Characterization of Mint mosaic virus, a new member of the genus Carlavirus

Characterization of Mint mosaic virus, a new member of the genus Carlavirus | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Characterization of Mint mosaic virus, a new member of the genus Carlavirus Marco Forgia, Marta Vallino, Monica Marra, Paolo Mussano, Anna Paola Lanteri, and 2 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-5404341/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 10 Jan, 2025 Read the published version in Archives of Virology → Version 1 posted 5 You are reading this latest preprint version Abstract In this work the complete genome of a new carlavirus causing mosaic on mint plants in Italy is reported and tentatively named Mint mosaic virus (MMV). Flexuous particles of around 600 nm were observed through TEM analysis and NGS sequencing was performed to determine MMV genome. MMV genome is 8558 nt long, excluding the poly(A) tail and shows the typical organization of a carlavirus. The putative proteins coded by MMV have homology percentages ranging between 44% and 56% against the closest viruses in the NCBI database, suggesting that MMVcan be considered a new species in the genus Carlavirus. MMV was detected in independent mint samples from different Italian regions collected in years 2023 and 2024, suggesting a broad diffusion, at least in the Italian area. The two MMV isolates characterized in 2023 (Me1) and in 2024 (Me2) were included in the plant virus collection PLAVIT. Figures Figure 1 Fulltext Mint (genus Mentha , family Lamiaceae ) is commonly cultivated throughout the Mediterranean region, including Italy. Its cultivation is related to the different applications of its aromatic compounds, both in food production and traditional medicine, as well as for their antimicrobial potential (1). Knowledge of the viruses associated with this important plant is limited to few case studies. In fact, to date, the only virus characterized in mint is a potexvirus called Mint virus X, which was detected in “Variegata” mint in 2006 (2). However, the stralarivirus Strawberry latent ringspot virus and the begomovirus Chilli leaf curl India virus have been detected on mint in the USA and in India, respectively (3, 4). In fall 2023, leaves from Mentha spicata displaying mosaic symptoms were collected in the Liguria region of Italy (Supp. Figure 1 ). To determine the cause of the observed symptoms, the plant sample was mechanically inoculated on test plants as a viral infection was suspected. After one week, systemic symptoms were detected on Nicotiana. benthamiana . Both the original mint sample and symptomatic N. benthamiana leaves were used for transmission electron microscopy analysis (TEM). TEM observations were conducted by applying raw leaf extract for a few minutes onto carbon and formvar-coated grids and staining it with 0.5% uranyl acetate. Observations were made on a CM 10 electron microscope (Philips, Eindhoven, Netherlands). TEM analysis revealed filamentous and flexuous particles of around 600 nm in length and 13–15 nm in diameter in both samples (Fig. 1 b). To determine the genome sequence of the isolated virus, Illumina sequencing of total RNA was performed. Total RNA was extracted from systemically infected N. benthamiana leaves grinded in a 2 ml O-ring tube with 0.3 mm glass beads using a FastPrep-24™ bead beating grinder (MP Biomedicals™, Irvine, CA, USA). Grinded leaves were then processed for RNA extraction using Spectrum™ Plant Total RNA Kit (Merk, Darmstadt, Germany). Ribosomal depletion, library construction and sequencing of the total RNA were performed by Macrogen Europe (Amsterdam, Netherlands) using the TruSeq Stranded Total RNA with Ribo-Zero Plant kit (Illumina) on a Novaseq 6000 platform. Sequencing results were cleaned with bbtools (sourceforge.net/projects/bbmap/) and assembled using Trinity (ver. 2.9.1) (5) as described in previous works (6). The obtained metatranscriptome was searched for viral-related contigs with a blast against the NCBI nr viral database (taxid: 2559587) using DIAMOND (7). Each contig giving a viral hit was blasted against the nr database to detect false positive, and the results were manually checked, retrieving a carlavirus-related contig with a poly(A) tail. The leaves of the plants mechanically inoculated with MMV were lyophilized and included in the Plant Virus Italy (PLAVIT) collection with the ID Me1, stored at -20 C°. To obtain the complete viral sequence, we performed 5’-rapid amplification of cDNA analysis (5’-RACE). RACE was performed using the RLM described in Forgia et al. 2022, the total RNA was ligated with a blocked DNA oligo (5’-Phosphate-GCATTCGACCCCGGGTT-AmC3-3’) using T4 RNA Ligase (Takara, Japan) and cDNA was then synthetized with a primer complementary to the adaptor (5’-AACCCGGGGTCGAATGC-3’). Obtained cDNA was used as template for RT-PCR using two specific primers (MMV 320 Rev: 5’-TCTTGCACACTGGGTGGGAA-3’, MMV 361 Rev: 5’-AAGCGGCGGAATGACTACATACAA-3’), the obtained amplicon was cloned in