Abstract:
High diarrheal disease burden remains an urgent concern in low to middle income countries, greatly affecting children under the age of 5 years old and those living with HIV and AIDS. The capital city Harare, Zimbabwe, has been at the center of recurrent typhoid and cholera outbreaks in the last few decades. Treatment of these diarrheal illnesses and other infectious diseases has also become increasingly difficult with the rapid rise of antimicrobial resistance in the country. Despite these major concerns, there is little understanding about the enteric pathogens and antimicrobial resistance that contribute to disease burden in Harare. Clinical surveillance provides advantages in characterizing select pathogens and their characteristics from patients, but this method is expensive, resource-intensive, and highly dependent on health-seeking behaviors and accessibility, providing limited understanding about pathogen distribution and community impacts. Environmental surveillance of wastewater can supplement these gaps because all residents on a sewage system contributes to the wastewater, providing simple, composite samples that can give an improved understanding about both pathogens and antimicrobial resistance in the community.
This study evaluated the effectiveness of environmental surveillance with shotgun metagenomics as a tool to characterize a wide range of enteric pathogens, antibiotic resistance genes (ARGs), and virulence factor genes (VFGs) in wastewater from Harare, Zimbabwe. Between April 19 and May 9, 2019, wastewater samples from three high density, low-income suburbs and three low density, high-income suburbs were collected and processed for next-generation sequencing (n=18). Diversity analyses of the cumulative metagenomes revealed a significant difference in alpha diversity (Shannon-Wiener diversity index) and no significant difference in beta diversity (Bray-Curtis dissimilarity distance) between samples from high-income suburbs and low-income suburbs. The top enteric pathogens detected in all wastewater samples include Arcobacter spp. and Aeromonas spp., common bacteria found in environmental water and associated with mild to moderate gastroenteritis. Pathogens of high antimicrobial resistance (AMR) and clinical concern, including Staphylococcus aureus, Pseudomonas aeruginosa, and Salmonella enterica, were detected in all wastewater samples. From 510 predicted ARGs, we identified 139 unique ARGs across all samples. Of the total number of predicted ARGs, 66 were chromosomal and 212 were plasmid, suggesting the majority of ARGs are on mobile elements. Out of 3580 curated genes within 1381 VFs in the Virulence Factor Database, we detected 412 genes within 50 VFs in the samples. The top three virulence factor function classes were delivery, adherence, and motility, which play a major role in toxin secretion, host cell colonization, immune modulation, and cell survival. The findings provide a foundation for future studies to explore the potential of environmental surveillance and shotgun metagenomics as a public health monitoring tool for enteric disease.