What was the main factor in successful control of ascariasis in Korea?

In the l950s, under the legacy of traditional agriculture, Ascaris lumbricoides, spread epidemically in the war-bitten society of Korea. Consensus on the parasite control was drafted in the Parasite Disease Prevention Act, which passed a parliamentary agreement in 1966, and established safe disposal of feces and mass chemotherapy as control strategies. Biannual stool examinations and treating infected schoolchildren were basic scheme of the control activity through which revenue could be secured for organized business. In the 27 years following 1969, a maximum of 16 million stool examinations had been done every year. Cellophane thick smear enabled the task. The infection declined remarkably in the 1970s when industrialization and green revolution proceeded. A population study of A. lumbricoides in the late 1970s helped us better understand its epidemiology. The data also settled down the understandable protest of teachers against the repeated stool examinations. In the 9 years following 1987, the target population was gradually reduced when the egg positive rate was below 0.1%. An article in the Korean Law, stipulating obligatory stool examinations, was made optional. Although the long-term Korean effort of Ascaris control was a success, the effect of mass chemotherapy was not as succinct in terms of lowering reinfection. In the period of control, Korean agricultural technology changed, and the economy grew and supplied sanitary facilities by which the vicious cycle was disconnected. Reduction of morbidity was a benefit of mass chemotherapy, which is the only control method feasible in economically difficult countries. The most important hurdle of parasite control in the 1960s was poverty of general population and limited financial resources in Korea but the society formed a consensus on the priority of intestinal helminthiasis control during the ordeal period. The national consensus in the 1960s was the critical milestone for Ascaris control in Korea. Under the social agreement, application of timely technical and research advancements in parasitology achieved the success of ascariasis elimination. The successful experience of ascariasis elimination in Korea can be a benchmark for countries where neglected tropical diseases are endemically recycled.

Effect of control strategies on prevalence, incidence and re-infection of clonorchiasis in endemic areas of China.

BACKGROUND: A pilot clonorchiasis control project was implemented to evaluate the efficacies of various chemotherapy strategies on prevalence, incidence and re-infection in Heilongjiang Province, China. METHODS AND FINDINGS: Seven intervention groups (14,139 residents, about 2000 in each group) in heavily or moderately endemic areas were subjected to repeated praziquantel administration from 2001 to 2004. In the selective chemotherapy groups, residents were examined for fecal eggs, and those who tested positive were treated with three doses of 25 mg/kg praziquantel at 5-hour-intervals in one day. However, all residents were treated in the mass chemotherapy groups. In heavily endemic areas, two mass treatments of all residents in 2001 and 2003 reduced the prevalence from 69.5% to 18.8%, while four annual mass treatments reduced the prevalence from 48.0% in 2001 to 8.4% in 2004. Selective annual treatments for egg-positive subjects reduced the egg-positive rates from 54.9% in 2001 to 15.0% in 2004 or from 73.2% in 2001 to 12.3% in 2004. Selective treatments every 6 months significantly reduced the prevalence from 59.5% in 2001 to 7.5% in 2004. All of the repeated treatments reduced EPG (eggs per gram of feces) significantly. The annual mass treatment and selective treatment every 6 months produced lower prevalence and re-infection rates and higher egg reduction rate than annual selective treatments did. In the moderate endemic areas, egg positive rates were 24.8% and 29.7% in 2001 but were 1.9% and 1.3% after 2 or 3 selective treatments. The prevalence, incidence, re-infection rates in a moderately endemic area were significantly lower than those of heavy endemic areas. CONCLUSIONS: Repeated mass treatment or selective treatment with praziquantel every 6 to 12 months is highly effective for clonorchiasis control in heavily endemic areas. In contrast, one or two selective treatments with health education is effective in moderately endemic areas.

Fifty years of the Korean Society for Parasitology.

