Negative immune regulation contributes to disease tolerance in Drosophila

Abstract

Disease tolerance is an infection phenotype where hosts show relatively high health despite harbouring elevated pathogen loads. Compared to the mechanisms of immune clearance our knowledge of the mechanisms underlying increased tolerance remains incomplete. Variation in the ability to reduce immunopathology may explain why some hosts can tolerate higher pathogen burdens with reduced pathology. Negative immune regulation would therefore appear to be a clear candidate for a mechanism underlying disease tolerance but this has not been tested directly for bacterial infections. Here, we examined how the negative regulation of the immune deficiency (IMD) pathway affects disease tolerance in Drosophila melanogaster when infected with the gram-negative bacterial pathogen Pseudomonas entomophila. We find that UASRNAi-mediated reduced expression of the negative regulators of IMD (pirk & caudal) severely reduced the ability to tolerate infection in both males and females across a wide range of infectious doses. While flies unable to regulate the IMD response exhibited higher expression of antimicrobial peptides and lower bacterial loads as expected, this was not accompanied by a proportional reduction in mortality. Instead, tolerance (measured as fly survival relative to its microbe load) was drastically reduced, likely due to the combination of increased immunopathology and cytotoxicity of elevated AMP expression. Our results therefore highlight that in addition to regulating an efficient pathogen clearance response, negative regulators of IMD also contribute to disease tolerance.

Ageing leads to nonspecific antimicrobial peptide responses in Drosophila melanogaster

“…Finally, we also found ageing-associated downregulation of the major negative regulators of Imd-signalling such as Caudal & Pirk in older flies (Fig. 5B; Table S12), which has been previously linked to the production of toxic levels of AMP production, causing reduced lifespan, locomotor defects and extensive neurodegeneration (Kounatidis et al, 2017; Prakash et al, 2021). Based on these results, we speculate that the observed expansion of AMP repertoire with age is therefore most likely to represent suboptimal body condition, characterized by poorly regulated immune system and increased physiological costs.…”

“…While pirk interferes with the interaction of PGRP-LC and -LE with the molecule Imd to limit the activation of the Imd pathway, caudal downregulates the expression of AMPs (Lee and Ferrandon, 2011). To examine whether non-specific expansion of AMP repertoire was associated with the lower expression of these negative regulators, we estimated their relative expression level in both young and old iso- w 1118 individuals infected with P. rettgeri at 24 hours post-infection, by using qPCR (as outlined in Prakash et al, 2021) (n= Total 15-21 flies in a group of 3 homogenized in Trizol reagent/ Infection treatment/ age-group and sex-combination).…”

“…To test whether ageing reduces host’s ability to supress the bacterial growth, we quantified bacterial load in each of the fly lines at 24-hours (or 20-hours) after Providencia rettgeri (or Pseudomonas entomophila ) infection across different age groups (3- vs 25-day-old) and sexes (as described in Prakash et al, 2021). First, we thoroughly washed individual flies (pooled in a group of 6 in 1.5ml centrifuge tube) with 70% ethanol for 30-seconds to remove the surface bacteria, followed by rinsing them twice with sterile distilled water.…”

“…The expression of major Imd-pathway regulators was quantified by qRT-PCR for both young and older individuals, as described previously in Prakash et al (2021). We randomly selected a subset of young and old iso- w 1118 flies infected with 0.1 OD P. rettgeri for RNA extraction at 24h post-infection.…”

The restriction factor pastrel is associated with host vigor, viral titer, and variation in disease tolerance during Drosophila C Virus infection

“…Pro-inflammatory cytokines tend to be associated with decreased tolerance to infection -for example, a tolerant house finch population (Haemorhous mexicanus) infected with a bacterial pathogen, Mycoplasma gallisepticum, exhibited lower cytokine expression compared with a less tolerant population [35]; mice receiving the anti-inflammatory drug Ibuprofen showed improved increased tolerance during Mycobacterium tuberculosis infection [39]; and lower levels of circulating pro-inflammatory cytokines are associated with tolerance of malaria after reexposure to the parasite [40]. Negative regulation of immune responses that minimize inflammation would therefore appear to be prime candidates for mechanisms that promote disease tolerance [34,[40][41][42]. This is supported by previous work showing that the epigenetic modifier, G9a, which regulates JAK-STAT signalling to prevent hyperactivation of the immune response, increases tolerance to RNA virus infection by limiting immunopathology [43,44].…”