Biomics

Densoviruses (Densovirinae) are a subfamily of small, non-enveloped, single-stranded DNA viruses which predominantly infect invertebrates, including insects, crustaceans, and echinoderms. Despite being structurally simple, they exhibita significant genomic diversity and are widespread in nature. The key nonstructural protein of these viruses is NS1, which plays a central role in viral DNA replication, regulation of gene expression, and induction of apoptosis in the infected cells. NS1 contains conserved domains characteristic of parvoviruses.

In the present study, we have performed a phylogenetic analysis of the NS1 protein of Clinch densovirus 1 (ClDV1) isolated from the freshwater mussel Actinonaias pectorosa during a mass die-off of the mollusks in the Clinch River. On the basis of UPGMA analysis of amino acid sequences, we have constructed a phylogenetic tree comprising 59 NS1 proteins isolated from various sources, including insects, crustaceans, echinoderms, birds, and bats.

The analysis has revealed that Clinch densovirus 1 clusters with unclassified Ambidensoviruses, confirming its assignment to the genus Ambidensovirus. In some cases, closely related viruses are detected in the organisms that are not their typical hosts (birds, bats), suggesting a possibility of passive accumulation of the viruses through food chains rather than their active replication in these animals. NS1 is found to be a highly conserved marker suitable for phylogenetic classification of densoviruses.

The practical significance of this work lies in the fact that some of the studied viruses infect commercially important aquaculture species (crayfish Cherax quadricarinatus) or insect pests, this opening prospects for using densoviruses as bioinsecticides. At the same time, viruses infecting mass-reared feeder insects (Zophobas morio) pose an economic threat. The study underscores the need for further investigation into the ecology and transmission mechanisms of densoviruses in natural ecosystems.

