Meat Isolates Research Articles

The European Union Summary Report on Antimicrobial Resistance in zoonotic and indicator bacteria from humans, animals and food in 2020/2021.

Antimicrobial resistance (AMR) data on zoonotic and indicator bacteria from humans, animals and food are collected annually by the EU Member States (MSs) and reporting countries, jointly analysed by EFSA and ECDC and presented in a yearly EU Summary Report. This report provides an overview of the main findings of the 2020-2021 harmonised AMR monitoring in Salmonella spp., Campylobacter jejuni and C. coli in humans and food-producing animals (broilers, laying hens and turkeys, fattening pigs and bovines under 1year of age) and relevant meat thereof. For animals and meat thereof, indicator E. coli data on the occurrence of AMR and presumptive Extended spectrum β-lactamases (ESBL)-/AmpC β-lactamases (AmpC)-/carbapenemases (CP)-producers, as well as the occurrence of methicillin-resistant Staphylococcus aureus are also analysed. In 2021, MSs submitted for the first time AMR data on E. coli isolates from meat sampled at border control posts. Where available, monitoring data from humans, food-producing animals and meat thereof were combined and compared at the EU level, with emphasis on multidrug resistance, complete susceptibility and combined resistance patterns to selected and critically important antimicrobials, as well as Salmonella and E. coli isolates exhibiting ESBL-/AmpC-/carbapenemase phenotypes. Resistance was frequently found to commonly used antimicrobials in Salmonella spp. and Campylobacter isolates from humans and animals. Combined resistance to critically important antimicrobials was mainly observed at low levels except in some Salmonella serotypes and in C. coli in some countries. The reporting of a number of CP-producing E.coli isolates (harbouring bla OXA-48, bla OXA-181, and bla NDM-5 genes) in pigs, bovines and meat thereof by a limited number of MSs (4) in 2021, requests a thorough follow-up. The temporal trend analyses in both key outcome indicators (rate of complete susceptibility and prevalence of ESBL-/AmpC- producers) showed that encouraging progress have been registered in reducing AMR in food-producing animals in several EU MSs over the last years.

PREVALENCE, PHENOTYPIC-GENOTYPIC RESISTANCE AND BIOFILM FORMATION OF Staphylococcus aureus IN CHICKEN MEAT WITH REFERENCE TO ITS PUBLIC HEALTH HAZARD

The purpose of this study was to determine the prevalence, antimicrobial resistance in S. aureus and presence of enterotoxigenic as well as biofilm-forming genes in S. aureus bacteria isolated from broiler meat from traditional shops and supermarkets in Sharkia, Egypt. For this determination, two hundred fresh raw chicken meat cotton swabs were collected from the breast and thighs (100 each). S. aureus was highlighted through a coagulase test, and then phenotypic and genotypic characterizations were studied. Uniplex PCR was used to identify the occurrence of enterotoxin genes in the selected isolates. Finally, they were subjected to a biofilm formation test using 96-well flat bottom polystyrene microtiter plates; besides, biofilm-forming genes were investigated. Nineteen isolates out of the 200 samples tested positive for S. aureus (9.5 percent), No Vancomycin resistance strains were obtained, nor did any ciprofloxacin isolates for S. aureus while, all isolates were 100 % resistant for streptomycin, amoxicillin- clavulanic acid and sulfamethoxazole-trimethoprim S. aureus isolates were found resistant to at least one antibiotic. The multiple antibiotic resistance (MAR) index of S. aureus isolates was ranged from 0.3 to 0.7 with an average of 0.4. In The mecicillin resistant S.aureus (MRSA) strains carried mecA gene with a percentage of 5%, while the blaZ gene was distributed with a percentage of 5.5%, 11/200). The obtained isolates gave 60% strong biofilm formation and 40% were non-formers, with nil results for moderate or weak production isolates. The icaA gene was 100%, in comparison to icaD which was zero. It should be noted that strong biofilm former strains were only 100% positive for sea and seb. This study pointed out the higher prevalence of MDR isolates of S. aureus isolates in chicken meat due to inadequate handling and insufficient sanitary equipment and post microbial contamination. This finding highlighted the importance of broiler meat as a reservoir for antimicrobial-resistant strains of S. aureus and biofilm-forming of Staphylococcus aureus.

Prevalence of Antimicrobial Resistance and Clonal Relationship in ESBL/AmpC-Producing Proteus mirabilis Isolated from Meat Products and Community-Acquired Urinary Tract Infection (UTI-CA) in Southern Brazil.

