Development and Validation of Fungal Extraction and Real -Time PCR Assay for the Diagnosis of Medically Important Fungal Infections in Blood Samples.

Brief Summary

Detailed Description

Invasive fungal infections are a major cause of morbidity and mortality in immunocompromised patients. Among them, patients with hematological malignancies, neutropenic patients, human immunodeficiency virus infected patients, diabetics, solid organ transplanted patients and patients admitted in an intensive care unit are particularly at high risk. The survival of these patients depends on early diagnosis and prompt appropriate antifungal treatment. The early diagnosis of these infections is difficult because of the lack of sensitive test methods, notably blood cultures. Its sensitivity is poor or close-zero for aspergillosis. In addition, the response time is several days. For these reasons, we decided to develop a real-time PCR (Polymerase Chain Reaction) assay on blood samples. It should allow rapid response to establish a positive or negative diagnosis of invasive fungal infection, it could contribute strongly to the decision of treating using antifungals, and it should monitor the effectiveness and the optimization of antifungal prescriptions. Methods: Our project is a multicentre prospective inter-regional collaborative work between Nice, Rennes and Toulouse Mycological and parasitological laboratories. Our objectives are: First, to validate an extraction method from blood infected by fungi species. The three laboratories will work together to determine the best extraction method, since there is no consensus method for the extraction of nucleic acids of fungal origin in the context of human infections. The numerous extraction techniques already used lead to differences in the PCR results. As a consequence, inter-laboratory comparisons are not easy. Secondly, we aim to develop three real-time PCR assays: A panfungal real-time PCR assay able to detect most fungal species responsible for human diseases; a real-time PCR assay targetting Candida albicans and one targeting Aspergillus fumigatus which are two clinically important pathogens. Then patient blood samples (classified according to EORTC consensus) will be collected during the study in order to evaluate and validate our method on clinical samples. Results will allow us to determine the sensitivity, specificity and reproducibility, negative and predictive values. Objectives Overall, our work aims to evaluate the clinical impact of real-time PCR in the early diagnosis of invasive fungal infections and on the initiation or stopping of antifungal therapy. The economic impact resulting from the use of this method will be evaluated. Indeed, an excellent predictive negative value of a panfungal real-time PCR assay could warrant a decrease in the use of empirical antifungal therapy

Primary Outcomes

Secondary Outcomes

• IN Nice, Rennes and Toulouse Teaching hospitals, patients suspected of invasive fungal infection will be included in order to classified in three categories: proven, probable and possible fungal infection, according to EORTC consensus.(1 year)