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ARS Home » Northeast Area » Beltsville, Maryland (BARC) » Beltsville Agricultural Research Center » Molecular Plant Pathology Laboratory » Research » Publications at this Location » Publication #409937
Research Project: Emerging Stress Challenges and Functional Genomics of Stress Responses in Alfalfa

Location: Molecular Plant Pathology Laboratory

Title: Complete genome sequence of alfalfa-associated picorna-like virus 2

Author
item Nemchinov, Lev
item Grinstead, Sam

Submitted to: Microbiology Resource Announcements
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 2/7/2024
Publication Date: 3/1/2024
Citation: Nemchinov, L.G., Grinstead, S.C. 2024. Complete genome sequence of alfalfa-associated picorna-like virus 2. Microbiology Resource Announcements. 13. Article e01052-23. https://doi.org/10.1128/mra.01052-23.
DOI: https://doi.org/10.1128/mra.01052-23

Interpretive Summary: Viruses are an integral part of a diverse community of pathogenic microbes associated with alfalfa. In this research, aimed at surveying alfalfa varieties grown in the U.S. to identify, characterize, and prevent the spread of novel and emerging viruses in the country, we report a discovery and characterization of a novel viral pathogen found in field alfalfa samples. It is expected that this research will be of interest to plant pathologists, extension specialists, alfalfa growers, and people working in the field of alfalfa improvement.

Technical Abstract: During our recent survey of alfalfa field samples we detected genomic fragments of a novel picorna-like virus provisionally named alfalfa associated picorna-like virus 2 (AAPLV 2). Here we report a complete genomic sequence of the AAPLV2 and confirm its phylogenetic relationship to the members of the family Iflaviridae. The AaPLV2 genome consists of 9235 nucleotides (nt), excluding poly (A) tail. This corresponds to the length of RNA genomes of other iflaviruses (9–11 kilobases). According to 5’RACE, the 5' non-coding region (5’NCR) of the virus contains 479 nt; 3’RACE indicated that the 3'NCR of the virus contains 83 nt without the poly (A) tail. The 5’ NCR of APLV2 appears to have multiple stem-loop structures predicted by the RNAfold tool. These structures can potentially serve as IRES elements to promote translation initiation. Reverse transcription-polymerase chain reaction performed with two sets of primers designed based on the predicted sequence of the viral genome led to amplification of the correct-size amplicons, verified by Sanger sequencing. AaPLV2 contains a single large open reading frame encoding a polyprotein of 2890 amino acids (aa). PSI-BLAST search using the viral polyprotein as a query resulted in a number of high-quality hits to different iflaviruses, although percent their identity did not exceed 38%. The InterPro-predicted viral domains correspond to the genome organization of Iflaviridae family with structural capsid proteins located at the N-terminal end of the polyprotein and non-structural proteins (Hel, Pro, and RdRp) at the C-terminal region. Location of the InterPro-predicted domain of the first structural protein 380 aa away from the putative start codon indicated a presence of a leader protein in this region of the viral genome. Phylogenetic analysis carried out by RAxML-NG using Clustal W alignment and maximum likelihood algorithm with 1000 replicates demonstrated that AaPLV 2 clustered with established and putative members of the genus Iflavirus rather than with several plant-infecting monopartite picorna-like viruses of the family Secoviridae. We therefore propose that AaPLV 2 is an iflavirus associated with alfalfa likely due to the localized contamination from insects feeding on the plant.