PGRP-LD mediates A. stephensi vector competency by regulating homeostasis of microbiota-induced peritrophic matrix synthesis

Abstract

Peptidoglycan recognition proteins (PGRPs) and commensal microbes mediate pathogen infection outcomes in insect disease vectors. Although PGRP-LD is retained in multiple vectors, its role in host defense remains elusive. Here we report that Anopheles stephensi PGRP-LD protects the vector from malaria parasite infection by regulating gut homeostasis. Specifically, knock down of PGRP-LD (dsLD) increased susceptibility to Plasmodium berghei infection, decreased the abundance of gut microbiota and changed their spatial distribution. This outcome resulted from a change in the structural integrity of the peritrophic matrix (PM), which is a chitinous and proteinaceous barrier that lines the midgut lumen. Reduction of microbiota in dsLD mosquitoes due to the upregulation of immune effectors led to dysregulation of PM genes and PM fragmentation. Elimination of gut microbiota in antibiotic treated mosquitoes (Abx) led to PM loss and increased vectorial competence. Recolonization of Abx mosquitoes with indigenous Enterobacter sp. restored PM integrity and decreased mosquito vectorial capacity. Silencing PGRP-LD in mosquitoes without PM didn’t influence their vector competence. Our results indicate that PGPR-LD protects the gut microbiota by preventing hyper-immunity, which in turn promotes PM structurally integrity. The intact PM plays a key role in limiting P. berghei infection.

Glucose transporter GLUT1 influences Plasmodium berghei infection in Anopheles stephensi

“…We also noticed the signi cant induction of caudal, and reduction of pgrps, pgrp-la, pgrp-lc, pgrp-ld, and the antimicrobial peptides, defensin in dsAsteglut1-treated mosquitoes, suggesting that Asteglut1 might be involved in regulation of immune responses [25,40,56]. PGRP-LA is a receptor of the mosquito immune de ciency pathway (Imd) [41].…”

“…Five immune related genes were differentially regulated. Caudal, the negative regulator of Imd pathway was signi cantly upregulated (P = 0.0337) [40], while the peptidoglycan recognition proteins, pgrp-la, -lc,ld, and the antimicrobial peptides, defensin were signi cantly downregulated (P = 0.0111, P = 0.0378, P = 0.0022) [25,[40][41][42][43]. These results indicate that Asteglut1 might control parasite infection by regulating mosquito immune responses.…”

“…Four-day-old mosquitoes were injected with 69 nl dsRNA (4 μg/μl) using a nanoject II microinjector (Drummond, Philadelphia, USA). The dsRNA-treated mosquitoes were collected two days post-treatment and knockdown e ciency was veri ed by qPCR as previously described [25].…”

“…One µg of total RNA was used to synthesize cDNA using 5´ Allin-One MasterMix (AccuRT Genomic DNA Removal Kit; ABM, Shanghai, China). The qPCR was performed using a Roche LightCycler 96 Real Time PCR Detection System using SYBR Green qPCR Master Mix (Biomake, Shanghai, China) according to a previously described protocol [25]. The data were processed and analyzed using the Roche LightCycler 96 software.…”

Glucose-mediated expansion of a gut commensal bacterium promotesPlasmodiuminfection through alkalizing mosquito midgut

“…Notably, glucose and trehalose supplementation significantly repressed the activity of Imd pathway ( Figure 4A and Figure S3A). The expression level of caudal, the negative regulator of IMD pathway was significantly up-regulated, while peptidoglycan recognition proteins, pgrp-lc, antimicrobial peptides, defensin, and cecropin were significantly down-regulated both prior to and post infectious blood meal (Cirimotich et al, 2010;Clayton et al, 2013;Clayton et al, 2014;Song et al, 2018)( Figure 4A). It is highly possible that increased susceptibility to P. berghei infection results from the down-regulation of Imd pathway.…”