Detection of methanogens in peri-appendicular abscesses: Report of four cases.

The aetiology of appendicular abscess is predominantly microbial with aerobic and anaerobic bacteria from gut flora. In this study, by using specific laboratory tools, we co-detected Methanobrevibacter oralis and Methanobrevibacter smithii among a mixture of enterobacteria including Escherichia coli, Enterococcus faecium and Enterococcus avium in four unrelated cases of postoperative appendiceal abscesses. These unprecedented observations raise a question on the role of methanogens in peri-appendicular abscesses, supporting antibiotics as an alternative therapeutic option for appendicitis, including antibiotics active against methanogens such as metronidazole or fusidic acid.

Methanobrevibacter smithii tonsillar phlegmon: a case report.

Untreated tonsillar phlegmon is a life-threatening condition commonly caused by Streptococcus pyogenes and Fusobacterium necrophorum, among other pathogens. Here, using specific laboratory tools, we detected Methanobrevibacter smithii in addition to S. pyogenes. This unprecedented observation questions the role of methanogens in phlegmon and the optimal treatment of this mixed infection.

Endocarditis in the Mediterranean Basin.

Infective endocarditis is a severe disease with high mortality. Despite a global trend towards an increase in staphylococcal aetiologies, in older patients and a decrease in viridans streptococci, we have observed in recent studies great epidemiologic disparities between countries. In order to evaluate these differences among Mediterranean countries, we performed a PubMed search of infective endocarditis case series for each country. Data were available for 13 of the 18 Mediterranean countries. Despite great differences in diagnostic strategies, we could classify countries into three groups. In northern countries, patients are older (>50 years old), have a high rate of prosthetic valves or cardiac electronic implantable devices and the main causative agent is Staphylococcus aureus. In southern countries, patients are younger (<40 years old), rheumatic heart disease remains a major risk factor (45-93%), viridans streptococci are the main pathogens, zoonotic and arthropod-borne agents are frequent and blood culture-negative endocarditis remains highly prevalent. Eastern Mediterranean countries exhibit an intermediate situation: patients are 45 to 60 years old, the incidence of rheumatic heart disease ranges from 8% to 66%, viridans streptococci play a predominant role and zoonotic and arthropod-borne diseases, in particular brucellosis, are identified in up to 12% of cases.

International experts' practice in the antibiotic therapy of infective endocarditis is not following the guidelines.

OBJECTIVE: The management of infective endocarditis (IE) may differ from international guidelines, even in reference centres. This is probably because most recommendations are not based on hard evidence, so the consensus obtained for the guidelines does not represent actual practices. For this reason, we aimed to evaluate this question in the particular field of antibiotic therapy. METHODS: Thirteen international centres specialized in the management of IE were selected, according to their reputation, clinical results, original research publications and quotations. They were asked to detail their actual practice in terms of IE antibiotic treatment in various bacteriological and clinical situations. They were also asked to declare their IE-related in-hospital mortality for the year 2015. RESULTS: The global compliance with guidelines concerning antibiotic therapy was 58%, revealing the differences between theoretical 'consensus', local recommendations and actual practice. Some conflicts of interest were also probably expressed. The adherence to guidelines was 100% when the protocol was simple, and decreased with the seriousness of the situation (Staphylococus spp. 54%-62%) or in blood-culture-negative endocarditis (0%-15%) that requires adaptation to clinical and epidemiological data. CONCLUSION: Worldwide experts in IE management, although the majority of them were involved and co-signed the guidelines, do not follow international consensus guidelines on the particular point of the use of antibiotics.

Antiphospholipid antibodies proposed in the diagnosis of infective endocarditis.

Antiphospholipid antibodies (aPL) may occur alone or associated with other diseases. To evaluate aPL, tested as anticardiolipin antibodies (IgG aCL) in infective endocarditis (IE) diagnosis, we investigated their prevalence in a cohort of 651 patients with IE suspicion. aPL was significantly associated with definite IE versus IE-rejected patients. Their mean levels were significantly higher in patients with definite IE versus possible IE. When applied as Duke minor criterion, they were significantly more often positive, and at higher levels, in patients with definite IE than in patients with possible or rejected IE. aPL could be helpful in difficult cases of IE diagnosis.

Genome analysis of Listeria ivanovii strain G770 that caused a deadly aortic prosthesis infection.

