Evaluation of in vitro protocols for the elimination of banana streak virus in tissue culture produced banana plantlets

Abstract

Tissue culture (TC) propagation of banana crop has been used in many laboratories for mass production of planting materials which are free from plant pathogens. The Jomo Kenyatta University of Agriculture and Technology (JKUAT) commercial banana TC laboratory which currently supplies banana planting material all over the country, and parts of East Africa, utilizes the TC technique for the mass propagation of banana plantlets for sale to farmers. However, the TC process does not eliminate banana viruses. Banana streak virus (BSV) reduces banana yield from 6% to 15% and is also the most prevalent banana virus in Kenya. This poses a big danger since the plantlets in supply could be BSV infected. Therefore, this research was done to evaluate in vitro protocols for elimination of BSV with the aim of producing BSV-free TC banana planting materials for farmers. Asymptomatic and symptomatic leaf and shoot samples were randomly collected from Kenya Agricultural Research Institute (KARI) Kisii, KARI Thika and JKUAT banana mother orchards that mainly supply JKUAT commercial TC laboratory with initiation materials. The leaf samples were screened for presence of BSV using polymerase chain reaction (PCR) technique, while the suckers were used for initiation materials which were taken through TC process up to the second subculture. The second subcultered plantlets were used in evaluation of the in vitro protocols for elimination of BSV. Three viral elimination techniques namely chemotherapy (ribavirin, and salicylic acid), thermotherapy and meristem tip culture were evaluated in attempts to eliminate BSV. All the costs incurred in the production of BSV-free plantlets were determined using the year 2012 market price in Kenya xiii shillings to determine which of the virus elimination methods was cost effective. Of the 30 samples collected from the three mother orchards, 15 asymptomatic and 15 symptomatic, 21 were detected with BSV using PCR. In chemotherapy, 0mg/l (control), 10 - 40 mg/l were used and they gave 0 - 90% virus elimination using ribavirin and 0 - 90% virus elimination using salicylic acid respectively. In thermotherapy, 27°C (control), 32 - 38°C all for 10 days were used which resulted in between 0% and 90% virus elimination. Meristem tip culture using tips ranging between 1, and 5mm (control) resulted in 0%, and 70% virus elimination. Best BSV elimination and explant regeneration rates were observed at 20mg/l of salicylic acid and at 36°C. However, thermotherapy can be time consuming and laborious in laboratories which do not have automated systems. Production of one BSV– explant using the four virus elimination methods was Ksh 125.5, 125.6, 127.9, and 130.4, using salicylic acid, ribavirin, meristem tip culture and thermotherapy, respectively. Chemotherapy using salicylic acid at 20mg/l was considered the most effective since it was cheaper, in terms of implementation, less laborious, and it had the highest regeneration and virus elimination rates. Therefore all infected initiation materials to be used in commercial laboratories should subjected to chemotherapy using salicylic acid (20mg/l) to ensure that all banana planting materials released for sale are BSVfree.