Integration of pathogenicity, thermotolerance, and gene expression analyses reveals a superior native Metarhizium anisopliae isolate for the biological control of the mustard aphid, Lipaphis erysimi

Introduction The mustard aphid, Lipaphis erysimi , is a notorious pest of cruciferous crops worldwide, causing up to 100% yield losses through direct phloem depletion and the dissemination of plant viruses. Currently, management of this pest relies heavily on synthetic insecticides, leading to significant adverse effects on both the environment and human health. This study aimed to evaluate the pathogenicity, conidial production, thermotolerance, and molecular characteristics of indigenous Metarhizium anisopliae isolates for biological control of L. erysimi . Methods Ten M. anisopliae isolates (MaCMUIPL 2, 14, 26, 37, 40, 43, 47, 48, 54, and 57) were collected from various natural habitats and initially screened for pathogenicity against Tenebrio molitor and L. erysimi using a contact plate assay and conidial spraying method (1 × 10 6 conidia mL -1 ), respectively. The highly pathogenic strain MaCMUIPL 57 was subjected to virulence tests against different conidial concentrations. The most virulent strains were subjected to detailed phenotypic evaluations for conidial yield over 14 days and critical thermotolerance at 45 °C for up to 120 minutes of exposure. Molecular characterizations included sequencing the conserved virulence gene–glycosylphosphatidylinositol-anchored surface protein ( GAS1 ) and conducting reverse transcription quantitative PCR (RT-qPCR) to profile the expression of conidiation-associated gene ( cag8 ) and serine/threonine protein kinase ( slt2 ) gene. Results Among ten indigenous isolates, MaCMUIPL 57 exhibited the highest pathogenicity with rapid insecticidal action against L. erysimi , achieving complete mortality within five days at 1 × 10 9 conidia mL -1 and 80% mortality at 1 × 10 5 conidia mL -1 . Phenotypic characterization revealed exceptional sporulation capacity, with MaCMUIPL 57 producing the highest conidial yield (52.2 × 10 6 conidia mL -1 ) on the 14th day, and maintained germination rates of 76.5%, 25.7%, 12.2%, and 2.0% after 30, 60, 90, and 120 min of exposure to 45 °C, respectively. Molecular characterization confirmed the conserved virulence gene GAS1 with 100% sequence identity to reference entomopathogen homologues. Furthermore, RT-qPCR profiling showed significant upregulation of stress-adaptation genes in MaCMUIPL 57, where cag8 expression strongly correlated with conidial thermotolerance and slt2 expression mirrored enhanced vegetative sporulation. Discussion These findings demonstrate that strain MaCMUIPL 57 effectively couples rapid insect lethality with physiological resilience to provide a sustainable biocontrol strategy against L. erysimi .