The Degradation Genes Of Polyvinyl Alcohol Degrading Bacteria QL-P4 And Optimization Of Degradation Conditions

Polyvinyl alcohol（PVA）is a superior water-soluble polymer material as it has been widely used and produced in industry.Consider the biodegradability of PVA is very limited,PVA effluent has caused increasingly serious environmental issues such as foam and viscosity increasing of the water,have adverse effect on aerobic microorganism activity and so on.Therefore,research of isolating and domesticating high-efficiency PVA degradation strains and studying their degradation pathway have theoretical and practical meaning to remediate contaminated water.This study takes a PVA degrading bacteria Stenotrophomonas rhizophila QL-P4 as the object.By studying its species identification,degradation system optimization,whole genome sequencing,gene knockout and phenotypic validation,PVA biodegrading bacteria resources has been expand,which establishes the foundation of exploitation,using and degrading mechanism of this bacteria.The main results of this research including:（1）Strain QL-P4 colony on LB medium presents a rounded and convex shape,with light yellow color,opaque,neat edge and smooth surface.The morphology observation under scanning electron microscope of strain QL-P4 showed typical bacilliform cells with the surface of irregular wrinkles,length is about 1.65μm,diameter is about 0.45μm.Combined with physiological,biochemical tests and 16S r RNA sequencing（Gen Bank NO.KT935058）and genome sequencing（Gen Bank NO.CP016294）,identified strain QL-P4 belongs to Stenotrophomonas rhizophila.（2）Through the lethal concentration detection of strain QL-P4 with different antibiotics,it was found that strain QL-P4 is resistant to tetracycline,Spectinomycin dihydrochloride,ampicillin and penicillin.And has no resistance to erythromycin,chloramphenicol,gentamicin and kanamycin（lethal concentration is 50mg/L）.（3）The optimal conditions for PVA biodegradation of strain QL-P4 were 25℃,1.0g/LPVA as the sole carbon source,p H7.0,and inorganic nitrogen source as Na NO₃and NH₄NO₃ each added 0.2g/L.（4）Building the co-culture system by strain QL-P4 with strain HK1 which was a PVA degrading bacteria isolated previously in our lab.And the co-culture system didn’t reveal better degradation ability as we expected.（5）Based on whole genome sequencing and alignment results,eight gene knock-out engineering bacteria was built.And six degrading-related genes（BAY15＿3292,BAY15＿3123,BAY15＿3143,BAY15＿0976,BAY15＿0291,BAY15＿0160）got its knock-out mutant.AndΔ3292＿QL-P4,Δ0160＿QL-P4 andΔ0291＿QL-P4 has significant difference with wild type QL-P4 on PVA degradation ability.All these results show that PVA degrading bacteria QL-P4 has a tremendous applicable value for its excellent biodegradability and mild culture conditions.The result of degrading genes’knock-out has not only verified degrading pathway which was predicted but also provides theoretical support for improve degrading ability on molecular level.