Dissertation > Excellent graduate degree dissertation topics show

Prevalence and Transfer Mechanism of 16S rRNA Methylase Genes Among Escherichia Coli and Klebiella Pneumonia Clinical Isolates

Author: YuFangYou
Tutor: ZuoDi
School: Fudan University
Course: Pathogen Biology
Keywords: Coli bacteria The large intestine Klebsiella Pneumonia 16S rRNA methylase Gene Plasmid ESBLs Class I integron Distributed Transfer mechanisms Homology
CLC: R378
Type: PhD thesis
Year: 2011
Downloads: 227
Quote: 0
Read: Download Dissertation

Abstract


Research background of Escherichia coli and Klebsiella pneumoniae clinical isolates of common pathogens can cause respiratory tract infections, urinary tract infections, sepsis, purulent infection, an important cause of hospital infection pathogens, clinical anti-infection treatment and hospital infection control object. With the extensive use of antimicrobial agents in clinical, caused by multidrug-resistant Escherichia coli and Klebsiella pneumoniae infection growing number of clinical anti-infection treatment and hospital infection control cause difficulties. Especially Escherichia coli and Klebsiella pneumoniae, produce extended-spectrum β-lactamase (Extended-spectrumβlactamases, ESBLs), resulting in the penicillins, cephalosporins, and single-ring antimicrobial resistance. Addition to ESBLs outside, Escherichia coli and Klebsiella pneumoniae also produce plasmid-mediated AmpC enzyme, resulting in more serious resistance. In recent years, also found in Escherichia coli and Klebsiella pneumoniae carbapenemases, including KPC enzymes and metalloenzymes, such enzymes can hydrolyze carbapenems including all β-lactam class of antibacterial drugs. Aminoglycoside antibiotics because of the ear and kidney toxicity, clinical application subject to certain restrictions, but the aminoglycoside antibiotics against Pseudomonas aeruginosa, Klebsiella pneumoniae, Escherichia coli and other common gram-negative bacilli post-antibiotic effect longer still for the treatment of aerobic gram-negative bacilli due to a serious infection. Used alone aminoglycoside antibiotic treatment may be poor efficacy often be combined with other antibacterial drugs, such as third-generation cephalosporins and fluoroquinolones drugs with powerful antibacterial activity against gram-negative bacilli. Bacteria can produce aminoglycoside inactivating enzymes, such as N-acetyltransferase, O-phosphate transferase and O-adenosine transferase, resulting in resistance to aminoglycosides, these modified enzyme is present in the plasmid or chromosome. These modified enzymes are usually acts only a structure similar to one or several of the aminoglycoside antibiotics, all the aminoglycoside antibiotics can not be inactivated. In recent years, found that a class of plasmid-mediated 16S rRNA methylase in clinical isolates of Gram-negative bacilli, the enzyme can protect the bacterial 30S ribosomal 16S rRNA aminoglycoside antibiotic combination, resulting in, including Abbe clinical card Star aminoglycoside antibiotic resistance, and high-level resistance. 16S rRNA methylase ArmA been found since 2003, so far, have been found in clinical isolates of Gram-negative bacilli ArmA, RmtA, RmtB, RmtC, RmtD and NpmA6 kinds of 16S rRNA methylase encoded a structural gene for these enzymes by the plasmid in the spread between the different strains. Recently, in Escherichia coli isolated from cattle body found 16S rRNA methylase RmtE, clinical isolates found RmtE. It is reported that, in many parts of the world by producing 16S rRNA methylase strains caused infection occurred. About our production 16S rRNA methylase strains cause infection epidemiological data are not comprehensive enough, this study 16S rRNA methylase gene transfer mechanism is not deep enough to be isolated from the First Affiliated Hospital of Wenzhou Medical colorectal coli bacteria and pneumonia Klebsiella clinical isolates 16S rRNA methylase gene screening study of 16S rRNA the methylase gene transfer mechanism. The first part of the 16S rRNA methylase gene of Escherichia coli and Klebsiella pneumoniae collection of First Affiliated Hospital of Wenzhou Medical Screening January 2006 to September 2007, 337 clinical isolates of Klebsiella pneumoniae, and 2006 January to July 2008, 680 clinical isolates of E. coli, all strains are non-repetitive strains. Automatic microbial analyzer for strain identification and susceptibility testing, screening gentamicin (GEN), tobramycin (TOB) and amikacin (AMK) either a drug-resistant strains as measured strains. Screened a total of 64 Klebsiella pneumoniae resistant to at least one kind of the aminoglycoside, was selected as the test strains. 