枯萎松树中滑刃科线虫的鉴定与致病性检测
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摘要
滑刃科线虫近些年来成为线虫学研究的热点,新种不断被发现,对致病性的研究也非常普遍。在对伞滑刃属线虫的调查研究中,常常能从病死树木或进境木包装材料中分离到滑刃属线虫。本实验室从辽宁省大连市老铁山发现了数株枯萎黑松,后将病木带回并从中分离到一种陌生的滑刃属线虫。本研究经形态观察测量、分子生物学实验,该线虫被证明是一个新种,以其发现的城市命名为大连滑刃线虫(Aphelenchoides dalianensis sp. nov.).该线虫的雌虫尾型非常特殊,具有形似触角状的尖端分叉的尾尖突。此外,4条侧线、雌虫的后阴子宫囊、雄虫的交合刺形状等特征可以将其与滑刃属其它线虫种进行区分。它的近似种是大核滑刃线虫(A. macronucleatus),但两者在雌虫尾尖突和雄虫热杀死形态上有明显区别。
同时,本实验室在江苏省张家港市的枯萎马尾松中分离到一种伞滑刃属线虫。本研究经过显微拍照、电镜观测、特征值测记等形态学分类鉴定方法,以及ITS-PCR-RFLP和DNA测序、序列比对等分子生物学研究方法,确定该种线虫为莱奴尔夫伞滑刃线虫(Bursaphelenchus rainulfi)。该线虫有2条侧线；雌虫的阴门有一小的阴门盖,尾端弯向腹面,末端钝圆；雄虫的交合刺很小,弯曲不明显,交合伞卵圆形,短小,仅包住尾部末端。
已有研究表明松材线虫以外的线虫对松树具有潜在的致病性。由于本实验室分离、鉴定出的Aphelenchoides dalianensis和Bursaphelenchus rainulfi都是枯死松树中的优势种,并未发现其它寄生性线虫,故而我们对两者的致病性产生兴趣并进行了研究。通过野外接种实验,检测了两种线虫对湿地松、马尾松和黑松的致病性。实验结果证实B. rainulfi无明显致病性。A. dalianensis对黑松表现出较强的致病性,并造成超过一半的马尾松枯黄衰弱,以及小部分的湿地松表现出一定的枯萎症状。依此结果可以推测出A. dalianensis与大连老铁山的枯死黑松有直接的联系,故而对A. dalianensis进行风险分析,查明分布状况和寄主范围等非常必要。
Nematodes of Aphelenchoididae have been a hot topic in recent years, quite a lot of new species have been found, and their pathogenicity has been studied. During the study of nematodes in Bursaphelenchus, nematodes in Aphelenchoides have often been found. Our lab found some wilt black pines in Lao Tieshan, Dalian City, Liaoning Province, and isolated a strange kind of nematodes of Aphelenchoides. It has been proved to be a new species through morphological and molecular experiments. It was named by the name of the city which it was found:Aphelenchoides dalianensis sp. Nov. A. dalianensis was characterized by 4 lateral lines, small spicule of male, female vulva post median and especially, female tail with a special mucronate structure which had a divarication, forming an antennae-like terminus. A. macronucleatus resembles the new species, especially in male spicule, but differs from female tail terminus and the shape of male after heat-killed.
Our lab has also isolated a suspicious species of nematode from mason pines in Zhangjiagang City, Jiangsu Province, in 2007. It has been proved to be B. rainulfi through morphological identification and ITS-PCR-RFLP, and DNA sequencing as well. This species has 2 lateral lines; female has a small vulval flap, its tail slightly curved ventrally, and has a round end. Male has a small spicule, an oval and small bursa, only wraps the end of the tail.
Researches have already showed that nematodes other than Bursaphelenchus xylophilus had potential pathogenicity on pines. B. rainulfi and A. dalianensis were found from wilt pines as dominant species without Bursaphelenchus xylophilus. As a result, we were interested in their pathogenicity and detected them through field experiment. The two species were inoculated into Pinus elliottii, P. massoniana and P. thunbergii. The result showed that B. rainulfi had little pathogenicity while A. dalianensis showed comparatively strong pathogenicity towords P. thunbergii, caused more than half of the P. massoniana wilt, and some symptoms on a small part of P. elliottii. Therefore, it is quiet necessary to carry out Pest Risk Analysis, to investigate its distribution, host range and risk.
同时,本实验室在江苏省张家港市的枯萎马尾松中分离到一种伞滑刃属线虫。本研究经过显微拍照、电镜观测、特征值测记等形态学分类鉴定方法,以及ITS-PCR-RFLP和DNA测序、序列比对等分子生物学研究方法,确定该种线虫为莱奴尔夫伞滑刃线虫(Bursaphelenchus rainulfi)。该线虫有2条侧线；雌虫的阴门有一小的阴门盖,尾端弯向腹面,末端钝圆；雄虫的交合刺很小,弯曲不明显,交合伞卵圆形,短小,仅包住尾部末端。
已有研究表明松材线虫以外的线虫对松树具有潜在的致病性。由于本实验室分离、鉴定出的Aphelenchoides dalianensis和Bursaphelenchus rainulfi都是枯死松树中的优势种,并未发现其它寄生性线虫,故而我们对两者的致病性产生兴趣并进行了研究。通过野外接种实验,检测了两种线虫对湿地松、马尾松和黑松的致病性。实验结果证实B. rainulfi无明显致病性。A. dalianensis对黑松表现出较强的致病性,并造成超过一半的马尾松枯黄衰弱,以及小部分的湿地松表现出一定的枯萎症状。依此结果可以推测出A. dalianensis与大连老铁山的枯死黑松有直接的联系,故而对A. dalianensis进行风险分析,查明分布状况和寄主范围等非常必要。
Nematodes of Aphelenchoididae have been a hot topic in recent years, quite a lot of new species have been found, and their pathogenicity has been studied. During the study of nematodes in Bursaphelenchus, nematodes in Aphelenchoides have often been found. Our lab found some wilt black pines in Lao Tieshan, Dalian City, Liaoning Province, and isolated a strange kind of nematodes of Aphelenchoides. It has been proved to be a new species through morphological and molecular experiments. It was named by the name of the city which it was found:Aphelenchoides dalianensis sp. Nov. A. dalianensis was characterized by 4 lateral lines, small spicule of male, female vulva post median and especially, female tail with a special mucronate structure which had a divarication, forming an antennae-like terminus. A. macronucleatus resembles the new species, especially in male spicule, but differs from female tail terminus and the shape of male after heat-killed.
Our lab has also isolated a suspicious species of nematode from mason pines in Zhangjiagang City, Jiangsu Province, in 2007. It has been proved to be B. rainulfi through morphological identification and ITS-PCR-RFLP, and DNA sequencing as well. This species has 2 lateral lines; female has a small vulval flap, its tail slightly curved ventrally, and has a round end. Male has a small spicule, an oval and small bursa, only wraps the end of the tail.
Researches have already showed that nematodes other than Bursaphelenchus xylophilus had potential pathogenicity on pines. B. rainulfi and A. dalianensis were found from wilt pines as dominant species without Bursaphelenchus xylophilus. As a result, we were interested in their pathogenicity and detected them through field experiment. The two species were inoculated into Pinus elliottii, P. massoniana and P. thunbergii. The result showed that B. rainulfi had little pathogenicity while A. dalianensis showed comparatively strong pathogenicity towords P. thunbergii, caused more than half of the P. massoniana wilt, and some symptoms on a small part of P. elliottii. Therefore, it is quiet necessary to carry out Pest Risk Analysis, to investigate its distribution, host range and risk.
引文
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