Analysis of Genetic Determinants Involved in Multiresistance in Clinical Strains Isolated from Renal Transplantation Recipients in Guangzhou, China
Lei Shi1), Yali Kou1), Lin Li1) and Shin-ichi Miyoshi2)
1) College of Food and Biological Engineering, South China University of Technology
2) Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama University
(Received October 30, 2006)
(Accepted December 25, 2006)
In the present study, we examined the antibiotic sensitivity of 19 bacterial strains [5 coagulase-negative Staphylococcus, 2 methicillin-resistant Staphylococcus aureus (S. aureus), 2 Enterococcus faecium (E. faecium), 5 Escherichia coli (E. coli), 3 Cedecea sp., 1 Klebsiella pneumoniae (K. pneumoniae), and 1 Burkholderia cepacia (B. cepacia)], which were isolated from renal transplantation patients using the Kirby-Bauer method. We also investigated the production of β-lactamase and extended-spectrum β-lactamase (ESBL), and the presence of the integrase gene (intI1) and resistance gene cassette. Among the 19 strains tested, all displayed severe multiresistance, and 12 strains produced β-lactamase, in which 6 strains were ESBL positive. Eleven strains were revealed to possess the class 1 integron; however, neither class 2 nor 3 was detected. Additionally, 3 drug resistance genes, aadA2, dfrA17, and aadA5, were found in some strains. The results indicate that the horizontal transfer of the β-lactamase gene and/or the class 1 integron may contribute significantly to the spread of multiresistant bacteria among renal transplantation patients.
Key words renal transplantation, multiresistance, β-lactamase, integron
Tampilkan postingan dengan label Biotecnology. Tampilkan semua postingan
Tampilkan postingan dengan label Biotecnology. Tampilkan semua postingan
Rabu, 20 Februari 2008
Selasa, 19 Februari 2008
Gene Cassette PCR: Sequence-Independent Recovery of Entire Genes from Environmental DNA
Gene Cassette PCR: Sequence-Independent Recovery of Entire Genes from Environmental DNA
H. W. Stokes,1,* Andrew J. Holmes,1,2 Blair S. Nield,1 Marita P. Holley,1,2 K. M. Helena Nevalainen,1 Bridget C. Mabbutt,3 and Michael R. Gillings1,2
Department of Biological Sciences,1 Key Centre for Biodiversity and Bioresources,2 and Department of Chemistry,3 Macquarie University, Sydney, New South Wales 2109, Australia
Received 24 May 2001/Accepted 20 August 2001
The vast majority of bacteria in the environment have yet to be cultured. Consequently, a major proportion of both genetic diversity within known gene families and an unknown number of novel gene families reside in these uncultured organisms. Isolation of these genes is limited by lack of sequence information. Where such sequence data exist, PCR directed at conserved sequence motifs recovers only partial genes. Here we outline a strategy for recovering complete open reading frames from environmental DNA samples. PCR assays were designed to target the 59-base element family of recombination sites that flank gene cassettes associated with integrons. Using such assays, diverse gene cassettes could be amplified from the vast majority of environmental DNA samples tested. These gene cassettes contained complete open reading frames, the majority of which were associated with ribosome binding sites. Novel genes with clear homologies to phosphotransferase, DNA glycosylase, methyl transferase, and thiotransferase genes were identified. However, the majority of amplified gene cassettes contained open reading frames with no identifiable homologues in databases. Accumulation analysis of the gene cassettes amplified from soil samples showed no signs of saturation, and soil samples taken at 1-m intervals along transects demonstrated different amplification profiles. Taken together, the genetic novelty, steep accumulation curves, and spatial heterogeneity of genes recovered show that this method taps into a vast pool of unexploited genetic diversity. The success of this approach indicates that mobile gene cassettes and, by inference, integrons are widespread in natural environments and are likely to contribute significantly to bacterial diversity.
from : http://aem.asm.org
H. W. Stokes,1,* Andrew J. Holmes,1,2 Blair S. Nield,1 Marita P. Holley,1,2 K. M. Helena Nevalainen,1 Bridget C. Mabbutt,3 and Michael R. Gillings1,2
Department of Biological Sciences,1 Key Centre for Biodiversity and Bioresources,2 and Department of Chemistry,3 Macquarie University, Sydney, New South Wales 2109, Australia
Received 24 May 2001/Accepted 20 August 2001
The vast majority of bacteria in the environment have yet to be cultured. Consequently, a major proportion of both genetic diversity within known gene families and an unknown number of novel gene families reside in these uncultured organisms. Isolation of these genes is limited by lack of sequence information. Where such sequence data exist, PCR directed at conserved sequence motifs recovers only partial genes. Here we outline a strategy for recovering complete open reading frames from environmental DNA samples. PCR assays were designed to target the 59-base element family of recombination sites that flank gene cassettes associated with integrons. Using such assays, diverse gene cassettes could be amplified from the vast majority of environmental DNA samples tested. These gene cassettes contained complete open reading frames, the majority of which were associated with ribosome binding sites. Novel genes with clear homologies to phosphotransferase, DNA glycosylase, methyl transferase, and thiotransferase genes were identified. However, the majority of amplified gene cassettes contained open reading frames with no identifiable homologues in databases. Accumulation analysis of the gene cassettes amplified from soil samples showed no signs of saturation, and soil samples taken at 1-m intervals along transects demonstrated different amplification profiles. Taken together, the genetic novelty, steep accumulation curves, and spatial heterogeneity of genes recovered show that this method taps into a vast pool of unexploited genetic diversity. The success of this approach indicates that mobile gene cassettes and, by inference, integrons are widespread in natural environments and are likely to contribute significantly to bacterial diversity.
