Acanthamoeba spp. are known to cause human diseases such as granulomatous amoebic encephalitis and keratitis. These protozoa can persist in aquatic environments for extended periods and exhibit resistance to disinfectants used in water treatment, posing a significant threat to public health. Traditional culture and molecular methods have limitations in the rapid detection of viable Acanthamoeba in water. Therefore, we optimized real-time PCR (qPCR) combined with propidium monoazide (PMA) and evaluated the applicability of PMA-qPCR for the rapid detection of viable Acanthamoeba throughout the tap water production process. Using a PMA concentration of 50 µM, we successfully differentiated viable from non-viable cells and selected a 180-bp fragment of the 18S rRNA gene to enhance qPCR efficiency while minimizing amplification from non-viable cells. Application of PMA-qPCR to samples collected from each stage of the water treatment process, from raw water to purified water, enabled the detection of viable Acanthamoeba in samples with turbidity levels between 0.03 and 13.7 nephelometric turbidity units. These results demonstrate that PMA-qPCR is a suitable tool for monitoring pathogenic Acanthamoeba in drinking water supplies and enabling prompt responses to potential waterborne contamination events.

Enteric protozoan infections—cryptosporidiosis, giardiasis, microsporidiosis, and coccidiosis—constitute a significant global burden on human and animal health. Yet comprehensive parasitological surveys of wildlife in the eastern Tibetan Plateau, where anthropogenic, livestock, and wildlife activities extensively intersect, remain deficient. We collected 359 fecal specimens from wild carnivores across 98 random quadrats and employed molecular techniques for host identification and protozoan screening. Seven carnivore species were identified, with Tibetan fox (Vulpes ferrilata) predominating. Intestinal protozoa were exclusively recovered from Vulpes species, yielding a prevalence of 19.37% (68/351; 95% CI, 15.24%–23.51%). Eimeria falciformis (7.41%, 26/351) and Eimeria banffensis (7.12%, 25/351) were the most frequently detected, followed by Enterocytozoon bieneusi (3.70%, 13/351). Cryptosporidium was confirmed in one V. ferrilata, and Giardia intestinalis in 2 samples (V. ferrilata and Vulpes vulpes). This investigation furnishes the first molecular evidence that wild carnivores, especially V. ferrilata, harbor zoonotic protozoa including G. intestinalis assemblage AII and E. bieneusi genotypes PigEBITS5 and a D-like variant. The concurrent detection of Cryptosporidium yak genotype and E. bieneusi subgroup 1i in foxes, pikas, yaks, and sheep implies potential interspecies transmission at the wildlife–livestock interface. These findings fill knowledge gaps in regional parasite diversity and emphasize the need for enhanced surveillance of wildlife-borne protozoa under a One Health framework, thereby providing scientific evidence for zoonotic risk assessment and public health planning.

Persistent malaria transmission remains a considerable challenge in Indonesia despite national elimination efforts, with the Menoreh Hills representing a residual transmission hotspot characterized by ecological and socio-behavioral complexity. This highland region, spanning the boundaries of the Kulon Progo, Purworejo, and Magelang districts presents a complex eco-epidemiological setting in which malaria transmission persists despite substantial reductions in incidence. Spatial analyses consistently reveal micro-foci and cross-district clusters, underscoring the structural and ecological interconnectedness of the Menoreh Hills, including Anopheles mosquito habitats. Anopheles mosquitoes exhibit ecological adaptability and behavioral plasticity that may contribute to residual transmission. This region features a heterogeneous landscape of secondary forests, agricultural lands, and rural settlements that foster diverse vector species and habitats, occupational exposure, and sustained human-vector contact associated with Plasmodium transmission. Hypnozoite-mediated relapse in P. vivax, prolonged gametocyte carriage in P. falciparum, and asymptomatic or submicroscopic infections may contribute to residual transmission. Surveillance is complicated by fragmented governance, variable funding, high human mobility across district borders, and logistical constraints imposed by rugged topography. This review emphasizes the necessity of strengthening integrated malaria surveillance systems that combine entomological, serological, genomic, and spatial data to enable early detection and targeted interventions. Cross-sectoral collaboration, community engagement, and adaptive, location-specific strategies addressing socio-ecological determinants alongside biological and environmental drivers are essential to interrupt persistent transmission in complex residual foci such as the Menoreh Hills.

