Decoupling Abundance from Centrality: Sexual Dimorphism and Geographical Structuring in the Hyalomma lusitanicum Microbiome Network

Background: The tick microbiome is shaped by environmental filters and host biological traits. Microbial co-occurrence networks offer a powerful framework to map inter-microbial interactions, node topological roles, and community stability beyond standard diversity metrics; however, how tick sex modulates these structural networks remains poorly understood. Methods: We sequenced the V3–V4 variable region of the 16S rRNA gene from 86 adult Hyalomma lusitanicum ticks (37 males and 49 females) collected across six ecogeographical localities in Andalusia during 2023. A refined dataset of 5570 high-confidence Amplicon Sequence Variants (ASVs) was analyzed. Alpha and beta diversity patterns were evaluated using PERMANOVA models, and sex-stratified co-occurrence networks were constructed using Spearman’s rank correlations and the Zi-Pi topological framework to assess network complexity and keystone roles. Results: The primary obligate endosymbiont Francisella dominated the bacteriome (60–90% relative abundance), followed by Candidatus Midichloria. PERMANOVA models confirmed that geographical locality (R2 = 0.111, p = 0.003) and host sex (R2 = 0.095, p = 0.001) exerted strong, independent influences on microbial composition. Female ticks harbored a highly streamlined bacterial profile. Conversely, males exhibited significantly higher alpha diversity (p < 0.001), reflecting host-sex physiological and behavioral differences that create a broader ecological niche allowing diverse environmental and transient taxa (Acinetobacter, Sphingomonas, Stenotrophomonas, Brevundimonas) to colonize. Network analysis revealed a striking sexual dimorphism: female networks were highly centralized and anchored by a single global hub (Roseomonas), whereas male networks lacked global hubs and displayed higher topological complexity, achieving stability through a decentralized architecture sustained by 68 inter-module connectors (compared to 24 in females). Despite these macro-topological differences, a core set of seven connector ASVs was conserved across both sexes. Conclusions: These findings demonstrate that host sex shapes the ecological niche of the tick microbiome and drives distinct network topological strategies to maintain community stability, providing new insights into the structural ecology of tick–microbe interactions.