Detection of Brucellae in Blood Cultures

Yagupsky, Pablo

doi:10.1128/jcm.37.11.3437-3442.1999

Cited by 230 publications

(106 citation statements)

References 30 publications

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“…Such amounts of DNA can be expected in any blood sample from a patient with active brucellosis. The precision of the assay, with an intra-assay variation of < 1.5% and an inter-assay variation of < 6.5%, was similar to or lower than that of other PCR assays based on similar principles [23], while the diagnostic yield was 91.9%, i.e., far higher than that for blood cultures [3,4]. Melting curve analysis increased the specificity of the assay by confirming that a positive fluorescence signal obtained during the real-time PCR was associated with an amplified product with a characteristic T m .…”

Section: Discussionmentioning

confidence: 64%

Rapid diagnosis of human brucellosis by SYBR Green I-based real-time PCR assay and melting curve analysis in serum samples

Queipo-Ortuño

Colmenero²,

Reguera³

et al. 2005

Clinical Microbiology and Infection

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The aim of this study was to develop a LightCycler-based real-time PCR (LC-PCR) assay and to evaluate its diagnostic use for the detection of Brucella DNA in serum samples. Following amplification of a 223-bp gene sequence encoding an immunogenetic membrane protein (BCSP31) specific for the Brucella genus, melting curve and DNA sequencing analysis was performed to verify the specificity of the PCR products. The intra- and inter-assay variation coefficients were 1.3% and 6.4%, respectively, and the detection limit was 5 fg of Brucella DNA (one genome equivalent). After optimisation of the PCR assay conditions, a standard curve was obtained with a linear range (correlation coefficient=0.99) over seven orders of magnitude from 10(7) to 10 fg of Brucella DNA. The LC-PCR assay was found to be 91.9% sensitive and 95.4% specific when tested with 65 negative control samples and 62 serum samples from 60 consecutive patients with active brucellosis. The assay is reproducible, easily standardised, minimises the risk of infection in laboratory workers, and has a total processing time of <2 h. It could therefore form a promising and practical approach for the rapid diagnosis of human brucellosis.

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Section: Discussionmentioning

confidence: 64%

Rapid diagnosis of human brucellosis by SYBR Green I-based real-time PCR assay and melting curve analysis in serum samples

Queipo-Ortuño

Colmenero²,

Reguera³

et al. 2005

Clinical Microbiology and Infection

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“…Attempts to isolate Brucella strains by blood culture were not successful due to the limited efficacy of Brucella isolation from blood samples processed after considerable delay. 23 However, Brucella DNA was detected by qPCR in 14 of 135 seroreactive plasma samples, 13 of which were confirmed by nested PCR and amplicon sequences of Brucella genome. As shown in Table 1, Brucella DNA-positive results were found mostly in samples reactive with RBPT, SAT, and ELISA or SAT, ELISA, and WB reactive (11/13) but two samples were reactive only with RBPT, suggesting that, despite its apparently poor specificity, RBPT could be utilized as primary screening assay and that SAT would not provide additional safety or increase specificity.…”

Section: Discussionmentioning

confidence: 92%

Potential risk of blood transfusion‐transmitted brucellosis in an endemic area of China

Wang

Liao

et al. 2014

Transfusion

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“…Osteomyelitis, although not as common as arthritis, is also observed in human patients with brucellosis [9,10,60,61], and the recovery of Brucella spp. from bone marrow in brucellosis patients is more common than recovery from the blood [62]. Minimal damage to cartilage was observed, and mice did not produce robust cytokine or antibody responses (data not shown) to CII collagen.…”

Section: Discussionmentioning

confidence: 96%

IFN-γ-deficient mice develop IL-1-dependent cutaneous and musculoskeletal inflammation during experimental brucellosis

Skyberg

Thornburg

Kochetkova

et al. 2012

Journal of Leukocyte Biology

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Human brucellosis exhibits diverse pathological manifestations that can affect almost any organ. In particular, osteoarticular complications are the most common focal manifestation of brucellosis and occur in 40-80% of patients. In immunocompetent mice, Brucella replication is generally restricted to the spleen, liver, and to a lesser extent, LNs, thereby limiting their use for study of focal inflammation often found in brucellosis. Here, we report that nasal, oral, or peritoneal infection of IFN-γ(-/-) mice with WT Brucella melitensis or Brucella abortus results in joint and periarticular tissue inflammation. Histological analysis of the affected joints revealed inflammatory infiltrates and debris within the joint space colocalizing with Brucella antigen. Osteoarthritis, necrosis, periarticular soft tissue inflammation, and substantial brucellae burdens were observed. Oral rifampicin was effective in clearing infection and halting further progression of focal inflammation from infected IFN-γ(-/-) mice, although some symptoms and swelling remained. Elevated IL-1 β, but not TNF-α, IL-6, or IL-17, was detected in joint homogenates from infected IFN-γ(-/-) mice. Whereas more susceptible to systemic infection, IL-1R(-/-) mice depleted of IFN-γ were more resistant to focal inflammation than WT mice similarly depleted of IFN-γ. Collectively, these results show IFN-γ(-/-) mice represent a potential model for study of focal inflammation attributed to Brucella infection and will allow evaluation of intervention strategies targeting IL-1, IL-1R, or other inflammatory mediators, with the potential to complement antibiotic-based therapies.

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Detection of Brucellae in Blood Cultures

Cited by 230 publications

References 30 publications

Rapid diagnosis of human brucellosis by SYBR Green I-based real-time PCR assay and melting curve analysis in serum samples

Rapid diagnosis of human brucellosis by SYBR Green I-based real-time PCR assay and melting curve analysis in serum samples

Potential risk of blood transfusion‐transmitted brucellosis in an endemic area of China

IFN-γ-deficient mice develop IL-1-dependent cutaneous and musculoskeletal inflammation during experimental brucellosis

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