Sarcocystis spp. are common protozoan parasites infecting a wide range of intermediate hosts, including wild cervids. In this study, we investigated the occurrence and molecular characteristics of Sarcocystis infection in wild Korean roe deer (Capreolus pygargus tianschanicus) from Jeju Island, Korea. Sarcocysts were identified in the heart and skeletal muscle of 18 out of 40 examined animals (45.0%) based on histopathological examination. The cysts were oval to spherical in shape, measuring 42.41–340.29 μm in length and 31.62–129.76 μm in width, and were not associated with significant inflammatory responses. Molecular analysis targeting the 18S rRNA gene yielded 2 representative sequences, which showed 99.82% identity and high similarity to Sarcocystis linearis, as well as closely related species including S. taeniata and S. grueneri. Phylogenetic analysis placed these sequences within a clade of ungulate-associated Sarcocystis spp., although species-level resolution was not achieved due to the conserved nature of the marker. These findings provide baseline data on Sarcocystis infection in wild roe deer from Jeju Island and highlight the limitations of 18S rRNA-based identification for closely related Sarcocystis species.

Dirofilaria spp. are mosquito-borne filarial nematodes that primarily infect mammals. Here, we describe a filarial nematode infection in a female little egret (Egretta garzetta) from Jeju, Korea. Necropsy revealed multiple slender nematodes (4–6 cm) in the subcutaneous tissues of both legs. Histopathological examination demonstrated granulomatous inflammation with intralesional parasites. Morphologically, the nematodes exhibited a thick multilayered cuticle with prominent longitudinal ridges, consistent with members of the genus Dirofilaria. Molecular analysis based on 18S rRNA and mitochondrial c oxidase subunit I (cox1) gene sequences indicated that the parasite was most closely related to D. repens; however, the level of sequence divergence precluded definitive species-level identification. Phylogenetic analysis further demonstrated that the isolates clustered within the Dirofilaria clade and were clearly separated from avian-associated filarioid nematodes. This study provides molecular and histopathological evidence of a Dirofilaria sp. infection in an avian host and supports incidental infection under natural exposure conditions.

Human echinococcosis, caused by Echinococcus spp., remains a significant public health concern in endemic regions and manifests primarily as cystic echinococcosis (CE) and alveolar echinococcosis (AE), both characterized by chronic infection and heterogeneous clinical outcomes. This review synthesizes current evidence on host genetic determinants that influence immune responses, susceptibility, and disease persistence in CE and AE. Available studies indicate that polymorphisms in genes encoding key immunoregulatory cytokines, particularly IL10 and TGFB1, are associated with an anti-inflammatory immune profile that favors parasite persistence through enhanced regulatory activity and suppression of protective effector responses. Cytokine pathways involving IL-4, IL-13, and IFN-γ further modulate the balance between Th2-mediated and Th1-mediated immune responses, thereby influencing infection outcomes. In addition, innate immune pathways, including Toll-like receptor signaling, affect early parasite recognition and downstream immune responses, although direct human genetic evidence for Toll-like receptor polymorphisms in CE and AE remains limited. Human leukocyte antigen class II polymorphisms may also contribute to inter-individual differences in antigen presentation and CE susceptibility, although reported associations remain population-specific and require further validation. Collectively, these findings support a mechanistic framework in which host immunogenetic variation shapes the balance between immune activation and tolerance and may influence disease trajectory. Integration of genetic markers with clinical and imaging data may improve risk stratification and support the development of more targeted therapeutic strategies. Future research should prioritize large-scale, multicenter studies, standardized phenotyping, and functional validation to clarify the clinical relevance of identified variants.

