Prevalence of Trichomonas vaginalis in Women Visiting 2 Obstetrics and Gynecology Clinics in Daegu,South Korea

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.     

Prevalence of Trichomoniasis by PCR in Women Attending Health Screening in Korea

Trichomoniasis is the most common curable sexually-transmitted infection (STI) worldwide. There are few reports on the prevalence of Trichomonas vaginalis in Korea. The purpose of this study was to examine the prevalence of trichomoniasis by PCR in Guri city, Korea. All adult women who visited Hanyang University Guri Hospital for health screening within the National Health Care Service were invited to participate in the study, and 424 women were enrolled between March and June 2011. PCR was used to detect Trichomonas vaginalis using primers based on a repetitive sequence cloned from T. vaginalis (TV-E650). Fourteen women (3.3%) were found to have T. vaginalis. All were over 50, and they were significantly older on average than the 410 Trichomonas-negative women (mean ages 63.4 vs 55.3 years). It seems that T. vaginalis infection is not rare in women receiving health screening, especially among those over 50.     

Inflammatory Responses in a Benign Prostatic Hyperplasia Epithelial Cell Line (BPH-1) Infected with Trichomonas vaginalis

Trichomonas vaginalis causes the most prevalent sexually transmitted infection worldwide. Trichomonads have been detected in prostatic tissues from prostatitis, benign prostatic hyperplasia (BPH), and prostate cancer. Chronic prostatic inflammation is known as a risk factor for prostate enlargement, benign prostatic hyperplasia symptoms, and acute urinary retention. Our aim was to investigate whether T. vaginalis could induce inflammatory responses in cells of a benign prostatic hyperplasia epithelial cell line (BPH-1). When BPH-1 cells were infected with T. vaginalis, the protein and mRNA of inflammatory cytokines, such as CXCL8, CCL2, IL-1β, and IL-6, were increased. The activities of TLR4, ROS, MAPK, JAK2/STAT3, and NF-κB were also increased, whereas inhibitors of ROS, MAPK, PI3K, NF-κB, and anti-TLR4 antibody decreased the production of the 4 cytokines although the extent of inhibition differed. However, a JAK2 inhibitor inhibited only IL-6 production. Culture supernatants of the BPH-1 cells that had been incubated with live T. vaginalis (trichomonad-conditioned medium, TCM) contained the 4 cytokines and induced the migration of human monocytes (THP-1 cells) and mast cells (HMC-1 cells). TCM conditioned by BPH-1 cells pretreated with NF-κB inhibitor showed decreased levels of cytokines and induced less migration. Therefore, it is suggested that these cytokines are involved in migration of inflammatory cells. These results suggest that T. vaginalis infection of BPH patients may cause inflammation, which may induce lower urinary tract symptoms (LUTS).     

Prevalence of Toxoplasma gondii Infection in Stray and Household Cats in Regions of Seoul, Korea

The principal objective of this study was to investigate the prevalence of toxoplasmosis in household and stray cats in Seoul, Republic of Korea. We collected blood samples from 72 stray and 80 household cats, and all samples were examined by ELISA and nested PCR. The overall positive rates of Toxoplasma gondii in stray cats were 38.9% (28/72), with 15.3% (11/72) in ELISA and 30.6% (22/72) in PCR. The positive rate in male stray cats was slightly higher than that of female stray cats. The highest positive rate of T. gondii infection was noted in Gangnam and Songpa populations in ELISA and in Gwangjin population in PCR. In household cats, however, we could not detect any specific antibodies or DNA for T. gondii. In conclusion, we recognized that the infection rate of toxoplasmosis in stray cats in Seoul was considerably high but household cats were free from infection.     

Prevalence of Dirofilaria immitis Infection in Stray Cats by Nested PCR in Korea

The purpose of this study was to conduct a survey of Dirofilaria immitis infection among stray cats in Korea using nested PCR. We included 235 stray cats (121 females and 114 males) and evaluated each for the presence of feline heartworm infection. Blood samples were collected from 135 cats in Daejeon, 50 cats in Seoul, and 50 cats from Gyeonggi-do (Province). Of the 235 DNA samples, 14 (6.0%) were positive for D. immitis. The prevalence of infection in male cats (8/114, 7.0%) tended to be higher than that in female cats (6/121, 5.0%), but the difference was not statistically significant. In each location, 8, 2, and 4 cats were positive for infection, respectively, based on DNA testing. No significant differences in the prevalence were observed among the geographic regions, although the rate of infection was higher in Gyeonggi-do (8.0%) than Daejeon (5.9%) and Seoul (4.0%). We submitted 7 of the 14 D. immitis DNA-positive samples for sequencing analysis. All samples corresponded to partial D. immitis cytochrome c oxidase subunit I gene sequences with 99% homology to the D. immitis sequence deposited in GenBank (accession no. {"type":"entrez-nucleotide","attrs":{"text":"FN391553","term_id":"295980069","term_text":"FN391553"}}FN391553). To the best of our knowledge, this is the first survey using nested PCR to analyze the prevalence of D. immitis in stray cats in Korea.     

