Genotyping of Giardia lamblia isolates from humans in China and Korea using ribosomal DNA Sequences.

Genetic characterization of a total of 15 Giardia lamblia isolates, 8 from Anhui Province, China (all from purified cysts) and 7 from Seoul, Korea (2 from axenic cultures and 5 from purified cysts), was performed by polymerase chain reaction amplification and sequencing of a 295-bp region near the 5' end of the small subunit ribosomal DNA (eukaryotic 16S rDNA). Phylogenetic analyses were subsequently conducted using sequence data obtained in this study, as well as sequences published from other Giardia isolates. The maximum parsimony method revealed that G. lamblia isolates from humans in China and Korea are divided into 2 major lineages, assemblages A and B. All 7 Korean isolates were grouped into assemblage A, whereas 4 Chinese isolates were grouped into assemblage A and 4 into assemblage B. Two Giardia microti isolates and 2 dog-derived Giardia isolates also grouped into assemblage B, whereas Giardia ardeae and Giardia muris were unique.

Infectivity and pathological changes in murine clonorchiasis: comparison in immunocompetent and immunodeficient mice.

The main complications of clonorchiasis are periportal inflammation, biliary hyperplasia, periductal fibrosis, and subsequently the development of biliary tumors in the liver. This study was undertaken to compare the infectivity and histopathologic changes between in immunocompetent FVB/NJ and BALB/cA strains, and immunodeficient severe combined immunodeficient (SCID) and athymic nude mice after the metacercariae of Clonorchis (C.) sinensis were infected. The experiment showed that C. sinensis was very infective in all strains studies, but the status of worm development, infectivity, recovery rate, and morphological changes of livers were very different in each strain. FVB/NJ mice showed more worm recovery than any other strain. Histopathologically the liver of FVB/NJ mice at 4 weeks postinfection showed marked cystic and fibrotic changes, in which C. sinensis was fully developed with ovum production, severe infiltration of inflammatory cells, mostly eosinophils, and high degrees of biliary hyperplasia. In SCID and nude mice, there were few foci of inflammatory cells even at 8 weeks postinfection in periportal areas of the liver, associated with no development into adult worm with ovum production. Fibrosis occurring at 4 weeks postinfection was highly correlated with inflammatory infiltration when each strain was compared. We suggest that massive infiltration of eosinophil and plasma cells caused by the infection might initiate cystic formation and fibrosis. These data demonstrate that the infection of C. sinensis might be related to pathologic consequences of inflammatory cell infiltration, cystic formation and fibrosis which might play a role in the defense mechanism against the parasitism in the liver of each strain. The FVB/NJ mouse model might be very helpful in elucidating the mechanism for human clonorchiasis.

Chemotherapeutic efficacy of praziquantel in rats with protective immunity to Clonorchis sinensis infection.

OBJECTIVE: To study the synergic effect of praziquantel (PZQ) and host acquired immunity on Clonorchis sinensis. METHODS: Acquired immunity to C. sinensis was induced by immunization with crude adult worm antigen (AW Ag) and excretory-secretory antigen (ES Ag) or infection with C. sinensis metacercariae. The effect was assessed by the worm reduction rate compared with the control groups after challenge infection with 50 metacercariae and treated orally with a subcurative dose of praziquantel (50 mg/kg). Significant test was performed by analysis of variance (ANOVA) and Npar1 way Kruskal-Wallis test. All calculations were performed by PC-SAS system. RESULTS: 1. PZQ was more effective against C. sinensis larvae than against adult worms in the control (P < 0.001), ES Ag (P < 0.01) or crude AW Ag immunization group (P < 0.001). 2. As compared with the control, the worm reduction rate after challenge infection was significantly higher (P < 0.001) in ES Ag immunized group (35.6%) and metacercaria infection group (97.5%) and less in crude AW Ag group (23.4%). The PZQ efficacy was significantly enhanced in ES Ag immunized group. CONCLUSION: The efficacy of PZQ against C. sinensis could be synergically enhanced in rats by inducing host acquired immunity.

Clonorchiasis: an update.

