Identification of marine fungus HL-3 with activity against Vibrio parahaemolyticus and separation of its active substances

Abstract:

[Background] At present, the prevention and control of aquatic pathogens such as Vibrio parahaemolyticus in aquaculture mainly rely on antibiotics, which are effective in a short term. However, long-term use of antibiotics causes the enhancement of drug resistance in pathogens and other problems. Therefore, it is urgent to explore safe and effective antibiotic alternatives. [Objective] To screen marine microorganisms with anti-V. parahaemolyticus activity, identify the species, optimize the culture conditions, and preliminarily separate the active substances. [Methods] Marine microorganisms were isolated and purified with spread plate method and streak plate method, and the strains with anti-V.parahaemolyticus activity were screened with Oxford cup method. The active strains were identified based on the morphological characteristics and ITS sequences. The medium type and salinity were tested to optimize the culture conditions. The compound was prepared by semi-preparative high performance liquid chromatography, and the structure of the compound was identified based on nuclear magnetic resonance spectroscopy data. [Results] A total of 76 strains were isolated from 11 types of samples, such as conch, little yellow croaker, and prawn, including 26 fungal strains. A fungal strain HL-3 with anti-V.parahaemolyticus activity was screened out and identified as Aspergillus flavus. strains HL-3 produced diverse metabolites and showed strong antibacterial activity in the modified Sabouraud’s medium. It synthesized only one metabolite and demonstrated high antibacterial activity on fungus No. 5 medium. The metabolite was identified as kojic acid, and the minimum inhibitory concentration of kojic acid against V.parahaemolyticus was 4.0 mg/mL. [Conclusion] The experimental results lay a theoretical basis for the further separation and purification of secondary metabolites of strains HL-3 against V.parahaemolyticus with various chromatographic methods.

Key words:marine fungus;Vibrio parahaemolyticus;antibacterial activity;strain identification;culture conditions