Incidence and molecular characterization of ESBL-producing and colistin-resistant Escherichia coli isolates recovered from healthy food-producing animals in Pakistan.

OBJECTIVES: To investigate the occurrence and molecular features of ESBL-producing and colistin-resistant Escherichia coli isolates recovered from healthy food-producing animals in Pakistan. METHODS: A total of 153 E. coli isolates were recovered from 250 faecal samples collected from livestock and poultry. The antibiotic susceptibility, resistant determinants and mobile genetic elements were determined for all the isolates. The clonal relatedness was analysed by MLST. Plasmids harbouring, localization and transferability of mcr-1 gene were carried out by Southern hybridization, S1-PFGE and transconjugation. RESULTS: Out of 153 E. coli strains, 49.01% isolates were ESBLs producers, whereas 18.95% were resistant to colistin and 84.31% of the isolates. Multidrug resistance was found in 84% of the isolates. The ESBL-producing E. coli in buffaloes, cattle, sheep, goat and broilers faecal samples were 60%, 74%, 54%, 50% and 68%, respectively. Among the ESBLs genes, blaCTX-M was the most prevalent group detected in 98.66%, while only mcr-1 of the colistin-resistant genes could be PCR amplified in 29 isolates. The common MGEs found were ISECP1 (35.13%), ISCR1 (33.78%), ISApl1 (20.27%) and Inti1 (58.10%). The most predominant Inc. types found were IncFIB 46.66%, followed by IncFIA 30.66%, IncFIC 26.66%, IncFrepB 26.66%, IncHI2 26.66%, IncP 22.66% and IncX4 21.33%. The most frequent sequence type detected was ST58. Southern blot and S1-PFGE confirmed the plasmid harbouring of mcr-1 gene. CONCLUSION: The co-occurrence of mcr-1 and ESBLs-encoding genes, along with MGEs in E. coli from healthy food animals in Pakistan, is a major concern. SIGNIFICANCE AND IMPACT OF STUDY: Antimicrobial resistance can be transferred from animals to humans by direct contact or via the food chain and environment. The prevalence and co-occurrence of ESBL and colistin resistance genes from food-producing animals is rare in Pakistan. To our knowledge, this is the first report to find ESBLs and mcr-1-harbouring E. coli from the faecal samples of the healthy food-producing animals in Pakistan. The presence of ARGs in association with MGEs, co-harbouring the virulence factors, as determined in the current study, is a severe threat to livestock and the human community as it has horizontally and food web transferability.

Microbiomes and Resistomes in Biopsy Tissue and Intestinal Lavage Fluid of Colorectal Cancer.

Aim: The gut microbiome plays a crucial role in colorectal cancer (CRC) tumorigenesis, but compositions of microorganisms have been inconsistent in previous studies due to the different types of specimens. We investigated the microbiomes and resistomes of CRC patients with colonic biopsy tissue and intestinal lavage fluid (IVF). Methods: Paired samples (biopsy tissue and IVF) were collected from 20 patients with CRC, and their gut microbiomes and resistomes were measured by shotgun metagenomics. Clinical and laboratory data were recorded. Bioinformatics (KneadData, Kraken2, and FMAP) and statistical analysis were done using the R (v4.0.2) software. Results: Bacterial diversity in IVF was higher than in tissue samples, and bacterial operational taxonomic units (OTUs) were 2,757 in IVF vs. 197 in tissue. ß-diversity showed distinct clusters in paired samples. The predominant bacteria in IVF were phylum Proteobacteria, while the predominant bacteria of tissue were phylum Actinobacteria. Twenty-seven representative bacteria were selected to form six bacterial clusters, which showed only Firmicutes Cluster 1, and the Bacteroidetes Cluster 1 were significantly more abundant in the IVF group than those in the tissue group (p < 0.05). The Firmicutes Cluster 2, Bacteroidetes Cluster 2, Pathogen Cluster, and Prevotella Cluster were not significantly different between IVF and tissue (p > 0.05). Correlation analysis revealed that some bacteria could have effects on metabolic and inflammatory parameters of CRC patients. A total of 1,295 antibiotic resistance genes (ARGs) were detected in the gut microbiomes, which conferred multidrug resistance, as well as resistance to tetracycline, aminoglycoside, and more. Co-occurrence patterns revealed by the network showed mainly ARG-carrying bacteria to be similar between IVF and tissue, but leading bacteria located in the hub differed between IVF and tissue. Conclusion: Heterogeneity of microbiota is particularly evident when studied with IVF and tissue samples, but bacterial clusters that have close relationships with CRC carcinogenesis are not significantly different, using IVF as an alternative to tissue for gut microbiome, and resistome assessment may be a feasible method.