Clonorchis sinensis, the Chinese or oriental liver fluke, is an important human parasite and is widely distributed in southern Korea, China (including Taiwan), Japan, northern Vietnam and the far eastern part of Russia. Clonorchiasis occurs in all parts of the world where there are Asian immigrants from endemic areas. The human and animal reservoir hosts (dogs, pigs, cats and rats) acquire the infection from the ingestion of raw fish containing infectious metacercariae. The first intermediate snail hosts are mainly species of Parafossarulus and Bithynia. Numerous species of freshwater fish serve as the second intermediate hosts of C. sinensis. Extensive studies of clonorchiasis during several decades in Japan, Korea, China and other countries have shown much progress in proving its morphological features including ultrastructure, biology, pathogenesis, epidemiology, clinical manifestations and chemotherapy. The present review deals with mainly current results obtained on the epidemiological, pathological and clinical aspects, as well as control measures in endemic areas. As for the complications of clonorchiasis, formation of calculi in the intrahepatic biliary passages is one of the most characteristic pathological features. It is sometimes accompanied by suppurative cholangitis, cholecystitis, cholangiohepatitis and ultimately can cause cholangiocarcinoma. Experimental results on the relationship to the occurrence of cholangiocarcinoma are presented. Clinical diagnosis by radiological findings including cholangiography, sonography and computerized tomography as well as magnetic resonance imaging for biliary or pancreatic ducts are outlined. Current studies on immunology and molecular biology of C. sinensis were introduced. Praziquantel is the drug of choice for clonorchiasis. The most effective regimen is 25 mg kg(-1) three times daily (total dose, 75 mg kg(-1)) administered orally at 5- to 6-h intervals over a single day. Prevention and control measures are also discussed.

Mixed infections with Opisthorchis viverrini and intestinal flukes in residents of Vientiane Municipality and Saravane Province in Laos.

Faecal examinations for helminth eggs were performed on 1869 people from two riverside localities, Vientiane Municipality and Saravane Province, along the Mekong River, Laos. To obtain adult flukes, 42 people positive for small trematode eggs (Opisthorchis viverrini, heterophyid, or lecithodendriid eggs) were treated with a 20-30 mg kg(-1) single dose of praziquantel and purged. Diarrhoeic stools were then collected from 36 people (18 in each area) and searched for helminth parasites using stereomicroscopes. Faecal examinations revealed positive rates for small trematode eggs of 53.3% and 70.8% (average 65.2%) in Vientiane and Saravane Province, respectively. Infections with O. viverrini and six species of intestinal flukes were found, namely, Haplorchis taichui, H. pumilio, H. yokogawai, Centrocestus caninus, Prosthodendrium molenkampi, and Phaneropsolus bonnei. The total number of flukes collected and the proportion of fluke species recovered were markedly different in the two localities; in Vientiane, 1041 O. viverrini (57.8 per person) and 615 others (34.2 per person), whereas in Saravane, 395 O. viverrini (21.9 per person) and 155207 others (8622.6 per person). Five people from Saravane harboured no O. viverrini but numerous heterophyid and/or lecithodendriid flukes. The results indicate that O. viverrini and several species of heterophyid and lecithodendriid flukes are endemic in these two riverside localities, and suggest that the intensity of infection and the relative proportion of fluke species vary by locality along the Mekong River basin.

Paragonimiasis: experimental and clinical experience with praziquantel in Korea.

The effect of praziquantel (2-cyclohexylcarbonyl-1,2,3,6,7,11b-hexahydro-4H-pyrazino[2,1-a]++ +isoquinolin- 4-one, EMBAY 8440, Biltricide) on the ultrastructure of Paragonimus westermani was investigated in vitro and in vivo. In in vitro experiments, P. westermani was less sensitive to praziquantel. Only very few vacuoles were formed in the tegumental syncytium after incubation in 100 micrograms/ml for 60 min. However, in vivo the parasite was successfully destroyed by praziquantel. In fine structural changes of P. westermani obtained from experimentally infected dogs after treatment, bleb-like structures were formed at the papillae near the two suckers and bubble-like vacuoles and vesicles in the tegumental syncytium were observed along the basement membrane being detached from the circular muscles of the worm. It was also observed that many host cells invaded the exposed muscular layer of the worm after exfoliation of the tegument and the interior of the parasite was heavily damaged by those host cells. In our clinical trials, complete cure was obtained in 10 patients given praziquantel at a dosage of 3 X 25 mg/kg body weight for 3 consecutive days and with a dosage of 3 X 25 mg/kg for 2 consecutive days, 18 (85.7%) of 21 patients were cured. On the other hand, the dosage of 3 X 25 mg/kg on one day cured 15 (71.4%) out of 21 patients. Nine uncured cases were treated again with dosages of 3 X 25 mg/kg for 2 or 3 consecutive days. Only two out of nine failed to be completely cured and a very small number of eggs were detected again in their sputum and faeces.(ABSTRACT TRUNCATED AT 250 WORDS)

Health education in control of fluke infections.

