湛江海域褐篮子鱼寄生虫多样性调查及种群生态学研究
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摘要
本文从2009.1-2009.12在湛江海域对675尾褐篮子鱼(Siganus fuscessens)的寄生虫展开多样性及种群动态调查,计发现10种寄生虫,隶属于4门7纲9目9科9属,包括扁形动物门3种(单殖吸虫2种、复殖吸虫1种)、线虫动物门2种、棘头动物门2种、节肢动物门3种,分别为:朦胧四叉虫Tetranclstrum nebulosi、未定种多唇虫Polylabris sp.、广东寡腺吸虫Oligolecithoides guangdongensis、内弯宫脂线虫Hysterothylacium aduncum、篮子鱼前驼形线虫Procamallanus sigani、未定种副细吻虫Parahadinorhynchus sp.、未定种新棘吻虫Neoechinorhynchus sp.、未定种鱼虱caligus sp.1、caligus sp.2,及未定种似柱颚虱Clavellopsis sp.。
分子生物学方法在物种鉴定中被证明具有有效、快速和区分度强等优点。本实验关于寄生虫的分类鉴定以形态学观察为首要依据,结合18S rDNA、5.8S rDNA、ITS-1 rDNA和ITS-2 rDNA序列数据等对物种进行鉴定。实验中分别扩增了10种寄生虫的以上序列的全长或接近全长,结果显示只用18S rDNA片段或ITS-1 + 5.8S + ITS-2 rDNA片段作为DNA序列,可以鉴定到属该分类阶元的有5种,鉴定到科的达9种,鉴定准确度分别都为50%、90%;用18S rDNA和ITS-1 + 5.8S + ITS-2 rDNA两种方法进行综合鉴定,鉴定到属的水平的为6种,鉴定到科亦为9种,准确率为60%、90%;且用ITS-1 + 5.8S + ITS-2 rDNA有一种鉴定到种。该结果表明,18S rDNA用于科间、属间鉴定具有较高的可靠性,而ITS-1+ 5.8S + ITS-2 rDNA可作为属间,甚至属内不同物种的鉴定的关键指标。结果中用ITS-1 + 5.8S + ITS-2 rDNA,只将其中的1个序列鉴定到种,作者认为原因与扩增得到的10种虫的18S rDNA、ITS-1 + 5.8S + ITS-2 rDNA序列都为新序列,GenBank中尚无该物种或该种所在的属的序列登记有关。
在调查中发现,只有朦胧四叉虫T.nebulosi和未定种多唇虫Polylabris sp.具有明显的季节性动态变化。朦胧四叉虫种群从1月到5月,感染率、平均密度和平均感染强度不断上升,5月达到峰值,此后开始下降,在10月份降到最低点,接近零感染,然后,在11、12月开始缓慢回升,但保持在较低水平。未定种多唇虫的感染率、平均密度的变化趋势除了感染率的峰值提前到3月与在9月、11月该虫种群各出现一次小的波动外,其它动态变化与朦胧四叉虫基本相似。
两种单殖吸虫种群在宿主种群中均呈聚集性分布。两种单殖吸虫在空间分布上,对宿主的左鳃或右鳃的选择没有显著差异性,但对鳃片大小和鳃片在鳃腔中的位置有偏好。朦胧四叉虫和未定种多唇虫都集中分布在第一对和第二对鳃片上,在第三对鳃、第四对鳃片分布的比例少,第一、第二对与第三、第四对鳃片的平均密度有显著差异。对结果进行分析表明朦胧四叉虫对鳃片位置偏好程度为第一对>第二对>第三对>第四对鳃。未定种多唇虫对鳃片位置的喜好则为第一>第二>第四>第三,分布在第四对鳃的平均密度稍大于第三对,但后两对差异性不显著。在鳃片分布区的喜好选择上,朦胧四叉虫集中寄生在鳃的中区,即Ⅳ、Ⅴ、Ⅵ三个位点,并偏好寄生于鳃片上靠腹侧的鳃丝;未定种多唇虫栖息于于外区,即鳃丝远端的第Ⅶ、Ⅷ、Ⅸ位点,并偏好寄生于鳃片上靠背侧的鳃丝。研究还发现朦胧四叉虫和未定种多唇虫种群的平均密度、平均感染强度、感染率与宿主体长呈正相关,不同月份的海水温度、盐度的动态变化是引起种群动态变化的主要因素。
In a survey of the parasites of Siganu fuscessenss, 10 species of parasites, belonging to 4 phylum 7 class 9 orders 9 families 9 genera, were indentified in 675 S. fucessenss in zhanjiang area of South China Sea during Jan 2009 to Dec 2009. The gill parasites include two Monogeneas (Tetranclstrum nebulosi and Polylabris sp.), one Copepoda (Caligus sp.1). The intestinal parasite found was one Digenea (Oligolecithoides guangdongensis). Two Nematodas (Procamallanus sigani and Hysterothylacium aduncum). And two Acanthocephalas (Parahadinorhynchus sp. and Neoechinorhynchus sp.).The body surface parasites were two Copepodas (Caligus sp.2 and Clavellopsis sp.).
