Acta virologica 66: 149 – 156, 2022

doi: 10.4149/av_2022_204

Artemisia capillaris nucleorhabdovirus 1,
a novel member of the genus Alphanucleorhabdovirus, identified
in the Artemisia capillaris transcriptome
Dongjin Choi1, Chaerim Shin1, Ken Shirasu2,3, Yasunori Ichihashi4, Yoonsoo Hahn1*
Department of Life Science, Chung-Ang University, Seoul 06974, South Korea; 2RIKEN Center for Sustainable Resource Science,
Yokohama, Kanagawa 230-0045, Japan; 3Graduate School of Science, The University of Tokyo, Bunkyo, Tokyo 113-0033, Japan;
4
RIKEN BioResource Research Center, 3-1-1 Koyadai, Tsukuba, Ibaraki, 305-0074, Japan

1

Received February 17, 2022; accepted April 5, 2022
Summary. – A novel, negative-sense, single-stranded RNA virus, Artemisia capillaris nucleorhabdovirus
1 (AcNRV1), was identified in the transcriptome data of Artemisia capillaris (commonly known as capillary
wormwood) root tissue. The AcNRV1 genome contains six open reading frames encoding a nucleocapsid(N), phosphoprotein, movement protein P3, matrix protein, glycoprotein, and polymerase (L). Sequencecomparison and phylogenetic analysis using L and N protein sequences revealed that AcNRV1 is a novel
member of the genus Alphanucleorhabdovirus, one of the six plant-infecting rhabdovirus genera of thefamily Rhabdoviridae. Wheat yellow striate virus and rice yellow stunt virus were identified as the closest known rhabdoviruses of AcNRV1. The conserved regulatory sequences involved in transcription
termination/polyadenylation (TTP) and transcription initiation (TI) of individual genes were identified
in the AcNRV1 genome with the consensus sequence 3'-(A/U)UUAUUUUU–GGG–UUG-5' (in the negativesense genome), whereby dashes separate the TTP, untranscribed intergenic spacer, and TI elements. The
AcNRV1 genome sequence will contribute to further understanding the genome structural evolution ofplant rhabdoviruses.
Keywords: Artemisia capillaris nucleorhabdovirus 1; plant virus; Alphanucleorhabdovirus; Rhabdoviridae

Introduction
Rhabdoviruses (the family Rhabdoviridae) infectanimals and plants and have negative-sense, singlestranded RNA genomes (Dietzgen et al., 2017; Walker etal., 2018; Bejerman et al., 2021). The family Rhabdoviridae
comprises three subfamilies, 40 genera, and more thanCorresponding author. E-mail: hahny@cau.ac.kr; phone: +822-820-5812.
Abbreviations: AcNRV1 = Artemisia capillaris nucleorhabdovirus 1; ORF = open reading frame; RdRp = RNA-dependent RNA
polymerase; RYSV = rice yellow stunt virus; SRA = Sequence
Read Archive; TI = transcription initiation; TTP = transcription
termination/polyadenylation; UIS = untranscribed intergenic
spacer; WYSV = wheat yellow striate virus*

200 species currently approved by the International Committee on Taxonomy of Viruses (https://talk.ictvonline.
org, last accessed on January 20, 2022). Plant-infecting
rhabdoviruses of the subfamily Betarhabdovirinae areclassified into six genera: Alphanucleorhabdovirus, Betanucleorhabdovirus, Cytorhabdovirus, Dichorhavirus, Gammanucleorhabdovirus, and Varicosavirus. They have either
monopartite (Alphanucleorhabdovirus, Betanucleorhabdovirus, Cytorhabdovirus, and Gammanucleorhabdovirus)
or bipartite (Dichorhavirus and Varicosavirus) genomes(Dietzgen et al., 2017). Insects, including aphids, leafhoppers, and planthoppers, serve as transmission vectors forplant rhabdoviruses (Ammar et al., 2009; Liu et al., 2018;
Whitfield et al., 2018; Bhat et al., 2020).
Plant rhabdoviruses with a monopartite genome havesix shared open reading frames (ORFs) encoding a nucle-

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ocapsid (N), phosphoprotein (P), movement protein 3 (P3),matrix protein (M), surface glycoprotein (G), and large
multi-functional protein (L), in the order of 3'-N-P-P3-MG-L-5' (Walker et al., 2018). The L protein, which contains
an RNA-directed RNA polymerase (RdRp) domain, isinvolved in the genome replication and transcription ofindividual ORFs (Jackson et al., 2005; Walker et al., 2018).
The N protein encloses the viral RNA genomic moleculein the protein shell. The P protein acts as a cofactor of the
L protein during transcription and replication and helps
N proteins during encapsidation of genomic molecules(Fang et al., 2019). The M protein is required for viralmaturation and budding (Sun et al., 2018). The G proteinfunctions in receptor binding and membrane fusion (Coll,
1995; Mann et al., 2016). The P3 protein enables viruses tomove from one cell to another (Zhou et al., 2019). Some
plant rhabdoviruses may have additional ORFs apart fromthe abovementioned six (Walker et al., 2011). For example,
wheat yellow striate virus (WYSV) and rice yellow stunt
virus (RYSV) have a seventh ORF, P6, which encodesa small protein between the G and L ORFs (Huang et al.,
2003; Liu et al., 2018).
Rhabdovirus genes are transcribed into separate
mRNAs from the negative-sense RNA genome througha transcriptase complex composed of the L and P proteins(Jackson et al., 2005; Walker et al., 2011). The gene junction
regions of rhabdoviruses contain conserved regulatory
sequences called transcription termination/polyadenylation (TTP) and transcription initiation (TI) elements thatmediate polyadenylation of the preceding gene transcript
and transcriptional initiation of the next gene, respectively (Jackson et al., 2005; Goh et al., 2020; Bejerman et al.,2021; Shin et al., 2021). One or several nucleotides locatedbetween the TTP and TI elements are not transcribed;
this region is called the untranscribed intergenic spacer(UIS). For example, the consensus sequences of the WYSVand RYSV conserved regulatory sequences are presented
as 3'-UAAAUUUUU–GGGG–UUG-5' and 3'-AUUAUUUUU–GGG–UUG-5', respectively, whereby dashes (–) separate
the TTP, UIS, and TI elements (Huang et al., 2003; Liu etal., 2018; Bejerman et al., 2021).
RNA-sequencing (RNA-seq) data are frequently obtained from plant tissues to investigate gene expression
patterns during interaction with other organisms andplant development and differentiation (Ichihashi et al.,2018; Yoshida et al., 2019). When plant tissues are latently
infected with RNA viruses, full-length virus genome
sequences can be obtained by assembling transcriptome
reads and identifying virus genome contigs (Bejerman etal., 2020; Choi et al., 2021; Goh et al., 2021; Park et al., 2021;
Park and Hahn, 2021). Many novel rhabdovirus genome
sequences, including those of Trichosanthes associatedrhabdovirus 1, Agave tequilana virus 1, and Zostera as-sociated varicosavirus 1, have previously been identified
in plant transcriptome data (Goh et al., 2020; Bejerman etal., 2021; Shin et al., 2021). In the present study, the genome
sequence of a novel rhabdovirus belonging to the genus
Alphanucleorhabdovirus (the family Rhabdoviridae) was
identified in Artemisia capillaris (commonly known ascapillary wormwood) root transcriptome data (Ichihashi
et al., 2018).

