Adapting the rhizome concept to an extended definition of viral quasispecies and the implications for molecular evolution.

The rhizome concept proposed by Gilles Deleuze and Félix Guattari offers a novel perspective on the organization and interdependence of complex constellations of heterogeneous entities, their mapping and their ruptures. The emphasis of the present study is placed on the dynamics of contacts and communication among such entities that arise from experimentation, without any favored hierarchy or origin. When applied to biological evolution, the rhizome concept integrates all types of heterogeneity resulting from "symbiotic" relationships among living beings (or their genomic material), horizontal genetic transfer, recombination and mutation, and breaks away from the approach that gives rise to the phylogenetic tree of life. It has already been applied to describe the dynamics and evolution of RNA viruses. Thus, here we introduce a novel framework for the interpretation the viral quasispecies concept, which explains the evolution of RNA virus populations as the result of dynamic interconnections and multifaceted interdependence between highly heterogeneous viral sequences and its inherently heterogeneous host cells. The rhizome network perspective underlines even further the medical implications of the broad mutant spectra of viruses that are in constant flow, given the multiple pathways they have available for fitness loss and gain.

Prevalence and molecular analysis of some important viruses in honey bee colonies in Türkiye: the status of multiple infections.

In this study, seven bee viruses of significant importance for bee health in Türkiye were investigated using one-step RT-PCR. For this purpose, larvae from 1183 hives and adult bees from 1196 hives were sampled from 400 apiaries in 40 provinces. The prevalence of viral infections in hives was as follows: acute bee paralysis virus (ABPV), 6.4%; black queen cell virus (BQCV), 77%; chronic bee paralysis virus (CBPV), 3.2%; deformed wing virus (DWV), 63.8%; Israel acute bee paralysis virus (IAPV), 7%; Kashmir bee virus (KBV), 2.7%; sacbrood virus (SBV), 49.7%. Moreover, 50 different combinations of viral infections were identified in the hives. While dual infections (36.1%) were the most common in hives, triple infections with BQCV, DWV, and SBV were found to have the highest prevalence (22.1%). At least one viral infection was detected in all of the apiaries tested. Phylogenetic analysis showed that the isolates from this study generally exhibited the highest similarity to previously reported Turkish isolates. When similarity ratios and the locations and types of amino acid mutations were analyzed, it was observed that the isolates from our study exhibited high similarity to isolates from various countries, including China, the United Kingdom, Syria, and Germany.

Decoding the RNA viromes in shrew lungs along the eastern coast of China.

Shrews being insectivores, serve as natural reservoirs for a wide array of zoonotic viruses, including the recently discovered Langya henipavirus (LayV) in China in 2018. It is crucial to understand the shrew-associated virome, viral diversity, and new viruses. In the current study, we conducted high-throughput sequencing on lung samples obtained from 398 shrews captured along the eastern coast of China, and characterized the high-depth virome of 6 common shrew species (Anourosorex squamipes, Crocidura lasiura, Crocidura shantungensis, Crocidura tanakae, Sorex caecutiens, and Suncus murinus). Our analysis revealed numerous shrew-associated viruses comprising 54 known viruses and 72 new viruses that significantly enhance our understanding of mammalian viruses. Notably, 34 identified viruses possess spillover-risk potential and six were human pathogenic viruses: LayV, influenza A virus (H5N6), rotavirus A, rabies virus, avian paramyxovirus 1, and rat hepatitis E virus. Moreover, ten previously unreported viruses in China were discovered, six among them have spillover-risk potential. Additionally, all 54 known viruses and 12 new viruses had the ability to cross species boundaries. Our data underscore the diversity of shrew-associated viruses and provide a foundation for further studies into tracing and predicting emerging infectious diseases originated from shrews.

Molecular characterization of a novel mitovirus from the plant-pathogenic fungus Nigrospora oryzae.

