Plasticity of Expression of Immunoregulatory Genes in Two Allopatric Populations of Drosophila ananassae: Novel Insight into Molecular Process Under Selection.

Evolutionary responses to persistent pathogen exposure can involve changes in immune regulation that vary among populations, sexes, and immune-related genes. We investigated the transcriptional response to long-term bacterial selection in two allopatric populations of Drosophila ananassae from the Indian subcontinent. Flies were subjected to continuous selection with Pseudomonas aeruginosa for 55 generations, while a separate relaxed-selection regime was initiated after generation 45 and maintained for a further 10 generations in the absence of bacterial challenge. The expression of nine immune-regulatory genes involved in melanization, immune signalling, and antimicrobial defence was quantified using RT-qPCR across generations, populations, and sexes. Continuous bacterial selection resulted in progressive changes in the expression of immune-related genes, although the magnitude and temporal pattern differed among genes, populations, and sexes. Different genes exhibited population specific trends in their changing expression level. Females generally showed greater transcriptional responses than males for several genes. Following removal of bacterial selection, expression of most genes declined toward control levels, although residual elevation persisted for several genes after ten generations of relaxed selection and the extent of reversion differed among genes and populations. These findings demonstrate that prolonged bacterial selection is associated with coordinated but gene-specific changes in immune gene expression in D. ananassae, together with population- and sex-dependent variation in transcriptional responses. The persistence and subsequent reversion of expression after removal of selection highlight the dynamic nature of immune regulation, while the underlying genetic, physiological, and epigenetic mechanisms remain to be determined. This study provides a molecular framework for investigating the evolution and regulation of immune responses under prolonged pathogen exposure and subsequent relaxation of selection.