Densovirinae, Clinch densovirus 1, UPGMA, NS1 protein, phylogenetic analysis

1. Aş Y, Eroğlu GB. Densovirinae: An Eco-Friendly Alternative in Biological Control. Zenodo. 2025. doi: 10.5281/zenodo.15877070
2. Batool K, Xiao J, Xu Y et al. Densovirus Oil Suspension Significantly Improves the Efficacy and Duration of Larvicidal Activity against Aedes albopictus. Viruses. 2022. 14(3). 475. doi: 10.3390/v14030475
3. Bochow S, Condon K, Elliman J, Owens L. First complete genome of an Ambidensovirus; Cherax quadricarinatus densovirus, from freshwater crayfish Cherax quadricarinatus. Marine Genomics. 2015. 24. 305-312. doi: 10.1016/j.margen.2015.07.009
4. Buck CB, Welch N, Belford AK et al. Widespread Horizontal Gene Transfer Among Animal Viruses. bioRxiv. 2024. doi: 10.1101/2024.03.25.586562
5. Canuti M, Verhoeven JTP, Munro HJ et al. Investigating the Diversity and Host Range of Novel Parvoviruses from North American Ducks Using Epidemiology, Phylogenetics, Genome Structure, and Codon Usage Analysis. Viruses. 2021. 13(2). 193. doi: 10.3390/v13020193
6. Dai Z, Wang H, Wu H et al. Parvovirus dark matter in the cloaca of wild birds. GigaScience. 2022. 12. giad001. doi: 10.1093/gigascience/giad001
7. Divekar G, Colmant AMG, Furlong MJ, Etebari K. Transcriptome Analysis Reveals a Diverse Range of Novel Viruses in Australian Sugarcane Soldier Fly (Inopus flavus) Larvae. Viruses. 2024. 16(4). 516. doi: 10.3390/v16040516
8. Erlandson MA. Insect Pest Control by Viruses. In: Mahy BWJ, Van Regenmortel MHV (eds.). Encyclopedia of Virology (Third Edition). Academic Press. 2008. 125-133. doi: 10.1016/B978-012374410-4.00432-5
9. Fahsbender E, Charlys da-Costa A, Gill DE et al. Plasma virome of 781 Brazilians with unexplained symptoms of arbovirus infection include a novel parvovirus and densovirus. PLoS One. 2020. 15(3). e0229993. doi: 10.1371/journal.pone.0229993
10. Felsenstein J. Confidence limits on phylogenies: An approach using the bootstrap. Evolution. 1985. 39. 783-791. doi: 10.1111/j.1558-5646.1985.tb00420.x
11. Grenet A-S, Salasc F, Francois S et al. The densovirus: A "massive attack" in the arthropods. Virologie. 2015. 19. 19-31. doi: 1684/vir.2015.0589
12. Guo H, Zhang J, Hu Y. Complete sequence and organization of Periplaneta fuliginosa densovirus genome. Acta virologica. 2001. 44. 315-322.
13. Guo Y, Zhao Y, Yang Y et al. Plants affect the horizontal transmission of a new densovirus infecting the green peach aphid Myzus persicae by modulating honeydew production. Insect Science. 2023. 31. doi: 10.1111/1744-7917.13235
14. Gutsche I, Coulibaly F, Voss JE et al. Secreted dengue virus nonstructural protein NS1 is an atypical barrel-shaped high-density lipoprotein. Proc Natl Acad Sci USA. 2011. 108(19). 8003-8008. doi: 10.1073/pnas.1017338108
15. Hu L, Zhang L, Shen C et al. The densovirus of Periplaneta fuliginosa (PfDNV) as an insect vector for persistent foreign gene expression in vivo. Biochemical and Biophysical Research Communications. 2007. 358. 976-982. doi: 10.1016/j.bbrc.2007.04.210
16. Jackson EW, Wilhelm RC, Johnson MR et al. Diversity of Sea Star-Associated Densoviruses and Transcribed Endogenous Viral Elements of Densovirus Origin. J Virol. 2020. 95(1). e01594-20. doi: 10.1128/JVI.01594-20
17. Jones DT, Taylor WR, Thornton JM. The rapid generation of mutation data matrices from protein sequences. Computer Applications in the Biosciences. 1992. 8. 275-282. doi: 10.1093/bioinformatics/8.3.275
18. Kapelinskaya TV, Martynova EU, Korolev AL et al. Transcription of the smoky brown cockroach densovirus BgDNV genome: alternative processing of viral RNAs. Doklady Akademii nauk. 2008. 421(2). 256-261. doi: 10.1134/S1607672908040042 (In Russian)
19. Kumar S, Stecher G, Li M et al. MEGA X: Molecular Evolutionary Genetics Analysis across computing platforms. Molecular Biology and Evolution. 2018. 35. 1547-1549. doi: 10.1093/molbev/msy096
20. Li Y, Ma Y, Liu X et al. Genome sequence of a new densovirus infecting the leaf-curl plum aphid Brachycaudus helichrysi. Archives of Virology. 2025. 170(4). 65. doi: 10.1007/s00705-025-06266-w
21. Lim FS, González-Cabrera J, Jehle JA et al. No longer uncertain: the validation of tenebrionid insects as hosts of Blattambidensovirus incertum1 isolates by phylogeny and infection studies. J Gen Virol. 2026. 107(2). 002211. doi: 10.1099/jgv.0.002211
22. Liu K, Li Y, Jousset FX et al. The Acheta domesticus densovirus, isolated from the European house cricket, has evolved an expression strategy unique among parvoviruses. J Virol. 2011. 85(19). 10069-10078. doi: 10.1128/JVI.00625-11
23. Maganga GD, Bourgarel M, Obame-Nkoghe J et al. Identification of an Unclassified Paramyxovirus in Coleura afra: A Potential Case of Host Specificity. PLoS One. 2014. 9. e115588. doi: 10.1371/journal.pone.0115588
24. McCallum B, Bakkeren G, Wang X et al. Comprehensive analysis of the wheat leaf virome associated with Puccinia triticina isolates across Canada reveals high viral diversity and novel mycoviruses. Phytopathol Res. 2026. 8. 64. doi: 10.1186/s42483-026-00443-8
25. Penzes JJ, Holm M, Yost SA, Kaelber JT. Cryo-EM-based discovery of a pathogenic parvovirus causing epidemic mortality by black wasting disease in farmed beetles. 2024. 187(20). 5604-5619.e14. doi: 10.1016/j.cell.2024.07.053
26. Pénzes J, Kaelber J. Identification by cryoEM of a densovirus causing mass mortality in mass-reared larval darkling beetles (Zophobas morio). bioRxiv. 2022. doi: 10.1101/2022.05.14.491968
27. Popov IV, Manakhov AD, Gorobets VE et al. Metagenomic Investigation of Intestinal Microbiota of Insectivorous Synanthropic Bats: Densoviruses, Antibiotic Resistance Genes, and Functional Profiling of Gut Microbial Communities. Int J Mol Sci. 2025. 26(13). 5941. doi: 10.3390/ijms26135941
28. Richard JC, Leis E, Dunn CD et al. Mass mortality in freshwater mussels (Actinonaias pectorosa) in the Clinch River, USA, linked to a novel densovirus. Scientific Reports. 2020. 10. 14498. doi: 10.1038/s41598-020-71459-z
29. Rudoy DV, Olshevskaya AV, Golovko LS et al. The most significant diseases of the Australian redclaw crayfish Cherax quadricarinatus (von Martens, 1868) during its cultivation in aquaculture conditions. Vestnik KrasGAU. 2025. (9). 123-140. doi: 10.36718/1819-4036-2025-9-123-140 (In Russian)
30. Sneath PHA, Sokal RR. Numerical Taxonomy. Freeman, San Francisco. 1973.
31. Sukhikh N, Golyaev V, Laboureau N et al. Deep Sequencing Analysis of Virome Components, Viral Gene Expression and Antiviral RNAi Responses in Myzus persicae Int J Mol Sci. 2024. 25(23). 13199. doi: 10.3390/ijms252313199
32. Wallace M, Coffman K, Gilbert C et al. The discovery, distribution and diversity of DNA viruses associated with Drosophila melanogaster in Europe. bioRxiv. 2020. doi: 10.1101/2020.10.16.342956