Evaluation of Meristem Tip Culture, Chemotherapy and Thermotherapy on the Reduction of Cassava Brown Streak Virus Ininfected Cassava (Guzo Variety) Cuttings

Abstract

Cassava brown streak disease (CBSD) is an economically important disease of cassava (Manihot esculenta crantz) caused by Cassava brown streak virus (CBSV) which was the target in this study and Ugandan cassava brown streak virus (UCBSV). This disease is spreading fast in several countries in the East and Central Africa. The general objective of this research was in vitro production of CBSV-free cassava from infected cassava. Specifically this research aimed at (i) evaluating the effect of meristem tip sizes (ii) determine the effects of varying concentration levels of selected chemicals (iii) determine the efficacy of combining thermotherapy with meristem tip culture and chemotherapy on the reduction of cassava brown streak virus from infected cassava. CBSV infected cuttings of Guzo variety collected from the Coast province of Kenya established and maintained in a greenhouse at the Plant Quarantine and biosecurity Station (PQBS) in Kenya Plant Health Inspectorate Service's (KEPHIS) in Muguga were used as test plants. Leaves were sampled from eighteen cassava plants of Guzo variety and virus indexing was done using (RT-PCR) with virus specific primers and those that tested positive for CBSV were used as initiation materials. From the in vitro plantlets established the second subcultured materials were subjected to the virus reduction procedures. In vitro meristems (0.5mm, 1mm, 2mm and 10mm (control) were obtained and cultured in modified Murashige and Skoog (MS) media. For chemotherapy, nodes were cultured in MS media supplemented with antivirals (ribavirin and salicylic acid) at 10mg/l, 20mg/l, 30mg/l and control of 0mg/l. In the combination treatments, plants were subjected to thermotherapy at 38ºC then meristem tips (1.0mm) were excised and heat treated plants at 38ºC were subjected to ribavirin treatments at (10mg/l, 20mg/l and 30mg/l). Data collected was analysed using genstat. The regeneration of plants established from 0.5mm was 63% while those from 2mm was 88% with the proportion of virus-free plants decreasing with increase in meristem tip size. Meristem tip size of 0.5mm resulted in the highest % (88.23) of virus-free obtained. In chemotherapy, survival of shoots was observed to decrease with increase in the antiviral concentrations. Ribavirin 10mg/l recorded the highest value of survivals compared to the other treatments. On the other hand salicylic acid exhibited the least survivals compared to ribavirin. The number of virus-free plants obtained was observed to increase with increase in concentration for both chemicals. At 30 mg/l of ribavirin and salicylic, 88.8% and 100% of virus-free plantlets were produced respectively. Thermotherapy (38°C) combined with meristem tips (1mm) resulted in 68% of regenerated plants with 84% being virus free. In vitro plants that had been thermo treated and then subjected to chemotherapy did not give the expected results since complete death of the plants was recorded. Based on efficiency of virus reduction in terms of CBSV-free plants obtained and the number of plants surviving after the treatment, thermotherapy at (38°C) combined with meristem tip culture can be used for production of CBSV-free cassava. Farmers are therefore encouraged to use in vitro raised materials that have been adequately diagnosed free from CBSV. This will ultimately reduce the risk of spreading CBSV to uninfected cassava fields. There is need for studies to be done that will show effects of these cleaning methods on the reduction of Ugandan cassava brown streak virus (UCBSV) from infected guzo plants and evaluate the effect of chemotherapy by subjecting virus infected cassava plants to lower antiviral concentrations while increasing duration of treatment to evaluate the effect of CBSV reduction from infected cassava.