进境木质包装中伞滑刃属线虫疫情分析

2003年1月至2006年6月，宁波口岸共检测进境木质包装8720批次，其中1772批次检出各种活线虫，疫情检出率高达20．3％。根据形态学特征和ITS—RFLP图谱鉴定伞滑刃属线虫19种（其中4种被鉴定为新种，1种尚未发表）342批次，占总检测批次的3．9％。不仅从疫区也从多个非疫区国家进境木质包装中检出松材线虫。     

进境木质包装中豆伞滑刃线虫的鉴定

豆伞滑刃线虫由宁波出入境检验检疫局植检室验室于2004年6月首次从台湾的针叶木质包装中截获并报道以来,至今共截获17批次,分别来自韩国（6批次）、中国台湾（4批次）,日本（4批次）,印度（1批次）,马来西亚（1批次）和南非（1批次）.该线虫属松材线虫组,与拟松材线虫、松材线虫等十分接近.其主要鉴定特征是：雌虫尾圆锥形略向腹面弯曲,其腹面的边线较平直,而背面的边线明显呈圆弧形;尾尖长约2～4μm,渐细,有时呈丝状,位于腹面;雄虫交合刺较大（约36～40μm）,中间部分几乎呈直线.该线虫很容易与拟松材线虫等混淆,当形态学鉴定有困难时,ITS-RFLP方法是十分有效的辅助手段.我国大陆尚未有该线虫发生的报道.     

无锡口岸进境木质包装中截获的两种伞滑刃线虫的形态鉴定

松材线虫病是林业上的一种毁灭性病害.在进境木质包装检疫中,除该病的病原物松材线虫(Bursaphelenchus xylophilus(steiner et buhrer)Nickel)外,还常常截获同属于伞滑刃线虫属(Bursaphelenchus spp.)的多种与之形态非常相似的线虫.本研究对进境木质包装检疫中截获的两种伞滑刃属线虫--B. rainulfi和B.conicaudatus进行了形态描述和鉴定.     

食菌伞滑刃线虫和阿苏里伞滑刃线虫的比较鉴定

宁波口岸在进境木质包装检疫中经常截获食菌伞滑刃线虫和阿苏里伞滑刃线虫,这2种线虫都属于fungivorus组,形态上十分近似。主要区别在于:前者唇区缢缩十分明显,而后者缢缩不明显;前者排泄孔与半月体平均距离较远(约11μm),而后者较近(约5μm);前者雄虫热杀死后尾部强烈向腹面弯曲(超过180°),后者尾部略腹弯(约90°);前者基顶与喙突相距约9.8μm,喙突高约2.7～3.5μm,后者则分别为8.4μm和1.5～2.8μm;前者雌虫尾多数腹弯十分明显,尾较短(长约50μm),后者尾一般略腹弯(长约68μm)。此外,ITS区的PCR-RFLP方法可区分上述2种线虫。     

俄罗斯木质包装中杨树伞滑刃线虫的鉴定

宁波口岸从来自俄罗斯的杨树木质包装中截获一种伞滑刃线虫,经形态学和分子生物学研究,鉴定为杨树伞滑刃线虫。该线虫与伪伞滑刃线虫十分近似,主要鉴定特征是:阴门盖明显腹弯,近阴门盖后半部的虫体明显凹陷;雄虫交合刺近远端边缘平直,无明显凹陷,远端盘状突十分显著;雌虫尾亚圆柱形或近圆锥形,尾端多数呈多变的尾尖突状,偶尔几乎钝圆。该线虫人工培养后雌虫尾形多数介于圆柱形和圆锥形之间,这与原始文献的报道有差异。这是我国口岸首次截获该线虫。     

宁波检验检疫局多次截获非洲伞滑刃组线虫

2006年来,宁波出入境检验检疫局植检实验室曾多次从进境木质包装中首次截获并报道非洲伞滑刃组（africanus group）线虫,它们分别为非洲伞滑刃线虫（Bursaphelenchus africanus Braasch, Gu, Burgermeister, Brandstetter &; Metge, 2006）、伯氏伞滑刃线虫（B. burgermeisteri Braasch, Gu, &; Brandstetter, 2007）和欧斐切伞滑刃线虫（B. obeche Gu, Braasch, Zhen, Burgermeister &; Lin, 2008）。这些线虫均为作者最先截获。非洲伞滑刃组线虫的主要鉴定特征是：侧线4条;尾乳突7个（第3对和第4对靠近并位于交合伞起始处,第3对略位于第4对前并更靠近腹面）;交合刺较直,中部有横纹,冠状体愈合（基顶和喙突不明显）,远端无盘状突;后阴子宫囊约占肛阴距1 /2。目前尚无关于该组线虫媒介昆虫及致病性的报道。     