Here, we characterized a novel mitovirus from the fungus Nigrospora oryzae, which was named "Nigrospora oryzae mitovirus 3" (NoMV3). The NoMV3 genome is 2,492 nt in length with a G + C content of 33%, containing a single large open reading frame (ORF) using the fungal mitochondrial genetic code. The ORF encodes an RNA-dependent RNA polymerase (RdRp) of 775 amino acids with a molecular mass of 88.75 kDa. BLASTp analysis revealed that the RdRp of NoMV3 had 68.6%, 50.6%, and 48.6% sequence identity to those of Nigrospora oryzae mitovirus 2, Suillus luteus mitovirus 6, and Fusarium proliferatum mitovirus 3, respectively, which belong to the genus Unuamitovirus within the family Mitoviridae. Phylogenetic analysis based on amino acid sequences supported the classification of NoMV3 as a member of a new species in the genus Unuamitovirus within the family Mitoviridae.

Spatiotemporal evolution of the distribution of Chronic bee paralysis virus (CBPV) in honey bee colonies.

Chronic bee paralysis virus (CBPV) is a Apis mellifera viral infectious disease, exhibiting dark and hairless abdomen in workers with tremors and ataxita. Clinical signs are also typically linked to adverse weather conditions and overcrowding in the hive. The disease occurs in spring but recently it has been observed cases increase and seasonality loss of the disease incidence. This study analyses the evolution of CBPV in Italy, through data collected from 2009 to 2023 within three monitoring projects comprising nationwide extended detection networks, aimed to investigate the evolution of the CBPV spatial distribution, identifying high-risk areas for the virus spread. This study highlights an increased risk over years. Prevalence increased from 4.3% during 2009-2010 to 84.7% during 2021-2023 monitoring years. CBPV outbreaks were irregular between investigated seasons, highlighting Spring and Autumn as the most susceptible seasons. Risk of CBPV infection has increased, reaching high-risk in last years of monitoring. Sequence analysis showed a high similarity to other isolated Italian CBPVs. The study offers an epidemiological insight into the aetiology of this disease. CBPV distribution is a prerequisite to predict its future spread and factors involved in its propagation not only in honey bees but also in other pollinators and environments.

Diversity of Picorna-Like Viruses in the Teltow Canal, Berlin, Germany.

The viromes of freshwater bodies are underexplored. The Picornavirales order, with 371 acknowledged species, is one of the most expansive and diverse groups of eukaryotic RNA viruses. In this study, we add 513 picorna-like viruses to the assemblage of more than 2000 unassigned picorna-like viruses. Our set of the aquatic Picornavirales virome of the Teltow Canal in Berlin, Germany, consists of 239 complete and 274 partial genomes. This urban freshwater body is characterized by the predominance of marna-like viruses (30.8%) and dicistro-like viruses (19.1%), whereas picornaviruses, iflaviruses, solinvi-like viruses, polycipi-like viruses, and nora-like viruses are considerably less prevalent. Caliciviruses and secoviruses were absent in our sample. Although presenting characteristic domains of Picornavirales, more than 100 viruses (20.8%) could not be assigned to any of the 9 Picornavirales families. Thirty-three viruses of the Marnaviridae-mostly locarna-like viruses-exhibit a monocistronic genome layout. Besides a wealth of novel virus sequences, viruses with peculiar features are reported. Among these is a clade of untypeable marna-like viruses with dicistronic genomes, but with the capsid protein-encoding open reading frame located at the 5' part of their RNA. A virus with a similar genome layout but clustering with dicistroviruses was also observed. We further detected monocistronic viruses with a polymerase gene related to aparaviruses. The detection of Aichi virus and five novel posa-like viruses indicates a slight burden in municipal wastewater.

Metatranscriptomic Sequencing of Sheath Blight-Associated Isolates of Rhizoctonia solani Revealed Multi-Infection by Diverse Groups of RNA Viruses.

