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Objective:To investigate the drug resistance genes carried on a carbapenem-resistant Acinetobacter spp. isolate. Methods:A retrospective study was conducted on one strain of carbapenem-resistant Acinetobacter spp, which was clinically isolated in July 2016. The strain was identified by Vitek2-compact and further confirmed by PCR detection of bla OXA-51-like, 16S-23S rRNA intergenic space sequencing and partial rpoB gene sequence analysis. Carbapenemase (NDM, KPC, VIM, IMP, SIM, SPM, GIM, OXA-23-like, OXA-24-like, OXA-58-like), 16S rRNA methylase gene (armA, rmtA, rmtB), Beta-lactamases (TEM, SHV, CTX-M, VEB, PER) and class 1 integron were amplified and detected by PCR, and plasmids were analyzed using Southern hybridization. Results:The Acinetobacter baylyi isolate showed the positive for bla NDM-1 only, and negative for the other genes. Plasmid analysis and Southern bybridization result revealed that the bla NDM-1 gene was located on unconjugatable plasmid (54 000-60 000 bp). Conclusions:Attention should be paid to the occurrence of carbapenem-resistant non-baumannii Acinetobacter spp. and the transmission mechanism of drug resistant genes.
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Objective To observe the application of multiple fluorescent PCR (Polymerase Chain Reaction) in the diagnosis and clinical detection of bloodstream infection. Methods 256 blood cultures were collected by the Laboratory Department of Yinzhou People′s Hospital from January 2018 to May 2018, and were detected by multiplex fluorescent PCR. The results of the PCR were compared with the traditional blood culture bacteria identification instrument (traditional blood culture method). The number of positive and negative samples and the number of corresponding samples of the two methods were counted. Then, they analyzed the specificity and sensitivity of multiplex fluorescence PCR in the diagnosis of bloodstream flow infections. Results A total of 18 pathogenic microbes are detected through blood culture and PCR. Multiple fluorescent PCR detects 142 positive samples and 114 negative samples. Among them, 132 samples also show positive through blood culture, and 111 samples show negative. The consistency rate between multiple PCR and traditional blood cultures is 91.8％ (235/256). The negative prediction rate of PCR is 97.4％ (111/114), sensitivity rate 97.8％ (132/135), specificity rate 91.7％ (111/121). 10 samples show positive through multiple fluorescence PCR but negative for blood culture, 3 samples show positive through blood culture but negative for PCR. Besides, there are 3 types of pathogens that exceed the detection range of PCR. Conclusions Multiplex PCR method can detect 17 pathogens in blood culture specimens of patients, which can not only optimize the traditional blood culture process, but also greatly shorten the reporting time and improve the detection rate of blood culture methods. Especially for patients treated with antibiotics, it can reduce missed detection and improve the diagnostic rate of bloodstream infections.
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Objective To identify Mycobacterium abscessus rapidly with HAIN molecular assay genotype kit、gene chip and hsp65 gene sequencing,and to assess the clinical value of these three methods.Methods 13 clinical non-tuberculous Mycobacterium (NTM) of in-patient samples were collected from January2014 to January 2015,in the Department of Clinical Laboratory,Yinzhou People's Hospital,and meanwhile,these strains were identified with HAIN molecular assay genotype Mycobacterium kit and gene chip respectively.The hsp65 gene sequencing was used as the standard method to be compared with HAIN and gene chip.Results The results of HAIN kit and hsp65 gene sequencing showed that all the 13 strains were subspecies Mycobacterium abscessus,while that of gene chip showed that these strains were Mycobacteriumchelonae complex strains,and the subtypescould not be identified.Conclusion These results obtained from the HAIN molecular assay genotype mycobacterium system are in agreement with those obtained from the hsp65 gene sequencing,whereas the HAIN kit method is easier to use.
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Objective To establish a method for detecting Aspergillus spp.by Loop-mediated isothermal amplification.Methods Aspergillus spp.specific primers were designed from the relative conservation region of the published sequence of 28S rRNA genes of A.fumigatus (GenBank accession number AY660917),A.terreus (GenBank accession number AF454183),A.flavus (GenBank accession number AF454158),and A.niger (GenBank accession number AF454169).Genomic DNA were extracted from Aspergillus standard strains,clinical control strains and clinical samples,and amplified by LAMP.The amplified results could be read with the naked eye by the coloring effect of fluorescent nucleotide dye without the DNA electrophoresis.Approximately 103 each Aspergillus conidia suspension was added to the serum from healthy volunteers,and detected these simulative clinical samples bv LAMP assay.Results The LAMP and PCR assays obtained positive results for all four Aspergillus species and 8 simulative clinical samples (double samples for each Aspergillus species) including 103 conidia,but negative in the remaining 15 non-Aspergillus species,human total blood genomic DNA,30 clinical serum samples infected with non-Aspergillus and 10 healthy volunteers.The LAMP assay had a minimum detection of 0.05-0.5pg,by means of detect different levels of 500,50,5,0.5,0.05 pg template in each reaction tube.Conclusion The results confirm that LAMP is a simple,rapid,sensitive and specific method,and can be used for detection of Aspergillus strains in clinical and environmental specimens.
