Biofilm-forming antimicrobial-resistant pathogenic Escherichia coli: A one health challenge in Northeast India.

This study aimed to investigate the prevalence of Shiga toxin-producing Escherichia coli (STEC), Enteropathogenic E. coli (EPEC), and Enterotoxigenic E. coli (ETEC) in common food animals (cattle, goats, and pigs) reared by tribal communities and smallholder farmers in Northeast India. The isolates were characterized for the presence of virulence genes, extended-spectrum beta-lactamases (ESBL) production, antimicrobial resistance, and biofilm production, and the results were statistically interpreted. In pathotyping 141 E. coli isolates, 10 (7.09%, 95% CI: 3.45%-12.66%) were identified as STEC, 2 (1.42%, 95% CI: 0.17%-5.03%) as atypical-EPEC, and 1 (0.71%, 95% CI: 0.02%-3.89%) as typical-EPEC. None of the isolates were classified as ETEC. Additionally, using the phenotypic combination disc method (ceftazidime with and without clavulanic acid), six isolates (46.1%, 95% CI: 19.22%-74.87%) were determined to be ESBL producers. Among the STEC/EPEC strains, eleven (84.6%, 95% CI: 54.55%-98.08%) and one (7.7%, 95% CI: 0.19%-36.03%) strains were capable of producing strong or moderate biofilms, respectively. PFGE analysis revealed indistinguishable patterns for certain isolates, suggesting clonal relationships. These findings highlight the potential role of food animals reared by tribal communities and smallholder farmers as reservoirs of virulent biofilm-forming E. coli pathotypes, with implications for food contamination and zoonotic infections. Therefore, monitoring these pathogens in food animals is crucial for optimizing public health through one health strategy.

Isolation and sero-genomo-epidemiological studies on Brucella infection in dairy cattle in Meghalaya, India.

In this study, we report the serological, bacteriological and whole genome sequencing data of a 6 years study of Brucella abortus in Meghalaya, India. Investigation of 3060 sera samples indicated overall prevalence of 6.4% by Rose Bengal Plate Test and 10.7% by ELISA. Considerably higher prevalence was observed among milk samples (17.5%, n = 362) and in blood samples (37.7%, n = 262) by direct PCR. Clinical samples (n = 94) from late abortion cases yielded 11 B. abortus isolates. Multi-locus sequence typing indicated circulation of single sequence type, ST1. Whole genome sequencing (n = 8) and phylogenomic analysis revealed close clustering of majority of isolates in two clusters alongwith genomes from other countries, indicating global relatedness among B. abortus. Taken together, the results of our study revealed the putative hotspot of infection in the dairy-dominant districts of the state and also calls for concerted One Health based action for prevention and control of this zoonotic disease.

Erratum to "Development of a novel polymerase spiral reaction (PSR) assay for rapid and visual detection of Clostridium perfringens in meat" [Heliyon 7 (1) (January 2021) Article e05941].

[This corrects the article DOI: 10.1016/j.heliyon.2021.e05941.].

Development of a novel polymerase spiral reaction (PSR) assay for rapid and visual detection of Clostridium perfringens in meat.

C. perfringens is a widespread foodborne pathogen and one of the major concerns in the meat industry. There is a need for a simple, rapid and equipment free detection system for C. perfringens as conventional anaerobic culture method is labour and resource intensive. Here, we applied a novel polymerase spiral reaction phenomenon to develop and evaluate an assay for effortless and visual detection of C. perfringens in meat foods employing pork as a representative model. Specificity of the assay was determined using 51 C perfringens and 20 non- C. perfringens strains. Analytical sensitivity of the developed test was 80 fg DNA per tube indicating 100 times more sensitivity than end-point PCR assay. The detection limits were 980 CFU/g and 9.8 × 104 CFU/g of pork for PSR and PCR assays, respectively. The operation time of the PSR assay including DNA extraction was 120 min. The developed PSR assay was accurate and effective in comparison to culture method, in detecting C. perfringens in 38 of 74 pork samples. Therefore the specificity, sensitivity, negative predictive value, positive predictive value and accuracy rate of the developed PSR assay were 100%. The developed PSR assay is easy to perform, rapid, affordable, permitting sophisticated-equipment free amplification and naked eye interpretation. This is the initial report in which the PSR assay was optimized for the detection of C. perfringens.

Dietary flaxseed oil improve boar semen quality, antioxidant status and in-vivo fertility in humid sub-tropical region of North East India.