337 Klebsiella pneumoniae GEN, TOB, and AMK resistance rates were 19.0% (64/337), 16.3% (55/337) and 8.3% (28/337); 28 bacteria resistant to GEN TOB and AMK, all 28 of AMK-resistant Klebsiella pneumoniae resistant TOB and GEN. 365 E. coli at least one of the above aminoglycoside resistance 44 GEN, AMK and TOB resistant, was selected as the test strain. 680 Escherichia coli GEN, TOB, and AMK resistance rates were 52.5% (357/680), 50.9% (346/680) and 6.5% (44/680). AMK-resistant Escherichia coli resistance GEN and TOB. AMK vitro antibacterial activity of Escherichia coli and Klebsiella pneumoniae clinical isolates. Confirmed by PCR and DNA sequencing, 21 Klebsiella pneumoniae 16S rRNA methylase gene-positive, the positive rate was 6.2% (21/337), of which three positive armA and, 13 positive for rmtB and 5 positive for both armA and rmlB. 16S rRNA methylase genes in AMK-resistant strains, TOB resistant strains and GEN-resistant strains were detected in 75.0% (21/28), 38.2% (21/55) and 32.8% (21 / 64). 37 strains of Escherichia coli 16S rRNA methylase gene-positive, the total positive rate was 5.4% (37/680), including 36 for rmtB positive, l strains for armA-positive. All tested strains rmtA, rmtC, rmtD, and npmA as negative. All 16S rRNA methylase gene-positive strains highly resistant to GEN, TOB, and AMK (MIC gt; 256mg / L). 19 (90.5%, 19/21) of the 16S rRNA methylase gene-positive Klebsiella pneumoniae strains producing ESBLs confirmed by PCR and DNA sequencing, 21, 15 and 19, respectively, for of blaTEM, blasHv and blaCTX- M gene positive. All blaTEM for blaTEM-1, blasHv for blaSHv-12, 19 strains of blaCTX-M-positive strains were 13 blaCTX-M-14-positive and six blaCTX-M-15-positive. 29 (78.4%, 29/37) of the 16S rRNA methylase gene positive Escherichia coli strains producing ESBLs were detected by PCR and DNA sequencing confirmed that all 37 were positive for blaTEM sequencing all blaTEM-1. 29 Zhu ESBLs producing strains and 2 the ESBLs-negative strains blaCTX-M gene positive blaCTX-M-14 and blaCTX-M-15. 16SrRNA methylase gene-positive Klebsiella pneumoniae and Escherichia coli strains class I integrase gene positive. 16S rRNA methylase gene has been isolated in the Wenzhou area Escherichia coli and Klebsiella pneumoniae clinical isolates disseminated to genotype and to rmtB mainly for rmtB and armA; the first discovered rmtB and armA common exist in the same strain; 16S rRNA methylase gene-positive strains highly resistant to GEN, TOB, and AMK, 16S rRNA methylase genes usually associated with ESBL genes and class I integrase gene co-exist in the same strain, resulting in multi-drug resistant. The second part of 16S rRNA methylase gene transfer mechanism of the 16S rRNA methylase gene-positive Escherichia coli and Klebsiella pneumoniae strains as the donor strain, sodium azide-resistant Escherichia coli J53 joined to the recipient strain test. 13 (61.9%, 13/21), Klebsiella pneumoniae by conjugation test successfully passed to the recipient strain carrying the 16S rRNA methylase gene plasmid, 2 for armA positive strains 7 rrntB positive strains The four and for armA and rmtB positive strains, but of 5 armA and rmtB simultaneously the positive strains zygote only rmtB positive. By trial and error, only four (10.8%, 4/37) rmtB positive E. coli strains by conjugation test successfully carry 16S rRNA methylase gene plasmid transfer to the recipient strain. All zygote is highly resistant to GEN, TOB, and AMK and the class I integrase gene all positive ESBL genotypes for bacteria. The amount of plasmid extraction kit to extract the non-bonding strain plasmid after transformation test drug plasmids were transformed into Escherichia coli DH5a transformants, all transformants were highly resistant to GEN, TOB, and AMK, and class I integrase enzyme Gene all positive, same carried by ESBL genotype for bacteria. DNA hybridization test positive at the same time on the of 5 armA and rmtB Klebsiella pneumoniae and zygote plasmid DNA hybridization, found rmtB located on an approximately 75Kb plasmid, and may be located on chromosome armA. By molecular hybridization test of of a rmtB positive and the other strains armA positive