The present study aimed to evaluate the prevalence of antimicrobial resistance and clonal relationships in Proteus mirabilis isolated from chicken meat, beef, pork, and community-acquired urinary tract infections (UTI-CA). Chicken meat isolates showed the highest multidrug resistance (MDR), followed by those from pork and UTI-CA, whereas beef had relatively few MDR strains. All sources had strains that carried blaCTX-M-65, whereas blaCTX-M-2 and blaCMY-2 were only detected in chicken meat and UTI-CA isolates. This indicates that chicken meat should be considered an important risk factor for the spread of P. mirabilis carrying ESBL and AmpC. Furthermore, ESBL/AmpC producing strains were resistant to a greater number of antimicrobials and possessed more resistance genes than non-producing strains. In addition, the antimicrobial resistance genes qnrD, aac(6')-Ib-cr, sul1, sul2, fosA3, cmlA, and floR were also found. Molecular typing showed a genetic similarity between chicken meat and UTI-CA isolates, including some strains with 100% similarity, indicating that chicken can be a source of P. mirabilis causing UTI-CA. It was concluded that meat, especially chicken meat, can be an important source of dissemination of multidrug-resistant P. mirabilis in the community.

Incidence of Antibiotic Resistant Bacteria in Poultry and Livestock in Dhaka City

Antimicrobial resistance is a concern for humans and animals all over the world. Indiscriminate use of antibiotics in livestock and poultry has become one of the major causes of antimicrobial resistance development in microorganisms. The aim of the study was to determine the antimicrobial resistance patterns of bacteria isolated from raw meat, intestine, feces and feed of farm chicken, cow and local chicken. Samples were collected from different retail shops at Malibagh area in Dhaka City, Bangladesh. Bacterial load was enumerated, potentially pathogenic bacteria were identified and antibiogram was determined following standard methods. A total of 43 bacterial isolates were identified from different samples which were Escherichia coli, Enterobacter, Citrobacter, Klebsiella, Providencia and Acinetobacter spp. Eight types of antibiotics such as, Ampicillin-10μg, Gentamicin-10μg, Amikacin-10μg, Amoxicillin-10μg, Ceftriaxone-30μg, Imipenem-10μg, Chloramphenicol-30μg and Tetracycline-30μg were used to determine the antibiotic sensitivity pattern of isolated bacteria. The antibiotic resistance pattern of bacteria isolated from feed and feces samples of cow and chicken were found to be similar. Isolated bacteria from chicken meats showed higher antimicrobial resistance (80-100%) against Ampicillin-10μg, Imipenem-10μg and Amoxicillin-10μg compared to cow meat isolates. The incidence of antibiotic resistant bacteria is a threat to animals, food handlers and consumers if they are being infected by these antibiotic resistant pathogenic bacteria. Stamford Journal of Microbiology, Vol.12 (1) 2022: 37-42

1367. Resistance to Ceftriaxone and Ciprofloxacin in Nontyphoidal Salmonella from Humans and Relatedness to Isolates from Animal Sources

Abstract Background Nontyphoidal Salmonella (NTS) is a leading cause of foodborne bacterial infections. Annually, ∼200,000 antimicrobial-resistant (AMR) NTS illnesses occur in the United States. NTS infections resistant to ceftriaxone and ciprofloxacin, drugs used for the treatment of severe infections, are a growing concern. Methods We identified patient NTS isolates submitted to the Pennsylvania Bureau of Laboratories by two hospitals from the same health system during 2018–2020. All were tested by broth microdilution for susceptibility to antibiotics tracked by the National Antimicrobial Resistance Monitoring System and analyzed by whole-genome sequencing. We identified resistance mechanisms and plasmids with ResFinder and PlasmidFinder, respectively, and compared each isolate in selected serotypes against the PulseNet database to identify genetic relatedness within ≤5 allele differences to non-human sequences uploaded during 2017–2021. Results Of 164 human NTS analyzed for susceptibility, 28 (17%) had decreased susceptibility to ciprofloxacin (DSC) [MIC ≥0.12 µg/mL] while 6 (3.7%) were ceftriaxone-resistant. AMR varied by serotype; 8 (50%) S. Infantis and 16 (42%) Enteritidis had DSC, representing 86% of all isolates with DSC (Figure). Six ceftriaxone-resistant isolates had genes that confer resistance to third-generation cephalosporins including blaCMY−2 in 1 Dublin and 2 Typhimurium and blaCTX-M-65 in 3 Infantis isolates. The 3 Infantis isolates also had a mutation in gyrA that results in DSC, plus a known transmissible plasmid IncFIB(pN55391) linked to multiple resistance genes. An Infantis isolate was related to a chicken meat isolate and a Hadar isolate was related to isolates from 3 animals and 3 meat sources. One Dublin isolate was related to 2 isolates from beef in Pennsylvania. Figure.Percent of antimicrobial resistance in nontyphoidal Salmonella (NTS) clinical isolates (n=164) submitted to public health authorities by two hospital systems in Pennsylvania, 2018–2020. Multidrug resistance (MDR) was defined as resistance to ≥3 of the 9 antimicrobial classes tested for NTS clinical isolates. Resistance to ceftriaxone (AXO), a third-generation cephalosporin preferred for treating severe infections in children, was over 5% and 19% in serotypes S. Typhimurium and S. Infantis, respectively. Half (8/16) of Infantis isolates and 42% (16/38) of Enteritidis isolates had decreased susceptibility to ciprofloxacin (DSC). Conclusion Ceftriaxone resistance and DSC in NTS from patients in Pennsylvania varied by serotype and some isolates harbored blaCMY−2 and blaCTX-M-65 genes. Dissemination of mechanisms that confer resistance to ceftriaxone and DSC is concerning as is the genetic relatedness of isolates from human and animal sources. Our study highlights the need for enhanced One Health AMR monitoring combined with a review of animal production and food processing practices. Disclosures All Authors: No reported disclosures.

