Direct identification of bacterial isolates in blood cultures by using a DNA probe

Davis, Thomas E.; Fuller, D.

doi:10.1128/jcm.29.10.2193-2196.1991

Cited by 74 publications

(21 citation statements)

References 23 publications

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“…Similar to our study, direct identification of bacterial isolates in blood cultures by chemiluminescent DNA probes that detect the rRNAs of certain target organisms has been reported previously (5). However, the method used in the present study is easier and more rapid.…”

Section: Discussionsupporting

confidence: 83%

Fluorescent In Situ Hybridization Allows Rapid Identification of Microorganisms in Blood Cultures

2000

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Using fluorescent in situ hybridization (FISH) with rRNA-targeted fluorescently labelled oligonucleotide probes, pathogens were rapidly detected and identified in positive blood culture bottles without cultivation and biotyping. In this study, 115 blood cultures with a positive growth index as determined by a continuous-reading automated blood culture system were examined by both conventional laboratory methods and FISH. For this purpose, oligonucleotide probes that allowed identification of approximately 95% of those pathogens typically associated with bacteremia were produced. The sensitivity and specificity of these probes were 100%. From all 115 blood cultures, microorganisms were grown after 1 day and identification to the family, genus, or species level was achieved after 1 to 3 days while 111 samples (96.5%) were similarly identified by FISH within 2.5 h. Staphylococci were identified in 62 of 62 samples, streptococci and enterococci were identified in 19 of 20 samples, gram-negative rods were identified in 28 of 30 samples, and fungi were identified in two of two samples. Thus, FISH is an appropriate method for identification of pathogens grown in blood cultures from septicemic patients.

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Section: Discussionsupporting

confidence: 83%

Fluorescent In Situ Hybridization Allows Rapid Identification of Microorganisms in Blood Cultures

2000

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“…The precise identi¢cation of enterococci to the species level is important in the management of patients with enterococcal infections [11]. Tests for identi¢cation of enterococci based on biochemical characteristics, analysis of whole cell protein pro¢les [12,30] or commercially available enterococcal ribosomal RNA (rRNA) based oligonucleotide DNA probes [31,32] and intragenic DNA probes for gyrA and aac(6P)-li [33] have been successfully used for the di¡erentiation of enterococcal species. Problems arise when isolates display unusual fermentation behavior or atypical phenotypic characteristics [11].…”

Section: E Faecium Efaa Sequencementioning

confidence: 99%

In vivo testing of an Enterococcus faecalis efaA mutant and use of efaA homologs for species identification

Singh¹

1998

FEMS Immunology and Medical Microbiology

105

136

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Disruption of the previously described efaA (from Enterococcus faecalis antigen A) gene was generated in E. faecalis strain OG1RF and loss of an V37-kDa immunoreactive band from the mutant was demonstrated in Western blots. In a mouse peritonitis model, mice infected with the efaA fs (fs=from Enterococcus faecalis) mutant showed more prolonged survival than mice infected with the parent strain OG1RF. These results suggest that efaA fs encodes a function important for infection of mice by enterococci. An efaA-like gene was also identified in E. faecium DNA libraries and its deduced amino acid sequence showed 73% similarity to EfaA of E. faecalis and 42^63% similarities to a group of streptococcal virulence and adhesion associated proteins that are components of ATP-binding cassette transport systems. Intragenic probes representing efaA fs , recA fs , efaA fm (fm=from E. faecium) and gyrA fm were tested for their ability to identify E. faecalis and E. faecium using colony lysates of 133 enterococci and one Streptococcus sp. Probes of E. faecium and E. faecalis origin hybridized to all isolates of E. faecium and E. faecalis, respectively, regardless of their clinical source but not to any of 29 other enterococci. These results suggest that the use of gene probes may prove helpful in identification of isolates of E. faecium and E. faecalis. z

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“…The usual time for detection of S. pneumoniae in a blood culture is 24 to 48 h (8). Identification of the cultured bacteria is conducted with modified conventional tests, commercial immunological kits, and commercial DNA probe kits (2)(3)(4)24). Multiple-step procedures are involved in all of these methods, usually adding an additional 24 h for final identification of the organism.…”

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confidence: 99%

Detection of Streptococcus pneumoniae in whole blood by PCR

et al. 1995

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Streptococcus pneumoniae is a major cause of bacteremia in both children and adults. Currently, the diagnosis of pneumococcal bacteremia relies on the isolation and identification of the bacteria from blood cultures. We have developed a sensitive assay for the detection of S. pneumoniae in whole blood by the PCR. A specific primer-probe set (JM201 and JM202 primers with JM204 probe) designed from the penicillin-binding protein 2B gene was demonstrated to reproducibly detect between 10 and 100 fg of input purified S. pneumoniae DNA. This assay system was shown to be inclusive for all strains of S. pneumoniae evaluated, including 15 different serotypes and a battery of penicillin-resistant and -sensitive strains. The specificity of this PCR-based assay was demonstrated by its inability to support amplification from a series of human, bacterial, and yeast genomic DNAs. A general specimen preparation method which should be suitable for the purification of DNA from any pathogens in whole blood was developed. With this protocol it was possible to detect S. pneumoniae-specific DNA from whole blood specimens inoculated with as little as 4 CFU/ml. Copurified human blood DNA, ranging from 0 to 4.5 g per PCR, did not affect the sensitivity of S. pneumoniae detection by PCR. A blinded clinical trial was used to compare the PCR-based assay with standard microbiological blood culture for the detection of S. pneumoniae bacteremia in 36 specimens obtained from pediatric patients seen in the emergency room of Children's Hospital of Pittsburgh. With culture as the ''gold standard,'' the PCR-based assay had a sensitivity of 80% (4 of 5 culture-positive specimens were PCR positive) and a specificity of 84% (26 of 31 culture-negative specimens were PCR negative). However, three patients whose specimens were PCR positive and culture negative had histories suggestive of bacteremia, including recent positive blood cultures, treatment with antibiotics, cellulitis, and multiple emergency room visits for fever within a 24-h period. These data suggest that PCR-based assays for S. pneumoniae may prove useful to augment current methods of detection for S. pneumoniae bacteremia.

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Direct identification of bacterial isolates in blood cultures by using a DNA probe

Cited by 74 publications

References 23 publications

Fluorescent In Situ Hybridization Allows Rapid Identification of Microorganisms in Blood Cultures

Fluorescent In Situ Hybridization Allows Rapid Identification of Microorganisms in Blood Cultures

In vivo testing of an Enterococcus faecalis efaA mutant and use of efaA homologs for species identification

Detection of Streptococcus pneumoniae in whole blood by PCR

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