Choclo virus (CHOV) recovered from deep metatranscriptomics of archived museum tissues

Background Hantaviruses are negative-stranded RNA viruses that can sometimes cause severe disease in humans; however, they are maintained in mammalian populations without causing harm. In Panama, sigmodontine rodents serve as hosts to transmissible hantaviruses. Due to natural and anthropogenic forces, these rodent populations are having increased contact with humans. Methods We extracted RNA and performed Illumina deep metatranscriptomic sequencing on Orthohantavirus seropositive museum tissues from rodents. We acquired sequence reads mapping to Choclo virus (CHOV, Orthohantavirus chocloense ) from the heart and kidney tissue of a two decade old sample from a Costa Rican pygmy rice rat ( Oligoryzomys costaricensis ) collected in Panama. Reads mapped to the CHOV reference were assembled and then validated by visualization of the mapped reads against the assembly. Results We recovered a 91% complete consensus sequence from a reference-guided assembly to CHOV with an average of 16X coverage. The S and M segments used in our phylogenetic analyses were nearly complete (98% and 99%, respectively). There were 1,199 ambiguous base calls of which 93% were present in the L segment. Our assembled genome varied 1.3% from the CHOV reference sequence resulting in 11 nonsynonymous mutations. Further analysis of all publicly available partial S segment sequences support a clear relationship between CHOV clinical cases and O. costaricensis acquired strains. Conclusions Viruses occurring at extremely low abundances can be recovered from deep metatranscriptomics of archival tissues housed in museums or biorepositories. Our efforts resulted in the second CHOV genome publicly available. This genomic data is important for future surveillance and diagnostic tools as well as understanding the evolution and pathogenicity of CHOV. Author Summary Hantavirus cardiopulmonary syndrome (HCPS) in Panama, caused by Choclo virus (CHOV, Orthohantavirus chocloense ), is intimately linked to the primary mammalian reservoir host, the Costa Rican pygmy rice rat ( Oligoryzomys costaricensis ). Although the prevalence of hantavirus disease is relatively low in Panama, over a quarter of the country has the agroecological conditions that favor this rodent. In addition, serologic evidence suggests infections are under-reported. Sequence data of the pathogen and host collected across temporal and spatial scales is necessary for diagnostics, surveillance, and forecasting; however, only one complete genome is available in NCBI GenBank. By leveraging deep metatranscriptomics of archived frozen mammal tissues, we generated a low-coverage genome using a reference-guided assembly approach. Sequence data can be used to develop pan-hantavirus diagnostic tools to facilitate acquisition of more detailed genetic data from archival samples to increase our understanding of the evolutionary and population dynamics of rare and neglected hantaviruses. Generating additional genomic sequence data will also be essential for developing a rigorous taxonomic framework to improve the understanding of hantavirus diversity and distribution. ### Competing Interest Statement The authors have declared no competing interest.