Elucidation of Ralstonia solanacearum Phylotypes causing Bacterial Wilt in Solanum lycopersicum (Cobra tomatoes) through Modified Agar Medium (MAM) and Multiplex-PCR.
Author(s)
Forkwa Etienne Yong , Eneke Esoeyang Tambe Bechem , Egbe Enow Andrew ,
Download Full PDF Pages: 15-34 | Views: 23 | Downloads: 8 | DOI: 10.5281/zenodo.14795035
Abstract
Tomatoes are major staples in global diets and are frequently threatened by the virulent soil-borne pathogen, Ralstonia solanacearum, which causes devastating bacterial wilt and poses a significant risk to food security. This study was designed to elucidate the Ralstonia solanacearum phylotypes associated with bacterial wilt in Solanum lycopersicum in Buea, Cameroon. One hundred and fifty symptomatic tomato plants were collected from five tomatoes producing sites in Buea (Bwitingi, Ewonda, Great Soppo, Molyko, and Muea). A modified agar medium (MAM) was used to minimize the presence of contaminating pathogens, followed by nutrient agar to further purify the 45 isolates. Subsequently, the isolates underwent biochemical tests (Kliger iron agar and Gram staining), pathogenicity tests, genomic DNA extraction, and Polymerase Chain Reaction (Multiplex PCR) using primers that targeted 16S and 23S rRNA. The symptoms observed included wilted and yellow leaves, discoloured stems, and adventitious roots. Spearman correlations showed the strongest associations between discoloured xylem and wilted leaves (r = 0.43) and the weakest between discoloured xylem and yellow leaves (r = 0.13). Kliger iron agar, Gram stain, and pathogenicity tests confirmed the presence of the pathogen, but no significant differences were found amongst the 45 isolates (P = 0.05), except for stem streaming (H= 11.82, P = 0.019). Genomic DNA was successfully extracted from all 45 isolates, and effectively amplified during PCR, yielding distinct bands of 372, 144, 213, and 91 base pairs, indicating the presence of phylotypes II, I, IV, and III, respectively, in the isolates. These findings provide valuable insights into the pathogen's diversity and identity in Cameroon, providing clues to stakeholders in the management of bacterial wilt disease in the tomato secto
Keywords
Ralstonia solanacearum, phylotypes, Modified agar medium, nutrient agar, Primers, PCR, and Gel electrophoresis
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