Milk supply chain as a reservoir of antimicrobial-resistant Staphylococcus species.

Milk is a source of essential nutrients, but food safety across the milk supply chain has emerged as an integral part of food trade. Of the several food safety hazards, antimicrobial-resistant Staphylococcus species have emerged as one of the major microbial hazards with significant public health concerns. The present crosssectional study was undertaken with the objective to isolate Staphylococcus species from the milk supply chain, characterize isolates for antimicrobial resistance, and trace the origin of isolates using molecular techniques. Samples collected from the formal and informal milk supply chains showed prevalence of Staphylococcus species of 4.3% (n=720); isolates were identified as coagulase-positive (S. aureus 67.7% and S. intermedius 6.4%) and coagulase-negative (S. lentus 9.6%, S. sciuri 3.2%, S. xylosus 3.2%, S. schleiferi 3.2%, S. felis 3.2%, and S. gallinarum 3.2%) species. Staphylococcus isolates showed antimicrobial resistance to methicillin (32.2%), ß-lactam (41.9%), and macrolide-lincosamide-streptogramin B (3.2%). Staphylococcus isolates phenotypically resistant to methicillin also carried the mecA gene and displayed diverse pulsed field gel electrophoresis (PFGE) profiles, indicating their diverse origins in the milk supply chain. Based on the similarity of PFGE profile, the origin of one of the Staphylococcus isolates was traced to the soil in contact with milch cows. The findings of this study highlight the need for more comprehensive microbial risk analysis studies across the milk supply chain, capacity building, creation of awareness among stakeholders about the judicious use of antimicrobials, and protection of public health using a One-Health approach.

Assessment of Knowledge and Biosecurity Practices Related to Avian Influenza Among Poultry Workers in a District of South India.

CONTEXT: Avian influenza, commonly known as bird flu, is a contagious disease that affects both animals and humans, posing a significant threat to public health, animal welfare, and the economy. This study aims to evaluate the knowledge of avian influenza among poultry farmworkers and evaluate the biosecurity practices implemented on their farms. OBJECTIVE: The study's primary objective was to assess the knowledge regarding avian influenza among poultry farmworkers and the biosecurity practices they follow at the farm. DESIGN: Cross-sectional study. SETTING: The study was conducted in a district of South India. PARTICIPANTS: This study included 105 poultry farmworkers across 70 poultry farms in the district. MAIN OUTCOME MEASURES: Assessment of the participants' knowledge related to avian influenza, and the association between knowledge levels, demographic, and farm-related factors such as working experience in the poultry farm, type of poultry farm, type of poultry reared, and biosecurity practices. RESULTS: In the study, 90% of participants were aware of avian influenza, with 36% correctly identifying the virus as its cause, whereas 5% wrongly cited it to be a bacteria. Although 90% knew avian influenza was infectious, only 18% recognized its potential transmission to humans; however, 82% understood prevention methods. Participants with an education level beyond high school displayed significantly higher awareness (P < .05), emphasizing importance of the education. CONCLUSIONS: The study showed diverse awareness levels among poultry farmworkers regarding avian influenza, emphasizing gaps in the knowledge, particularly about its transmission to humans. This underscores the need for targeted awareness campaigns focusing on zoonotic risks to improve the level of understanding and implement effective preventive measures against avian influenza.

Carcass characteristics, meat quality, and nutritional profiles of Mithun (Bos frontalis) meat reared under a semi-intensive system.

The present study aimed to determine carcass characteristics, meat quality, nutrient profiles, and sensory characteristics of Mithun meat. Sixteen Mithun were selected and divided into four groups, MM-4 (male; n = 4; <4 years of age), MM-47 (male; n = 4; 4-7 years of age), MF-4 (female; n = 4; <4 years of age), MF-47 (female; n = 4; 4-7 years of age). Carcass characteristics showed that adult males (MM-47) have significantly higher (P < 0.05) live weight, carcass weight, and meat-to-bone ratio. Fat (%) was significantly higher (P < 0.05), and deboned meat (%) was lower in MF-4 and MF-47, while marginal differences were observed in bone (%), dressing percentage, and offal yield between groups. Visible marbling increased with age and varied from "slight" to "small" in all groups. Nutrient profiling revealed a significantly higher (P < 0.05) fat percentage and cholesterol in MF-4 and MF-47. Fatty acid profile, amino acid profile, water-soluble vitamins, and minerals content did not differ between groups. However, lysine and leucine (essential amino acids) and glutamic acid and aspartic acid (nonessential amino acids) were most abundant. Effect of age was significant (P < 0.05) on juiciness, tenderness, and connective tissue residue scores. In conclusion, results indicate mithun meat is nutrient-rich regardless of the animal's age or sex.

