Nuning Winaris
1,2 
, Loeki Enggar Fitri
1,2* , Aulia Rahmi Pawestri
1,2 , Didi Candradikusuma
3,4 , Iraky Mardya Rakhmadhan
4,5 , Firman Arief
4,5 , Adam Adam
4,5 , Abdurachman Omar Baabdullah
4,5 , Sara Hayati
6 , Syarellia Safira Putri
6 , Nectarine Natasya Regitta Yasmin
6 , Levina Auryn Noor Putri Joe
6 , Hanifa Rizky Rahmawati
7 , Maria Mardalena Martini Kaisar
8,9 1 Department of Clinical Parasitology, Faculty of Medicine, Universitas Brawijaya, Malang, Indonesia
2 AIDS, Toxoplasma, Opportunistic Disease and Malaria Research Group, Faculty of Medicine, Universitas Brawijaya, Malang, Indonesia
3 Department of Internal Medicine, Faculty of Medicine, Universitas Brawijaya, Malang, Indonesia
4 Medical Staff Group of Internal Medicine Dr. Saiful Anwar General Hospital, Malang, Indonesia
5 Residency Training Program, Department of Internal Medicine, Faculty of Medicine, Universitas Brawijaya, Malang, Indonesia
6 Bachelor of Medicine Program, Faculty of Medicine, Universitas Brawijaya, Malang, Indonesia
7 Master Program in Biomedical Science, Faculty of Medicine, Universitas Brawijaya, Malang, Indonesia
8 Master in Biomedicine Study Program, School of Medicine and Health Sciences, Atma Jaya Catholic University of Indonesia, Jakarta, Indonesia
9 Department of Parasitology, School of Medicine and Health Sciences, Atma Jaya Catholic University of Indonesia, Jakarta, Indonesia
Abstract
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.