Simultaneous single-tube PCR assay for the detection of Neisseria meningitidis, Haemophilus influenzae type b and Streptococcus pneumoniae

Abstract

Rapid, accurate and inexpensive diagnosis of bacterial meningitis is critical for patient management. This study describes the development and evaluation of a multiplex PCR assay for the detection of Neisseria meningitidis, Streptococcus pneumoniae and Haemophilus influenzae type b, which globally account for 90% of cases of bacterial meningitis. The single-tube assay, based on the ctrA, ply and bex targets, respectively, enabled detection of 5-10 pg DNA. When the assay was tested with clinical samples (n = 425), its sensitivity for the three targets was 93.9%, 92.3% and 88%, respectively, while the overall specificity and positive predictive value of the assay was 100%. The negative predictive value was 99.1-99.5%. The methodology permits rapid and accurate detection of the three main pathogens that cause bacterial meningitis.

DNA bacterial load in children and adolescents with pneumococcal pneumonia and empyema

“…[17] have published an accurate meta-analysis and found that the studies were highly heterogeneously including different type of PCR methods [18,19], different target genes [20], most used stored frozen [21] and different samples (i.e. whole blood or plasma) [22], as well as different patients' characteristics and different clinical syndromes [23,24]. The authors concluded that "the lack of appropriate reference standard might have caused underestimation of the performance of the PCR" and that "currently available methods for PCR with blood samples for the diagnosis of IPD lack the sensitivity and specificity necessary for clinical practice".…”

Simultaneous Detection of Bacterial Meningitis in Suspected Cases of Meningitis in Children Using PCR Assay, In Taif, Saudi Arabia

“…This failure to isolate bacteria by CSF culture and the risk of false negative result were high because either only a small number of microorganisms may be present in the specimens or the low quality of CSF samples and the use of antibiotics prior to the lumbar puncture. It might be due also to differences in hospital practices with respect to infection control, difference in methodology, patient characteristics and spectrum of bacterial pathogens [20,27]. Hereby, biochemical analyzes support meningitis in 102 out of 300 samples (whose culture was negative) however bacterial meningitis were detected, by PCR, only in 12 cases.…”

“…The specific gene targets, the primer sets and mPCR conditions were as previously described [18][19][20]. The PCR products were visualized on 2% agarose gels stained with ethidium bromide.…”

Neuro-Intensive Care of Patients with Acute CNS Infections

“…PCR assays for common bacterial ribosomal sequences exhibit promise as a rapid diagnostic tool, but are subject to false-positive results [27,28]. PCR assays may be useful in cases of bacterial meningitis with negative Gram stain and culture or in cases with prior antibiotic therapy.…”