Update on the Detection and Characterization of Bacterial Pathogens by Nucleic Acid Amplification

Abstract

Real-time-based platforms currently offer numerous advantages over conventional nucleic acid amplification techniques (NAATs) such as higher speed, less handling of PCR products, decreased risk of false-positive results due to carryover contamination, and the capability to quantify results. This chapter reviews some updates on the detection by NAAT of some relevant individual agents alone, and associated with clinical syndromes, with emphasis on the detection of respiratory agents, particularly the atypical pathogens Mycobacterium pneumoniae, Chlamydophila pneumoniae, Legionella spp., and Bordetella pertussis but also Streptococcus pneumoniae, for which also quantitative real-time amplification is discussed. Given the many alternative amplification protocols proposed for some applications, such studies are clearly needed. Although such studies are often lacking, new-generation molecular techniques are gradually replacing tissue culture and even conventional PCRs as the gold standard for the diagnosis of respiratory infections and for the detection of particular individual bacterial pathogens, as is further reviewed in this chapter. A recent development is the application of PCR for detecting the family of Haemophilus integrating conjugative elements among antibiotic-resistant Haemophilus influenzae type b directly to cerebrospinal fluid (CSF) to diagnose Haemophilus influenzae type b meningitis and predict the organism's susceptibility, irrespective of culture results. The potential of pyrosequencing after broad-range PCR was successfully investigated by Jordan for the identification of bacterial pathogens responsible for sepsis in neonates.