First Report of Passion fruit woodiness virus Associated with Passion Fruit Woodiness Disease of Passion Fruit in Nigeria

Passion fruit (Passiflora edulis) is an important horticultural crop grown as a sole crop in north-central and southeastern regions of Nigeria and in homesteads and backyard gardens throughout the country. Passion fruit cultivation is gaining popularity with the proliferation of juice making industries in Nigeria. In 2016, passion fruit plants with virus symptoms such as systemic mosaic, mottling leaf distortion, upward curling, and puckering of lamina were observed in three farms in Oyo state. Fruits of symptomatic plants were misshapen with corky skin and knobby growth similar to the symptoms of passion fruit woodiness disease (Ochwo-Ssemakula et al. 2012). To confirm infectivity, leaf samples from symptomatic plants were used as a source for mechanical sap inoculation onto 21-day-old healthy passion fruit seedlings maintained in an insect-proof screenhouse. Newly emerging leaves of inoculated plants developed mild chlorotic mottle symptoms approximately 14 days after inoculation. Leaf samples from infected field samples as well as experimentally infected plants in direct antigen coating ELISA (Hobbs et al. 1987) reacted with the rabbit polyclonal antibodies to Bean common mosaic virus-black eye cowpea mosaic strain (BCMV-BlCM, genus Potyvirus; available at the International Institute of Tropical Agriculture). To further confirm the virus identity, total nucleic acid extracts were subjected to reverse-transcription polymerase chain reaction (RT-PCR) using a generic potyvirus primer pair (POTY CP-D-F 5′-TGGTGYATGANAAYGGNACNT-3′ and POTY-D-R 5′-CCCAGTCACGACTTTTTTTTTTTTTTT-3′) specific to the 3′ end of the coat protein and linked untranslated region, as detailed in Sudheera et al. (2014). Positive amplification of an expected ∼700-bp product was obtained from symptomatic plants only. The RT-PCR products were cloned in pGEM-T Easy vector (Promega), and two independent clones were sequenced in both orientations using plasmid-specific primers. The 668-bp nucleotide sequences of both clones of the Nigerian isolate were identical (GenBank accession no. KY284619), and the BLASTN searches in NCBI GenBank revealed the highest identity of 88%, and 94% similarity of the deduced amino acid (137 aa) sequence, with an isolate of Passion fruit woodiness virus (PWV) (FR694180) reported from Uganda (Ochwo-Ssemakula et al. 2012). The virus also showed serological cross-reaction with BCMV-BlCM antibody in ELISA. The maximum-likelihood phylogenetic tree constructed using MEGA6 placed the PWV Nigerian isolate on a separate subcluster compared with PWV isolates (FR694180 to FR694182) infecting passion fruit in East Africa, indicating that the Nigerian isolate could be a distinct strain of PWV. Passion fruit woodiness disease in Nigeria was first described in 1958 (Martini 1962); however, the identity of an associated aphid-transmitted flexuous rod-shaped virus named then as Passion fruit mosaic virus was not established. To our knowledge, this is the first report of PWV in Nigeria. PWV affects fruit quality and has high potential to reduce market value. Further surveys are necessary to assess prevalence, incidence, and diversity of PWV and to identify the principal aphid species transmitting PWV in Nigeria.