The genome of Gryllus bimaculatus nudivirus indicates an ancient diversification of baculovirus-related nonoccluded nudiviruses of insects

Abstract

The Gryllus bimaculatus nudivirus (GbNV) infects nymphs and adults of the cricket Gryllus bimaculatus (Orthoptera: Gryllidae). GbNV and other nudiviruses such as Heliothis zea nudivirus 1 (HzNV-1) and Oryctes rhinoceros nudivirus (OrNV) were previously called "nonoccluded baculoviruses" as they share some similar structural, genomic, and replication aspects with members of the family Baculoviridae. Their relationships to each other and to baculoviruses are elucidated by the sequence of the complete genome of GbNV, which is 96,944 bp, has an AT content of 72%, and potentially contains 98 predicted protein-coding open reading frames (ORFs). Forty-one ORFs of GbNV share sequence similarities with ORFs found in OrNV, HzNV-1, baculoviruses, and bacteria. Most notably, 15 GbNV ORFs are homologous to the baculovirus core genes, which are associated with transcription (lef-8, lef-9, lef-4, vlf-1, and lef-5), replication (dnapol), structural proteins (p74, pif-1, pif-2, pif-3, vp91, and odv-e56), and proteins of unknown function (38K, ac81, and 19kda). Homologues to these baculovirus core genes have been predicted in HzNV-1 as well. Six GbNV ORFs are homologous to nonconserved baculovirus genes dnaligase, helicase 2, rr1, rr2, iap-3, and desmoplakin. However, the remaining 57 ORFs revealed no homology or poor similarities to the current gene databases. No homologous repeat (hr) sequences but fourteen short direct repeat (dr) regions were detected in the GbNV genome. Gene content and sequence similarity suggest that the nudiviruses GbNV, HzNV-1, and OrNV form a monophyletic group of nonoccluded double-stranded DNA viruses, which separated from the baculovirus lineage before this radiated into dipteran-, hymenopteran-, and lepidopteran-specific clades of occluded nucleopolyhedroviruses and granuloviruses. The accumulated information on the GbNV genome suggests that nudiviruses form a highly diverse and phylogenetically ancient sister group of the baculoviruses, which have evolved in a variety of highly divergent host orders.

FIG. 1.

Restriction endonuclease profiles of GbNV DNA for the restriction endonucleases as shown above each lane. Molecular sizes are shown on the left.

FIG. 2.

Linear map of the GbNV genome. ORFs and their transcriptional directions are indicated as arrows.

FIG. 3.

Alignments of the putative LEF-5 zinc ribbon domains from AcMNPV, HzNV-1, OrNV, and GbNV. Identical and biochemically similar amino acid residues are shaded in black and gray, respectively. Potential Zn-coordinating amino acids are marked with asterisks.

FIG. 4.

Unrooted ME phylogenetic tree based on 2,518 sites of the DNA polymerases of 20 viruses from various families. Distances were calculated using Poisson correction. The homogeneous substitution pattern among lineages with the gamma-distributed rate among sites (Gamma parameter 2.02) is employed for reconstruction of the tree. Gaps and missing data are included for calculation of informative sites using pairwise deletion. The robustness of the tree was tested using bootstrap analyses (1,000 replicates), and the percent values (ME, NJ, MP) are given next to the nodes. The names of the selected virus families are indicated on the tree. The GbNV, OrNV, and HzNV-1 clade is indicated on a gray background. The GenBank accession numbers of the viral DNA polymerase (from top to bottom) are YP_142676, NP_048532, NP_077578, YP_293784, NP_045328, NP_050219, AAC54632, NP_149500, NP_078724, NP_478036, NP_042783, NP_064832, NP_063712, ABF93350, ABO45345, NP_690550, NP_203396, YP_025217, NP_054095, and NP_148895. The scale bar represents a distance of 50%.

FIG. 5.

(A) Midpoint-rooted NJ phylogenetic tree based on the concatenated amino acid sequences (1,805 sites) of the lef-4, lef-5, dnapol, pif-2, and ac81 genes from GbNV, OrNV, HzNV-1, and four selected baculoviruses. Distances were calculated using Poisson correction. Homogeneous substitution patterns among lineages with uniform rates among sites are employed for reconstruction of the tree. Gaps and missing data are excluded from analyses. The robustness of the tree was tested using bootstrap analyses (1,000 replicates), and the percent values (NJ) are given next to the nodes. ME and MP analyses revealed similar topologies for the tree. The group of GbNV, OrNV, and HzNV-1 is indicated on a gray background. (B) Evolutionary relationships of the orders of insects that are infected by the baculoviruses and nudiviruses shown in panel A (12). The branching line style indicates the group affiliations of the insect hosts. The scale bar represents a distance of 20%.