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Selection of reliable reference genes in Colletotrichum scovillei during different growth stages, host interactions, and plant extract treatment for qRT-PCR

Summary

This research identifies the best internal control genes to use when measuring gene activity in a fungus that causes pepper disease. Scientists tested eight different reference genes under various conditions like fungal growth and pepper infection, plus when treating the fungus with plant extracts. Different genes worked best for different situations: one gene was best during fungal growth, another during infection, and a third when treated with plant extracts. These findings will help future studies better understand how this disease-causing fungus works and develop better control methods.

Background

Colletotrichum scovillei, a member of the C. acutatum species complex, is a significant pathogen causing anthracnose in chili peppers across multiple countries. Quantitative real-time PCR (qRT-PCR) is a sensitive method for evaluating gene expression, but requires reliable internal control genes. Stable reference genes have not yet been identified for C. scovillei under various experimental conditions.

Objective

To identify and validate reliable reference genes in C. scovillei during different growth stages (conidia, conidial germination, mycelium growth), host interactions with chili peppers, and treatment with 0.5% DMSO and antifungal plant extracts. Eight candidate reference genes were selected and evaluated using multiple statistical approaches.

Results

CsPP2A demonstrated the best stability during growth stages, CsTUB showed superior stability during host interactions, and CsUCE was most stable following DMSO and plant extract treatment. No single reference gene was suitable under all combined conditions. The validation experiments confirmed the suitability of selected reference genes for respective experimental conditions.

Conclusion

This study provides the first identification of suitable reference genes for C. scovillei under different conditions, essential for accurate gene expression analysis. CsPP2A is recommended for growth stage studies, CsTUB for host interaction studies, and CsUCE for plant extract treatment studies. These findings will facilitate future molecular studies of C. scovillei pathogenesis and disease management strategies.
  • Published in:BMC Plant Biology,
  • Study Type:Experimental Research,
  • Source: 10.1186/s12870-025-06733-1, PMID: 40615801
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