Studies On Establishment And Characteristics Of Fusants Of Aerompunctta F. Intestinalis And Myxococcs Piscicola Sp.nov

This experiment used Aeromonas punctta f. intestinalis 58-20-9 strains and Myxococcs piscicola sp.nov G4 strains to operate as arch-strains to conduct protoplast fusion,established fusants and studied their morphological configuration physiologic biochemical,enzyme character and pathogenicity,antigenicity and so on . The main results were presented as follows:1. Screening of fusants' genetic markers:Using the qualitative and quantitative antimicrobial susceptibility experiment to screen the genetic markers of protoplast fusion between Aeromonas punctta f. intestinalis 58-20-9 strains and Myxococcs piscicola sp.nov 64 strains,they are Emr(Erythomycin resistance) and Gmr(Gentamicin resistance) respectively . In this protoplast fusion experiment between two bacteria,the applying density in selective culture medium of Erythomycin and Gentamicin are 3000 lu / ml and 50 lu / ml.2. Establishment of fusants:When the temperature of enzymolysis was 37C,we singled out the best enzymolysis time to be 40 minute and the best lysozyme density to be 4 mg/ml . In the condition of PEG operating as agent of accelerating fusion,Aeromonas punctta f. intestinalis 58-20-9 strains and Myxococcs piscicola sp.nov G4 strains conducted protoplast fusion,the protoplast formation rates of Aeromonas punctta f. intestinalis 58-20-9 strains are 54.7 % and the protoplast regenerative rates of 58-20-9 strains were 18.2%,the protoplast formation rates of Myxococcs piscicola sp.nov 64 strains were 50.8% and the protoplast regenerative rates of 64 strains were 12.2% . Arch-screening protoplast fusion rates were 4 . Breeding respectively 500 fusants colony of arch-screening on duplexing repel select culture medium and generating continuously 12 times,derived a steady fusant conoly . The fusants steady rates were 0.2% .3. Judging of the characters of fusants on morphological and configuration physics and chemical:The screened fusants are short rhabditiform . they are shorterthan 64 strains,but more grossness than 58-20-9 strains .They are near round .and their average macro-axis are 5.60um,average brachy-axis are 0.67um,average volume are 1.34um3 .Gram female,no flagellum,no motion .The colour of giga-colony which was cultured 30 days is scream white and have brilliancy,the colony's center is thicker than rim .The colony is bulging and it's rim is orderliness .The fusants' characters of enzyne and physiologic and biochemical exist between 58-20-9 strains and 64 strains .4. Studies on pathogenicity and antigenicity:Using fusant strains to infect manually to health grass carps which were born in just that year .After observing two weeks,the mortality rates of grass carps were 95%,but the control group's survival rates were 100%,the results showed that the fusant strains also had pathogenicity .Examined the disease and dead grass carps further,we found that the grass carps which were infected fusant strains had part symptom not only AKromonu.s punctta f. intestinalis 58-20-9 strains but also Myxococcs piscicola sp.nov G4 strains . Studied on antigenicity of fusant strains,the results showed that after the grass carps which were immunized by fusant strains infecting 58-20-9 strains and G4 strains,their survival rates run to 76.6% and 80.0%,ralative percents survival were 74.9% and 75.5%,valency of antibody upmost to 1:322.5 and 1:435.5 on the fourth week.In summary,under the present conditions,Aeromonas punctta f. intestinalis 58-20-9 strains and Myxococcs piscicola sp.nov G4 strains protoplasts could fusion and form steady fusants,the fusants also have two parents' pathogenicity and antigenicity .