基于环境DNA技术检测木材中松材线虫的应用研究OA

[Objective]This study aims to use environment DNA(eDNA)technology to screen for efficient filters suitable for enriching Bursaphelenchus xylophilus eDNA and to establish a method capable of rapidly detecting B.xylophilus from large quantities of wood.[Methods]Using B.xylophilus tissue homogenate as the material,the enrichment efficiency of mixed cellulose ester(MCE),polyethersulfone(PES),and cellulose acetate(CA)membranes for B.xylophilus acids was compared.The enriched eDNA was detected by Quantitative real-time PCR(qPCR)to evaluate the enrichment efficiency of different membranes.Further,the best screened filter membrane was used to enrich eDNA from five wood samples of different sources(50 g each),and qPCR detection was performed with B.xylophilus specific primers and probes to verify the practical applicability of this method.[Result]MCE,PES,and CA membranes can all enrich B.xylophilus eDNA,with MCE and PES showing significantly better enrichment than CA,while there is no significant difference between the two;under low nematode concentration conditions,the MCE membrane exhibited the highest recovery rate.Based on this,MCE membranes were selected for eDNA enrichment and detection in actual wood samples.The results showed that the eDNA detection results of all wood samples were completely consistent with the molecular detection results of their corresponding wood shavings,confirming the reliability of this method.[Conclusion]The method established in this study,based on eDNA technology and MCE membrane enrichment,can effectively detect B.xylophilus from a large number of infected pine trees.It is suitable for high-throughput and rapid screening of pine wood nematodes in wood samples,providing a feasible technical approach for early monitoring and control of B.xylophilus.