Glutamate dehydrogenase 2 is required for virulence by facilitating fungal growth in the host hemocoel

Metarhizium robertsii, a fungal pathogen employed in pest biocontrol, can alkalize the surrounding environment, although the biological implications remain unclear. Here, we found that M. robertsii glutamate dehydrogenase 2 (MrGDH2) was responsible for alkalization during fungal growth on insect wings or media containing cuticle powder. Loss of MrGDH2 function resulted in significantly reduced virulence during both cuticle-passing and cuticle-bypassing infections but did not affect appressorium formation or cuticle penetration. Deletion of MrGDH2 failed to alkalize amino acid-containing media under carbon deprivation, leading to impaired mycelia growth and conidiation. Hemolymph carbohydrates were decreased during M. robertsii infection, and the ΔMrgdh2 mutant exhibited delayed fungal growth and impaired alkalization in hemolymph cultures. Notably, expression of PacC, a pH-responsive transcription factor critical for virulence, was downregulated in hyphal bodies of the ΔMrgdh2 mutant. Contrary to the established models in plant and human fungal pathogens, we demonstrate that Gdh2 activity is dispensable for appressorium formation but essential for fungal colonization in insect hemocoel during M. robertsii infection.