Genotypic virulence profiles and associations in Salmonella isolated from meat samples in wet markets and abattoirs of Metro Manila, Philippines

BackgroundSalmonella are pathogenic foodborne bacteria with complex pathogenicity from numerous virulence genes housed in Salmonella pathogenicity islands (SPIs), plasmids, and other gene cassettes. However, Salmonella virulence gene distributions and mechanisms remain unestablished. In the Philippines, studies mainly report Salmonella incidences and antimicrobial resistance, but little to none on virulence profiles, their associations to animal sources, collection sites and Salmonella serogroups. Hence, a total of 799 Salmonella isolates, previously obtained from pig, cow, and chicken meat samples in wet markets and abattoirs (wet markets: 124 chicken, 151 cow, and 352 pig meat isolates; abattoirs: 172 pig tonsil and jejunum isolates) in Metro Manila, Philippines, were revived and confirmed as Salmonella through invA gene polymerase chain reaction (PCR). Isolates were then screened for eight virulence genes, namely avrA, hilA, sseC, mgtC, spi4R, pipB, spvC and spvR, by optimized multiplex PCR and significant pair associations between virulence genes were determined through Fisher’s exact test. Gene frequency patterns were also determined. Salmonella serogroups in addition to animal sources and location types were also used to predict virulence genes prevalence using binary logistic regression.ResultsHigh frequencies (64 to 98%) of SPI virulence genes were detected among 799 Salmonella isolates namely mgtC, pipB, avrA, hilA, spi4R and sseC, from most to least. However, only one isolate was positive for plasmid-borne virulence genes, spvC and spvR. Diversity in virulence genes across Salmonella serogroups for 587 Salmonella isolates (O:3 = 250, O:4 = 133, O:6,7 = 99, O:8 = 93, O:9 = 12) was also demonstrated through statistical predictions, particularly for avrA, hilA, sseC, and mgtC. mgtC, the most frequent virulence gene, was predicted by serogroup O:9, while sseC, the least frequent, was predicted by serogroup O:4 and chicken animal source. The highest virulence gene pattern involved SPIs 1-5 genes which suggests the wide distribution and high pathogenic potential of Salmonella. Statistical analyses showed five virulence gene pair associations, namely avrA and hilA, avrA and spi4R, hilA and spi4R, sseC and spi4R, and mgtC and pipB. The animal sources predicted the presence of virulence genes, sseC and pipB, whereas location type for hilA and spi4R, suggesting that these factors may contribute to the type and pathogenicity of Salmonella present.ConclusionThe high prevalence of virulence genes among Salmonella in the study suggests the high pathogenic potential of Salmonella from abattoirs and wet markets of Metro Manila, Philippines which poses food safety and public health concerns and threatens the Philippine food animal industry. Statistical associations between virulence genes and prediction analyses across Salmonella serogroups and external factors such as animal source and location type and presence of virulence genes suggest the diversity of Salmonella virulence and illustrate determining factors to Salmonella pathogenicity. This study recommends relevant agencies in the Philippines to improve standards in food animal industries and increase efforts in monitoring of foodborne pathogens.

The Occurrence and Genomic Characteristics of mcr-1-Harboring Salmonella from Retail Meats and Eggs in Qingdao, China.

Salmonella are widely distributed foodborne pathogens and are often associated with food animal products. Colistin resistance mediated by mcr-1 is an increasing threat; however, data on the characteristics of mcr-1-harboring Salmonella among retail foods are still lacking. In this study, retail meats from 24 supermarkets and eggs from nine markets in Qingdao city were investigated to determine the presence and genomic characteristics of mcr-1-harboring Salmonella. We found the retail meats and eggs were highly contaminated by Salmonella, with detection rates of 17.5% (31/177) and 12.3% (16/130), respectively. A total of 76 Salmonella isolates were obtained in this study, and 77.6% showed multidrug resistance (MDR). The MDR proportion of egg isolates (97.5%) was significantly higher than that in meat isolates (55.6%) (p < 0.05). The most prevalent Salmonella serotypes were Typhimurium (56.6%) and Enteritidis (17.1%). Of the 76 Salmonella isolates, 40 possessed mcr-1. All 40 mcr-1-positive isolates were ST34 S. Typhimurium and were from eggs of eight brands. Different mcr-1-harboring isolates existed in the same egg, and some isolates from different egg samples or brands showed clonal relationships. The mcr-1 was located on similar IncHI2/HI2A MDR non-conjugative plasmids lacking transfer region, resulting in the failure of conjugation. The phylogenetic tree using genome sequences showed that the mcr-1-positive isolates from eggs clustered together with mcr-1-positive isolates from chicken and humans in China, revealing that mcr-1-positive egg-borne Salmonella might be derived from chicken and could potentially trigger outbreaks in humans. The high occurrence of mcr-1-harboring Salmonella in fresh eggs is alarming, and there is an urgent need to monitor mcr-1-harboring Salmonella in retail meats and eggs. We report for the first time the role of retail eggs in disseminating mcr-1-positive Salmonella and the risk of transmission of these MDR pathogens from retail food to humans should be evaluated comprehensively.

