Beatrice Hill Virus Represents a Novel Species in the Genus Tibrovirus (Mononegavirales: Rhabdoviridae)

ABSTRACT The rhabdoviral genus Tibrovirus currently has three official members assigned to two species: Bivens Arm virus and Tibrogargan virus (species Tibrogargan tibrovirus) and Coastal Plains virus (species Coastal Plains tibrovirus). Here, we report the complete genome sequence of a new putative member of this genus, Beatrice Hill virus. Although relatively closely related to the three classified viruses, Beatrice Hill virus represents a novel Tibrovirus species.

Beatrice Hill virus (BHV) was first reported in 1984 as a novel virus of biting midges (Culicoides peregrinus) that had been collected at Beatrice Hill, Northern Territory, Australia (11). In 2016, Huang et al. reported a 5,734 nt-long contig of the BHV genome, which indicated that this virus most likely falls into the Tibrovirus clade (12). To determine the taxonomic position of BHV, we obtained a historical sample of brain tissue from a laboratory mouse that had been infected intracranially with BHVcontaining material. Viral RNA from this sample was processed and sequenced to obtain the complete genome using a sequence-independent single-primer amplification (SISPA) protocol that included rapid amplification of cDNA ends (RACE) (13). Resulting libraries were sequenced on an Illumina MiSeq desktop sequencer. Illumina and SISPA-RACE adapter sequences were trimmed from the sequencing reads using Cutadapt version 1.2.1 (14), quality filtering was conducted with Prinseq-lite (15), and reads were assembled into contigs using Ray Meta with a k-mer length of 25 (16). Resultant contigs were aligned to the National Center for Biotechnology Information (NCBI) sequence database using BLAST. Reads were aligned back to the assembled BHV sequence using Bowtie2 (17), and custom scripts were used to generate a final consensus sequence.
Using PASC (18), the determined complete BHV genome sequence was found to be 72.79% identical to that of TIBV (GenBank accession no. GQ294472.1) and 72.48% identical to that of BAV (KP688373.1). The genome organization matches that of all tibroviruses, therefore confirming Huang et al.'s hypothesis that BHV is a tibrovirus. Our data furthermore suggest that this virus should be assigned to a novel species ("Beatrice Hill tibrovirus") based on its divergence from TIBV and BAV.
Accession number(s). The GenBank accession number of Beatrice Hill virus isolate CSIRO 25 is KY073493.

ACKNOWLEDGMENTS
We thank Toby S. George (Australia) for sharing his knowledge and historical reports on tibroviruses and Laura Bollinger (IRF-Frederick, Frederick, MD, USA) for editing the manuscript.
The content of this publication does not necessarily reflect the views or policies of the U.S. Department of the Army, the U.S. Department of Defense, or the U.S. Department of Health and Human Services or of the institutions and companies affiliated with the authors. This work was funded in part through Battelle Memorial Institute's prime contract with the U.S. National Institute of Allergy and Infectious Diseases (NIAID) under contract no. HHSN272200700016I. Subcontractors to Battelle Memorial Institute who performed this work are Y.C. and J.H.K., employees of Tunnell Government Services, Inc.