Lavender Harbors More Viruses than Previously Thought: First Report of Raspberry Ringspot Virus and Phlox Virus M in Lavandula × intermedia.

Plants of the genus Lavandula are thought to be rarely infected by viruses. To date, only alfalfa mosaic virus, cucumber mosaic virus, tobacco mosaic virus, and tomato spotted wilt virus have been reported in this host. In this study, we identified for the first time raspberry ringspot virus (RpRSV) and phlox virus M (PhlVM) in lavender using herbaceous indexing, enzyme-linked immunosorbent assay, and high-throughput sequencing. Nearly complete genome sequences for both viruses were determined. Phylogenetic and serological characterizations suggest that the obtained RpRSV isolate is a raspberry strain. A preliminary survey of 166 samples indicated RpRSV was spread only in the lavender cultivar 'Grosso', while PhlVM was detected in multiple lavender cultivars. Although RpRSV raspberry strain may have spread throughout Auckland and nearby areas in New Zealand, it is very likely restricted to the genus Lavandula or even to the cultivar 'Grosso' due to the absence or limited occurrence of the nematode vector. Interestingly, all infected lavender plants, regardless of their infection status (by RpRSV, PhlVM, or both) were asymptomatic. RpRSV is an important virus that infects horticultural crops including grapevine, cherry, berry fruits, and rose. It remains on the list of regulated pests in New Zealand. RpRSV testing is mandatory for imported Fragaria, Prunus, Ribes, Rosa, Rubus, and Vitis nursery stock and seeds for sowing, while this is not required for Lavandula importation. Our study revealed that lavender could play a role not only as a reservoir but also as an uncontrolled import pathway of viruses that pose a threat to New Zealand's primary industries.

Detection, Characterization, and Distribution of the First Case of Pepino Mosaic Virus in Aotearoa New Zealand.

Tomato (Solanum lycopersicum L., family Solanaceae) represents one of the most economically valuable horticultural crops worldwide. Tomato production is affected by numerous emerging plant viruses. We identified, for the first time in New Zealand (NZ), Pepino mosaic virus (PepMV) in greenhouse grown tomato crops using a combination of methods from electron microscopy and herbaceous indexing to RT-qPCR and high-throughput sequencing. Phylogenetic and genomic analysis of a near-complete PepMV genome determined that the detected strain belonged to the mild form of the CH2 lineage of the virus. Subsequently, a delimiting survey of PepMV was conducted, and PepMV was detected at four additional locations. PCR-derived sequences obtained from samples collected from different greenhouses and from herbaceous indicator plants were identical to the original sequence. Since PepMV has never been reported in NZ before, seed pathways are speculated to be the most likely source of entry into the country.

First report of Fig virus B in Ficus carica in New Zealand.