E. coli transformants plasmid DNA hybridization, found the the Escherichia coli rmtB and armA located approximately 54Kb plasmid. Walking sequencing method to obtain the Klebsiella pneumoniae rmtB gene surrounding sequence approximately 11Kb size of the fragment, found upstream near rmtB genes exist a Tn3 the transposon sequences rmtB genes exist between the two insertion sequence IS26 transposase tnpA gene present downstream of a coding with the bonding functional traI gene. The Escherichia coli rmtB the upstream and Klebsiella pneumoniae rmtB gene upstream, but downstream a quinolones efflux pump genes qepA of. Escherichia coli and Klebsiella pneumoniae armA gene upstream existence the the two transposase gene, respectively, as the transposase gene tnpA and tnpU of, there are a LSCR1 insert sequence upstream of the transposase gene tnpU; in armA downstream a transposase gene tnpD. Exist in the Klebsiella pneumoniae rmtB genes are usually co-located with the ESBL gene and class I integron transfer plasmid plasmid transfer between different strains. 5 armA rmtB positive strains armA may be located on the chromosome, the rest of the strains armA located on the plasmid. The presence of E. coli armA rmtB gene is usually co-located in the non-engagement of the plasmid with ESBL gene and class I integron The around armA and rmtB transfer of the inserted sequence, and the transposase gene may occur through the transposon. Klebsiella pneumoniae carrying rmtB plasmid transfer occurs through bonding due to rmtB downstream to a bonding traI gene and the presence of E. coli rmtB gene downstream the lack of traI gene is not plasmid By engaging the reason for the shift occurred. The gene downstream Escherichia coli rmtB the with qepA gene, cause rmtB gene-positive strains simultaneously resistant to ciprofloxacin and levofloxacin reason. The third part of the 16S rRNA methylase gene-positive strains homology analysis using PFGE of 21 16S rRNA methylase gene-positive Klebsiella pneumoniae and 37 16SrRNA methylase gene-positive E. coli homology analysis. 21 Klebsiella pneumoniae divided into 14 PFGE type, type A and type I; five belong to type A, four of them isolated from the ICU and a the separation from neurosurgery rmtB positive strains; 4 belongs to I type, of which three separation from thoracic surgery and isolated from one of Urology, 4 strains simultaneously for armA rmtB positive, the rest are for the plant belonging to single - the PFGE types and distribution in different sections. 37 16S rRNA methylase gene-positive E. coli are divided into 19 PFGE type, where H 12 3 A and N-type, G, I, and P-type 2, type the rest of each a. Same PEGE type strain exist in various departments, and there is more than one PFGE type and the same department. Klebsiella pneumoniae and Escherichia coli carrying 16S rRNA methylase genes clone disseminated, but most of the strains dispersed genetic clones; same clone cause infection in different wards spread; same ward there are several scattered clone; rmtB, and armA gene positive strains disseminated clone, and can also be spread through the horizontal. Summary In summary, the main conclusions of this study are the following: 1. Amikacin in vitro antibacterial activity against Escherichia coli and Klebsiella pneumoniae clinical isolates, screening 16S rRNA methylation ideal gene drugs; Wenzhou area of ??clinical isolates of Escherichia coli and Klebsiella pneumoniae 16S rRNA methylase gene-positive rate, and the rate of the high rmtB gene positive armA positive, with Contrary to foreign reports; first discovered simultaneously rmtB, and armA gene in E. coli 16S rRNA methylase gene usually ESBL gene and class I integron located in a non-bonding in the same strain; resistance plasmid. Klebsiella pneumoniae, 16S rRNA methylase genes usually associated with ESBL gene and class I integron located on a self-transfer bonding plasmid; 5.rmtB, and armA genes often located between insertion sequence by transposon transferred. Exist in clinical isolates of Escherichia coli and Klebsiella pneumoniae armA the same sequence of genes around armA sources and transfer mechanisms may exists in both bacteria around the rmtB gene sequence, two bacteria rmtB of sources, transfer mechanisms and the resistance phenotype may vary. 6.16S rRNA methylase gene-positive E. coli and Klebsiella pneumoniae bacteria can be spread, clone can also be spread through the horizontal.