2种常见伞滑刃线虫的特异PCR检测方法

包括松材线虫和椰子红环腐线虫在内的伞滑刃属线虫形态上相似性高,形态鉴定难度较大。本文通过对伞滑刃属常见线虫群体ITS2区序列比对、分析,设计了木质包装材料检疫中常见的泰国伞滑刃线虫和豆伞滑刃线虫特异引物对各1对,并研究了这2种常见伞滑刃线虫的PCR检测方法,使用本文特异引物可分别特异、快速检测出这两种线虫。     

韩国入境木质包装箱中阿苏里伞滑刃线虫的鉴定

广东出入境检验检疫局植检实验室于2007年12月自韩国木质包装中截获一种伞滑刃属线虫,经形态学观察、测量,以及rDNA-ITS区扩增测序分析,鉴定为阿苏里伞滑刃线虫.该线虫除韩国和中国台湾外,还未见在其他地方报道.     

伞滑刃属线虫专家鉴定系统

近年来,口岸检验检疫机构多次在进境木质包装中截获活体线虫,其中多次截获到伞滑刃属的松材线虫和与其形态相似的拟松材线虫.由于线虫形体细小,种的形态差别不大,一般非专业的工作人员难以进行鉴定.     

厦门口岸首次从木包装中截获瓦里西伞滑刃线虫

厦门口岸于2011年6月从来自希腊的荒料石垫木中截获一种伞滑刃属线虫。经形态学比较、测量,该线虫虫体细长,侧区有4条侧线。雌虫有小阴门盖,尾圆锥形,略腹弯,尾端尖。雄虫交合刺成对,不愈合,显著腹弯,长约13～15μm;基顶前伸,端部圆滑,略背弯;喙突显著,末端钝尖,远端有小而明显的盘状突;雄虫尾部乳突7个。结合ITS-RFLP指纹图谱比对,进一步将该线虫鉴定为瓦里西伞滑刃线虫(Bursaphelenchus vallesianus Braasch,Schonfeld,Polomski,et al,2004)。     

Validation of morphological keys for identification of Bursaphelenchus xylophilus (Nematoda,Parasitaphelenchidae) to group and species level

Official analyses undertaken in the framework of an official survey, or import controls need reliable results. This can be achieved by using validated methods. For morphological tests, this validation process is rarely illustrated despite guidance provided in EPPO PM 7/98 (1) Specific requirements for laboratories preparing accreditation for a plant pest diagnostic activity. This paper presents validation for the morphological identification of Bursaphelenchus xylophilus, with the evaluation results of published identification keys and of internally designed identification keys at B. xylophilus group and species levels. For published identification keys some criteria were shown not to be reliable for routine use: excretory pore position and number of caudal papillae. The key designed in the laboratory for xylophilus group identification was shown to be sensitive and specific when one male and one female were observed. The key designed for B. xylophilus species identification is sensitive, specific and reproducible if only one female of B. xylophilus is observed. The tools designed were validated as simple and reliable for routine analysis. The advantages and limitations of the validation process for morphological tools are discussed for process improvement.     

一种锥尾伞滑刃线虫形态和RFLP图谱变化研究

宁波口岸植检实验室在来自台湾的榕属木质包装中截获一种伞滑刃线虫,经形态学观察、测量,ITS-RFLP图谱比对及序列分析,鉴定为锥尾伞滑刃线虫(Bursaphelenchus conicaudatus Kanzaki,Tsuda&Futai,2000)。该线虫属松材线虫组,与鲍嘉伞滑刃线虫最近似。本文着重对该线虫与近似种的形态特征区别及ITS-RFLP图谱种内变化进行讨论。     

伞滑刃线虫属分种检索

依据2004年以前发表的所有伞滑刃属新种原始报道的资料,分析了伞滑刃属73个相关种的所有形态学分类特征,编制了64个伞滑刃属有效种的分类检索表.     

