Conventional and Molecular Methods for Verification of Results Obtained with BacT/Alert Nonvent Blood Culture Bottles

Marlowe, Elizabeth M.; Gibson, Linda; Hogan, James J.; Kaplan, Shannon; Bruckner, David A.

doi:10.1128/jcm.41.3.1266-1269.2003

Cited by 14 publications

(12 citation statements)

References 14 publications

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“…Currently, inoculation of blood cultures (BC) is the standard method for microbiological diagnosis of bloodstream infections. However, the limitations of BC include relatively low sensitivities and a long time-to-result for detection and identification of the pathogen, generally over 2 days, and even longer for fastidious organisms (13,20,27).In contrast, DNA-based procedures may offer faster and more reliable diagnoses (3, 30). PCR amplification of microbial genes, followed by detection of amplified products by gel electrophoresis or real-time PCR monitoring using fluorescent dyes or target-directed fluorescent probes, is a quick process allowing pathogen detection within a few hours (18).…”

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confidence: 99%

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Diagnosis of Bacteremia in Whole-Blood Samples by Use of a Commercial Universal 16S rRNA Gene-Based PCR and Sequence Analysis

et al. 2009

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In a prospective, multicenter study of 342 blood samples from 187 patients with systemic inflammatory response syndrome, sepsis, or neutropenic fever, a new commercial PCR test (SepsiTest; Molzym) was evaluated for rapid diagnosis of bacteremia. The test comprises a universal PCR from the 16S rRNA gene, with subsequent identification of bacteria from positive samples by sequence analysis of amplicons. Compared to blood culture (BC), the diagnostic sensitivity and specificity of the PCR were 87.0 and 85.8%, respectively. Considering the 34 BC-positive patients, 28 were also PCR positive in at least one of the samples, resulting in a patient-related sensitivity of 82.4%. The concordance of PCR and BC for both positive and negative samples was (47 ؉ 247)/342, i.e., 86.0%. In total, 31 patients were PCR/sequencing positive and BC negative, in whom the PCR result was judged as possible or probable to true bacteremia in 25. In conclusion, the PCR approach facilitates the detection of bacteremia in blood samples within a few hours. Despite the indispensability of BC diagnostics, the rapid detection of bacteria by SepsiTest appears to be a valuable tool, allowing earlier pathogen-adapted antimicrobial therapy in critically ill patients.Bloodstream infection is a life-threatening condition with a high mortality rate, especially in intensive care and neutropenic patients (5,19,35,38). Pathogenic bacteria are the most frequent causes of bloodstream infection, although fungi can also be isolated in a minority of patients (7,17,21,32,34). Currently, inoculation of blood cultures (BC) is the standard method for microbiological diagnosis of bloodstream infections. However, the limitations of BC include relatively low sensitivities and a long time-to-result for detection and identification of the pathogen, generally over 2 days, and even longer for fastidious organisms (13,20,27).In contrast, DNA-based procedures may offer faster and more reliable diagnoses (3, 30). PCR amplification of microbial genes, followed by detection of amplified products by gel electrophoresis or real-time PCR monitoring using fluorescent dyes or target-directed fluorescent probes, is a quick process allowing pathogen detection within a few hours (18). Identification of microorganisms can be performed by PCR algorithms, taxon-specific oligonucleotide microarrays, or sequencing amplicons (30).PCR amplification of conserved regions of the bacterial genome, in particular the 16S rRNA gene, combined with sequence analysis is a well-established technique for the identification of bacterial pathogens (18). The main advantages of targeting the 16S rRNA gene are the broad range of pathogens detectable and the independence of this method from the in vitro viability of strains (6). The high sensitivity of detection by PCR of bacterial DNA (15) suggests its use in the diagnosis of bacteremia (16). Initial disadvantages of PCR, notably the incidence of false-positive results from bacterial DNA contaminating PCR reagents (4, 39), have been counteracted by the devel...