Extracellular vesicles (EVs) secreted by Trichomonas vaginalis are now recognized as important mediators of host-parasite communication and play crucial roles in the development of trichomoniasis. These membrane-enclosed vesicles, which include exosomes and microvesicles, carry a diverse array of cargo, including proteins, lipids, and nucleic acids, as well as virulence-associated components such as adhesion molecules, proteases, and tRNA-derived small RNAs. Functionally, EVs enhance parasite attachment to vaginal epithelial cells, facilitate colonization, and contribute to host tissue damage. They also influence host immune responses, exhibiting both pro-inflammatory and immunosuppressive activities. Furthermore, the composition of EVs can be modulated by endosymbionts such as Trichomonasvirus, thereby influencing host-pathogen interactions. In addition, EVs mediate parasite-parasite communication, thereby promoting parasite adaptation and survival. This review summarizes current knowledge on the composition and functional roles of T. vaginalis EVs, highlighting their involvement in adhesion, immune modulation, and infection, as well as their potential utility as diagnostic biomarkers and therapeutic targets.

Coxiella burnetii is an environmentally stable intracellular bacterium responsible for severe global outbreaks of the zoonotic disease, Q fever. Q fever is transmitted by the inhalation of aerosols contaminated with the birth products and excretions of infected animals, mainly of sheep and goats. A 28-year-old cattle raiser presented with headache and fever without a significant medical history, although his cattle herd had previously been diagnosed with brucellosis. He spent time outdoors but reported no tick bites or eschar. Initial evaluation revealed mild thrombocytopenia and elevated aspartate aminotransferase, alanine aminotransferase, and C-reactive protein levels. On day 2 after disease onset, his blood tested positive for the IS1111 gene of C. burnetii by nested PCR. After amplifying C. burnetii in severe combined immunodeficient mice, the pathogen was isolated using a cell culture system. The isolated CH12 strain was confirmed via PCR, nucleotide sequence analysis, and whole-genome sequencing. This study reports a case of acute Q fever in Korea, in which C. burnetii was isolated and characterized using whole-genome sequencing.

Accurate identification of tick species is essential for understanding their role in pathogen transmission and guiding vector control strategies. Although morphological classification is widely used, it can be limited by specimen damage or immature developmental stages. In this study, we comparatively evaluated mitochondrial 16S and 12S ribosomal DNA (rDNA) markers for species-level identification and morphological verification of hard ticks and rodent-associated ectoparasites collected from vegetation, wild rodents, and patients in Gwangju, Republic of Korea. A total of 74 morphologically identified adult ticks—Haemaphysalis longicornis, Haemaphysalis flava, and Ixodes nipponensis—along with 3 damaged ticks removed from patients, were analyzed. PCR and sequencing success varied among species and markers. The 16S rDNA marker showed a higher amplification rate but lower sequencing success, whereas 12S rDNA showed more consistent performance. I. nipponensis showed the most balanced performance (89.5% for 16S and 78.9% for 12S), H. flava showed moderate variation, and H. longicornis demonstrated strong marker dependence, with low 16S but complete 12S success. Importantly, 16S rDNA identified 3 severely damaged ticks from patients—2 I. nipponensis and 1 Amblyomma testudinarium—demonstrating its diagnostic value when morphology was uninformative. Phylogenetic analyses of both markers yielded well-supported clades consistent with morphological identification. Additionally, 12S rDNA identified 3 flea specimens (Ctenophthalmus sp. and Neopsylla specialis) obtained during tick collection from wild rodents, indicating exploratory applicability to other rodent-associated ectoparasites. These findings support mitochondrial 16S and 12S rDNA markers as reliable tools for tick identification, especially when morphological assessment is limited by damage or developmental stage.