Human trematode infections are largely confined to the tropical and oriental countries. Especially schistosomiasis, clonorchiasis, paragonimiasis, heterophydiasis and fasciolopsiasis are great public health problems in certain countries. The control of clonorchiasis, paragonimiasis and intestinal fluke infections theoretically is very simple, as the infection is transmitted to man mostly by eating fish and crabs serving as intermediate hosts and harbouring metacercariae. It is different with schistosomiasis where, by contact with contaminated water, man gets infected with cercarial larvae penetrating the intact skin. Therefore, prophylactic measures are far more difficult to apply successfully. Thus, except in schistosomiasis, the most practical method of preventing human trematode infections is to avoid eating raw, freshly pickled or imperfectly cooked fresh-water fish, crayfish or crabs, etc. It is, however, exceedingly difficult to enforce such simple measures in face of century old traditions, to which relatively primitive populations cling with great tenacity. Therefore, in areas where the consumption of raw fish or crabs is customary, insistent and persistent educational propaganda stressing the importance of thoroughly cooking all fresh-water fish or crabs appears to be the most effective means of preventing such diseases. Such educational efforts should be directed primarily toward school children because they are less entrenched in their food habits. Thus, the educational process could involve various levels to successively change knowledge, attitudes, behaviour, habits and customs of their lives. However, this could better be accomplished by supporting health education by successful chemotherapy.

Clonorchiasis in Korea.

Since the first report on the incidence of egg positiveness of Clonorchis sinensis by Matsumoto in 1915, clonorchiasis has been recognized as one of the most important endemic diseases in Korea. Most of the works on clonorchiasis after then had been studied by Japanese workers until 1945. After the Korean War in 1950-1953, health problems of Koreans were extremely aggravated. This gave us attentions for the prevention of contagious diseases as well as parasitic infections in public health point of view. It was greatly recognized that not only ascariasis, hookworm infections, and trichuriasis, but also clonorchiasis and paragonimiasis constitute the important helminthic diseases in Korea. In the late 1950s, several Korean workers conducted studies of C. sinensis on its biology, epidemiology, pathology, clinical symptoms as well as treatment. Thereafter overall epidemiological surveys were carried out actively throughout the country since 1960s. From these surveys, it became clear that endemic areas of clonorchiasis are scattered all over the country along the Nakdong, Kum, Mankyong, Yeongsan and Han rivers. The most extensive and intensive endemic regions were found mainly along the Nakdong river. Since then, clonorchiasis has been one of the most extensively studied subjects during the past 30 years. The present review mainly deals with current results obtained by Korean investigators on the prevalence of clonorchiasis, as well as its biology, epidemiology, pathogenesis, clinical manifestations, diagnosis, treatment, prevention and control.

Therapy of fluke infections in the past. A review.

Chemotherapy of trematode infections has been studied intensively in the past. The compounds so far used for the treatment of those parasitic infections were not satisfactory, however, because of their poor tolerability and/or their limited anthelmintic efficacy. Since the introduction of tartar emetic in 1918, a number of antimonial as well as non-antimonial preparations have been used for the treatment of schistosomiasis. In the treatment of clonorchiasis with antimony preparations, gentian violet, emetine HCl, chloroquine diphosphate and dithiazanine iodide, only temporary clinical improvement and negative or reduced egg counts could be achieved, but complete cure was not obtained. Although Hetol (or hexachloroparaxylol), prolonged administration of oral dehydroemetine and only two doses of niclofolan showed therapeutic effect on Clonorchis infection, side effects were observed in most of the treated cases. On the other hand, bithionol proved to be a drug of choice for paragonimiasis. However, the need of multiple doses and the many adverse reactions hampered its use in the field. Recently the newly developed praziquantel (2-cyclohexylcarbonyl-1,2,3,6,7,11b-hexahydro-4H-pyrazino-[2,1-a]+ ++isoquinolin -4-one, EMBAY 8440, Biltricide) proved to be safe and effective against trematodes and appears to be the drug of choice for infections due to all species of Schistosoma, Clonorchis sinensis, Opisthorchis viverrini, Paragonimus spp. and Metagonimus yokogawai in man.

Morphologic descriptions of Taenia asiatica sp. n.

Among taeniid tapeworms infecting humans through pork or beef, Taenia solium Linnaeus 1758 and Taenia saginata Goeze 1782 have already been known. Based on the morphologic characteristics of adult and metacestodes of Asian Taenia saginata, the third kind of human taeniid tapeworm known to distribute in Asian countries, a new species name of Taenia asiatica is proposed. In addition to the known biology in their intermediate hosts, T. asiatica was different morphologically from Taenia saginata Goeze 1782 in having the unarmed rostellum on the scolex of adult, the large number of 'uterine twigs' and the existence of 'posterior protuberance'. These structures in the gravid proglottids were used as taxonomic keys in taeniid tapeworms for the first time. T. asiatica metacestode (Cysticercus viscerotropica) was different morphologically from T. saginata metacestode (Cysticercus bovis) in having wartlike formations on the external surface of the bladder wall.

Clinical evaluation of the therapeutic efficacy of praziquantel against human cysticercosis.