Materials and Methods
Transcriptome data analysis. RNA-seq data were previously
obtained from the root parasitic plant Thesium chinense andits host plants (A. capillaris, Eragrostis curvula, and Lespedeza
juncea) to understand transcriptional regulation during haustorium formation (Ichihashi et al., 2018). Sequence data aredeposited in the Sequence Read Archive (SRA) of the National
Center for Biotechnology Information (NCBI) under SRA ProjectAcc. No. SRP114897. RNA-seq data (14 sequencing runs from T.
chinense and six from its hosts) were filtered to collect highquality reads using the sickle program (version 1.33; https://github.com/najoshi/sickle), with the parameter “-q 30 -l 55.”
High-quality reads from the 20 sequencing runs were separately
assembled into contigs using the SPAdes Genome Assembler(version 3.15.3; http://cab.spbu.ru/software/spades), with theparameter “--rnaviral” (Bushmanova et al., 2019).
The assembled transcriptome contigs were compared withknown viral RdRp domain sequences. A total of 2620 RdRp
domain sequences were obtained from the Pfam database(release 34.0; https://pfam.xfam.org). The Pfam Acc. Nos. are
PF00602, PF00603, PF00604, PF00680, PF00946, PF00972,
PF00978, PF00998, PF02123, PF03035, PF03431, PF04196,
PF04197, PF05788, PF05919, PF06317, PF06478, PF07925, PF08467,
PF08716, PF08717, PF12426, and PF17501. For sequence similarity searches, the DIAMOND program (version 2.0.4.142; https://github.com/bbuchfink/diamond) in the blastx mode was used
(Buchfink et al., 2015).
Viral genome annotation. RNA-seq read depth of the virus
genome was examined by mapping high-quality reads to theviral contig sequence using the bwa-mem2 program (version
2.0pre2; https://github.com/bwa-mem2/bwa-mem2). Putative
ORFs were predicted using the getorf program of the EMBOSS
package (version 6.6.0.0; http://emboss.open-bio.org), with theparameter “-find 1.” Putative conserved domains in the viralprotein sequences were identified using the InterPro web
server (release 87.0; https://www.ebi.ac.uk/interpro). Signal
peptides and transmembrane domains were predicted using
SignalP (version 6.0; https://services.healthtech.dtu.dk/service.
php?SignalP-6.0) and TMHMM (version 2.0; https://services.
healthtech.dtu.dk/service.php?TMHMM-2.0), respectively.
Conserved regulatory sequences in gene junction regions
were predicted using the MEME webtool (version 5.4.1; https://

CHOI, D. et al.: NOVEL NUCLEORHABDOVIRUS IDENTIFIED IN ARTEMISIA CAPILLARISmeme-suite.org/meme/tools/meme). The WebLogo application(version 3; http://weblogo.threeplusone.com) was used to createa sequence logo representation of the conserved sequences.
Phylogenetic analysis. Known viral protein sequences wereobtained using NCBI's BLAST (https://blast.ncbi.nlm.nih.gov/
Blast.cgi). The needle program of the EMBOSS package wasused to calculate pairwise identities of orthologous viral proteins. The MAFFT program (version 7.475; https://mafft.cbrc.
jp/alignment/software) with the parameter “--auto” was used
to generate multiple sequence alignments. Gap-rich segmentsof multiple alignments were removed using trimAl (version 1.4.
rev22; http://trimal.cgenomics.org) with the parameter “-automated1” (Capella-Gutierrez et al., 2009). Maximum-likelihood
phylogenetic trees were generated using the IQ-TREE program(version 2.1.3; http://www.iqtree.org) using the parameter “-B1000” to calculate bootstrap support values from 1000 replicates(Minh et al., 2020).

Results and Discussion
High-quality RNA-seq reads obtained from the parasitic plant T. chinense and its host plants, A. capillaris,
E. curvula, and L. juncea, were assembled to generatetranscriptome contigs (Ichihashi et al., 2018). Sequencecomparisons with known viral RdRp sequences revealed
that several plant transcriptome contigs may containa viral RdRp domain. One contig that was assembled fromtranscriptome data (SRA Acc. No. SRR5917890) derived

151

from A. capillaris root tissue showed substantial sequencesimilarities with the RdRp domains of WYSV and RYSV
(Huang et al., 2003; Maurino et al., 2018). Sequence similarity search of the A. capillaris contig against the NCBI
protein database confirmed that it encodes multiple
proteins similar to those of known members of the genus
Alphanucleorhabdovirus; the highest sequence similarity scores with WYSV and RYSV of this genus. Therefore,
the A. capillaris contig was assumed to be the genome
sequence of a novel member of Alphanucleorhabdovirusof the family Rhabdoviridae and tentatively named Artemisia capillaris nucleorhabdovirus 1 (AcNRV1). The
AcNRV1 genome sequence has been deposited in NCBI
under the Acc. No. OM372677.
The AcNRV1 genome is 13939 nt in length and has six
complete ORFs: N, encoding a 479-amino acid (aa) nucleocapsid protein; P, a 365-aa phosphoprotein; P3, a 295-aa
movement protein; M, a 269-aa matrix protein; G, a 655-aa
glycoprotein; and L, a 1960-aa polymerase (Fig. 1). N, P, M, G,
and L are canonical rhabdovirus proteins shared by members of the family Rhabdoviridae, whereas P3 is present inmembers of the five plant rhabdovirus genera Alphanucleorhabdovirus, Betanucleorhabdovirus, Cytorhabdovirus,
Dichorhavirus, and Gammanucleorhabdovirus (Walker
et al., 2011; Walker et al., 2018; Shin et al., 2021). AcNRV1
proteins have functional domains commonly found inother rhabdoviruses. The N protein has a “Rhabdovirus
nucleoprotein” domain (InterPro Acc. No. IPR004902). The
L protein has a “Mononegavirales RNA-directed RNA poly-Fig. 1Comparison of the genome organization of AcNRV1 and related viruses
Genome organization of AcNRV1 and two closely related viruses (WYSV and RYSV) are presented. The RNA-seq read depth of the AcNRV1
genome contig is shown at the top. Protein-coding ORFs are indicated with light green (AcNRV1) or white (WYSV and RYSV) arrows in
the 3'-to-5' direction. Yellow regions in the WYSV N and P3 ORFs indicate genomic segments that were incorrectly predicted as being partof the ORF. Light blue shades connect orthologous ORFs of the three viruses (note that AcNRV1 lacks the P6 ORF). Predicted functional
domains of the AcNRV1 proteins are marked by blue lines above the ORF: Nuc, “Rhabdovirus nucleoprotein” (InterPro Acc. No. IPR004902);
SP, signal peptide; TM, transmembrane domain; Pol, “Mononegavirales RNA-directed RNA polymerase catalytic domain” (IPR014023);
Cap, “Mononegavirales mRNA-capping region V” (IPR026890). Red diamonds indicate putative conserved regulatory sequences in thegene junction regions.

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merase catalytic domain” (IPR014023) and a “Mononegavirales mRNA-capping region V” domain (IPR026890).
The G protein has a signal peptide and a transmembrane
domain near the N- and C-termini, respectively.
WYSV and RYSV are the closest known rhabdoviruses
based on comparisons of the L and N protein sequences(Table 1). Further sequence comparisons using the otherfour proteins of AcNRV1 and their orthologs in WYSV and
RYSV confirmed that they are closely related. Multiplealignments of all six proteins are presented in Supplementary Data S1. The WYSV and RYSV genomes have anadditional ORF (called P6) located between the G and L
ORFs, in the order of 3'-N-P-P3-M-G-P6-L-5' (Fig. 1) (Huang
et al., 2003; Liu et al., 2018). The P6 proteins of WYSV and
RYSV share 29.5% aa sequence identity, implying that they
descended from a common ancestral protein. However, no
ORF equivalent to P6 was found present in the genome of
AcNRV1 or that of any other known rhabdovirus, indicating that the P6 ORF is specific to WYSV and RYSV.
In rhabdovirus genomes, the gene junction regions
have a conserved regulatory sequence involved in thetranscription termination and polyadenylation of thepreceding gene transcript (the TTP element) as well as
the transcriptional initiation and capping of the following gene transcript (the TI element) (Jackson et al., 2005;
Ogino and Green, 2019; Bejerman et al., 2021; Shin et al.,2021). The gene junction regions of the AcNRV1 genome,
together with those of the WYSV and RYSV genomes, wereanalyzed to predict conserved sequences. Seven (3'–N,
N–P, P–P3, P3–M, M–G, G–L, and L–5' for AcNRV1) or eight(3'–N, N–P, P–P3, P3–M, M–G, G–P6, P6–L, and L–5' for WYSVand RYSV) gene junction regions were extracted. A motif
search analysis using 23 gene junction region sequences
revealed shared motif sequences 20- or 21-nt long (Fig. 2).
The deduced consensus sequence of the AcNRV1 genejunction regions was 3'-(A/U)UUAUUUUU–GGG–UUG-5',
where dashes (–) separate TTP, UIS, and TI elements.
The TTP element in the gene junction region inducestranscription termination of the preceding gene. Since
the AcNRV1 genome sequence was assembled from a large
number of viral transcripts and a small number of viralgenomic RNAs, the sequencing depths near the conserved
sequences in gene junction regions would be lower thanin other regions. The read depths of the AcNRV1 genome
sequence sharply decreased in regions where conserved
regulatory sequences were predicted (Fig. 1). Similar sharp
drops in sequencing depths at the conserved sequenceswere observed in other rhabdoviruses (Shin et al., 2021).
The TI element of a conserved gene junction sequence
mediates the transcriptional initiation of the downstream
gene. In AcNRV1, WYSV, and RYSV gene junction regions,
the TI element has a 3'-UUG-5' sequence. When the positions of ORFs and conserved regulatory sequences ofFig. 2
Conserved sequences in the gene junction regions
Putative conserved regulatory sequences for polyadenylation andtranscriptional initiation, identified in the gene junction regions of
AcNRV1, WYSV, and RYSV, are shown in the 3'-to-5' orientation. A sequence logo representation generated from 23 sequences is shownat the bottom. TTP, transcription termination/polyadenylation;
UIS, untranscribed intergenic spacer; TI, transcription initiation.