Pest survey of softwood boxes,with special emphasis on pinewood nematode,Bursaphelenchus xylophilus (Nematoda: Parasitaphelenchidae)

Wood packaging material (WPM) is an important pathway for the spread of non‐native plant pests. To reduce the likelihood of plant pest movement with WPM, the International Standard for Phytosanitary Measures No. 15: “Regulation of Wood Packaging Material in International Trade” (ISPM 15) was developed in the framework of the International Plant Protection Convention. To be compliant with this Standard, WPM shipped internationally must be either heat‐treated or fumigated, regardless of any specific characteristics of the WPM. The objective of this survey was to determine if Bursaphelenchus xylophilus (Nematoda: Parasitaphelenchidae) or insects, especially Monochamus spp. (Coleoptera: Cerambycidae), were present in a specific subset of softwood boxes. While not compliant with ISPM 15, these boxes had been treated with wood preservatives, stored indoors for over five years, and, in addition to meeting various quality standards, almost all had a moisture content below 20 percent. United States Department of Agriculture staff inspected a sample of 630 softwood boxes, focusing on those showing signs of possible pest infestation. Based on a binomial distribution with an efficacy of detection equal to 0.95, a sample size of 630 provides a 95 percent confidence of detecting pests if >0.5 % of boxes are infested. No B. xylophilus or insect pests were found in the boxes, though other nematode genera (Aphelenchoides, Aphelenchus, and Filenchus), which feed on decomposing fungi in wood and do not damage trees, were found in 21 boxes. This study demonstrates that not all types of WPM present a high phytosanitary risk. It may be worthwhile to consider an amendment to ISPM 15 to differentiate between various risk categories of WPM in order to minimize costs and environmental impacts associated with treatments currently prescribed in ISPM 15.     

A new Huffmanela species, H. schouteni sp. n. (Nematoda: Trichosomoididae) from flying fishes in Cura?ao

A new nematode species, Huffmanela schouteni sp. n., has been established on the basis of its egg morphology and biological characters (adult nematodes are unknown). The dark-shelled eggs of this histozoic parasite occur in masses in the abdominal cavity, serose covers of internal organs and in the liver of the flying fishes Hirundichthys affinis Günther (type host) and Cypselurus cyanopterus Cuvier et Valenciennes in Cura?ao. The eggs of H. schouteni sp. n. differ from those in other congeneric species mainly in the absence of small spines on the surface of the transparent envelope enclosing the egg proper, measurements (size of eggs 0.069-0.075 x 0.027-0.030 mm) and their localization in the host. A key to Huffmanela species based on egg morphology has been provided.     

Two new species of Rhabdochona (Nematoda: Rhabdochonidae) from freshwater fishes in Thailand

Two new species ofrhabdochonid nematodes are described from the intestine of freshwater fishes in Chiang Mai Province, northern Thailand: Rhabdochona (Rhabdochona) pseudomysti sp. n. from the catfish Pseudomystus siamensis (Regan) (Bagridae, Siluriformes) in the Fang Brook, a tributary of the Kok River (the Mekong River basin), Fang District and Rhabdochona (Globochona) thaiensis sp. n. from the cyprinid Mystacoleucus marginatus (Valenciennes) (Cyprinidae, Cypriniformes) in the Ping River (the Chao Phraya River basin), Muang District. Rhabdochona pseudomysti is mainly characterized by simple, leaf-like oval deirids (a unique feature among Rhabdochona spp.), a prostom with 14 anterior teeth, the presence of basal prostomal teeth, the length ratio of the muscular and glandular portions of oesophagus (1:2.1-2.6), an unusually long left spicule (1.10-1.22 mm), length ratio of spicules (1:11.5-14.7), arrangement of genital papillae, and conspicuously elevated vulval lips. Rhabdochona thaiensis differs from other representatives of the subgenus Globochona Moravec, 1972 possessing caudal projections on the tail tip in that it has only 2 claw-shaped projections located ventrally on the tail tip of both males and females; the species is mainly characterized by the presence of distinct pseudolabia, 8 anterior prostomal teeth, absence of basal teeth, bifurcated deirids, length ratio of the muscular and glandular portions of oesophagus (1:11.3-11.9), conspicuously short (135-141 microm) left spicule, arrangement of genital papillae, and somewhat elevated vulval lips. Fully developed eggs of R. pseudomysti and R. thaiensis remain unknown. These are the first nominal species of Rhabdochona reported from Thailand.