Occurrence, identification and characterization of viruses infecting passion fruit in Uganda

Horticulture is one of the fastest growing sub sectors in Uganda with an estimated annual growth rate of about 20%. Although its contribution to the economy has never been quantified, the role of horticulture in creating employment and generating foreign exchange earnings are known. In addition, its potential to alleviate poverty both in rural and urban regions has been noted. Promotion of horticultural production, as such, directly contributes to the Government of Uganda’s Poverty Eradication Action Plan and the Plan for Modernization of Agriculture. In the mid 1990’s, the diversification of export crops promoted was passion fruit. Against the backdrop of falling prices for traditional export crops such as coffee and cotton, passion fruit was seen as an alternate export for which high foreign exchange earnings could be attained to boost economic growth and enhance rural livelihood. This expectation was initially met, with export revenue steadily increasing from $0.04m in 1995 to $0.09m in 1998. Production then took a downward trend and initiated widespread enterprise abandonment across the country. Preliminary investigations into contributory factors as prominent. Among the biological factors, virus infection emerged as the single most limiting factor to productivity. Causing 40% yield loss and up to 100% crop loss in some parts of the country. However, disease distribution was not well described. Many interventions were initiated by both the National Agricultural Research Organization (NARO) and private partners but these had limited impact. The interventions included pathogen identification, which confirmed viral infection in Central Uganda and gave a preliminary identification of the prevalent causal agent as a potyvirus similar to Passion fruit woodiness virus (PWV). With data paucity as the instigating factor, this research was initiated to supplement the information available and facilitate on-going disease management and crop improvement effrots at the Horticultural Research Program of the National Crops Resources Research Institute (NaCRRI) and in the decentralized districts, with the aim of revitalizing the passion fruit industry in Uganda. Surveys were conducted in 15 major passion fruit growing districts during 2003 and 2995. Viral disease indicators were assembled and evaluated using geographical information systems software to generate spatial patterns. These patterns revealed high incidence and severity in central Uganda. an area once considered the heart of commercial passion fruit production. The districts of Mukono and Masaka in particular recorded 67% and 61% incidence, respectively. Masaka district also scored the highest mean disease severity 3.75 (Scale of 1-5). The eastern and western highlands of Uganda, regarded as Centre of diversity for purple passion fruit, were relatively disease-free and recorded average incidence levels between 0% and 2% in the districts of Sironko, Mbala, and Kabarole. In addition, Sironko district had the lowest mean severity score of 1. With the aid of electron microscopy, the presence of flexuos rods, typical of potyviruses, was confirmed in infected plant tissue. In addition, about 45% of 161 infected tissue samples tested with polyclonal antiserum for two potyvirus species in enzymes-linked immunosorbent assays (ELISA), namely, Cowpea aphid borne mosaic virus (CABMV) and passion fruit ringspot virus (PFRSV), over a two year period yielded positive responses. Immunoblots were also positive for (PFRSV), with a few variant isolates reacting with polyclonal antibodies for CABMV. A single coat protein sub unit of a molecular weight 33kDa was detected within the range for CABMV, PFRSV and PW. Further analysis of this protein unit yielded a derivative molecular weight of 31 kDa, only compared to CABMV isolates. Biological assays on 24 indicator plnat species in Chenopodium, Datura, Lycopersicon, Phaseolus, Nicotiana and Vigna genera were neither typical of CABMV nor PFRSV though variation in host range and infection also occurred among virus isolates from Uganda, indicating possible occurrence of strains. Still, the genetic similarity values among the potyvirus isolates ranged from 87 to 100% for derived amino acid sequences over the complete coat protein gene. These values are above the threshold of 85% approved by the International Committee for Taxonomy of Viruses as the benchmark for species demarcation. The Ugandan virus isolates recorded lower similarity values with four potyvirus species that showed closest similarity in BLAST searches, including Ben common mosaic virus (72-73%), cowpea aphid borne mosaic virus (72-76%). Of these species, the closest virus was the South African Passiflora strain of CABMV at 76%. The commercial diagnostic tools used for detection of the potyvirus infecting Ugandan passion fruit were not conclusive enough to discriminate this virus from CABMV at the species levels. Erratic results were observed in ELISA, while molecular primers available for detection of CABMV and related potyvirues such as PWV often gave variant banding patterns depending on the region of the virus genome used in primer development. As part of this study, molecular primers were thus designed for the Ugandan passion fruit potyvirus and evaluated with five isolates of the species. Preliminary results offer promise for detection of the virus both at species and sub-group level. One virus species was detected infecting passion fruit in Uganda, with a largely uniform molecular profile. The species was different from other viruses that cause similar disease symptoms on passion fruit, including CABMV, EAPV, and PWV. The name Ugandan Passiflora potyvirus is proposed for the species. On the basis of biological variation within the species, the occurrence of viral strains is further advanced. Nevertheless, further research is necessary to study the population genetics of the species. The molecular primers and protocol developed will also have to be more widely tested. Development of a simpler and cheaper bio-assay for virus detection would also go a long way in facilitating pathogen monitoring and disease management in mother gardens within the decentralized districts of Uganda.