Kumar Gokhlesh, Abd-Elfattah Ahmed, El-Matbouli Mansour
Clinical Division of Fish Medicine, Department for Farm Animals and Veterinary Public Health, University of Veterinary Medicine, Veterinärplatz 1, 1210, Vienna, Austria.
Parasitol Res. 2015 Mar;114(3):929-39. doi: 10.1007/s00436-014-4258-1. Epub 2015 Jan 7.
Tetracapsuloides bryosalmonae Canning et al., 1999 (Myxozoa) is the causative agent of proliferative kidney disease in various species of salmonids in Europe and North America. We have shown previously that the development and distribution of the European strain of T. bryosalmonae differs in the kidney of brown trout (Salmo trutta) Linnaeus, 1758 and rainbow trout (Oncorhynchus mykiss) Walbaum, 1792, and that intra-luminal sporogonic stages were found in brown trout but not in rainbow trout. We have now compared transcriptomes from kidneys of brown trout and rainbow trout infected with T. bryosalmonae using suppressive subtractive hybridization (SSH). The differentially expressed transcripts produced by SSH were cloned, transformed, and tested by colony PCR. Differential expression screening of PCR products was validated using dot blot, and positive clones having different signal intensities were sequenced. Differential screening and a subsequent NCBI-BLAST analysis of expressed sequence tags revealed nine clones expressed differently between both fish species. These differentially expressed genes were validated by quantitative real-time PCR of kidney samples from both fish species at different time points of infection. Expression of anti-inflammatory (TSC22 domain family protein 3) and cell proliferation (Prothymin alpha) genes were upregulated significantly in brown trout but downregulated in rainbow trout. The expression of humoral immune response (immunoglobulin mu) and endocytic pathway (Ras-related protein Rab-11b) genes were significantly upregulated in rainbow trout but downregulated in brown trout. This study suggests that differential expression of host anti-inflammatory, humoral immune and endocytic pathway responses, cell proliferation, and cell growth processes do not inhibit the development of intra-luminal sporogonic stages of the European strain of T. bryosalmonae in brown trout but may suppress it in rainbow trout.
四极虫（Tetracapsuloides bryosalmonae）Canning等人，1999年（粘孢子虫纲）是欧洲和北美的各种鲑科鱼类中增殖性肾脏病的病原体。我们之前已经表明，欧洲株四极虫在褐鳟（Salmo trutta）林奈，1758年和虹鳟（Oncorhynchus mykiss）瓦尔鲍姆，1792年的肾脏中的发育和分布有所不同，并且在褐鳟中发现了管腔内孢子生殖阶段，而在虹鳟中未发现。我们现在使用抑制性消减杂交（SSH）比较了感染四极虫的褐鳟和虹鳟肾脏的转录组。SSH产生的差异表达转录本被克隆、转化并通过菌落PCR进行测试。使用斑点印迹法验证PCR产物的差异表达筛选，并对具有不同信号强度的阳性克隆进行测序。对表达序列标签的差异筛选和随后的NCBI-BLAST分析揭示了两种鱼类之间有9个克隆表达不同。这些差异表达基因通过在感染的不同时间点对两种鱼类的肾脏样本进行定量实时PCR来验证。抗炎（TSC22结构域家族蛋白3）和细胞增殖（前胸腺素α）基因的表达在褐鳟中显著上调，但在虹鳟中下调。体液免疫反应（免疫球蛋白μ）和内吞途径（Ras相关蛋白Rab-11b）基因的表达在虹鳟中显著上调，但在褐鳟中下调。这项研究表明，宿主抗炎、体液免疫和内吞途径反应、细胞增殖和细胞生长过程的差异表达不会抑制欧洲株四极虫在褐鳟管腔内孢子生殖阶段的发育，但可能会在虹鳟中抑制它。
