| Yeonchul Hong | 31 Articles |
Glucose-6-phosphate dehydrogenase (G6PD) deficiency is an X-linked recessive genetic disorder that can cause severe anemia in affected individuals exposed to oxidative stress. This risk is particularly relevant in patients treated with the antimalarial drug primaquine. In Myanmar, primaquine has been widely administered as a Plasmodium vivax malaria treatment; however, prevalence of G6PD deficiency among the population remains insufficiently characterized. This study investigated the prevalence of G6PD variants among various minority ethnic subgroups residing in Kachin State, Myanmar. Blood samples from 440 participants were analyzed; however, the Mahidol variant (G487A) was identified in 21 individuals (4.8%). A major limitation of this study was the absence of G6PD enzyme activity data to confirm whether the Mahidol variant induces G6PD deficiency.
Acanthamoeba is an opportunistic pathogen responsible for granulomatous amoebic encephalitis and amoebic keratitis. Despite its clinical significance, effective treatments remain challenging due to a limited understanding of its pathogenic mechanism. This study developed a genetic manipulation system in Acanthamoeba to facilitate gene function and drug screening studies. We applied the Cre/loxP system to integrate the gene encoding the tdTomato fluorescent protein into the genome of Acanthamoeba castellanii via homologous recombination. The polyubiquitin gene and its untranslated regions were identified and verified, after which the tdTomato gene was cloned between the untranslated regions of the polyubiquitin gene. The construct was then introduced into the Acanthamoeba genome using a modified pLPBLP vector containing loxP sites. Cre recombinase was utilized to remove the neomycin resistance cassette flanked by loxP sites, and genetically modified cells were selected by clonal dilution. The integration of the tdTomato gene, confirmed through PCR and fluorescence microscopy, showed stable expression in both trophozoites and cysts without the need for antibiotic selection. We demonstrated the feasibility of antibiotic-free reporter gene expression in Acanthamoeba. The system provides a valuable tool for functional genomics, allowing us to explore gene functions in Acanthamoeba and develop reliable drug screening models. Furthermore, the ability to express genes without the continuous use of selection markers opens up new possibilities for studying the pathobiology of this pathogen and advancing the development of novel therapeutic strategies against Acanthamoeba infections.
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Plasmodium vivax variant interspersed repeats (vir) refer to the key protein used for escaping the host immune system. Knowledge in the genetic variation of vir genes can be used for the development of vaccines or diagnostic methods. Therefore, we evaluated the genetic diversity of the vir genes of P. vivax populations of several Asian countries, including Pakistan, which is a malaria-endemic country experiencing a significant rise in malaria cases in recent years. We analyzed the genetic diversity and population structure of 4 vir genes (vir 4, vir 12, vir 21, and vir 27) in the Pakistan P. vivax population and compared these features to those of the corresponding vir genes in other Asian countries. In Pakistan, vir 4 (S=198, H=9, Hd=0.889, Tajima’s D value=1.12321) was the most genetically heterogenous, while the features of vir 21 (S=8, H=7, Hd=0.664, Tajima’s D value =-0.63763) and vir 27 (S =25, H =11, Hd =0.682, Tajima’s D value=-2.10836) were relatively conserved. Additionally, vir 4 was the most genetically diverse among Asian P. vivax populations, although within population diversity was low. Meanwhile, vir 21 and vir 27 among all Asian populations were closely related genetically. Our findings on the genetic diversity of vir genes and its relationships between populations in diverse geographical locations contribute toward a better understanding of the genetic characteristics of vir. The high level of genetic diversity of vir 4 suggests that this gene can be a useful genetic marker for understanding the P. vivax population structure. Longitudinal genetic diversity studies of vir genes in P. vivax isolates obtained from more diverse geographical areas are needed to better understand the function of vir genes and their use for the development of malaria control measures, such as vaccines.
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Acanthamoeba infection is associated with keratitis in humans; however, its association with keratitis in dogs remains unclear. To investigate this possibility, we collected 171 conjunctival swab samples from dogs with eye-related diseases (65 with keratitis and 106 without keratitis) at Chungbuk National University Veterinary Teaching Hospital, Korea, from August 2021 to September 2022. Polymerase chain reaction identified 9 samples (5.3%) as Acanthamoeba positive; of these, 3 were from dogs with keratitis (4.6%) and 6 were from dogs without keratitis (5.7%). Our results indicated no significant association between Acanthamoeba infection and keratitis, season, sex, or age. All Acanthamoeba organisms found in this study had the genotype T4, according to 18S ribosomal RNA analysis. Acanthamoeba infection in dogs might have only a limited association with keratitis.