PCR for diagnosis of male Trichomonas vaginalis infection with chronic prostatitis and urethritis

The aim of this study was to assess the usefulness of PCR for diagnosis of Trichomonas vaginalis infection among male patients with chronic recurrent prostatitis and urethritis. Between June 2001 and December 2003, a total of 33 patients visited the Department of Urology, Hanyang University Guri Hospital and were examined for T. vaginalis infection by PCR and culture in TYM medium. For the PCR, we used primers based on a repetitive sequence cloned from T. vaginalis (TV-E650). Voided bladder urine (VB1 and VB3) was sampled from 33 men with symptoms of lower urinary tract infection (urethral charge, residual urine sensation, and frequency). Culture failed to detect any T. vaginalis infection whereas PCR identified 7 cases of trichomoniasis (21.2%). Five of the 7 cases had been diagnosed with prostatitis and 2 with urethritis. PCR for the 5 prostatitis cases yielded a positive 330 bp band from bothVB1 and VB3, whereas positive results were only obtained from VB1 for the 2 urethritis patients. We showed that the PCR method could detect T. vaginalis when there was only 1 T. vaginalis cell per PCR mixture. Our results strongly support the usefulness of PCR on urine samples for detecting T. vaginalis in chronic prostatitis and urethritis patients.     

Loop-Mediated Isothermal Amplification Targeting Actin DNA of Trichomonas vaginalis

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.     

Serologic Survey of Toxoplasmosis in Seoul and Jeju-do, and a Brief Review of Its Seroprevalence in Korea

Knowledge of the prevalence of human Toxoplasma gondii infection is required in the Republic of Korea. In this study, we surveyed the seroprevalence of T. gondii infection and analyzed the risk factors associated with seropositivity among residents in 2 administrative districts; Seoul and the island of Jeju-do, which have contrasting epidemiologic characteristics. Sera and blood collected from 2,150 residents (1,114 in Seoul and 1,036 in Jeju-do) were checked for IgG antibody titers using ELISA and for the T. gondii B1 gene using PCR. In addition, participants completed a questionnaire that solicited information on gender, age, occupation, eating habits, history of contact with animals, and travel abroad. The T. gondii B1 gene was not detected in all residents examined. However, ELISA showed 8.0% (89 of 1,114 sera) positive for IgG antibodies against T. gondii in Seoul and 11.3% (117 of 1,036 sera) in Jeju-do. In both districts, the positive rates were higher in males than in females, and those 40-79 years of age showed higher rates than other ages. In Seoul, residents older than 70 years of age showed the highest positive rate, 14.9%, whereas in Jeju-do the highest prevalence, 15.6%, was in those in their sixties. The higher seropositive rate in Jeju-do than in Seoul may be related to eating habits and occupations. The present results and a review of related literature are indicative of an increased seroprevalence of T. gondii in Korea in recent years.     

Prevalence of Toxoplasma gondii Infection in Household Cats in Korea and Risk Factors

Several epidemiological surveys have reported the prevalence of Toxoplasma gondii infection in stray cats in Korea, but little information is available on T. gondii infection in household cats. The aim of the present study was to assess the prevalence and risk factors of T. gondii infection among household cats reared in Seoul, Korea. A total of 474 blood samples were collected from clinically healthy household cats. All samples were tested using ELISA and PCR. The risk factor analysis was based on a questionnaire filled out by the owners. The overall positive rate for ELISA and PCR assays was 2.2% (10/437) and 2.1% (10/474), respectively. With regard to the origin of cats, the positive rates among cats adopted from the animal shelter and veterinary clinic for stray cats were significantly different (P<0.05). Our study demonstrated that the positive rate of T. gondii infection in household cats was low and that this low prevalence was assumed to be associated with keeping the cats indoors and restriction of eating raw food and uncooked meat. Therefore, we suggest that the owners check the origin of the cats prior to adoption to prevent infection of other animals, including humans.     

