XI International Meeting on Paracoccidioidomycosis

XI International Meeting on Paracoccidioidomycosis

Resume:149-1

Poster (Painel)

149-1

LATEX AGGLUTINATION TEST FOR DETECTION OF SPECIFIC ANTIBODY ANTI-GP43 FOR RAPID DIAGNOSIS OF PARACOCCIDIOIDOMYCOSIS

Authors:

Priscila Oliveira dos Santos (UNIFESP - Federal University of São Paulo) ; Anderson Messias Rodrigues (UNIFESP - Federal University of São Paulo) ; Geisa Ferreira Fernandes (UNIFESP - Federal University of São Paulo) ; Cristiane Candida do Amaral (UNIFESP - Federal University of São Paulo) ; Alexandre Augusto Sasaki (UNIFESP - Federal University of São Paulo) ; Silvia Helena Marques-da-silva (IEC/SVS/MS - Evandro Chagas Institute, Bacteriology and Mycology Section.) ; Zoilo Pires de Camargo (UNIFESP - Federal University of São Paulo)

Abstract

Paracoccidioides brasiliensis is the causal agent of paracoccidiodomycosis (PCM), a systemic granulomatous disease, which is most prevalent in Latin America, especially in Brazil. Diagnosis can be obtained by direct visualization, culture, and serologic tests. Serologic procedures are time consuming and lack specificity. In case of PCM, most serologic diagnostic assays use crude antigenic preparations, that give cross-reactions. Therefore, there is a need for a faster, sensitive and specific diagnosis of PCM to initiate appropriate management as well as to evaluate the use of new therapeutics. Gp43 is the immunodominant antigen recognized by up to 100% of patients with PCM. Also, gp43 stimulates the humoral immune response and is useful in diagnosis and post therapeutic control. In this study, a latex agglutination test (LAT) was established for detection of anti-gp43 antibody in serum samples. Carboxylated latex particles were washed in appropriate wash buffer. Then, the microspheres were resuspended in coupled buffer and gp43. The mixture was incubated overnight at room temperature. The mixture was centrifuged and pellet was resuspended in buffer containing 0,1% bovine serum albumin and incubated for 4 hours at room temperature with gentle mixing. The supernatant was removed and the sensitized latex particles (SLPs) were resuspended in storage buffer and stocked at 4ÂºC until use. Latex agglutination tests were performed by mixing 25 Âµl of SLPs with 25 µl of serum sample on a black-coated slide. The slide was then rotated manually for 30 sec. The results were scored as ++++ (rapid agglutination of 100% SLPs), +++ (agglutination of 75% of SLPs), ++ (agglutination of approximately 50% of SLPs), + (agglutination of 25% of SLPs), and â€“ (no visible agglutination). The sample was considered positive if agglutination of 25% SLPs occurred within 15 min at 37ºC. Sera from 118 patients with PCM, 49 clinical samples from patients with other mycosis and 10 normal patients were tested. We observed a sensitivity and specificity of 94.91% and 92.45%, respectively. These results indicate that the latex test couples with gp43 is sensitive, specific, economical, and fast for serodiagnosis of PCM.਀ This study was supported by FAPESP, CNPq, and CAPES.਀

Keyword: paracoccidioidomycosis, gp43, latex agglutination, antibody

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