We sequenced the genome of Listeria ivanovii strain G770, which caused a deadly infection of the thoracic aortic prosthesis of a 78-year-old man. The 2.9 Mb genome exhibited 21 specific genes among L. ivanovii strains, including five genes encoding a type I restriction modification system and one glycopeptide resistance gene.

Complementarity between targeted real-time specific PCR and conventional broad-range 16S rDNA PCR in the syndrome-driven diagnosis of infectious diseases.

Molecular tools have shown an added value in the diagnosis of infectious diseases, in particular for those caused by fastidious intracellular microorganisms, or in patients receiving antibiotics before sampling. If 16S rDNA amplification had been gradually implemented in microbiology laboratories, specific real-time polymerase chain reaction (PCR) would have permitted an increase in the sensitivity of molecular methods and a reduction of contamination. Herein, we report our experience in the diagnosis of infectious diseases over two years, during which 32,948 clinical samples from 18,056 patients were received from France and abroad. Among these samples, 81,476 PCRs were performed, of which 1,192 were positive. Molecular techniques detected intracellular microorganisms in 31.3 % of respiratory samples, 27.8 % of endocarditis samples and 51.9 % of adenitis samples. Excluding intracellular bacteria, 25 % of the positive samples in this series were sterile in culture. Conventional broad-range PCR permitted the identification of fastidious and anaerobic microorganisms, but specific real-time PCR showed a significant superiority in the diagnosis of osteoarticular infections, in particular for those caused by Kingella kingae and Staphylococcus aureus, and for endocarditis diagnosis, specifically when Streptococcus gallolyticus and Staphylococcus aureus were involved. The sensitivity of conventional broad-range PCR was 62.9 % concerning overall diagnoses for which both techniques had been performed. These findings should lead microbiologists to focus on targeted specific real-time PCR regarding the clinical syndrome. Finally, syndrome-driven diagnosis, which consists of testing a panel of microorganisms commonly involved for each syndrome, permitted the establishment of 31 incidental diagnoses.

Staphylococcus caprae bone and joint infections: a re-emerging infection?

Staphylococcus caprae has been recently classified as a human pathogen, but the incidence of S. caprae in human bone and joint infections (BJIs) is under-reported. In this study, we report 25 cases of S. caprae BJI, and we review the 31 cases published in the literature. Molecular techniques and matrix-assisted laser desorption ionization time-of-flight mass spectrometry improved the identification of clinically relevant S. caprae strains. In this study, 96% of S. caprae BJIs were localized to the lower limbs, and 88% of the cases involved orthopaedic device infections. S. caprae joint prosthesis infections (JPIs), internal osteosynthesis device infections (I-ODIs) and BJIs without orthopaedic device infections were recorded in 60%, 28% and 12% of cases, respectively. Ten (40%) S. caprae BJIs were polymicrobial infections. These infections were associated with past histories of malignancy (p 0.024). Of the 14 bacterial species related to S. caprae BJI, 57% were staphylococci. I-ODIs were significantly associated with polymicrobial infections (p 0.0068), unlike JPIs, which were monomicrobial infections (p 0.0344). Treatment with rifampicin and fluoroquinolone was recorded in 40% of cases. Surgical treatment was performed in 76% of cases, e.g. prosthesis removal (36%), osteosynthesis device removal (24%), and surgical debridement (16%). Thirty per cent of cases were not treated. Relapses were observed mainly in the patients treated by surgical debridement only (p 0.033). In summary, S. caprae BJI is an underestimated hospital-acquired emerging infection. S. caprae BJI is correlated with infections in orthopaedic devices, which must be removed to control the infection.

Real-time laboratory surveillance of sexually-transmissible infections in Marseille University hospitals reveals rise of gonorrhoea, syphilis and human immunodeficiency virus seroconversions in 2012.

Real-time systematic monitoring of the number of infections diagnosed in our clinical microbiology laboratory in Marseille recently drew attention to the fact that the incidence of gonorrhoea was 10-fold greater from September through December 2012 than during same months of previous years. We also found an increase in the annual incidence of syphilis and human immunodeficiency virus seroconversion. Our system allowed timely identification of an increase in sexually-transmitted infections in Marseille for the whole year of 2012.

