Cytochrome b Gene-Based Assay for Monitoring the Resistance of Colletotrichum spp. to Pyraclostrobin

Resistance to pyraclostrobin due to a single nucleotide polymorphism at 143rd amino acid position on the cy-tochrome b gene has been a major source of concern in red pepper field infected by anthracnose in Korea. Therefore, this study investigated the response of 24 isolates of C. acutatum and C. gloeosporioides isolated from anthracnose infected red pepper fruits using agar dilution method and other molecular techniques such as cytochrome b gene sequencing, polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP), and allele-specific polymerase chain re-action (PCR). The result showed that four isolates were resistant to pyraclostrobin on agar dilution method and possessed GCT (alanine) codon at 143rd amino acid position, whereas the sensitive isolates possessed GGT (glycine). Furthermore, this study illustrated the differ-ence in the cytochrome b gene structure of C. acutatum and C. gloeosporioides. The use of cDNA in this study suggested that the primer Cacytb-P2 can amplify the cytochrome b gene of both C. acutatum and C. gloeo-sporioides despite the presence of various introns in the cytochrome b gene structure of C. gloeosporioides. The use of allele-specific PCR and PCR-RFLP pro-vided clear difference between the resistant and sensitive isolates. The application of molecular technique in the evaluation of the resistance status of anthracnose pathogen in red pepper provided rapid, reliable, and accurate results that can be helpful in the early adop-tion of fungicide-resistant management strategies for the strobilurins in the field.

Automated summary

This study investigated the response of C. acutatum and C. gloeosporioides isolates from anthracnose infected red pepper fruits to pyraclostrobin. Molecular techniques such as cytochrome b gene sequencing, PCR-RFLP, and allele-specific PCR were used. The results showed that four isolates were resistant to pyraclostrobin and possessed a specific mutation in the cytochrome b gene. The study also observed structural differences in the cytochrome b gene between the two species. The application of molecular techniques in evaluating the resistance status provided rapid and accurate results for fungicide-resistant management strategies.