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To elucidate the localization of oomycetes (water molds) and bacteria in a single egg of chum salmon, Oncorhynchus keta, eggs infected with oomycetes were examined by bright-field microscopy, scanning electron microscopy (SEM), and fluorescent in situ hybridization (FISH) using oomycete universal and bacterial species-specific probes. Furthermore, DNA barcoding for oomycetes and bacteria was performed on the chorion and cytoplasm of the eggs using genomic DNA extracted from the sections of eggs used for FISH analyses. Using bright-field microscopy and SEM, it was shown that oomycete hyphae penetrated the chorion and invaded the cytoplasm of the eggs. Based on DNA barcoding, two oomycetes (Pythium monospermum and Pythium sp. SE-OP1), and three bacterial species (Flavobacterium plurextorum, Flavobacterium sp. SE-BF1 and Undibacterium pigrum) were predominantly detected in the infected eggs. FISH analyses indicated that localization of the oomycetes was not noticeable different between the chorion and cytoplasm of the eggs. In contrast, FISH analyses using bacterial species-specific probes demonstrated clear differences in their localization. Flavobacterium sp. SE-BF1 and U. pigrum were predominantly found inside the chorion and around the oomycete hyphae invading the cytoplasm, respectively. These results are the first to indicate the possibility that U. pigrum has a specific relationship with oomycetes and coordinately invades eggs with oomycetes. However, it seems that Flavobacterium sp. SE-BF1 invades by a mechanism different from that of U. pigrum. Although this study is preliminary and the interaction between oomycetes and U. pigrum associated with chum salmon egg infection needs to be elucidated in detail, this finding may provide new insights into the infection mechanism of bacteria in salmon eggs, which is a longstanding problem in salmon hatcheries. |
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