AcNRV1, WYSV, and RYSV were compared, two conserved
sequences of WYSV associated with the N and P3 ORFs
were found located within the respective ORF (Fig. 1). The
predicted TI element possibly associated with the N ORF
is located at positions 209–211 of the WYSV genome sequence, whereas the start codon AUG is at positions 77–79.
The second in-frame AUG codon is present at positions
230–232, which is closely positioned after the predicted TI
element, supporting the possibility that the second AUG
codon is the actual start codon. Sequence comparison of
N proteins of AcNRV1, WYSV, and RYSV showed that the153

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Table 1. Sequence comparison of the L and N proteins of AcNRV1 and representative Rhabdoviridae viruses
No

Genus

Virus

1

Alphanucleorhabdovirus

Wheat yellow striate virus

Acronym

Genomea

Lb

Nb

WYSV

NC_055484.1

937/1992 (47.0%)

199/588 (33.8%)

2

Rice yellow stunt virus

RYSV

NC_003746.1

932/2012 (46.3%)

200/525 (38.1%)

3

Joa yellow blotch-associated virus

JYBaV

MW014292.1

706/2097 (33.7%)

122/535 (22.8%)

4

Potato yellow dwarf virus

PYDV

NC_016136.1

689/2085 (33.0%)

140/510 (27.5%)

5

Physostegia chlorotic mottle virus

PhCMoV

NC_055466.1

717/2096 (34.2%)

119/504 (23.6%)

6

Eggplant mottled dwarf virus

EMDV

NC_025389.1

710/2080 (34.1%)

119/521 (22.8%)

7

Peach virus 1

PeV1

MN520414.1

720/2102 (34.3%)

152/505 (30.1%)
129/502 (25.7%)

8

Maize Iranian mosaic virus

MIMV

NC_036390.1

659/2102 (31.4%)

9

Maize mosaic virus

MMV

NC_005975.1

655/2131 (30.7%)

129/523 (24.7%)

10

Taro vein chlorosis virus

TaVCV

NC_006942.1

679/2106 (32.2%)

135/546 (24.7%)

11

Morogoro maize-associated virus

MMaV

NC_055512.1

654/2078 (31.5%)

130/513 (25.3%)

12

Agave tequilana virus 1

ATV1

BK014297.1

677/2113 (32.0%)

138/507 (27.2%)

13

Gammanucleorhabdovirus

Maize fine streak virus

MFSV

NC_005974.1

590/2213 (26.7%)

121/554 (21.8%)

14

Betanucleorhabdovirus

Alfalfa-associated nucleorhabdovirus

AaNV

MG948563.1

595/2261 (26.3%)

102/524 (19.5%)

15

Sonchus yellow net virus

SYNV

NC_001615.3

577/2291 (25.2%)

111/524 (21.2%)

16

Cardamom vein clearing virus

CdVCV

MN273311.1

605/2321 (26.1%)

116/543 (21.4%)

17Black currant-associated rhabdovirus

BCaRV

MF543022.1

583/2356 (24.7%)

99/545 (18.2%)

18

Datura yellow vein virus

DYVV

NC_028231.1

607/2343 (25.9%)

98/524 (18.7%)

Sowthistle yellow vein virus

SYVV

MT185675.1

624/2335 (26.7%)

121/523 (23.1%)

Orchid fleck virus

OFV

NC_009609.1,
NC_009608.1

590/2244 (26.3%)

110/544 (20.2%)

21

Citrus leprosis virus N

CiLV-N

NC_052231.1,
NC_052230.1

591/2132 (27.7%)

104/522 (19.9%)

22

Citrus chlorotic spot virus

CiCSV

NC_055209.1,
NC_055208.1

586/2149 (27.3%)

116/524 (22.1%)

23

Clerodendrum chlorotic spot virus

ClCSV

NC_043649.1,
NC_043648.1

579/2121 (27.3%)

113/507 (22.3%)

24

Coffee ringspot virus

CoRSV

NC_038755.1,
NC_038756.1

561/2146 (26.1%)

115/522 (22.0%)
88/574 (15.3%)

19
20

25

Dichorhavirus

Northern cereal mosaic virus

NCMV

NC_002251.1

543/2340 (23.2%)

26

Barley yellow striate mosaic virus

BYSMV

NC_028244.1

561/2333 (24.0%)

93/526 (17.7%)

27

Rice stripe mosaic virus

RSMV

NC_040786.1

537/2320 (23.1%)

83/613 (13.5%)

28

Tomato yellow mottle-associated virus

TYMaV

NC_034240.1

519/2303 (22.5%)

86/528 (16.3%)

29

Lettuce necrotic yellows virus

LNYV

NC_007642.1

544/2293 (23.7%)

89/583 (15.3%)

Trichosanthes associated rhabdovirus 1

TrARV1

BK011194.1

554/2391 (23.2%)

72/566 (12.7%)

Lettuce big-vein associated virus

LBVaV

NC_011558.1,
NC_011568.1

516/2273 (22.7%)

95/559 (17.0%)

32

Alopecurus myosuroides varicosavirus 1

AMVV1

NC_026801.1,
NC_026798.1

483/2345 (20.6%)

98/626 (15.7%)

33

Red clover associated varicosavirus

RCaVV

MF918568.1,
MF918569.1

492/2278 (21.6%)

94/552 (17.0%)

34

Zostera associated varicosavirus 1

ZaVV1

BK014484.1,
BK014485.1

529/2259 (23.4%)

89/562 (15.8%)

Cytorhabdovirus

30
31

Varicosavirus

NCBI Acc. No. for genome sequences (note that dichorhaviruses and varicosaviruses have two segments); bamino acid sequence identity
to the AcNRV1 L and N proteins in the format of “identical residues/aligned length (percent identity).”a

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(a) L

(b) N

Fig. 3Phylogenetic relationships of AcNRV1 and known plant rhabdoviruses
Maximum-likelihood phylogenetic trees were inferred based on the L (a) and N (b) protein sequences of AcNRV1 and 34 representativeplant rhabdoviruses. AcNRV1 formed a subclade with WYSV and RYSV within the genus Alphanucleorhabdovirus. Bootstrap support
percentages, 95% or greater, calculated from 1000 replicates, are shown at the respective nodes.

