Genome comparison of long-circulating field CnmeGV isolates from the same region.

Cnaphalocrocis medinalis granulovirus (CnmeGV), belonging to Betabaculovirus cnamedinalis, can infect the rice pest, the rice leaf roller. In 1979, a CnmeGV isolate, CnmeGV-EP, was collected from Enping County, China. In 2014, we collected another CnmeGV isolate, CnmeGV-EPDH3, at the same location and obtained the complete virus genome sequence using Illumina and ONT sequencing technologies. By combining these two virus isolates, we updated the genome annotation of CnmeGV and conducted an in-depth analysis of its genome features. CnmeGV genome contains abundant tandem repeat sequences, and the repeating units in the homologous regions (hrs) exhibit overlapping and nested patterns. The genetic variations within EPDH3 population show the high stability of CnmeGV genome, and tandem repeats are the only region of high genetic variation in CnmeGV genome replication. Some defective viral genomes formed by recombination were found within the population. Comparison analysis of the two virus isolates collected from Enping showed that the proteins encoded by the CnmeGV-specific genes were less conserved relative to the baculovirus core genes. At the genomic level, there are a large number of SNPs and InDels between the two virus isolates, especially in and around the bro genes and hrs. Additionally, we discovered that CnmeGV acquired a segment of non-ORF sequence from its host, which does not provide any new proteins but rather serves as redundant genetic material integrated into the viral genome. Furthermore, we observed that the host's transposon piggyBac has inserted into some virus genes. Together, dsDNA viruses could acquire non-coding genetic material from their hosts to expand the size of their genomes. These findings provide new insights into the evolution of dsDNA viruses.

Genome analysis of Psilogramma increta granulovirus and its intrapopulation diversity.

The complete genome of Psilogramma increta granulovirus (PsinGV), isolated from P. increta (Lepidoptera: Sphingidae), was ultra-deep sequenced with a Novaseq PE150 platform and de novo assembled and annotated. The PsinGV genome is a circular double-stranded DNA, 103,721 bp in length, with a G+C content of 33.0%, the third lowest G+C content in present sequenced baculoviruses. It encodes 123 putative open reading frames, including 38 baculovirus core genes, 42 lepidopteran baculovirus conserved genes, 38 betabaculovirus conserved genes, and 5 genes unique to PsinGV. Meanwhile, 3 homologous repeated regions with the core sequence TTGCAA and 3 direct repeated sequences were identified within the PsinGV genome. Kimura two-parameters distance analysis confirmed that Psilogramma increta granulovirus is a representative of a prospective new species of the genus Betabaculovirus. Phylogenetic analysis based on the baculovirus core genes showed that PsinGV is closely related to Choristoneura fumiferana granulovirus, Clostera anastomosis granulovirus-B, and Erinnyis ello granulovirus. These four species therefore share a common ancestor residing in the Betabaculovirus genus. The genome of the PsinGV isolate contained two p10 copies: p10 and p10-2. Phylogenetic reconstruction of P10 suggests a transfer event of the p10-2 gene from an alphabaculovirus to the aforementioned common ancestor. Analysis of genomic diversity showed that 203 intrahost variants, including 182 single nucleotide variants and 21 short insertions/deletions, are present within the PsinGV isolate. Meanwhile, allele frequency indicated that the isolate contains three major genotypes, implying the archived isolate consists of several P. increta carcasses killed by PsinGV with different genotypes.

The wide distribution and horizontal transfers of beta satellite DNA in eukaryotes.

Beta satellite DNA (satDNA), also known as Sau3A sequences, are repetitive DNA sequences reported in human and primate genomes. It is previously thought that beta satDNAs originated in old world monkeys and bursted in great apes. In this study, we searched 7821 genome assemblies of 3767 eukaryotic species and found that beta satDNAs are widely distributed across eukaryotes. The four major branches of eukaryotes, animals, fungi, plants and Harosa/SAR, all have multiple clades containing beta satDNAs. These results were also confirmed by searching whole genome sequencing data (SRA) and PCR assay. Beta satDNA sequences were found in all the primate clades, as well as in Dermoptera and Scandentia, indicating that the beta satDNAs in primates might originate in the common ancestor of Primatomorpha or Euarchonta. In contrast, the widely patchy distribution of beta satDNAs across eukaryotes presents a typical scenario of multiple horizontal transfers.

Autographa californica Multiple Nucleopolyhedrovirus ac75 Is Required for the Nuclear Egress of Nucleocapsids and Intranuclear Microvesicle Formation.

Autographa californica multiple nucleopolyhedrovirus (AcMNPV) orf75 (ac75) is a highly conserved gene of unknown function. In this study, we constructed an ac75 knockout AcMNPV bacmid and investigated the role of ac75 in the baculovirus life cycle. The expression and distribution of the Ac75 protein were characterized, and its interaction with another viral protein was analyzed to further understand its function. Our data indicated that ac75 was required for the nuclear egress of nucleocapsids, intranuclear microvesicle formation, and subsequent budded virion (BV) formation, as well as occlusion-derived virion (ODV) envelopment and embedding of ODVs into polyhedra. Western blot analyses showed that two forms, of 18 and 15 kDa, of FLAG-tagged Ac75 protein were detected. Ac75 was associated with both nucleocapsid and envelope fractions of BVs but with only the nucleocapsid fraction of ODVs; the 18-kDa form was associated with only BVs, whereas the 15-kDa form was associated with both types of virion. Ac75 was localized predominantly in the intranuclear ring zone during infection and exhibited a nuclear rim distribution during the early phase of infection. A phase separation assay suggested that Ac75 was not an integral membrane protein. A coimmunoprecipitation assay revealed an interaction between Ac75 and the integral membrane protein Ac76, and bimolecular fluorescence complementation assays identified the sites of the interaction within the cytoplasm and at the nuclear membrane and ring zone in AcMNPV-infected cells. Our results have identified ac75 as a second gene that is required for both the nuclear egress of nucleocapsids and the formation of intranuclear microvesicles.IMPORTANCE During the baculovirus life cycle, the morphogenesis of both budded virions (BVs) and occlusion-derived virions (ODVs) is proposed to involve a budding process at the nuclear membrane, which occurs while nucleocapsids egress from the nucleus or when intranuclear microvesicles are produced. However, the exact mechanism of virion morphogenesis remains unknown. In this study, we identified ac75 as a second gene, in addition to ac93, that is essential for the nuclear egress of nucleocapsids, intranuclear microvesicle formation, and subsequent BV formation, as well as ODV envelopment and embedding of ODVs into polyhedra. Ac75 is not an integral membrane protein. However, it interacts with an integral membrane protein (Ac76) and is associated with the nuclear membrane. These data enhance our understanding of the commonalities between nuclear egress of nucleocapsids and intranuclear microvesicle formation and may help to reveal insights into the mechanism of baculovirus virion morphogenesis.