Lactobacillus rhamnosus bacteremia: an emerging clinical entity.

Lactobacillus spp. are ubiquitous commensals of the normal human flora that are only occasionally found in clinical infections. Their role in human disease is established for infectious endocarditis but is controversial for other infections. We sought to characterize clinically associated Lactobacillus spp. We conducted a retrospective study, which consisted of the screening of Lactobacillus isolates obtained in our laboratory from January 2004 to December 2009. The polymerase chain reaction (PCR) assay was selected as the gold standard method. The isolates were first identified using API Coryne strips, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), and 16S rRNA gene sequencing. Lactobacillus tuf gene-based identification was used when the 16S rRNA results were inconclusive. Among the 60 strains of Lactobacillus spp. obtained in our laboratory, L. rhamnosus was the most commonly isolated species and was found in blood cultures from 16 patients. Combined with 45 patients reported in the literature, we found that patients presenting with L. rhamnosus bacteremia experienced nosocomial infections associated with both immunosuppression (66 %) and catheters (83 %).

Clinical characteristics and outcome of infective endocarditis in adults with bicuspid aortic valves: a multicentre observational study.

OBJECTIVE: To analyse characteristics and outcomes of infective endocarditis (IE) on bicuspid aortic valves (BAV) and to compare the risk of death according to the presence or absence of BAV. DESIGN: 5-year observational study. SETTING: Population of 856 patients with definite IE according to the Duke criteria from two tertiary centres (Amiens and Marseille, France). PATIENTS: 310 consecutive patients with definite native aortic valve IE enrolled between 1991 and 2007. INTERVENTIONS: Patients underwent transthoracic and transoesophageal echocardiography during hospitalisation. Surgery was performed on a case-by-case basis according to conventional guidelines. MAIN OUTCOME MEASURES: In-hospital mortality and 5-year overall mortality. RESULTS: Patients with BAV IE (n=50, 16%) were younger, had fewer comorbidities and a higher frequency of aortic perivalvular abscess (50%). Presence of BAV (OR 3.79 (1.97-7.28); p<0.001) was independently predictive of abscess formation. Early surgery was performed in 36 BAV patients (72%) with a peri-operative mortality of 8.3%, comparable to that of patients with tricuspid aortic valve IE (p=0.89). BAV was not independently predictive of in-hospital mortality (OR 0.89 (0.28-2.85); p=0.84) or 5-year survival (HR 0.71 (0.37-1.36); p=0.30). Age, comorbidities, heart failure, Staphylococcus aureus and uncontrolled infection were associated with increased 5-year mortality in BAV patients. CONCLUSION: BAV is frequent in adults with native aortic valve IE. Patients with BAV IE incur high risk of abscess formation and require early surgery in almost three-quarters of cases. IE is a severe complication in the setting of BAV and warrants prompt diagnosis and treatment.

Evaluation of the LightCycler SeptiFast test in the rapid etiologic diagnostic of infectious endocarditis.

The SeptiFast test (Roche Diagnostics) is a new commercial molecular technique that has emerged for the detection of bacteria in blood. We compared in this study the sensitivity of blood culture to a commercially available broad-range real-time polymerase chain reaction (PCR) assay for the detection in blood of 19 bacterial species and six fungal species (SeptiFast test, Roche Diagnostics) in 63 patients with infectious endocarditis (IE). The SeptiFast test is not more sensitive for organisms such as Streptococci, Enterococci, and Staphylococcus aureus (11/29 versus 12/29 for blood culture). It has detected less commonly coagulase-negative Staphylococci (0/15 versus 3/15, P = 0.2) and significantly fewer other microorganisms (0/6 versus 4/6, P = 0.03). However, bacteria were detected from three IE treated by antibiotics, with blood culture negative on admission. The SeptiFast test may be useful in cases of IE in patients treated with antibiotics before admission.

[Microbiological diagnosis: Etiologic driven and syndrome driven]. / Diagnostic microbiologique : du diagnostic par étiologie au diagnostic par syndrome.

OBJECTIVES: Over the past decade, the introduction of techniques of molecular biology and the automation have undergone important changes in the practice of clinical microbiological laboratories. METHODS: Commonly, physician prescriptions are etiology driven for emerging infectious agent. The improving in test standardization and rapidity and the best communication between physician and clinical microbiologist lead to profound changes in microbiology laboratories. RESULTS AND CONCLUSION: The present review summarizes the recent development in multiplexed microbiological technology adapted to syndrome driven diagnosis, including sampling strategy, DNA microarray and antigenic microarray.

