厚垣普奇尼亚菌Pochonia chlamydosporia产生的几丁质酶对南方根结线虫卵孵化的影响

Effect of chitinases produced by Pochonia chlamydosporia on egg-hatching of Meloidogyne incognita

【目的】卵寄生真菌厚垣普奇尼亚菌Pochonia chlamydosporia产生的几丁质酶对定居性根结类和胞囊类线虫的卵壳消解起重要促进作用。线虫卵寄生真菌产生几丁质酶的特性是评价食线虫真菌生物防治潜力的重要生化指标之一。【方法】利用NAG法和pNP法分别测定7株不同来源厚垣普奇尼亚菌Pochonia chlamydosporia几丁质酶的降解酶系和外切酶活性。【结果】7个菌株都能够产生外切酶,其中QNAV97-2、NRRL13094、CFCC84963和CFCC80919菌株能够产生几丁质降解酶系,几丁质酶活性染色检测分别有2条和4条具有几丁质酶活性的蛋白谱带,其中CFCC80919和QNAV97-2分别产生的38.9kD和39.8kD几丁质酶可能是已知的CHI43几丁质酶。具有几丁质降解酶活性的4个厚垣普奇尼亚菌对根结线虫的卵孵化抑制率为40.32%～55.15%,未测定出几丁质降解酶活性的3个菌株对根结线虫的卵孵化抑制率不足20%。显微观察处理的南方根结线虫卵壳变形和破坏。【结论】厚垣普奇尼亚菌系产生的几丁质酶能够消解南方根结线虫卵壳,特别是胚前发育期未成熟卵,抑制卵孵化或杀死卵,且在消解卵的过程中起重要作用。厚垣普奇尼亚菌的几丁质酶活性水平可能是造成该类菌杀线活性差异的原因之一。

【Objective】 Chitinases produced by egg-parasitic fungus Pochonia chlamydosporia have been suggested to play an important role in digestion of root knot and cyst nematode eggshells. The egg-parasitic fungi with chitinase activity is one of the most important biochemical characters in suppression of egg hatching,and can be used for evaluation of the biocontrol potential of those fungi. 【Method】 The chitinolytic systems and exochitinase produced by 7 isolates of P. chlamydosporia were measured by means of NAG（N-acetyglucosamine） and pNP（ρ-nitrophenol） respectively. 【Result】 The results showed that all the isolates had different levels of exochitinase activity,however only the isolates QNAV97-2,NRRL13094,CFCC84964 and CFCC80919 had activity of chitinolytic systems. On SDS-PAGE analysis with 0.01% glycol chitin,two and four protein bands with chitinase activity were detected,the chitinases of 38.9kDa and 39.8kDa produced by CFCC80964 and QNAV97-2 were believed to be the CHI43 purified in early studies. Inhibitory rates to egg hatching of Meloidogyne incognita by 4 isolates with chitinolytic activity were ranged from 40.32% to 55.15%,however,those by 3 isolates without chitinolytic activity were lower than 20%. Microscopy observations demonstrated that the eggshells of M. incognita was deformed and destroyed in the treatment of culture filtrates from P. chlamydosporia. 【Conclusion】 It is concluded that the chitinases produced by P. chlamydosporia causes lysis of the eggshell of M. incognita,especially immature eggs at early embryonic development stage,and result in egg hatching inhibition and/or egg kill. The digestion of the eggshell by chitinase activity plays an important role in the control of M. incognita. Difference in chitinase activity levels produced by P. chlamydosporia may result in their difference in nematicidal activity to nematodes.