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Research Support, Non-U.S. Gov't
- Development of SCAR Primers Based on a Repetitive DNA Fingerprint for Escherichia coli Detection
- Aphidech Sangdee , Sitakan Natphosuk , Adunwit Srisathan , Kusavadee Sangdee
- J. Microbiol. 2013;51(1):31-35. Published online March 2, 2013
- DOI: https://doi.org/10.1007/s12275-013-2244-4
- 45 View
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- 4 Scopus
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Abstract
- The present study aimed to use enterobacterial repetitive intergenic consensus (ERIC) fingerprints to design SCAR primers for the detection of Escherichia coli. The E. coli strains were isolated from various water sources. The primary presumptive identification of E. coli was achieved using MacConkey agar. Nineteen isolates were selected and confirmed to be E. coli strains based on seven biochemical characteristics. ERIC-PCR with ERIC 1R and ERIC 2 primers were used to generate DNA fingerprints. ERIC-PCR DNA profiles showed variant DNA profiles among the tested E. coli strains and distinguished all E. coli strains from the other tested bacterial strains. A 350 bp band that predominated in five E. coli strains was used for the development of the species-specific SCAR primers EC-F1 and EC-R1. The primers showed good specificity for E. coli, with the exception of a single false positive reaction with Sh. flexneri DMST 4423. The primers were able to detect 50 pg and 100 CFU/ml of genomic DNA and cells of E. coli, respectively.
Journal Articles
- Use of rpoB Sequences and rep-PCR for Phylogenetic Study of Anoxybacillus Species
- Kadriye Inan , Yusuf Bektas , Sabriye Canakci , Ali Osman Belduz
- J. Microbiol. 2011;49(5):782-790. Published online November 9, 2011
- DOI: https://doi.org/10.1007/s12275-011-1136-8
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- 11 Scopus
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Abstract
- This study was conducted to investigate the applicability of rpoB, which encodes the β subunit of RNA polymerase, to be used as an alternative to 16S rRNA gene sequence similarity analysis in the thermophilic genus Anoxybacillus. Partial rpoB sequences were generated for the 14 type strains of Anoxybacillus species and 6 other strains of four Anoxybacillus species. The sequences and the phylogenetic tree of rpoB were compared with those obtained from 16S rRNA gene analysis. The rpoB gene was found to provide a better resolution for Anoxybacillus species, with lower interspecies sequence similarities. The rpoB sequence similarity analysis permitted a more accurate discrimination of the species within the Anoxybacillus genus than the more commonly used 16S rRNA gene. Furthermore, rapid and reproducible repetitive extragenic palindromic fingerprinting techniques (REP-, ERIC-, and BOX-PCR) were employed for the specimens of genus Anoxybacillus. Through comparison of the three methods, it was found that the BOX-PCR method generated more informative results than REP-PCR for the studied strains; BOX-PCR profiles were more distinct for the different strains, including a higher number of bands. Rapid and reproducible repetitive extragenic palindromic fingerprinting techniques (rep-PCR) constitute a suitable molecular approach for the validation and maintenance of taxonomy within the Anoxybacillus genus. The results of this study show that rpoB and rep-PCR provide rapid and reliable methods for molecular typing of Anoxybacillus species.
- Molecular Characterization of Vibrio cholerae Isolates from Cholera Outbreaks in North India
- Joseph J. Kingston , Kuruvilla Zachariah , Urmil Tuteja , Sanjay Kumar , Harsh Vardhan Batra
- J. Microbiol. 2009;47(1):110-115. Published online February 20, 2009
- DOI: https://doi.org/10.1007/s12275-008-0162-7
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- 12 Scopus
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Abstract
- Vibrio cholerae isolates recovered from cholera outbreaks in Bhind district of Madhya Pradesh and Delhi, Northern India were characterized. The O1 serogroup isolates from Bhind outbreak were of Inaba serotype whereas both Ogawa and Inaba serotypes were recovered from Delhi. PCR analysis revealed that only O1 serogroup V. cholerae isolates carried the virulence-associated genes like ctxA, tcpA, ace, and zot. Molecular typing by repetitive sequence based ERIC, VCR1, and VC1 PCR’s revealed similar DNA profile for both Inaba and Ogawa serotypes. A discrete VC1-PCR band identified among the El Tor strains had greater similarity (>97%) to the V. cholerae genome sequence and therefore has the potential to be used as a marker for the identification of the V. cholerae strains. Non-O1 strains recovered from Bhind region differed among themselves as well as from that of the O1 isolates. All the O1 serogroup isolates possessed SXT element and were uniformly resistant to the antibiotics nalidixic acid, polymyxin-B, furazolidone, cloxacilin, trimethoprim-sulfamethaxazole, and vibriostatic agent O129. Inaba strains from both Delhi and Bhind differed from Ogawa strains by their resistance to streptomycin despite sharing similar DNA patterns in all the three rep-PCRs. Though Delhi and Bhind are separate geographical regions in Northern India, Inaba strains from both these places appear to be closely related owing to their similarity in antibiogram and genetic profile.
Research Support, Non-U.S. Gov't
- Outbreaks of Imipenem-Resistant Acinetobacter baumannii Producing Carbapenemases in Korea
- Seok Hoon Jeong , Il Kwon Bae , Kwang Ok Park , Young Jun An , Seung Ghyu Sohn , Seon Ju Jang , Kwang Hoon Sung , Ki Suk Yang , Kyungwon Lee , Dongeun Young , Sang Hee Lee
- J. Microbiol. 2006;44(4):423-431.
- DOI: https://doi.org/2410 [pii]
- 43 View
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Abstract
- Among 53 Acinetobacter baumannii isolates collected in 2004, nine imipenem-resistant isolates were obtained from clinical specimens taken from patients hospitalized in Busan, Korea. Nine carbapenemase-producing isolates were further investigated in order to determine the mechanisms underlying resistance. These isolates were then analyzed via antibiotic susceptibility testing, microbiological tests of carbapenemase activity, pI determination, transconjugation test, enterobacterial repetitive consensus (ERIC)-PCR, and DNA sequencing. One outbreak involved seven cases of infection by A. baumannii producing OXA-23 β-lactamase, and was found to have been caused by a single ERIC-PCR clone. During the study period, the other outbreak involved two cases of infection by A. baumannii producing IMP-1 β-lactamase. The two clones, one from each of the outbreaks, were characterized via a modified cloverleaf synergy test and an EDTA-disk synergy test. The isoelectric focusing of the crude bacterial extracts detected nitrocefin-positive bands with pI values of 6.65 (OXA-23) and 9.0 (IMP-1). The PCR amplification and characterization of the amplicons via direct sequencing showed that the clonal isolates harbored blaIMP-1 or blaOXA-23 determinants. The two clones were characterized by a multidrug resistance phenotype that remained unaltered throughout the outbreak. This resistance encompassed penicillins, extended-spectrum cephalosporins, carbapenems, monobactams, and aminoglycosides. These results appear to show that the imipenem resistance observed among nine Korean A. baumannii isolates could be attributed to the spread of an IMP-1- or OXA-23-producing clone. Our microbiological test of carbapenemase activity is a simple method for the screening of clinical isolates producing class D carbapenemase and/or class B metallo-β-lactamase, in order both to determine their clinical impact and to prevent further spread.
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