pGEM-T Easy (Promega, Madison, Wisconsin, USA) and sequenced through Sanger method by Biofab s.r.l. (Rome, Italy). From the 6 clones sequenced, we reconstructed the complete genome of the virus, tentatively named mint mosaic virus (MMV) which is 8558 nt long excluding the poly(A) and encode 6 putative proteins (Fig. 1 a). The putative protein sequences were retrieved using ORFinder and the conserved domains were investigated with the NCBI conserved domain search tool ( https://www.ncbi.nlm.nih.gov/Structure/cdd/wrpsb.cgi ) from 5’ to 3’. The first ORF encodes a putative protein of 1,967 aa that shows a Betaflexiviridae_RdRp domain, a Viral_helicase1 domain, a Vmethyltransf domain, a Peptidase_C23 domain, an AlkB domain and an OTU_RDRP-like domain. The second, third and fourt ORFs encode proteins of 231, 110 and 280 aa long and exhibit the typical conserved domains of carlavirus triple gene block (Viral_helicase1 for TGB1, Plant_vir_prot for TGB2 and TGBp3 domain for TGB3). The fifth ORF encodes a 280 aa long protein showing a conserved Flexi_CP domain, while the last ORF codes for a 111 aa protein with a conserved Carla_C4 domain. Each putative protein was blasted against the NCBI nr database to detect homology against the closest known viruses. Results in Table 1 show that the replicase protein, along with the proteins from triple gene block 1 and 2, and the coat protein, have Poplar mosaic virus (PopMV) as the closest hit, with a homology percentage ranging between 45 and 56%. The protein from triple gene block 3 and the C4 protein show similar homology percentages against Apple stem pitting foveavirus (ASPV) and Cowpea mild mottle virus (CMMV), respectively. To perform phylogenetic analysis, the replicase of all the viruses accepted by the ICTV in the Betaflexiviridae family ( https://ictv.global/msl ) were retrieved and manually selected to remove partial and redundant sequences. The selected proteins (listed in Supp. Table 1) were aligned with MAFFT (8) including the ORF1 replicase sequence of MMV and the multiple alignment obtained was submitted to IQ-TREE 2 (9) using the automatic model selection (10) and ultrafast bootstrap with 1000 replicates (11) to build a maximum likelihood phylogenetic tree of the whole viral family. Results in Fig. 1 c collocate the new virus in the genus Carlavirus , forming a branch with PopMV as already suggested by the blast analysis (Table 1). Taken together these results and considering the species demarcation criteria for the genus Carlavirus (Distinct species have less than about 72% nt identity or 80% aa identity between their CP or polymerase genes, https://ictv.global/report_9th/RNApos/Betaflexiviridae ) MMV should be accomodated in a new species in the genus, for which we tentatively propose the name Carlavirus menthae. In summer 2024, a new mint sample showing mosaic symptoms was collected in the Calabria region of Italy. TEM analysis suggested a possible infection by MMV (not shown) thus, the virus was isolated in greenhouse as previously described and qRT-PCR primers were designed to validate an infection from MMV (MMV_RT_For: 5’-CATTCCGCCGCTTTTGGATA-3’, MMV_RT_Rev: 5’-AAATGAGGGAGAGGTTGCGA-3’). qRT-PCR was performed in 10 µl of total volume using the iTaq Universal SYBR Green Supermix on a CFX apparatus (Biorad, Hercules, CA, USA). Results of the qRT-PCRconfirmed the presence of MMV also in the sample from 2024, demonstrating that the virus is present in different Italian regions and could be detected consecutively for two years. The MMV viral accession from 2024 was included in the PLAVIT collection with the ID Me2, stored as dried leaf at -20 C°. In conclusion, this work characterized a new carlavirus infecting mint that was tentatively named mint mosaic virus. The virus was detected in two independent sampling in Italy and the two isolates identified are stored and maintained in the plant virus collection PLAVIT at the IPSP-CNR institute. Results suggest that that surveillance on symptomatic plants is still important to detect and characterize new viral pathogens or potential cross species infection by viral agents. Declarations Acknowledgements This research was funded by the European Virus Archive GLOBAL (EVA-GLOBAL) project that has received funding from the European Union’s Horizon 2020 research and innovation program under grant agreement No. 871029, and by the SUS-MIRRI.IT project, granted by the European Commission—NextGenerationEU, code n. IR0000005. Data availability The complete viral genome of MMV Me1 accession is deposited in the NCBI database with the accession number PQ562895. References Gupta S, Kumar A, Gupta AK, Jnanesha AC, Talha M, Srivastava A. Industrial mint crop revolution, new opportunities, and novel cultivation ambitions: A review. Ecological Genetics and Genomics. 2023;27. Tzanetakis IE, Postman JD, Martin RR. Mint virus X: a novel potexvirus associated with symptoms in ‘Variegata’ mint. Archives of Virology. 