Development of monoclonal antibodies to recombinant terrelysin and characterization of expression in Aspergillus terreus.
Nayak-AP; Green-BJ; Friend-S; Beezhold-DH
J Med Microbiol 2012 Apr; 61(4):489-499
Aspergillus terreus is an emerging pathogen that mostly affects immunocompromised patients with infections that are often difficult to manage therapeutically. Current diagnostic strategies are limited to the detection of fungal growth using radiological methods or biopsy that often does not enable species-specific identification. As a result, there is a critical need for diagnostic techniques to enable early and specific identification of the causative agent. In this study, we describe monoclonal antibodies (mAbs) developed to a previously described recombinant terrelysin. Sixteen hybridomas of various IgG isotypes were generated to the recombinant protein, of which seven demonstrated reactivity to the native protein in hyphal extracts. Cross-reactivity analysis using hyphal extracts from 29 fungal species, including 12 Aspergillus species and 5 different strains of A. terreus showed that 3 mAbs (13G10, 15B5 and 10G4) were A. terreus-specific. Epitope analysis demonstrated mAbs 13G10 and 10G4 recognize the same epitope 'PSNEFE', while mAb 15B5 recognized the epitope 'LYEGQFHS'. Time course studies showed that terrelysin expression was highest during early hyphal growth and dramatically reduced after mycelial expansion. Immunolocalization studies demonstrated that terrelysin is localized not only within the cytoplasm of hyphae but appeared to be more abundant at the hyphal tip. These findings were confirmed in cultures grown at room temperature as well as at 37 degrees C. Additionally, terrelysin was detected in the supernatant of A. terreus cultures. These observations suggest terrelysin may be a candidate biomarker for A. terreus infection.
Pathogens; Pathology; Infectious-diseases; Diagnostic-tests; Fungal-diseases; Fungal-infections; Fungi; Immune-system; Immune-system-disorders; Immunologic-disorders; Immunology; Diagnostic-techniques;
Author Keywords: Aspergillus terreus; terrelysin; monoclonal antibodies
Ajay P Nayak, Allergy and Clinical Immunology Branch, Health Effects Laboratory Division, National Institute for Occupational Safety and Health, Centers for Disease Control and Prevention, Morgantown, WV 26505
Healthcare and Social Assistance; Services
Journal of Medical Microbiology