Detection of Streptococcus pneumoniae and Haemophilus influenzae type B by real-time PCR from dried blood spot samples among children with pneumonia: a useful approach for developing countries

PLoS One. 2013 Oct 8;8(10):e76970. doi: 10.1371/journal.pone.0076970. eCollection 2013.

Abstract

Background: Dried blood spot (DBS) is a reliable blood collection method for storing samples at room temperature and easily transporting them. We have previously validated a Real-Time PCR for detection of Streptococcus pneumoniae in DBS. The objective of this study was to apply this methodology for the diagnosis of S. pneumoniae and Haemophilus influenzae b (Hib) in DBS samples of children with pneumonia admitted to two hospitals in Mozambique and Morocco.

Methods: Ply and wzg genes of S. pneumoniae and bexA gene of Hib, were used as targets of Real-Time PCR. 329 DBS samples of children hospitalized with clinical diagnosis of pneumonia were tested.

Results: Real-Time PCR in DBS allowed for a significant increase in microbiological diagnosis of S. pneumoniae and Hib. When performing blood bacterial culture, only ten isolates of S. pneumoniae and none of Hib were detected (3·0% positivity rate, IC95% 1·4-5·5%). Real-Time PCR from DBS samples increased the detection yield by 4x fold, as 30 S. pneumoniae and 11 Hib cases were detected (12·4% positivity rate, IC95% 9·0-16·5%; P<0·001).

Conclusion: Real-Time PCR applied in DBS may be a valuable tool for improving diagnosis and surveillance of pneumonia caused by S. pneumoniae or Hib in developing countries.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • ATP-Binding Cassette Transporters / genetics
  • Bacterial Proteins / genetics
  • Blood Specimen Collection / methods
  • Child
  • Child, Preschool
  • DNA, Bacterial / blood
  • DNA, Bacterial / genetics
  • Developing Countries
  • Female
  • Haemophilus Infections / blood
  • Haemophilus Infections / diagnosis
  • Haemophilus Infections / microbiology*
  • Haemophilus influenzae type b / genetics*
  • Haemophilus influenzae type b / isolation & purification
  • Hospitals
  • Humans
  • Infant
  • Male
  • Morocco
  • Mozambique
  • Pneumonia, Pneumococcal / blood
  • Pneumonia, Pneumococcal / diagnosis
  • Pneumonia, Pneumococcal / microbiology*
  • Real-Time Polymerase Chain Reaction
  • Reproducibility of Results
  • Sensitivity and Specificity
  • Streptococcus pneumoniae / genetics*
  • Streptococcus pneumoniae / isolation & purification
  • Streptolysins / genetics

Substances

  • ATP-Binding Cassette Transporters
  • Bacterial Proteins
  • CpsA protein, Streptococcus pneumoniae
  • DNA, Bacterial
  • Streptolysins
  • plY protein, Streptococcus pneumoniae
  • bexA protein, Haemophilus influenzae

Grants and funding

This work was supported in part by Fondo de Investigaciones Sanitarias [PI10/02058 and a Research Grant to QB], Agència de Gestió d'Ajuts Universitaris i de Recerca [SGR00136 and FI-ICIP 2012 to LS] and Fundació Sant Joan de Déu [Research Grant to LS]. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.