Genetic diversity of pathogenic bacteria and incidence of selected antibiotic resistant genes in water used by eateries and open markets in Juja, Kenya.

Abstract

Water is an important bacterial habitat and a major avenue of dissemination of antibiotic- resistant bacteria between and among different environments. Contamination with antibiotic-resistant bacteria in water used in eateries and open markets poses a direct threat to human health. This is because antimicrobial resistance monitoring is not currently a routine standard for drinking water. This study aimed to investigate the occurrence of ARGs and variation in the composition of bacterial communities in eighty-two water samples collected from eateries and open markets in Juja. The purposeful sampling technique was used to collect water samples used in eateries and open markets in Wakulima, Gachororo, and Mwerevu in Juja. The samples were collected from water containers placed at the entrance of eateries, taps at the market entrance, and jerricans used by traders within the market to wash groceries. Special media of MacConkey, Triple Sugar Iron Medium, Thioglycollate, and Lauryl Tryptose Broth media were used for the isolation of Enterobacteriaceae, the detection and numeration of F. streptococci, the differentiation and enumeration of E. coli, as well as the detection of bacterial coliforms, respectively. Culture-dependent and independent techniques were used to profile the bacteria and detect antibiotic-resistant bacteria (ARBs) and antibiotic-resistant genes (ARGs) present in the samples. Metagenomics sequencing and analysis were applied to investigate the ARB genetic profiles. Data was analysed using R software and Quantitative Insights into Microbial Ecology (QIIME2, version 2021.4) software. Heatmaps showed the morphological and biochemical characterization relationships, antibiotic susceptibility, and detection of ARGs in the culturable bacteria. The DNA extracted from the eighty-two raw water samples was pooled into six tubes according to the site of collection: Wakulima Eateries (WE), Wakulima Open Market (WOM), Gachororo Eateries (GE), Gachororo Open Market (GOM), Mwerevu Eateries (ME), and Mwerevu Open Markets (MOM). There is an abundance of the bacterial phylum Proteobacteria (above 86%) across all the collected water samples from the open markets and eateries. The dominant genera belonging to Proteobacteria were: WE (Acinetobacter 44.180%), WOM (Duganella 28.201%), GE (Acinetobacter 15.189%), GOM (Acinetobacter 30.675%), ME (Acinetobacter 40.823%), and MOM (Curvibacter 48.785%). This study reported the presence of antibiotic-resistant priority pathogens from the WHO list: Priority 1 (critical), where Pseudomonas spp. was detected in samples ME (12.163%) and WOM (0.323%), Acinetobacter baumannii was detected in samples GE (2.975%) and WE (5.495%), and Enterobacteriaceae was detected in samples WE (0.125%), MOM (0.326%), WOM (0.875%), GE (0.452%), and ME(0.206%). The genus of Escherichia coli, Klebsiella, Raoultella, and Enterobacter dominate the Enterobacteriaceae. Priority 2 (high) pathogen detected was the genus Staphylococcus in samples MOM (1.826%) and GE (0.962%). The ARGs of qnrD and sul2 were detected in all six samples; int1 and FloR genes were present in five samples, while those of strB, catA, and blaTEM were found in one sample each using qualitative polymerase chain reaction (PCR) assays. The findings of this study form a critical reference point for policy development and rapid mitigation strategies to prevent a possible outbreak of water-borne diseases in the area. Key words: antibiotic-resistant bacteria (ARBs); antibiotic-resistant genes (ARGs); genetic diversity; pathogenic bacteria