Natural variation in the contribution of microbial density to inducible immune dynamics

Abstract

Immune responses evolve to balance the benefits of microbial killing against the costs of autoimmunity and energetic resource use. Models that explore the evolution of optimal immune responses generally include a term for constitutive immunity, or the level of immunological investment prior to microbial exposure, and for inducible immunity, or investment in immune function after microbial challenge. However, studies rarely consider the functional form of inducible immune responses with respect to microbial density, despite the theoretical dependence of immune system evolution on microbe‐ versus immune‐mediated damage to the host. In this study, we analyse antimicrobial peptide (AMP) gene expression from seven wild‐caught flour beetle populations (Tribolium spp.) during acute infection with the virulent bacteria Bacillus thuringiensis (Bt) and Photorhabdus luminescens (P.lum) to demonstrate that inducible immune responses mediated by the humoral IMD pathway exhibit natural variation in both microbe density‐dependent and independent temporal dynamics. Beetle populations that exhibited greater AMP expression sensitivity to Bt density were also more likely to die from infection, while populations that exhibited higher microbe density‐independent AMP expression were more likely to survive P. luminescens infection. Reduction in pathway signalling efficiency through RNAi‐mediated knockdown of the imd gene reduced the magnitude of both microbe‐independent and dependent responses and reduced host resistance to Bt growth, but had no net effect on host survival. This study provides a framework for understanding natural variation in the flexibility of investment in inducible immune responses and should inform theory on the contribution of nonequilibrium host‐microbe dynamics to immune system evolution.

Decomposing virulence to understand bacterial clearance in persistent infections

“…It has been suggested that the SPBL will remain at around the bacterial load at which the infection was controlled [19, 21]. Given that insects can show dose dependent inducible immune activation [34], and given that the antimicrobial peptide Drosocin has been shown to control E. cloacae infections and that a combination of Drosocin, Attacins and Diptericins control Pr. burhodogranariea infections [47], one could hypothesise that these AMPs are to some degree involved.…”

“…The initial exposure dose will also determine the outcome of infection, partly because microbe density at the beginning of an infection can determine the strength of the immune response [34]. Not only is dose-dependent survival frequently reported in response to bacterial infections [19, 26, 35], but bacterial load later in the infection has been demonstrated to correlate with the initial injection dose [19, 21].…”

Larvae and adults exhibit contrasting patterns of immune gene expression and infection resistance in wild flour beetle populations

“…The initial collection of wild beetle populations and analysis of variation in immune gene expression among larvae from these populations has been described in detail in [15]. The current study compares these larval data (available from Data Dryad [16]) to data from adults processed at the same time (data available i).…”

“…All injected beetles were sacrificed at 8 hours post injection by flash freezing, and stored at -80°C. We extracted RNA, created cDNA, and quantified gene expression via RT-qPCR as previously described [15] using 96 degenerate primers capable of amplifying transcripts from both flour beetle species with equal 97 efficiency for four immune genes: attacin-1, defensin-1, pgrp-sc2, and ddc. In addition, we 98 quantified the expression of host reference gene rps18 and a fragment of the Bt 16s gene that has 99 been previously validated as a proxy for bacterial CFU [17].…”

“…To address this question, we turned to wild and lab-derived populations of flour beetles ( Tribolium castaneum and T. confusum ). Previous research in lab-reared T. castaneum has suggested that the regulation of immune [13] and cuticular [14] gene expression differs among life stages in uninfected individuals, while, in larvae specifically, different populations of both species exhibit natural variation in immune gene expression and resistance to bacterial infection [15]. To test whether immune gene expression and infection resistance in these populations differs among life stages, we injected larvae and adults from seven populations (four T. castaneum , three T. confusum ) with one of four increasing doses of the entomopathogenic bacterium Bacillus thuringiensis (Bt) or sterile media as a wounding control.…”