Antimicrobial Resistant E. coli in Pork and Wild Boar Meat: A Risk to Consumers.

Antimicrobial-resistant foodborne microorganisms may be transmitted from food producing animals to humans through the consumption of meat products. In this study, meat that was derived from farmed pigs and wild boars was analyzed and compared. Escherichia coli (E. coli) were isolated and tested phenotypically and genotypically for their resistance to quinolones, aminoglycosides and carbapenems. The co-presence of AMR-associated plasmid genes was also evaluated. A quinolone AMR phenotypic analysis showed 41.9% and 36.1% of resistant E. coli derived from pork and wild boars meat, respectively. A resistance to aminoglycosides was detected in the 6.6% of E. coli that was isolated from pork and in 1.8% of the wild boar meat isolates. No resistant profiles were detected for the carbapenems. The quinolone resistance genes were found in 58.3% of the phenotypically resistant pork E. coli and in 17.5% of the wild boar, thus showing low genotypic confirmation rates. The co-presence of the plasmid-related genes was observed only for the quinolones and aminoglycosides, but not for the carbapenems. Wild boar E. coli were the most capable to perform biofilm production when they were compared to pork E. coli. In conclusion, the contamination of pork and wild boar meat by AMR microorganisms could be a threat for consumers, especially if biofilm-producing strains colonize the surfaces and equipment that are used in the food industry.

Phenotypic and genotypic characterization of multi-drug resistant, biofilm forming, human invasive strain of Salmonella Typhimurium SMC25 isolated from poultry meat in India

Emergence of highly virulent and multi-drug resistant (MDR) strains of Salmonella in food products significantly impacts public health and demands continuous monitoring for their presence in the food chain. The ability of Salmonella to form biofilms under harsh environmental conditions accompanied by MDR serotypes underscores an important food safety threat. This study aimed to isolate, identify and characterize MDR, biofilm-forming Salmonella from local Indian dairy and meat products (n = 60). All of the 24 isolates of Salmonella produced biofilm and were categorized as strong (16.6%), moderate (58.3%), and weak (25%) biofilm producers. Multiple antimicrobial resistance (MAR) index of all the Salmonella isolates was ≥0.2. The strongest biofilm forming poultry meat isolate, Salmonella SMC25 demonstrated intermediate to complete resistance to 14 of 22 different antibiotics tested. Epifluorescence microscopy showed that biofilm formation initiated as early as 4 h, reaching zenith within 96 h and much denser and robust biofilm is formed on rough stainless steel (SS316) surface compared to smooth glass surface. The results corroborated with increased temporal production of exopolysaccharides (EPS), high cell surface hydrophobicity and upregulation of marker genes vital to biofilm-formation in Salmonella. Significantly, SMC25 was found to adhere and invade mammalian cell lines Caco2 and HepG2, thus posing a serious food safety threat. This study is important in comprehending the prevalence of multidrug resistant, biofilm-forming, invasive strains of foodborne Salmonella in Indian food products and is important for effective risk assessment besides ensuring better food safety and public health.

Biofilm formation, antibiotic resistance and genotyping of Shiga toxin-producing Escherichia coli isolated from retail chicken meats

