Utility of a novel multiplex TaqMan PCR assay for metallo-β-lactamase genes plus other TaqMan assays in detecting genes encoding serine carbapenemases and clinically significant extended-spectrum β-lactamases

The prevalence of CTX-M-producing members of the Enterobacteriaceae is increasing worldwide. A novel, multiplex, real-time TaqMan PCR assay to detect and type bla CTX-M genes is described which is an improvement on previously described techniques with respect to reduced assay time, elimination of the need for protracted post-PCR processing and the convenience of a single reaction vessel. Based on β-lactam antibiogram and MIC data, 478 of 1279 Enterobacteriaceae isolates from clinical blood and urine culture specimens were selected and tested for extended-spectrum β-lactamase (ESBL) production using phenotypic methods. The new TaqMan assay detected and typed bla CTX-M genes in 21 of 28 ESBL-producing isolates.

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Detection of Genes Encoding of Extended-Spectrum and AmpC β-Lactamases in Klebsiella pneumoniae Isolates from Clinical Specimens

Journal of Al-Nahrain University-Science ◽

10.22401/jnus.18.2.16 ◽

2015 ◽

Vol 18 (2) ◽

pp. 125-132

Author(s):

Ghusoon A. Abdulhasan ◽

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Hula Y. Fadhil ◽

Kifah A. Jasem ◽

◽

...

Keyword(s):

Klebsiella Pneumoniae ◽

Clinical Specimens ◽

Extended Spectrum ◽

Genes Encoding

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Real-time TaqMan PCR for rapid detection of genes encoding five types of non-metallo- (class A and D) carbapenemases in Enterobacteriaceae

International Journal of Antimicrobial Agents ◽

10.1016/j.ijantimicag.2011.03.010 ◽

2011 ◽

Vol 38 (1) ◽

pp. 35-38 ◽

Cited By ~ 55

Author(s):

R.L. Swayne ◽

H.A. Ludlam ◽

V.G. Shet ◽

N. Woodford ◽

M.D. Curran

Keyword(s):

Real Time ◽

Rapid Detection ◽

Class A ◽

Genes Encoding ◽

Taqman Pcr

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Draft Genome Sequences of Two Enterobacter cloacae subsp. cloacae Strains Isolated from Australian Hematology Patients with Bacteremia

Genome Announcements ◽

10.1128/genomea.00756-17 ◽

2017 ◽

Vol 5 (33) ◽

Cited By ~ 1

Author(s):

Lex E. X. Leong ◽

David Shaw ◽

Lito Papanicolas ◽

Diana Lagana ◽

Ivan Bastian ◽

...

Keyword(s):

Gut Microbiota ◽

Draft Genome ◽

Enterobacter Cloacae ◽

Opportunistic Pathogen ◽

Healthy Individuals ◽

Genome Sequences ◽

Content Type ◽

Extended Spectrum ◽

Genes Encoding

ABSTRACT Enterobacter cloacae is a common member of the gut microbiota in healthy individuals. However, it is also an opportunistic pathogen, capable of causing bacteremia. We report the draft genomes of two Enterobacter cloacae subspecies cloacae strains isolated from hematology patients with bacteremia. Both isolates carry genes encoding extended-spectrum β-lactamases.

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Automated 5′ Nuclease PCR Assay for Identification of Salmonella enterica

Journal of Clinical Microbiology ◽

10.1128/jcm.38.9.3429-3435.2000 ◽

2000 ◽

Vol 38 (9) ◽

pp. 3429-3435 ◽

Cited By ~ 160

Author(s):

J. Hoorfar ◽

P. Ahrens ◽

P. Rådström

Keyword(s):

Salmonella Enterica ◽

Relative Sensitivity ◽

Positive Control ◽

Taqman Assay ◽

Diagnostic Specificity ◽

Pcr Assay ◽

Sample Tube ◽

Simple Alternative ◽

Taqman Pcr ◽

Internal Positive Control

A simple and ready-to-go test based on a 5′ nuclease (TaqMan) PCR technique was developed for identification of presumptiveSalmonella enterica isolates. The results were compared with those of conventional methods. The TaqMan assay was evaluated for its ability to accurately detect 210 S. enterica isolates, including 100 problematic “rough” isolates. An internal positive control was designed to use the same Salmonella primers for amplification of a spiked nonrelevant template (116 bp) in the sample tube. The PCR test correctly identified all the Salmonellastrains by resulting in positive end-point fluorescence (FAM) signals for the samples and positive control (TET) signals (relative sensitivity [ΔRn], >0.6). The diagnostic specificity of the method was assessed using 120 non-Salmonella strains, which all resulted in negative FAM signals (ΔRn, ≤0.5). All 100 roughSalmonella strains tested resulted in positive FAM and TET signals. In addition, it was found that the complete PCR mixture, predispensed in microwell plates, could be stored for up to 3 months at −20°C. Thus, the diagnostic TaqMan assay developed can be a useful and simple alternative method for identification ofSalmonella, particularly in large reference laboratories.

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Identification of Streptococci to Species Level by Sequencing the Gene Encoding the Manganese-Dependent Superoxide Dismutase

Journal of Clinical Microbiology ◽

10.1128/jcm.36.1.41-47.1998 ◽

1998 ◽

Vol 36 (1) ◽

pp. 41-47 ◽

Cited By ~ 201

Author(s):

Claire Poyart ◽

Gilles Quesne ◽

Stephane Coulon ◽

Patrick Berche ◽

Patrick Trieu-Cuot

Keyword(s):

Superoxide Dismutase ◽

Pcr Assay ◽

Degenerate Primers ◽

Gene Encoding ◽

Genes Encoding ◽

Type Strains ◽

Internal Fragment ◽

Rrna Sequences ◽

16S Rrna Sequences

We have used a PCR assay based on the use of degenerate primers in order to characterize an internal fragment (sodAint ) representing approximately 85% of the genes encoding the manganese-dependent superoxide dismutase in various streptococcal type strains (S. acidominimus,S. agalactiae, S. alactolyticus, S. anginosus, S. bovis, S. constellatus,S. canis, S. cricetus, S. downei,S. dysgalactiae, S. equi subsp.equi, S. equi subsp. zooepidemicus,S. equinus, S. gordonii, S. iniae,S. intermedius, S. mitis, S. mutans, S. oralis, S. parasanguis,S. pneumoniae, S. porcinus, S. pyogenes, S. salivarius, S. sanguis,S. sobrinus, S. suis, S. thermophilus, and S. vestibularis). Phylogenetic analysis of these sodAint fragments yields an evolutionary tree having a topology similar to that of the tree constructed with the 16S rRNA sequences. We have shown that clinical isolates could be identified by determining the positions of theirsodAint fragments on the phylogenetic tree of the sodAint fragments of the type species. We propose this method for the characterization of strains that cannot be assigned to a species on the basis of their conventional phenotypic reactions.

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