Acinetobacter Baummanii As Nosocomial Pathogens: Epidemiological Survey And Resistant Mechanisms

Objective To epidemiologically survey on Acinetobacter baummanii nosocomial infections and to study the risk factors of the infections. To establish the molecular typing methods for epidemic strains of A. baumannii. Detection of β-lactamses and study on quinolone resistant mechanisms in A. baumannii.Methods 1.The 140 patiens with A. baummanii nosocomial infections from a teaching hospital were surveyed, and were performed to identify risk factors by a case control comparison. The rates of antibiotic resistance were analyzed by WHONET4 software. 2. In our study for typing A. baummanii isolates from our hospital, the repetitive element ERIC and the core region of bacteriophage M13 had been used as single primers to compare their discriminatory results. The method of pulsed-field gel electrophoresis for intact chromosomal DNA had been used to analyze for typing A. baummanii. 3.Theβ-lactamases of 23 A. baummanii strains was screened by Nitrocefin. A modified three-dimensional extract test and the double-disc diffusion test were used to detect phenotypically isolates which produced AmpC β-lactamases and extended- spectrum β-lactamases, and the stability of antibiotics to β-lactamase was studied. The isoelectric points of β-lactamases was measured. The plasmids and chromosomal DNA were amplificated by primers of bla-TEM and ampC coding region. 4.According to MICs of ciprofloxacin, isolates were classified to high-resistance strains, resistance strains and susceptibility strains. We analyzed outer memberane proteins, accumulation of ofloxacin, morphological ultrastructures and gyrA, parC genes of amplification in different strains. Detection of point mutation in their genes ofquinolone-resistant isolates was dependant on PCR-RFLP , PCR-SSCP and the samples were processed for DNA sequencing. Results 1.The A. baummanii infections associated morbidity was high ( 5.4‰)in our hospital, particularly in trauma ward (ICU) and respiratory ward. Except imipenem,the rates of A. baummanii resistance various of antibiotics were over 50％ in vitro susceptibility testing. By calculating ,the odds ratios(OR) were: state of an illness (OR=8.691), using immunosuppressant (OR=4.85), mechanical ventilation(OR=3.680),treatment with antibiotics more than 3 kinds (OR=3.014). 2.Analysis of theDNA figerprintings indicated that ERIC-PCR was divided into 5 profiles and M13-PCR was divided into 9 profiles. Restriction fragment length polymorphisms were generated from intact chromosomal DNA of A. baummanii. 3.All of isolates producedβ-lactamases. Although none of them possessed ESBLs, half of isolates hyperexpressed AmpC enzymes. The measurement of isoelectric points showed the one of PI value was 5.4 ,another were above 9.0. Results of PCR amplification revealed that 23 A. baummanii strains had ampC structure gene in chromosomal DNA and bla-TEM in plasmids which they carried.4.Results of MICs to ciprofloxacin showed that there were 9 strains were high-resistance (≥32μg/ml), 3 resistance strains (≥4μg/ml), and 11 susceptibility strains(＝0.25μg/ml).The change of outer membrane proteins was observed in RH7. The accumulation of ofloxacin in RH7 decreased. After treatment with carbonyl cyanide-m-chlorophenyl hydrazone(CCCP), the drug uptake increased to the normal level. There wasn't any change in ultrastructures of RH7. All 9 high-resistance isolates showed a change at gyrA gene, and 5 of them had a change of parC gene. 2 of 3 resistance isolates showed some changes at gyrA gene.Conclusions 1.There was highly morbidity of A. baummanii nosocomial infections in our hospital, particularly in trauma ward (ICU) and respiratory ward. Such infections are often extremely difficult for the clinician to treat because of the widespread resistance of these bacteria to themajor groups of antibiotics. 2. Risk factors of nosocomial infections with A. baummanii were associated with severe illness, immunosuppressant, mechanical ventilation , treatment with antibiotics. 3...