Comparison of the new InoDiag automated fluorescence multiplexed antigen microarray to the reference technique in the serodiagnosis of atypical bacterial pneumonia.

The aetiological diagnosis of pneumonia depends largely on culture-, antigen- or PCR-based tests. Atypical agents of pneumonia include Coxiella burnetii, Chlamydophila pneumoniae, Chlamydia psittaci, Legionella pneumophila, Francisella tularensis and Mycoplasma pneumoniae. In these cases, serological tests are commonly used for diagnosis. All of the above species were comparatively screened for by using the InoDiag multiplexed automatic immunofluorescence assay and established reference techniques. The InoDiag assay required 5 microL of serum, took 76 min per serum sample, and required an incubator, a fluorescence reader and interpretation software. In total, 248 single sera from patients were tested, for the diagnosis of pneumonia, and the results obtained with selected serum samples were compared with results obtained with the reference method. It was shown that, for the detection of Coxiella burnetii IgM, the automated assay had a sensitivity and specificity of 100%. For the detection of M. pneumoniae IgM, sensitivity was 100% and specificity was 98%. For the detection of Chlamydophila pneumoniae and Chlamydia psittaci IgG, sensitivity was 81% and specificity was 94%. For the detection of L. pneumoniae IgG, sensitivity was 63% and specificity was 98%. For the detection of F. tularensis IgG and IgM, sensitivity was 100% for both, and specificity was 95% and 100%, respectively. The performance of this serological assay was comparable to that of other assays reported in the literature. This preliminary study shows that the automatic InoDiag assay opens the way to immunofluorescence assay standardization.

Multiplexed whole bacterial antigen microarray, a new format for the automation of serodiagnosis: the culture-negative endocarditis paradigm.

The serological diagnosis of blood culture-negative endocarditis due to Coxiella burnetii, Bartonella spp., Brucella melitensis and Legionella pneumophila is based on a manual immunofluorescence assay (IFA), which is taken to be the reference method. The automated IFA InoDiag multiplexed antigenic microarray, which includes a slide with all the above bacteria and four internal controls, an incubator, a fluorescent reader and software with an algorithm of interpretation for infectious endocarditis (IE) was evaluated. A single serum dilution at 1/128 was used. Eleven patients with Bartonella spp. IE and ten with C. burnetii IE, diagnosed using the modified Duke criteria, as well as one patient with B. melitensis infection and three patients with L. pneumophila IE were tested. In total, 236 sera were used as negative controls, with the reference method. The results of IgG detection were: C. burnetii phase I, 'sensitivity (Se) = 88% and specificity (Sp) = 99%', and C. burnetii phase II, Se = 88% and Sp = 99%; for Bartonella henselae, Se = 100% and Sp = 100%; for Bartonella quintana, Se = 78% and Sp = 96%; for B. melitensis, Se = 100% and Sp = 99%; and for L. pneumophila, Se = 100% and Sp = 99%. With the algorithm interpretation, the negative and positive predictive values of the test 'were 100% for the diagnosis of IE caused by the four bacteria tested. These results were confirmed by two other assays, one using triplicate testing and one blind testing performed by another centre. This multiplexed test is therefore a valuable tool for the rapid diagnosis of blood-culture negative IE.

[Staphylococcus aureus isolated from a case of lymph node botryomycosis]. / Une botryomycose ganglionnaire àStaphylococcus aureus.

Botryomycosis is an uncommon chronic bacterial infection, which usually involves the skin and the subcutaneous tissues. Visceral involvement often occurs in immunodepressed patients. We describe the case of a 4-year-old boy who presented a right inguinal inflammatory mass associated with pruritic papules without fever. Blood analyses showed a substantial inflammatory syndrome and blood hypereosinophilia. Histological examination of the right inguinal lymph node showed granulomatous adenitis with a Splendore-Hoeppli phenomenon surrounding Gram-positive bacteria, corresponding to Staphylococcus aureus after biopsy culture. The diagnosis of S. aureus lymph node botryomycosis was made and the child was successfully treated with a double dose of oxacillin over 3 weeks.