Fig (Ficus carica) has been cultivated since ancient times, and is now grown worldwide, both for its fruit and as an ornamental plant. Several viruses and viroids are associated with Fig mosaic disease (FMD), a disease complex occurring worldwide (Preising et al. 2021). Fig mosaic virus (FMV), fig leaf mottle-associated virus 1 (FLMaV-1), fig mild mottle-associated virus (FMMaV), and fig badnavirus 1 (FBV-1) are known to infect fig in New Zealand (Minafra et al. 2012; Veerakone et al. 2015). In December 2020, leaf samples from a fig tree growing on the roadside at St Heliers, Auckland, showing dieback with foliar chlorotic mosaic symptoms, was received for virus testing. Total nucleic acid was extracted from the symptomatic leaves using a KingFisher™ mL Purification System (Thermofisher Scientific, Waltham, MA) with an InviMag Plant DNA Mini Kit (Invitek Molecular GmbH, Germany) and subjected to high-throughput sequencing on an Oxford Nanopore Technologies MinION device using the method described in Liefting et al. 2021. All sequence analysis was performed using Geneious Prime 2021.1.1 (https://www.geneious.com). A total of 355,858 reads that passed quality check were subjected to BLASTn search against the NCBI nt database as described in Liefting et al. 2021. The following viruses produced hits: FMV, FBV-1, FMMaV and a fig closterovirus. The presence of FMV, FBV-1 and FMMaV were confirmed by species specific RT-PCRs. To identify the closterovirus, reads were mapped to closteroviruses reported in fig including the recently identified tentative species fig virus A (FiVA; GenBank accession no MN817232) and fig virus B (FiVB; GenBank accession no. MN817233). Five viral contigs ranging from 939 to 2,340 nucleotides (nt) were obtained from mapping to FiVB. Subsequently, a 6.4 kb sequence (GenBank accession no. OQ968551) from the 3' region of the NZ isolate was amplified by overlapping RT-PCR using primers designed from the contig sequences. The sequence shared 79.5% nucleotide (nt) identity with FiVB The original sample and a further 25 symptomatic and 10 asymptomatic fig samples, collected from the Auckland area between 2016 and 2021, were tested using FiVB specific RT-PCR and Sanger sequencing using primers FiVB-F1 (5'-GAGGGAGAGATGTAGATGC-3') and FiVB-R2 (5'-TGTCGTCGATATCGTTGTGT-3'), designed to amplify a 725 nt fragment in the 70 kDa heat shock protein (HSP70) ORF. Products of the expected size were amplified from the original sample and three symptomatic samples and their sequences found to be identical. BLAST searches showed that the sequence (GenBank accession no. ON553403) shared 82.7% nt and 87.3% amino acid (aa) identity to an isolate of FiVB (GenBank accession no MN817233). These additional positive samples were collected from a small home nursery where the plants were propagated from cuttings and have been distributed locally, suggesting the virus is very likely to have a limited spread throughout the Auckland area. All three FiVB infected samples were also positive for FMV. However, the association of FiVB with FMD symptoms is unknown. FiVB was first identified from a latex sample exuded from a fig tree collected from Japan (Park et al. 2021) and is the only report of FiVB in the world to date. Although an identical sequence from Argentina, named fig closterovirus 1, was submitted to GenBank, the origin of this isolate is not known. To our knowledge, this is the first report of FiVB in New Zealand.

First report of Grapevine Algerian latent virus in carnation in New Zealand.

Carnation (Dianthus caryophyllus) is a popular ornamental plant widely used as a cut flower and in landscaping. In New Zealand, several viruses are known to infect plants of the genus Dianthus: arabis mosaic virus, carnation etched ring virus (CERV), carnation latent virus, carnation mottle virus, carnation necrotic fleck virus, carnation ringspot virus, carnation vein mottle virus and cucumber mosaic virus (Veerakone et al. 2015). In October 2020, a carnation sample with leaf chlorotic spots and distortion from a home garden in Auckland, New Zealand was submitted to the Plant Health and Environment Laboratory (PHEL) for virus testing. Leaf tissue of the sample was mechanically inoculated onto a range of herbaceous species using the method described in Tang et al. (2013). Chenopodium amaranticolor and C. quinoa plants developed local necrotic pinpoint spots while Nicotiana benthamiana, N. clevelandii, and N. occidentalis plants exhibited systemic leaf mosaic symptoms 7 days post-inoculation. The carnation plant and all five symptomatic indicator species tested positive for tombusviruses using an in-house designed generic RT-qPCR (available on request). Direct sequencing of the ~140 bp PCR product revealed the presence of grapevine Algerian latent virus (GALV). To further characterise the detected sequence, forward (5'-GTAGCGATGTATTGGGATAAGGA-3') and reverse (5'-TGCCGACACCCCGAAAGGT-3') primers were designed based on an alignment of the conserved region in the coat protein (CP) of 19 GALV isolates deposited in GenBank. Products of the expected size of 406 bp were amplified from all infected plants and their sequences found to be identical (GenBank accession No. OM891837). BLAST searches showed that the CP region of the sequence shared 97.0% (nucleotide) and 97.8% (amino acid) identity to the type isolate of GALV (GenBank accession no. NC_011535). GALV was first reported in Italy from a symptomless Algerian grapevine (Vitis vinifera) (Gallitelli et al., 1989), and is the only report of GALV in Vitis in the world. Since then, GALV has been reported in Germany, the Netherlands and Japan in several ornamental plant species including Alstroemeria sp. (Tomitaka et al., 2016), Gypsophila paniculata, Limonium sinuatum (Koenig et al., 2004, Fujinaga et al., 2009) and Solanum mammosum (Ohki et al., 2006). These infected ornamental host plants were reported to show various types of foliar symptoms, including chlorotic leaf spots. The GALV-infected carnation plant in this study was tested by PCR for all viruses that are known to infect D. caryophyllus in New Zealand, and CERV was identified. It is therefore unclear if the observed symptoms were induced by either GALV or CERV, or if they were the results of a synergistic interaction between GALV and CERV. Samples from a further 11 plants, comprised of nine symptomatic Dianthus spp. and two asymptomatic Alstroemeria spp. were collected from the same address and tested individually using the GALV-specific RT-PCR. GALV (along with CERV) was detected from all Dianthus plants while the Alstroemeria samples were negative. To our knowledge, this is the first report of GALV in New Zealand, and the first report in the host Dianthus in the world. Given the GALV-infected carnation plants were purchased from a local garden centre between 2007-2020, and plants from this garden centre have been widely distributed over this period of time to various customers, the virus is very likely to have spread throughout the country.