ABSTRACT 1. The Shiga toxin-producing Escherichia coli (STEC) is a hazardous zoonotic agent for chicken meat consumers. This study determined the serogroups and evaluated the virulence genes, antibiotic resistance, biofilm-forming profiles and genetic relationships of STEC isolates in chicken meat. 2. A total of 100 samples belonging to dressed-whole chicken and different parts of the chicken (wing, breast, thigh, drumstick) were collected between September and November 2019 from different retail markets in Kayseri, Türkiye. 3. Phenotypic (identification, disc diffusion test, Congo red agar and microtitre plate tests) and molecular tests (identification, serogrouping, virulence factors, biofilm, antibiotic susceptibility, 16S rRNA sequencing and enterobacterial repetitive intergenic consensus-PCR for typing of the isolates) were carried out. 4. E. coli was isolated from 35% of the samples and 35% of the samples harboured at least one STEC. Among 35 STEC isolates, 3 (8.5%), 6 (17.1%), 2 (5.7%) and 3 (8.5%) were found to be positive for fliCH2, fliCH8, fliCH11, fliCH19 genes, respectively. Out of 35 STEC positive isolates, 4 (11.4%) were identified as E. coli O157, from which 2 (5.7%) were E. coli O157:H7. E. coli O157 was detected in two (10%), one (5%), one (5%) of the thigh, drumstick and whole chicken samples, respectively. 5. Biofilm-forming ability was reported in 33 (94.2%) of 35 E. coli isolates, whilst the biofilm-associated genes detected among 35 STEC isolates included csgA (88.5%), fimH (88.5%), bcsA (85.7%), agn43 (14.2%) and papC (8.5%). The STEC strains showed resistance against ampicillin (88.5%) and erythromycin (88.5%), followed by tetracycline (74.2%) and gentamicin (25.7%). However, the distribution of isolates harbouring bla CMY, ere(A), tet(A) and aac(3)-IV antibiotic resistance genes was found to be 17.1%, 11.4%, 85.7% and 5.7%, respectively. 6. ERIC-PCR showed that E. coli strains obtained from different parts and whole of chicken samples had genetic diversities. ERIC-PCR patterns grouped strains of 35 STEC into eight clusters designated A-H, with 73% similarity. Proper hygiene measures and staff training are essential for public health during poultry processing and in retail stores to control STEC.

Genomic Diversity, Antimicrobial Resistance, and Virulence Gene Profiles of Salmonella Serovar Kentucky Isolated from Humans, Food, and Animal Ceca Content Sources in the United States.

Salmonella serovar Kentucky is frequently isolated from chickens and dairy cattle, but recovery from humans is comparatively low based on the U.S. National Antimicrobial Resistance Monitoring System (NARMS) reports. We aimed to better describe the genetic diversity, antimicrobial resistance, and virulence determinants of Salmonella Kentucky isolates from humans, food animal ceca, retail meat and poultry products, imported foods and food products, and other samples. We analyzed the genomes of 774 Salmonella Kentucky isolates and found that 63% (54/86) of human isolates were sequence type (ST)198, 33% (29/86) were ST152, and 3.5% (3/86) were ST314. Ninety-one percent (570/629) of cecal isolates and retail meat and poultry isolates were ST152 or ST152-like (one allele difference), and 9.2% (58/629) were ST198. Isolates from imported food were mostly ST198 (60%, 22/37) and ST314 (29.7%, 11/37). ST198 isolates clustered into two main lineages. Clade ST198.2 comprised almost entirely isolates from humans and imported foods, all containing triple mutations in the quinolone resistance-determining region (QRDR) that confer resistance to fluoroquinolones. Clade ST198.1 contained isolates from humans, ceca, retail meat and poultry products, and imported foods that largely lacked QRDR mutations. ST152 isolates from cattle had a lineage (Clade 2) distinct from ST152 isolates from chicken (Clade 4), and half of ST152 human isolates clustered within two other clades (Clades 1 and 3), largely distinct from Clades 2 and 4. Although clinical illness associated with Salmonella Kentucky is low, ST198 appears to account for most human infections in the Unites States but is uncommon among ceca of domestic food animals and retail meat and poultry products. These findings, combined with human exposure data, suggest that fluoroquinolone-resistant ST198 infections may be linked to the consumption of food products that are imported or consumed while traveling. We also found unique differences in the composition of virulence genes and antimicrobial resistance genes among the clades, which may provide clues to the host specificity and pathogenicity of Salmonella Kentucky lineages.

Effects of Sublethally Injured Campylobacter jejuni in Mice.

ABSTRACTGlobally, Campylobacter spp. are the most common food-associated bacterial cause of human gastrointestinal disease. Campylobacteriosis is primarily associated with the consumption of contaminated chicken meat. Chemical decontamination of chicken carcasses during processing is one of the most effective interventions to mitigate Campylobacter contamination. Following exposure to sanitizers, however, sublethally injured populations of bacteria may persist. The risk that sublethally injured Campylobacter pose for public health is unknown. Furthermore, the virulence potential of sublethally injured Campylobacter jejuni during prolonged storage in relation to host pathogenesis and the host immune response has not been well established. Therefore, we evaluated the effects of sublethally injured C. jejuni on the host, after storage in chicken meat juice. C57BL/6 mice were infected with two C. jejuni chicken meat isolates or the ATCC 33291 strain that had been stored in the chicken meat juice, after exposure to chlorine or acidified sodium chlorite (ASC). Although chlorine exposure was unable to reduce intestinal colonization by C. jejuni, exposure to ASC significantly reduced the intestinal colonization and tissue translocation in mice. The expression of pro- and anti-inflammatory cytokine genes for interleukin-6 (IL-6), IL23a, and IL-10, Toll-like receptor 2 (TLR2) and TLR4 genes, and host stress response genes (CRP, MBL1, and NF-κB1) were significantly reduced following the exposure to ASC. Our results demonstrated that sublethally injured C. jejuni has reduced virulence potential and colonization in mice. The data contribute toward clarification of the importance of chemical decontamination during processing to minimize human campylobacteriosis.IMPORTANCE Campylobacter is the most common cause of bacterial gastrointestinal disease, and consumption of contaminated poultry is frequently identified as the source of bacteria. The survivability and virulence potential of sublethally injured Campylobacter following exposure to chemicals which are commonly used to eliminate Campylobacter during the poultry meat processing are of concern to the food industry, government health officials, and consumers. Here, we demonstrate that sublethally injured Campylobacter jejuni has reduced bacterial virulence and colonization potential in mice.