2005;151(1):143 − 53. Tzanetakis IE, Postman JD, Gergerich RG, Martin RR. A virus between families: nucleotide sequence and evolution of S trawberry latent ringspot virus . Virus Research. 2006;121(2):199–204. Saeed ST, Kumar B, Shasany AK, Samad A. Molecular identification of Chilli leaf curl India virus along with betasatellite molecule causing leaf curl disease of menthol mint (Mentha arvensis var. Kosi) in India. Journal of General Plant Pathology. 2017;83(5):333-6. Haas BJ, Papanicolaou A, Yassour M, Grabherr M, Blood PD, Bowden J, et al. De novo transcript sequence reconstruction from RNA-seq using the Trinity platform for reference generation and analysis. Nature protocols. 2013;8(8):1494 − 512. Forgia M, Chiapello M, Daghino S, Pacifico D, Crucitti D, Oliva D, et al. Three new clades of putative viral RNA-dependent RNA polymerases with rare or unique catalytic triads discovered in libraries of ORFans from powdery mildews and the yeast of oenological interest Starmerella bacillaris. Virus Evolution. 2022;8(1). Buchfink B, Xie C, Huson DH. Fast and sensitive protein alignment using DIAMOND. Nature Methods. 2015;12(1):59–60. Katoh K, Rozewicki J, Yamada KD. MAFFT online service: multiple sequence alignment, interactive sequence choice and visualization. Briefings in Bioinformatics. 2019;20(4):1160-6. Minh BQ, Research School of Computer Science ANU, Canberra, ACT, Australia, Department of Ecology and Evolution RSoB, Australian National University, Canberra, ACT, Australia, Schmidt HA, Center for Integrative Bioinformatics Vienna MPL, University of Vienna and Medical University of Vienna, Vienna, Austria, Chernomor O, et al. IQ-TREE 2: New Models and Efficient Methods for Phylogenetic Inference in the Genomic Era. Molecular Biology and Evolution. 2024;37(5):1530-4. Kalyaanamoorthy S, Bui Quang M, Wong TKF, von Haeseler A, Jermiin LS. ModelFinder: fast model selection for accurate phylogenetic estimates. Nature Methods. 2017;14(6):587-+. Hoang DT, Chernomor O, Von Haeseler A, Minh BQ, Vinh LS. UFBoot2: improving the ultrafast bootstrap approximation. Molecular Biology and Evolution. 2017;35(2):518 − 22. Tables Table 1. Blastp first hit for each putative protein coded by MMV. Putative protein lenght Blastp hit Query Cover E value Per. Ident Accession replicase 1967aa Poplar mosaic virus 99% 0.0 52.31% WBG54312.1 TGB 1 231aa Poplar mosaic virus 100% 3,00E-56 45.26% Q02109.2 TGB 2 110aa Poplar mosaic virus 99% 5,00E-32 55.96% CAH55775.1 TGB 3 72aa Foveavirus mali 91% 4,00E-07 44.12% ASS35940.1 CP 280aa Poplar mosaic virus 97% 3,00E-99 52.38% WBG54313.1 C4 111aa Cowpea mild mottle virus 84% 6,00E-21 50.00% AFS33781.1 Supplementary Files CompleteSequenceMMV.fasta Submission2889379.txt Supp.Mat.docx Cite Share Download PDF Status: Published Journal Publication published 10 Jan, 2025 Read the published version in Archives of Virology → Version 1 posted Editorial decision: Major Revision 11 Nov, 2024 Reviewers agreed at journal 07 Nov, 2024 Reviewers invited by journal 07 Nov, 2024 Editor assigned by journal 07 Nov, 2024 First submitted to journal 06 Nov, 2024 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-5404341","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":375264004,"identity":"5d8ce092-8754-4381-b684-562d51f1e55d","order_by":0,"name":"Marco Forgia","email":"","orcid":"","institution":"IPSP CNR: Istituto per la Protezione Sostenibile delle Piante Consiglio Nazionale delle Ricerche","correspondingAuthor":false,"prefix":"","firstName":"Marco","middleName":"","lastName":"Forgia","suffix":""},{"id":375264005,"identity":"dda46949-f1a6-48f6-802f-5b01544d81d5","order_by":1,"name":"Marta Vallino","email":"","orcid":"","institution":"IPSP CNR: Istituto per la Protezione Sostenibile delle Piante Consiglio Nazionale delle Ricerche","correspondingAuthor":false,"prefix":"","firstName":"Marta","middleName":"","lastName":"Vallino","suffix":""},{"id":375264006,"identity":"0976c794-2820-4f64-849f-ac8cade5ffd2","order_by":2,"name":"Monica Marra","email":"","orcid":"","institution":"Universita degli Studi di Bari Aldo Moro Dipartimento di Scienze del Suolo della Pianta e degli Alimenti","correspondingAuthor":false,"prefix":"","firstName":"Monica","middleName":"","lastName":"Marra","suffix":""},{"id":375264007,"identity":"91ff3d0e-64fe-4874-b73f-f2a9056e9a92","order_by":3,"name":"Paolo Mussano","email":"","orcid":"","institution":"IPSP CNR: Istituto per la Protezione Sostenibile delle Piante Consiglio Nazionale delle Ricerche","correspondingAuthor":false,"prefix":"","firstName":"Paolo","middleName":"","lastName":"Mussano","suffix":""},{"id":375264008,"identity":"8d83751f-a106-48f4-a915-9f749eb273f5","order_by":4,"name":"Anna Paola Lanteri","email":"","orcid":"","institution":"CeRSAA Centro di Sperimentazione e Assistenza Agricola","correspondingAuthor":false,"prefix":"","firstName":"Anna","middleName":"Paola","lastName":"Lanteri","suffix":""},{"id":375264009,"identity":"83e6d4e7-4bd0-4d0d-9671-80da76dadfa0","order_by":5,"name":"Gian Paolo Accotto","email":"","orcid":"","institution":"IPSP CNR: Istituto per la Protezione Sostenibile delle Piante