Etiology And Histopathology Study For White Porphyritic Ulcer Disease Of Hippocampus Japonicus

Ulcer disease will always occurs to the cultured Hippocampus japonicus, whichcould trigger skin ulcers, erosion of the liver and intestinal whitish. Meanwhile H.japonicus could all die without timely treatment. The paper maily research thepathogenic bacteria. We isolated the pathogenic bacteria from dying H. japonicus’slesion sites and classified pathogen bacteria by it’s morphological, physiological andbiochemical characteristics, and16S rDNA sequence analysis. With the experiment ofthe artificial infection, the infectious bacteria to the H. japonicus could be evaluated.Dominant strain HM16and HC2were found from the infected H. japonicus’s ulcer skin,liver and intestinal tract. A Gram-negative and rod shaped with flagellum under electronmicroscope. Strain HM16gave positive reactions for lipase, phospholipase, gelatinaseand catalase, but was negative for caseinase, VP test, M.R, Urease and amylase. In orderto confirm the identification result, strain HM16’s16S rDNA was amplified and blast inthe EzTaxon date bank. The result showed that strain HM16has the highest similarity toVibrio campbellii, with99.508%identity. Furthermore the test shows that HM16is verysensitive to ceftazidime, ceftriaxone sodium, streptomycin, chloramphenicol anderythromycin. The infection experiments prove that H. japonicus will be easily infectedof HM16. The mount of LD50, which could cause the death of H. japonicus is just7.9×102cfu/g. Strain HC2gave positive reaction for gelatinase, catalase and urease, butwas negative for VP test, M.R, amylase, lipase, phospholipase and caseinase. In order toconfirm the identification result, strain HC2’s16S rDNA was amplified and blast in theEzTaxon date bank. The result showed that strain HC2has the highest similarity toPseudomonas sp., with99.508%identity. Furthermore the drug sensitivity test resultshowed that strain HC2was highly sensitive to seven antibiotics （penbritin,carbenicillinNa2, piperacillin, ceftazidime, ceftriaxone sodium, neomycin andofloxacin）. The artificial infection experiments prove that H. japonicus will be easilyinfected of strain HC2. The mount of LD50, which could cause the death of H. japonicuswas just1.6×103cfu/ind.Antagonistic endophytic strains with strongly inhibitory activity to enteritis diseaseof H. japonicus were isolated from H. japonicus. We optimize the fermentationconditions of produce antimicrobial active substances.56strains were isolated from intestinal tract of healthy H. japonicus and examined for their antagonistic activitiesagainst pathogenic HC2by the dot-inoculating method. Strain HS17was identified bymorphological, physiological and biochemical characteristics, and16S rDNA sequenceanalysis. Via the choice of the optimal culture medium, orthogonal optimization ofmedia components, single factor test to confirm the optimization of fermentationconditions. Strain HS17was a antagonistic bacteria and the morphological,physiological and biochemical characteristics, and16S rDNA sequence analysis provedthat strain HS17belonged to Bacilus sp.. The optimized growth carbon source, nitrogensource and phosphate of strain HS17were phosphate, maltose and KH2PO4, respectively.The optimal fermentation conditions were liquid volume30mL in100mL flask,180r/min at28℃, inoculation size3%for48h with a medium of phosphate(20.0g/L),maltose(10.0g/L), KH2PO4(1.5g/L), NaCl(10.0g/L), MgSO4·7H2O(0.2g/L) at initial pHof7.0. Strain HS17shows stronger antagonistic action to pathogenic HC2underoptimized fermentation conditions．Histopathological study showed that the H. japonicus’s epidermis was completelycome off, connective tissuse serious deformation, and produced a large number of acidicsubstances. The tail of H. japonicus was ulcer, serious damage of cartilage, muscleserious deformation, nucleus concentration, and produced a large number of acidicsubstances. The pigment layer of skin of H. japonicus falled off, and the yellow pigmentcells and red pigment cells were disappeared in the dermis. The liver’s blood vessel wasruptured, appeared red blood cells infiltration and a large number of vacuoles, freednucleus and produced a large number of acidic substances.