Related Dissertations

  1. Research and Implementation on Knowledge Representation of Gene Regulatory Network,Q78
  2. Research on Combinatorial Regulation of Multiple Transcription Factors,Q78
  3. Study on Channel Allocation of Multi-Channel MAC Protocol in Ad-Hoc Network,TN929.5
  4. Research of Fault Injection for a Distributed System,TP338.8
  5. The Research of Fault-Tolerant Techniques for Parallel/Distributed Network Simulator PDNS,TP302.8
  6. Research and Implementation of Retrieval System on Massive Mail,TP393.098
  7. Retuospective and Follow up Study on the Treatement of Pneumonia in Zhe Elderly by Integrated Tradi-Tional Chinese and Western Medicine,R563.1
  8. Cloning and Expression of CHS and CHI Genes and Their Regulation on the Accumulation of Flavonoids in ’Cara Cara’ Navel Orange (Citrus Sinensis Osbeck) and ’Guoqing NO.4’ Satsuma Mandarin (Citrus Unshiu Marcow),S666.4
  9. The Expression of Let-7 microRNA during the Process of the Mouse Embryonic Lung Development and the Construction of the Adenovirus Shuttle Plasmid,Q78
  10. Chemical Structures and Mechanism Research of Anti-aging Leading Compounds from Natural Products,R285.5
  11. CYP2C19 Gene Polymorphism in Patients with Ischemic Cerebrovascular,R743
  12. The Expression Survey of BMP Signalling Pathway in the Human Embryonic Tooth Germ,R78
  13. Purification of Endogenous Proteases from (Litopenaeus Vannamei) Shrimp Head and Their Enzymology Properties,S985.21
  14. Cloning and Expression of Growth Hormone Gene in Cobia (Rachycentron Canadus),Q786
  15. Breeding of 1, 3-propanediol-overproducing Strain by Genome Shuffling,TQ923
  16. Effects of Different Reconciled Corn Oils on Antioxidative Stress, Lipid Metabolism Enzymes, and Abundance of Immune-related Genes of Broilers,S831.5
  17. Polyethylenimine Conjugated Stearic Acid-g-chitosan Oligosaccharide Micelles for Antitumor Gene Therapy,R450
  18. Research and Design of One Kind of Paper’s QCS That Based on Embedded System,TP368.1
  19. An Approach for Identifying a Plant Resistance Gene Based on the Random Forest,Q943
  20. Micro- grid with distributed power control strategy research,TM61
  21. Functional Analysis of Proteins Encoded by RNA2 of Wheat Yellow Mosaic Virus,S435.121

CLC: > Medicine, health > Basic Medical > Medical Microbiology ( pathogenic bacteriology,pathogenic microbiology ) > Pathogenic bacteria
© 2012 www.DissertationTopic.Net  Mobile