Optimization of Trichomonas vaginalis Diagnosis during Pregnancy at a University Hospital,Argentina

The aim of this study was to evaluate different methods for Trichomonas vaginalis diagnosis during pregnancy in order to prevent maternal and perinatal complications. A total of 386 vaginal exudates from pregnant women were analyzed. T. vaginalis was investigated by 3 types of microscopic examinations direct wet mount with physiologic saline solution, prolonged May-Grunwald Giemsa (MGG) staining, and wet mount with sodium-acetate-formalin (SAF)/methylene blue method. PCR for 18S rRNA gene as well as culture in liquid medium were performed. The sensitivity and specificity of the microscopic examinations were evaluated considering the culture media positivity or the PCR techniques as gold standard. The frequency of T. vaginalis infection was 6.2% by culture and/or PCR, 5.2% by PCR, 4.7% by culture, 3.1% by SAF/methylene blue method and 2.8% by direct wet smear and prolonged MGG staining. The sensitivities were 83.3%, 75.0%, 50.0%, and 45.8% for PCR, culture, SAF/methylene blue method, and direct wet smear-prolonged MGG staining, respectively. The specificity was 100% for all the assessed methods. Microscopic examinations showed low sensitivity, mainly in asymptomatic pregnant patients. It is necessary to improve the detection of T. vaginalis using combined methods providing higher sensitivity, such as culture and PCR, mainly in asymptomatic pregnant patients, in order to prevent maternal and perinatal complications.     

Toxoplasma gondii B1 Gene Detection in Feces of Stray Cats around Seoul,Korea and Genotype Analysis of Two Laboratory-Passaged Isolates

The increasing prevalence of Toxoplasma gondii infection in the human population in the Republic of Korea (= Korea) is due to various reasons such as an increase in meat consumption. However, the importance of cats in transmitting T. gondii infection through oocysts to humans has seldom been assessed. A total of 300 fecal samples of stray cats captured around Seoul from June to August 2013 were examined for T. gondii B1 gene (indicating the presence of oocysts) using nested-PCR. Fourteen (4.7%) of 300 cats examined were positive for B1 gene. Female cats (7.5%) showed a higher prevalence than male cats (1.4%). Cats younger than 3 months (5.5%) showed a higher prevalence than cats (1.5%) older than 3 months. For laboratory passage of the positive samples, the fecal suspension (0.2 ml) of B1 gene positive cats was orally inoculated into experimental mice. Brain tissues of the mice were obtained after 40 days and examined for the presence of tissue cysts. Two isolates were successfully passaged (designated KNIH-1 and KNIH-2) and were molecularly analyzed using the SAG5D and SAG5E gene sequences. The SAG5D and SAG5E gene sequences showed high homologies with the ME49 strain (less virulent strain). The results indicated the importance of stray cats in transmitting T. gondii to humans in Korea, as revealed by detection of B1 gene in fecal samples. T. gondii isolates from cats were successfully passaged in the laboratory for the first time in Korea.     

Trichomonas vaginalis: Investigation of a novel diagnostic method in urine samples using cysteine proteinase 4 gene and PCR technique

Trichomonas vaginalis is the agent of a highly prevalent sexually transmitted disease that leads to vaginitis, urethritis, ectocervicitis and has been associated with human immunodeficiency virus (HIV). Detection of T. vaginalis based on wet-mount microscopy and culture methods is insensitive and time consuming, respectively. Thus the quest for reliable PCR techniques of T. vaginalis in vaginal discharge and urine sample is more importance. In this study, 500 urine and vaginal-discharge samples were collected from women referred to Sexual Transmitted Disease Clinic of Mirzakuchakkhan Hospital in Tehran, Iran between May 2008 and March 2009. Wet-mount and culture methods were done on the vaginal discharges, and PCR assay targeting cysteine proteinase 4 (CP4) was performed on the urine samples. The present study demonstrated 16 (3.2%) of patients were infected with T. vaginalis using culture and wet-mount, whereas PCR assay using CP4 could detect 12 (2.4%) positivity. Sensitivity and specificity of urine PCR assay compared to culture were 80% (95% CI, 54-96) and 99.6% (95% CI, 98.96-100), respectively. These results indicate that using urine-based detection method for T. vaginalis may not be appropriate in women.     