Analysis of 90 Listeria monocytogenes contaminated in poultry and livestock meat through whole-genome sequencing

Listeria monocytogenes is a food-borne pathogen that can cause human listeriosis. The main cause of L.monocytogenes poisoning is the consumption of contaminated or processed foods, of which meat is the main one. At present, it is not clear whether the listeria contaminated in different meatis genetically diverse. In this study, a total of 90 L.monocytogenes isolates including 28 isolates in livestock meat and 62 isolates in poultry meat were obtained from chicken, duck, pork, and beef respectively in Beijing. And whole-genome sequencing (WGS) based analysis was performed to investigate the genetic diversity of L.monocytogenes between poultry and livestock meat isolates. As a result, we found the prevalence of the multilocus sequence typing (MLST)-derived clonal complexes (CCs) were quite different between two different types of meat. The number of resistance genes and virulence genes in most of the L.monocytogenes isolated from the four meat species were close, but the sequences of eight resistance genes and 19 virulence genes were significantly varied between poultry meat and livestock meat. The phylogenetic and collinear analysis of these isolates revealed the isolates in the same meat have a similar genome. These results showed that the genetic variation of L.monocytogenes in livestock and poultry meat were significantly different isolates.

Genomic Analysis of Multidrug-Resistant Salmonella enterica Serovar Kentucky Isolates from Humans, Turkey, and Food in the Republic of Serbia.

Owing to the emerging resistance to antimicrobials in Salmonella Kentucky isolates around the globe, the genomic comparison of all the registered multidrug-resistant Salmonella Kentucky isolates in Serbia (five from humans, one from turkey flock, and one from meat) was done. Most of the isolates were isolated from patients returning from Egypt or Tunisia or originated from imported turkey flock and turkey meat. The comparative analysis of resistance and virulence genes was done. All isolates belonged to sequence type-ST198 and were resistant to ciprofloxacin (Cip). The resistance to Cip was mediated by target mutations of the gyrA and parC genes, which encode topoisomerase I and II, respectively. Multidrug-resistant phenotype to aminoglycosides, β-lactam antibiotics, sulfonamides, and tetracyclines was detected in five isolates. However, none of the isolates was pan-resistant to antimicrobials. The number of single nucleotide polymorphisms between isolates varied from 8 to 43 and phylogenomics revealed the genetic proximity of the human isolate 10475/11 and the turkey meat isolate 5264/14, indicating a possible meat-to-human transfer. All isolates belonged to the main Salmonella Kentucky MDR lineage, carrying the Salmonella genomic island 1 (SGI1-K) subtype. The SGI1-K of Serbian isolates showed mosaicism attributed to rapid intraclonal evolution. Many virulence factors were detected in all the isolates, including SPI-1, SPI-2, SPI-3, SPI-4, SPI-5, SPI-9, and C63PI. Although Salmonella Kentucky has rarely been isolated from humans, food, and animals in Serbia, further surveillance is needed to diminish the risk of the spreading of resistant clones and their meat-to-human transmission.

Prevalence, species diversity, and antimicrobial susceptibility of Campylobacter strains in patients with diarrhea and poultry meat samples: one-year prospective study.

Source tracking of antimicrobial resistance in Campylobacter is useful for control measures. In this study, Campylobacter-associated diarrhea and homology in antimicrobial resistance of humans and poultry meat isolates were investigated. A total of 400 stools of patients and 100 poultry meat samples were analyzed. Susceptibility of the isolates was detected by disk diffusion, Etest, and agar dilution methods. Mismatch amplification mutation assay was used for the detection of mutations in the gyrA quinolone resistance determining region (QRDR). Campylobacter spp., including C. jejuni, C. coli, and C. lari, were detected in 35% of the chicken meat and 6.75% of the stool samples, respectively. The QRDR mutation was detected in most of the stool and chicken meat samples. Although the frequency of resistance to tetracycline (53.5% and 62.8%), erythromycin (39.2% and 37.1%), and gentamicin (32.1% and 31.4%) was relatively similar, higher frequency of resistance to ciprofloxacin (51.4% vs 28.6%) and nalidixic acid (42.15% vs 28.6%) among the chicken meat, and ampicillin (50% and 17.1%) among the human stool was detected. High percentage of poultry meat samples is contaminated with different Campylobacter species, which shows homology with the patients' isolates in Tehran.

