Correlation-based analysis of gene expression in Cardinium , Wolbachia , and Tyrophagus putrescentiae reveals distinct host-symbiont interactions and parallel evolutionary expansion of infection-associated genes

Cardinium and Wolbachia are phylogenetically distinct intracellular symbionts that have convergently evolved mechanisms to infect a wide range of arthropod and nematode hosts. Here, we investigated the interaction dynamics of two basal strains, Cardinium (cTPut) and Wolbachia (wTPut), in the stored-product mite Tyrophagus putrescentiae (Acari: Astigmata). The KEGG-annotated gene expression profiles of mite samples were analyzed in single infections ( Cardinium or Wolbachia ) and mixed infections across three time points (2, 4, and 6 months). Temporal effects on gene expression were marginal; therefore, correlation-based network analyses were used to characterize host-symbiont interactions. The symbionts exhibited distinct host-targeting strategies: Wolbachia displayed a “broad” interaction pattern characterized by numerous weak correlations with host KEGG genes, whereas Cardinium showed a “focused” strategy involving fewer but stronger correlations. These patterns were consistent across global KEGG data sets, immune and regulatory pathways (including AMPK, autophagy, endocytosis, Hippo, lysosome, MAPK, mTOR, and Notch), receptor subsets, and metabolic pathways. Consistent with these interaction patterns, orthologous analyses across Cardinium and selected Wolbachia genomes revealed that infection-associated genes are not broadly conserved but are enriched in lineage-specific, duplicated gene families, particularly in T. putrescentiae -associated symbionts. These findings indicate that infection-related traits evolve primarily through lineage-specific gene family expansion rather than conservation and suggest that basal Cardinium and Wolbachia employ fundamentally distinct host interaction strategies. IMPORTANCE The study provides novel insight into host-symbiont interactions: Cardinium , Wolbachia infecting mite Tyrophagus putrescentiae , an important producer of allergens (i) Cardinium gene expression responds to the presence of Wolbachia , whereas Wolbachia gene expression shows no reciprocal response; (ii) the two symbionts employ contrasting host interaction strategies with the host, with Cardinium exhibiting fewer but stronger associations and Wolbachia showing widespread but weaker correlations; (iii) distinct host pathways are differentially associated with each symbiont; and (iv) we identified 40 of the most correlated bacterial genes to the host that include candidates of viral origin, suggesting a potential role of horizontally acquired elements in the host. Furthermore, evolutionary analyses demonstrate that infection-associated genes are primarily lineage expanded rather than broadly conserved across symbiont genomes. Given the allergenic relevance of T. putrescentiae , these findings suggest that symbiont-specific interaction strategies can indirectly influence host physiology and allergen production.