Rice sheath blight, caused by the soil-borne fungus Rhizoctonia solani (teleomorph: Thanatephorus cucumeris, Basidiomycota), is one of the most devastating phytopathogenic fungal diseases and causes yield loss. Here, we report on a very high prevalence (100%) of potential virus-associated double-stranded RNA (dsRNA) elements for a collection of 39 fungal strains of R. solani from the rice sheath blight samples from at least four major rice-growing areas in the Philippines and a reference isolate from the International Rice Research Institute, showing different colony phenotypes. Their dsRNA profiles suggested the presence of multiple viral infections among these Philippine R. solani populations. Using next-generation sequencing, the viral sequences of the three representative R. solani strains (Ilo-Rs-6, Tar-Rs-3, and Tar-Rs-5) from different rice-growing areas revealed the presence of at least 36 viruses or virus-like agents, with the Tar-Rs-3 strain harboring the largest number of viruses (at least 20 in total). These mycoviruses or their candidates are believed to have single-stranded RNA or dsRNA genomes and they belong to or are associated with the orders Martellivirales, Hepelivirales, Durnavirales, Cryppavirales, Ourlivirales, and Ghabrivirales based on their coding-complete RNA-dependent RNA polymerase sequences. The complete genome sequences of two novel RNA viruses belonging to the proposed family Phlegiviridae and family Mitoviridae were determined.

Sacbrood viruses and select Lake Sinai virus variants dominated Apis mellifera colonies symptomatic for European foulbrood.

European foulbrood (EFB) is a prevalent disease in the European honey bee (Apis mellifera) in the United States, which can lead to colony decline and collapse. The bacterial components of EFB are well-studied, but the diversity of viral infections within infected colonies has not been explored. In this study, we use meta-transcriptomics sequencing of 12 honey bee hives, symptomatic (+, n = 6) and asymptomatic (-, n = 6) for EFB, to investigate viral infection associated with the disease. We assembled 41 viral genomes, belonging to three families (Iflaviridae, Dicistroviridae, and Sinhaliviridae), all previously reported in honey bees, including Lake Sinai virus, deformed wing virus, sacbrood virus, Black queen cell virus, and Israeli acute paralysis virus. In colonies with severe EFB, we observed a higher occurrence of viral genomes (34 genomes) in contrast to fewer recovered from healthy colonies (seven genomes) and a complete absence of Dicistroviridae genomes.We observed specific Lake Sinai virus clades associated exclusively with EFB + or EFB - colonies, in addition to EFB-afflicted colonies that exhibited an increase in relative abundance of sacbrood viruses. Multivariate analyses highlighted that a combination of site and EFB disease status influenced RNA virome composition, while EFB status alone did not significantly impact it, presenting a challenge for comparisons between colonies kept in different yards. These findings contribute to the understanding of viral dynamics in honey bee colonies compromised by EFB and underscore the need for future investigations to consider viral composition when investigating EFB.IMPORTANCEThis study on the viromes of honey bee colonies affected by European foulbrood (EFB) sheds light on the dynamics of viral populations in bee colonies in the context of a prevalent bacterial brood disease. The identification of distinct Lake Sinai virus and sacbrood virus clades associated with colonies affected by severe EFB suggests a potential connection between viral composition and disease status, emphasizing the need for further investigation into the role of viruses during EFB infection. The observed increase in sacbrood viruses during EFB infection suggests a potential viral dysbiosis, with potential implications for honey bee brood health. These findings contribute valuable insights related to beekeeping practices, offering a foundation for future research aimed at understanding and mitigating the impact of bacterial and viral infection in commercial honey bee operations and the management of EFB.

Molecular investigation and infection patterns of seven viruses of honey bee (Apis mellifera L, 1758) populations from southeastern Morocco.

An epidemiological survey of honey bee viruses was conducted on 87 clinically healthy beehives located in southeastern Morocco. The sampled colonies were analyzed by reverse transcriptase (RT)-PCR / Real Time RT-qPCR with the aim of detecting and / or quantifying the following viruses: acute bee paralysis virus (ABPV), chronic bee paralysis virus (CBPV), deformed wing virus (DWV), sacbrood virus (SBV), black queen cell virus (BQCV), Kashmir bee virus (KBV) and Israeli acute paralysis virus (IAPV). With the exception of the last two of these viruses, all the other five were detected with different prevalence rates. DWV showed the highest prevalence rate (89.65 %), followed by BQCV (17.24 %), ABPV (8.04 %), CBPV (4.59 %), and SBV (2.29 %). This study represents the first molecular detection of BQCV in the country. Among all investigated colonies, only eight were virus free (9.2 %). By contrast, single infection was detected in 64.37 % of colonies, 21.8 % showed mixed infection with two viruses, while 4.6 % showed three. Nucleotide sequences of a portion of the DWV polyprotein gene obtained for six honey bee samples showed the greatest nucleotide identity with sequences of DWV from Sweden and Ireland. The negative effect of migratory beekeeping as opposed to stationary beekeeping was highlighted given that stationary beehives showed infection with up to three viruses only, while migratory beehives showed up to five viruses. The results of this study are of crucial importance as they shed light on the current status of honey bee health in southeastern Morocco.