Whole-Genome Sequencing Provides Insight Into Antimicrobial Resistance and Molecular Characteristics of Salmonella From Livestock Meat and Diarrhea Patient in Hanzhong, China.

Salmonella is a major zoonotic pathogen, which usually contaminates food resulting in salmonellosis in humans. Exploring the characteristics and origins of Salmonella is essential in formulating prevention and control measures for Salmonella infection. We used slide agglutination, antimicrobial susceptibility testing, and whole-genome sequencing to analyze and compare Salmonella’s phenotype, genotyping diversity, and genetic relatedness from livestock meat and diarrhea patients in Hanzhong, China, from 2018 to 2020. Totally 216 Salmonella enterica isolates were screened from frozen whole chicken carcasses (44.3%, 70/158), frozen raw ground pork (36.2%, 59/163), and diarrhea patients (4.4%, 87/1964). Salmonella Typhimurium was the dominant serotype. Notably, compared with other sources, isolates obtained from frozen whole chicken carcasses showed significant resistance to third-generation cephalosporin and fluoroquinolones (p < 0.05). All strains were assigned into 36 sequence types (STs) and two novel STs, and an excellent consistency was observed between ST and serotype. Genomic data revealed that extended-spectrum β-lactamase genes were responsible for third-generation cephalosporin resistance in 52 Salmonella strains, and the most predominant resistance determinant was blaCTX–M. Furthermore, of the 60 ciprofloxacin-resistant isolates, five single-base mutations in quinolone resistance-determining regions were identified in gyrA or parC, and the plasmid-mediated quinolone resistance gene aac(6’)Ib-cr was most often detected. The cgMLST clusters show that five clusters among four serotypes (including S. Typhimurium, S. London, S. Derby, and S. Agona) cover samples from diarrhea patients and livestock meat pathway isolate, indicating a possibility of cross-host transmission. In conclusion, the livestock meat isolates have a higher level of resistance than diarrhea patients’ isolates and could be an essential source of human Salmonella infection.

Antibiotic Resistance in Campylobacter spp. Isolated from Broiler Chicken Meat and Human Patients in Estonia.

Poultry meat is considered the most important source of Campylobacter spp. Because of rising antimicrobial resistance in Campylobacter spp., this study investigated the antimicrobial resistance of Campylobacter isolates from fresh broiler chicken meat originating from the Baltic countries sold in Estonian retail settings. Additionally, human clinical isolates obtained from patients with Campylobacter enteritis in Estonia were analysed. The aim of this study was to investigate the susceptibility of Campylobacter spp. to nalidixic acid, ciprofloxacin, tetracycline, streptomycin, erythromycin and gentamicin. The broth microdilution method with the EUCAMP2 panel was used for MIC determination and antimicrobial mechanisms were analysed using WGS data. A total of 46 Campylobacter strains were analysed, of which 26 (42.6%) originated from Lithuanian, 16 (26.2%) from Latvian, and 4 (6.6%) from Estonian fresh broiler chicken meat. In addition, 15 (24.6%) Campylobacter strains of patients with campylobacteriosis were tested. The antimicrobial resistance patterns of Campylobacter spp. isolated from fresh broiler chicken meat samples of Estonian, Latvian and Lithuanian origin collected in Estonian retail, and from patients with Campylobacter enteric infections, were determined. A total of 46 (75%) of the isolates tested were C. jejuni and 15 (25%) were C. coli. Campylobacter resistance was highest to nalidixic acid (90.2% of strains) and ciprofloxacin (90.2%), followed by tetracycline (57.4%), streptomycin (42.6%) and erythromycin (6.6%). All strains were sensitive to gentamicin. Additionally, antimicrobial resistance genes and point mutations were detected in 27 C. jejuni and 8 C. coli isolates previously assigned as resistant with the phenotypic method. A high antibiotic resistance of Campylobacter spp. in Lithuanian- and Latvian-origin broiler chicken meat and Estonian clinical isolates was found. Similar antibiotic resistance patterns were found for broiler chicken meat and human Campylobacter isolates.

