Virus Prospecting in Crickets-Discovery and Strain Divergence of a Novel Iflavirus in Wild and Cultivated Acheta domesticus

Abstract

Orthopteran insects have high reproductive rates leading to boom-bust population dynamics with high local densities that are ideal for short, episodic disease epidemics. Viruses are particularly well suited for such host population dynamics, due to their supreme ability to adapt to changing transmission criteria. However, very little is known about the viruses of Orthopteran insects. Since Orthopterans are increasingly reared commercially, for animal feed and human consumption, there is a risk that viruses naturally associated with these insects can adapt to commercial rearing conditions, and cause disease. We therefore explored the virome of the house cricket Acheta domesticus, which is both part of the natural Swedish landscape and reared commercially for the pet feed market. Only 1% of the faecal RNA and DNA from wild-caught A. domesticus consisted of viruses. These included both known and novel viruses associated with crickets/insects, their bacterial-fungal microbiome, or their plant food. Relatively abundant among these viral Operational Taxonomic Units (OTUs) was a novel Iflavirus, tentatively named Acheta domesticus Iflavirus (AdIV). Quantitative analyses showed that AdIV was also abundant in frass and insect samples from commercially reared crickets. Interestingly, the wild and commercial AdIV strains had short, extremely divergent variation hotspots throughout the genome, which may indicate specific adaptation to their hosts' distinct rearing environments.

Keywords: Acheta domesticus; Acheta domesticus Iflavirus; cricket; cricket rearing; metagenome; strain evolution; virome.

Figure 1

Taxonomic composition of Acheta domesticus frass. (A) Full composition of the wild Acheta domesticus frass sample at the level of Domain (Archaea, Bacteria, Eukarya, and Viruses) as determined by RNA sequencing. (B) Composition of the 1% virus subfraction, separated by primary host: bacteria (yellow), fungi (grey), plants (green), insects (orange), invertebrates (red), and miscellaneous viruses (blue) whose host status is unclear. (C) Cladogram of the viral reads obtained through RNA sequencing, with the primary hosts of the virus identified by coloured dots, as for Figure 1B. The composite circles represent the host distribution at higher virus taxonomic categories. The numbers indicate the number of unique reads assigned to the corresponding taxon.

Figure 2

Genome organization and phylogenetic placement of Acheta domesticus Iflavirus. (A) Genome organization of Acheta domesticus Iflavirus (AdIV), showing the location and size of the structural (VP2, VP4, VP3, VP1; dark grey) and non-structural (L protein, helicase, 3C-protease, and RNA dependent RNA polymerase; light grey) genes, separated by the location of putative 3C-protease cleavage sites. The chart below the genome map summarizes the nature of the nucleotide differences between the two strains in a moving 90-nucleotide window for: the proportion of all changes that are transitions (Ts/(Ts+Tv): grey); the overall rate of synonymous and non-synonymous change, (d(N+S): blue), and the rate of non-synonymous change, (d(N): red). The blue arrows identify the location and size of the AdIV diagnostic RT-qPCR assay. The red arrows identify the location and size of the high variability hotspots. (B) Location of the wild (AdIV-w) and commercial (AdIV-c) strains of AdIV (red branch) in the Iflavirus phylogenetic tree, based on analysis of the full polyprotein amino acid sequences of a representative selection of Iflaviruses (Table S2). The insect icons indicate the Families of their primary hosts. The tree is draw to scale, measured in amino acid substitutions per site. Also included is a high-resolution phylogeny of the six positive samples (Table 1), based on 684 nucleotides in the helicase region. Groups of taxa that cluster significantly are identified by the white, grey, and black (red) circles on the respective branching nodes.