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.jpg): Failed to open stream: No such file or directory in C:\Inetpub\vhosts\kidney.de\httpdocs\pget.php on line 117 J+Biomol+Tech
2016 ; 27
(4
): 132-137
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Establishment of a Simple and Quick Method for Detecting Extended-Spectrum
?-Lactamase (ESBL) Genes in Bacteria
#MMPMID27672351
Han ST
; Fei Y
; Huang JY
; Xu M
; Chen LC
; Liao DJ
; Tan YJ
J Biomol Tech
2016[Dec]; 27
(4
): 132-137
PMID27672351
show ga
Extended-spectrum ?-lactamase (ESBL) genes that render bacteria resistant to
antibiotics are commonly detected using phenotype testing, which is time
consuming and not sufficiently accurate. To establish a better method, we used
phenotype testing to identify ESBL-positive bacterial strains and conducted PCR
to screen for TEM (named after the patient Temoneira who provided the first
sample), sulfhydryl reagent variable (SHV), cefotaxime (CTX)-M-1, and CTX-M-9,
the 4 most common ESBL types and subtypes. We then performed multiplex PCR with 1
primer containing a biotin and hybridized the PCR products with gene-specific
probes that were coupled with microbeads and coated with a specific fluorescence.
The hybrids were linked to streptavidin-R-phycoerythrins (SA-PEs) and run through
a flow cytometer, which sorted the fluorescently dyed microbeads and quantified
the PEs. The results from single PCR, multiplex PCR, and cytometry were
consistent with each other. We used this method to test 169 clinical specimens
that had been determined for phenotypes and found 154 positive for genotypes,
including 30 of the 45 samples that were negative for phenotypes. The CTX-M
genotype tests alone, counting both positive and negative cases, showed 99.41%
(168/169) consistency with the ESBL phenotype test. Thus, we have established a
multiplex-PCR system as a simple and quick method that is high throughput and
accurate for detecting 4 common ESBL types and subtypes.
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