Characterization of Staphylococcus aureus biofilms via crystal violet binding and biochemical composition assays of isolates from hospitals, raw meat, and biofilm-associated gene mutants

Staphylococcus aureus (SA) is a gram-positive coccus and an opportunistic pathogen of humans. The ability of SA to form biofilms is an important virulence mechanism because biofilms are protected from host immune responses and antibiotic treatment. This study examines the relative biofilm strength of a variety of hospital and meat-associated strains of SA, using a crystal violet (CV) staining assay. Biofilms were treated with either DNase or proteinase K prior to CV staining, and compared to mock-treated results, to better understand the biochemical composition. Biofilm polysaccharide concentration was also measured using the phenol sulfuric-acid assay which was normalized to base biofilm strength. We found that hospital-associated isolates have biofilms that bind significantly more CV than for meat isolates and are significantly more protein and polysaccharide-based while meat isolates have significantly more DNA-based biofilms. This study also investigates the effects that biofilm-related genes have on biofilm formation and composition by analyzing specific transposon mutants of genes previously shown to play a role in biofilm development. agrA, atl, clfA, fnbA, purH, and sarA mutants produce significantly weaker biofilms (bind less CV) as compared to a wild-type control, whereas the acnA mutant produces a significantly stronger biofilm. Biofilms formed from these mutant strains were treated (or mock-treated) with DNase or proteinase K and tested with phenol and sulfuric acid to determine what role these genes play in biofilm composition. The acnA, clfA, fnbA, and purH mutants showed significant reduction in biofilm staining after either proteinase K or DNase treatment, agrA and sarA mutants showed significant biofilm reduction after only proteinase K treatment, and an atl mutant did not show significant biofilm reduction after either proteinase K or DNase treatment. These data suggest that biofilms that form without acnA, clfA, fnbA, and purH are DNA- and protein-based, that biofilms lacking agrA and sarA are mainly protein-based, and biofilms lacking atl are mainly polysaccharide-based. These results help to elucidate how these genes affect biofilm formation and demonstrate how mutating biofilm-related genes in SA can cause a change in biofilm composition.

Comparative Analysis of L-Fucose Utilization and Its Impact on Growth and Survival of Campylobacter Isolates

Campylobacter jejuni and Campylobacter coli were previously considered asaccharolytic, but are now known to possess specific saccharide metabolization pathways, including L-fucose. To investigate the influence of the L-fucose utilization cluster on Campylobacter growth, survival and metabolism, we performed comparative genotyping and phenotyping of the C. jejuni reference isolate NCTC11168 (human isolate), C. jejuni Ca1352 (chicken meat isolate), C. jejuni Ca2426 (sheep manure isolate), and C. coli Ca0121 (pig manure isolate), that all possess the L-fucose utilization cluster. All isolates showed enhanced survival and prolonged spiral cell morphology in aging cultures up to day seven in L-fucose-enriched MEMα medium (MEMαF) compared to MEMα. HPLC analysis indicated L-fucose utilization linked to acetate, lactate, pyruvate and succinate production, confirming the activation of the L-fucose pathway in these isolates and its impact on general metabolism. Highest consumption of L-fucose by C. coli Ca0121 is conceivably linked to its enhanced growth performance up to day 7, reaching 9.3 log CFU/ml compared to approximately 8.3 log CFU/ml for the C. jejuni isolates. Genetic analysis of the respective L-fucose clusters revealed several differences, including a 1 bp deletion in the Cj0489 gene of C. jejuni NCTC11168, causing a frameshift in this isolate resulting in two separate genes, Cj0489 and Cj0490, while no apparent phenotype could be linked to the presumed frameshift in this isolate. Additionally, we found that the L-fucose cluster of C. coli Ca0121 was most distant from C. jejuni NCTC11168, but confirmation of links to L-fucose metabolism associated phenotypic traits in C. coli versus C. jejuni isolates requires further studies.

Characterization of Salmonella enterica Contamination in Pork and Poultry Meat from São Paulo/Brazil: Serotypes, Genotypes and Antimicrobial Resistance Profiles.

Salmonellosis is a zoonosis of major relevance to global public health. Here we present the assessment of Salmonella enterica contamination in pork and poultry meat sold at retail markets in São Paulo, Brazil. A total of 780 meat samples (386 poultry meat and 394 pork samples) were collected from 132 markets. From these, 57 samples (7.3%) were positive for S. enterica isolation, including 32 (8.3%) poultry meat and 25 (6.3%) pork samples. S. enterica isolates were further characterized for serotyping, antimicrobial resistance and genotyping by amplified fragment length polymorphism and pulsed field gel electrophoresis. Antimicrobial resistance analysis demonstrated two main profiles: pork isolates were more resistant to macrolides, β-lactams, tetracycline, phenicols, and fluoroquinolones, and poultry meat isolates presented higher resistance to fluoroquinolones, sulfonamides, tetracycline, and β-lactams. A total of 72.4% of poultry meat isolates were identified as S. Heidelberg, while most of pork isolates were S. Typhimurium (31.7%) and S. Give (16.7%). Genotyping resulted in most clusters consisting exclusively of pork or poultry meat, no cross-contamination was detected, and a tendency to differentiate isolates according to their serotypes and markets of origin. High resistance rates to critically important antimicrobials reinforce the importance of controlling Salmonella contamination in meat production chains.