Exploring bacterial communities and genetic diversity haboured by Amblyomma hebraeum and Rhipicephalus evertsi evertsi ticks from livestock in North West Province, South Africa

Abstract

Ticks are important vectors of pathogens that impact both livestock and human health. Among the significant tick species in South Africa, transmitting economically important diseases to livestock are Amblyomma hebraeum and Rhipicephalus evertsi evertsi. Amblyomma hebraeum is a vector for Ehrlichia ruminantium, causing heartwater in cattle, sheep, and goats. It also transmits Rickettsia africae, responsible for tick-bite fever in human. R. e evertsi transmits Coxiella spp. in sheep, and Anaplasma marginale, causing anaplasmosis in cattle. This study investigated the bacterial diversity and veterinary important pathogens harboured by A. hebraeum and R. e. evertsi ticks in sheep and goats from the North West Province of South Africa. Ixodid ticks were collected from different farms in Mahikeng and Matlwang village and were identified using morphological and molecular techniques. Molecular identification targeting the 16S rRNA and 18S rRNA genes sequences was performed on the A. hebraeumand R. e. evertsi ticks. Bacterial communities associated with these tick species were investigated using next-generation sequencing (NGS) technques performed using highthroughput sequencing of the V3-V4 hypervariable regions of the 16S rRNA gene on the Illumina MiSeq platform. Key differences in bacterial communities among these tick species were further exploited. The ticks collected from Mahikeng were morphologically identified as A. hebraeum. They were characterized by their legs, which had distinctive red and white rings, and a notably ornate scutum with yellow, green, brown, and black markings. Meanwhile, ticks collected frm Matlwang were identified as R. e. evertsi, characterized by their dark brown bodies, reddish-orange lateral sides and reddish-orange legs. Amplicon gene sequencing revealed significant sequence similarity, with 18S rRNA gene sequences exhibiting 99.36-99.87% identity to known GenBank sequences of the investigated tick species. Phylogenetic analysis based on 16S and 18S rRNA gene sequences demonstrated strong monophyletic clustering, clearly distinguishing between the two tick genera. Next-generation sequencing showed that both tick species were primarily associated with Proteobacteria, with A. hebraeumfrom sheep showing a high abundance of 82.8%, indicating significant reliance on this phylum for survival. Amblyomma hebraeum was mainly composed of Rickettsia spp., which comprised 61.42% of its bacterial composition. Meanwhile, R. e. evertsi exhibited greater bacterial diversity, including various taxa such as Acinetobacter and Pseudomonas species, particularly from sheep, hosting 8,425 unique taxa (27.8%) compared to A. hebraeum's 4,402 (9%). Both species shared 1,534 bacterial taxa, representing 63.1% of their core microbiome, pointing to conserved microbial functions. Their predicted functional pathways related to energy metabolism, carbon, and nitrogen cycling highlight their ecological adaptability. The study's findings demonstrate that host species significantly influence the tick microbiome, with ticks from sheep exhibiting more stable and less variable bacterial communities compared to those from goats. The R. e. evertsi ticks demonstrated greater bacterial diversity, including more Actinobacteria in goat-derived ticks. The molecular identification of ticks provided a genetic library of local or regional tick fauna, especially in underrepresented areas of the local farming system, where tick diversity is high yet under-documented. Data generated in this study will contribute to guiding the development of tick surveillance systems, risk mapping and disease control efforts in South African livestock production systems.