Non-diarrhoeic pigs as source of highly virulent and multidrug-resistant non-typhoidal Salmonella.

Food-producing animals act as reservoirs of non-typhoidal Salmonella (NTS) serovars with potential food safety and public health implications. The present cross-sectional study aimed at determining the prevalence of Salmonella serotypes in non-diarrhoeic pigs and characterizing the isolates using molecular tools. Salmonella isolates (n = 22) recovered from faecal samples of 194 randomly selected pigs were characterized for virulence and antimicrobial resistance and subtyped using XbaI-PFGE. The prevalence of Salmonella in apparently healthy non-diarrhoeic pigs was 11.3% (95%CI, 4.3-19.5%), with S. Weltevreden (81.8%) and S. Enteritidis (18.2%) being the serotypes detected. Salmonella isolates harboured virulence genes such as invA (100%), stn (100%), spvR/spvC (86.3%) and fimA (22.7%). Phenotypically, isolates showed sensitivity to chloramphenicol, levofloxacin and ciprofloxacin and resistance to tetracycline and ampicillin (100%), streptomycin (86.4%), amoxicillin-clavulanate (63.6%), cefotaxime (22.7%) and ceftriaxone (9.1%). Notably, 18.2% isolates were multidrug-resistant (≥ 3 antimicrobial class) with multiple antimicrobial resistance (MAR) index of 0.56-0.67 (18.2%), 0.44 (45.5%), 0.33 (31.8%) and 0.22 (4.5%). Genotypically, isolates carried various antibiotic resistance genes: ESBL (blaTEM and blaOXA), aminoglycoside (strA, strB and aadA1), sulphonamide (sul1, sul2 and dfrA1), tetracycline (tetA and tetB) and plasmid AmpC beta-lactamase (ACC, FOX, MOX, DHA, CIT and EBC). The present investigation emphasizes the epidemiological significance of PFGE typing in the detection of emerging strains of highly virulent and multidrug-resistant S. Weltevreden and S. Enteritidis in non-diarrhoeic pigs that pose serious public health implications in the pork supply chain environment. More extensive longitudinal study is warranted to provide epidemiological links between environmental reservoirs and animal and human infections in piggery settings.

Identification of sheep (Ovis aries) meat by alkaline lysis-loop mediated isothermal amplification technique targeting mitochondrial D-loop region.

Identification of meat species origin using reliable techniques is a critical requirement for ensuring label compliance, protection of consumer preference and prevention of fraudulence in the meat trade. Although a plethora of protein and DNA based meat species identification techniques are in vogue, need for rapid test suitable for under-resourced laboratories catering point-of-care (PoC) services was construed. Present study deals with development of rapid sheep (Ovis aries) meat identification technique using DNA extraction by alkaline lysis (AL) and loop-mediated isothermal amplification (LAMP) technique. The AL-LAMP specifically amplifies sheep-specific signal of mitochondrial D loop region under an isothermal temperature of 60 °C with an analytical sensitivity of 0.5 ng sheep DNA. The test was highly specific to sheep and performed well even in the presence of DNA of closely related meat animal species such as goat, cattle, buffalo and chicken. The novel primers designed for the AL-LAMP successfully detected sheep meat in raw and cooked meat samples heated up to 121 °C for 30 min. Sheep-specific AL-LAMP assay could detect 0.1% mutton-in-beef adulteration. Novel AL-LAMP assay being simple, rapid and reliable for sheep meat authentication in just 120 min; hence, it could be conveniently used by terminal laboratories engaged in rendering on-site or PoC services.

Studies on occurrence, characterisation and decontamination of emerging pathogenic Escherichia coli (STEC, ETEC and EIEC) in table eggs.

