| The bacterial diseases of silkworm or called flacherie outbreaks frequently and often make great damage to sericultural industry. On the basis of pathogenesis flacherie could be divided into three groups: septicemia, toxicosis, and gastroenteric disease. To date, a wide variety of bacterias were found pathogenic to silkworms, including Bacillus, Enterobacter, Serratia, Aeromonas, Streptococcus, Pseudomonas, and Staphyloccus .In this study, a pathogenic bacterial strain, ST-1, was isolated from a naturally infected silkworm. Color of the strain's colony was similar to that of a particular aurantium. ST-1 bacteria was gram-negative and 0.70.9×1.31.5μm long, short rods with rounded ends. And it was motile. Physiological and biochemical identification tests of the ST-1 strain were prepared. The results showed that strain ST-1 was an aerobic bacteria which its peroxidase reaction was positive. It could impel nitrate deoxidization, and make gluten liquefactions. Voges-Proskauer reaction, Methyl Red test, Indole test, H2S production and Phenylalanine deaminase of strain ST-1 was negative. Also it could utilize sodium citrate, malonate, D-glucose, sucrose, D-fructose, D-mannose, and L-arabinose. The sequence analysis of its 16S rRNA gene was used to further research. Finally ST-1 strain was identified as Ps. chlororaphis subsp. aurantiaca. The symptoms preceding fifth instar larvae of silkworm death appeared to be consistent with those of bacterial septicemia. Pathogenicity of ST-1 for silkworm could be depicted as a linear regression of the logarithm (y) of ST-1 concentration against probability (x) (y = 0.4040 + 0.0600x). The median lethal concentration (LC50) was 2.12×104 cfu/mL. This is the first strain of P.aurantiaca which has been identified as againsting the silkworm (Bombyx mori).The pathogenic fungi of silkworm yellow muscardine was once thought to belong to the species Stilbaceae, Isaria, Isaria farinasa. But the silkworm white muscardine(Beauveria bassiana) was recently classified in species Ascomycota, Sordariomycetes, Hypocreomycetidae, Hypocreales, Cordycipitaceae, Cordyceps. Recent studies suggested that the pathogen of yellow muscardine had only minor morphological differences with Beauveria bassiana. Most researchers believed that its type should be Beauveria bassiana and only had diverse serotype.In our study, yellow muscardine Strain STH1 was isolated from a naturally infected silkworm. Most STH1 conidia were sphere, and smaller than Beauveria bassiana SHB. To observe the growth diameter of colony, STH1 was grown on PDA plate for 12 days. Strain STH1 was 4.25±0.81 mm·d-1, and SHB was 4.75±1.00 mm·d-1. This meaned that the grower rate of STH1 was slower than that of SHB. After 15 days, the conidia output of strain STH1 was 9.82±3.49×108spores/cm2, and SHB was 4.01±2.12×108spores/cm2. Theα=0.01 which means there are extremely significant difference between the grower rate two kinds of strains.The ITS gene of strain STH1 and SHB was all 549 bp, and had high similarity with eachother (equal to 100%). Phylogenetic tree showed that STH1 was a member of Beauveria bassiana clade. The complete sequence of 18S rRNA gene of STH1 was 1 766 bp and a insertion sequence of group I introns (1 866 bp) encode homing endonucleases within the gene. The phylogenetic analyse of 18S rRNA, 28S rRNA and IGS gene of STH1 and SHB indicated that they had approximately 100% similarity. So we believed the pathogenic fungi STH1 was from the species Cordyceps bassiana, and it was a novel strain that contains group I introns.Study on the phylogenetic identification and characteritions of strain ST-1 and STH1, could define their taxonomic status, and lay a foundation for the prevention and therapy of diseases caused by ST-1 and STH1. |