Complete genome analysis of a novel hypovirus in the phytopathogenic fungus Monilinia fructicola.

Monilinia fructicola is one of the most devastating fungal diseases of rosaceous fruit crops, both in the field and postharvest, causing significant yield losses. Here, we report the discovery of a novel positive single-stranded RNA virus, Monilinia fructicola hypovirus 3 (MfHV3), in a strain (hf-1) of the phytopathogenic fungus Monilinia fructicola. The complete genome of MfHV3 is 9259 nucleotides (nt) in length and contains a single large open reading frame (ORF) from nt position 462 to 8411. This ORF encodes a polyprotein with three conserved domains, namely UDP-glycosyltransferase, RNA-dependent RNA polymerase (RdRp), and DEAD-like helicase. The MfHV3 polyprotein shares the highest similarity with Colletotrichum camelliae hypovirus 1. Phylogenetic analysis indicated that MfHV3 clustered with members of the genus Betahypovirus within the family Hypoviridae. Taken together, the results of genomic organization comparisons, amino acid sequence alignments, and phylogenetic analysis convincingly show that MfHV3 is a new member of the genus Betahypovirus, family Hypoviridae.

Complete genome sequence of a novel botourmiavirus infecting the fungus Phomopsis asparagi.

Here, we report a novel ourmia-like mycovirus, named "Phomopsis asparagi magoulivirus 1" (PaMV1), derived from the phytopathogenic fungus Phomopsis asparagi. The genome of PaMV1 consists of a positive-sense single-stranded RNA (+ ssRNA) that is 2,639 nucleotides in length, with a GC content of 57.13%. It contains a single open reading frame (ORF) encoding a putative RNA-dependent RNA polymerase (RdRp) consisting of 686 amino acids with a molecular mass of 78.57 kDa. Phylogenetic analysis based on RdRp sequences revealed that PaMV1 grouped together with Diaporthe gulyae magoulivirus 1 (DgMV1) in a distinct clade. Sequence comparisons and phylogenetic analysis suggest that PaMV1 is a novel member of the genus Magoulivirus, family Botourmiaviridae.

Varroa destructor shapes the unique viral landscapes of the honey bee populations of the Azores archipelago.

The worldwide dispersal of the ectoparasitic mite Varroa destructor from its Asian origins has fundamentally transformed the relationship of the honey bee (Apis mellifera) with several of its viruses, via changes in transmission and/or host immunosuppression. The extent to which honey bee-virus relationships change after Varroa invasion is poorly understood for most viruses, in part because there are few places in the world with several geographically close but completely isolated honey bee populations that either have, or have not, been exposed long-term to Varroa, allowing for separate ecological, epidemiological, and adaptive relationships to develop between honey bees and their viruses, in relation to the mite's presence or absence. The Azores is one such place, as it contains islands with and without the mite. Here, we combined qPCR with meta-amplicon deep sequencing to uncover the relationship between Varroa presence, and the prevalence, load, diversity, and phylogeographic structure of eight honey bee viruses screened across the archipelago. Four viruses were not detected on any island (ABPV-Acute bee paralysis virus, KBV-Kashmir bee virus, IAPV-Israeli acute bee paralysis virus, BeeMLV-Bee macula-like virus); one (SBV-Sacbrood virus) was detected only on mite-infested islands; one (CBPV-Chronic bee paralysis virus) occurred on some islands, and two (BQCV-Black queen cell virus, LSV-Lake Sinai virus,) were present on every single island. This multi-virus screening builds upon a parallel survey of Deformed wing virus (DWV) strains that uncovered a remarkably heterogeneous viral landscape featuring Varroa-infested islands dominated by DWV-A and -B, Varroa-free islands naïve to DWV, and a refuge of the rare DWV-C dominating the easternmost Varroa-free islands. While all four detected viruses investigated here were affected by Varroa for one or two parameters (usually prevalence and/or the Richness component of ASV diversity), the strongest effect was observed for the multi-strain LSV. Varroa unambiguously led to elevated prevalence, load, and diversity (Richness and Shannon Index) of LSV, with these results largely shaped by LSV-2, a major LSV strain. Unprecedented insights into the mite-virus relationship were further gained from implementing a phylogeographic approach. In addition to enabling the identification of a novel LSV strain that dominated the unique viral landscape of the easternmost islands, this approach, in combination with the recovered diversity patterns, strongly suggests that Varroa is driving the evolutionary change of LSV in the Azores. This study greatly advances the current understanding of the effect of Varroa on the epidemiology and adaptive evolution of these less-studied viruses, whose relationship with Varroa has thus far been poorly defined.

