Comparative evaluation of Aspergillus terreus DNA and galactomannan in serum and bronchoalveolar lavage of intravenously infected mice

Abstract number: 1733_1472

Ahmad S., Theyyathel A., Khan Z.

Objective: Invasive aspergillosis is a growing problem in immunocompromised individuals. Although Aspergillus fumigatus is the principal aetiologic agent of the disease, infections caused by Aspergillus terreus have shown an increased occurrence in recent years. Additionally, A. terreus is more resistant to amphotericin B and is probably also associated with higher mortality in humans. Therefore, an early diagnosis is imperative for reduced mortality and better prognosis. The aim of this study was to develop sensitive and specific diagnosis of invasive aspergillosis by detecting A. terreus-specific DNA by nested PCR (nPCR) in serum and bronchoalveolar lavage (BAL) specimens of intravenously infected mice and to compare the results with galactomannan (GM) levels.

Methods: Sixty BALB/c mice, immunosuppressed with four intraperitoneal injections of cyclophosphamide (200 mg/kg) on day -1, 0, +1, +3 were infected intravenously with 1×10⁶ conidia of A. terreus. The mice were sacrificed on day 1, 3, 5, 7 and 9 post-infection in groups of twelve each and their BAL, blood, and lungs were cultured. The A. terreus-specific DNA and GM in serum and BAL were detected by nPCR and Platelia Aspergillus (BioRad, Marnes-la-Coquette, France), respectively.

Results: The nPCR developed with primers derived from internally transcribed spacer (ITS)-1 and ITS-2 regions of rDNA was specific for A. terreus as genomic DNA from other Aspergillus species or other fungi was not amplified. The lung homogenates of all the infected mice yielded A. terreus in culture, while the blood and BAL specimens were uniformly negative. The % positivity of serum samples for GM and nPCR were 78% and 73%, respectively. In contrast, among BAL samples, 71% were positive for GM and 81% for A. terreus-specific DNA. The combined detection of A. terreus DNA and/or GM enhanced the positivity to 95% in serum and 98% in BAL.

Conclusions: The data suggest that GM and A. terreus DNA are easily detected in both, serum and BAL, during the entire course of infection and their combined detection could be useful in the early diagnosis of invasive aspergillosis.

Supported by Kuwait University Research Grant MI04/02