RESUMO
A rapid and sensitive loop-mediated isothermal amplification (LAMP) method was developed for the specific detection of Pseudocercospora eumusae. The LAMP primers designed based on the specific SCAR (Sequence Characterized Amplified Region) marker sequence of P. eumusae proved highly specific to P. eumusae and there was no cross reactivity with closely related Pseudocercospora spp. (P. fijiensis and P. musicola) and 17 other leaf spot causing fungal pathogens of banana. The developed LAMP method exhibited greater sensitivity as the minimum detectable concentration of P. eumusae genomic DNA was 10 Æ¿g/µl which was 100 times lower than that of conventional PCR (1 ng/µl).This method also detected the target pathogen from crude DNA of the mycelium and single leaf spot tissues which eliminates laborious purification steps in DNA isolation and requires less operational time. To the best of our knowledge, this is the first report on the development of this LAMP method to specifically detect and diagnose P. eumusae pathogen from pure cultures and leaf spot tissues.
RESUMO
To find out the genetic diversity of Indian Foc isolates of banana, a total of 107 isolates of Fusarium which includes 98 Foc isolates obtained from different banana growing regions of India and seven Foc isolates belong to all known VCGs obtained from Australia and two non-pathogenic Fusarium oxysporum (npFo) isolates were subjected to ISSR analysis. In the initial screening of ISSR primers, out of 34, 10 primers which generated more polymorphic bands were selected for further analysis. The Phylogenetic analysis carried out based on the fingerprints obtained through ISSR analysis indicated the presence of wide genetic diversity among the Foc isolates of India and also its polyphyletic nature. Totally, seven different clusters were obtained and these clusters differentiated the Foc isolates of India based on the races/VCGs. Besides, the cluster analysis clearly distinguished the freshly emerged Foc strain obtained from cv. Grand Naine (Cavendish-AAA) and Poovan (Mysore-AAB) from the other Foc isolates. The non-pathogenic F. oxysporum isolates which have been included for comparison purpose also clustered separately. All these above said findings indicates for the first time the discriminatory power of ISSR to clearly distinguish and separate the Foc isolates according to its race/VCGs and also its virulence. This study would be useful not only to design and develop effective management strategies but also useful for quarantine purposes.