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Glucose-6-phosphate dehydrogenase (G6PD) deficiency is caused by X-linked recessive disorderliness. It induces severe anemia when a patient with G6PD deficiency is exposed to oxidative stress that occurs with administration of an antimalarial drug, primaquine. The distribution of G6PD deficiency remains unknown while primaquine has been used for malaria treatment in Myanmar. This study aimed to investigate the prevalence of G6PD deficiency and its variants in Chin State, Myanmar. Among 322 participants, 18 (11 males and 7 females) demonstrated a G6PD deficiency. Orissa variant was dominant in the molecular analysis. This would be related to neighboring Indian and Bangladeshi population, in which Orissa variant was also reported as the main mutation type. The screening test for G6PD deficiency before primaquine treatment appears to be important in Myanmar.
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Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1), encoded by the polymorphic var multigene family, is a highly polymorphic antigen that plays a crucial role in the pathology of malaria. The contribution of the genetic diversity of var toward the immune escape of P. falciparum has not yet been fully elucidated. This study aimed to characterize the diversity of var repertoires by screening P. falciparum Duffy-binding-like α domain (PfDBLα) among field isolates from central Myanmar. Genetic analysis revealed that the D-H segments of var in Myanmar populations have an extensive polymorphic repertoire, with high numbers of unique sequence types in each individual. However, var genes from the global population, including Myanmar, shared close genetic lineages regardless of their geographic origins, indicating that they have not undergone rapid evolutionary changes.
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The encystation of Acanthamoeba leads to the development of metabolically inactive and dormant cysts from vegetative trophozoites under unfavorable conditions. These cysts are highly resistant to anti-Acanthamoeba drugs and biocides. Therefore, the inhibition of encystation would be more effective in treating Acanthamoeba infection. In our previous study, a sirtuin family protein—Acanthamoeba silent-information regulator 2-like protein (AcSir2)—was identified, and its expression was discovered to be critical for Acanthamoeba castellanii proliferation and encystation. In this study, to develop Acanthamoeba sirtuin inhibitors, we examine the effects of sirtinol, a sirtuin inhibitor, on trophozoite growth and encystation. Sirtinol inhibited A. castellanii trophozoites proliferation (IC50=61.24 μM). The encystation rate of cells treated with sirtinol significantly decreased to 39.8% (200 μM sirtinol) after 24 hr of incubation compared to controls. In AcSir2-overexpressing cells, the transcriptional level of cyst-specific cysteine protease (CSCP), an Acanthamoeba cysteine protease involved in the encysting process, was 11.6- and 88.6-fold higher at 48 and 72 hr after induction of encystation compared to control. However, sirtinol suppresses CSCP transcription, resulting that the undegraded organelles and large molecules remained in sirtinol-treated cells during encystation. These results indicated that sirtinol sufficiently inhibited trophozoite proliferation and encystation, and can be used to treat Acanthamoeba infections.
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Human sparganosis is a zoonotic disease caused by infection and migration of the plerocercoid of Spirometra spp. Although sparganosis were reported from most parts of the body, the sparganum parasitizing inside cerebral artery is remarkably uncommon. We report a case of cerebral intravascular sparganosis in an elderly patient with acute ischemic stroke who was diagnosed by retrieving sparganum during mechanical thrombectomy. Finally, the parasites were identified as Spirometra erinaceieuropaei using multiplex PCR and cox1 gene sequencing.
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From October 2015 to August 2018, tapeworm proglottids were obtained from 10 patients who were residents of Daegu and Gyeongbuk provinces and had a history of raw beef consumption. Most of them had no overseas travel experience. The gravid proglottids obtained from the 10 cases had 15-20 lateral uterine branches. A part of internal transcribed spacer 1 (ITS1) DNA of the 10 cases, amplified by polymerase chain reaction (PCR) and digested with AleI restriction enzyme, produced the same band pattern of Taenia saginata, which differentiated from T. asiatica and T. solium. Sequences of ITS1 and cytochrome c oxidase subunit 1 (cox1) showed higher homology to T. saginata than to T. asiatica and T. solium. Collectively, these 10 cases were identified as T. saginata human infections. As taeniasis is one of the important parasitic diseases in humans, it is necessary to maintain hygienic conditions during livestock farming to avoid public health concerns.
