Abstract
Background: In patients with HIV, conventional tests are of low sensitivity; therefore, a new diagnostic test with hydrogel nanoparticles with reactive blue dye is proposed, which allows the capture, conservation and concentration of ESAT-6 in urine. NIPAs are copolymers that capture low molecular weight proteins and protect against enzymatic degradation. Using an immunoassay, it is possible to detect and quantify ESAT-6 present in urine as a possible marker of active TB. Design/Methods: Study in Lima, Peru, HIV+ participants, ≥18 years with and without tuberculosis (TB). Smear microscopy, culture in solid medium and urine immunoassay were performed. The reference was the diagnosis of TB by radiological or clinical microbiological criteria (indication for TB treatment). There were 2 preanalytical processes: untreated and treated urine (centrifuged, heated), then incubation with NIPAm. After washing, elution, sonication, heat and centrifugation, the final eluate was obtained. This was placed on nitrocellulose membranes, which by means of fixation and incubation processes with anti-ESAT-6 and anti-IgG antibodies, revelation with C-DiGit® Blot Scanner and FluorChem R FR0001. Calibration curves were included in the membranes, density was measured using Image J software. ROC curves, sensitivity and specificity were obtained. Results: The result by groups was HIV+ patient: ROC: 0.75, cut-off point ≥24.06 ng/ml, sensitivity 76.32%, specificity 68.89%, patients ≤200 cells CD4 mm3/ml, ROC: 0.78, cutoff point ≥26.20 ng/ml, sensitivity 75.86%, specificity 71.88%, patients >200 CD4 mm3 cells/ml, ROC: 0.73, cutoff point ≥24.6 CD4 mm3/ml, sensitivity 73.68%, specificity 73.68%. Conclusions: The ESAT-6 detection assay using NIPAm was effective, with higher rates in patients with ≤200 CD4 cells/mm3, the test being more sensitive than smear and culture, but less specific.
Highlights
Tuberculosis (TB) and human immunodeficiency virus (HIV) infection are two of the leading causes of infectious mortality worldwide [1]
Patients HIV positive are highly susceptible to tuberculosis infection, due to their compromised immune system, and low immune response, which does not form caverns, his patients had a decreased proliferation of bacilli, that difficult the conventional test like direct bacilli observation, culture and nucleic acid amplification [6] [7] [8]
The ESAT-6 protein in urine was present in all study groups, with TB patients having a higher concentration
Summary
Tuberculosis (TB) and human immunodeficiency virus (HIV) infection are two of the leading causes of infectious mortality worldwide [1]. Patients HIV positive are highly susceptible to tuberculosis infection, due to their compromised immune system, and low immune response, which does not form caverns, his patients had a decreased proliferation of bacilli, that difficult the conventional test like direct bacilli observation, culture and nucleic acid amplification [6] [7] [8]. This is the main reason for late diagnosis and treatment initiation, increasing the probability of M. Results: The result by groups was HIV+ patient: ROC: 0.75, cut-off point ≥24.06 ng/ml, sensitivity 76.32%, specificity 68.89%, patients ≤200 cells CD4 mm3/ml, ROC: 0.78, cutoff point ≥26.20 ng/ml, sensitivity 75.86%, specificity 71.88%, pa-
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