Proinflammatory Cytokine and Nitric Oxide Production by Human Macrophages Stimulated with Trichomonas vaginalis

Trichomonas vaginalis commonly causes vaginitis and perhaps cervicitis in women and urethritis in men and women. Macrophages are important immune cells in response to T. vaginalis infection. In this study, we investigated whether human macrophages could be involved in inflammation induced by T. vaginalis. Human monocyte-derived macrophages (HMDM) were co-cultured with T. vaginalis. Live, opsonized-live trichomonads, and T. vaginalis lysates increased proinflammatory cytokines, such as TNF-α, IL-1β, and IL-6 by HMDM. The involvement of nuclear factor (NF)-κB signaling pathway in cytokine production induced by T. vaginalis was confirmed by phosphorylation and nuclear translocation of p65 NF-κB. In addition, stimulation with live T. vaginalis induced marked augmentation of nitric oxide (NO) production and expression of inducible NO synthase (iNOS) levels in HMDM. However, trichomonad-induced NF-κB activation and TNF-α production in macrophages were significantly inhibited by inhibition of iNOS levels with L-NMMA (NO synthase inhibitor). Moreover, pretreatment with NF-κB inhibitors (PDTC or Bay11-7082) caused human macrophages to produce less TNF-α. These results suggest that T. vaginalis stimulates human macrophages to produce proinflammatory cytokines, such as IL-1, IL-6, and TNF-α, and NO. In particular, we showed that T. vaginalis induced TNF-α production in macrophages through NO-dependent activation of NF-κB, which might be closely involved in inflammation caused by T. vaginalis.     

Influence of 120 kDa Pyruvate:Ferredoxin Oxidoreductase on Pathogenicity of Trichomonas vaginalis

Trichomonas vaginalis is a flagellate protozoan parasite and commonly infected the lower genital tract in women and men. Iron is a known nutrient for growth of various pathogens, and also reported to be involved in establishment of trichomoniasis. However, the exact mechanism was not clarified. In this study, the author investigated whether the 120 kDa protein of T. vaginalis may be involved in pathogenicity of trichomonads. Antibodies against 120 kDa protein of T. vaginalis, which was identified as pyruvate:ferredoxin oxidoreductase (PFOR) by peptide analysis of MALDI-TOF-MS, were prepared in rabbits. Pretreatment of T. vaginalis with anti-120 kDa Ab decreased the proliferation and adherence to vaginal epithelial cells (MS74) of T. vaginalis. Subcutaneous tissue abscess in anti-120 kDa Ab-treated T. vaginalis-injected mice was smaller in size than that of untreated T. vaginalis-infected mice. Collectively, the 120 kDa protein expressed by iron may be involved in proliferation, adhesion to host cells, and abscess formation, thereby may influence on the pathogenicity of T. vaginalis.     

The antimicrobial peptide,shrimp anti-lipopolysaccharide factor (SALF), inhibits proinflammatory cytokine expressions through the MAPK and NF-κB pathways in Trichomonas vaginalis adherent to HeLa cells

Trichomonas vaginalis is a parasitic protozoan that causes sexually transmitted infections (STIs) worldwide. The infection is dangerous and easily spreads within a community. Also, some cases of drug resistance were reported. Previously, we reported that the shrimp anti-lipopolysaccharide factor (SALF), an antimicrobial peptide of 24 amino acids, modulates inflammatory responses and inhibits T. vaginalis growth. To date, there is no report on the mechanism of SALF's actions in T. vaginalis’ adherence to HeLa cells. In this research using an ELISA, we found that the SALF downregulated the release of proinflammatory cytokines (tumor necrosis factor (TNF)-α, interleukin (IL)-1α, IL-6, IL-8, and monocyte chemoattractant protein (MCP)-1) secreted by T. vaginalis which was adhering to HeLa cells. We also performed real-time PCR experiments to determine the roles of the SALF in the expressions of several proinflammatory genes. Through a Western blot analysis, we determined that SALF treatment inhibited T. vaginalis-treated HeLa cells through the p38 and NF-κB pathways. Furthermore, we used different inhibitors to confirm the pathway by ELISA and Western blotting. Taken together, it is apparent that the SALF suppresses T. vaginalis-induced secretion of proinflammatory cytokines by HeLa cells by acting through the p38 and NF-κB pathways.     