WYSV N protein has an extended region at its N-terminuscompared with the AcNRV1 and RYSV N proteins, supporting that the WYSV N ORF was overpredicted (yellowhighlighted regions in Fig. 1 and Supplementary Data S1a)and that the second methionine is the true N-terminus.
Similarly, the predicted TI element associated with the
WYSV P3 ORF is located at positions 3395–3397, whereas
its annotated start codon is at positions 3348–3350. The
P3 ORF has a second AUG codon at positions 3468–3470,
which is in good agreement with the N-termini of the
AcNRV1 and RYSV P3 proteins, indicating that the second
methionine is the actual N-terminus (Supplementary
Data S1c).
Overprediction of an ORF may hinder the identificationof any associated conserved regulatory sequence. Indeed,
the originally deduced regulatory sequences in the WYSV
3'–N and P–P3 regions were incorrect and markedly different from the typical consensus sequence because they

were predicted from incorrectly defined untranslated
gene junction regions (Liu et al., 2018). Therefore, for thecorrect annotation of ORFs and conserved regulatory sequences in a rhabdovirus genome sequence, the locational
relationships between ORFs and conserved gene junction
sequences must be carefully examined.
To determine the phylogenetic position of AcNRV1,
the L and N protein sequences of 34 viruses belonging to
the six plant rhabdovirus genera were collected (Table 1).
The AcNRV1 L and N proteins showed 30.7%–47.0% and22.8%–38.1% aa sequence identities, respectively, withthose of other members of the genus Alphanucleorhabdovirus. The L and N proteins of other rhabdoviruses showed
20.6%–27.7% and 12.7%–23.1% aa sequence identities with
the AcNRV1 L and N proteins, respectively. Maximumlikelihood phylogenetic trees constructed from the L and
N protein sequences confirmed that AcNRV1 belongs to
the genus Alphanucleorhabdovirus and is closely related

CHOI, D. et al.: NOVEL NUCLEORHABDOVIRUS IDENTIFIED IN ARTEMISIA CAPILLARIS
to WYSV and RYSV, whereby the three form a strong subclade (Fig. 3).
In conclusion, the genome sequence of AcNRV1, a novel,negative-sense, single-stranded RNA virus, was identified
in A. capillaris transcriptome data. The AcNRV1 genome
has six protein-coding ORFs that are shared with membersof the five plant rhabdovirus genera. Phylogenetic analysis revealed that AcNRV1 is a novel species of the genus
Alphanucleorhabdovirus of the family Rhabdoviridae. The
AcNRV1 genome sequence may be a useful resource forstudying the evolution of genomic organization of plant
rhabdoviruses.
Acknowledgment. This work was supported by the National
Research Foundation of Korea (NRF) grants funded by the Government of Korea (2018R1A5A1025077 and 2020R1A2C1013403).
Supplementary information is available in the online versionof the manuscript.

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Acta virologica 66: 138 – 145, 2022

doi: 10.4149/av_2022_204

Supplementary information
Artemisia capillaris nucleorhabdovirus 1,
a novel member of the genus Alphanucleorhabdovirus, identified
in the Artemisia capillaris transcriptome
Dongjin Choi1, Chaerim Shin1, Ken Shirasu2,3, Yasunori Ichihashi4, Yoonsoo Hahn1*
Department of Life Science, Chung-Ang University, Seoul 06974, South Korea; 2RIKEN Center for Sustainable Resource Science,
Yokohama, Kanagawa 230-0045, Japan; 3Graduate School of Science, The University of Tokyo, Bunkyo, Tokyo 113-0033, Japan;
4
RIKEN BioResource Research Center, 3-1-1 Koyadai, Tsukuba, Ibaraki, 305-0074, Japan

1

Received February 17, 2022; accepted April 5, 2022

Corresponding author. E-mail: hahny@cau.ac.kr; phone: +82-2-820-5812.

*

S2

Choi, D. et al.: Novel nucleorhabdovirus identified in Artemisia capillaris
Data S1. Multiple sequence alignments of the homologous proteins of AcNRV1, WYSV, and RYSV(a) N (potentially overpredicted region is highlighted in yellow)
AcNRV1WYSV
RYSV------------------------------------------------------MSDLSAYISTPLSGTFPLFRNRDNVPTVGNEAQEEPYIYQSYITWLMDTSLSGSQKYLGPRKRRHIPNHGKNITHRQGASCSSISHFKNPNNNFTGTMAQNPNVANYANAAPLPRFEGLGDRENLAPIGNEAVEIPYQKEAYLAWI----------------------------------------------------MANDNVSDYANAAPFARFANLQNRETLNPIGNEAKEIPYNRDQYLTWL::: * .:.
* : :*:.: .:**** * ** : *::*:

46
100
48

AcNRV1WYSV
RYSV

NDGNTFALERLTDDQVIASWTAIEASMEGNTFSESNMRALVQCALNLRGVIGDNRPLYRELPENAARRYAPAPSTAPIYGGEAMQAGMLMPDMQIRQ--DNEGRVFQVNQLTDEQMIQMWETVKTSMQGNTFSEQHMRDIVQMACNLKGVDPATKPLYRQYEMPENGRWADAPSQDPIFSGQQ-VAGVIVPLQEAQP--LAEGKLFQIGALTDAEIVAAWTTIKTAMGNNTFSETHMRSIVKIACNLRGITPGSTPLLVTYNPPQSATWAPAPSTDAIYSGTP-VAGVIIPQNTGAGGED:*. * : *** ::: * :::::* .***** :** :*: * **:*:. **:* *** .*:.***:::*

144
197
147

AcNRV1WYSVRYSV

QESNTSRAKAICFICAYLVRLITKMDTHLEKSIEDIKKQYGKLYNTQSSLLNNWSPNITWAKRVKTAFDTYPWLKSTVAFAVGEADSSLERNTLDYGLCRVEDVSGKARAIGFICGFLLRFIVKTEEHLNNSLANLKLQFSRIYGVQSATINQWNPTTTWASRIKLAFDTYLTLRATVALHVALADGNLNADNVNFGLCRTETEASKARAIAFICCYLLRFIVKTEEHLTNSLGNLKLQYSRLYSAQSATLSNWNPSNTWASRVKLGFDTYLTLRATVAYNIASADALLVPENVNYGLCR* :.:*:** *** :*:*:*.* : ** :*: ::* *:.::*..**: :.:*.*. ***.*:* .**** *::*** :. **. * :.:::****

244
297
247

AcNRV1WYSVRYSV

MLLFQHLELSGMQIYKMTMTLIAHLNLIEPERFLEWICLPASTETVQKIYQIGITYDGPNHQANKHWKYAKLAKPNYWLSVNVRHNKFLAYLLGDLQVAY
344
MLVFQHLELSGLQLYKMTMTLISHLNLISPAKFLSWVYDPLAEKPITQIYTIATTHDTRDRQDQKHWKYAKLARGQYWLDTTVKRNQFFAYVLADLEVRY
397
MLVFQHLELSGLQLYKMAMTLIAHFKLIEPNKFLSWIYDPLSEASIDQIYKIAVNYDNVNSKTHKHWKYAKLARGQYWLNTTVKRNQFLAYILADLELKY
347**:********:*:***:****:*::**.* :**.*: * : .: :** *. .:* : : :*********: :***...*::*:*:**:*.**:: *AcNRV1WYSV
RYSV

GLGGKTEYSDPRNMGALEGMPENQKATAIKVSDTIVKFYEAIEKQQSGGSGPAWRAAMGLAAA---------------------PMRAQDRQD------GLGGKTEYSNPKRMKALDGTPVETRNDAESVAAAFQQMYRVIEDEKRREAGAAFRLARGMPPP----------PN-------PPPAAGGQGGG------GLAGKSDYSSPKRMKALSGMPVERMTEAETISKAVEQMYTAIESAKRVDAGAAYRLAKKLGPPRANAHSRRKEPNNSRQHRDKQPNSKQQGRDKRNKHQV**.**::**.*:.* **.* * :* .:: :. ::* .**. ::*.*:* * : ..
*: .