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Malarial infection induces tissue hypoxia in the host through destruction of red blood cells. Tissue hypoxia in malarial infection may increase the activity of HIF1α through an intracellular oxygen-sensing pathway. Activation of HIF1α may also induce vascular endothelial growth factor (VEGF) to trigger angiogenesis. To investigate whether malarial infection actually generates hypoxia-induced angiogenesis, we analyzed severity of hypoxia, the expression of hypoxia-related angiogenic factors, and numbers of blood vessels in various tissues infected with Plasmodium berghei. Infection in mice was performed by intraperitoneal injection of 2×106 parasitized red blood cells. After infection, we studied parasitemia and survival. We analyzed hypoxia, numbers of blood vessels, and expression of hypoxia-related angiogenic factors including VEGF and HIF1α. We used Western blot, immunofluorescence, and immunohistochemistry to analyze various tissues from Plasmodium berghei-infected mice. In malaria-infected mice, parasitemia was increased over the duration of infection and directly associated with mortality rate. Expression of VEGF and HIF1α increased with the parasitemia in various tissues. Additionally, numbers of blood vessels significantly increased in each tissue type of the malaria-infected group compared to the uninfected control group. These results suggest that malarial infection in mice activates hypoxia-induced angiogenesis by stimulation of HIF1α and VEGF in various tissues.
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PCR is known to be the most sensitive method for diagnosing Trichomonas vaginalis infections. This study aimed to compare the sensitivity of a PCR assay for trichomoniasis (HY-PCR) developed in Hanyang University with the use of a Seeplex Ace Detection Kit®, using urine collected from four Korean men with prostatic disease. Overall, HY-PCR was more sensitive than the Seeplex Kit. The use of Chelex 100 is recommended for DNA isolation in order to increase the sensitivity of the PCR test.
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Acanthamoeba spp. are free-living protozoa that are opportunistic pathogens for humans. Cysteine proteases of Acanthamoeba have been partially characterized, but their biochemical and functional properties are not clearly understood yet. In this study, we isolated a gene encoding cysteine protease of A. castellanii (AcCP) and its biochemical and functional properties were analyzed. Sequence analysis of AcCP suggests that this enzyme is a typical cathepsin L family cysteine protease, which shares similar structural characteristics with other cathepsin L-like enzymes. The recombinant AcCP showed enzymatic activity in acidic conditions with an optimum at pH 4.0. The recombinant enzyme effectively hydrolyzed human proteins including hemoglobin, albumin, immunoglobuins A and G, and fibronectin at acidic pH. AcCP mainly localized in lysosomal compartment and its expression was observed in both trophozoites and cysts. AcCP was also identified in cultured medium of A. castellanii. Considering to lysosomal localization, secretion or release by trophozoites and continuous expression in trophozoites and cysts, the enzyme could be a multifunctional enzyme that plays important biological functions for nutrition, development and pathogenicity of A. castellanii. These results also imply that AcCP can be a promising target for development of chemotherapeutic drug for Acanthamoeba infections.
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A fly larva was recovered from a boil-like lesion on the left leg of a 33-year-old male on 21 November 2016. He has worked in an endemic area of myiasis, Uganda, for 8 months and returned to Korea on 11 November 2016. The larva was identified as Cordylobia anthropophaga by morphological features, including the body shape, size, anterior end, posterior spiracles, and pattern of spines on the body. Subsequent 28S rRNA gene sequencing showed 99.9% similarity (916/917 bp) with the partial 28S rRNA gene of C. anthropophaga. This is the first imported case of furuncular myiasis caused by C. anthropophaga in a Korean overseas traveler.