Partial biological and molecular characterization of a novel citrivirus from Nandina domestica.

We report the complete genome sequence of a novel virus isolated from Nandina domestica 'Firepower' in Auckland, New Zealand. It was mechanically transmitted to Nicotiana species, although all of these infections were symptomless. The complete genome of the new virus is 8892 nucleotides (nt) long, excluding the 3' poly(A) tail, contains three open reading frames (ORF), and is most closely related to citrus leaf blotch virus (CLBV) Actinidia isolate (CLBV-Act; 72% nt sequence identity), a member of the genus Citrivirus. Replicase and coat proteins, encoded by genome ORFs 1 and 3 respectively, shared 81-83% and 76-79% amino acid (aa) sequence identity, respectively, with CLBV-Act. Computer-based analysis suggests that this novel virus is the result of recombination between CLBV-Act and an unknown virus, highlighting the importance of this phenomenon for betaflexivirus evolution.

The complete nucleotide sequence and genome organisation of a novel member of the family Betaflexiviridae from Actinidia chinensis.

We report the complete genome sequence of a novel virus, tentatively named "actinidia seed-borne latent virus" (ASbLV), isolated from Actinidia chinensis in Auckland, New Zealand. The complete genome of ASbLV is 8,192 nucleotides long, excluding the 3' poly(A) tail, contains four open reading frames, and is most closely related to Caucasus prunus virus (56% nucleotide sequence identity), a member of the genus Prunevirus. Based on the demarcation criteria of the family Betaflexiviridae, ASbLV is a new member of the genus Prunevirus.

Opium poppy mosaic virus, a new umbravirus isolated from Papaver somniferum in New Zealand.

A novel virus, tentatively named "opium poppy mosaic virus" (OPMV), was isolated from Papaver somniferum (opium poppy) with leaf mosaic and mottling symptoms in Auckland, New Zealand, in 2006. The virus was mechanically transmitted to herbaceous plants of several species, in which it induced local and/or systemic symptoms. No virus particles were observed by electron microscopy in the diseased P. somniferum or any of the symptomatic herbaceous plants. The complete genomic sequence of 4230 nucleotides contains four open reading frames (ORF) and is most closely related (59.3 %) to tobacco bushy top virus, a member of the genus Umbravirus. These data suggest that OPMV is a new umbravirus.