[Do the new molecular assays-microarray and realtime polymerase chain reaction-for dermatophyte detection keep what they promise?] / Halten die neuen molekularen Teste ­ Microarray und Realtime-Polymerasekettenreaktion ­ zum Dermatophytennachweis das, was sie versprechen?

In this study, a novel real-time polymerase chain reaction (PCR) assay (DermaGenius®2.0, PathoNostics BV, Maastricht, The Netherlands) and a recently developed microarray test (EUROArray Dermatomycosis, Euroimmun, Lübeck, Germany) were evaluated regarding their diagnostic specificity to identify dermatophyte DNA. The tests were compared to conventional methods and sequencing. The microarray Dermatomycosis test allows the detection of 50 dermatophytes and definitive identification of 23 dermatophyte species, 6 yeasts and moulds combined in one test. In comparison, real-time PCR is able to identify 11 dermatophytes and one yeast at the species level. Using the EUROArray, 22 out of 24 dermatophyte species were correctly identified. Using real-time PCR, 9 out of the 11 different dermatophytes included in the test kit were correctly identified. Both molecular tests for detection and differentiation of dermatophytes are useful tools for daily clinical practice. The real-time PCR test does not detect as many species, and specificity is slightly lower. However, real-time PCR is a very fast and easy to perform test, especially since no post-PCR step is necessary. Real-time PCR detects the most frequent dermatophytes like T. rubrum, T. interdigitale, and M. canis without any problems. The EUROArray is more elaborate to perform in the lab, due to the hybridization step. However, the EUROArray shows higher specificity and can detect a much broader range of causative agents, including rare species, in dermatomycology.

PCR-based detection of Aspergillus fumigatus Cyp51A mutations on bronchoalveolar lavage: a multicentre validation of the AsperGenius assay® in 201 patients with haematological disease suspected for invasive aspergillosis.

OBJECTIVES: In patients with invasive aspergillosis (IA), fungal cultures are mostly negative. Consequently, azole resistance often remains undetected. The AsperGenius® multiplex real-time PCR assay identifies clinically relevant Aspergillus species and four resistance-associated mutations (RAMs; TR34/L98H/T289A/Y121F) in the Cyp51A gene. This multicentre study evaluated the diagnostic performance of this assay on bronchoalveolar lavage (BAL) fluid and correlated the presence of RAMs with azole treatment failure and mortality. METHODS: Stored BAL samples from patients with haematological diseases with suspected IA were used. BAL samples that were galactomannan/culture positive were considered positive controls for the presence of Aspergillus. Azole treatment failure and 6 week mortality were compared in patients with and without RAMs that had received ≥5 days of voriconazole monotherapy. RESULTS: Two hundred and one patients each contributed one BAL sample, of which 88 were positive controls and 113 were negative controls. The optimal cycle threshold cut-off value for the Aspergillus species PCR was <38. With this cut-off, the PCR was positive in 74/88 positive controls. The sensitivity, specificity, positive predictive value and negative predictive value were 84%, 80%, 76% and 87%, respectively. 32/74 BAL samples were culture negative. Azole treatment failure was observed in 6/8 patients with a RAM compared with 12/45 patients without RAMs (P = 0.01). Six week mortality was 2.7 times higher in patients with RAMs (50.0% versus 18.6%; P = 0.07). CONCLUSIONS: The AsperGenius® assay had a good diagnostic performance on BAL and differentiated WT from Aspergillus fumigatus with RAMs, including in culture-negative BAL samples. Most importantly, detection of RAMs was associated with azole treatment failure.

Interspecies discrimination of A. fumigatus and siblings A. lentulus and A. felis of the Aspergillus section Fumigati using the AsperGenius® assay.

The AsperGenius® assay detects several Aspergillus species and the A. fumigatus Cyp51A mutations TR34/L98H/T289A/Y121F that are associated with azole resistance. We evaluated its contribution in identifying A. lentulus and A. felis, 2 rare but intrinsically azole-resistant sibling species within the Aspergillus section Fumigati. Identification of these species with conventional culture techniques is difficult and time-consuming. The assay was tested on (i) 2 A. lentulus and A. felis strains obtained from biopsy proven invasive aspergillosis and (ii) control A. fumigatus (n=3), A. lentulus (n=6) and A. felis species complex (n=12) strains. The AsperGenius® resistance PCR did not detect the TR34 target in A. lentulus and A. felis in contrast to A. fumigatus. Melting peaks for L98H and Y121F markers differed and those of the Y121F marker were particularly suitable to discriminate the 3 species. In conclusion, the assay can be used to rapidly discriminate A. fumigatus, A. lentulus and A. felis.

