold air treatments at -90°C were effective in reducing the mean contamination level of Campylobacter by 0.4-0.5 log10 CFU/ml at all exposure times (p < 0.05). Hot water treatments showed a decreasing trend on TCC by 0.6-0.9 log10 CFU/ml (p < 0.05). TCC counts were not significantly affected by cold air treatment. For E. coli no statistically significant reductions were observed by hot water treatment. The cold air treatment at -90°C for 20 and 40 s led to a reduction of E. coli by 0.4 and 0.8 log10 CFU/ml (p < 0.05), respectively. Treatment of carcasses with higher bacterial levels tended to show higher reduction. The research demonstrated that the efficacy of physical treatments for decontamination of broiler carcasses was more pronounced for hot water immersion than for cold air exposure. In conclusion, the results shed light on the potential application of these physical treatments in practice to reduce the quantitative load of contaminating pathogens to enhance food safety in the broiler meat production.