Consiglio Nazionale delle Ricerche","correspondingAuthor":false,"prefix":"","firstName":"Gian","middleName":"Paolo","lastName":"Accotto","suffix":""},{"id":375264010,"identity":"20dd50bf-3806-4e45-8261-6ec19472af73","order_by":6,"name":"Marina Ciuffo","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA/UlEQVRIiWNgGAWjYJCCAyCCjYHxwQeGCgm4KA8RWpgNZzCcQdKCVw8EALUwtiHxcWnRbT/78HBBDYM8n0QyY8PHeRZ58rMbGD8X7mGQscehxexMusHhGccYDNuAWhpnbpMoNrhzgFl6xjPcDjM7kMZwmIeNIYFNIv/4Y95tEokbJBLYmHkO4NFy/hlQyz+QlmTGZt45EonzZxDScgNoC28bTEuDRGLDDYJagLbw9kkYtvE8ZmyccQzkl4PN0jwHJHiA2nA4LI35M883G3n5dmCIfaipA4ZY88HPPAds7NkbcFgDAYgYTGCQYGxAESEIEkhRPApGwSgYBSMDAAD3oU9mAl8bUgAAAABJRU5ErkJggg==","orcid":"https://orcid.org/0000-0003-0422-408X","institution":"Consiglio Nazionale delle Ricerche","correspondingAuthor":true,"prefix":"","firstName":"Marina","middleName":"","lastName":"Ciuffo","suffix":""}],"badges":[],"createdAt":"2024-11-06 16:18:30","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-5404341/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-5404341/v1","draftVersion":[],"editorialEvents":[{"content":"https://doi.org/10.1007/s00705-025-06222-8","type":"published","date":"2025-01-10T15:57:58+00:00"}],"editorialNote":"","failedWorkflow":false,"files":[{"id":69862973,"identity":"a60a4c16-62bf-45e9-9d9b-8ddc547be28f","added_by":"auto","created_at":"2024-11-26 06:13:10","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":319401,"visible":true,"origin":"","legend":"\u003cp\u003ea) schematic representation of MMV genome. b) TEM picture of MMV particles. Scale bar: 100 nm. c) Maximum likelihood phylogenetic tree of the replicase protein from members of the family Betaflexiviridae. Members of the families Deltaflexiviridae and Gammaflexiviridae were used as outgroups.\u003c/p\u003e","description":"","filename":"OnlineFigure1Final.png","url":"https://assets-eu.researchsquare.com/files/rs-5404341/v1/9d1183ee048d5a930a75235c.png"},{"id":73694237,"identity":"8a572804-ffb5-4cf3-abd9-0762b79fc599","added_by":"auto","created_at":"2025-01-13 16:12:40","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":734796,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-5404341/v1/1b049475-7ecf-4619-9750-083cc31a9f83.pdf"},{"id":69863065,"identity":"2f0925c0-42ea-4a89-a233-64f95181a737","added_by":"auto","created_at":"2024-11-26 06:21:11","extension":"fasta","order_by":6,"title":"","display":"","copyAsset":false,"role":"supplement","size":8591,"visible":true,"origin":"","legend":"","description":"","filename":"CompleteSequenceMMV.fasta","url":"https://assets-eu.researchsquare.com/files/rs-5404341/v1/d558bef48c67447766e7136c.fasta"},{"id":69862974,"identity":"588821e3-4f52-43ca-becf-7cf9637d139f","added_by":"auto","created_at":"2024-11-26 06:13:10","extension":"txt","order_by":7,"title":"","display":"","copyAsset":false,"role":"supplement","size":17579,"visible":true,"origin":"","legend":"","description":"","filename":"Submission2889379.txt","url":"https://assets-eu.researchsquare.com/files/rs-5404341/v1/cf12b1d630881140bd074537.txt"},{"id":69862976,"identity":"86a7a959-1eba-4582-b8e6-006a1fcb66ad","added_by":"auto","created_at":"2024-11-26 06:13:11","extension":"docx","order_by":8,"title":"","display":"","copyAsset":false,"role":"supplement","size":285960,"visible":true,"origin":"","legend":"","description":"","filename":"Supp.Mat.docx","url":"https://assets-eu.researchsquare.com/files/rs-5404341/v1/8aea91ca70fe7d5ff2adce23.docx"}],"financialInterests":"","formattedTitle":"Characterization of Mint mosaic virus, a new member of the genus Carlavirus","fulltext":[{"header":"Fulltext","content":"\u003cp\u003eMint (genus \u003cem\u003eMentha\u003c/em\u003e, family \u003cem\u003eLamiaceae\u003c/em\u003e) is commonly cultivated throughout the Mediterranean region, including Italy. Its cultivation is related to the different applications of its aromatic compounds, both in food production and traditional medicine, as well as for their antimicrobial potential (1). Knowledge of the viruses associated with this important plant is limited to few case studies. In fact, to date, the only virus characterized in mint is a potexvirus called Mint virus X, which was detected in \u0026ldquo;Variegata\u0026rdquo; mint in 2006 (2). However, the stralarivirus Strawberry latent ringspot virus and the begomovirus Chilli leaf curl India virus have been detected on mint in the USA and in India, respectively (3, 4). In fall 2023, leaves from \u003cem\u003eMentha spicata\u003c/em\u003e displaying mosaic symptoms were collected in the Liguria region of Italy (Supp. Figure\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). To determine the cause of the observed symptoms, the plant sample was mechanically inoculated on test plants as a viral infection was suspected. After one week, systemic symptoms were detected on \u003cem\u003eNicotiana. benthamiana\u003c/em\u003e. Both the original mint sample and symptomatic \u003cem\u003eN. benthamiana\u003c/em\u003e leaves were used for transmission electron microscopy analysis (TEM). TEM observations were conducted by applying raw leaf extract for a few minutes onto carbon and formvar-coated grids and staining it with 0.5% uranyl acetate. Observations were made on a CM 10 electron microscope (Philips, Eindhoven, Netherlands). TEM analysis revealed filamentous and flexuous particles of around 600 nm in length and 13\u0026ndash;15 nm in diameter in both samples (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eb). To determine the genome sequence of the isolated virus, Illumina sequencing of total RNA was performed. Total RNA was extracted from systemically infected \u003cem\u003eN. benthamiana\u003c/em\u003e leaves grinded in a 2 ml O-ring tube with 0.3 mm glass beads using a FastPrep-24\u0026trade; bead beating grinder (MP Biomedicals\u0026trade;, Irvine, CA, USA). Grinded leaves were then processed for RNA extraction using Spectrum\u0026trade; Plant Total RNA Kit (Merk, Darmstadt, Germany). Ribosomal depletion, library construction and sequencing of the total RNA were performed by Macrogen Europe (Amsterdam, Netherlands) using the TruSeq Stranded Total RNA with Ribo-Zero Plant kit (Illumina) on a Novaseq 6000 platform. Sequencing results were cleaned with bbtools (sourceforge.net/projects/bbmap/) and assembled using Trinity (ver. 2.9.1) (5) as described in previous works (6). The obtained metatranscriptome was searched for viral-related contigs with a blast against the NCBI nr viral database (taxid: 2559587) using DIAMOND (7). Each contig giving a viral hit was blasted against the nr database to detect false positive, and the results were manually checked, retrieving a carlavirus-related contig with a poly(A) tail. The leaves of the plants mechanically inoculated with MMV were lyophilized and included in the Plant Virus Italy (PLAVIT) collection with the ID Me1, stored at -20 C\u0026deg;. To obtain the complete viral sequence, we performed 5\u0026rsquo;-rapid amplification of cDNA analysis (5\u0026rsquo;-RACE). RACE was performed using the RLM described in Forgia et al. 2022, the total RNA was ligated with a blocked DNA oligo (5\u0026rsquo;-Phosphate-GCATTCGACCCCGGGTT-AmC3-3\u0026rsquo;) using T4 RNA Ligase (Takara, Japan) and cDNA was then synthetized with a primer complementary to the adaptor (5\u0026rsquo;-AACCCGGGGTCGAATGC-3\u0026rsquo;). Obtained cDNA was used as template for RT-PCR using two specific primers (MMV 320 Rev: 5\u0026rsquo;-TCTTGCACACTGGGTGGGAA-3\u0026rsquo;, MMV 361 Rev: 5\u0026rsquo;-AAGCGGCGGAATGACTACATACAA-3\u0026rsquo;), the obtained amplicon was cloned in pGEM-T Easy (Promega, Madison, Wisconsin, USA) and sequenced through Sanger method by Biofab s.r.l. (Rome, Italy). From the 6 clones sequenced, we reconstructed the complete genome of the virus, tentatively named mint mosaic virus (MMV) which is 8558 nt long excluding the poly(A) and encode 6 putative proteins (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ea). The putative protein sequences were retrieved using ORFinder and the conserved domains were investigated with the NCBI conserved domain search tool (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://www.ncbi.nlm.nih.gov/Structure/cdd/wrpsb.cgi\u003c/span\u003e\u003cspan address=\"https://www.ncbi.nlm.nih.gov/Structure/cdd/wrpsb.cgi\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e) from 5\u0026rsquo; to 3\u0026rsquo;. The first ORF encodes a putative protein of 1,967 aa that shows a Betaflexiviridae_RdRp domain, a Viral_helicase1 domain, a Vmethyltransf domain, a Peptidase_C23 domain, an AlkB domain and an OTU_RDRP-like domain. The second, third and fourt ORFs encode proteins of 231, 110 and 280 aa long and exhibit the typical conserved domains of carlavirus triple gene block (Viral_helicase1 for TGB1, Plant_vir_prot for TGB2 and TGBp3 domain for TGB3). The fifth ORF encodes a 280 aa long protein showing a conserved Flexi_CP domain, while the last ORF codes for a 111 aa protein with a conserved Carla_C4 domain. Each putative protein was blasted against the NCBI nr database to detect homology against the closest known viruses. Results in Table\u0026nbsp;1 show that the replicase protein, along with the proteins from triple gene block 1 and 2, and the coat protein, have Poplar mosaic virus (PopMV) as the closest hit, with a homology percentage ranging between 45 and 56%. The protein from triple gene block 3 and the C4 protein show similar homology percentages against Apple stem pitting foveavirus (ASPV) and Cowpea mild mottle virus (CMMV), respectively. To perform phylogenetic analysis, the replicase of all the viruses accepted by the ICTV in the \u003cem\u003eBetaflexiviridae\u003c/em\u003e family (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://ictv.global/msl\u003c/span\u003e\u003cspan address=\"https://ictv.global/msl\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e) were retrieved and manually selected to remove partial and redundant sequences. The selected proteins (listed in Supp. Table\u0026nbsp;1) were aligned with MAFFT (8) including the ORF1 replicase sequence of