[Use of indicators of fluid responsiveness in septic shock: a survey in public emergency departments]. / Indices de réponse au remplissage vasculaire lors de la prise en charge d'un choc septique : utilisation par les médecins urgentistes.

INTRODUCTION: Fluid therapy is one of the major elements of severe sepsis and septic shock management. A systematic initial fluid bolus is recommended before evaluation of left ventricular filling pressure by the use of indicators of fluid responsiveness, preferentially dynamic ones. A massive fluid therapy could be damaging for the patient. Dynamic indicators of fluid responsiveness are not often relevant in the emergency department. This study was aimed to evaluate the use of indicators of fluid responsiveness by emergency practitioners during septic shock management. STUDY DESIGN: Cross sectional survey using anonymous self-questionnaire. METHODS: We included all practitioners working in public emergency department of Languedoc-Roussillon (France). Primary-end point was the use of one indicator of fluid responsiveness at least. Uni- and multivariate analysis was conducted to find associated factors. RESULTS: Of 232 practitioners included, we collected 149 questionnaires (response rate=64%). Hundred and eight practitioners (72% [64-79%]) used at least one indicator of fluid responsiveness. Fifty-six practitioners (38% [30-46%]) used echocardiography, 54 practitioners (36% [29-44%]) used blood lactate concentration, 45 practitioners (30% [23-38%]) used passive leg raising. The use of indicators of fluid responsiveness is associated with easy access to echography device (odd ratio=2.94 [1.16-7.62], P=0.03). CONCLUSION: Emergency practitioners use preferentially less invasive and less time-consuming indicators of fluid responsiveness.

[Reliability of the pulse contour analysis for cardiac output measurement for assessing the fluid responsiveness]. / La mesure du débit cardiaque par analyse du contour de l'onde de pouls n'est pas fiable pour diagnostiquer une réponse au remplissage vasculaire.

BACKGROUND: The cardiac output (CO) is classically measured in intensive care unit patients. pulse contour (PC) method allows monitoring of CO. OBJECTIVE: The aim of the present study was to assess the ability of PC to assess the fluid responsiveness defined as an increase in CO more than or equal to 15% after 500 mL hydroxyethyl starch over 20 minutes. PATIENTS AND METHODS: In this observational prospective study, patients in shock in whom a PC method was inserted were included. CO was measure using the PC and thermodilution methods before and after a fluid challenge indicated by the physician. The correlation coefficient was measured, the diagram of Bland and Altman was built and the percentage of error (Critchley and Critchley method) was calculated. The ability of PC to diagnose fluid responsiveness was assessed using a receiver operating characteristics (ROC) curve. RESULTS: Sixty-two fluid challenges were performed in 37 included patients. After fluid challenge, r(2) was 0.05 (P<0.01), the bias between PC and thermodilution was 0.3 ± 1.2L/min and the percentage of error was 36%. The area of the ROC curve was 0.601 [0.468-0.723]. CONCLUSION: In ICU patients with shock, PC cannot replace thermodilution to diagnose fluid responsiveness.

Reduction of the severe sepsis or septic shock associated mortality by reinforcement of the recommendations bundle: a multicenter study.

INTRODUCTION: We determined whether the implementation of a bundle of 10 recommendations leads to the reduction of mortality in ICU patients with severe sepsis or septic shock. METHODS: All patients with severe sepsis or septic shock during two consecutive phases: a 6-month quality control period (observational) and secondly a 6-month intervention period based on the implementation of a bundle of 10 recommendations adapted from the Surviving Sepsis Campaign guidelines (initial bacteriological samples and initiating antibiotics, measurement of arterial lactate, volume expansion > or =20 ml/kg, targeted mean arterial pressure > or =65 mmHg and the assessments of central venous pressure and Scv(O2); glucose control, low doses of corticosteroids, a tidal volume < or =8 ml/kg in mechanically ventilated patients with ALI; adequate use of recombinant human activated protein C) were evaluated in 15 ICUs. The primary endpoint was the 28-day mortality rate and the secondary endpoint was the compliance with the recommendations of the care bundle. MEASUREMENT AND RESULTS: Four hundred and forty-five patients (230 and 215 in the observational and intervention periods, respectively) were included. In the two periods, the patients had similar characteristics. The 28-day mortality rate significantly decreased from 40% in the observational period to 27% in the intervention period (P=0.02). According to each recommendation, compliance with the care bundle was achieved in 9 to 100% of patients. CONCLUSION: The implementation of a care bundle adapted from the Surviving Sepsis Campaign guidelines decreases the 28-day mortality rate in patients with severe sepsis and/or septic shock.