Molecular biology methods used for the identification of parasite species proved to be cost-effective, rapid and discriminatory. In this study, species identification principally based on morphological characteristics and tightly combined with molecular dates of 18S rDNA, 5.8S rDNA, ITS-1 rDNA and ITS-2 rDNA sequence. The whole or nearly whole length of the18S ribosomal DNA fragment sequences, 5.8S ribosomal DNA fragment sequences, ITS-1 and ITS-2 rDNA gene of ten species of parasites were amplified using PCR techniques.The results indicated that 5 speicies,9 families can be accurately identified in the level of genus only using either of the 18S rDNA sequence or 5.8S rDNA together with ITS-1 and ITS-2 sequence fragmet, each precision rate of the two reached 50 percentage and 90 percentage respectively in the level of genus and family. While 6 speicies, 9 families can be correctly identified in the level of genus by combining both the two sequence date with ITS-1 and ITS-2 and got percentage precision of 60 percentage and 90 percentage . Results certified that both 18S rDNA and 5.8S rDNA together with ITS-1 and ITS-2 as important indexs to analyse different genera of the family or between different family was cost-effective. Moreover, the later method still was suited to discriminate the closely–related taxa within genus. Only one speices was indentified in the level of species,not because of low degree of accuracy by our methods but the sequence dates of 18S rDNA, 5.8S rDNA, ITS-1 rDNA and ITS-2 rDNA were first found and the lack of dates of corresponding genus in GenBank at prensent.
Only two parasites T. nebulosi and Polylabris sp. obviously dispaly the seasonal population dynamics. The prevalence,the abundance and intensity of T.nebulosi population in the wild host increased from January, and attained peak in May then declined quickly in spite of being at a high leve in June, and it nearly reached to zero infection till October. The prevalence,the abundance and intensity tended to increase but still keep on a low level of infection since November.The prevalence,the abundance and intensity dynamics of Polylabris sp. population were very similar with the T.nebulosi population except the prevalence atained peak to shift to an earlier time in March,and embraced a small peak of infection respectively in September and November. Both two Monogenea presented aggregated frequency distributions in the host population.The sites selection of two monogeneain frapopulations on host were as follows. There were no statistically significant diferences for both of them covering the sides of gills, but every type of parasites shew noticeable preferences in the front two pairs of gills and only few distributed in the two back pairs of gills. The site preference in gill decreased from the 1st, 2nd ,3th to the 4th pairs in T.nebulosi, while the seuence1st, 2nd , 4th ,3th pairs in Polylabris sp.
As to the distributions of two parasites on the branchial plate,T.nebulosi generally occupiedⅣ,Ⅴ,Ⅵsectors and prefered gastro- to dorso gill. While Polylabris sp. distributed inⅦ,Ⅷ,Ⅸsectors and prefered dorso to gastro- gill. In addition, we found the prevalence,the abundance and intensity of both two Monogenea positively correlated with the host's length ,and the dynamic rule may mainly induced by the variation of sea water salinity and tempture.
分子生物学方法在物种鉴定中被证明具有有效、快速和区分度强等优点。本实验关于寄生虫的分类鉴定以形态学观察为首要依据,结合18S rDNA、5.8S rDNA、ITS-1 rDNA和ITS-2 rDNA序列数据等对物种进行鉴定。实验中分别扩增了10种寄生虫的以上序列的全长或接近全长,结果显示只用18S rDNA片段或ITS-1 + 5.8S + ITS-2 rDNA片段作为DNA序列,可以鉴定到属该分类阶元的有5种,鉴定到科的达9种,鉴定准确度分别都为50%、90%;用18S rDNA和ITS-1 + 5.8S + ITS-2 rDNA两种方法进行综合鉴定,鉴定到属的水平的为6种,鉴定到科亦为9种,准确率为60%、90%;且用ITS-1 + 5.8S + ITS-2 rDNA有一种鉴定到种。该结果表明,18S rDNA用于科间、属间鉴定具有较高的可靠性,而ITS-1+ 5.8S + ITS-2 rDNA可作为属间,甚至属内不同物种的鉴定的关键指标。结果中用ITS-1 + 5.8S + ITS-2 rDNA,只将其中的1个序列鉴定到种,作者认为原因与扩增得到的10种虫的18S rDNA、ITS-1 + 5.8S + ITS-2 rDNA序列都为新序列,GenBank中尚无该物种或该种所在的属的序列登记有关。
在调查中发现,只有朦胧四叉虫T.nebulosi和未定种多唇虫Polylabris sp.具有明显的季节性动态变化。朦胧四叉虫种群从1月到5月,感染率、平均密度和平均感染强度不断上升,5月达到峰值,此后开始下降,在10月份降到最低点,接近零感染,然后,在11、12月开始缓慢回升,但保持在较低水平。未定种多唇虫的感染率、平均密度的变化趋势除了感染率的峰值提前到3月与在9月、11月该虫种群各出现一次小的波动外,其它动态变化与朦胧四叉虫基本相似。
