Decoding interactions of the nematophagous fungus, pochonia chlamydosporia, with black pepper

Abstract

Biological control agents like Pochonia chlamydosporia are gaining prominence as eco-friendly alternatives to chemical pesticides in sustainable agriculture. This nematophagous fungus not only parasitizes nematode eggs but also promotes plant growth, making it valuable for crops like black pepper. This study investigates the biocontrol potential of P. chlamydosporia through its secondary metabolites, assessing their antifungal and nematicidal properties, while exploring the fungus’s ability to enhance plant growth and defence. The nematophagous fungus Pochonia chlamydosporia was evaluated for its plant growth-promoting properties in black pepper (Piper nigrum L.). The fungal isolate, P. chlamydosporia strain IISR-MTCC5412, was tested for its ability to produce plant growth-promoting substances, including indole-3-acetic acid (IAA), ammonia, and its capacity to solubilize inorganic phosphate and zinc. Results showed that the strain produced IAA (9.8 μg/ml), ammonia, and solubilised both phosphate (1.3 ± 0.07 response units) and zinc (1.1 ± 0.01 response units). The fungus also exhibited indirect plant growth-promoting effects, such as siderophore production (2 ± 0.04 response units) and the synthesis of extracellular enzymes, including α-amylases, cellulases, and pectinases. Pot culture studies demonstrated that P. chlamydosporia significantly enhanced shoot and root growth, biomass, and nutrient uptake (nitrogen, phosphorus, potassium, calcium, magnesium, copper, manganese, iron and zinc) in black pepper plants. These findings underscore the agronomic potential of P. chlamydosporia strain IISR-MTCC5412 as both a biocontrol agent and a plant growth promoter. Secondary metabolites of P. chlamydosporia were extracted using ethyl acetate, and their activity against pathogens and plant parasitic nematodes was tested. The crude extract showed strong antifungal activity against pathogens such as Pythium myriotylum (99.54%) and Phytophthora spp. (99.99%) while the lowest percentage inhibition was observed with Macrophomina spp. (40%). The ethyl acetate extract also exhibited nematicidal activity against Radopholus similis, with the highest mortality observed at the crude extract concentration. High-resolution UPLC- (ESI)- QToF-MS analysis identified several bioactive compounds, including bisevertinol, enniatin I, and enniatin H. To identify potential lead compounds, an in-silico virtual screening, including molecular docking and binding free energy computation, was performed. The results indicated that three compounds - bisevertinol, enniatin I, and enniatin H - could serve as lead molecules for the development of nematicidal chemicals against plant parasitic nematodes. Furthermore, transcriptome analysis was conducted to study the interaction between P. chlamydosporia and black pepper roots at 14 and 28 days post-inoculation (dpi). Using next generation sequencing (NGS) technologies (HiSeq 500 platforms), 2,03,092 transcripts and 65,354 differentially expressed genes (DEG) were identified, revealing significant changes in gene expression related to plant growth promotion and induced systemic resistance (ISR). Genes involved in phytohormone biosynthesis (auxin, ethylene, cytokinin, abscisic acid, and gibberellin), nutrient uptake (nitrogen, phosphorous, potassium, calcium, iron, boron, and zinc) and plant defence-related genes (including pathogenesis-related (PR) proteins, lipid-transfer protein (DIR1), endochitinase, germin-like proteins, late embryogenesis abundant (LEA) proteins, LURP1, thaumatin and Major Latex-like Proteins (MLPs)) were upregulated in black pepper plants colonised by P. chlamydosporia. Functional gene ontology (GO) analysis showed enrichment in stress resistance and ISR signalling pathways, indicating that the fungus enhances the plant’s ability to withstand biotic and abiotic stress. KEGG enrichment analysis also highlighted upregulation of pathways related to plant hormone signal transduction. This transcriptomic data provides molecular evidence that P. chlamydosporia promotes plant growth and strengthens defence mechanisms of black pepper, offering new insights into its dual role as a biocontrol agent and endophyte. This study confirms P. chlamydosporia as a promising biocontrol agent with dual functions: suppressing plant pathogens and promoting growth in black pepper. Key bioactive compounds, such as bisevertinol, enniatin I, and enniatin H, were identified with potential for nematicidal use. Transcriptomic analysis provided molecular insights into how the fungus boosts plant growth and defence, emphasising its potential in sustainable pest management and crop resilience strategies.