High prevalence and diversity of Bartonella in small mammals from the biodiverse Western Ghats

Abstract

Bartonella species are recognized globally as emerging zoonotic pathogens. Small mammals such as rodents and shrews are implicated as major natural reservoirs for these microbial agents. Nevertheless, in several tropical countries, like India, the diversity of Bartonella in small mammals remain unexplored and limited information exists on the natural transmission cycles (reservoirs and vectors) of these bacteria. Using a multi-locus sequencing approach, we investigated the prevalence, haplotype diversity, and phylogenetic affinities of Bartonella in small mammals and their associated mites in a mixed-use landscape in the biodiverse Western Ghats in southern India. We sampled 141 individual small mammals belonging to eight species. Bartonella was detected in five of the eight species, including three previously unknown hosts. We observed high interspecies variability of Bartonella prevalence in the host community. However, the overall prevalence (52.5%) and haplotype diversity (0.9) was high for the individuals tested. Of the seven lineages of Bartonella identified in our samples, five lineages were phylogenetically related to putative zoonotic species-B. tribocorum, B. queenslandensis, and B. elizabethae. Haplotypes identified from mites were identical to those identified from their host species. This indicates that these Bartonella species may be zoonotic, but further work is necessary to confirm whether these are pathogenic and pose a threat to humans. Taken together, these results emphasize the presence of hitherto unexplored diversity of Bartonella in wild and synanthropic small mammals in mixed-use landscapes. The study also highlights the necessity to assess the risk of spillover to humans and other incidental hosts.

Fig 1. Map of the study area and sampling sites.

(A) World map with India marked inside the red box. (B) Map of India with the Karnataka state and the Kadumane landscape marked. (C) The enlarged land-use land cover map of Kadumane. Red circles indicate sampling locations. The shapefile for the world was downloaded from DIVA-GIS (http://www.diva-gis.org/gdata) and the shapefile for India from Community Created Maps of India (http://projects.datameet.org/maps/). The land-use map of Kadumane was created by Nature Conservation Foundation. All layers are in WGS84 geographic coordinate system and were processed and plotted using QGIS3.10 (https://qgis.org/en/site/), an open-access software.

Fig 2. Prevalence of Bartonella in the four most common small mammal species.

This figure summarizes our predictions about the high prevalence of Bartonella in the small mammal community. The dots represent calculated prevalence and the bar represents 95% confidence intervals. The sample size of the other species was too low to calculate confidence intervals. The silhouette images of animals were downloaded from PhyloPic (http://phylopic.org), an open-access database that stores reusable silhouette images of organisms.

Fig 3. Haplotype diversity.

(A) Haplotype network showing the relationship among Bartonella sequences from small mammal host species in Kadumane, South India. An integer neighbor joining network was constructed using 657 bp of rpoB using PopArt. The numbers between nodes indicate the number of nucleotide differences between haplotypes. (B) The haplotype accumulation curve (solid line) shows the cumulative haplotype richness (rpoB) sampled across 63 positive small mammals. The extrapolated richness (dotted lines) saturates around the estimated requirement to sample 160 positive individuals to detect all possible haplotypes in this community.

Fig 4. Bayesian phylogenetic inference of Bartonella based on 773 bp of rpoB sequences.

Sequences from this study, zoonotic Bartonella and other Bartonella are colored purple, brown, and black respectively. Nodes are colored based on posterior probability and monophyletic clades are collapsed for easy visualization. The tree was rooted with one representative of Brucella melitensis. The original phylogenetic tree and accession numbers (MT787669-MT787733) are provided in the S1 Fig and S2 Table respectively. The silhouette images of animals were downloaded from PhyloPic (http://phylopic.org), an open-access database that stores reusable silhouette images of organisms.