In 1959, the Korean Society for Parasitology was founded by clinical scientists, specialists of public health, and 5 core parasitologists with experience in American science and medicine. The Society this year celebrates its 50th anniversary. Due to public health importance at the time of foundation, medical parasitology was the main stream for next 3 decades. Domestic problems of niche parasitic diseases, unlisted in 6 tropical diseases of major importance, had been studied by own efforts. To cope with the demand of parasite control, evaluation system for control activity was built up. Control activity against soil-transmitted nematodes, conducted for almost 3 decades, was evaluated as a success. Evaluation of praziquantel efficacy for clonorchiasis, paragonimiasis, and neurocysticercosis, population dynamics of Ascaris lumbricoides infection in a situation of continuous reinfections, diagnostic modalities of antibody tests combined with brain imaging developed for helminthiasis of the central nervous system and researches on intestinal trematodes were achievements in the first 30 years. During the recent 2 decades, science researches, such as cell and molecular biology of parasites and immunology of parasitic infections have been studied especially on parasitic allergens and proteolytic and anti-oxidant enzymes. Experiences of international cooperation for world health have been accumulated and would be expanded in the future.

PwRn1, a novel Ty3/gypsy-like retrotransposon of Paragonimus westermani: molecular characters and its differentially preserved mobile potential according to host chromosomal polyploidy.

BACKGROUND: Retrotransposons have been known to involve in the remodeling and evolution of host genome. These reverse transcribing elements, which show a complex evolutionary pathway with diverse intermediate forms, have been comprehensively analyzed from a wide range of host genomes, while the information remains limited to only a few species in the phylum Platyhelminthes. RESULTS: A LTR retrotransposon and its homologs with a strong phylogenetic affinity toward CsRn1 of Clonorchis sinensis were isolated from a trematode parasite Paragonimus westermani via a degenerate PCR method and from an insect species Anopheles gambiae by in silico analysis of the whole mosquito genome, respectively. These elements, designated PwRn1 and AgCR-1 - AgCR-14 conserved unique features including a t-RNATrp primer binding site and the unusual CHCC signature of Gag proteins. Their flanking LTRs displayed >97% nucleotide identities and thus, these elements were likely to have expanded recently in the trematode and insect genomes. They evolved heterogeneous expression strategies: a single fused ORF, two separate ORFs with an identical reading frame and two ORFs overlapped by -1 frameshifting. Phylogenetic analyses suggested that the elements with the separate ORFs had evolved from an ancestral form(s) with the overlapped ORFs. The mobile potential of PwRn1 was likely to be maintained differentially in association with the karyotype of host genomes, as was examined by the presence/absence of intergenomic polymorphism and mRNA transcripts. CONCLUSION: Our results on the structural diversity of CsRn1-like elements can provide a molecular tool to dissect a more detailed evolutionary episode of LTR retrotransposons. The PwRn1-associated genomic polymorphism, which is substantial in diploids, will also be informative in addressing genomic diversification following inter-/intra-specific hybridization in P. westermani populations.

Hepatic visceral larva migrans of Toxocara canis: CT and sonographic findings.

OBJECTIVE: The aim of this study was to describe the CT and sonographic findings of hepatic visceral larva migrans of Toxocara canis. MATERIALS AND METHODS: Fifty-four patients (44 men, 10 women; age range, 30-80 years; mean age, 53 years) with serologically confirmed visceral larva migrans of Toxocara canis underwent evaluation of the liver with CT (n = 25), sonography (n = 48), or both. Two radiologists used consensus for retrospective evaluation of CT and sonographic findings. Correlation between the presence and severity of hepatic abnormalities on images and the degree of peripheral eosinophilia was assessed. RESULTS: Seventeen (68%) of 25 patients who underwent CT had single or multiple ill-defined, oval or elongated, small, low-attenuating lesions in the liver. Eighteen (38%) of 48 patients who underwent sonography had single or multiple small, poorly defined, oval or elongated, hypoechoic scattered focal lesions in the liver. In the 19 patients who underwent both CT and sonography, the two techniques had no significant difference in rate of detection of hepatic lesions (p = 0.375, McNemar test). The lesion numbers on CT and sonography showed excellent linear correlation (r = 0.844, p = 0.001) by Pearson's correlation test. An independent samples t test showed that eosinophil count and percentage in the peripheral blood were significantly higher in patients with hepatic lesions on CT and sonography than in patients without lesions. CONCLUSION: CT and sonographic findings of hepatic visceral larva migrans of T. canis are multiple, ill-defined, oval or elongated, small, nodular lesions scattered in the liver parenchyma. The presence of hepatic lesions on images was associated with higher peripheral eosinophil count and percentage.