First Report of Feline Intestinal Trichomoniasis Caused by Tritrichomonas foetus in Korea

Feline intestinal tritrichomoniasis by Tritrichomonas foetus was first recognized in USA in 1999 and has so far been reported from UK, Norway, Switzerland, and Australia, but not from the Far East Asian countries. In November 2008, 2 female and male littermate Siamese cats, 6-month old, raised in a household in Korea were referred from a local veterinary clinic with a history of chronic persistent diarrhea. A direct smear examination of fecal specimens revealed numerous trichomonad trophozoites which were isolated by the fecal culture in InPouch™ TF-Feline medium. A PCR testing of the isolate based on the amplification of a conserved portion of the T. foetus internal transcribed spacer (ITS) regions (ITS1 and ITS2) and the 5.8S rRNA gene, and the molecular sequencing of the PCR amplicons confirmed infection with T. foetus. This is the first clinical case of feline intestinal trichomoniasis caused by T. foetus in Korea.     

A Survey of Intestinal Protozoan Infections among Gastroenteritis Patients during a 3-Year Period (2004-2006) in Gyeonggi-do (Province), South Korea

The incidence and etiology of parasite-associated gastroenteritis during 2004-2006 in Gyeonggi-do (province), South Korea was determined by means of antigen detection ELISA on 6,071 stool specimens collected from 6 general hospitals. At least 1 parasitic agent was detected in 3.4% (208/6,071) of the stool samples. Among these, Giardia lamblia was the most numerous (152 cases; 2.5%), followed by Entamoeba histolytica (25 cases; 0.4%), Cryptosporidium parvum (23 cases; 0.4%), and mixed infections (8 cases; 0.1%). Patients aged 1-5 years had the largest proportion (69.2%; 144/208) of parasite-positive stool specimens. Parasite-mediated gastroenteritis was most common from June to September. The detection rate gradually increased from 2004 to 2006. This study shows that parasite-mediated gastroenteritis may be significant among children in Korea and that parasite infection surveillance should be constantly performed.     

Detection and Molecular Characterization of Cryptosporidium spp. from Wild Rodents and Insectivores in South Korea

In order to examine the prevalence of Cryptosporidium infection in wild rodents and insectivores of South Korea and to assess their potential role as a source of human cryptosporidiosis, a total of 199 wild rodents and insectivore specimens were collected from 10 regions of South Korea and screened for Cryptosporidium infection over a period of 2 years (2012-2013). A nested-PCR amplification of Cryptosporidium oocyst wall protein (COWP) gene fragment revealed an overall prevalence of 34.2% (68/199). The sequence analysis of 18S rRNA gene locus of Cryptosporidium was performed from the fecal and cecum samples that tested positive by COWP amplification PCR. As a result, we identified 4 species/genotypes; chipmunk genotype I, cervine genotype I, C. muris, and a new genotype which is closely related to the bear genotype. The new genotype isolated from 12 Apodemus agrarius and 2 Apodemus chejuensis was not previously identified as known species or genotype, and therefore, it is supposed to be a novel genotype. In addition, the host spectrum of Cryptosporidium was extended to A. agrarius and Crosidura lasiura, which had not been reported before. In this study, we found that the Korean wild rodents and insectivores were infected with various Cryptosporidium spp. with large intra-genotypic variationa, indicating that they may function as potential reservoirs transmitting zoonotic Cryptosporidium to livestock and humans.     

Fasciola hepatica in Snails Collected from Water-Dropwort Fields using PCR

Fasciola hepatica is a trematode that causes zoonosis mainly in cattle and sheep and occasionally in humans. Fascioliasis has been reported in Korea; however, determining F. hepatica infection in snails has not been done recently. Thus, using PCR, we evaluated the prevalence of F. hepatica infection in snails at 4 large water-dropwort fields. Among 349 examined snails, F. hepatica-specific internal transcribed space 1 (ITS-1) and/or ITS-2 markers were detected in 12 snails and confirmed using sequence analysis. Morphologically, 213 of 349 collected snails were dextral shelled, which is the same aperture as the lymnaeid snail, the vectorial host for F. hepatica. Among the 12 F. hepatica-infected snails, 6 were known first intermediate hosts in Korea (Lymnaea viridis and L. ollula) and the remaining 6 (Lymnaea sp.) were potentially a new first intermediate host in Korea. It has been shown that the overall prevalence of the snails contaminated with F. hepatica in water-dropwort fields was 3.4%; however, the prevalence varied among the fields. This is the first study to estimate the prevalence of F. hepatica infection using the vectorial capacity of the snails in Korea.