Aras Part Medical International Press International Journal of Medical Parasitology and Epidemiology Sciences 2766-6492 7 2 2026 06 29 Detection of Intestinal Protozoa in HIV Patients: A Comparative Study of Microscopy and Multiplex Real-Time PCR 101 110 10.34172/ijmpes.6268 EN Nuning Winaris https://orcid.org/0000-0001-5941-7095 Loeki Enggar Fitri https://orcid.org/0000-0002-4880-1048 Aulia Rahmi Pawestri https://orcid.org/0000-0002-8308-9930 Didi Candradikusuma https://orcid.org/0000-0002-0577-7279 Iraky Mardya Rakhmadhan https://orcid.org/0000-0003-2277-1685 Firman Arief https://orcid.org/0009-0008-9298-2799 Adam Adam https://orcid.org/0009-0005-4842-5520 Abdurachman Omar Baabdullah https://orcid.org/0009-0009-6710-8351 Sara Hayati https://orcid.org/0009-0006-9535-6668 Syarellia Safira Putri https://orcid.org/0009-0004-3034-408X Nectarine Natasya Regitta Yasmin https://orcid.org/0009-0008-6119-890X Levina Auryn Noor Putri Joe https://orcid.org/0009-0005-9123-296X Hanifa Rizky Rahmawati https://orcid.org/0009-0007-1242-773X Maria Mardalena Martini Kaisar https://orcid.org/0000-0002-7515-7673 Journal Article 10.34172/ijmpes.6268 2026 03 19 2026 06 05 Introduction: Intestinal protozoa are traditionally diagnosed through microscopic examination, which is labor-intensive and often requires repeated sampling. Recent molecular approaches, particularly multiplex Real-Time PCR (RT-PCR), provide a faster alternative for parasite detection. This study aimed to compare the sensitivity and specificity of direct smear and multiplex RT-PCR methods against trichrome staining for intestinal protozoa detection. Methods: A cross-sectional study was conducted using 51 stool samples collected from HIV-naïve patients, HIV patients on therapy, and healthy volunteers at General Hospital Dr. Saiful Anwar, Malang, Indonesia. The samples consisted of 41 HIV-positive individuals (17 HIV-naïve and 24 on therapy) and 10 healthy volunteers. Stool specimens were examined using direct smear, trichrome staining, and multiplex RT-PCR. Results: Microscopic examination identified Entamoeba histolytica, Blastocystis hominis, coccidian parasites, and non-pathogenic amoebae. In contrast, multiplex RT-PCR detected only B. hominis in both HIV and non-HIV samples. Compared with trichrome staining, direct smear demonstrated 36.6% sensitivity and 80.0% specificity, while multiplex RT-PCR showed higher sensitivity (63.4%) but lower specificity (40.0%). Differences among methods may be influenced by limited sample size, population characteristics, clinical status, and technical or biological factors affecting microscopy and molecular examinations. Conclusion: Combining molecular diagnostics with conventional microscopy may improve the diagnostic accuracy of intestinal protozoa infections and support more effective treatment strategies and better clinical outcomes in HIV patients.