The dynamics of the microbiome in Ixodidae are shaped by tick ontogeny and pathogens in Sarawak, Malaysian Borneo

Tick -borne diseases have recently been considered a potential emerging public health threat in Malaysia; however, fundamental studies into tick -borne pathogens and microbiome appear limited. In this study, six tick species (Ixodes granulatus, Haemaphysa-lis hystricis, Haemaphysalis shimoga, Dermacentor compactus, Dermacentor steini and Dermacentor atrosignatus) collected from two primary forests and an oil palm plantation in Sarawak, Malaysian Borneo, were used for microbiome analysis targeting bacterial 16S rDNA using next-generation sequencing (NGS). In addition, bacterial species were further characterized in con-ventional PCRs to identify potential pathogens. Sequences generated from NGS were first filtered with the Decontam package in R before subsequent microbial diversity analyses. Alpha and beta analyses revealed that the genus Dermacentor had the highest microbial diversity, and H. shimoga significantly differed in microbial composition from other tick species. Alpha and beta diversities were also significantly different between developmental stages of H. shimoga. Furthermore, we observed that some bacterial groups were significantly more abundant in certain tick species and developmental stages of H. shimoga. We tested the relative abundances using pairwise linear discriminant analysis effect size (LEfSe), which also revealed significant microbial composition differences between Borrelia-positive and Borrelia-negative I. granulatus ticks. Finally, pathogenic and potentially pathogenic bacteria circulating in different tick species, such as Rickettsia heilongjiangensis, Ehrlichia sp., Anaplasma sp. and Bartonella spp. were characterized by PCR and sequencing. Moreover, Coxiella and Francisella- like potential symbionts were identified from H. shimoga and D. steini, respectively. More studies are required to unravel the factors associated with the variations observed in this study.