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Objective To investigate the distribution of virulence-related genes in multidrug resistant Escherichia coli.Methods Seven virulence genes papA,cnf1,cnf2,cfaB,ipaB,hofQ and ompT were detected by PCR in 20 strains of multidrug resistant Escherichia coli clinically isolated,and the positive genes were further searched in 31 strains of Escherichia coli in BioCyc database whose genomies had been fully sequenced.Results Virulence genes hofQ and ompT were detected in 20 strains of Escherichia coli with a positive rate of 95.0％ (19/20) and 55.0％ ( 11/20),respectively.Among 31 strains of Escherichia coli in BioCyc,21 (67.7％) were positive for hofQ gene and 15 (48.4％) were positive for ompT gene.Conclusion hofQ and ompT genes are prevalent in multidrug resistant Escherichia coli.
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OBJECTIVE To study the mechanisms of chromosome and plasmid-mediated quinolones resistance in Escherichia coli(ECO).METHODS Clinical isolates of ECO were collected from clinical specimens at ICU of Yinzhou People′s Hospital from Dec 2007 to Jun 2008.To detect the susceptibility to 30 types of antibiotics K-B disk diffusion method was used.The susceptibicity to ciprofloxacin and levofloxacin were detected by agar dilution testing.Then six type genes gyrA,qnrA,qnrB,qnrS,aac(6′)-Ⅰb-Cr,and qepA were investigated by PCR.In the meantime,the PCR products were sequenced.RESULTS The alterations in gyrA were found in all 20 tested strains.Two new subtypes were found in ECO 13 and ECO 15.Three strains were found acc(6′)-Ⅰb-Cr and ECO 2 was detected qurA gene.CONCLUSIONS The mutations in gyrA play a dominant role in the resistance to quinolones in ECO.Both aac(6′)-Ⅰb-Cr and qepA may responsible for quinolones resistance in ECO too.
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OBJECTIVE To investigate the antibiotics resistance of Escherichia coli(ECO) and distrubtion of aminoglycoside-modifying enzymes and 16S RNA methylase genes in ICU.METHODS The samples of 20 ECO isolates were collected from Dec 2007 to Jun 2008 of patients in ICU.To determine the sensitivity to the 30 antibacterials K-B method was used and aminoglycoside-modifying enzymes and 16S RNA methylase genes were analyzed by polymerase chain reaction(PCR).RESULTS In among the 20 ECO isolates,8 strains carried aac(3)-Ⅱ(40%),3 carried aac(6′)-Ⅰb(15%) and ant(3″)-Ⅰ(15%),1 be found aph(3′)-Ⅰ(10%)and 13 be found aadA4/5 aminoglycoside-modifying enzymes genes,no strain carried 16S RNA methylase genes.CONCLUSIONS aac(3)-Ⅱ、aac(6′)-Ⅰb,ant(3″)-Ⅰ,aph(3′)-Ⅰ and aadA4/5 aminoglycoside-modifying enzymes genes exist in ECO widely,they should be the main cause inducing the high rate of drug-resistance to aminoglycosides.
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OBJECTIVE To investigate the antibiotics resistance of multi-resistant Acinetobactor baumannii(ABA) and genotypes of beta-lactamases in ICU.METHODS The samples of 20 A.baumannii isolates were collected from Oct 2007 to Jul 2008 from patients in ICU.To determine the sensitivity to the 32 kinds of antibacterials,K-B method was used and the detection of ESBLs and AmpC beta-lactamases was performed by three dimensional test and 21 types of beta-lactamases genes were analyzed by polymerase chain reaction(PCR).RESULTS Among the 20 ABA isolates,all carried TEM beta-lactamases gens(100%),10 carried OXA-23 beta-lactamases gens(50%) and 15 strains carried ADC beta-lactamases gene(75%),50% strains produced TEM,OXA-23 and ADC beta-lactamases simultaneously.Through determining sequence of one PCR product from TEM,OX23 and ADC respectively,we found TEM-116,OXA-73 and ADC-25 type beta-lactamases genes.CONCLUSIONS The antibiotics resistance of ABA is very serious.TEM,OXA-23 and ADC exist in multi-resistant A.baumannii widely.It should be the main causes as high rate of drug-resistantce to beta-lactamantibiotics.
