Detection of Sweet Potato Viruses in Western Kenya, Development of a Multiplex PCR Technique for Detection of Major Viruses and Evaluation of Medicinal Plants for Antifungal and Antibacterial Activities against the Crop Pathogens

Abstract

Sweet potato is an important food crop worldwide since it is drought tolerant and acts as a famine relief crop. However, its production is limited by viral, fungal and bacterial infections. Lack of rapid and sensitive techniques for detection of infections inhibits their control. Use of synthetic chemicals to manage microbial infections cm.is!s is discouraged since they ar non-biodegradable. The aim of this study was to identify viruses infecting sweet potato in western Kenya, develop a multiplex PCR protocol for detection of major viruses of the crop, and evaluate efficacy of medicinal plants for antimicrobial activity against the crop pathogens. Symptomatic sweet potato vines obtained from farmers' fields were testes for ten viruses using NCM-ELISA. The mPCR protocol for detection of SPFMV and SPCSV was optimized through variation of test parameters under standard PCR conditions. Extracts from Warburgia ugandensis, Elaeodendron schweinfurthianum and Terminalia brownii, which are traditionally used to manage microbial infections, were fractionated using chromatographic methods to obtain pure compounds. Structures of isolates were determined using spectroscopic and physical methods. Extracts and isolates were tested for antimicrobial activity against Alternaria spp, Aspergillus niger, Fusarium oxysporum, F. solanum, Rhizopus stolonifer (fungi), Ralstonia solanacearum and Streptomyces ipomoeae (bacteria). Five viruses namely SPFMV, SPCSV, SPMMV, SPCFV and CMV were detected. Eighty nine percent of samples had viral infection with over 80% showing multiple infectious. Occurrence of CMV in Kenya was recorded for the first time. An mPCR protocol was successfully developed that detects SPFMV and SPCSV. Phytochemical studies afforded 26 compounds including 7uacetylugandensolide that was isolated from W Ugandensis for the first time. All the methanol, ethyl acetate and n-hexane extracts of the three plants were active against one or more of the test organisms. Methanol extract of W ugandensis exhibited significantly higher activity (24.50 mrn) than positive controls against R. stolonifer. Eighteen out of the twenty six compounds isolated exhibited antimicrobial activity against one or more of the test pathogens at concentrations :S200 ug/rnl. Polygodial, warbuganal, mukaadial, ugandensidial, ugandensolide, deacetoxyugandensolide and muzigadial, exhibited low MIC (MIC :S 50 ug/rnl) against one or more pathogens that was comparable to those exhibited by standard drugs. This study provided a quantitative assessment of viruses infecting sweet potato in western Kenya and a simplified mPCR protocol for routine rapid detection of SPFMV and SPCS V. Scientific proof for the efficacy of extracts of W ugandensis, E. schweifurthianum and T. brownii as antimicrobial agents against sweet potato pathogens and perhaps other crop pat~ogens was also provided.