Impact of a Molecular Approach to Improve the Microbiological Diagnosis of Infective Heart Valve Endocarditis

Abstract

Background-Even today, infective endocarditis (IE) remains a severe and potentially fatal disease demanding sophisticated diagnostic strategies for detection of the causative microorganisms. Despite the use of appropriate laboratory techniques, classic microbiological diagnostics are characterized by a high rate of negative results. Methods and Results-Broad-range polymerase chain reaction (PCR) targeting bacterial and fungal rDNA followed by direct sequencing was applied to excised heart valves (nϭ52) collected from 51 patients with suspected infectious endocarditis and from 16 patients without any signs of IE during an 18-month period. The sensitivity, specificity, and the positive and negative predictive values for the bacterial broad-range PCR were 41.2%, 100.0%, 100.0%, and 34.8%, respectively, compared with 7.8%, 93.7%, 80.0%, and 24.2% for culture and 11.8%, 100.0%, 100.0%, and 26.2% for Gram staining. Without exception, database analyses allowed identification up to the (sub)species level comprising streptococcal (nϭ13), staphylococcal (nϭ4), enterococcal (nϭ2), and other signature sequences such as Bartonella quintana and Nocardia paucivorans. Fungal ribosomal sequences were not amplified. All valve tissues of the reference group were negative for both PCR and conventional methods, except one sample that was contaminated by molds. Conclusions-Culture-independent molecular methods substantially improve the diagnostic outcome of microbiological examination of excised heart valves. Importantly, this was true not only for fastidious, slow-growing, and/or nonculturable microorganisms but also for easy-to-culture pathogens such as streptococci and staphylococci. Both patient management and empiric antibiotic therapy of IE are likely to benefit from improved knowledge of the spectrum of pathogens now causing IE.

Laboratory Diagnosis of Infective Endocarditis

“…Although broad-range bacterial PCR has been applied to blood sources, sensitivity is superior when performed on excised valve tissue, with an organism detected in 66% (150/227) of endocarditis cases in one study compared to just 14% (35/257) of cases when performed on EDTA blood (7). Broad-range bacterial PCR performed on valve tissue has a reported sensitivity of 33% to 90%, while sensitivity of valve culture in the same studies was 8% to 33% (7,(27)(28)(29)(30)(31). Differing patient populations and assay designs likely account for variations in sensitivities between studies.…”

“…In cases of blood culture-negative endocarditis, an organism was identified by broad-range bacterial PCR of valve tissue in 60% to 100% of cases across five studies (28-30, 32, 33). Broad-range bacterial PCR assays have dem-onstrated high specificity (77% to 100%), with detection of contaminating organisms being rare (27,29). While sensitivity may not be ideal in patients with infective endocarditis who undergo surgical excision of the affected valve and for whom no etiologic agent has yet been identified, we recommend testing valvular tissue by broad-range bacterial PCR when histopathologic examination of excised tissue shows acute inflammation.…”

LightCycler SeptiFast assay as a tool for the rapid diagnosis of sepsis in patients during antimicrobial therapy

“…Routine aetiological diagnosis of sepsis relies on standard culture methods that take at least 24 h or more to give the initial information. Therefore, quick identification of pathogens causing sepsis within the clinically relevant time frame should be based on state-of-the-art molecular methods that target bacterial and fungal DNA (Abdul-Redha et al, 2007;Breitkopf et al, 2005;Dombrovskiy et al, 2007). In this letter, we describe the clinical utility of the first standardized, CE-certified, multiplex real-time PCR assay for the molecular diagnosis of sepsis that has been approved for in vitro diagnostic use (LightCycler SeptiFast assay; Roche Diagnostics).…”

Usefulness of 16S rDNA sequencing for the diagnosis of infective endocarditis caused by Corynebacterium diphtheriae

“…These methods often fail because of previous antimicrobial therapy or the involvement of fastidious, slow-growing or non-cultivable micro-organisms. To overcome this, culture independent molecular techniques based on the amplification and direct sequencing of ribosomal sequences have been developed (Breitkopf et al, 2005).…”