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Protein arginine methyltransferase (PRMT) is an important epigenetic regulator in eukaryotic cells. During encystation, an essential process for Acanthamoeba survival, the expression of a lot of genes involved in the encystation process has to be regulated in order to be induced or inhibited. However, the regulation mechanism of these genes is yet unknown. In this study, the full-length 1,059 bp cDNA sequence of Acanthamoeba castellanii PRMT1 (AcPRMT1) was cloned for the first time. The AcPRMT1 protein comprised of 352 amino acids with a SAM-dependent methyltransferase PRMT-type domain. The expression level of AcPRMT1 was highly increased during encystation of A. castellanii. The EGFPAcPRMT1 fusion protein was distributed over the cytoplasm, but it was mainly localized in the nucleus of Acanthamoeba. Knock down of AcPRMT1 by synthetic siRNA with a complementary sequence failed to form mature cysts. These findings suggested that AcPRMT1 plays a critical role in the regulation of encystation of A. castellanii. The target gene of AcPRMT1 regulation and the detailed mechanisms need to be investigated by further studies.
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Encystation mediating cyst specific cysteine proteinase (CSCP) of Acanthamoeba castellanii is expressed remarkably during encystation. However, the molecular mechanism involved in the regulation of CSCP gene expression remains unclear. In this study, we focused on epigenetic regulation of gene expression during encystation of Acanthamoeba. To evaluate methylation as a potential mechanism involved in the regulation of CSCP expression, we first investigated the correlation between promoter methylation status of CSCP gene and its expression. A 2,878 bp of promoter sequence of CSCP gene was amplified by PCR. Three CpG islands (island 1-3) were detected in this sequence using bioinformatics tools. Methylation of CpG island in trophozoites and cysts was measured by bisulfite sequence PCR. CSCP promoter methylation of CpG island 1 (1,633 bp) was found in 8.2% of trophozoites and 7.3% of cysts. Methylation of CpG island 2 (625 bp) was observed in 4.2% of trophozoites and 5.8% of cysts. Methylation of CpG island 3 (367 bp) in trophozoites and cysts was both 3.6%. These results suggest that DNA methylation system is present in CSCP gene expression of Acanthamoeba. In addition, the expression of encystation mediating CSCP is correlated with promoter CpG island 1 hypomethylation.
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Variant surface antigens (VSAs) encoded by pir families are considered to be the key proteins used by many Plasmodium spp. to escape the host immune system by antigenic variation. This attribute of VSAs is a critical issue in the development of a novel vaccine. In this regard, a population genetic study of vir genes from Plasmodium vivax was performed in the Republic of Korea (ROK). Eighty-five venous blood samples and 4 of the vir genes, namely vir 27, vir 21, vir 12, and vir 4, were selected for study. The number of segregating sites (S), number of haplotypes (H), haplotype diversity (Hd), DNA diversity (π and Θw), and Tajima’s D test value were conducted. Phylogenetic trees of each gene were constructed. The vir 21 (S=143, H=22, Hd=0.827) was the most genetically diverse gene, and the vir 4 (S=6, H=4, Hd=0.556) was the opposite one. Tajima’s D values for vir 27 (1.08530, p>0.1), vir 12 (2.89007, p<0.01), and vir 21 (0.40782, p>0.1) were positive, and that of vir 4 (-1.32162, p>0.1) was negative. All phylogenetic trees showed 2 clades with no particular branching according to the geographical differences and cluster. This study is the first survey on the vir genes in ROK, providing information on the genetic level. The sample sequences from vir 4 showed a clear difference to the Sal-1 reference gene sequence, whereas they were very similar to those from Indian isolates.
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Trichomoniasis caused by Trichomonas vaginalis is a common sexually transmitted disease. Its association with several health problems, including preterm birth, pelvic inflammatory disease, cervical cancer, and transmission of human immunodeficiency virus, emphasizes the importance of improved access to early and accurate detection of T. vaginalis. In this study, a rapid and efficient loop-mediated isothermal amplification-based method for the detection of T. vaginalis was developed and validated, using vaginal swab specimens from subjects suspected to have trichomoniasis. The LAMP assay targeting the actin gene was highly sensitive with detection limits of 1 trichomonad and 1 pg of T. vaginalis DNA per reaction, and specifically amplified the target gene only from T. vaginalis. Validation of this assay showed that it had the highest sensitivity and better agreement with PCR (used as the gold standard) compared to microscopy and multiplex PCR. This study showed that the LAMP assay, targeting the actin gene, could be used to diagnose early infections of T. vaginalis. Thus, we have provided an alternative molecular diagnostic tool and a point-of-care test that may help to prevent trichomoniasis transmission and associated complications.