Critical roles for excretory-secretory cysteine proteases during tissue invasion of Paragonimus westermani newly excysted metacercariae.

Paragonimus westermani is a trematode parasite, which causes pulmonary and/or extrapulmonary granulomatous disease in humans. Successful invasion of the host tissue is critical for the survival of this tissue-invasive parasite. The enzymatic hydrolysis of host proteins is clearly a prerequisite of this process. In this study, we have investigated the functional roles of the excretory-secretory cysteine proteases of P. westermani newly excysted metacercariae (PwNEM) in tissue invasion. The 27 and 28 kDa enzymes (PwMc27 and PwMc28) purified from PwNEM excretory-secretory products (ESP), preferentially degraded fibrillar proteins, but not globular proteins. PwMc28 significantly facilitated the invasion of PwNEM into mouse peritoneum, whereas a diffusible cysteine protease inhibitor, trans-epoxysuccinyl-L-leuciloamido-(4-guanidino) butane (E-64) inhibited this process dose-dependently. Two distinct isoforms of PwMc28 (PwMc28a and PwMc28b), which exhibited two amino acid differences in their mature domains, were identified by tandem mass spectrometry and sequence analysis. Both enzymes were localized at the tegument on the anterior border and on the oral sucker, which suggests excretion-secretion via exocytosis or via the excretory canal network. The mRNA transcripts of PwMc28a and b were expressed abundantly during the active invasion/migration through the host's tissues, suggesting their relevant function to tissue invasion/migration in the definitive host.

Identification of immunodominant excretory-secretory cysteine proteases of adult Paragonimus westermani by proteome analysis.

Paragonimus westermani causes inflammatory lung disease in humans. The parasite excretes a host of biologically active molecules, which are thought to be involved in pathophysiological and immunological events during infection. Analyses of the 2-DE protein profiles of the excretory-secretory products (ESP) of adult P. westermani revealed approximately 147 protein spots, at least 15 of which were identified as cysteine proteases (CPs), at pHs between 4.5 and 8.5, and molecular weights (MWs) between 27 and 35 kDa. An additional three CPs (designated as PwCP-3, -8 and -11) were newly recognized by TOF/TOF MS. Their molecular biological information, which shared a high level sequence homology, was elucidated. The majority of the CPs reacted strongly with sera from paragonimiasis patients. When we observed the chronological changes in the antibody responses of the respective CPs against canine sera collected serially at 1, 3, 5, 7, 11 and 14 wk after experimental infection, these molecules exhibited a multiplicity of distinct immune recognition patterns. Our results clearly showed that P. westermani adult ESP were principally composed of excretory-secretory CPs, and that these CPs may exert effects not only on host tissue degradation and nutrient uptake, but also on the immune-regulating cells via synergistic and independent interactions.

A seroepidemiological survey of Taenia solium cysticercosis in Nabo, Guangxi Zhuang Autonomous Region, China.

