Divergent Immune Strategies of Colorado Potato Beetle Larvae against the Entomopathogenic Fungi Metarhizium robertsii and Beauveria bassiana. A Comparative Transcriptomic Analysis. Supplementary data.

Abstract
The entomopathogenic fungi Beauveria bassiana and Metarhizium robertsii are natural pathogens of the Colorado potato beetle (CPB) and hold promise for biocontrol. However, the insect’s immune responses to fungal infections remain poorly understood. In this study, we performed transcriptomic analyses of hemocytes and fat body tissue from CPB larvae topically infected with B. bassiana and M. robertsii at the stage of pathogen encapsulation. Quantitative PCR validation of gene expression, along with complementary physiological and biochemical assays, were also conducted. Reference gene annotations were expanded using both the classical bioinformatic tool InterProScan and the AI-based tool ProteInfer. For the first time, pathogen-specific differences in CPB tissue responses to fungal infections were revealed. Gene Ontology enrichment and expression profiles of immune-related genes indicated a stronger antifungal response to M. robertsii, whereas infection with B. bassiana was associated with enhanced protection against self-damage. Pathogenspecific responses were linked to the expression of genes encoding certain receptors, serine proteases, serpins, Toll signaling components, antimicrobial peptides, chitin deacetylases, chitin synthases, laccases, as well as to the production of phenoloxidases and reactive oxygen species. The most diverse and pathogen-specific gene expression changes occurred in hemocytes rather than in the fat body. We propose that the observed differences in immune responses are mainly driven by the fungal enzymatic machinery, secondary metabolites, and pigments. The differentially expressed genes identified in this study provide novel insights into insect–pathogen interactions and represent promising molecular targets for advancing both fundamental research on insect immunity and the applied development of innovative biocontrol strategies.
Supplementary tables:

Table S1. List of genes and primers for qPCR.Table S2. Extended gene annotation and the results of differential gene expression analysis in hemocytes and the fat body of Colorado potato beetle larvae in response to fungal infections.Table S3. Gene Ontology enrichment analysis of differentially expressed genes in hemocytes and the fat body of Colorado potato beetle larvae in response to fungal infections.Table S4. Differential expression of immune-related genes in hemocytes and the fat body of Colorado potato beetle larvae in response to fungal infections. Extended gene annotation is shown.