In this study, eight Escherichia coli isolates were obtained from milk samples of dairy cattle suffering from clinical/subclinical mastitis. Isolates were characterized for antimicrobial resistance traits and virulence genes. Results revealed that one isolate was harbouring New Delhi metallo-beta-lactamase gene (blaNDM ). Cloning and sequencing of the PCR amplicon confirmed the identity of the gene (GenBank accession no. KC769583) having 100% homology with blaNDM-5 (GenBank accession no. JN104597.1), and this isolate was susceptible to colistin, chloramphenicol and tetracycline only. Moreover, another isolate carried extended-spectrum beta-lactamase (ESBL) gene - blaCTX-M , and all isolates possessed blaTEM gene. Of the eight isolates, only one isolate was positive for shiga toxin gene (stx2), and none were harbouring stx1 gene. Occurrence of New Delhi metallo-beta-lactamase (blaNDM ) in one E. coli isolate and ESBL genes in other isolates poses a potential threat to human health following possible entry and spread through food chain.

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Aeromonas species are important pathogens of fishes and aquatic animals capable of infecting humans and other animals via food. Due to the paucity of pan-genomic studies on aeromonads, the present study was undertaken to analyse the pan-genome of three clinically important Aeromonas species (A. hydrophila, A. veronii, A. caviae). Results of pan-genome analysis revealed an open pan-genome for all three species with pan-genome sizes of 9181, 7214 and 6884 genes for A. hydrophila, A. veronii and A. caviae, respectively. Core-genome: pan-genome ratio (RCP) indicated greater genomic diversity for A. hydrophila and interestingly RCP emerged as an effective indicator to gauge genomic diversity which could possibly be extended to other organisms too. Phylogenomic network analysis highlighted the influence of homologous recombination and lateral gene transfer in the evolution of Aeromonas spp. Prediction of virulence factors indicated no significant difference among the three species though analysis of pathogenic potential and acquired antimicrobial resistance genes revealed greater hazards from A. hydrophila. In conclusion, the present study highlighted the usefulness of whole genome analyses to infer evolutionary cues for Aeromonas species which indicated considerable phylogenomic diversity for A. hydrophila and hitherto unknown genomic evidence for pathogenic potential of A. hydrophila compared to A. veronii and A. caviae.

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Campylobacter is a major cause of foodborne illnesses worldwide. Campylobacter infections, commonly caused by ingestion of undercooked poultry and meat products, can lead to gastroenteritis and chronic reactive arthritis in humans. Whole genome sequencing (WGS) is a powerful technology that provides comprehensive genetic information about bacteria and is increasingly being applied to study foodborne pathogens: e.g., evolution, epidemiology/outbreak investigation, and detection. Herein we report the complete genome sequence of Campylobacter coli strain YH502 isolated from retail chicken in the United States. WGS, de novo assembly, and annotation of the genome revealed a chromosome of 1,718,974 bp and a mega-plasmid (pCOS502) of 125,964 bp. GC content of the genome was 31.2% with 1931 coding sequences and 53 non-coding RNAs. Multiple virulence factors including a plasmid-borne type VI secretion system and antimicrobial resistance genes (beta-lactams, fluoroquinolones, and aminoglycoside) were found. The presence of T6SS in a mobile genetic element (plasmid) suggests plausible horizontal transfer of these virulence genes to other organisms. The C. coli YH502 genome also harbors CRISPR sequences and associated proteins. Phylogenetic analysis based on average nucleotide identity and single nucleotide polymorphisms identified closely related C. coli genomes available in the NCBI database. Taken together, the analyzed genomic data of this potentially virulent strain of C. coli will facilitate further understanding of this important foodborne pathogen most likely leading to better control strategies. The chromosome and plasmid sequences of C. coli YH502 have been deposited in GenBank under the accession numbers CP018900.1 and CP018901.1, respectively.

Aims:  The aim of the study was to characterize 16S rDNA of Aeromonas spp. to rapidly identify clinically important species of these bacteria. Methods and Results:  Sequence analysis of published 16S rDNA for unique restriction sites revealed prospect of species identification. Extraction of genomic DNA followed by amplification and step‐by‐step restriction endonuclease digestion of 16S rDNA was able to identify Aeromonas spp. of medical significance. Validation of the method was performed by subjecting 53 Aeromonas strains of multiple origin to similar treatment. Results of the study were in agreement with corresponding species of the isolates. Conclusions:  The method developed offers an easily interpretable tool for the identification of Aeromonas spp. of clinical relevance. Significance and Impact of the Study:  The developed methodology should facilitate routine laboratory diagnosis of Aeromonas spp. from clinical cases to species level.

Aim: To investigate the incidence and virulence properties of E. coli in fresh fish and ready-to-eat fish products from retail markets of the Ludhiana the present study was conducted. Materials and Methods:Total of 184 samples comprising 96 raw fish and 88 ready-to-eat (RTE) fish products were collected from Ludhiana and other parts of Punjab and were subjected to suitable microbiological methods for E. coli isolation. E. coli isolates were subjected for haemolytic activity and indicators of plausible cytotoxicity (lecithinase, protease and gelatinase production), congo red dye biding assay. To assess virulence potential isolates were molecularly screened for stx 1 and 2 genes.Results: From raw fish samples 47(48.95%), E. coli, were isolated. From RTE fish products 7(12.96%), E. coli were isolated. Overall incidence for E. coli was 54 (29.34%). In vitro virulence characterization of isolates exhibited that all E. coli isolates were haemolytic while indicators of plausible cytotoxicity ( lecithinase, protease and gelatinase production) were in the range of 16.67% to 35.19% indicated that though the isolates were haemolytic they were perhaps less likely to be cytotoxic. Congo Red binding assay for E. coli isolates revealed that majority (88.89%) of the isolates failed to uptake the dye and only few (11.11%) could bind the dye. Results of serotyping revealed a total of 15 different serotypes among the E. coli isolates. More variation was observed among isolates from raw fish samples (12 serotypes) while RTE fish products harboured only 5 different serotypes. Molecular characterization of E. coli isolates revealed that PCR screening of isolates revealed that total 39 (72.22%) samples out of 54 E. coli isolates were positive for stx1 gene and 28 (51.85%) of isolates were positive for stx2 gene. Sources wise, 36 (66.66%) of isolates from raw fish and 3(5.55%) of isolates from RTE fish products were positive for stx1 while and stx2 gene could be detected in 24(44.44%) isolates from raw fish and 4(7.4%) isolates from RTE fish products.Interestingly, about 20% (37.03%) isolates were positive for both stx1 and stx2 genes. Among these multivirulent isolates majority (n=18) belonged to raw fish samples compared to a few (n=2) from RTE fish products. Conclusion:The results of the present study highlighted the possible risks to consumers of fish and fish products in the region that demand action to address this public health concern.