We have observed the seropositive rate of Taenia solium cysticercosis in residents at Nabo Village, Tiandong County, Guangxi Zhuang Autonomous Region, China by enzyme linked immunosorbent assay. The village had been found to be a relatively high endemic area of porcine cysticercosis among roaming pigs. Of 202 persons examined four males aged 15, 25, 35 and 41 year-old exhibited absorbance (abs) at 0.18, 0.20, 0.35 and 0.55, respectively. In addition, two females whose ages were 35 and 39 years revealed specific antibody levels of abs 0.26 and 0.41 in their sera. Overall positive rate among the people was 2.97%. All of these persons agreed that they had ingested the pork infected with T. solium metacestode (TsM), while history of proglottid discharge was not noticed from all of them. Three males and one female complained of intermittent headache. Our findings reinforced not only that the prevalence of cysticercosis might be related with roaming pigs infected with TsM but also that behavioral and environmental practices in local community constituted risk factors for transmission of the infection.

Correlation between sonographic findings and infection intensity in clonorchiasis.

This prospective study was performed to evaluate the diagnostic usefulness of sonography in terms of the intensity of Clonorchis sinensis infection. Total 1,384 residents were subjected to this study at an endemic area in China, in which a clonorchiasis control program had been performed. History taking, fecal examination, and sonography were performed, and 87.8% of the subjects were found to be egg positive. Sonography showed intrahepatic bile duct dilatation (IHDD) in 68.4% of the subjects, increased periductal echogenicity (IPDE) in 24.5%, and gallbladder sludge (GBS) in 20.8%. Moreover, the grades of these sonographic findings were higher in subjects with heavier infections as determined by egg counts. IHDD was found to be the most accurate sonographic indicator of any infection degree (76.3), whereas IPDE and GBS were useful only in cases of heavy infection. Subjective symptoms were found to be only poorly correlated with sonographic findings or infection intensity.

Evaluation of Clonorchis sinensis recombinant 7-kilodalton antigen for serodiagnosis of clonorchiasis.

The diagnostic applicability of the Clonorchis sinensis recombinant 7-kDa protein was evaluated. In enzyme-linked immunosorbent assays and immunoblots, the protein showed high sensitivities (81.3 and 71.9%, respectively) and specificities (92.6 and 89.7%, respectively) for sera obtained from various helminthic infections. Some paragonimiasis sera showed cross-reactions. The antigen might be valuable in the serodiagnosis of human clonorchiasis.

Sonographic findings of active Clonorchis sinensis infection.

PURPOSE: The aim of this study was to document the characteristic sonographic findings of clonorchiasis for the diagnosis of active infection in an endemic area. METHODS: In a village in northeastern China, residents underwent fecal examinations for detection of Clonorchis sinensis eggs. Shortly thereafter, residents were examined with abdominal sonography. An experienced radiologist performed the sonographic examinations and analyzed the findings. Subjects whose fecal examinations were positive for eggs were considered to have active clonorchiasis; those whose examinations were negative for eggs were used as control subjects. The distinguishing sonographic features of active clonorchiasis were identified by stepwise logistic regression analysis. RESULTS: The study population comprised 457 subjects; fecal examinations revealed C. sinensis eggs in 316 and no eggs in 141. Four sonographic findings distinguished subjects with active clonorchiasis from control subjects: increased periductal echogenicity (p < 0.001; R = 0.11; sensitivity, 35%; specificity, 91%), floating echogenic foci in the gallbladder (p < 0.001; R = 0.09; sensitivity, 28%; specificity, 94%), diffuse dilatation of the intrahepatic bile ducts (p < 0.01; R = 0.03; sensitivity, 67%; specificity, 48%), and gallbladder distention (p < 0.05; R = 0.02; sensitivity, 3%; specificity, 100%), in decreasing order of significance. Among these 4 sonographic findings, increased periductal echogenicity and floating echogenic foci in the gallbladder were more significantly associated with active infection than were the other 2. CONCLUSIONS: Increased periductal echogenicity and floating echogenic foci in the gallbladder were identified as the 2 most significant findings for the sonographic diagnosis of active C. sinensis infection.

Prevalence of intestinal parasite infections on a national scale among primary schoolchildren in Laos.

