Functional genomics of a Spiroplasma associated with the carmine cochineals Dactylopius coccus and Dactylopius opuntiae

Abstract

Background Spiroplasma is a widely distributed endosymbiont of insects, arthropods, and plants. In insects, Spiroplasma colonizes the gut, hemolymph, and reproductive organs of the host. Previous metagenomic surveys of the domesticated carmine cochineal Dactylopius coccus and the wild cochineal D. opuntiae reported sequences of Spiroplasma associated with these insects. However, there is no analysis of the genomic capabilities and the interaction of this Spiroplasma with Dactylopius. Results Here we present three Spiroplasma genomes independently recovered from metagenomes of adult males and females of D. coccus, from two different populations, as well as from adult females of D. opuntiae. Single-copy gene analysis showed that these genomes were > 92% complete. Phylogenomic analyses classified these genomes as new members of Spiroplasma ixodetis. Comparative genome analysis indicated that they exhibit fewer genes involved in amino acid and carbon catabolism compared to other spiroplasmas. Moreover, virulence factor-encoding genes (i.e., glpO, spaid and rip2) were found incomplete in these S. ixodetis genomes. We also detected an enrichment of genes encoding the type IV secretion system (T4SS) in S. ixodetis genomes of Dactylopius. A metratranscriptomic analysis of D. coccus showed that some of these T4SS genes (i.e., traG, virB4 and virD4) in addition to the superoxide dismutase sodA of S. ixodetis were overexpressed in the ovaries. Conclusion The symbiont S. ixodetis is a new member of the bacterial community of D. coccus and D. opuntiae. The recovery of incomplete virulence factor-encoding genes in S. ixodetis of Dactylopius suggests that this bacterium is a non-pathogenic symbiont. A high number of genes encoding the T4SS, in the S. ixodetis genomes and the overexpression of these genes in the ovary and hemolymph of the host suggest that S. ixodetis use the T4SS to interact with the Dactylopius cells. Moreover, the transcriptional differences of S. ixodetis among the gut, hemolymph and ovary tissues of D. coccus indicate that this bacterium can respond and adapt to the different conditions (e.g., oxidative stress) present within the host. All this evidence proposes that there is a strong interaction and molecular signaling in the symbiosis between S. ixodetis and the carmine cochineal Dactylopius.

Highly transmissible cytoplasmic incompatibility by the extracellular insect symbiont Spiroplasma

“…The s Dis symbiont encodes complete ATP synthase and glycolysis pathways, as well as transport components for glucose, fructose, GlcNAc, glycerol, and possibly mannose ( Figure 5 ). Amino acid and lipid biosynthesis pathways are incomplete or absent, as reported previously for hemolymph-dwelling Spiroplasma species, including those in the Ixodetis clade ( Martin et al., 2020 ; Vera-Ponce León et al., 2021 ; Yeoman et al., 2019 ). Incomplete pathway components were identified for lysine ( dapE ), valine, leucine, and isoleucine ( ilvA ), and arginine ( OTC ) synthesis.…”

Male-killing mechanisms vary between Spiroplasma species

“…The present study confirmed that H. magnanima was infected with the MK S. ixodetis strain s Hm that harbored two distinct 16S rRNA gene sequences in its genome ( Figure 2 ). Moreover, Spiroplasma strains often encode multiple ribosomal RNA gene sets in their genome ( Chang et al, 2014 ; Tsai et al, 2018 ; Vera-Ponce León et al, 2021 ).…”

“… 2 Based on Martin et al (2020) . 3 Based on Vera-Ponce León et al (2021) . 4 Based on Pollmann et al (2022) .…”

“…Protein homology between different Spiroplasma strains was analyzed using S. apis B31 (CP006682.1), S. citri strain BLH-MB (CP047437.1–CP047446.1), S. syrphidicola strain EA-1 (NC_021284.1), D. melanogaster endosymbiont S. poulsonii MSRO (CM020866.1–CM020867.1) ( s Mel, MK strain, Masson et al, 2018 ), Danaus chrysippus (Nymphalidae) endosymbiont S. ixodetis (NZ_CADDIL010000001.1–NZ_CADDIL010000012.1) ( s Da, MK strain, Martin et al, 2020 ), Lariophagus distinguendus (Pteromalidae) endosymbiont S. ixodetis (NZ_JALMUW010000001.1–NZ_JALMUW010000198.1) [ s Dis, cytoplasmic incompatibility (CI) strain, Pollmann et al, 2022 ], and Dactylopius coccus (Dactylopiidae) endosymbiont S. ixodetis (JACSER010000001.1–JACSER010000358.1) ( s Coc, non-MK strain, Vera-Ponce León et al, 2021 ) with OrthoVenn2. 2 Homology of s Hm genes and proteins with spaid from strain s Mel ( Harumoto and Lemaitre, 2018 ), ankyrin genes from S. ixodetis ( Yeoman et al, 2019 ; Martin et al, 2020 ; Vera-Ponce León et al, 2021 ), and the Wolbachia MK candidate factor responsible for WO-mediated killing (Wmk, presumed helix-turn-helix transcriptional regulator, Perlmutter et al, 2019 ; Arai et al, 2022b ) was evaluated using both BLASTn and BLASTp. Moreover, to verify whether MK microbes of H. magnanima carried conserved genes, the genes on the MK-associated prophage region WO w Hm-t76 of MK Wolbachia w Hm-t ( Arai et al, 2022b ) and those of the Partiti-like virus OGVs ( Fujita et al, 2021 ) were compared to the s Hm genes using both BLASTn and BLASTp.…”

“…1 Phage WO infections were annotated using PHASTER (Arndt et al, 2016). Protein homology between different Spiroplasma strains was analyzed using S. apis B31 (CP006682.1), S. citri strain BLH-MB (CP047437.1-CP047446.1), S. syrphidicola strain EA-1 (NC_021284.1), D. melanogaster endosymbiont S. poulsonii MSRO (CM020866.1-CM020867.1) (sMel, MK strain, Masson et al, 2018) (Harumoto and Lemaitre, 2018), ankyrin genes from S. ixodetis (Yeoman et al, 2019;Martin et al, 2020;Vera-Ponce León et al, 2021), and the Wolbachia MK candidate factor responsible for WO-mediated killing (Wmk, presumed helixturn-helix transcriptional regulator, Perlmutter et al, 2019;Arai et al, 2022b) was evaluated using both BLASTn and BLASTp. Moreover, to verify whether MK microbes of H. magnanima carried conserved genes, the genes on the MK-associated prophage region WOwHm-t76 of MK Wolbachia wHm-t (Arai et al, 2022b) and those of the Partiti-like virus OGVs (Fujita et al, 2021) were compared to the sHm genes using both BLASTn and BLASTp.…”