Study On Detection Of Molecular Typing,biofilm And Transcriptomics Of Staphylococcus Aureus Isolated From Cow In Ningxia

Staphylococcus aureus is one of the important pathogenic bacteria that can cause diseases in human and animal,and it is also the main pathogenic bacteria causing clinical and subclinical mastitis in dairy cows.S.aureus can produce a variety of toxins and invasive enzymes.With the widespread and improper use of antibacterial drugs,the resistance of S.aureus increased,which has exacerbated the occurrence of diseases and increased the cost of treatment.S.aureus has multiple resistance mechanisms.The formation of biofilm can increase bacteria toxicity and the resistance of antibacterial drugs,and increase tolerance to nutritional deprivation,so many persistent infections are caused,making treatment more difficult.Therefore,it will help to find effective anti-S.aureus biofilm drugs and targets and it is of great significance for the prevention and treatment of infections caused by S.aureus to study the formation and regulation mechanism of biofilms.1.A micro-semi-quantitative method was used to detect 234 strains of bovine-derived S.aureus in Ningxia,combined with a variety of microscopes to observe the biofilm and its formation process,and the surface hydrophobicity of bacteria was analyzed by MATH test.The results showed that 185 strains of S.aureus could form a biofilm with a positive rate of 79.06%.The BF-negative strain with hydrophilicity was higher than the BF-positive strain.As the adhesion of BF increases,the hydrophilicity of bacteria gradually decreases and the hydrophobicity gradually increases.With the enhancement of the biofilm formation ability,the biofilm is gradually thickened,and with the increase of culture time,its structure becomes more and more compact.2.According to the broth microdilution method recommended by the CLSI,the MICs for the S.aureus of two states 15 antibacterial drugs were determinated.The results showed that planktonic S.aureus had the highest resistance to penicillin,reaching 94.87%,followed by the resistance rate to sulfisoxazole(82.05%)and ampicillin(85.90%),and was completely sensitive to vancomycin and telmicoxin.In the state of biofilm,the resistance rate of S.aureus to vancomycin was 34.37%,the resistance rates to the remaining 14 antibacterial drugs reached more than 90%.The MIC value of S.aureus in the biofilm state increased by 4-1024 times compared with S.aureus in the planktonic state.3.The PCR technology was used to amplify 15 biofilm-related genes.The results showed that the detection rates of clfA,clfB,sigB,atlE,fib,ebps and icaD reached more than 90%,the detection rates of fizbpB,bap and sarA were low,and no bbp gene was detected.The detection rates of atlE,icaA,bap,fnbpB,icaD and SaG genes of biofilm positive strains were significantly higher than that of negative bacteria(P<0.05).4.The molecular epidemiological characteristics of S.aureus from cow in Ningxia were analyzed using RAPD,agr and spa typing techniques.The results showed that 23 strains did not amplify the bands in RAPD typing,and the remaining 211 strains of S.aureus were divided into 6 types,mainly type ?,accounting for 29.06%.Agr typing is mainly agr I,accounting for 61.11%.234 S.aureus were divided into 20 spa types and the main type is t224,followed by t518,t359,t519 and t5100.5.The RNA-Seq sequencing technology was used to study the differences between planktonic state and biofilm state of S.aureus from the transcription level.The results showed that there were 1512 gene expression differences between the two states of bacteria,of which 760 genes were significantly up-regulated and 752 genes were significantly down-regulated.The significantly different genes were annotated to 1,825 GO terms,and the annotated genes were related to 79 pathways.In summary,this study investigated the molecular epidemiology,and detected the biofilm and drug resistance of cow mastitis in Ningxia region.Simultaneously,the differentially expressed genes in the two states were compared,the genes and pathways related to biofilm formation were excavated,which can establish the foundation for clarifying the mechanism of biofilm formation and regulation of S.aureus.