两株松材线虫拮抗细菌的筛选和鉴定

【目的】筛选对松材线虫具有较高杀线活性的拮抗细菌。【方法】以分离自南京、洛阳、上海的松树茎部的137株内生细菌为研究对象,采用培养滤液和菌悬液浸渍法对这些菌株进行杀线活性的测定,并对筛选的高效拮抗细菌进行菌种鉴定。【结果】通过培养滤液浸渍试验筛选出3株细菌对松材线虫有较高的杀线活性,使用3种菌滤液处理线虫48 h后,线虫死亡率均达到100%,其中菌株LYMC-3的培养滤液还可使线虫虫体出现消解。将3株菌的培养滤液分别稀释2倍、4倍和10倍后处理松材线虫,随稀释倍数的增加,培养滤液的杀线活性逐渐降低。处理线虫48 h后,菌株LYMC-3的10倍稀释滤液与其他两株菌滤液相比对线虫的致死率最高,为94.7%;在菌悬液浸渍试验中筛选出菌株NJSZ-13对松材线虫有较高杀线活性,105cfu/mL浓度的菌悬液处理线虫48 h后,线虫死亡率达81.5%。通过对LYMC-3和NJSZ-13菌株的形态特征、生理生化特征、Biolog鉴定和16S rDNA序列以及系统发育分析,确定菌株LYMC-3、NJSZ-13分别为短小芽孢杆菌(Bacillus pumilus)和蜡样芽孢杆菌(Bacillus cereus)。【结论】从松树体内筛选到两株对松材线虫有高效拮抗活性的菌株LYMC-3和NJSZ-13,对生物防治松材线虫病具有潜在的应用价值。

The screening and identification of two bacterial strains with nematicidal activity against Bursaphelenchus xylophilus

Objective] The aim of the present study was to screen bacteria with nematicidal activity against the pine wood nematode (Bursaphelenchus xylophilus).Method] A total of 137 bacteria strains,which were isolated from the stems of pine trees in Nanjing,Luoyang and Shanghai,China,were assessed for nematicidal activity by immersion test with cultural filtrate and buffer suspension.Result] Cultural filtrates of three strains had stronger nematicidal activity than the other strains,and nematodes treated with these strains reached 100％ mortality after 48 h and even exhibited decomposition when treated with the cultural filtrate of strain LYMC-3.The cultural filtrates of three strains,which were diluted 2,4 and 10 times still showed high nematicidal activity.Nematicidal activity decreased with increasing dilution,and when nematodes were treated with 10× diluted filtrates for 48 h,the greatest observed mortality was 94.7％ withstrain LYMC-3.Moreover,the buffer suspension of strain NJSZ-13 had strong nematicidal activity against B.xylophilus,and 81.5％ nematode mortality was achieved after 48 h treatment with 105 cfu/mL buffer suspension of strain NJSZ-13.Based on morphology,physiological and biochemical characteristics,Biolog system identification,16SrDNA sequences,and a phylogenetic tree,the strains LYMC-3 and NJSZ-13 were identified as Bacillus pumilus and B.cereus,respectively.Conclusion] Two strains of bacteria LYMC-3 and NJSZ-13 with high nematicidal activity which were isolated from the stems of pine trees could provide potential solution for biological control of Bursaphelenchus xylophilus.