New Insights into the Morphological Diversity of Saprolegnia parasitica (Oomycota) Strains under In Vitro Culture Conditions

Abstract

Saprolegnia parasitica Coker, 1923 is a primary fish pathogen and one of the most common water molds in freshwater ecosystems. In our study, nineteen strains of S. parasitica were isolated, identified, and characterized using morphological and genetic markers. On the basis of the abundance of zoosporangia, gemmae, the formation of gemma chains, and the induction of zoospore release, three morphotypes were differentiated. A species-level molecular identification of isolates was performed using the ITS 1 and 2 regions. A total of six genotypes were distinguished based on partial DNA sequences of the genes RNA polymerase II subunit B (RPB2) and serine hydroxymethyltransferase (SHMT). In five settings of in vitro culture conditions differing in the mineral content and the temperature of water and in the presence of a host or bait, we found that the addition of fish skin extract boosted the formation of asexual reproductive and persistent vegetative structures in cultures, whereas an unfavorable environment did not support the formation of these structures in vitro.

Description of Saprolegnia velencensis sp. n. (Oomycota), a novel water mold species from Lake Velence, Hungary

“…Strains SAP239 and SAP241 maintained on GY+P+S agar medium were sub-cultured under four different in vitro conditions at two temperatures (+20–21°C vs. +6–7°C) and at different pH (pH 6.0–7.2 vs. pH 9.0) to increase the chance of obtaining reproductive structures for morphological characterization [ 17 ]. Four-mm agar plugs were cut from the plates with sterile biopsy punch, and one plug per well was placed into 24-well plates in 1.5 mL volume of different medium in triplicates.…”

“…Mold swabs from each well were regularly examined under light microscope (Zeiss AxioStar Plus, Zeiss). Zoospore production was induced using the method described by Erdei et al [ 17 ]. Colonized agar plugs were incubated at 24-well plates in 1.5 mL GY+P+S broth for three days.…”

“…oogonia and antheridia) in vitro, which would be necessary for the morphology-based identification [5,10,14]. Some studies have suggested that changing favorable and unfavorable conditions can induce the production of sexual reproductive structures of Saprolegnia strains in vitro, however it could not be applied successfully in every case [11,[15][16][17].…”