Microsporidian infection of mosquito larvae changes the host-associated microbiome towards the synthesis of antimicrobial factors

Abstract

Background: Microsporidians (Microsporidia) are a group of obligate intracellular parasites that commonly infect mosquitoes. Recently, it has been shown that infection by these parasites can alter the composition and functionality of the mosquito-associated microbiome. The host-associated microbiome of the mosquito can play a pivotal role in various physiological processes of this host, including its vector competence for pathogens. Thus, understanding how microsporidians shape the mosquito microbiome may be crucial for elucidating interactions between these parasites and their mosquito hosts, which are also vectors for other parasites and pathogens.

Methods: The effects of microsporidian infection on the microbiome structure and functionality of Culex pipiens and Culex torrentium larvae under semi-natural conditions were examined. The host-associated microbiome of Cx. pipiens (n = 498) and Cx. torrentium (n = 465) larvae, including that of the 97 infected individuals of these samples, was analysed using 16S DNA profiling and functional prediction analysis.

Results: Microbiome network analysis revealed that, in the microsporidian-positive larvae, host microbial communities consistently grouped within a common bacterial module that included Aerococcaceae, Lactobacillaceae, Microbacteriaceae, Myxococcaceae, and Polyangiaceae. Indicator species analysis revealed two strong positive correlations between microsporidian infection and the presence of Weissella cf. viridescens and Wolbachia pipientis. Functional predictions of microbiome content showed enrichment in biosynthetic pathways for ansamycin and vancomycin antibiotic groups in infected larvae. Furthermore, the MexJK-OprM multidrug-resistance module was exclusively present in the infected larvae, while carbapenem- and vancomycin-resistance modules were specific to the microsporidian-free larvae.

Conclusions: Our results demonstrate that microsporidian infection alters the microbial community composition in mosquito larvae. Moreover, they show that microsporidian infection can increase the antimicrobial capabilities of the host-associated microbiome. These results provide novel insights into host microbiome-parasite interactions and have potential implications for the vector competencies of mosquitoes.

Keywords: Weissella viridescens; Wolbachia pipientis; Antimicrobial factors; Biosynthetic pathways; Disease vectors; Host–microbe interactions; Microbiota; Microsporidia; Mosquitoes; Parasites.

Fig. 1

Boxplot showing the relative abundances of the phyla identified in the examined samples

Fig. 2

Hierarchical clustering heatmap showing the prevalence (%) of bacterial families in Culex pipiens and Culex torrentium samples positive (M+) or negative (M−) for microsporidians, and in water samples. Colour intensity corresponds to prevalence, with light blue indicating low prevalence and dark blue indicating high prevalence

Fig. 3

Principal coordinate (PC) analysis of Bray–Curtis dissimilarity based on microbial community composition. A Samples grouped by microbiota composition from early (light green) and late (dark green) larval stages of Culex pipiens, early (light blue) and late (dark blue) larval stages of Culex torrentium, and water (red). B Samples colour coded by ‘Microsporidium’ sp. PL03 infection status, where microsporidian-positive individuals (M+) are shown in green (Cx. pipiens), blue (Cx. torrentium), and red (water), while non-infected individuals (M−) are shown in grey

Fig. 4

A Co-occurrence networks of microbial communities in Culex pipiens and Culex torrentium, from ‘Microsporidium’ sp. PL03-positive individuals. Nodes represent bacterial families, with significant co-occurrence relationships (P < 0.05) are shown as edges. B Presence or absence of bacterial families is shown within the red module for both Cx. pipiens and Cx. torrentium. Colour intensity indicates prevalence, with dark blue denoting presence and light blue denoting absence of a given bacterial family. Taxa identified in both species are marked with a red frame. C Sankey diagram showing the systematics of the common taxa in the module containing'Microsporidium'sp. PL03

Fig. 5

Relationship between the number of'Microsporidium'sp. PL03 ribosomal DNA copies and A contribution of Weissella cf. viridescens and B Wolbachia between the Culex spp. Dashed red lines represent fitted regression models, with shaded areas indicating the 95% confidence intervals

Fig. 6

A Hierarchical clustering heatmap showing the prevalence (%) of the most prevalent functional metabolic profiles at level 3 of the microbiome in Culex pipiens and Culex torrentium infected by ‘Microsporidium’ sp. PL03 (M+) and non-infected ones (M−). Colour intensity reflects prevalence, with light blue indicating low prevalence and dark blue indicating high prevalence. Raincloud plots for the proportion of sequences assigned to the activity of the ansamycin (B) and vancomycin biosynthesis pathways (C) in Cx. pipiens and Cx. torrentium infected by ‘Microsporidium’ sp. PL03 (M+) and non-infected ones (M-)