Glucose acquisition is essential for the long-term survival of adult Clonorchis sinensis within the host bile duct. Although several glucose transporter-related proteins have been identified in C. sinensis, the molecular and functional properties of facilitative glucose transport proteins, including the protein designated as glucose transporter protein 5 (CsGTP5) in this study, have not been characterized. In this study, CsGTP5 was cloned from adult C. sinensis and functionally analyzed using the Xenopus laevis oocyte expression system. The CsGTP5 open reading frame comprised 1,575 bp and encoded a 524-amino-acid protein with 12 predicted transmembrane domains, consistent with the topology of facilitative glucose transporter-like proteins. CsGTP5-expressing oocytes showed significantly increased uptake of [3 H] deoxy-D-glucose compared with water-injected control oocytes, whereas no significant uptake was observed for other tested substrates, including arginine, α-ketoglutarate, p-aminohippurate, taurocholate, and tetraethylammonium. CsGTP5-mediated [3 H] deoxy-D-glucose uptake increased in a time-dependent manner and was not affected by replacement of extracellular Na+ with Li+ or choline, indicating sodium-independent transport. Uptake was saturable, with an apparent Km of 4.0 mM and a Vmax of 300.0 pmol/oocyte/h. Competition assays showed that deoxy-D-glucose and glucose strongly inhibited CsGTP5-mediated uptake, whereas other monosaccharides had little effect. Molecular docking analysis further supported the predicted glucose-recognition capacity of CsGTP5. These findings demonstrate that CsGTP5 mediates sodium-independent deoxy-D-glucose uptake in a heterologous oocyte expression system and provide a basis for further investigation of its physiological role in C. sinensis.

Dense granule protein 16 (GRA16), an effector protein from Toxoplasma gondii, has been reported to enhance antitumor responses in several malignancies. However, whether GRA16 can enhance the inhibitory effects of standard anticancer drugs across diverse cancer types remains unclear. Gemcitabine, oxaliplatin, and irinotecan continue to play a key role in clinical oncology; however, their efficacy is limited by persistent tumor survival-promoting pathways. In this study, we stably expressed GRA16 in HepG2, PANC-1, and HCT116 cells and evaluated their responses to gemcitabine, oxaliplatin, and irinotecan under 50% inhibitory concentration-normalized dosing conditions. In vector control cells, baseline drug sensitivity was quantified using the CCK-8 assay across 0–64 μM after 48 h of exposure, and 50% inhibitory concentration was determined for each drug and cell line. Time-dependent proliferation assays (0–72 h) demonstrated that GRA16 reduced the basal growth rate and enhanced the growth inhibitory effects of all 3 drugs in all 3 cell models. Representative microscopy images at 48 h were consistent with reduced cell density in GRA16-expressing cultures under each treatment condition. These in vitro data indicate that GRA16 enhances the inhibitory effects of standard chemotherapeutic agents in hepatic, pancreatic, and colorectal cancer cell lines. This study provides a cross-cancer, multi-drug evaluation of GRA16-associated enhancement of drug response and includes, to our knowledge, the first assessment in a pancreatic cancer model. The underlying mechanism may involve pathway modulation, as suggested by previous studies and supported by the pancreatic cancer model although it was not comprehensively examined across all cell lines.

The present study was performed to investigate the infection status of zoonotic intestinal helminths in field mice captured from 7 local sites of Jeollabuk-do, Korea. A total of 78 Apodemus agrarius were collected in 7 survey sites, i.e., Sunchang-gun, Wanju-gun, Imsil-gun, Muju-gun, Gochang-gun, Jeonju-si, and Namwon-si, with Sherman live traps (Bioquip) for the surveillance of rodent-borne diseases through 2 times in 2019. The small intestine of each mouse was resected and longitudinally opened using scissors. The intestinal contents were washed with 0.85% saline until the supernatant became clear. The helminthic worms were collected with naked eyes and/or under a stereomicroscope from the sediment of the intestinal content. A total of 3 species of zoonotic intestinal helminths (1 trematode sp. and 2 cestode spp.) were collected. Plagiorchis muris were found in the small intestines of 7 (9.0%) mice (from Sunchang, Muju, Jeonju, and Namwon). Two species of tapeworm, Hymenolepis nana (from all survey sites except Wanju) and Hymenolepis diminuta (from Wanju, Imsil, and Namwon) were detected in 14 (17.9%) and 4 (5.1%) mice, respectively. The ventral view of P. muris was morphologically described. In this study, it was confirmed that 3 species of zoonotic intestinal helminths, P. muris, H. nana, and H. diminuta, are prevalent in striped field mice from Jeollabuk-do, Korea.