AcNRV1WYSV
RYSV-PTPEEIRLQQEEAERARILEEERLRQERAANNQGGEGGGGGDVNMADPQ----------PDDRALLADMLGIQ
VGAQGLGNVQAPPGAGGQVIPPGNLPPPPAAPPAGAAGQAPG---QQDQQINPPGQVPMDLDQRARDAAALGLVLGRHSKPQAQGPPNKQQDLGPHNNKPKEASRPPQQDRQQLAQPWRLTRRQRGARGHRTQTLSGMFCKGTCQCIQ
. *
:
.
:
*
.
.
:

416
473
447

479
544
521

(b) P
AcNRV1WYSV
RYSV

MSGENNTQPKSNANVN-------TRSKTKHDKNAYDTAPPPPPIPPVTGRYSGFNPGSAQIISNDNRDSNAKTFETGAPPTLPPPPDQTLKRQEALAAPLMADSLDTDRGRTTRSGSKLKAQTTHRVGTSSGSVRGGKPYEKDFKAVEARFHNFDPISASIIRGDQEGTKADLIMSDSSVTKATAP------AEAAAPPPMSGSGSEQTPRLTRSSS-----RSTLTGVASGRVEKIRSSPKSLDRIAKKYKDFDPETAKIIRADLEETQADKTMEVGGSTQESA--------------*:.. . :
:. .
:
. .
.
: : :: .*:* :*.** * . ::*.
. * ..

AcNRV1WYSV
RYSV

PANSSSEESGASSEERNQARQSPGDKRAKSASQSSKADISNKRFKMDTEMASLNLAFRDQEMLPMDKKDGASIVKTFCDTIKTQQKEITVADIVNIFAAG193
PPQPAAVPLTPPASATTQGQKRPGDPETLEEGNAAKRVQRAN---------KVNNLLTTQGIGEPSKSAISQYVVARLNANNVEHDDAMVAECANMAVYG
185--------------QQITGAKRPNEEDQGGAQEAAKRVNRSN---------KVNSLLTSNGVTDPAKSKISNYIVGRLNANNIEADSVMVAECTNIAIHA
157. : *.::::*:.:* : : :*. :. ::: : : .. **: .*:.

AcNRV1WYSVRYSVYKLARKSIPTDILHQISGTLPELITELRTVSGSISLGVGNLDKSIHDMTTHMSNSFDLLLENDRIKNACETTLLATAQQIYQNSVKETKVDFINNYIQKSWKEGKKFVSTQILNQATTVIPDLITSMVTNANLITNAANALNSIPDKVAGAIRTNIEHVAFQTGSKATKRDTLVRTAESIYQNAAAESKVDFINNFIISAWKEGKKYLDDKIISQATTTIPTLITNLVSNANTLSNVIASLNNVPDKLVSDIRTQVENVSNQTGQKAAKRDVLLKSSESIYNNAVKESKVDFINNYLTSS:* .:* : .*: * : .:* ***.: : :. ::*:. ..:. : ...: : :* : . .*: :::.**:*:. *:*******:: .:AcNRV1WYSV
RYSV

KIDLSILKTNIDTYKAVYHAIIPEYTVLILSGEGESYPGLVSKIKDNMALIMNIARSSLPRSVYQATTSYARGINIQEVKRNVANYRTIANAIIKRNTVLNIINETTEHQALLTQVQGNKDDIRNIARGLSASYVI-------GVNVDELRKDSHHYRTVVSRIEKKYTVLVMMPEHEEHHTLKEKVATNRTFVKESAQTLSQKYVTQ------:::. :: :*:::* . *** : * .: * :: *: : *:
*

365
349
322

93
94
80

293
285
257

Choi, D. et al.: Novel nucleorhabdovirus identified in Artemisia capillaris

S3(c) P3 (potentially overpredicted region is highlighted in yellow)
AcNRV1WYSV
RYSV----------------------------------MEDSKYFEGPIKAGKITLLNGKASQINLKLRGSFFSNMSLIGTKKRQSWSVGDTT-------TITN
MYSKFTYVSLLYLKTPTEIYSLHSRQPNSEERPKDRKTTSMASEAKNSQKFSFRNTEEEVDLSISKFALFKLKLKQSRVVTMFGNDDKVDPANCYINMTS-----------------------------------------MGEGKNHQSFSFKNADDEIDLSLSKFSLFKLKMAKSKIIKVFGQNDPVDPNNCYINMRS. * ::.. .:::*.:: ::.: :::. .* .
.: .

59
100
59

AcNRV1WYSVRYSVVTLKVRSILHVLSDEYIKATITLDSSGKTTRVGWALWKSSFSGNIVFQTELGICKTHQGVYTPYSINIETSTGEELKGVSIDMKVVIREPGHNKLNSGEG
IKIVTSSVLPESDPRFMVWEMSYKTDEENHKLGQLAWKASYNGSFVIKSTYAMMVVNGDLYTPYTASITTSDGSEIKGVKVNVILNWTPSDVRPSNAKMG
IKITTSSVLPESDPKYLIWEMSYKTDEEDHTLGQLAWKASYNGTFIVTTTYAMMVTGGELYTPYTAVIRSSDGNEIKGVKVKVTLSWDPANDRPSKARMG:.: . *:*
. .:::: .:. ::***:*:.*.::. : .: .
:****: * :* *.*:***.:.: :.. . :: *

159
200
159

AcNRV1WYSV
RYSVSAILDVYS-LMTGDNSHYHPVLAWYVNKEGYAKQRILSTEPEAIMAGYIPVFEAFPTLQGQEPILHYENLLYFILEDEELKVFNNIIGDRRRGMSPRQSD
GFMSDIYCNNPSNGKTQIPPMVAWYMGKDEQRYCKIINKLATELSSENIYPMMELISSPQTSLNPILNRLISSSLEKGDRDRISQMTKAVGGGMSLNKAD
GFIQDMYCKTITNGKTQISPMVGWYIGQDERRYCKVLNKSALEFSSEGIYPLMELVSGADSVINPLINKLISGMLNDEEKRRVSLYTSTVGAGTSLTQSE. : *:*.
:..::: *::.**:.:::::.. . : : * : : :
:.*:
*:. :
..
* * :::

258
300
259

AcNRV1WYSV
RYSV

IEHGMRILTSQKLTCLIEFLKRASPKIKGDVDFFHKVISYLKGIIEKKSSSALVTFLMRASTELGVEVYIDG-KLLLKKLVESKTGSGLVQFLMRACKELGTDVYLEA-:: .:. : *: ** **. :: :* :

295
335
294

(d) M
AcNRV1WYSV
RYSV

M-TSEQEITIIRPTSQKSNAGIENDGVVVDSQYHLPPIVNYSLSFMIEISFYDQESCKMFLDKKMKTVELFGALKENWINSTGAVKYHTPVEPTDEQAHLMAHSKIRITGAGLDGVSYLERIRKSLSLNPADYHDDSAVRISLSFYIKISFHDVSDYDVFIREGVTTTELFDALKTSWKRNPENVRYIDGMTHTDDEIDTMANKKIRVTGAEAEQPSILKRISGALTLNPLDYHLDYSKLISLNFMVKISFHDASDYDLFVREGITPVELFEALASNWSTDSGEVHYVDGNTHDKDEIDT* .: .:*.
*::****.* ::***:* .. .:*: : :...*** ** .* .. *:*
.:: .

99
100
100

AcNRV1WYSV
RYSV

VIDICCLIAVLRELPFVEGIIQTHFHRNTTVLRMSFGNVLPQTVHTSIVPISSQLQLPLIFHGNLRYEFVEDTGKIHASVMMGIRATAFPKGNSPPVMDTTILLCELVTILKDLSLHRSDERTHYSLMSTSLTLGFGDQITQPHDNAVIPIVTTKVIPRYMHTVIQYEYPRVSGGIAASVCAGICIRSPPIGKCPPIMKTSVKLCELITIIKDLPFHKSEESTHFSILSTSLTLGFGDQILQKHDNSVIPIITERSLPQYMHAIIQYEYPRVSGGIAATICAGICIRSPPIGNCPPIMKP: :* *::::::*.: ..
**::* * :.**: : * ..:::** ::* :* ::**: . :* * *:: **: * *:.**:*..