1. Escherichia coli is one of the most common facultative anaerobic species present in the gastrointestinal tract of animals and human beings. Usually they occur as commensals, but some serotypes can cause significant illnesses in humans as well as mammals and birds. 2. The occurrence of E. coli in different categories of table eggs collected from markets was evaluated. Isolates were analysed for the presence of virulence genes, antibiotic susceptibility pattern and efficacy of peracetic acid and chlorine for the purpose of decontaminating table eggs. 3. Significant differences were observed in the occurrence of E. coli between different groups viz. processed (cleaned, washed, sanitised and packed eggs), unprocessed (un-cleaned, un-sanitised and loose eggs) and free range (eggs obtained from backyard poultry) table eggs. Overall, E. coli occurred in table eggs at 28.6% with 22.9, 29.2 and 50.0% occurrence in processed, unprocessed and free-range table eggs, respectively. 4. A total of 24 isolates of E. coli were obtained and screened for virulence genes viz. STH, SLT1/2 and INVE genes. Of the 24 isolates recovered, 10 typeable isolates belonged to O141, O119, O9, O120 and O101 serotypes, while the remaining 14 were untypeable. Antibiograms of the isolates showed multiple antimicrobial resistance (MAR) index in the range of 0.13-0.40. 5. Peracetic acid (PAA) and chlorine (CL) were studied for their sanitisation efficacy; concentrations of 100 mg/kg of PAA and 200 mg/kg of CL completely inactivated E. coli over the egg surface and also resulted in 2.58 and 2.38 log reduction in total viable counts (TVC), respectively. 6. The presence of virulence-associated shiga-like toxin (SLT1/2) and invasion E (INVE) genes and antimicrobial resistance among the emerging serotypes of pathogenic E. coli isolated from table eggs has public health implications. It underscores the need to implement better management practices across the production systems and marketing channels to produce E. coli-free wholesome eggs for consumers.

Breed traceability of buffalo meat using microsatellite genotyping technique.

Although buffalo has emerged as a major meat producing animal in Asia, major research on breed traceability has so far been focused on cattle (beef). This research gap on buffalo breed traceability has impelled development and validation of buffalo breed traceability using a set of eight microsatellite (STR) markers in seven Indian buffalo breeds (Bhadawari, Jaffaarabadi, Murrah, Mehsana, Nagpuri, Pandharpuri and Surti). Probability of sharing same profile by two individuals at a specific locus was computed considering different STR numbers, allele pooling in breed and population. Match probabilities per breed were considered and six most polymorphic loci were genotyped. Out of eight microsatellite markers studied, markers CSSMO47, DRB3 and CSSM060 were found most polymorphic. Developed technique was validated with known and unknown, blood and meat samples; wherein, samples were genetically traced in 24 out of 25 samples tested. Results of this study showed potential applications of the methodology and encourage other researchers to address the problem of buffalo traceability so as to create a world-wide archive of breed specific genotypes. This work is the first report of breed traceability of buffalo meat utilizing microsatellite genotyping technique.

Authentication of beef, carabeef, chevon, mutton and pork by a PCR-RFLP assay of mitochondrial cytb gene.

Authentication of meat assumes significance in view of religious, quality assurance, food safety, public health, conservation and legal concerns. Here, we describe a PCR-RFLP (Polymerase Chain Reaction- Restriction Fragment Length Polymorphism) assay targeting mitochondrial cytochrome-b gene for the identification of meats of five most common food animals namely cattle, buffalo, goat, sheep and pig. A pair of forward and reverse primers (VPH-F & VPH-R) amplifying a conserved region (168-776 bp) of mitochondrial cytochrome-b (cytb) gene for targeted species was designed which yielded a 609 bp PCR amplicon. Further, restriction enzyme digestion of the amplicons with Alu1 and Taq1 restriction enzymes resulted in a distinctive digestion pattern that was able to discriminate each species. The repeatability of the PCR-RFLP assay was validated ten times with consistent results observed. The developed assay can be used in routine diagnostic laboratories to differentiate the meats of closely related domestic livestock species namely cattle from buffalo and sheep from goat.

Detection of pork adulteration by highly-specific PCR assay of mitochondrial D-loop.

We describe a highly specific PCR assay for the authentic identification of pork. Accurate detection of tissues derived from pig (Sus scrofa) was accomplished by using newly designed primers targeting porcine mitochondrial displacement (D-loop) region that yielded an unique amplicon of 712 base pairs (bp). Possibility of cross-amplification was precluded by testing as many as 24 animal species (mammals, birds, rodent and fish). Suitability of PCR assay was confirmed in raw (n = 20), cooked (60, 80 and 100 °C), autoclaved (121 °C) and micro-oven processed pork. Sensitivity of detection of pork in other species meat using unique pig-specific PCR was established to be at 0.1%; limit of detection (LOD) of pig DNA was 10 pg (pico grams). The technique can be used for the authentication of raw, processed and adulterated pork and products under the circumstances of food adulteration related disputes or forensic detection of origin of pig species.