In order to investigate the epidemiological situation of intestinal parasite infections in Laos, parasitological surveys were carried out on a national scale including 17 provinces and the Vientiane Municipality. A total of 29,846 stool specimens were collected from primary schoolchildren from May 2000 to June 2002 and examined once with the cellophane thick smear technique. The cumulative egg positive rate for intestinal helminths was 61.9%. By species, the rate for Ascaris lumbricoides was 34.9%, hookworm 19.1%, Trichuris trichiura 25.8%, Opisthorchis viverrini 10.9%, Taenia spp. 0.6% and Hymenolepis spp. 0.2%. The northern mountainous regions such as Phongsaly, Huaphan or Saysomboune Province showed a higher prevalence (over 70%) of soil- transmitted helminths. The regions along the Mekong River such as Khammuane, Saravane or Savannakhet Province showed a higher prevalence (over 20%) of fish-borne parasites. On the other hand, Schistosoma mansoni eggs were detected in 1.7% of schoolchildren only in Champassak Province, a previously endemic area. The highest prevalence was noted in Phongsaly Province (96.0%) and the lowest in Bolikhamxay Province (27.5%). An additional small-scale survey by cellophane anal swab detected Enterobius vermicularis eggs in 35.7% of 451 schoolchildren aged 6-8 years in Khammuane, Vientiane, Champassak Province and the Vientiane Municipality. Meanwhile, the mean blood haemoglobin level of hookworm-infected children was not lower than that of children not infected with hookworm, suggesting that nutritional factors are more important than parasite infection per se. Nevertheless, the above results indicate that a nationwide parasite control project is necessary to reduce possible morbidity due to parasitic diseases in the country.

Taenia solium: identification of specific antibody binding regions of metacestode 10-kDa protein.

Taenia solium neurocysticercosis (NCC) represents one of the major public health problems associated with several neurological manifestations worldwide. We previously identified a recombinant 10-kDa protein of T. solium metacestode (CyDA) specific to active NCC. Immunoblottings with sera from active NCC patients and from animals experimentally infected with larval T. solium (pig), T. saginata (pig), T. asiatica (pig), and T. crassiceps (mouse) strongly recognized CyDA, while sera from patients infected only with adult worms did not. Mapping of antigenic sites using deletion mutants revealed that amino acids (aa) residues 30-34, Asn-Met-Thr-Val-Met (NMTVM), reacted only with sera from active stage T. solium cysticercosis cases. Recognition of CyDA aa 30-34 resided almost exclusively in the IgG4 isotype. Competitive immunoprecipitation with synthetic peptides confirmed the specificity of anti-sera for this penta-peptide. These results demonstrated that aa residues NMTVM in CyDA comprise the core sequence for an active stage NCC-related antigenic determinant. ligand binding protein, HLBP; Cyst fluid, CF; Pooled serum of 10 active NCC patients, serum-pool.

Excretory bladder: the source of cysteine proteases in Paragonimus westermani metacercariae.

The cysteine proteases of Paragonimus westermani metacercariae are involved in metacercarial excystment, host immune modulation, and possibly in tissue penetration. In order to clarify the origin of the enzymes, 28 and 27 kDa cysteine proteases in metacercarial excretory-secretory products were purified through the FPLC system using Mono Q column chromatography. The polyclonal antibodies to the enzymes were produced in BALB/c mice. Immunolocalization studies revealed that both cysteine proteases were distributed at the linings of excretory bladder and excretory concretions of the metacercariae. It was suggested that the excretory epithelium of P. westermani undertake the secretory function of metacercarial cysteine proteases, in addition to its role as a route for eliminating waste products.

Recombinant Paragonimus westermani yolk ferritin is a useful serodiagnostic antigen.

