Differential Diagnosis of Fungal Pneumonias vs. Tuberculosis in AIDS Patients by Using Two New Molecular Methods

Abstract

Opportunistic fungal pneumonias (OFP) are the main cause of death in AIDS patients worldwide. Diagnosis of these infections is often late as tuberculosis (TB) is frequently the first suspicion. In addition, diagnostic tools have limitations and are unavailable in disadvantaged regions. To perform the differential diagnosis of the main fungi causing OFP in AIDS patients (Histoplasma capsulatum, Cryptococcus neoformans/C. gattii and Pneumocystis jirovecii) vs. the Mycobacterium tuberculosis complex (MTBC), two new assays were developed: (i) a multiplex real-time PCR (MRT-PCR) and (ii) a simple and cost-effective method based on real-time PCR and the analysis of melting curves after amplification (MC-PCR). Both of the techniques were optimized and standardized “in vitro”, showing a suitable reproducibility (CV ranged between 1.84 and 3.81% and 1.41 and 4.83%, respectively), a 100% specificity and detection limits between 20 and 2 fg of genomic DNA per 20 µL of reaction. A validation study was performed by retrospectively using 42 clinical samples from 37 patients with proven fungal infection or TB, and 33 controls. The overall sensitivity for the MRT-PCR assay and the MC-PCR assay was 88% and 90.4%, respectively. Both techniques were fast, sensitive and reproducible, allowing for the detection of these pathogens and the performance of a differential diagnosis.

Molecular Diagnosis of Endemic Mycoses

“…In non-endemic countries, where the availability of some other useful techniques, as antigen detection, is limited, qPCR-based molecular assays have been developed to this purpose, extending their usefulness to difficult-to-diagnose forms of infection [ 34 , 37 ]. The introduction of multiplex formats also allows for performing a differential diagnosis with other pathogens causing similar clinical patterns reducing costs [ 118 ]. In endemic areas, especially in resource-limited settings, cost-effective molecular methods such as LAMP could be a promising alternative.…”

Identification of the shared hub gene signatures and molecular mechanisms between HIV-1 and pulmonary arterial hypertension

“…However, there is still a significant disparity between those infected with HIV-1 and the general population in terms of life expectancy and quality of life, which can largely be attributed to complications caused by HIV-1 infection 4 . As survival time of HIV-1 infections, the incidence of chronic complications associated with HIV-1 has significantly increased 5 , 6 , particularly chronic lung disease 7 and cardiovascular disease 8 .…”