A novel betapartitivirus isolated from Cordyceps militaris, an edible-medicinal mushroom.

In this study, we identified a novel partitivirus, named "Cordyceps militaris partitivirus 1" (CmPV1), in Cordyceps militaris strain RCEF7506. The complete genome of CmPV1 comprises two segments, dsRNA1 and dsRNA2, each encoding a single protein. dsRNA1 (2,206 bp) encodes an RNA-dependent RNA polymerase (RdRp), and dsRNA2 (2,256 bp) encodes a coat protein (CP). Sequence analysis revealed that dsRNA1 has the highest similarity to that of Bipolaris maydis partitivirus 2 (BmPV2), whereas dsRNA2 shows the highest similarity to human blood-associated partitivirus (HuBPV). Phylogenetic analysis based on RdRp sequences suggests that CmPV1 is a new member of the genus Betapartitivirus of the family Partitiviridae. This is the first documentation of a betapartitivirus infecting the entomopathogenic fungus C. militaris.

Identification of a novel member of the genus Laulavirus (family Phenuiviridae) from the entomopathogenic ascomycete fungus Cordyceps javanica.

The virus family Phenuiviridae (order Hareavirales, comprising segmented negative-sense single stranded RNA viruses) has highly diverse members that are known to infect animals, plants, protozoans, and fungi. In this study, we identified a novel phenuivirus infecting a strain of the entomopathogenic fungus Cordyceps javanica isolated from a small brown plant hopper (Laodelphax striatellus), and this virus was tentatively named "Cordyceps javanica negative-strand RNA virus 1" (CjNRSV1). The CjNRSV1 genome consists of three negative-sense single stranded RNA segments (RNA1-3) with lengths of 7252, 2401, and 1117 nt, respectively. The 3'- and 5'-terminal regions of the RNA1, 2, and 3 segments have identical sequences, and the termini of the RNA segments are complementary to each other, reflecting a common characteristic of viruses in the order Hareavirales. RNA1 encodes a large protein (∼274 kDa) containing a conserved domain for the bunyavirus RNA-dependent RNA polymerase (RdRP) superfamily, with 57-80% identity to the RdRP encoded by phenuiviruses in the genus Laulavirus. RNA2 encodes a protein (∼79 kDa) showing sequence similarity (47-63% identity) to the movement protein (MP, a plant viral cell-to-cell movement protein)-like protein (MP-L) encoded by RNA2 of laulaviruses. RNA3 encodes a protein (∼28 kDa) with a conserved domain of the phenuivirid nucleocapsid protein superfamily. Phylogenetic analysis using the RdRPs of various phenuiviruses and other unclassified phenuiviruses showed CjNRSV1 to be grouped with established members of the genus Laulavirus. Our results suggest that CjNRSV1 is a novel fungus-infecting member of the genus Laulavirus in the family Phenuiviridae.

[Comparative analysis of the taxonomic classification criteria for a number of groups of pathogenic DNA and RNA viruses based on genomic data].

