In Vitro Inhibition of Saprolegnia sp. by an Antifungal Peptide from Pseudomonas protegens XL03

Abstract

Saprolegniasis, a fungal fish disease, is an important challenge in the aquaculture industry. To find a natural agent to control saprolegniasis, an anti-Saprolegnia bacterium, named XL03, was isolated from the sediment of the Baimang River in Xili, Guangzhou, China. The bacterium was identified as Pseudomonas protegens by means of 16S rDNA gene sequence and its fluorescent character. Saprolegnia sp. was used to evaluate the antifungal activity of the P. protegens strain XL03. The cell-free disruption supernatant (CFDS) showed an antifungal zone of 19.34 ± 1.20 mm against the Saprolegnia hyphae on the potato dextrose agar (20% potato extract, 2% glucose, 2% agar) medium plate and depicted a minimum inhibitory concentration (MIC = 12.5 mg/mL) against the Saprolegnia spore germination in vitro. To purify the antifungal substance from the CFDS, ammonium sulphate precipitation, gel filtration, and ion-exchange chromatography were applied, and the purified peptide exhibited a single band at molecular weight about 58 kDa on a SDS-PAGE gels. Furthermore, the MIC of the purified peptide was identified as 0.0625 mg/mL against Saprolegnia spores, and the activity of peptide showed thermal stability between 30°C and 60°C but retained activity only in a narrow pH range of 5.0–9.0. These findings demonstrate that P. protegens strain XL03 has the potential to be used in biocontrol of saprolegniasis in aquaculture.

Received June 21, 2016; accepted October 24, 2016 Published online February 23, 2017

ACKNOWLEDGMENTS

This work was supported by Science and Technology Project of Guangdong Province (grant number: 2014A020208024) and the Marine and Fishery Special Project of Fish Diseases Treatment and Prevention in Guangdong Province (YueCaiNong [2016] number 11), as well as the Marine and Fishery Special Project of Science and Technology in Guangdong Province (grant numbers: A201301B05 and A201501B09).