199
200
200

AcNRV1WYSV
RYSV

VHLEHLLPHHGIREIQPQLPSSTSSNQAQDGDETGSTERRRKREGNDAGALVPSMKKAATSLANRLLRRTVNVEVLAFHYGLDAGQGPQLEETNATPKEEELKKINGFKRMTTLNKSSRVIKKPSKEGVGAALSRMLSWKLHLELLCYHYGLKMSG------DAPSPAEG---KIGRIKRPTERKEDTPSMTKRLKGGVGATISRMLSWKE:::* * *:*:.. :. . :* .
:.: :* .. : .*:* .

269
270
262

(e) G
AcNRV1WYSV
RYSV

MSYARVLVSLTTLIILTQVFLLLEEACSEISYWE----------KDGKWYAQWVPSPATSVEVPAPSPERIPAPPSPTLDKLPAVHQTTYG--TIHVG-V
MRTTTRMLTIIICMLFGLYMIMLGENMILAFGPVDGDDD----SQENRPFTLHPAHGAIPVLPSSQPSQKVSPDDTNSLIRLTTSEQLWHN-HPQDIGPKMNIIKRAGVFIGLVYFTVLLIIYGEHIVRSYATEDCMPAGLYYYAEGAAYTNLPVCKDEPKSSYKGTELFPTSPNIDKPLRMTTDIMVHPISSTSDLGPM
*
:
: :
::: *
:. ::
.
.
..
. ::.:
. .:*

87
95
100

AcNRV1WYSVRYSV

PTDTDLYPIYYCPNVSIYDLLPVWYSGCLSMCSITSVQDLVDISVFQINSTLGSTDLYKVSVRSVTKYSHVSVFGSCTVSVTADDYVSPPSDKLSEWVTK
DVPADMYPIYSCPNLSNAYLLPIWYGSCLDACQITTPKHTVNVKLWTINSSVTDVDGYQIDVYYDTKFSHVGPFGGCSVSLSESVTKEPKQEDIMIWKSR
DVPPDMYPMYSCPNLSNSSPLPLWYGSCINSCQITSSKETMAVTVWAINSTVDIIMGYKLTTFFETKFSHVGPFGGCSISLVGSEPRSAPEALISGWRSK. .*:**:* ***:***:**..*:. *.**: :. : :.:: ***::*:: .
**:***. **.*::*: .
.. . : * ::

187
195
200

AcNRV1WYSV
RYSV

LSIDRPSGSQEWDYSSDPPCNYFSDEYSTGYKLYVTHYKVEISVDSAQNRYLRDPTNGLYMSYEQDYYVDGSTFYHWVQNSDVSQEVCYFREIGEDSCTVLVSKPVNDVESWIMYDEPSCNYFSDEYSSGFRLVITRTKLKLMIDSVGNLYIADLRPGSYDTYKTGYAIHGSTAWIWDTDDSMNHGMCYFKQTDDTYCDYILSQGNIQDAAWYLYNDPECNYFSDIYSSGFFVRIDRIKLTVLIDAVGNLYLSDLVAGSYDSYDKGFATHGNNVWIWDTDRVKVDNNCYFKQTDDTYCDY: .
*.:* ****** **:*: : : : *: : :*:. * *: ** * :*. .: .*.. : * :. ***:: .:

287
295
300
*

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Choi, D. et al.: Novel nucleorhabdovirus identified in Artemisia capillaris

AcNRV1WYSV
RYSV

DSSSGYISCRVLGITFKKDIKNIITSSCAGSLNVSSDGVIYQVLSAYNKTSLSDRLSGLLSS--NINMNQLIHVINDALISIEDAYCSSTCDVMEIILSNDNNTKYMFCKMSGVSFDTTVQQRITSSCAGDLNISTDGVIYQIGDSGDTASTQQRLSDILHQNVELGMQSLVSLINDVFINIESSYCTGVCDIMEVIVSN
DNTTGYILCKTIGVSFRSELQTRVESPCAGHLNISTDGVIYRLEGTADTPSTQDRLSSILKNNVNLGMESLISLINDVFTNIESSYCTGMCDLMEVILSN*..: *: *: *::* . :: : *.*** **:*:*****:: .: :..* .:***.:* . ::.*:.*: :***.: .**.:**:. **:**:*:**

385
395
400

AcNRV1WYSVRYSVYPTATTVIETPIGPWLPLVAKGNTVMTACSADTEWYLKLPIEYCAEKNMMKVVHNSTKSEEWWRIMNTYVVSGESCNSSDSGWDEILVEKMQSKREIVFHYPTATTVLETPIGPWLPITSDGHTIMTPCMADVNWIIQTPIVYCFSKEMIKVINKDTRKEAWWRIVNSYIILNETCSDTNSTALEILRDRMSKRRDIVYSYPTATTVLETPIGPWLPITTNGHTVMTACSSDPNWVIKTPVSYCYSKKMIKVVNKDTMREAWWRIVNSYIILNETCTDENTTSFNLVKERMELRQDITYS*******:*********:.:.*:*:**.* :* :* :: *: ** .*:*:**:::.* * ****:*:*:: .*:*.. ::::: ::*. :::*.:

485
495
500

AcNRV1WYSV
RYSV

FWRGDLTLVYPYNVSHWTLYKDEKIQRSSKWFPKLQYMMLDKEIYIDNVTQALHNYTQDIYNHYMGNQTASTGLGSIIPRILDRITLSVGNAVKLVVKISFWRGDLIVSYPYNKSRWITYKDEKIQRSSKWFDKLVDLKYKHPITLDNITSQLVNHTADLYEWHMGDKNGTAGQ-TTFSDLLGRVEKAGTNVIKGCVKMTFWRGDLIVSYPYNKSRWITYKDEKIQRSSKWFDKLLPLRYNHPITLDNITMELINHTRDIYDVHVYPSQGSTSK-RPMSDVIGRVGAAGSNLIKHLVKGI****** : **** *:* ************* ** : .: * :**:* * *:* *:*: :: . .::.
:. ::.*: : * :* **

585
594
599

AcNRV1WYSVRYSVGEVILFIMSHIHLIGDIIITVVCMVIGYYLIYIPTRWVRKRRGD-TSARITAVSNSYEMVPSSRTTRKDILGNLLIWITSHIEMIGDMLIIIVCLIGGYYVLIIPYGFLRRGRGPGTVTEVQQSNSQFRSPLIPRTYL---GETFFWVTQHIELICDILIIIVCVVIGYCVVLKPILYIT-MRGREKQQPVIVMRDEGRSHLFSRSTATNAL*: :::: .**.:* *::* :**:: ** :: * ::
** .
:
.. .
.*:

655
661
669

(f) L
AcNRV1WYSV
RYSV--------MSDRDSDDDGADYSGRYEEDDEDSNYKRHGEKADMVEGGDYHLKSALKCEADLEMDHVYLKEKNAMMAELGDLGDLIYPHSVFQFIEKISHMMDVEEPTYWAHDEDDDYWWDADEYLGEEEEDEIYEDTED--ELVEGGDFHLKSALRGEEDMLQNPIYKKERDSMVEELGSLGTLMNHIDVVSMLNLIGTRMDDEGHGYWQDYDEDESWLDAEN---DVFDDDIFEEAEDNEHLVEGGDFHLKSALRGEADMLTNPIYEKEREKLQEDVGDLGVALGHLNVRPFLEKMGER. :.*:* .
: :*. ::
: .:*****:******: * *: : :* **:: : ::*.** :.* ::: :.

