Abstract
BackgroundParacoccidioidomycosis (PCM) is a neglected systemic mycosis caused by a dimorphic fungus of the Paracoccidioides genus. The standard diagnosis is based on isolation of the fungi in culture, and by microscopic visualization of characteristic multiple budding yeast cells in biological samples. However, in some situations, access to the site of injury prevents the collection of biological material. A variety of immuno-serological techniques has proven useful for allowing inferring diagnosis with a certain degree of certainty, thus optimizing time. The aim of this study was to standardize and validate the Dot-ELISA (DE) assay, comparing it with the serological standard, double immunodiffusion (DI).MethodsIn order to standardize the DE assay, 143 serum samples were used. Out of those, 23 were from apparently healthy patients, 77 were from patients with confirmed PCM and 43 were from patients with other lung infections (tuberculosis, aspergillosis and histoplasmosis). To validate the DE technique, 300 serum samples from patients with PCM clinical suspicion (probable and possible cases) were employed, and these results were compared with those of DI.ResultsThe DE assay showed sensitivity of 91%, specificity of 95.4%, positive predictive value of 96%, negative predictive value of 98.2%, accuracy of 93%, and great precision (k = 0.93). In addition, the nitrocellulose membranes have proved to be viable for using at least 90 days after P. brasiliensis B-339 antigen sensitization.ConclusionDot-ELISA method was found to be an extremely promising tool as serologic screening technique, because of its high sensitivity. Furthermore, Dot-ELISA shows the prospect of being transferred to laboratories of mycoserology including those with fewer resources or even to be used directly in the field. It has an excellent shelf life – membranes coated with antigen can be used for testing without changes in the pattern of reactivity among laboratories – and presents reliable values of sensitivity, specificity, predictive values, accuracy and a high correlation with the serological standard methodology. Based on the present findings, it possible to state that this technique constitutes a remarkable option to be used in routine diagnosis for public health centers.
Highlights
Paracoccidioidomycosis (PCM) is a neglected systemic mycosis caused by a dimorphic fungus of the Paracoccidioides genus
Employment of culture filtrate obtained from the B339 strain of P. brasiliensis instead of culture filtrate obtained from Pb 113 isolate
The intrinsic parameters were calculated for both methodologies (DE and double immunodiffusion (DI)): sensitivity of 91% (70/77) and 72.7% (56/77), specificity of 95.4% (63/66) and 98.5% (65/66), accuracy of 93% and 84.6% (121/143), positive and negative predictive values were 96% (70/73) and 98.2% (56/57), 90% (63/70) and 75.6% (65/86), respectively (Table 1)
Summary
Paracoccidioidomycosis (PCM) is a neglected systemic mycosis caused by a dimorphic fungus of the Paracoccidioides genus. Paracoccidioidomycosis (PCM) is the most important systemic mycosis of Latin America caused by the thermally dimorphic fungus Paracoccidioides spp. The definitive diagnosis of P. brasiliensis infection consists in the direct microscopic examination of biological specimens and their culturing followed by macro and microscopic observation for the fungus identification [10, 11]. Serological techniques are usually simpler than culture and are employed in early diagnosis of PCM, they are useful for monitoring its evolution and response to treatment [3, 10,11,12,13,14]
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