Detection of pathogens in ticks collected from Tanzania using specific Polymerase Chain Reaction and Next-Generation Sequencing

Abstract

Background: Ticks are blood sucking arthropods that play a vital role in the transmission of a variety of pathogens to humans and animals. This tick also carries several other pathogens that cause human disease, including agents of Anaplasmosis, Babesiosis, Borreliosis, Rickettsia, Coxiella spp and others. Thus, tick borne diseases detection approaches using next-generation sequencing (NGS) of 16S rRNA gene amplicon enhance the efficiency of diagnosis and control strategies. In this study, detection of bacteria and protozoa pathogen was performed in ticks collected from wild animals from Serengeti National Park, Tanzania. Methods: Total 136 hard ticks were collected from wild animals (wildebeest, buffalo, zebra, and lion) in Tanzania in 2014-2016. Hard (Ixodidae) tick’s DNA was extracted from the ticks and pathogen-specific PCR was performed. In addition, microbiome study using NGS on of 16S rRNA gene amplification was performed. Result: In this pathogen specific analysis, 72 out of total 136 tick samples were positive for any potential pathogens and the detection rate 52.94%. The detection rate of pathogen in ticks from wildebeest, buffalo, zebra, lion were 64.7%, 60.6%, 54.24%, and 33.33%, respectively. The commonly detected potential pathogen was Coxiella spp. (38.24%), followed by Rickettsia spp. (13.24%), and Theileria spp. (0.74%). While Ana-plasma spp, Bartonella spp, and Borrelia spp were not detected in ticks. Microbiome study was performed on 16 tick samples. The number of bacterial species identified in ticks ranged from 70 to 122 among samples. The number of identified bacterial species and bacterial composition were not different between groups. The average relative abundances of Coxiella spp. in wildebeest, buffalo, and zebra were found as 0.01%, 24.33% and 26.93% respectively. The average relative abundances of Rickettsia spp. in wildebeest, buffalo, and zebra were found to be 0%, 0.28%, and 0.76%. Other potential pathogens were detected. All tick samples positive by NGS approach were found to be positive in pathogen-specific PCR approach in this study. Conclusion: Detection and analysis of ticks collected from wild animals demonstrated that Rickettsia and Coxiella pathogen detection rate were high in this study ,among the targeted pathogens. The microbiomes of bacterial composition varied between tick’s host animals, and the most occurrence microbiota from the members of Coxiellaceae, Francisellaceae, and Rickettsiaceae families (Phylum: Proteobacteria) were the most abundant. Therefore, Potential pathogens were detected in tick samples collected from wild animals in Tanzania using specific PCR and NGS approaches. In the future, NGS application for detection of pathogens could be considered since it is accurate and time saving. Key words: Tick, PCR, 16S rRNA, NGS, Microbiomes.