title: Development of an Anti-Elicitin Antibody-Based Immunohistochemical Assay for Diagnosis of Pythiosis.
abstract: Pythiosis is an emerging and life-threatening infectious disease of humans and animals living in tropical and subtropical countries and is caused by the fungus-like organism Pythium insidiosum. Antifungals are ineffective against this pathogen. Most patients undergo surgical removal of the infected organ, and many die from advanced infections. Early and accurate diagnosis leads to prompt management and promotes better prognosis for affected patients. Immunohistochemical assays (IHCs) have been developed using rabbit antibodies raised against P. insidiosum crude extract, i.e., culture filtrate antigen (CFA), for the histodiagnosis of pythiosis, but cross-reactivity with pathogenic fungi compromises the diagnostic performance of the IHC. Therefore, there is a need to improve detection specificity. Recently, the elicitin protein, ELI025, was identified in P. insidiosum, but it was not identified in other human pathogens, including true fungi. The ELI025-encoding gene was successfully cloned and expressed as a recombinant protein in Escherichia coli. This study aims to develop a new IHC using the rabbit anti-ELI025 antibody (anti-ELI) and to compare its performance with the previously reported anti-CFA-based IHC. Thirty-eight P. insidiosum histological sections stained positive by anti-ELI-based and anti-CFA-based IHCs indicating 100% detection sensitivity for the two assays. The anti-ELI antibody stained negative for all 49 negative-control sections indicating 100% detection specificity. In contrast, the anti-CFA antibody stained positive for one of the 49 negative controls (a slide prepared from Fusarium-infected tissue) indicating 98% detection specificity. In conclusion, the anti-ELI based IHC is sensitive and specific for the histodiagnosis of pythiosis and is an improvement over the anti-CFA-based assay.
authors.affiliation: Department of Pathology, Faculty of Medicine, Ramathibodi Hospital, Mahidol University, Bangkok, Thailand.
authors.forename: Ruchuros
authors.initials: R
authors.lastname: Inkomlue
authors.affiliation: Department of Pathology, Faculty of Medicine, Ramathibodi Hospital, Mahidol University, Bangkok, Thailand.
authors.forename: Noppadol
authors.initials: N
authors.lastname: Larbcharoensub
authors.affiliation: Department of Pathology, Faculty of Medicine, Ramathibodi Hospital, Mahidol University, Bangkok, Thailand.
authors.forename: Patcharee
authors.initials: P
authors.lastname: Karnsombut
authors.affiliation: Department of Pathology, Faculty of Medicine, Ramathibodi Hospital, Mahidol University, Bangkok, Thailand Molecular Medicine Program, Multidisciplinary Unit, Faculty of Science, Mahidol University, Bangkok, Thailand.
authors.forename: Tassanee
authors.initials: T
authors.lastname: Lerksuthirat
authors.affiliation: Department of Pathology, Faculty of Medicine, Ramathibodi Hospital, Mahidol University, Bangkok, Thailand.
authors.forename: Rangsima
authors.initials: R
authors.lastname: Aroonroch
authors.affiliation: Research Center, Faculty of Medicine, Ramathibodi Hospital, Mahidol University, Bangkok, Thailand.
authors.forename: Tassanee
authors.initials: T
authors.lastname: Lohnoo
authors.affiliation: Research Center, Faculty of Medicine, Ramathibodi Hospital, Mahidol University, Bangkok, Thailand.
authors.forename: Wanta
authors.initials: W
authors.lastname: Yingyong
authors.affiliation: Department of Pathology, Faculty of Medicine, Ramathibodi Hospital, Mahidol University, Bangkok, Thailand.
authors.forename: Pitak
authors.initials: P
authors.lastname: Santanirand
authors.affiliation: Department of Pathology, Faculty of Medicine, Chulalongkorn University, Bangkok, Thailand.
authors.forename: Lalana
authors.initials: L
authors.lastname: Sansopha
authors.affiliation: Department of Pathology, Faculty of Medicine, Ramathibodi Hospital, Mahidol University, Bangkok, Thailand mr_en@hotmail.com.
authors.forename: Theerapong
authors.initials: T
authors.lastname: Krajaejun
mesh_terms: D000818:Animals; D000906:Antibodies / Q000276:immunology*; D006801:Humans; D007150:Immunohistochemistry / Q000379:methods*; D058968:Pythiosis / Q000175:diagnosis*; D011775:Pythium / Q000276:immunology* / Q000302:isolation & purification*; D012680:Sensitivity and Specificity
chemical_list: D000906:Antibodies