Etiological diagnostic of blood culture negative endocarditis

Abstract

Streptococci in patients who received previous antibiotic treatment and infection of the right heart and specifically on pace makers. The diagnostic strategy of BCNIE starts by serological testing of fastidious agents2. At the minimum Coxiella burnetii and Bartonella should be tested. A single serum is sufficient, as endocarditis is a chronic disease associated with high level IgG antibodies. Q fever endocarditis is associated with antibody titers to C. burnetii phase I above 800 and Bartonella endocarditis to high levels ( 800) of IgG2. Patients with Q fever may have cross-reacting antibodies to Bartonella. Other organisms may be tested depending on the epidemiological situation including Brucella, Legionella pneumophila and Aspergillus2. Antibodies to Chlamydia usually result from cross-reactions with Bartonella, and there are few evidences that it can cause IE7. Detection by PCR of organisms in the blood is a promising technique8. Real time PCR has been successfully developed for the diagnosis of Bartonella IE9 and Q fever10. A preliminary study using broad spectrum primers was also performed11 and a PCR amplification of blood for the diagnosis of Whipple’s diseases has also been proposed12. However currently these techniques frequently lack specificity. PCR when performed on the valve has been recently reported in several studies. The use of universal primers amplifying any bacteria (based on 16S rRNA) or fungi (based on 18S rRNA) has been reported13. The detected microorganisms are mainly Streptococcus in patients with previous antibiotics treatment, and fastidious bacteria including Granulicatella, Abiotrophia, Bartonella, Coxiella burnetii and Tropheryma whipplei. DNA from the causative agent can persist months to years after clinical cure and the link between the current IE episode and the amplified DNA need to be carefully checked14. Moreover PCR can easily be contaminated and controls are necessary. Personally I recommend not using positive controls that may be confused with the causative agents, as carry over contamination is common. DNA from a microorganism very unlikely causing IE may be a good positive control. Negative controls are critical. They should be placed any 3 to 7 tested samples and be negative. Any positive amplicon should be sequenced to identify the causative agent. Some sequences usually result from contamination and are easily recognized such as DNA commonly found in the water (Pseudomonas) or the reagents (E. coli). A similar sequence found in the same round of PCR in a sample from another patient may result from contamination. In the other hand a sequence found for the first time in a laboratory reflect usually a true positive result. When the results have a low predictive value, amplification of a second gene is critical to confirm the etiological diagnosis. Infective endocarditis (IE) remained a diagnostic challenge. Etiological diagnosis is critical to select an appropriate treatment, as the fatality rate remains high1. The proportion of IE without etiological diagnosis varies from country to country and from different centers in the same country. These variations reflect the used diagnostic criteria, the early use of antibiotics in patients without documentation and the sampling strategy for etiological diagnosis2. Moreover the role of epidemiology which as been underestimated appears critical as in a recent series from Algeria zoonotic agents may caused as many as 25% of IE3. The diagnostic of IE is currently based on the commonly modified Duke Criteria4. Using this strategy requires obtaining high quality echocardiography and blood cultures sampled before antibiotic administration. It also requires testing serologically at the minimum Coxiella burnetii and Bartonella species. Duke criteria may not be perfectly efficient for blood culture negative IE, as they may be chronic, afebrile and, eventually such as for Q fever, may lack valvular vegetation. Moreover underlying lesions, and specifically aortic bicuspidy, may not be detected by echocardiography. In my own experience in 2 cases with evidence of chronic Q fever, two successive transthoracic echocardiography failed to detect aortic bicuspidy. In such patients without fever, vegetation and obvious underlying valve lesion IE diagnosis remains a real challenge. Culture of blood is critical for the diagnosis of IE It is clear that 3 samples containing 40 ml of blood, obtained in a 4 hours time is enough to detect usual organisms2. This allows starting empiric treatment 4 hours after the first sampling when IE is considered. Extensive blood culture and subculturing is not necessary5, and HACEK group organism are recovered in the regular 5 days incubation. Specialized culture methods are useful when regular blood cultures fail to recover the etiological agents. We have successfully used the shell vial cell culture assay for this purpose sampling blood or valves of patients with blood culture negative IE6. This allows recovering C. burnetii, Bartonella, Tropheryma whipplei and Brucella. However this technique is restricted to specialized laboratories. Blood culture negative IE (BCNIE) is defined as an endocarditis without etiology after 3 blood samples inoculated on standard medium. The causative agents of BCNIE are fastidious organisms (zoonotic agents and fungi)