92
98
97

AcNRV1WYSVRYSV

WSTDR-ASGLSRWVTVNPDSRLD-MSNTKRLLGAEISLVCPDVNPTDVGGWLTSGYDAFAARFSHLEHTMKILLILLPILNNIKNLRNGMGVDNAINEYIVAQDKRPSGGHTFLHEGGFNIIP-LQENVKLIKAELMSVLPEMAGMNIDQVLSGSYHLFMADHPYVTCTISTVLFILSVLNNVKNIRQGIEVGTLIGRILISTDHTNTPLIENIKQGGFGAIHHMKSTARLIVAEAATVTETMLTSGVDTTLAGAYHFFEQNHPHMKCTVNMVLMFLTILNNVKNIRLNVDLHQILKNNI: *: :: . . : :... :*: ***:.:. *:..*. *..:: *:. :*::*.:***:**:* .: :
: . :

190
197
197

AcNRV1WYSV
RYSV

TINSGLVSVDMNSVSVVRMNTDIVVFDCPDERYYLPKDYFLNMCDKLQERVNIYLYTEVCSQTWVPLAPKIELVSRVIAWGDEVLKKLGNDGYRVIGIYEYRKGNLVCLSLAAATLCYLSTDILVFDVDGSRHYMPKTYFLNGCDKVQERFNIMLYSYMAESLGVPGSCPSQIVKRTISWGDNVLANMGNEGYNVIGLYE
SIKGSTVCMYITVATVAYFSTDIVVFDIDGQKYNTPKTYFLNACDKIQERFNVILYSYLAEGLSIPGSPPTYIVNRIIDWGDSILYQMGNDGYDVIALYE:.. *.: : .:: :.***:*** ..:: ** **** ***:***.*: **: :..
:* ::*.* * ***.:* ::**:** **.:**

290
297
297

AcNRV1WYSV
RYSV

ATLVGIILNRDDPSLITNPDRPGFLEALKEDLGGVEIRFIDQLVNMLSTVDVRTIIDLHGLYRIWGHPIIDIDGGVKKLQKVTQEKKEVDPDSEESINARAILVGMILDRDDEALSPTPRSESFLSNILTGLSSEEQGYAKRLITTLQDMSPAQLADLHGLYRIWGHPIIDIDGGVRKLQRVTREEKGDINAKPESRETVATVVGVILSRDDKDLSPTSGGGDFLENIQQDLGPVQQNHIRYLIALLHDMTPHQLADLHGLYRIWAHPIIDIDGGVKKLQKVTQSLKGDINSKPESQETV* :**:**.*** * ...
.**. : .*. : .
*: * :: *********.**********:***:**:. *. ** ::

390
397
397

AcNRV1WYSVRYSV

RSFRKIFCKNYFKKNGFYPPLKLIDRTTPCYLSRCLLEGVEVMENHMDYKFQQWDNVALQKAFSIPYSWNIVHLAKDKSISPTRSELYSLLSKTGNIFSARSFRRLFTTEYFKKHSIYPPMEITSKFN-TYLGNCIRTGKEIDEKHINYQFSDWDYIELQETFSIPYSWNVLHLAKDKAISPTRNEIYNMLLSKGRIFNARSFRRLFVMDYFVKHQFYPPITLPEKSN-CYIGNCIRTSKKIDESHINYNFSDWDLVELGGAFSIPYSWNVLHLAKDKAISPTRSEMYTMLCKTKRVFNA****::* :** *: :***: : .: . *:..*: . :: *.*::*:*.:** : * :********::******:*****.*:*.:* .. .:*.*

490
496
496

AcNRV1WYSVRYSV

NLRRGILKLLDSKLIPLRDFLTKVAAEGLHVDDCIIGLFPKEREIKILARFFSLMSFNMRLYFTSTESLLGDKVLKYFPQITMNLNLLDMQEKMSSMSKSELRRGVLKLMTTTLIPLRDFLKKVASDGLDIDDCIIGLFPKERELKILARFFALLSFNMRLYFTSTEELLGSKLLKYFPQITMSSNLLEMQEKMASMSKEELRRGVLKLMNTTLTPLREFLTEVAEHGLDIDDCIIGLFPKERELKIMARFFALLSFKMRLYFTATEELLGSKLLRYFPQITMSSNLLDMQEKMSSMSRD:****:***: :.* ***:**.:** .**.:*************:**:****:*:**:******:**.***.*:*:*******. ***:*****:***:.

590
596
596

AcNRV1WYSV
RYSV

LKEQSDSVTYVVNMDFVKWNQQMREEICRSVFAELDAIFGLPGLYTRSHQIFKQSILYLADGTRELHSDPHTGVHVDGKYCWRDDGAGKEGIRQKAWTIMLRTQNRSVTYVINMDFVKWNQQMRESTCRGVFTELDKLFGLKGLYTRSHQIFKDSVLYIADGTRRILPDPISGVMIDDNACWTDDGAGKEGIRQKAWTIMLESQNKSVTYVINMDFVKWNQQMRESTCEGFLKNWSKLFGLPGLYSRSHQIFRDSILYIADGTRDLTPDPDTGIMVDNNVCWIDDGAGKEGIRQKAWTIM*. *. *****:*************. *...: : . :*** ***:******::*:**:***** : .** :*: :*.: ** *****************

690
696
696

Choi, D. et al.: Novel nucleorhabdovirus identified in Artemisia capillaris

S5

AcNRV1WYSV
RYSV

TVCDIETVAQGHTGQFQLVGGGDNQVLTVTYKTNNIDGQGDITEEGKAKIKRKMENFLADLERHFQRRGLPLKTSETWCSTSLFMYNKHIYYKGAATRAPTVCDIAAVARHHPGKFHLVGGGDNQVLTITYHTNQVDTQGVITEAGKQKIKSKVKRFLTDLEKHFSERGLPLKTTETWCSTSLFMYNKYMYYQGSPLRSPTVCDIAAVARHHPGVFHLVGGGDNQVLTVTYHTNQIDTDGNITEEGKSKIKAKVKKFIEALETHFAARGLPLKTSETWCSTSLFMYNKFMYYKGVPLRSP***** :**: *.* *:***********:**:**::* :* *** ** *** *::.*: ** ** *******:*************.:**:* . *:*

790
796
796

AcNRV1WYSV
RYSV

LKQVSRMFPYSNNTSMSISSMSQCLGTTLRSICQKDYVPFPALFMRNYWAQLLCWIISYCHPMMMSIS-KNIKQGVGGICRRGRRLEVPVEGSCSKHLMLLKQVSRLFPFSNNTSMTLQSMAQCLGTGLRSVAQKELSHIPALMMRNIWGSILTWIVTVCHPMLLSISNQDSLKGDGIITRGKKEIKMKVESVESAPLALLKQVSRLFPYSNNTSMTLQSMAQCLGTGLRSAAQKEVSHIALLFMRNIWGSVLGWIILYCHPMLPSISSETCNSGVSTIVRGRKAINMKTHRIDLVALIL******:**:******::.**:***** *** .**::. *:*** *..:* **: ****: *** :.* . * * : ::: ..
* *

889
896
896

AcNRV1WYSVRYSV

RLAYLPGHFGGPGMINIYQAMMRGFPDPITEGICFLRNMTQALKRRDDILYSFFRDLSGVSFSLSRSMEPLIEDVTSLNNDAPRTGASEHREAAKEILTKKIIYLPGHFGGPGLVNLLQMTMRGFPDPITEGIYFLMAMKKNTARLGSAYTSLFQRMAGVSFSRSRNYEPLVEDVCSLNTDAPRSGTSEQREVARKILLKKILYLPGQLGGPGLVNIYQMTMRGFPDPVTEAICFLRKFKNYLICVGSSYSSHLARMAGVSFSASRSYEPLIEDVCSLNLDTPRSGTGGKREVPRKILLK:: ****::****::*: * *******:**.* ** :.:..
* : ::***** **. ***:*** *** *:**:*:. :**..::** *

989
996
996

AcNRV1WYSVRYSV

SKLGKNLELKSLISIMSGDLETQFYRSLASGPSLDVKVLHEIASATLYSITNTLTSRIDKTSTLSRLTHKISLLKKMALAEESYLKYLMIRDSRKHDLSFSKLGGNENLRDLLRIMEGDNEKNFYKALTASRVLDIRVLHEIASATLYAITNTFTSRVDRTATLKRLTLRYSMIKSLAESERKFLRYLLVRDLKQHDIMF
SRLGGNQHLKELLGIMKGPSEGEFYRAISSGKVLDVRVMHEITSSTLYAVTNTFTSRVDKTATLKRLTFKFSMLESLADAELKYIRYLSVRDDKTHDIMF*:** * .*:.*: **.* * :**:::::. **::*:***:*:***::***:***:*:*:**.*** : *:::.:* :* .:::** :** : **: *

