Research Of Resistance Mechanism Of Pseudomonas Aeruginosa For Aminoglycosides And Analysis Of Homology For Pseudomonas Aeruginosa

Ojective:1. To explore drug-resistant phenotype of pseudomonasaeruginosa for aminoglycosides as well as distribution and relationship ofAminoglycosides modifying enzymes.2. Aminoglycoside resistant strains weretyped to monitor the prevalence of resistant strains so that antibiotics can beused rationally and hospital infection can be controled scientificly. Methods:1.Pseudomonas aeruginosa which were isolated clinically were collected fromSeptember2011to June2012.2. Pseudomonas aeruginosa were identified byMicroScan WalkAway96SI automated microbial identification/susceptibilitytest system and resistance of pseudomonas aeruginosa to15antimicrobialagents were obtained at the same time. In addiation, aminoglycoside resistancestrains were screened.3. Polymerase chain reaction was used to detect cases ofsix kinds of aminoglycoside modifying enzyme genes foraminoglycoside-resistant pseudomonas aeruginosa strains.4. The positive geneamplification products were sent to Shanghai Sangon to sequence. Thesequencing results were compared with the GenBank database.5. Multi-genecluster analysis was carried out based on molecular markers of sixaminoglycoside modifying enzyme genes in order to understand the prevalenceof pseudomonas aeruginosa in our hospital.6. Molecular typing forpseudomonas aeruginosa was analysed by repetitive extragenic palindromic- polymerase chain reaction.7. Surface enhanced laser desorption/ionization-timeof flight mass spectrometry was used to type pseudomonas aeruginosa. Results:1.82strains of pseudomonas aeruginosa were collected with a resistant rate of85.4%for imipenem. The resistant rate were between39%-68.9%for Amikacin,gentamicin, tobramycin, ceftriaxone, ceftazidime, cefotaxime, cefepime,ciprofloxacin, gatifloxacin, levofloxacin, piperacillin/methimazole behalfcardamoxicillin/clavulanic acid, piperacillin and aztreonam.2. There were49strains were resistant to aminoglycoside antibacterials and33strains weresensitive to aminoglycoside antibacterials in the82strains.3.5kinds ofaminoglycoside modifying enzyme genes were detected in which aac (3)–IIwere19strains (38.8%), aac (6’)-I were19strains (38.8%), aac (6’)-Ⅱwere18strains (36.7%), ant (2")-I were35strains (71.4%) and ant (3)-I were13strains (26.5%).4. The homology were98%to100%between5kinds ofpositive aminoglycoside modifying enzyme genes and the corresponding genesequences in GenBank which had been submitted.5.49aminoglycoside-resistant pseudomonas aeruginosa strains were divided into3groups7subgroups and11types,3groups6subgroups and13types as well as3group6subgroups12types by Multi-gene clustering analysis, rep-PCRtechnology and SELDI-TOF MS technology respectively. In addition, therewere clone epidemic strains.6.25protein peaks were found by SELDI-TOFMS technology analysing aminoglycoside resistant strains andaminoglycoside-resistant strains and the peaks of nucleus ratio for9900.024, 7600.041,9101.253,10372.874were significantly different. Conclusions:1. Inthe49aminoglycoside-resistant pseudomonas aeruginosa, the carrying rate ofant (2")-I was the highest. These starins carried Ant (2")-I or ant (2")-I andaac (6’)-I mainly.2. In addition to the presence of aminoglycosides modifyingenzyme resistance mechanisms, there were other mechanisms of resistance forthe49aminoglycoside resistant strains.3. The ability of molecular typingmethod for multi-gene cluster analysis, REP-PCR technology and SELDI-TOFMS technology are roughly the same and each of them has their ownadvantages. The combination use of these technologies not only can expand theability of typing for isoforms, but also can keep understanding the resistancesituation of these various strains.