Design and optimization of an improved qPCR assay for the detection of Histoplasma capsulatum.

Autor:	da Silva BKNI; 1Graduate Program in Pharmaceutical Sciences, Universidade Federal do Amazonas-UFAM, Manaus, Brazil., Alves Cortez AC; 2Mycology Laboratory, Instituto Nacional de Pesquisas da Amazonia-INPA, Manaus, Brazil., Oliveira LA; 2Mycology Laboratory, Instituto Nacional de Pesquisas da Amazonia-INPA, Manaus, Brazil., de Almeida JDR; 2Mycology Laboratory, Instituto Nacional de Pesquisas da Amazonia-INPA, Manaus, Brazil., de Souza ÉS; 3Superior School of Technology, Universidade do Estado do Amazonas -UEA, Manaus, Brazil., Cruz KS; 4Medical Mycology Laboratory, Fundação de Medicina Tropical Doutor Heitor Vieira Dourado-FMT-HVD, Manaus, Brazil., Jackisch Matsuura AB; 5Microbial Diversity Laboratory of the Amazon with Health Importance, Leônidas and Maria Deane Institute-Fiocruz-AM, Manaus, Amazonas, Brazil., Ogusku MM; 6Mycobacteriology Laboratory, Instituto Nacional de Pesquisas da Amazonia-INPA, Manaus, Brazil., Solórzano Chavarría KJ; 7Graduate Program in Tropical Medicine, Fundação de Medicina Tropical Doutor Heitor Vieira Dourado-FMT-HVD, Manaus, Brazil.; 8Carlos Borborema Clinical Research Unit, Fundação de Medicina Tropical Doutor Heitor Vieira Dourado-FMT-HVD, Manaus, Brazil., Baía-da-Silva D; 1Graduate Program in Pharmaceutical Sciences, Universidade Federal do Amazonas-UFAM, Manaus, Brazil.; 7Graduate Program in Tropical Medicine, Fundação de Medicina Tropical Doutor Heitor Vieira Dourado-FMT-HVD, Manaus, Brazil.; 8Carlos Borborema Clinical Research Unit, Fundação de Medicina Tropical Doutor Heitor Vieira Dourado-FMT-HVD, Manaus, Brazil.; 9Instituto Leônidas Maria Deane- ILMD, Fiocruz-AM, Manaus, Brazil.; 10Pharmacy Faculty, Universidade Nilton Lins, Manaus, Brazil., Bassat Q; 11ISGlobal, Hospital Clínic - Universitat de Barcelona, Barcelona, Spain.; 12Centro de Investigação em Saúde de Manhiça (CISM), Maputo, Mozambique.; 13ICREA, Pg. Lluís Companys 23, 080100 Barcelona, Spain.; 14Pediatrics Department, Hospital Sant Joan de Déu, Universitat de Barcelona, Esplugues, Barcelona, Spain.; 15Consorcio de Investigación Biomédica en Red de Epidemiología y Salud Pública (CIBERESP), Madrid, Spain., Guimarães Lacerda MV; 7Graduate Program in Tropical Medicine, Fundação de Medicina Tropical Doutor Heitor Vieira Dourado-FMT-HVD, Manaus, Brazil.; 8Carlos Borborema Clinical Research Unit, Fundação de Medicina Tropical Doutor Heitor Vieira Dourado-FMT-HVD, Manaus, Brazil.; 9Instituto Leônidas Maria Deane- ILMD, Fiocruz-AM, Manaus, Brazil.; 18University of Texas Medical Branch, Galveston, USA., Frickmann H; 16Institute for Medical Microbiology, Virology and Hygiene, University Medicine Rostock, Rostock, Germany.; 17Department of Microbiology and Hospital Hygiene, Bundeswehr Hospital, Hamburg, Germany., de Souza JVB; 2Mycology Laboratory, Instituto Nacional de Pesquisas da Amazonia-INPA, Manaus, Brazil.
Jazyk:	angličtina
Zdroj:	European journal of microbiology & immunology [Eur J Microbiol Immunol (Bp)] 2023 Sep 21. Date of Electronic Publication: 2023 Sep 21.
DOI:	10.1556/1886.2023.00026
Abstrakt:	Background: This study aimed at improving a real-time polymerase-chain-reaction (qPCR) assay for the detection of Histoplasma capsulatum, a fungal pathogen that can cause severe respiratory infections in humans, in clinical and soil samples. Methods: Primer and probes were in-silico designed, in-silico and in-vitro evaluated including clinical biopsy materials and finally subjected to a real-world application with collected soil samples. Results: Applying the qPCR assay with liver and lung biopsies from 71 patients each, including 59 patients infected with human immunodeficiency virus (HIV), as well as with Sabouraud (SAB) agar culture as the diagnostic reference standard, diagnostic accuracy of the qPCR assay of 100% (5/5) sensitivity and 96% (63/66) specificity for liver samples and 100% (4/4) sensitivity and 94% (63/67) specificity for the lung samples was recorded. When applying the assay with soil samples from caves near of Presidente Figueiredo city, Amazonas, Brazil, one sample from the Maroaga cave was confirmed as positive. Conclusions: The improved qPCR assessed in this study was successful in detecting H. capsulatum with high efficiency and accuracy in in-vitro evaluation, including the identification of the target pathogen in both clinical and environmental samples.
Databáze:	MEDLINE
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