1089
1096
1096

AcNRV1WYSV
RYSV

DACSRVLADKARSTSWRKQIIGVTVPTPFEYLVCKSGQDHVCNGDAIIARVDEV-DRTEMMTELGPCKVYLGTYTREKFKMTEIAAAYGEEDILAKSIQLEKCSRVTADECRTLGWGKQIIGVTVATPFEYLSVSLREKHVCDGNNIIVRLSSSGNKKQLGEVLGPCKPYLGTYTKEKFKMTEVAAAYGDEDVLTKALRIDGCSRIIADECRTKGWGKPVLGVTVPTPFEYLQISWTDEHICDNNHITVRISNQ-DRQVTTETLGPCKPYLGAYTKEKFKMTEVAAAYGDEDVLSKSLRI: ***: **:.*: .* * ::****.****** . :.*:*:.: * .*:.. ::***** ***:**:*******:*****:**:*:*::::

1188
1196
1195

AcNRV1WYSVRYSV

MKLINWRYPEGSQMAEILKCPLRAVTDLNPELLIQNEVLTKGDYDHRRKLDSRVHGGIPNFVTTSTSHTSLCTSTWVSHARGGKNENIHFQACLISSQFRMKIINWRFPEGSTMSEILKAPFRAVTDIDPQRMIQESSVTKGDYDHRRKMDSRVHGGIPNFVITPLSHMSICTSTWYKHARGGKNENIHFQACIIQTMYKLKIINWRYQDGSTMSEIIKAPFRAVTDIDPQRMVQESTVTKGDYDHRRKMDARVHGGIPNFVTTPLSHISISTSTWYKHARG-KNENIHFQACIIQTMYQ:*:****: :** *:**:*.*:*****::*: ::*:. :**********:*:********** *. ** *:.**** .**** **********:*.: ::

1288
1296
1294

AcNRV1WYSV
RYSV

ILCMRYWGILTSRVLHFHESCQTCIQALEQP--APGTTTPSIIPAFPTLRANRLVFVPSQEVEFDFEDQERVTRAQFLGLVDCEKKNVYDWADLYSSLAW
IVMLLMNNGKCDEIIHAHESCSTCICEITEP--VLDLVTPMMDLIFPQLQNNSLVYIPEQAISFDHSRMKEVDYARRVGMLDCHRDTDYTWLDTYSSLSW
IIIRTMSNIHSQEKLHVHECCQTSISEIQEPRANLDLTTPQMIPTFPTLMGNPLVYIPEQSITFDYSRTQEVDYSRRVGSLDCQRSTDYGWVDAYSSLSW*:.
.. :* **.*.*.* : :*. .** :** * * **::*.* : **.. :.* :: :* :**.:.. * * * ****:*

1386
1394
1394

AcNRV1WYSV
RYSV

TIICNMLGITSMSPSFYLIMRDNVNHHCLGLYLAMFWRCLRKLGALESTPNINWSPMIYTFSCEEGMRVLQEEMGLVITGSVEGGFVMGTGDSCDTLDLL
LIMCDVIGWTRMPHSFYFMIQNDVDHYLLSMYLICLWKVMRKEFELINT-QLDWEPMIKVYSTQEGITVLSNQMGLVVGGSLEGGLEVDMKDICSAFNTLLIMCDVIGWTKMPDSFYIMQQERVDHYLLSMYIISIWKVLKSEFDLHKT-MLDWGPLLKVYSSDTGVQVLSQSMGIVVLGKLEGGLSVNMMDICNSLTGL*:*:::* * *. ***:: :: *:*: *.:*: :*: ::.
* .* ::* *:: .:* : *: **.:.**:*: *.:***: :. * *.:: *

1486
1493
1493

AcNRV1WYSV
RYSV

LIDEMPPMTCNIFCPMIKWQVCVWWYSQSSFVSGCILCMRNIKSMWMSSNMLHRVDEDLRCSSHMTNEMKPVMINAHIENLRQYGKYSTEEPHTDIDTVC
TITQIPPFSVNLALPKIHKQVAAWWMLMDDNTLLCIDCHNNLNGYWDGTSINRRVDNDLRCYIHADGDVSPRIINAHISNLSQG----LIRLTPGGTSQSTSNQIPPFSVNLALPRIHKQVATWWKLTCSDTMYCIQCSNILKGYWEGTLINTRIDNDLRCSECADGGISPRIVNAHISNLSDH----LIKIIPDREEVQ::**:: *: * *: **..**. . ** * . ::. * .: : *:*:****. :.* ::****.** :
. ..

1586
1589
1589

AcNRV1WYSVRYSV

LPISKDLVEEEVGMIRRMISLPSEYPDEGSVPPVIRYGGDVDNVTLFRMILESLSLLNPVHIVLGPDINMLRNVVAGCSLTSLEMEIDTLMIDIVISALS
HPIIIG--GDKLNNLETVPSVEDEWPDVESITLLNQIGETNDIDGRLRFIIESMSVIMPDIVVVRPDMLDITLVKTARELLKCDDGSK-LIIYILVEEPSLPLLAG--GEDLNNLEVVLSLEDEWPDSDDITILNRIGEMNDIDGRMKNYLEVMALLNPDVVIIRPEILELEILRRVCEVVRLASEGK-LIIHILVEEPS*: .
:.:. :. : *: .*:** .:. : : **:: :* :::: * ::: *:: : :.:. *:* *::. *

1686
1686
1686

AcNRV1WYSVRYSVYQESDTFYELKAVVPKRMSRRIRFCMNAPQEGFCRFFIKPDRKLLKYLIAGDWVLCDCHELIEMNNTLKPDEVYYLPCTPLRW-LLLVKCGTGVFGPGDLYLESGTIYEMERAFPNSETVDVQFDYRKAPGGLHKLWINPREESIQRVEVGDWICISFTDLIKHHDSILADTVLTMKERRLGR-QFLVNKGVGSFKPGDLYLESGAIYEMERAFPRHNLFHVQFSFRKPPEGLHKLWLLPQDITLARADVGDWLVIPFTQLLDFHDTITPDTRLHHQREAIRQDVFWSTKEQGGLGLVVL* **.::**:: ..*.
::* . . *: :::: *:.***::*:. :::: .*
:
: .
* :
*

1785
1785
1786

S6

Choi, D. et al.: Novel nucleorhabdovirus identified in Artemisia capillaris

AcNRV1WYSVRYSVIETLSKPHYEP---RFWKIDHCIISAFSCGKLSGGGFTELRRKLSWSLFSNKYELLTICHRVTNQMNRSKAYGGLKQWSRNYREGIIVFILSVLMRSENHGKYIFSSELNMSYKQSRQVDYEINKVMIQNPTLGGKFMILKRRCSWALFSSRYEMLRAVERIKKNMTVEGRSTKTKEWSKTHQDDVLILIISMLMRADEKQHICTKTELSYRLDRSK-VDVEINKMMIKDPVMGGKYVILKRRCSWAIFSTKYELLCSAERIKGNLSADGKKITSKDFSRTYKEDIIIFIISVLMRSLDC
.
... .
:
:* * . : .
** : *:*: **::**.:**:*.*:. ::. .
*::*:.:::.::::*:*:***: :AcNRV1WYSVRYSV

DLHLITTLSYAEMDLTHLTITPKFNKAGVVSIRYLPYIWFFKRYYDTKRCHVDIEFATAIKSMNLGSPRGSGLNIIML---DESEIFTLGYVNCDVENLVMYPKFSSTQVVSLRYLDYFWFFKRMYEYQEKTFAIPYTEIRTSLNIGSPKRSGYNTINSSE-DEKRVMTLSYVECLPELLTLKPRFNKSGVISVRYLDYFWFFKRVYSYNNPEVAIPYSAVIKGVNLGSPRKEGYSLASSDGAP*: **.*.:*.: *:*..: *:*:*** *:***** *. :. . * ::..:*:***: .* .

1960
1965
1967

1882
1885
1885