A recombinant protein of Paragonimus westermani yolk ferritin was bacterially produced from a previously cloned complementary DNA and was used as an antigen for an enzyme-linked immunosorbent assay (ELISA) against paragonimiasis- and other helminth-infected sera to evaluate its serodiagnostic potential. The ELISA revealed that paragonimiasis westermani had 88.2% sensitivity and 100% specificity. The positive and negative predictive values of the ELISA were calculated to be 100% and 97.1%, respectively. Sera from cats experimentally infected with P. westermani began to produce immunoglobulin G antibodies against the yolk ferritin at 13 weeks after infection, which suggests that the corresponding antigen was derived from the vitellaria in accordance with maturation of P. westermani. These results indicate that the recombinant P. westermani yolk ferritin is a potent serodiagnostic reagent for paragonimiasis westermani from an early stage of the infection.

Clonorchis sinensis: immunolocalization of 26 kDa glutathione S-transferase in adult worms.

A mu-class glutathione S-transferase (Cs26GST) of molecular mass 26 kDa was characterized from Clonorchis sinensis. In adult C. sinensis, the distribution of the Cs26GST was investigated by immuno-histochemistry and electron microscopy. Cs26GST was localized to the tegument and parenchyma. Immunogold labeling was strong in the tegumental cell bodies and moderate in the tegument and ova in the oviduct. It is suggested that Cs26GST plays a role in the metabolism and fecundity of C. sinensis.

Paragonimus westermani: molecular cloning, expression, and characterization of a recombinant yolk ferritin.

Ferritin is an intracellular protein involved in iron metabolism. A cDNA PwYF-1 cloned from the adult Paragonimus westermani cDNA library encoded a putative polypeptide of 216 amino acids homologous with ferritins of vertebrates and invertebrates. Febinding motifs identified in PwYF-1 polypeptide were conserved and predicted to form a ferroxidase center. PwYF-1 polypeptide contained an extended peptide of 45 amino acids at its C-terminus. Recombinant PwYF-1 protein, expressed and purified from Escherichia coli, showed iron-uptake ability and ferroxidase activity. Ferroxidase activity of recombinant PwYF-1 protein was reactivated by secondary addition of apotransferrin to assay mixture. Mouse immune serum raised against the recombinant PwYF-1 protein recognized specifically 24 kDa protein from adult P. westermani lysate. PwYF-1 protein was localized to vitelline follicles and the eggs of P. westermani. Collectively, PwYF-1 protein was identified as a P. westermani yolk ferritin.

Immunoblot observation of antigenic protein fractions in Paragonimus westermani reacting with human patients sera.

In order to observe the antigenic fractions in saline extract of adult Paragonimus westermani, proteins in the crude extract were separated by sodium dodecyl sulfate-polyacylamide gel electrophoresis (SDS-PAGE) in reducing conditions. The separated protein fractions were transferred to nitrocellulose paper on which 20 sera from human paragonimiasis were reacted and immunoblotted. Out of 15 stained protein bands in SDS-PAGE, 7 reacted with the sera. Of 14 reacted bands, 30 kilodalton(kDa) band was the most frequently reacted (95%) and was a strong antigen. Protein bands of 23 and 46 kDa were also strong antigens. Bands of over 150 kDa, 120 kDa, 92 kDa, 86 kDa, 74 kDa, 62 kDa, 51 kDa, 32 kDa, 28 kDa, 16.5 kDa and 15.5 kDa were also reactive but their frequencies of the reaction were variable.

Antigenic protein fractions reacting with sera of sparganosis patients.

To observe the antigenic protein fractions in saline extract of Spirometra mansoni plerocercoid (sparganum), the crude extract was separated in reducing conditions of sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The proteins, transferred by electrophoresis to nitrocellulose paper, were reacted with sera from 15 surgically confirmed sparganosis and 24 cysticercosis patients for immunoblotting. Out of 30 identified protein bands in the extract, bands of 29 and 36 kilodaltons (kDa) were the strongest and the most frequently reacting with specific antibody (IgG) in sparganosis sera. Bands of higher molecular weight also reacted with the sera but their frequency of reactions was lower. Sera of cysticercosis reacted with different protein bands in saline extract of sparganum, but the cross reactions were observed in strong antigenic bands of 29 and 36 kDa.