Laboratory Diagnosis of 37 Cases of <i>Bartonella</i> Endocarditis Based on Enzyme Immunoassay and Real-Time PCR

Abstract

<i>Bartonella</i> spp., mostly <i>Bartonella quintana</i> and <i>B. henselae</i>, are a common cause of culture-negative endocarditis. Serology using immunofluorescence assay (IFA) and PCR performed on cardiac tissues are the mainstays of diagnosis. We developed an enzyme immunoassay (EIA) and a novel multiplex real-time PCR assay, utilizing <i>Bartonella</i> genus-specific, <i>B. henselae</i>-specific, and <i>B. quintana</i>-specific SimpleProbe probes, for diagnosis of <i>Bartonella</i> endocarditis. We aimed to evaluate the performance of these assays. Thirty-seven patients with definite endocarditis, 18 with <i>B. henselae</i>, 18 with <i>B. quintana</i>, and 1 with <i>B. koehlerae</i>, were studied. Diagnosis was confirmed by conventional PCR and DNA sequencing of surgical cardiac specimens. Similar to the case with IFA, anti-<i>Bartonella</i> IgG titers of ≥1:800 were found in 94% of patients by EIA; cross-reactivity between <i>B. henselae</i> and <i>B. quintana</i> precluded species-specific serodiagnosis, and frequent (41%) but low-titer cross-reactivity between <i>Coxiella burnetii</i> antibodies and <i>B. henselae</i> antigen was found in patients with Q fever endocarditis. Low-titer (1:100) cross-reactivity was uncommonly found also in patients with brucellosis and culture-positive endocarditis, particularly <i>Enterococcus faecalis</i> endocarditis. Real-time PCR performed on explanted heart valves/vegetations was in complete agreement with results of sequence-based diagnosis with characteristic melting curves. The genus-specific probe identified five additional endocarditis-associated <i>Bartonella</i> spp. at the genus level. In conclusion, EIA coupled with a novel real-time PCR assay can play an important role in <i>Bartonella</i> endocarditis diagnosis and expand the diagnostic arsenal at the disposal of the clinical microbiologist. Since serology remains a major diagnostic tool, recognizing its pitfalls is essential to avoid incorrect diagnosis.