Isolation and identification of Lactarius deliciosus and the interaction with Pinus massoniana
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摘要: 【目的】开展松乳菇的分离鉴定及松乳菇与马尾松的共培养研究，以期通过与马尾松共培养实现松乳菇的规模栽培。【方法】试材松乳菇子实体采自湖北恩施天然马尾松次生林，使用组织分离法获得松乳菇菌株，采用rDNAITS分子鉴定分离菌株的种属，利用One-way ANOVA分析松乳菇接种对马尾松菌根侵染率、苗木生长及氮（N）、磷（P）、钾（K）含量的影响，并利用Duncan法分析马尾松菌根侵染率与松乳菇子实体生长指标的相关性。【结果】 rDNAITS分子鉴定3株真菌均为松乳菇，其在松木屑、棉籽壳等基质上能实现菌丝体的良好生长，属于兼性腐生菌根真菌。松乳菇与马尾松共培养结果显示，3株松乳菇均与马尾松形成外生菌根，在共培养11个月后，马尾松的菌根侵染率均在95%以上，外源接种松乳菇显著增加马尾松的干重、株高、地径、主根长和侧根数（P<0.05，下同），与CK相比，接种处理马尾松干重增加5.41%~19.25%，株高增加11.15%~20.59%，主根长增加12.58%~28.90%，侧根数增加34.88%~46.42%，地径增加幅度最大，比CK增加52.63%~68.42%；外源接种松乳菇显著提高马尾松根系和叶片的N和P含量，有利于马尾松苗木的培育。L-1无论对马尾松苗木的菌根侵染率、生长及N、P养分吸收影响效果均优于L-3和L-2处理。在共培养13个月后，获得24个松乳菇子实体，平均出菇数为2.67个/m2，产量为80.67 g/m2，其平均出菇数、总鲜重与马尾松菌根侵染率之间不显著相关（P>0.05）。【结论】 rDNA ITS方法可准确鉴定野生松乳菇，外源接种松乳菇能促进马尾松苗木的生长及对N、P等元素的利用，菌根侵染率的提高有利于松乳菇子实体形成，湖北省恩施市境内的松乳菇资源可作为驯化栽培松乳菇的潜在优良菌株。Abstract: 【Objective】To conduct isolation and identification of Lactarius deliciosus and cocultivation of L. deliciosus and Pinus massoniana, so as to realize large-scale cultivation of L. deliciosus by the co-cultivation.【Method】The fruiting body of Lactarius matsutake was collected from the natural masson pine secondary forest in Enshi, Hubei Province, and the Lactarius matsutake strain was obtained by tissue separation, genus and species of the strain were isolated through rDNA ITS. Influences of L. deliciosus inoculation on growth, mycorrhizal infection rate, nitrogen(N), phosphorus(P) and Kalium(K) contents of P. massoniana were analyzed using one-way ANOVA, and correlation analysis was conducted using Duncan method to explore correlation between the mycorrhizal infection rate of P. massoniana and the growth of L. deliciosus.【Result】According to the results of molecular identification of rDNA ITS sequences, three stains of fungi were identified as L. deliciosus. The L. deliciosus strains grew well on media such as pine sawdust and cottonseed shells. L. deliciosus strains were partially saprobe. Co-cultivation of L. deliciosus and P. massoniana showed that 3 L. deliciosus strains had developed mycorrhizae with P. massoniana. After co-cultivation for 11 months, the mycorrhizal infection rate of P. massoniana with L. deliciosus inoculation were all above 95%. Dry weight, height, basal diameter, length of main root and number of lateral roots were significantly increased(P<0.05, the same below). Compared with CK, the dry weights, plant heights, basal diameters, lengths of main roots and numbers of lateral roots of P. massoniana in treatments were increased by 5.41%-19.25%, 11.15%-20.59%, 12.58%-28.90%, 34.88%-46.42% and 52.63%-68.42%, respectively. Contents of N and P elements of P. massoniana were promoted with exogenous inoculation of L. deliciosus, which was good for cultivation of P. massoniana seedling growth. Generally, inoculation with L-1 had better effect on mycorrhizal infection rate, growth and absorption of N and P in P. massoniana than L-3 and L-2. After co-cultivation for 13 months, twenty-four fruiting bodies were obtained, with an average output of 2.67 mushrooms/m2 and a yield of 80.67 g/m2. However, the correlations of average number and total weight of fruiting bodies of L. deliciosus with the mycorrhizal ratio of P. massoniana were not significant(P>0.05).【Conclusion】rDNA ITS can be used to identify wild L. deliciosus accurately. The growth of P. massoniana and utilization of N and P were promoted by L. deliciosus exogenous inoculation. At the same time, the increase of mycorrhizal infection rate is helpful for the formation of fruiting body of L. deliciosus. The L. deliciosus in Enshi, Hubei Province can be used as a potential species for cultivation of L. deliciosus.
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Keywords:
- Lactarius deliciosus /
- ectomycorrhizal fungi /
- co-cultivation /
- Pinus massoniana
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