Vet World   Vol.18   November-2025  Article - 27 

First molecular evidence of Hepatocystis infection in non-human primates from Indonesia using fecal DNA: Implications for wildlife surveillance and One Health

Background and Aim: Hepatocystis, a malaria-like hemoparasite closely related to Plasmodium, infects non-human primates (NHPs), bats, and other mammals, yet remains understudied in Indonesia. Although Plasmodium detection in primates has been extensively reported, molecular confirmation of Hepatocystis in Indonesian wildlife is lacking. This study aimed to screen NHP fecal samples for Plasmodium infection and to identify any malaria-like parasites using molecular methods.

Materials and Methods: A total of 227 fecal samples from captive and rescued NHPs, representing multiple Macaca species and other primates, were collected from Tasikoki Wildlife Rescue Center, Manado, Indonesia, in 2019 and 2021. Genomic DNA was extracted using a QIAamp Fast DNA Stool Mini Kit (Qiagen, Germany) and screened for Plasmodium using polymerase chain reaction (PCR) targeting the mitochondrial small subunit ribosomal RNA gene. Positive amplicons were purified, sequenced, and analyzed using the basic local alignment search tool and phylogenetic reconstruction with MEGA X.

Results: Eight (3.5%) of 227 samples yielded positive PCR bands of approximately 600 bp, differing from the expected 467 bp for Plasmodium. Sequencing of four representative samples (MNig-01, MNig-17, MNig-18, and HM-160) revealed >99.7% identity with Hepatocystis spp. (GenBank: KY653782.1). Multiple sequence alignment confirmed complete nucleotide conservation among the four isolates, and phylogenetic analysis clustered them within the Hepatocystis clade, closely related to Hepatocystis spp. ex Pteropus hypomelanus from Malaysia. All positive detections were from 2019 samples, suggesting temporal variation in infection or vector activity.

Conclusion: This study provides the first molecular evidence of Hepatocystis infection in Indonesian NHPs using fecal DNA, expanding current knowledge of parasite distribution and host range. The successful detection of Hepatocystis through non-invasive sampling underscores the potential of fecal-based PCR for wildlife disease surveillance. These findings highlight the importance of integrating molecular diagnostics into conservation and One Health frameworks to monitor zoonotic parasites and understand host–vector–pathogen interactions in natural ecosystems.

Keywords: fecal DNA, Hepatocystis, Indonesia, non-human primates, One Health, Plasmodium, small subunit ribosomal RNA.

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How to cite this article: Hasan F, Siregar JE, Pravitasari NE, Rizki AFM, Azmi WA, Artika IM, and Kuswanda W (2025) First molecular evidence of Hepatocystis infection in non-human primates from Indonesia using fecal DNA: Implications for wildlife surveillance and One Health, Veterinary World, 18(11): 3651–3669.

Received: 28-07-2025   Accepted: 31-10-2025   Published online: 29-11-2025

Corresponding author: Josephine Elizabeth Siregar    E-mail: jose001@brin.go.id

DOI: 10.14202/vetworld.2025.3651-3669

Copyright: Hasan, et al. This article is an open access article distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http:// creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.