Background
A nematophagous fungus, Esteya vermicola , is recorded as the first endoparasitic fungus of pine wood nematode (PWN), Bursaphelenchus xylophilus , in last century. E. vermicola exhibited high infectivity toward PWN in the laboratory conditions and conidia spraying of this fungus on Japanese red pine, Pinus densiflora , seedlings in the field protected the pine trees from pine wilt disease to some extent, indicating that it is a potential bio-control agent against PWN. Previous research had demonstrated that the living fungal mycelia of E. vermicola continuously produced certain volatile organic compounds (VOCs), which were responsible for the PWN attraction. However, identity of these VOCs remains unknown.
Methodology/Principal Findings
In this study, we report the identification of α-pinene, β-pinene, and camphor produced by living mycelia of E. vermicola , the same volatile compounds emitted from PWN host pine tree, as the major VOCs for PWN attraction using gas chromatography-mass spectrometry (GC-MS). In addition, we also confirmed the host deception behavior of E. vermicola to PWN by using synthetic VOCs in a straightforward laboratory bioassay.
Conclusions/Significance
This research result has demonstrated that the endoparasitic nematophagous fungus, E. vermicola , mimics the scent of PWN host pine tree to entice PWN for the nutrient. The identification of the attractive VOCs emitted from the fungus E. vermicola is of significance in better understanding parasitic mechanism of the fungus and the co-evolution in the two organisms and will aid management of the pine wilt disease.
Pine wilt disease (PWD), caused by the pine wood nematode (PWN,Bursaphelenchus xylophilus), is one of the most serious threats to pine forests worldwide. The fungusEsteya vermicola, with its lunate conidia capable of parasitizing the PWN, has shown promise as an efficient biological control agent against PWD. Solid-state fermentation (SSF) is preferred for large-scale applications in the field, as it facilitates microbial agent transport and ensures a long shelf life. However, research on enhancing the yield of lunate conidia fromE. vermicolathrough SSF is limited. In this study, we initially achieved a yield of 3.04 × 108conidia/g using a basic SSF medium composed of wheat bran, corn flour, and soybean flour. To improve this yield, we employed an orthogonal experimental design (OED) to identify the optimal medium composition, which required a wheat bran-to-corn flour-to soybean flour ratio of 7:2:1 (w/w/w), a substrate-to-water ratio of 1:0.7 (w/v), and the addition of 1.33% (w/w) glucose, 1.33% (w/w) yeast extract fermentation, and 1.33% (w/w) MgSO4. Using the response surface methodology (RSM), we calculated the optimal fermentation conditions, which were 24.9 °C, 78.0% relative humidity (RH), an inoculation volume of 16.3% (v/w), and a fermentation time of 7.1 days. Under these conditions, the yield of lunate conidia reached a maximum of 16.58 × 108conidia/g, a 4.45-fold increase after optimization. This study improved the yield ofE. vermicolalunate conidia and provides insights for developing biopesticides based on this strain.
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