EPIDEMIOLOGY AND CHARACTERISATION OF Groundnut Ringspot Virus (GRSV) INFECTING GROUNDNUTS AND OTHER PLANTS IN WESTERN KENYA
Abstract
Groundnut (Arachis hypogaea L.) is an annual oilseed legume crop grown by small holder farmers in Kenya for its economic and nutritive value. However, its yield has declined upto 680 kg ha-1 than its genetic potential of 1690 kg ha-1 attributed to abiotic and biotic stressors. Viruses are among biotic stressors for yield reduction globally. These include; Groundnut ringspot virus (GRSV), Tomato spotted wilt virus (TSWV), among others. GRSV was reported in South Africa, Ghana, Brazil and USA infecting groundnuts, soybeans and others. GRSV and TSWV have similar biological symptoms but differentiated using serological tests. Typical Symptoms for GRSV appears on groundnuts and other plants in western Kenya but no report had been documented on the occurrence of the virus nor its management strategies Kenya. The general objective of this study was to determine the occurrence, distribution and characterisation of GRSV on groundnuts and other host plants in western Kenya. Survey on prevalence of GRSV, was conducted in short and long rain seasons of the years 2019 and 2020 in western Kenya. Simple random sampling (SRS) used in selecting farms visited in groundnut growing regions and disease incidence/ severity recorded and data collected analyzed using post-hoc analysisANOVA. Serological analysis was done on samples collected using polyclonal and monoclonal antisera against GRSV and TSWV respectively. Field trials on the effect of intercropping other legumes with groundnuts on GRSV incidences were laid on a randomized complete block design (RCBD) and replicated three times. Viral incidence and severity recorded and symptomatic leaf samples collected for GRSV ELISA tests. Health tested seeds to GRSV of groundnut varieties and other plant species were planted in plastic pots of a mixture of sterilized loam, sand and organic manure at a ratio of 2:1:1 respectively in greenhouse to screen for their response and host range to GRSV and inoculated with GRSV inoculum. Plants symptomatic development observed at an interval of 5 days for 8 weeks and plant samples for each variety/species collected for GRSV ELISA Tests. Total RNA of Kenyan plant isolates extracted using CTAB and purified by DCC™-5 purification kit then amplified using target primers GRSVnR (5’-GCGGTCTACAGTGTTGCACTT3’)andGRSVnF(5’TCTTGTGCATCATCCATTGT-3’) using Rt-PCR at 614-bp fragment of the nucleocapsid gene of GRSV corresponding to the part of the nucleocapsid (N) gene. The RT-PCR product taken for Sanger sequencing. Sequence readings trimmed using Bio-edit software and phylogenetic analysis done in MEGA-X. New primers from GRSV sequences of western Kenya was designed using primer3plus software, synthesized and validated using PCR tests. GRSV occurs in surveyed regions with variant incidence; Chwele having the highest incidence (45.04 %) while Kapkateny having the lowest incidence (17.75 %) with significant difference of (P < 0.05). Groundnuts planted in pure stand had lowest disease incidence (4%) while intercropped groundnuts had the highest (28%). Screened groundnuts showed Homabay variety being more susceptible with incidence of 31 %, followed by ICGV-9991 with incidence of 28 %. SM99568 variety was tolerant to the virus. Varieties ICGV-90704, ICGV-99048 and ICGV-99019 were resistant to the virus.Screened plants; Pigeon peas, Bambara nut, peas, Chenopodium album, Galinsoga parviflora among others, revealed being as host range for the virus. Kenyan GRSV isolates clustered with USA, Ghanaian and South African isolates in GenBank. One of developed primers formed clear bands in a PCR tests with positive samples of western Kenya. GRSV occurs in surveyed counties of western Kenya, which should be a big concern to all stakeholders. Introgression of resistant genes into local groundnuts to gain resistance to the virus with urgency. Farmers should avoid intercropping groundnuts with alternative hosts to reduce transmission of the virus.