Comparison of real-time florescence quantitative PCR measurements of VAD1 mRNA with three conventional methods in diagnosis and follow-up treatment of Cryptococcus neoformans infection

摘要

This study was to develop a real-time florescence quantitative PCR (RT-FQ-PCR) assay to measure virulence-associated DEAD-box RNA helicase (VAD1) mRNA from Cryptococcus neoformans and evaluate its potential use in diagnosis and follow-up treatment of C. neoformans meningitis (CNM). Cryptococcus neoformans was detected using RT-FQ-PCR, ink staining, fungal culturing and C. neoformans antigen detection in CNM compared with a normal control. VAD1 mRNA was measured in both acute and stable CNM patients. The sensitivity of RT-FQ-PCR (96%) is higher than ink staining (72%) and culture culturing (64%) (P<0.05, P<0.05 respectively), but its sensitivity is the same as antigen detection (96%, P>0.05). The levels of VAD1 mRNA in the acute and stable phase of a C. neoformans infection are 3.042±0.906 and 2.187±0.665 respectively (P<0.01). The levels of VAD1 mRNA are correlated to the numbers of C. neoformans, intracranial pressure and glucose concentration in cerebrospinal fluid (CSF; P<0.01, P<0.01 and P<0.05 respectively). The levels of expression of VAD1 mRNA in the group of patients who received an AmB/5-FC/FZC drug regimen decreased more than in patients taking a 5-FC/AmB or 5-FC/FCZ drug combination. Quantitative measurements of VAD1 mRNA are valuable and reliable in diagnosing C. neoformans infection and evaluating a therapy response.