Isolation, Identification And Biological Characteristics Of A Pathogenic Saprolegnia Sp. From The Crucian Carp Egg

Saprolegniasis is one of the most serious diseases in artificial propagation of aquatic animals and often causes significant economic losses. It is significant and necessary to find a suitable way of controlling the disease. A pathogenic strain JL1 was isolated and identificated by characteristics of morphological and molecular biological. Then effect of three selective drugs against saprolegnia on different developmental stages of saprolegnia was studied, and effect of preparation on the control eggs saprolegnia was tested, which was selected from three drugs. The main results are as follows:1 Identification and biological characteristics of a pathogenic saprolegnia sp. from the egg of Pengze Crucian carpThree filamentous fungal strains were isolated form Carassius auratus eggs with Saprolegniasis, and strain JL1 was proved to be pathogenic to Carassius auratus eggs by artificial infection. Therefore, morphology and growth characteristics of strain JL1 were studied, and the phylogenetic analysis based on its ITS rDNA sequence was further conducted. The experimental results showed that the hyphae of strain JL1 were aseptate, transparent and seldom branched. Its zoosporangia were often clavate and renewed internally. Primary zoospore was multi-row arrangement in zoosporangia and discharged in Saprolegnia fashion. Spherical oogonia were attached by monoclinous or diclinous antheridium hyphae.The ITS rDNA sequences of strain JL1 was naturally clustered with ITS rDNA sequences of Saprolegnia sp. submitted to GenBank with 99% of homology, and had closest relationship with S. diclina,S. ferax,S. longicaulis. Combined morphological characterization with phylogenetic analysis based on ITS rDNA sequence, strain JL1 was identified as Saprolegnia sp..2 The effect of temperature and nutrition on production and germination of strain JL1 zoosporeThe results show: strain JL1 could grow at 5°C-30°C and pH 4-11, its optimum growing temperature and pH were 25°C-30°C and 6-9, respectively. Strain JL1 was sensitive to sodium chloride, its growth could be completely inhibited by 2% NaCl, which could serve as a foundation for the Saprolegniasis control.Strain JL1 could produce zoospores at 15°C-30°C, and can't at 35°C because of mycelium growth inhibited. The optimum temperature range is 20°C-25°C. The number of zoospores reached peak at 25°C after 36h cultured in water, while at 20°C after 48h. The zoospores of Strain JL1 could germinate at 15°C-30°C and couldn't at 35°C. The optimum temperature range was 20°C-25°C. The mycelium of strain JL1 grew vigorously and zoospores didn't produce in nutrient-rich conditions, while nutrients could promote the germination of zoospores.3 The effect of 3 selected drugs on different developmental stages of strain JL1The effect of 3 drugs obtained by screening on different developmental stages of strain JL1 was tested. The results show: Zoospores were more sensitive than hypha to 3 drugs. The minimum inhibition concentration of compound H on mycelium growth, sporangium formation, zoospore discharge, zoospore swimming and germination were 10 mg/L,5 mg/L,10 mg/L,2.5 mg/L,5 mg/L respectively. The minimum inhibition concentration of Pseudolarix extract were 5 mg/mL,1 mg/mL,5 mg/mL,5 mg/mL,2.5 mg/mL respectively. The minimum inhibition concentration of Methylene bisthiocyanate (MBT) on mycelium growth, sporangium formation, zoospore discharge and germination were 1 mg/L,0.5 mg/L,1 mg/L,0.5mg/L respectively. The effect of MBT on zoospore swimming was weak.4 The prevention and treatment effects of anti-Saprolegnia drug on fish eggsConsidering all the factors, such as effectiveness, security, cost, process, etc. compound H was taken as the major component of preparation H, adding appropriate synergists, fillers, dispersant, etc. The result of experiment of preparation H on carp eggs showed that the hatching rate and survival rate of carp eggs had decreased slightly in the concentration of 50 mg/L, while the hatching rate and survival rate of carp eggs had no significant differences(p<0.05) in the concentration of 20 mg/L. The moldy rate had reduced and the hatching rate had improve by using preparation H with short-term exposure at 100mg/L once 12h and 3 consecutive times. The moldy rate had reduced and the hatching rate had improve by using preparation H with prolonged exposure to eggs at 15mg/L once a day and 3 consecutive times.