The basis for criteria of the taxonomic classification of DNA and RNA viruses based on data of the genomic sequencing are viewed in this review. The genomic sequences of viruses, which have genome represented by double-stranded DNA (orthopoxviruses as example), positive-sense single-stranded RNA (alphaviruses and flaviviruses as example), non-segmented negative-sense single-stranded RNA (filoviruses as example), segmented negative-sense single-stranded RNA (arenaviruses and phleboviruses as example) are analyzed. The levels of genetic variability that determine the assignment of compared viruses to taxa of various orders are established for each group of viruses.

Identification and molecular characterization of Wanghe virus: A novel member of the Solinviviridae family from mosquitoes, China.

Arboviruses have always been a significant public health concern. Metagenomic surveillance has expanded the number of novel, often unclassified arboviruses, especially mosquito-borne and mosquito-specific viruses. This report presents the first description of a novel single-stranded RNA virus, Wanghe virus, identified from mosquitoes that were collected in Shandong Province in 2022. In this study, a total of 4,795 mosquitoes were collected and then divided into 105 pools according to location and species. QRT-PCR and nested PCR were performed to confirm the presence of Wanghe virus, and its genomic features and phylogenetic relationships were further analyzed. Our results revealed that Wanghe virus was detected in 9 out of the 105 mosquito pools, resulting in a minimum infection rate (MIR) of 0.19 % (9/4,795). One complete genome sequence and three viral partial sequences were obtained from the Wanghe virus-positive pools. Pairwise distance analysis indicated that these amplified sequences shared high nucleotide identity. Phylogenetic analysis demonstrated that Wanghe virus is most closely related to Guiyang Solinvi-like virus 3, which belongs to Solinviviridae. Further analyses indicated that Wanghe virus is a new, unclassified member of Solinviviridae.

Highly divergent and diverse viral community infecting sylvatic mosquitoes from Northeast Brazil.

Mosquitoes can transmit several pathogenic viruses to humans, but their natural viral community is also composed of a myriad of other viruses such as insect-specific viruses (ISVs) and those that infect symbiotic microorganisms. Besides a growing number of studies investigating the mosquito virome, the majority are focused on few urban species, and relatively little is known about the virome of sylvatic mosquitoes, particularly in high biodiverse biomes such as the Brazilian biomes. Here, we characterized the RNA virome of 10 sylvatic mosquito species from Atlantic forest remains at a sylvatic-urban interface in Northeast Brazil employing a metatranscriptomic approach. A total of 16 viral families were detected. The phylogenetic reconstructions of 14 viral families revealed that the majority of the sequences are putative ISVs. The phylogenetic positioning and, in most cases, the association with a high RNA-dependent RNA polymerase amino acid divergence from other known viruses suggests that the viruses characterized here represent at least 34 new viral species. Therefore, the sylvatic mosquito viral community is predominantly composed of highly divergent viruses highlighting the limited knowledge we still have about the natural virome of mosquitoes in general. Moreover, we found that none of the viruses recovered were shared between the species investigated, and only one showed high identity to a virus detected in a mosquito sampled in Peru, South America. These findings add further in-depth understanding about the interactions and coevolution between mosquitoes and viruses in natural environments. IMPORTANCE: Mosquitoes are medically important insects as they transmit pathogenic viruses to humans and animals during blood feeding. However, their natural microbiota is also composed of a diverse set of viruses that cause no harm to the insect and other hosts, such as insect-specific viruses. In this study, we characterized the RNA virome of sylvatic mosquitoes from Northeast Brazil using unbiased metatranscriptomic sequencing and in-depth bioinformatic approaches. Our analysis revealed that these mosquitoes species harbor a diverse set of highly divergent viruses, and the majority comprises new viral species. Our findings revealed many new virus lineages characterized for the first time broadening our understanding about the natural interaction between mosquitoes and viruses. Finally, it also provided several complete genomes that warrant further assessment for mosquito and vertebrate host pathogenicity and their potential interference with pathogenic arboviruses.

The hidden RNA viruses in Blattodea (cockroaches and termites).

