Hallett Sascha L, Atkinson Stephen D, Erséus Christer, El-Matbouli Mansour
Institute of Zoology, Fish Biology and Fish Diseases, University of Munich, Kaulbachstrasse 37, DE-80539 Munich, Germany.
Syst Parasitol. 2004 Jan;57(1):1-14. doi: 10.1023/B:SYPA.0000010682.90311.91.
Thirty-nine freshwater tubificid oligochaetes were isolated, each of which harboured a triactinomyxon infection. Spore characteristics include the typical triactinomyxon anchor shape, eight germ cells within the sporoplasm and three unequal (two long and one shorter) caudal processes with square tips. Despite morphological similarities between the spores from the different hosts, their morphometrical data varied considerably; significantly, the ranges of dimensions of the smallest and largest exemplars were mutually exclusive. In order to ascertain the true number of spore types present, molecular methods were employed. Samples of waterborne spores, including the smallest and largest representatives, were selected from 11 host oligochaetes (all Tubifex tubifex Müller) and a nested PCR-RFLP 'riboprint' analysis performed. The small subunit ribosomal DNA gene (18S rDNA) was targetted and amplified through two rounds of PCR, then digested with the restriction enzymes Dde I and Hha I. The resultant major cleavage patterns produced by both enzymes indicated a single triactinomyxon form; however, the pattern of several less intense bands varied between the samples. From a subset of five samples drawn from across the full spectrum of spore sizes, a 327 bp region near the 5' was sequenced and was identical for all five samples. Comparison of this 327 bp region with that of 12 other triactinomyxons in GenBank showed 68.7-96.9% similarity (at least 9 base differences). A further 469 bp generated for each of the smallest, largest and mid-range (= reference) spore samples was identical also. The reference sample was sequenced further to yield 1,554 bp of 18S rDNA (GenBank accession number AY162270); comparison with other Myxozoa indicated this sequence was novel. The morphometrics of our triactinomyxon did not correlate with any published description. The morphometrical variation exhibited by spores of the triactinomyxon type in this study raises questions about the validity of using morphometrical data to distinguish spore types and suggests that there could be taxonomic redundancy in the diversity of actinosporeans recorded in the literature. The additional information provided by molecular data in this study was pivotal in the clarification of morphometrical variation exhibited by morphologically similar triactinomyxon spores released from different oligochaete hosts.
分离出39条淡水颤蚓寡毛类，每条都感染了三极虫。孢子特征包括典型的三极虫锚状形态、孢子质内有8个生殖细胞以及3个不等长（2个长的和1个短的）且末端为方形的尾突。尽管来自不同宿主的孢子在形态上相似，但其形态测量数据差异很大；值得注意的是，最小和最大样本的尺寸范围相互排斥。为了确定存在的孢子类型的真实数量，采用了分子方法。从11种宿主寡毛类（均为颤蚓属的颤蚓Müller）中选取了包括最小和最大代表的水传播孢子样本，并进行了巢式PCR-RFLP“核糖体印记”分析。以小亚基核糖体DNA基因（18S rDNA）为靶标，通过两轮PCR进行扩增，然后用限制性内切酶Dde I和Hha I进行消化。两种酶产生的主要切割模式表明为单一的三极虫形态；然而，几个较弱条带的模式在样本之间有所不同。从涵盖所有孢子大小范围的5个样本子集中，对5'端附近的一个327 bp区域进行了测序，所有5个样本的该区域均相同。将该327 bp区域与GenBank中其他12种三极虫的该区域进行比较，相似度为68.7 - 96.9%（至少有9个碱基差异）。为最小、最大和中等大小（=参考）孢子样本各自生成的另外469 bp也相同。对参考样本进一步测序，得到1554 bp的18S rDNA（GenBank登录号AY162270）；与其他粘孢子虫的比较表明该序列是新的。我们的三极虫的形态测量数据与任何已发表的描述均不相关。本研究中三极虫类型孢子所表现出的形态测量变异引发了关于使用形态测量数据区分孢子类型有效性的问题，并表明文献中记录的放线孢子虫多样性可能存在分类冗余。本研究中分子数据提供的额外信息对于阐明从不同寡毛类宿主释放的形态相似的三极虫孢子所表现出的形态测量变异至关重要。
