| Tigecycline(Tgc)is the first FDA-approved glycylcycline antibiotic,recognized as a "last resort" antibiotic to treat multidrug-resistant(MDR)bacterial infections,eapecially in complicated intra-abdominal infections,skin infections,and community-acquired pneumoniae.But more and more Tgc-resistant(Tgc-R)bacteria and infections were reported in recent years,raising a great challenge and risk hindering the optimal clinic treatment outcome.FDA approved Tgc to use in clinic in 2007,and Tgc was introduced into Chinese hospitals in late 2012.Bacteria in this research were retrived from clinic lab-reported Tgc-R A.baumannii in a tertiary hospital since late 2012 after Tgc was used in clinic.A.baumannii is an important opportunistic pathogen for hospital-acquired infections that may due to the advantages of its ability to rapidly acquire resistance genes,and survive on dry surfaces,with microcide or exposure to radioactivity.According to Mohnarin,the isolation rate of clinic MDR A.baumannii has been over 50%in the past five years,and the number of extensively drug-resistant(XDR)and pandrug-resistant(PDR)A.baumannii is increasing.The present study investigated the molecular epidemiology of clinic A.baumannii strains,and through a combined comparative genomic and transcriptomic approach,genomic variations as well as transcriptomic regulations related with the development of bacterial Tgc resistance were idientified.Moreover,the antibiotic resistance genes(ARGs)and virulence factors(VFs)of Tgc-R A.baumannii were comparatively analyzed and their relationship was investigated.In the first chapter of the thesis,the clinic lab-reported Tgc-R A.baumannii isolates were collected in the main lab and measured for their in vitro minimal inhibition concentrations(MICs)of Tgc by broth microdilution method.Their sequence types(STs)were examined by multilocus sequence typing(MLST)method and Eric-PCR.Totally 19 Tgc-R and 37 Tgc-intermediate(Tgc-I)A.baumannii strains were identified.The according clinic information including sample types,wards,age and sex of patients,and antibiotic usage history were retrived and analyzed,to reveal molecular and clinical characteristics of Tgc non-susceptible A.baumannii.This part of work also provided Tgc non-susceptible strains for comparative genomic studies in the second chapter.In the second chapter of the thesis,totally 6 A.baumannii strains were selected from three patients,and strains within each group(patient)were isolated at different times during antibiotic treatment,and sharing the same STs but with different Tgc-resistance level.The strains’ genomes were sequenced and comparatively analyzed for their phylogenetic relationship,genomic sequence and structure variations,and functional characteristics.Importantly,a Tgc-R strain may result from a putative homologous recombination event between two strains isolated in the same patient but with lower Tgc resistance.The shifted genomic region included six adjacent genes including OprD,a sugar-type MFS permease,a LuxR-type transcriptional regulator and a hypothetic membrane protein.Then the strains’ transcriptomes were collected under different Tgc concentrations(0μg/mL,0.5μg/mL and 1μg/mL)and comparatively analyzed to identify the "significantly differential-expressed genes".The data was further verified by semi-quantitative qRT-PCR.Overall,genomic mutations as well as transcriptional regulations may together play important roles in the development of Tgc resistance in A.baumannii.Meanwhile,genomes of 19 Tgc-R A.baumannii strains were comparatively analyzed for genomic plasticity,distribution of ARGs and virulence factors(VFs),and phylogenetic relationship.It was found that the 19 Tgc-R A.baumannii strains were phylogenetically close despite of different STs,with various ARGs and VFs.By copy number variations of ARGs or VFs,the 19 Tgc resistant strains can be divided into four groups.Different groups of strains,and other GeneBank-retrieved A.baumannii isolates,can be differentiated by marker genes. |
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