Biological control of plant-parasitic nematodes by the fungus Clonostachys rosea

Plant diseases caused by plant-parasitic nematodes are serious constraints to sustainable crop production due to high yield losses, the persistent nature of these nematodes and a lack of efficient control methods. Biological control is a promising approach to reduce plant diseases caused by nematodes. This study investigated the effect of the fungus Clonostachys rosea strain IK726 on nematode populations in a naturally nematode-infested soil planted with wheat in a climate chamber under controlled conditions. Populations of plant-parasitic nematodes extracted from soil and roots were 40 to 73% lower in soils when C. rosea was applied than in untreated soils, whereas non-parasitic nematodes were unaffected. Soil inoculation with C. rosea increased the shoot weight and shoot length of wheat plants by 20 and 24%, respectively. Light microscopy of in vitro C. rosea–nematode interactions did not reveal evidence of direct parasitism; however, culture filtrates of C. rosea grown in potato dextrose broth (PDB) exhibited toxicity towards nematodes and immobilized nematodes. A genome-wide analysis of protease genes showed that C. rosea contains more protease genes than other studied biocontrol fungi. A computational analysis of gene family evolution revealed a high gene copy number of serine protease subfamilies S8A, S9X and S33 in C. rosea, suggesting the involvement of these proteases in biotic interactions. A genome-wide association analysis of 53 strains of C. rosea further identified 279 single-nucleotide polymorphism markers that were significantly associated with the in vitro antagonism trait against plant-parasitic nematodes. Two non-ribosomal peptide synthetase (NRPS) genes (nps4 and nps5) were identified in genomic regions associated with nematicidal activity whereas nps1 was included based on previous published reports of functional studies. Gene deletion strains of nps1, nps4 and nps5 were generated and showed increased growth and conidiation rates. Culture filtrates from C. rosea Δnps1, Δnps4 and Δnps5 strains exhibited reduced nematicidal activity and immobilized lower numbers of nematodes compared with the wild type after 24 h of incubation. However, NRPS deletion strains still possessed some nematicidal activity compared with the PDB control treatment, which may be due to the presence of additional nematicidal compounds or enzymes (e.g., serine proteases). Furthermore, Δnps1, Δnps4 and Δnps5 strains showed reduced biocontrol efficacy in a naturally nematode-infested soil in a pot experiment and failed to reduce populations of nematodes in soil or in roots of wheat as efficiently as the wild type strain. This study demonstrates that C. rosea can control plant-parasitic nematodes and improve the growth of plants at the same time. The antagonistic potential of C. rosea could be used to control plant-parasitic nematodes, which may contribute to reduced applications of chemicals as part of an integrated pest management programme