MMV and the multiple alignment obtained was submitted to IQ-TREE 2 (9) using the automatic model selection (10) and ultrafast bootstrap with 1000 replicates (11) to build a maximum likelihood phylogenetic tree of the whole viral family. Results in Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ec collocate the new virus in the genus \u003cem\u003eCarlavirus\u003c/em\u003e, forming a branch with PopMV as already suggested by the blast analysis (Table\u0026nbsp;1). Taken together these results and considering the species demarcation criteria for the genus \u003cem\u003eCarlavirus\u003c/em\u003e (Distinct species have less than about 72% nt identity or 80% aa identity between their CP or polymerase genes, \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://ictv.global/report_9th/RNApos/Betaflexiviridae\u003c/span\u003e\u003cspan address=\"https://ictv.global/report_9th/RNApos/Betaflexiviridae\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e) MMV should be accomodated in a new species in the genus, for which we tentatively propose the name \u003cem\u003eCarlavirus menthae.\u003c/em\u003e In summer 2024, a new mint sample showing mosaic symptoms was collected in the Calabria region of Italy. TEM analysis suggested a possible infection by MMV (not shown) thus, the virus was isolated in greenhouse as previously described and qRT-PCR primers were designed to validate an infection from MMV (MMV_RT_For: 5\u0026rsquo;-CATTCCGCCGCTTTTGGATA-3\u0026rsquo;, MMV_RT_Rev: 5\u0026rsquo;-AAATGAGGGAGAGGTTGCGA-3\u0026rsquo;). qRT-PCR was performed in 10 \u0026micro;l of total volume using the iTaq Universal SYBR Green Supermix on a CFX apparatus (Biorad, Hercules, CA, USA). Results of the qRT-PCRconfirmed the presence of MMV also in the sample from 2024, demonstrating that the virus is present in different Italian regions and could be detected consecutively for two years. The MMV viral accession from 2024 was included in the PLAVIT collection with the ID Me2, stored as dried leaf at -20 C\u0026deg;.\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003cp\u003eIn conclusion, this work characterized a new carlavirus infecting mint that was tentatively named mint mosaic virus. The virus was detected in two independent sampling in Italy and the two isolates identified are stored and maintained in the plant virus collection PLAVIT at the IPSP-CNR institute. Results suggest that that surveillance on symptomatic plants is still important to detect and characterize new viral pathogens or potential cross species infection by viral agents.\u003c/p\u003e"},{"header":"Declarations","content":"\u003ch2\u003eAcknowledgements\u003c/h2\u003e \u003cp\u003eThis research was funded by the European Virus Archive GLOBAL (EVA-GLOBAL) project that has received funding from the European Union\u0026rsquo;s Horizon 2020 research and innovation program under grant agreement No. 871029, and by the SUS-MIRRI.IT project, granted by the European Commission\u0026mdash;NextGenerationEU, code n. IR0000005.\u003c/p\u003e\u003ch2\u003eData availability\u003c/h2\u003e \u003cp\u003eThe complete viral genome of MMV Me1 accession is deposited in the NCBI database with the accession number PQ562895.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n \u003cli\u003e\u003cspan\u003eGupta S, Kumar A, Gupta AK, Jnanesha AC, Talha M, Srivastava A. Industrial mint crop revolution, new opportunities, and novel cultivation ambitions: A review. Ecological Genetics and Genomics. 2023;27.\u003c/span\u003e\u003c/li\u003e\n \u003cli\u003e\u003cspan\u003eTzanetakis IE, Postman JD, Martin RR. Mint virus X: a novel potexvirus associated with symptoms in \u0026lsquo;Variegata\u0026rsquo; mint. Archives of Virology. 2005;151(1):143\u0026thinsp;\u0026minus;\u0026thinsp;53.\u003c/span\u003e\u003c/li\u003e\n \u003cli\u003e\u003cspan\u003eTzanetakis IE, Postman JD, Gergerich RG, Martin RR. A virus between families: nucleotide sequence and evolution of S\u003cem\u003etrawberry latent ringspot virus\u003c/em\u003e. Virus Research. 2006;121(2):199\u0026ndash;204.\u003c/span\u003e\u003c/li\u003e\n \u003cli\u003e\u003cspan\u003eSaeed ST, Kumar B, Shasany AK, Samad A. Molecular identification of Chilli leaf curl India virus along with betasatellite molecule causing leaf curl disease of menthol mint (Mentha arvensis var. Kosi) in India. Journal of General Plant Pathology. 2017;83(5):333-6.\u003c/span\u003e\u003c/li\u003e\n \u003cli\u003e\u003cspan\u003eHaas BJ, Papanicolaou A, Yassour M, Grabherr M, Blood PD, Bowden J, et al. De novo transcript sequence reconstruction from RNA-seq using the Trinity platform for reference generation and analysis. Nature protocols. 2013;8(8):1494\u0026thinsp;\u0026minus;\u0026thinsp;512.\u003c/span\u003e\u003c/li\u003e\n \u003cli\u003e\u003cspan\u003eForgia M, Chiapello M, Daghino S, Pacifico D, Crucitti D, Oliva D, et al. Three new clades of putative viral RNA-dependent RNA polymerases with rare or unique catalytic triads discovered in libraries of ORFans from powdery mildews and the yeast of oenological interest Starmerella bacillaris. Virus Evolution. 