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Diagnosis of Bacteremia in Whole-Blood Samples by Use of a Commercial Universal 16S rRNA Gene-Based PCR and Sequence Analysis

et al. 2009

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“…Based on literature survey, and as confirmed in this study, this probe set identifies Ͼ98.5% of Staphylococcus samples containing positive blood cultures to the species or group level (13,16,19,27,38,42). The limits of detection (LOD) for all staphylococci detected with species-specific probes were 10 to 1,000-fold below the level required to give positive results in automated blood culture systems using pure cultures (22,24). In mixed cultures, limits of detection were observed to be up to 10-to 30-fold higher.…”

Section: Discussionmentioning

confidence: 86%

Staph ID/R: a Rapid Method for Determining Staphylococcus Species Identity and Detecting the mecA Gene Directly from Positive Blood Culture

Pasko¹,

Hicke²,

Dunn³

et al. 2012

J Clin Microbiol

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Rapid diagnosis of staphylococcal bacteremia directs appropriate antimicrobial therapy, leading to improved patient outcome. We describe herein a rapid test (<75 min) that can identify the major pathogenic strains of Staphylococcus to the species level as well as the presence or absence of the methicillin resistance determinant gene, mecA. The test, Staph ID/R, combines a rapid isothermal nucleic acid amplification method, helicase-dependent amplification (HDA), with a chip-based array that produces unambiguous visible results. The analytic sensitivity was 1 CFU per reaction for the mecA gene and was 1 to 250 CFU per reaction depending on the staphylococcal species present in the positive blood culture. Staph ID/R has excellent specificity as well, with no cross-reactivity observed. We validated the performance of Staph ID/R by testing 104 frozen clinical positive blood cultures and comparing the results with rpoB gene or 16S rRNA gene sequencing for species identity determinations and mecA gene PCR to confirm mecA gene results. Staph ID/R agreed with mecA gene PCR for all samples and agreed with rpoB/16S rRNA gene sequencing in all cases except for one sample that contained a mixture of two staphylococcal species, one of which Staph ID/R correctly identified, for an overall agreement of 99.0% (P < 0.01). Staph ID/R could potentially be used to positively affect patient management for Staphylococcus-mediated bacteremia.

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“…Organon Tecnica replaced its vented BacT/Alert aerobic blood culture bottle with a redesigned nonvent blood culture bottle in 2000, so the BacT/Alert FAN nonvent aerobic blood culture bottle was adopted in our blood culture routine in February 2001. These two types of bottles (nonvent and vented) have, however, been shown to be equivalent (10,19).…”

Section: Methodsmentioning

confidence: 99%

Direct Identification of Gram-Positive Cocci from Routine Blood Cultures by Using AccuProbe Tests

Lindholm

Sarkkinen

2004

J Clin Microbiol

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Rapid and reliable identification of bacteria directly from blood cultures is important in clinical practice to guide appropriate antibiotic therapy. In this study, the performance of the AccuProbe (Gen-Probe, Inc., San Diego, Calif.) in direct identification of Staphylococcus aureus, Streptococcus pneumoniae, enterococci, and group A and B streptococci from positive blood culture bottles was evaluated by using 6-year routine clinical laboratory blood culture material from Päijät-Häme Central Hospital, Lahti, Finland. With the enterococcal and group A and B streptococcal probes, the diagnostic performance of the test was excellent at a cutoff value of 50,000 relative light units (RLU) as recommended by the manufacturer. However, with the S. aureus probe, although the specificity was very high (99.8%), the sensitivity was low (72.4%). To improve the clinical usability of the direct AccuProbe identification, optimal cutoff values for the individual AccuProbe tests were defined by using receiver-operating characteristic analysis. Consequently, cutoff values for S. aureus and S. pneumoniae tests were adjusted to 30,000 RLU and for enterococci and to 55,000 RLU for group A and B streptococci. With these adjustments, the performance of the AccuProbe tests, especially that for S. aureus, was significantly improved.Rapid and reliable identification of bacteria from blood culture is indisputably one of the most important functions of a clinical microbiology laboratory. Bloodstream infection is a potentially life-threatening condition that requires rapid, appropriate therapy. The increasing rate of nosocomial infections during the past decade (21), together with the threat from multiresistant infectious agents, further highlight the importance of early identification and tailored antimicrobial therapy.

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Conventional and Molecular Methods for Verification of Results Obtained with BacT/Alert Nonvent Blood Culture Bottles

Cited by 14 publications

References 14 publications

Diagnosis of Bacteremia in Whole-Blood Samples by Use of a Commercial Universal 16S rRNA Gene-Based PCR and Sequence Analysis

Diagnosis of Bacteremia in Whole-Blood Samples by Use of a Commercial Universal 16S rRNA Gene-Based PCR and Sequence Analysis

Staph ID/R: a Rapid Method for Determining Staphylococcus Species Identity and Detecting the mecA Gene Directly from Positive Blood Culture

Direct Identification of Gram-Positive Cocci from Routine Blood Cultures by Using AccuProbe Tests

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