Characterization of Bacillus anthracis from anthrax outbreaks in Kruger National Park (2014-2016) and the role of vultures in dissemination

Abstract

Background: Bacillus anthracis is the causal agent of anthrax. Dissemination aspects of this well-known zoonotic diseases are poorly understood. In South Africa, anthrax is endemic in the Ghaap Plateau and Northern Kruger National Park (KNP). The aim of this study was to use multiloci variable number of tandem repeat (VNTR) analysis (MLVA) to determine genetic diversity and track the distribution of B. anthracis isolated during the 2014 - 2018 outbreaks. In addition to the genotyping; camera traps and GPS collars were used on vultures to investigate their dissemination role in the, environment. Material and Methods: Bacillus anthracis isolates (n=62) were sampled from carcasses and the environment (n=38) during outbreaks in the Pafuri and Singita regions of KNP and from 24 vultures. DNA profiles of each B. anthracis strain were established using 31 VNTR markers (MLVA-31). Amplicons were used to generate Minimum Spanning Trees (MST) and unweighted pair group method with arithmetic mean (UPGMA) phylogenetic data using Bionumeric v6.6.5. The genotyping data was coupled with observed activity at carcass sites from camera images and video, preexisting isolates (n=107), as well as GPS movement maps for the vultures. Results: MST and UPGMA cluster analysis of MLVA-31 revealed 3 dominant clonal genotypes with a further 29 unique genotypes out of 169 isolates (i.e. 32 genotypes out of 169 isolates). With regards to the role of vultures, , a spore diluting role was observed from the data as at carcass sites that vultures fed on, low soil spore counts were observed. In contrast untouched carcass sites demonstrated higher soil spore contamination. For vultures B. anthracis was isolated from beaks, talons, feathers and cloacal swabs. Bacterial genotyping could link vultures to anthrax outbreaks. Beaks and feathers had clonal genotype suggesting that vultures fed during clonal outbreaks and the unique genotypes were from cloacal swabs (digested meals) representing diverse isolates from carcass sites not sampled during this study. MLVA has proven to be a useful tool to distinguish and determine genetic diversity of B. anthracis strains in KNP and vultures played the role of diminishing the pathogen load in the environment.