Microbial networks and pathogen detection: Insights from ticks and patients with acute febrile illness

Ticks are vectors of diverse pathogens, with transmission risk elevated in regions of close human-livestock-wildlife contact. In rural South Africa, non-malarial acute febrile illness (AFI) is common, yet the microbial diversity of ticks and human patients remains underexplored. We applied microbiome sequencing (16S rDNA for bacteria; 18S rDNA for eukaryotes) to four sample groups from Bushbuckridge, Mpumalanga, South Africa: Rhipicephalus sanguineus s.l. (n_16S = 175, n_18S = 30), Amblyomma hebraeum (n_16S = 74, n_18S = 21), human AFI patients (n_16S = 168, n_18S = 14), and apparently healthy humans (n_16S = 67, n_18S = 3). Bacterial alpha diversity did not differ significantly between groups (Shannon p = 0.81, Inverse-Simpson p = 0.36), whereas eukaryotic diversity was lower in human patients (Shannon p = 0.00026, Inverse-Simpson p = 0.00415). Beta diversity exhibited clear differences between groups, with ticks showing greater variability (bacteria p = 0.001; eukaryotes p = 0.001). Co-occurrence networks revealed structured clusters reflecting organised community interactions. Pathogen DNA detected included Plasmodium falciparum in an AFI patient, and sequences similar to several spotted fever group Rickettsia species, including the first report of Rickettsia barbariae in ticks from South Africa. This result requires confirmation by multi-gene sequence analysis. Novel Rickettsiales with uncertain pathogenicity were detected across sample groups. Opportunistic pathogens were detected including Acanthamoeba, Aspergillus and Malassezia. These findings provide baseline evidence on microbial diversity in a rural community in South Africa, underscoring the need for integrated surveillance to address AFI burden within a One Health framework.