两种单殖吸虫种群在宿主种群中均呈聚集性分布。两种单殖吸虫在空间分布上,对宿主的左鳃或右鳃的选择没有显著差异性,但对鳃片大小和鳃片在鳃腔中的位置有偏好。朦胧四叉虫和未定种多唇虫都集中分布在第一对和第二对鳃片上,在第三对鳃、第四对鳃片分布的比例少,第一、第二对与第三、第四对鳃片的平均密度有显著差异。对结果进行分析表明朦胧四叉虫对鳃片位置偏好程度为第一对>第二对>第三对>第四对鳃。未定种多唇虫对鳃片位置的喜好则为第一>第二>第四>第三,分布在第四对鳃的平均密度稍大于第三对,但后两对差异性不显著。在鳃片分布区的喜好选择上,朦胧四叉虫集中寄生在鳃的中区,即Ⅳ、Ⅴ、Ⅵ三个位点,并偏好寄生于鳃片上靠腹侧的鳃丝;未定种多唇虫栖息于于外区,即鳃丝远端的第Ⅶ、Ⅷ、Ⅸ位点,并偏好寄生于鳃片上靠背侧的鳃丝。研究还发现朦胧四叉虫和未定种多唇虫种群的平均密度、平均感染强度、感染率与宿主体长呈正相关,不同月份的海水温度、盐度的动态变化是引起种群动态变化的主要因素。
In a survey of the parasites of Siganu fuscessenss, 10 species of parasites, belonging to 4 phylum 7 class 9 orders 9 families 9 genera, were indentified in 675 S. fucessenss in zhanjiang area of South China Sea during Jan 2009 to Dec 2009. The gill parasites include two Monogeneas (Tetranclstrum nebulosi and Polylabris sp.), one Copepoda (Caligus sp.1). The intestinal parasite found was one Digenea (Oligolecithoides guangdongensis). Two Nematodas (Procamallanus sigani and Hysterothylacium aduncum). And two Acanthocephalas (Parahadinorhynchus sp. and Neoechinorhynchus sp.).The body surface parasites were two Copepodas (Caligus sp.2 and Clavellopsis sp.).
Molecular biology methods used for the identification of parasite species proved to be cost-effective, rapid and discriminatory. In this study, species identification principally based on morphological characteristics and tightly combined with molecular dates of 18S rDNA, 5.8S rDNA, ITS-1 rDNA and ITS-2 rDNA sequence. The whole or nearly whole length of the18S ribosomal DNA fragment sequences, 5.8S ribosomal DNA fragment sequences, ITS-1 and ITS-2 rDNA gene of ten species of parasites were amplified using PCR techniques.The results indicated that 5 speicies,9 families can be accurately identified in the level of genus only using either of the 18S rDNA sequence or 5.8S rDNA together with ITS-1 and ITS-2 sequence fragmet, each precision rate of the two reached 50 percentage and 90 percentage respectively in the level of genus and family. While 6 speicies, 9 families can be correctly identified in the level of genus by combining both the two sequence date with ITS-1 and ITS-2 and got percentage precision of 60 percentage and 90 percentage . Results certified that both 18S rDNA and 5.8S rDNA together with ITS-1 and ITS-2 as important indexs to analyse different genera of the family or between different family was cost-effective. Moreover, the later method still was suited to discriminate the closely–related taxa within genus. Only one speices was indentified in the level of species,not because of low degree of accuracy by our methods but the sequence dates of 18S rDNA, 5.8S rDNA, ITS-1 rDNA and ITS-2 rDNA were first found and the lack of dates of corresponding genus in GenBank at prensent.
Only two parasites T. nebulosi and Polylabris sp. obviously dispaly the seasonal population dynamics. The prevalence,the abundance and intensity of T.nebulosi population in the wild host increased from January, and attained peak in May then declined quickly in spite of being at a high leve in June, and it nearly reached to zero infection till October. The prevalence,the abundance and intensity tended to increase but still keep on a low level of infection since November.The prevalence,the abundance and intensity dynamics of Polylabris sp. population were very similar with the T.nebulosi population except the prevalence atained peak to shift to an earlier time in March,and embraced a small peak of infection respectively in September and November. Both two Monogenea presented aggregated frequency distributions in the host population.The sites selection of two monogeneain frapopulations on host were as follows. There were no statistically significant diferences for both of them covering the sides of gills, but every type of parasites shew noticeable preferences in the front two pairs of gills and only few distributed in the two back pairs of gills. The site preference in gill decreased from the 1st, 2nd ,3th to the 4th pairs in T.nebulosi, while the seuence1st, 2nd , 4th ,3th pairs in Polylabris sp.
As to the distributions of two parasites on the branchial plate,T.nebulosi generally occupiedⅣ,Ⅴ,Ⅵsectors and prefered gastro- to dorso gill. While Polylabris sp. distributed inⅦ,Ⅷ,Ⅸsectors and prefered dorso to gastro- gill. In addition, we found the prevalence,the abundance and intensity of both two Monogenea positively correlated with the host's length ,and the dynamic rule may mainly induced by the variation of sea water salinity and tempture.
引文
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