Metatranscriptomic Analysis of the Bacterial Symbiont Dactylopiibacterium carminicum from the Carmine Cochineal Dactylopius coccus (Hemiptera: Coccoidea: Dactylopiidae)

Abstract

The scale insect Dactylopius coccus produces high amounts of carminic acid, which has historically been used as a pigment by pre-Hispanic American cultures. Nowadays carmine is found in food, cosmetics, and textiles. Metagenomic approaches revealed that Dactylopius spp. cochineals contain two Wolbachia strains, a betaproteobacterium named Candidatus Dactylopiibacterium carminicum and Spiroplasma, in addition to different fungi. We describe here a transcriptomic analysis indicating that Dactylopiibacterium is metabolically active inside the insect host, and estimate that there are over twice as many Dactylopiibacterium cells in the hemolymph than in the gut, with even fewer in the ovary. Albeit scarce, the transcripts in the ovaries support the presence of Dactylopiibacterium in this tissue and a vertical mode of transmission. In the cochineal, Dactylopiibacterium may catabolize plant polysaccharides, and be active in carbon and nitrogen provisioning through its degradative activity and by fixing nitrogen. In most insects, nitrogen-fixing bacteria are found in the gut, but in this study they are shown to occur in the hemolymph, probably delivering essential amino acids and riboflavin to the host from nitrogen substrates derived from nitrogen fixation.

Keywords: Opuntia; betaproteobacteria; endosymbiont; nitrogen-fixation: gut microbiota; polysaccharide catabolism; scale insect.

Figure 1

Dorsal view of female second instar nymph of Dactylopius coccus with wax removed with 96% ethanol.

Figure 2

Number of Dactylopiibacterium expressed transcripts in different Dactylopius tissues. Reads were mapped to Dactylopiibacterium NFE1 reference genome.

Figure 3

Predicted metabolism and cellular features of Dactylopiibacterium in the insect host (Dactylopius). Red boxes represent expressed metabolism genes in the gut and hemolymph.

Figure 4

Flagellar and secretion systems expressed by Dactylopiibacterium in their insect host. (a) Heat map showing the average expression level of Dactylopiibacterium genes for flagellar production and motility in Dactylopius. (b) Flagellar diagram showing the expression (highlighted) of different flagellar components by Dactylopiibacterium in different insect tissues. (c) Heat map showing the average expression level of Dactylopiibacterium genes for secretion systems 1 and 2 in Dactylopius. (d) Bacterial secretion system diagram showing the expression (highlighted) of different Dactylopiibacterium genes for SS1 and SS2 in different insect tissues.

Figure 5

Expressed CAZymes and metabolic genes involved in pectin and rhamnogalacturonan metabolism of Dactylopiibacterium in the insect gut.

Figure 6

Illustrative proposed functional markers depicting Dactylopiibacterium differentially expression in different tissues: in blue, from the quantitative analysis; in red, from NOISeq; and in black, from both analyses.