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Encystation is an essential process for Acanthamoeba survival under nutrient-limiting conditions and exposure to drugs. The expression of several genes has been observed to increase or decrease during encystation. Epigenetic processes involved in regulation of gene expression have been shown to play a role in several pathogenic parasites. In the present study, we identified the protein arginine methyltransferase 5 (PRMT5), a known epigenetic regulator, in Acanthamoeba castellanii. PRMT5 of A. castellanii (AcPRMT5) contained domains found in S-adenosylmethionine-dependent methyltransferases and in PRMT5 arginine-N-methyltransferase. Expression levels of AcPRMT5 were increased during encystation of A. castellanii. The EGFP-PRMT5 fusion protein was mainly localized in the nucleus of trophozoites. A. castellanii transfected with siRNA designed against AcPRMT5 failed to form mature cysts. The findings of this study lead to a better understanding of epigenetic mechanisms behind the regulation of encystation in cyst-forming pathogenic protozoa.
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This study explored epidemiological trends in trichomoniasis in Daegu, South Korea. Wet mount microscopy, PCR, and multiplex PCR were used to test for Trichomonas vaginalis in vaginal swab samples obtained from 621 women visiting 2 clinics in Daegu. Of the 621 women tested, microscopy detected T. vaginalis in 4 (0.6%) patients, PCR detected T. vaginalis in 19 (3.0%) patients, and multiplex PCR detected T. vaginalis in 12 (1.9%) patients. Testing via PCR demonstrated high sensitivity and high negative predictive value for T. vaginalis. Among the 19 women who tested positive for T. vaginalis according to PCR, 94.7% (18/19) reported vaginal signs and symptoms. Notably, more than 50% of T. vaginalis infections occurred in females younger than 30 years old, and 58% were unmarried. Multiplex PCR, which simultaneously detects pathogens from various sexually transmitted infections, revealed that 91.7% (11/12) of patients were infected with 2 or more pathogens. Mycoplasma hominis was the most prevalent co-infection pathogen with T. vaginalis, followed by Ureaplasma urealyticum and Chlamydia trachomatis. Our results indicate that PCR and multiplex PCR are the most sensitive tools for T. vaginalis diagnosis, rather than microscopy which has been routinely used to detect T. vaginalis infections in South Korea. Therefore, clinicians should take note of the high prevalence of T. vaginalis infections among adolescent and young women in order to prevent persistent infection and transmission of this disease.
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The parasite Plasmodium falciparum causes severe malaria and is the most dangerous to humans. However, it exhibits resistance to their drugs. Farnesyltransferase has been identified in pathogenic protozoa of the genera Plasmodium and the target of farnesyltransferase includes Ras family. Therefore, the inhibition of farnesyltransferase has been suggested as a new strategy for the treatment of malaria. However, the exact functional mechanism of this agent is still unknown. In addition, the effect of farnesyltransferase inhibitor (FTIs) on mitochondrial level of malaria parasites is not fully understood. In this study, therefore, the effect of a FTI R115777 on the function of mitochondria of P. falciparum was investigated experimentally. As a result, FTI R115777 was found to suppress the infection rate of malaria parasites under in vitro condition. It also reduces the copy number of mtDNA-encoded cytochrome c oxidase III. In addition, the mitochondrial membrane potential (ΔΨm) and the green fluorescence intensity of MitoTracker were decreased by FTI R115777. Chloroquine and atovaquone were measured by the mtDNA copy number as mitochondrial non-specific or specific inhibitor, respectively. Chloroquine did not affect the copy number of mtDNA-encoded cytochrome c oxidase III, while atovaquone induced to change the mtDNA copy number. These results suggest that FTI R115777 has strong influence on the mitochondrial function of P. falciparum. It may have therapeutic potential for malaria by targeting the mitochondria of parasites.
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Ascidian soft tunic syndrome (AsSTS) caused by Citations Citations to this article as recorded by
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Autophagy-related protein 8 (Atg8) is an essential component of autophagy formation and encystment of cyst-forming parasites, and some protozoa, such as, Citations Citations to this article as recorded by
Amoebic keratitis (AK) caused by Citations Citations to this article as recorded by
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Rodent malaria parasites, such as
Autophagy is a catabolic process involved in the degradation of a cell's own components for cell growth, development, homeostasis, and the recycling of cellular products. Autophagosome is an essential component in the protozoan parasite during differentiation and encystation. The present study identified and characterized autophagy-related protein (Atg) 3, a member of Atg8 conjugation system, in Citations Citations to this article as recorded by
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