The insect order Blattodea (cockroaches and termites) has drawn substantial research attention for their dietary habits and lifestyle of living with or around humans. In the present study, we focused on the discovery of RNA viruses hidden in Blattodea insects using the publicly available RNA sequencing datasets. Overall, 136 distinctive RNA viruses were identified from 36 Blattodea species, of which more than 70 % were most closely related to the invertebrate-associated viral groups within Picornavirales, Sobelivirales, Bunyaviricetes, Jingchuvirales, Durnavirales, Lispiviridae, Orthomyxoviridae, Permutotetraviridae, Flaviviridae and Muvirales. Several viruses were associated with pathogens of vertebrates (Paramyxoviridae), plants (Tymovirales), protozoa (Totiviridae), fungi (Narnaviridae) and bacteria (Norzivirales). Collectively, 93 complete or near-complete viral genomes were retrieved from the datasets, and several viruses appeared to have remarkable temporal and spatial distributions. Interestingly, the newly identified Periplaneta americana dicistrovirus displayed a remarkable distinct bicistronic genome arrangement from the well-recognized dicistroviruses with the translocated structural and non-structural polyprotein encoding open reading frames over the genome. These results significantly enhance our knowledge of RNA virosphere in Blattodea insects, and the novel genome architectures in dicistroviruses and other RNA viruses may break our stereotypes in the understanding of the genomic evolution and the emergence of potential novel viral species.

Ambiviricota, a novel ribovirian phylum for viruses with viroid-like properties.

Fungi harbor a vast diversity of mobile genetic elements (MGEs). Recently, novel fungal MGEs, tentatively referred to as 'ambiviruses,' were described. 'Ambiviruses' have single-stranded RNA genomes of about 4-5 kb in length that contain at least two open reading frames (ORFs) in non-overlapping ambisense orientation. Both ORFs are conserved among all currently known 'ambiviruses,' and one of them encodes a distinct viral RNA-directed RNA polymerase (RdRP), the hallmark gene of ribovirian kingdom Orthornavirae. However, 'ambivirus' genomes are circular and predicted to replicate via a rolling-circle mechanism. Their genomes are also predicted to form rod-like structures and contain ribozymes in various combinations in both sense and antisense orientations-features reminiscent of viroids, virusoids, ribozyvirian kolmiovirids, and yet-unclassified MGEs (such as 'epsilonviruses,' 'zetaviruses,' and some 'obelisks'). As a first step toward the formal classification of 'ambiviruses,' the International Committee on Taxonomy of Viruses (ICTV) recently approved the establishment of a novel ribovirian phylum, Ambiviricota, to accommodate an initial set of 20 members with well-annotated genome sequences.

Identification and Genome Characterization of a Novel Nege-like Virus Isolated from Aphids (Aphis gossypii) in Yunnan Province.

Negeviruses are insect-specific enveloped RNA viruses that exhibit a wide geographic distribution. A novel nege-like virus, tentatively named Aphis gossypii nege-like virus (AGNLV, GenBank: OR880429.1), was isolated from aphids (Aphis gossypii) in Lijiang City, Yunnan, China. AGNLV has a genome sequence of 9258 nt (excluding the polyA tail) encoding three open reading frames (ORFs). ORF1 (7149 nt) encodes a viral methyltransferase, a viral RNA helicase, and an RNA-dependent RNA polymerase. ORF2 (1422 nt) encodes a DiSB-ORF2_chro domain and ORF3 encodes an SP24 domain. The genome sequence of AGNLV shares the highest nucleotide identity of 60.0% and 59.5% with Wuhan house centipede virus 1 (WHCV1) and Astegopteryx formosana nege-like virus (AFNLV), respectively. Phylogenetic analysis based on the RNA-dependent RNA polymerase shows that AGNLV is clustered with other negeviruses and nege-like viruses discovered in aphids, forming a distinct "unclassified clade". Interestingly, AGNLV only encodes three ORFs, whereas AFNLV and WHCV1 have four ORFs. Structure and transmembrane domain predictions show the presence of eight alpha helices and five transmembrane helices in the AGNLV ORF3. Translational enhancement of the AGNLV 5' UTR was similar to that of the 5' UTR of plant viruses. Our findings provide evidence of the diversity and structure of nege-like viruses and are the first record of such a virus from a member of the genus Aphis.