Isolation, identification and pathogenicity of Nocardia seriolae from Micropterus salmoides

Abstract:

[Background] Nocardia seriolae is the causative agent of chronic granulomatous disease in cultured fish characterized by skin ulcers, bleeding and nodular formation in tissues, which seriously harms aquaculture industry. So far, there is no effective prevention and treatment method for the disease. [Objective] To clarify the pathogen causing sarcoidosis of Micropterus salmonoides in an aquaculture farm in Linquan county, Anhui province and to explore its pathogenicity, so as to provide a scientific basis for the effective prevention and treatment of the disease. [Methods] The liver nodules were inoculated with TSB medium to isolate dominant bacterial strain, which was then identified by phenotypic examination combined with test of molecular biology. The pathogenicity of the isolated strain was analyzed through virulence gene identification, LD₅₀ to Micropterus salmonoides as well as the histopathological changes and tissue bacterial load. [Results] A dominant strain NI was isolated from diseased fish and identified as N. seriolae based on its phenotypic characteristics, the 16S rRNA gene sequence identity between the strain NI and N. seriolae reference strains, and the result of specific PCR amplification. Strain NI carried virulence genes gapA, ibeA and mip, and its LD₅₀ to Micropterus salmonoides was 2.58×10⁶ CFU/fish. The clinical signs of Micropterus salmonoides after artificial infection with the strain NI were in accordance with those of natural cases. Histopathologically, chronic granulomatous lesions in head kidney, heart, liver, stomach and spleen of diseased fish, loose intestinal muscle layer and shortened intestinal villi, loose and widened muscle fibers in the muscle tissue were observed. qPCR analysis found the descending order of N. seriolae load in the following infected tissues: head kidney, heart, liver, stomach, spleen, intestine and muscle. [Conclusion] Nocardia seriolae is the pathogen causing the outbreak of sarcoidosis in Micropterus salmonoides in the local aquaculture farm. Results of the study on pathogenicity of the isolated strain provided a theoretical basis for the prevention and control of nocardiosis in Micropterus salmonoides.