Negeviruses are a proposed group of insect-specific viruses that can be separated into two distinct phylogenetic clades, Nelorpivirus and Sandewavirus. Negeviruses are well-known for their wide geographic distribution and broad host range among hematophagous insects. In this study, the full genomes of two novel negeviruses from each of these clades were identified by RNA extraction and sequencing from a single dungfly (Scathophaga furcata) collected from the Arctic Yellow River Station, where these genomes are the first negeviruses from cold zone regions to be discovered. Nelorpivirus dungfly1 (NVD1) and Sandewavirus dungfly1 (SVD1) have the typical negevirus genome organization and there was a very high coverage of viral transcripts. Small interfering RNAs derived from both viruses were readily detected in S. furcata, clearly showing that negeviruses are targeted by the host antiviral RNA interference (RNAi) pathway. These results and subsequent in silico analysis (studies) of public database and published virome data showed that the hosts of nege-like viruses include insects belonging to many orders as well as various non-insects in addition to the hematophagous insects previously reported. Phylogenetic analysis reveals at least three further groups of negeviruses, as well as several poorly resolved solitary branches, filling in the gaps within the two sub-groups of negeviruses and plant-associated viruses in the Kitaviridae. The results of this study will contribute to a better understanding of the geographic distribution, host range, evolution and host antiviral immune responses of negeviruses.

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The cabbage whitefly (A. proletella) (Hemiptera: Aleyrodidae) is a major agricultural pest that primarily targets cruciferous crops, such as cabbage, broccoli, and kale, causing extensive damage through feeding and honeydew. However, its associated virome has received limited research attention. In this study, we collected cabbage whiteflies in Xinjiang Agricultural University (43.80833 N, 87.56778 E, 882.3 m), systematically identified the RNA virome of the A. proletella and successfully identified three novel iflaviruses (Aleyrodes proletella iflavirus 1 (APIV1), Aleyrodes proletella iflavirus 2 (APIV2) and Aleyrodes proletella iflavirus 3 (APIV3)). APIV1–3 all have a 5′ structural protein region and a 3’ non-structural protein region. Phylogenetic and sequence identity analyses suggest that APIV1–3 are novel members of the family Iflaviridae. Structural modeling using AlphaFold3 revealed a conserved protein core region and a variable outer loop region. This study provides valuable insights into the virome diversity of A. proletella, establishing a foundation for future research on virus–host interactions and the potential for biocontrol applications in sustainable agriculture.