2022;8(1).\u003c/span\u003e\u003c/li\u003e\n \u003cli\u003e\u003cspan\u003eBuchfink B, Xie C, Huson DH. Fast and sensitive protein alignment using DIAMOND. Nature Methods. 2015;12(1):59\u0026ndash;60.\u003c/span\u003e\u003c/li\u003e\n \u003cli\u003e\u003cspan\u003eKatoh K, Rozewicki J, Yamada KD. MAFFT online service: multiple sequence alignment, interactive sequence choice and visualization. Briefings in Bioinformatics. 2019;20(4):1160-6.\u003c/span\u003e\u003c/li\u003e\n \u003cli\u003e\u003cspan\u003eMinh BQ, Research School of Computer Science ANU, Canberra, ACT, Australia, Department of Ecology and Evolution RSoB, Australian National University, Canberra, ACT, Australia, Schmidt HA, Center for Integrative Bioinformatics Vienna MPL, University of Vienna and Medical University of Vienna, Vienna, Austria, Chernomor O, et al. IQ-TREE 2: New Models and Efficient Methods for Phylogenetic Inference in the Genomic Era. Molecular Biology and Evolution. 2024;37(5):1530-4.\u003c/span\u003e\u003c/li\u003e\n \u003cli\u003e\u003cspan\u003eKalyaanamoorthy S, Bui Quang M, Wong TKF, von Haeseler A, Jermiin LS. ModelFinder: fast model selection for accurate phylogenetic estimates. Nature Methods. 2017;14(6):587-+.\u003c/span\u003e\u003c/li\u003e\n \u003cli\u003e\u003cspan\u003eHoang DT, Chernomor O, Von Haeseler A, Minh BQ, Vinh LS. UFBoot2: improving the ultrafast bootstrap approximation. Molecular Biology and Evolution. 2017;35(2):518\u0026thinsp;\u0026minus;\u0026thinsp;22.\u003c/span\u003e\u003c/li\u003e\n\u003c/ol\u003e"},{"header":"Tables","content":"\u003cp\u003e\u003cstrong\u003eTable 1.\u0026nbsp;\u003c/strong\u003eBlastp first hit for each putative protein coded by MMV.\u003c/p\u003e\n\u003ctable border=\"0\" cellspacing=\"0\" cellpadding=\"0\" width=\"763\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 18.1937%;\"\u003e\n \u003cp\u003e\u003cstrong\u003ePutative protein\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 9.94764%;\"\u003e\n \u003cp\u003e\u003cstrong\u003elenght\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 21.0733%;\"\u003e\n \u003cp\u003e\u003cstrong\u003eBlastp hit\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 12.4346%;\"\u003e\n \u003cp\u003e\u003cstrong\u003eQuery Cover\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11.1257%;\"\u003e\n \u003cp\u003e\u003cstrong\u003eE value\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 12.4346%;\"\u003e\n \u003cp\u003e\u003cstrong\u003ePer. Ident\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 14.7906%;\"\u003e\n \u003cp\u003e\u003cstrong\u003eAccession\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 18.1937%;\"\u003e\n \u003cp\u003ereplicase\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 9.94764%;\"\u003e\n \u003cp\u003e1967aa\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 21.0733%;\"\u003e\n \u003cp\u003ePoplar mosaic virus\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 12.4346%;\"\u003e\n \u003cp\u003e99%\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11.1257%;\"\u003e\n \u003cp\u003e0.0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 12.4346%;\"\u003e\n \u003cp\u003e52.31%\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 14.7906%;\"\u003e\n \u003cp\u003eWBG54312.1\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 18.1937%;\"\u003e\n \u003cp\u003eTGB 1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 9.94764%;\"\u003e\n \u003cp\u003e231aa\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 21.0733%;\"\u003e\n \u003cp\u003ePoplar mosaic virus\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 12.4346%;\"\u003e\n \u003cp\u003e100%\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11.1257%;\"\u003e\n \u003cp\u003e3,00E-56\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 12.4346%;\"\u003e\n \u003cp\u003e45.26%\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 14.7906%;\"\u003e\n \u003cp\u003eQ02109.2\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 18.1937%;\"\u003e\n \u003cp\u003eTGB 2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 9.94764%;\"\u003e\n \u003cp\u003e110aa\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 21.0733%;\"\u003e\n \u003cp\u003ePoplar mosaic virus\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 12.4346%;\"\u003e\n \u003cp\u003e99%\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11.1257%;\"\u003e\n \u003cp\u003e5,00E-32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 12.4346%;\"\u003e\n \u003cp\u003e55.96%\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 14.7906%;\"\u003e\n \u003cp\u003eCAH55775.1\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 18.1937%;\"\u003e\n \u003cp\u003eTGB 3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 9.94764%;\"\u003e\n \u003cp\u003e72aa\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 21.0733%;\"\u003e\n \u003cp\u003eFoveavirus mali\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 12.4346%;\"\u003e\n \u003cp\u003e91%\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11.1257%;\"\u003e\n \u003cp\u003e4,00E-07\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 12.4346%;\"\u003e\n \u003cp\u003e44.12%\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 14.7906%;\"\u003e\n \u003cp\u003eASS35940.1\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 18.1937%;\"\u003e\n \u003cp\u003eCP\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 9.94764%;\"\u003e\n \u003cp\u003e280aa\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 21.0733%;\"\u003e\n \u003cp\u003ePoplar mosaic virus\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 12.4346%;\"\u003e\n \u003cp\u003e97%\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11.1257%;\"\u003e\n \u003cp\u003e3,00E-99\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 12.4346%;\"\u003e\n \u003cp\u003e52.38%\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 14.7906%;\"\u003e\n \u003cp\u003eWBG54313.1\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 18.1937%;\"\u003e\n \u003cp\u003eC4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 9.94764%;\"\u003e\n \u003cp\u003e111aa\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 21.0733%;\"\u003e\n \u003cp\u003eCowpea mild mottle virus\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 12.4346%;\"\u003e\n \u003cp\u003e84%\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11.1257%;\"\u003e\n \u003cp\u003e6,00E-21\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 12.4346%;\"\u003e\n \u003cp\u003e50.00%\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 14.7906%;\"\u003e\n \u003cp\u003eAFS33781.1\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"archives-of-virology","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"arvi","sideBox":"Learn more about [Archives of Virology](https://www.springer.com/journal/705)","snPcode":"705","submissionUrl":"https://submission.nature.com/new-submission/705/3","title":"Archives of Virology","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false},"keywords":"","lastPublishedDoi":"10.21203/rs.3.rs-5404341/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-5404341/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"In this work the complete genome of a new carlavirus causing mosaic on mint plants in Italy is reported and tentatively named Mint mosaic virus (MMV). Flexuous particles of around 600 nm were observed through TEM analysis and NGS sequencing was performed to determine MMV genome. MMV genome is 8558 nt long, excluding the poly(A) tail and shows the typical organization of a carlavirus. The putative proteins coded by MMV have homology percentages ranging between 44% and 56% against the closest viruses in the NCBI database, suggesting that MMVcan be considered a new species in the genus Carlavirus. MMV was detected in independent mint samples from different Italian regions collected in years 2023 and 2024, suggesting a broad diffusion, at least in the Italian area. The two MMV isolates characterized in 2023 (Me1) and in 2024 (Me2) were included in the plant virus collection PLAVIT.","manuscriptTitle":"Characterization of Mint mosaic virus, a new member of the genus Carlavirus","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2024-11-26 06:13:06","doi":"10.21203/rs.3.rs-5404341/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Major Revision","date":"2024-11-11T07:12:29+00:00","index":"","fulltext":""},{"type":"reviewerAgreed","content":"","date":"2024-11-07T11:03:56+00:00","index":0,"fulltext":""},{"type":"reviewersInvited","content":"","date":"2024-11-07T10:42:39+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2024-11-07T05:06:33+00:00","index":"","fulltext":""},{"type":"submitted","content":"Archives of Virology","date":"2024-11-06T11:17:31+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"archives-of-virology","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"arvi","sideBox":"Learn more about [Archives of Virology](https://www.springer.com/journal/705)","snPcode":"705","submissionUrl":"https://submission.nature.com/new-submission/705/3","title":"Archives of Virology","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false}}],"origin":"","ownerIdentity":"df17147e-f927-4574-bb7e-1ffa7dae4c39","owner":[],"postedDate":"November 26th, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[],"tags":[],"updatedAt":"2025-01-13T16:07:56+00:00","versionOfRecord":{"articleIdentity":"rs-5404341","link":"https://doi.org/10.1007/s00705-025-06222-8","journal":{"identity":"archives-of-virology","isVorOnly":false,"title":"Archives of Virology"},"publishedOn":"2025-01-10 15:57:58","publishedOnDateReadable":"January 10th, 2025"},"versionCreatedAt":"2024-11-26 06:13:06","video":"","vorDoi":"10.1007/s00705-025-06222-8","vorDoiUrl":"https://doi.org/10.1007/s00705-025-06222-8","workflowStages":[]},"version":"v1","identity":"rs-5404341","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-5404341","identity":"rs-5404341","version":["v1"]},"buildId":"qtupq5eGEP_6zYnWcrvyt","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}