Environmental stress in the form of high temperature humidity index (THI) in tropical and sub-tropical region negatively affects semen quality and fertility of boar. Therefore, the present study was done to evaluate the effect of supplementing flaxseed oil (FLO) to boar's diet on its semen quality, antioxidant status, fatty acid composition of seminal plasma and fertility under sub-tropical climate. For this purpose, six Hampshire crossbreed (50% Hampshire and 50% Gunghroo) boars were divided into two groups i.e control (CON) and treatment (FLO). In FLO and CON group, flaxseed and vegetable oil, respectively, was top dressed at the rate of 3% in basal diets for each boar on daily basis for 16 weeks during monsoon season. A total of 60 ejaculates, comprising 30 ejaculates from each group (ten ejaculates from each boar) were collected. Semen samples were evaluated for sperm quality parameters (SQPs: motility, viability, abnormality, acrosomal integrity and Hypo-osmotic swelling test) and velocity attributes by computer assisted semen analysis (CASA) at fresh and after 72 h of preservation at 17 °C. Antioxidant (glutathione peroxidase; GPx, catalase; CAT, total antioxidant capacity; TAC and malondialdehyde; MDA) were analyzed in seminal plasma and serum. Fatty acid compositions of seminal plasma were analyzed by gas chromatography-mass spectrometry (GC-MS). In-vivo fertility study was also conducted. Reaction time and false mounts were significantly (p < 0.05) reduced in FLO group as compared to CON group. Semen quality parameters were significantly (p < 0.05) improved at fresh stage and after 72 h of liquid storage in FLO group as compared to CON group. Velocity attributes (VAP, VSL, VCL, ALH, BCF and LIN) were significantly (p < 0.05) higher in FLO group. Flaxseed oil supplementation significantly (p < 0.01) enhanced serum GPx and CAT concentration. Serum and seminal plasma MDA concentration decreased significantly (p < 0.01) in FLO group. Similarly, GPx, TAC and CAT were significantly (p < 0.01) elevated in seminal plasma of FLO group. The study revealed that feeding of flaxseed oil altered the fatty acid composition of seminal plasma and significantly (p < 0.05) improved the farrowing rate. In summary, flaxseed oil supplementation improved the semen quality parameters and fertility of boars in sub-tropical climate by improving the antioxidant capacity and altering the fatty acid composition of seminal plasma.

Molecular Characterization and Computational Modelling of New Delhi Metallo-ß-Lactamase-5 from an Escherichia coli Isolate (KOEC3) of Bovine Origin.

Emergence of antimicrobial resistance mediated through New Delhi metallo-ß-lactamases (NDMs) is a serious therapeutic challenge. Till date, 16 different NDMs have been described. In this study, we report the molecular and structural characteristics of NDM-5 isolated from an Escherichia coli isolate (KOEC3) of bovine origin. Using PCR amplification, cloning and sequencing of full blaNDM gene, we identified the NDM type as NDM-5. Cloning of full gene in E. coli DH5α and subsequent assessment of antibiotic susceptibility of the transformed cells indicated possible role of native promoter in expression blaNDM-5. Translated amino acid sequence had two substitutions (Val88Leu and Met154Leu) compared to NDM-1. Theoretically deduced isoelectric pH of NDM-5 was 5.88 and instability index was 36.99, indicating a stable protein. From the amino acids sequence, a 3D model of the protein was computed. Analysis of the protein structure elucidated zinc coordination and also revealed a large binding cleft and flexible nature of the protein, which might be the reason for broad substrate range. Docking experiments revealed plausible binding poses for five carbapenem drugs in the vicinity of metal ions. In conclusion, results provided possible explanation for wide range of antibiotics catalyzed by NDM-5 and likely interaction modes with five carbapenem drugs.

Expression analysis of genes responsible for amino acid biosynthesis in halophilic bacterium Salinibacter ruber.

The degeneracy of the genetic code allows for multiple codons to encode the same amino acid. However, alternative codons and amino acids are used unevenly among genes, a phenomenon termed codon-usage bias. Genes regulating amino acid biosynthesis of Salinibacter ruber, an extremely halophilic bacterium were studied in order to determine the synonymous codon usage patterns. Factors responsible for codon usage variation among the genes were investigated using codon usage indices and multi-variate statistical approach. Overall codon usage data analysis indicated that codons ending in G and/or C were predominant among the genes. Multi-variate statistical analysis showed that there was a single major trend in the codon usage variation among the genes, which had a strong positive correlation (r = 0.93, P < 0.01) with (G + C) content of the genes. Further, correlation analysis indicated that genes with higher expression level and showing a greater degree of codon usage bias were GC-rich and preferred codons with C or G nucleotides at the third position. A set of thirteen codons were identified through Chi-square test as optimal codons, which were preferred in highly expressed genes. It could be concluded that mutational bias had a profound effect on codon usage pattern. In addition, translational selections also operated with a proper balance, making the genes translationally more efficient. The frequency of these codons appeared to be correlated with the level of gene expression and might be a useful indicator in the case of genes (or open-reading-frames) whose expression levels are unknown.