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OBJECTIVE To investigate the antibiotic resistance of multi-resistant Acinetobacter baumannii(ABA) and distribution of aminoglycoside-modifying enzymes and 16S rRNA methylase genes in ICU in Yinzhou People′s Hospital in Ningbo. METHODS The samples of 20 ABA isolates were collected from Oct 2007 to Jul 2008 in ICU.K-B method was used to determine the sensitivity to 32 antibacterials and the aminoglycoside-modifying enzymes and 16S rRNA methylase genes were analyzed by polymerase chain reaction(PCR). RESULTS From 20 ABA isolates,8 strains carried aminoglycoside-modifying enzymes genes aac(3)-Ⅰ,aac(3)-Ⅱ,aac(6′)-Ⅰb,and ant(3″)-Ⅰ,their positive rate was 10%,15%,30% and 25%,respectively;no strain carried 16S rRNA methylase genes. CONCLUSIONS The antibiotics resistance of A.baumannii is very serious in Yinzhou People′s Hospital in Ningbo.Aminoglycoside-modifying enzymes genes aac(3)-Ⅰ,aac(3)-Ⅱ,aac(6′)-Ⅰb and ant(3″)-Ⅰ exist in multi-resistant A.baumannii widely.They would be the main causes of high drug-resistantce to aminoglycosides.
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OBJECTIVE To study the existence status of class Ⅰ integron and transposable element of multi-resistant Acinetobactor baumannii isolated form ICU of Yinzhou People′s Hospital in Ningbo. METHODS The samples of 20 A.baumannii isolates were collected from Oct 2007 to Jul 2008.The susceptibility to 32 antibiotics of the isolates was measured.The genetic markers of integron qacE△1-sul1 and transposable element tnpU were analyzed by polymerase chain reaction(PCR).The PCR products of tnpU or qacE△1-sul1 were sequenced for determination. RESULTS In the 20 ABA isolates,the positive rate of class Ⅰ integron qacE△1-sull was 75%,and the positive rate of transposable element tnpU was 55.0%. CONCLUSIONS The positive rate of the integron qacE△1-sul1 and transposable element tnpU for multi-resistant A.baumannii is high in Yinzhou People′s Hospital in Ningbo.It should be reevaluated the preventative role of chlorhexidine for operation.
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OBJECTIVE To investigate the distribution and drug susceptibility of microorganism isolated from patients with infection of biliary tract,and provide informations for clinical reasonable antibiotics selection. METHODS A total of 957 bile specimens of patients with infection of biliary tract were cultured and drug susceptibility test was performed. RESULTS From the 398 strains(66.67%) of microorganism were cultured.The main pathogenic bacteria were Escherichia coli(33.09%),Klebsiella pneumoniae(11.19%),Enterococcus faecalis(8.67%),Enterococcus faecium(5.78%) and Pseudomonas aeruginosa(4.69%).Meanwhile,the main pathogenic fungus was Candida albicans(7.22%).The susceptibility of Enterobacteriaceae to amikacin,imipenem and meropenem were 95.48%,100.00% and 100.00%,respectively.Susceptibility of Enterococcus to ampicillin and high concentration gentamicin were 73.61% and 84.72%,respectively. CONCLUSIONS E.coli,K.pneumoniae,E.faecalis and C.albicans are the main pathogens resulted in biliary tract infections.The strategy of amikacin combined with ampicillin may be recommended for empirical treatment in biliary tract infection.
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OBJECTIVE To analyze the results of antimicrobial susceptibility for clinically isolated Corynebacterium striatum strains and investigate the distribution of minimal inhibitory concentration(MIC) of the antibiotics commonly used in clinic against C.striatum.METHODS C.striatum was identified with API Coryne System(bioMerieux,France).Antimicrobial susceptibility test was conducted by agar dilution method.RESULTS MIC90 of imipenem,vancomycin and rifampin against C.striatum was 8,0.5 and 2 ?g/ml,and that of amikacin,gentamicin,tobramycin,and minocycline was 8-16?g/ml,but MIC90 of the other antibiotics was 32-≥256 ?g/ml.The main source of 32 isolates was from bronchofibroscopic secretion.CONCLUSIONS The activity of vancomycin,rifampin and imipenem against C.striatum shows stable susceptibility,but of amikacin,gentamicin,tobramycin and minocycline shows inferiority.However,the susceptibility to other antibiotics is low.So vancomycin,rifampin and imipenem are the optimal antibiotics to treat the infections caused by C.striatum.
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OBJECTIVE To investigate mycoplasma infection in ICU patients.METHODS Sixty-five samples from blood,respiratory tract and genitourinary tract of patients were collected respectively from Oct 2007 to July 2008 in ICU.Mycoplasma pneumoniae(Mp),Urealasma urealytium(Uu) M.fermentans(Mf) and M.penetrans(Mpe) were cultivated by modified mycoplasma fluid and solid medium.Mf and Mpe positive isolates were verified by nested polymerase chain reaction(rPCR),Mp and Uu were confirmed by fluorescent quantitative PCR.RESULTS It was found that the positive detection rate for Mp was 12.3%(8/65)in blood and 35.4%(23/65) in respiratory tract excreta and for Mu 1.5%(1/65) and 26.2%(17/65) in blood or Genitourinary tract,respectively.Mpe and Mf did not detected.CONCLUSIONS The state of mycoplasma infection is very severe,and often accompanies bacterial infection.It is necessary to consider mycoplasma when chose antibiotics.