Detection of the intranuclear microsporidium Nucleospora salmonis in naturally and experimentally exposed Chinook salmon Oncorhynchus tshawytscha by in situ hybridization.

International audienceNucleospora salmonis, an intranuclear microsporidian parasite of salmonid fish, is often difficult to observe in histological sections or wet mount preparations from lightly infected tissues because of its small size and location within the nuclei of lymphoblast-type cells. Diagnosis of infections by conventional light microscopy is directly dependent upon distinguishing different stages of the parasite from host cell nuclear material or vacuoles. To assist detection of stages of the parasite in tissues of its primary host, the Chinook salmon (Oncorhynchus tshawytscha), we developed a nonradioactive in situ hybridization (ISH) method. The new method was then used to detect N. salmonis among Chinook salmon after both natural and experimental exposures to the parasite. Probes derived from the small subunit ribosomal DNA (ssu-rDNA) sequence of the microsporidium were labeled with digoxigenin deoxyuridine triphosphate (DIG-dUTP) and hybridized to parasite DNA present in infected tissues. The ISH procedure effectively identified merogonic and spore stages of N. salmonis in paraffin-embedded tissues of clinically and subclinically infected fish. A Nucleospora-like microsporidium was also detected by ISH in tissues of a nonsalmonid fish, the English sole (Pleuronectes vetulus), using probes designed to a region of the ssu-rDNA of N. salmonis

In situ hybridization: A detection tool for fish pathogens and its application on recent advances on whirling disease research

A non-radioactive in situ hybridization (ISH) protocol was used as a diagnostic tool for several fish pathogens including Candidatus Xenohaliotis californiensis, the etiologic agent of withering syndrome, which is a new disease in wild and cultured abalone Haliotis spp., the microsporidian Nucleospora salmonis and the myxozoan parasite Tetracapsula bryosalmonae (previously referred to as PKX) that causes proliferative kidney disease in salmonids. Most applications of the ISH protocol in our -laboratory were used in pathogenesis studies of whirling disease caused by the myxosporean Myxobolus cerebralis