A multiplex PCR for detection of Mycoplasma pneumoniae, Chlamydophila pneumoniae, Legionella pneumophila, and Bordetella pertussis in clinical specimens

McDonough, Erin A.; Barrozo, Christopher P.; Russell, Kevin L.; Metzgar, David

doi:10.1016/j.mcp.2005.05.002

Cited by 47 publications

(13 citation statements)

References 35 publications

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“…However, the study specimens showed virtually no carriage of the other nine bacterial and viral pathogens. The findings of this work, combined with those in previous reports showing good sensitivities and specificities for symptomatic patients (11,20,24,25,29,40), suggest that molecular assays testing for these nine pathogens in respiratory secretions will have good utility in clinical practice. The low rate of detection of NA in serum is reassuring in the presence of comparably high rates of detection in specimens from the nasopharynx and skin, sites that have been implicated previously as potentially contributing to the detection of pathogens in serum.…”

Section: Discussionsupporting

confidence: 67%

Detection of 11 Common Viral and Bacterial Pathogens Causing Community-Acquired Pneumonia or Sepsis in Asymptomatic Patients by Using a Multiplex Reverse Transcription-PCR Assay with Manual (Enzyme Hybridization) or Automated (Electronic Microarray) Detection

et al. 2008

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Community-acquired pneumonia (CAP) and sepsis are important causes of morbidity and mortality. We describe the development of two molecular assays for the detection of 11 common viral and bacterial agents of CAP and sepsis: influenza virus A, influenza virus B, respiratory syncytial virus A (RSV A), RSV B, Mycoplasma pneumoniae, Chlamydophila pneumoniae, Legionella pneumophila, Legionella micdadei, Bordetella pertussis, Staphylococcus aureus, and Streptococcus pneumoniae. Further, we report the prevalence of carriage of these pathogens in respiratory, skin, and serum specimens from 243 asymptomatic children and adults. The detection of pathogens was done using both a manual enzyme hybridization assay and an automated electronic microarray following reverse transcription and PCR amplification. The analytical sensitivities ranged between 0.01 and 100 50% tissue culture infective doses, cells, or CFU per ml for both detection methods. Analytical specificity testing demonstrated no significant cross-reactivity among 19 other common respiratory organisms. One hundred spiked "surrogate" clinical specimens were all correctly identified with 100% specificity (95% confidence interval, 100%). Overall, 28 (21.7%) of 129 nasopharyngeal specimens, 11 of 100 skin specimens, and 2 of 100 serum specimens from asymptomatic subjects tested positive for one or more pathogens, with S. pneumoniae and S. aureus giving 89% of the positive results. Our data suggest that asymptomatic carriage makes the use of molecular assays problematic for the detection of S. pneumoniae or S. aureus in upper respiratory tract secretions; however, the specimens tested showed virtually no carriage of the other nine viral and bacterial pathogens, and the detection of these pathogens should not be a significant diagnostic problem. In addition, slightly less sensitive molecular assays may have better correlation with clinical disease in the case of CAP.

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Section: Discussionsupporting

confidence: 67%

Detection of 11 Common Viral and Bacterial Pathogens Causing Community-Acquired Pneumonia or Sepsis in Asymptomatic Patients by Using a Multiplex Reverse Transcription-PCR Assay with Manual (Enzyme Hybridization) or Automated (Electronic Microarray) Detection

et al. 2008

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“…This involves development of assays that are symptom based and can detect multiple respiratory pathogens, including B. pertussis (55)(56)(57)(58). A disadvantage of the latter approach can be that multiple sets of primers may reduce the sensitivity of detection.…”

Section: Pcr Assaysmentioning

confidence: 99%

Laboratory Diagnosis of Pertussis

2015

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SUMMARYThe introduction of vaccination in the 1950s significantly reduced the morbidity and mortality of pertussis. However, since the 1990s, a resurgence of pertussis has been observed in vaccinated populations, and a number of causes have been proposed for this phenomenon, including improved diagnostics, increased awareness, waning immunity, and pathogen adaptation. The resurgence of pertussis highlights the importance of standardized, sensitive, and specific laboratory diagnoses, the lack of which is responsible for the large differences in pertussis notifications between countries. Accurate laboratory diagnosis is also important for distinguishing between the several etiologic agents of pertussis-like diseases, which involve both viruses and bacteria. If pertussis is diagnosed in a timely manner, antibiotic treatment of the patient can mitigate the symptoms and prevent transmission. During an outbreak, timely diagnosis of pertussis allows prophylactic treatment of infants too young to be (fully) vaccinated, for whom pertussis is a severe, sometimes fatal disease. Finally, reliable diagnosis of pertussis is required to reveal trends in the (age-specific) disease incidence, which may point to changes in vaccine efficacy, waning immunity, and the emergence of vaccine-adapted strains. Here we review current approaches to the diagnosis of pertussis and discuss their limitations and strengths. In particular, we emphasize that the optimal diagnostic procedure depends on the stage of the disease, the age of the patient, and the vaccination status of the patient.

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“…Upon thawing nucleic acids were extracted from 0.2 mL of each NPA specimen using the High Pure Viral Nucleic Acid kit (Roche Diagnostics, Australia), according to the manufacturer's instructions. Monospecific PCR and reverse transcription PCR (RT-PCR) method were used to detect M. pneumoniae, C. pneumoniae, 13 rhinoviruses, enteroviruses, coronaviruses, 14 and human metapneumovirus (hMPV), 15 whereas multiplex PCR and RT-PCR was used to detect adenovirus, parainfluenza (1, 2, 3), influenza (A and B), and respiratory syncytial virus (RSV). 16 All these methods have been previously validated in our virology laboratory.…”

Section: Study Protocolmentioning

confidence: 99%

The Impact of Viral Respiratory Infection on the Severity and Recovery From an Asthma Exacerbation

Chang¹,

Clark²,

Acworth³

et al. 2009

Pediatric Infectious Disease Journal

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A multiplex PCR for detection of Mycoplasma pneumoniae, Chlamydophila pneumoniae, Legionella pneumophila, and Bordetella pertussis in clinical specimens

Cited by 47 publications

References 35 publications

Detection of 11 Common Viral and Bacterial Pathogens Causing Community-Acquired Pneumonia or Sepsis in Asymptomatic Patients by Using a Multiplex Reverse Transcription-PCR Assay with Manual (Enzyme Hybridization) or Automated (Electronic Microarray) Detection

Detection of 11 Common Viral and Bacterial Pathogens Causing Community-Acquired Pneumonia or Sepsis in Asymptomatic Patients by Using a Multiplex Reverse Transcription-PCR Assay with Manual (Enzyme Hybridization) or Automated (Electronic Microarray) Detection

Laboratory Diagnosis of Pertussis

The Impact of Viral Respiratory Infection on the Severity and Recovery From an Asthma Exacerbation

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