Prolyl endopeptidase activity in bronchoalveolar lavage fluid: a novel diagnostic biomarker in a guinea pig model of invasive pulmonary aspergillosis. (August 2013)

• Record Type: Journal Article
• Authors: Jambunathan, Kalyani; Watson, Douglas S.; Najvar, Laura K.; Wiederhold, Nathan P.; Kirkpatrick, William R.; Patterson, Thomas F.; Askew, David S.; Kodukula, Krishna; Galande, Amit K.
• Is Part Of: Medical mycology. Volume 51:Number 6(2013)
• Page Start: 592
• Page End: 602
• Publication Date: 2013-08
• Subjects: Medical mycology -- Periodicals; Veterinary mycology -- Periodicals; Mycology -- Periodicals; Mycoses -- Periodicals; Pathogenic fungi -- Periodicals; 616.969005
• DOI: 10.3109/13693786.2012.761360
• Languages: English
• ISSNs: 1369-3786

Abstract

<abstract> <title>Abstract</title> <p>Improved diagnostics are needed to detect invasive pulmonary aspergillosis, a life-threatening infection caused by the pathogenic fungus <italic>Aspergillus fumigatus</italic>. We are investigating secreted fungal proteases as novel biomarkers for the diagnosis of this disease. Although the <italic>A. fumigatus</italic> genome encodes a multitude of secreted proteases, few have been experimentally characterized. Here, we employed an unbiased combinatorial library of internally quenched fluorogenic probes to detect infection-associated proteolysis in the lungs of guinea pigs experimentally infected with <italic>A. fumigatus</italic>. Comparative protease activity profiling revealed a prolyl endopeptidase activity that is reproducibly induced during infection but is not observed in healthy animals. This proteolytic activity was found in four independent animal experiments involving two <italic>A. fumigatus</italic> isolates. We synthesized a small, focused fluorogenic probe library to define the substrate specificity of the prolyl endopeptidase substrate motif and to identify optimal Probe sequences. These efforts resulted in the identification of a panel of six individual substrate-based fluorescent probes capable of detecting infection in guinea pigs with high statistical significance (<italic>P</italic><0.005 in most cases). Receiver operating characteristic analyses demonstrated that this fluorogenic assay could detect <italic>A. fumigatus</italic> infection-associated proteolysis with comparable sensitivity and specificity as existing diagnostic procedures, suggesting that further optimization of the methodology may lead to improved diagnostics options for invasive pulmonary aspergillosis.</p> </abstract>