Rinosh Mani1, Raquel Vilela2,3, Niesa Kettler1, Martin I Chilvers4, Leonel Mendoza3,5. 1. Veterinary Diagnostic Laboratory, College of Veterinary Medicine, Michigan State University, Michigan, USA. 2. Faculty of Pharmacy, Federal University of Minas Gerais, Minas Gerais, Brazil. 3. Biomedical Laboratory Diagnostics, Michigan State University, Michigan, USA. 4. Department of Plant, Soil and Microbial Sciences, Program in Ecology, Evolutionary Biology and Behavior, Michigan State University, Michigan, USA. 5. Microbiology and Molecular Genetics, Michigan State University, Michigan, USA.
Abstract
PURPOSE: Pythiosis is an infection of humans and other animals caused by the fungal-like pathogen Pythium insidiosum. This pathogen causes life-threatening infection in the infected hosts. Culture, histopathology, serology and molecular tools are used to diagnose its infections. Successful management of pythiosis is directly linked to an early diagnosis. Thus, a rapid identification of putative cultures developing submerged sparsely septate hyphae is of extreme importance. However, few laboratories are familiar with the culture identification of this unique pathogen and its differential diagnosis with similar filamentous fungi. METHODOLOGY: We have evaluated the use of matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF) on 53 isolates of P. insidiosum collected from cases of human and animal pythiosis in the USA and around the world. To assess the specificity of the approach, 18 pathogenic and saprotrophic filamentous fungal and fungal-like microbes were also tested. RESULTS: MALDI-TOF in-house spectra correctly identified the 53 P. insidiosum isolates (score range 1.93-2.51). MALDI-TOF based identification within P. insidiosum isolates showed protein spectra variation between geographical diverse isolates. A mass spectrometry approach was able to discriminate P. insidiosum from the 18 filamentous fungal and fungal-like microbes in this study, including four Pythium spp. and Phytopythium litorale plant pathogenic species. CONCLUSION: The data showed MALDI-TOF could be used for the accurate and rapid culture identification of P. insidiosum in the clinical laboratory.
PURPOSE: Pythiosis is an infection of humans and other animals caused by the fungal-like pathogen Pythium insidiosum. This pathogen causes life-threatening infection in the infected hosts. Culture, histopathology, serology and molecular tools are used to diagnose its infections. Successful management of pythiosis is directly linked to an early diagnosis. Thus, a rapid identification of putative cultures developing submerged sparsely septate hyphae is of extreme importance. However, few laboratories are familiar with the culture identification of this unique pathogen and its differential diagnosis with similar filamentous fungi. METHODOLOGY: We have evaluated the use of matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF) on 53 isolates of P. insidiosum collected from cases of human and animal pythiosis in the USA and around the world. To assess the specificity of the approach, 18 pathogenic and saprotrophic filamentous fungal and fungal-like microbes were also tested. RESULTS: MALDI-TOF in-house spectra correctly identified the 53 P. insidiosum isolates (score range 1.93-2.51). MALDI-TOF based identification within P. insidiosum isolates showed protein spectra variation between geographical diverse isolates. A mass spectrometry approach was able to discriminate P. insidiosum from the 18 filamentous fungal and fungal-like microbes in this study, including four Pythium spp. and Phytopythium litorale plant pathogenic species. CONCLUSION: The data showed MALDI-TOF could be used for the accurate and rapid culture identification of P. insidiosum in the clinical laboratory.