from : http://aem.asm.org
PCR isolation of catechol 2,3-dioxygenase gene fragments from environmental samples and their assembly into functional genes
PCR isolation of catechol 2,3-dioxygenase gene fragments from environmental samples and their assembly into functional genes
Akiko Okuta, Kouhei Ohnishi and Shigeaki Harayama*
Marine Biotechnology Institute, 3-75-1 Heita, Kamaishi, Iwate 026, Japan
Received 1 December 1997; revised 9 March 1998; accepted 10 March 1998 Available online 16 June 1998.
A. Nakazawa.
Abstract
A method was developed to isolate central segments of catechol 2,3-dioxygenase (C23O) genes from environmental samples and to insert these C23O gene segments into nahH (the structural gene for C23O encoded by catabolic plasmid NAH7) by replacing the corresponding nahH sequence with the isolated segments. To PCR-amplify the central C23O gene segments, a pair of degenerate primers was designed from amino acid sequences conserved among C23Os. Using these primers, central regions of the C23O genes were amplified from DNA isolated from a mixed culture of phenol-degrading or crude oil-degrading bacteria. Both the 5′ and 3′ regions of nahH were also PCR-amplified by using appropriate primers. These three PCR products, the 5′-nahH and 3′-nahH segments and the central C23O gene segments, were mixed and PCR-amplified again. Since the primers for the amplification of the central C23O gene segments were designed so that the 20 nucleotides at both ends of the segments are identical to the 3′ end of the 5′-nahH segment and the 5′ end of the 3′-nahH segment, respectively, the central C23O gene segments could anneal to both the 5′- and 3′-nahH segments. After the second PCR, hybrid C23O genes in the form of (5′-nahH segment—central C23O gene segment—3′-nahH segment) were amplified to full length. The resulting products were cloned into a vector and used to transform Escherichia coli. This method enabled divergent C23O sequences to be readily isolated, and more than 90% of the hybrid plasmids expressed C23O activity. Thus, the present method is useful to create, without isolating bacteria, a library of functional hybrid genes.
from : http://www.sciencedirect.com
Akiko Okuta, Kouhei Ohnishi and Shigeaki Harayama*
Marine Biotechnology Institute, 3-75-1 Heita, Kamaishi, Iwate 026, Japan
Received 1 December 1997; revised 9 March 1998; accepted 10 March 1998 Available online 16 June 1998.
A. Nakazawa.
Abstract
A method was developed to isolate central segments of catechol 2,3-dioxygenase (C23O) genes from environmental samples and to insert these C23O gene segments into nahH (the structural gene for C23O encoded by catabolic plasmid NAH7) by replacing the corresponding nahH sequence with the isolated segments. To PCR-amplify the central C23O gene segments, a pair of degenerate primers was designed from amino acid sequences conserved among C23Os. Using these primers, central regions of the C23O genes were amplified from DNA isolated from a mixed culture of phenol-degrading or crude oil-degrading bacteria. Both the 5′ and 3′ regions of nahH were also PCR-amplified by using appropriate primers. These three PCR products, the 5′-nahH and 3′-nahH segments and the central C23O gene segments, were mixed and PCR-amplified again. Since the primers for the amplification of the central C23O gene segments were designed so that the 20 nucleotides at both ends of the segments are identical to the 3′ end of the 5′-nahH segment and the 5′ end of the 3′-nahH segment, respectively, the central C23O gene segments could anneal to both the 5′- and 3′-nahH segments. After the second PCR, hybrid C23O genes in the form of (5′-nahH segment—central C23O gene segment—3′-nahH segment) were amplified to full length. The resulting products were cloned into a vector and used to transform Escherichia coli. This method enabled divergent C23O sequences to be readily isolated, and more than 90% of the hybrid plasmids expressed C23O activity. Thus, the present method is useful to create, without isolating bacteria, a library of functional hybrid genes.
from : http://www.sciencedirect.com
Langganan:
Komentar (Atom)
Nature
Spirulina
Mikroalga kaya protein
undersea explorer
