Endophytic Bacteria from Alstonia scholaris (L.) R. Br Latex as Novel Biocontrol Agents Against Phytopathogens

Latex-producing plants harbor unique microbial communities that may play important roles in host defense; however, their diversity and biocontrol potential remain largely unexplored. Characterizing these communities provides opportunities to identify novel microbial-derived antifungal agents for sustainable crop protection. Bacterial strains were isolated from the latex of Alstonia scholaris (L.) R. Br. and identified using 16S rRNA gene sequencing. Antifungal activity was evaluated against four phytopathogens: Fusarium graminearum, Colletotrichum musae, Colletotrichum gloeosporioides, and Glomerella cingulata. Bioassay-guided fractionation, size-exclusion chromatography, SDS-PAGE, and LC-MS/MS were used to characterize antifungal proteins. Nine bacterial strains were isolated, including eight Bacillus spp. and one Enterococcus faecalis. Among them, Bacillus sp. AsL-2 exhibited the strongest broad-spectrum antifungal activity, inhibiting fungal growth by up to 80%. The antifungal activity of its crude extract remained stable over a wide temperature range. Further characterization identified a novel endo-β-1,3-1,4-glucanase enzyme (~23 kDa) as the major antifungal protein. This study reveals A. scholaris latex as an underexplored microbial niche and identifies Bacillus sp. AsL-2, affiliated with the B. velezensis–B. amyloliquefaciens species complex, as a promising biocontrol candidate. The identified antifungal enzyme represents a potential natural alternative to synthetic fungicides for sustainable agricultural disease management.