Coinfection with Coxiella burnetii in infectious endocarditis.

Currently, most cases of Q fever endocarditis are diagnosed serologically, when levels of anti-phase I antibodies are ‡1:800 [1]. In Marseille, France, we proposed a systematic serological testing for all pathogens that could cause negative blood culture IE for all patients suspected to have IE. We demonstrated that this approach is useful in establishing the aetiological diagnosis of IE [2]. Thanks to this systematic serological testing, we could diagnose four patients with IE due to a typical microorganism and high antibody titres to C. burnetii. In a retrospective study, we enrolled patients, hospitalised in Marseille, between April 1994 and January 2007, with C. burnetti phase I IgG ‡1:800 and IE caused by another microorganism. The modified Duke criteria were used to define cases of endocarditis [3]. Each patient underwent testing that was guided by a diagnostic kit as described previously [2]. Titres of IgG, IgM and IgA antibodies in serum samples were determined by the indirect immunofluorescence assay [2]. Valvular surgical samples or heparinised blood samples were inoculated onto human embryonic lung fibroblasts grown on a coverslip in a shell vial. After a 7-day incubation period at 37 C, cultivated bacteria were detected by a direct immunofluorescence assay incorporating rabbit monoclonal antibodies to C. burnetii. A DNA extract was prepared from the valvular surgical samples with the QIAmp tissue kit (Qiagen, Hilden, Germany) and the QIAamp Blood Kit (Qiagen) for blood samples according to the manufacturer’s instructions. Bacterial DNA was detected by PCR amplification with universal primers and sequencing of the amplified fragment, as previously described [4]. For C. burnetii, the PCR was performed using a LightCycler apparatus (Roche), using two different target genes: the IS1111 repeated intervening sequence and the IS30a intervening sequence. Valvular surgical samples, when available, were embedded in paraffin, and sections were stained with hematoxylin-eosin-safran for histology. For detection of C. burnetii in tissues, immunochemistry was done on deparaffinised sections by use of an anti-C. burnetii rabbit polyclonal antibody, as previously described [5]. Four patients (6.5& of patients with IE tested) had IE according to Duke’s criteria due to common endocarditis pathogens (Streptococcus gallolyticus for patients 1 and 2, Streptococcus mitis for patient 3, and Enterococcus faecalis for patient 4) and anti-phase I C. burnetii antibodies ‡1:800. Characteristics of the patients and microbiological results are presented in Table 1. Only patient 1 had a past history of previous fever and asthenia suggestive of Q fever. All these patients underwent valvular surgery in the course of the disease. The valvular sample of patient 1 was positive for C. burnetii for the two targeted genes. Patient 3 had Q fever serology very suggestive of chronic infection with high titres of IgG1 1:102 400 and IgA1 1:12 800. Patients 2 and 4 had serology IgG1 of 1:800. For these two patients there was no other evidence of chronic Q fever. All the patients were treated for bacterial IE according to local guidelines and for Q fever endocarditis. All the patients improved and remain healthy at present. We report here four patients with IE due to typical microorganisms and Q fever serology with IgG1 ‡800. We could recover C. burnetii DNA in valvular samples in patient 1. Patient 3 had very high titres (1:102 400). We concluded that these two patients surely had an ongoing chronic Q fever. The two other patients with IgG1 1:800 had no evidence of ongoing illness. Patient 1 and patient 3 were included in a prospective study evaluating the contribution of systematic serological testing in the diagnosis of IE [2]. As witnessed in these four cases, the diagnosis of Q fever may Corresponding author and reprint requests: Prof. Didier Raoult, Unite des Rickettsies, CNRS UMR6020, IFR48, Faculte de Medecine, 27, Boulevard Jean Moulin, 13385 Marseille, France E-mail: didier.raoult@univmed.fr