Thirty-three patients with confirmed cysticercosis, with or without cerebral involvement, were treated with praziquantel at a daily dose of 75 mg/kg tid for 3 to 10 consecutive days and evaluated for tolerance and therapeutic effects. For dermal cysticercosis, a daily dose of 75 mg/kg tid for 3 days is effective. The nodules disappeared in one to 12 months after treatment. Twenty out of 33 patients with cysticercosis had cerebral cysticercosis. Among these 20 cases, 13 had epileptic seizures, 6 had a combination of both epileptic seizures and intracranial hypertension, and one had headache. The efficacy of the treatment of cerebral cysticercosis was assessed by the frequency of convulsions before and after treatment, and by the disappearance or decreased densities of the cystic lesions at brain CT scanning (6 months follow up). A daily dose of 75 mg/kg tid for 7 to 10 days (total 525 to 750 mg/kg) seems to be effective for the treatment of cerebral cysticercosis. The concomitant oral administration of dexamethasone during the course of treatment was effective for preventing and minimizing side effects. Praziquantel was very well tolerated.

Epidemiological understanding of Taenia tapeworm infections with special reference to Taenia asiatica in Korea.

In endemic areas of Taenia tapeworms in Korea, most of the reports showed that T. saginata was dominant over T. solium, but eating pigs is the dominant habit over eating cattle. Why do they have more T. saginata despite lower consumption of beef? This problem actually has long been recognized but until recently there has been no intensive trial to give a scientific explanation on this epidemiological enigma. By summing up the data published between the years 1963 and 1999, the ratio of armed versus unarmed tapeworms in humans was estimated at approximately 1:5. The ratio of pig-eaters versus cattle-eaters, however, was approximately 5:1. This inconsistency could be explained with the recently described T. asiatica, which infects humans through the eating of pig's viscera. We re-evaluate the importance of the consumption of visceral organ of pigs. leading us to an improved epidemiological understanding of the T. asiatica infection together with co-existing T. saginata and T. solium in Korea.

Natural infections of Asian Taenia saginata metacestodes in the livers of Korean domestic pigs.

The Taenia species in East Asia, hitherto identified as Taenia saginata, has been recognized as differing from the classical descriptions of life cycles and was recently named Taenia saginata taiwanensis (tentatively until 1992). Major differences between T. saginata and the newly recognized Asian T. saginata are their intermediate hosts and the infected tissues. Asian T. saginata metacestodes are found in the livers of pigs rather than in the muscles of cattle. In this study, we observed the natural infection status of Asian T. saginata metacestodes in the livers of 25,358 pigs at an abattoir in Cheongju City, Korea, from 1989 to 1990. Total 256 livers (1.01%) were infected with 1-96 (mean 2.5 per capita) metacestodes. Most of the metacestodes, however, were either calcified (87.1%) or highly degenerated (12.9%). Living metacestodes were found in only 0.01% (3/25, 358) of the examined livers. And these were distributed randomly in each lobe of the livers. The liver of pigs in Korea was confirmed as an organ hosting the Asian T. saginata metacestode. But its epidemiological significance as a source of human infections should be properly evaluated because of the rarity of the living metacestodes.

Experimental human infection with Asian Taenia saginata metacestodes obtained from naturally infected Korean domestic pigs.

The infectivity of metacestodes of Asian Taenia saginata, now tentatively called Taenia saginata taiwanensis, in human host was confirmed. The metacestodes used in experimental infection were collected from the livers of naturally infected domestic pigs at an abattoir in Cheongju City, Korea. The first gravid proglottid was spontaneously discharged 76 days after infection. Two worms were recovered two years later by chemotherapy. The scolex was unarmed. The number of main uterine branches, varying from 16 to 21, was similar to that of classical Taenia saginata. The liver of pigs was confirmed to be an infection source of Asian T. saginata in Korea.

Second case of human infection with Mesocestoides lineatus in Korea.

The second case of human infection with Mesocestoides lineatus in Korea was reported. The patient, a farm worker, complained of abdominal pain and massive discharge of sesame-like proglottids in his stool for several months. Worms, recovered by chemotherapy with niclosamide, consisted of 32 strobilae. This may be the heaviest worm burden in human infection ever reported. The infected man habitually ate the raw viscera of chickens.

Trematodes of the genus Haematoloechus (Digenea: Plagiorchiidae) from frogs in Korea.

Total 242 Rana nigromaculata and 9 R. catesbeiana were collected from the various localities in Korea from February 1989 to July 1991, and their lungs were examined. Five species in genus Haematoloechus, i.e., H. sibiricus japonicus (Yamaguti, 1936), H. nanchangensis Hsiung, 1934, H. variegatus (Rudolphi, 1819), H. lobatus (Seno, 1907), H. lobatus koreanus, were identified in this study. Among them, H. lobatus koreanus was a new subspecies and H. variegatus and H. lobatus were newly recorded from Korea.

Experimental infection of pigs and cattle with eggs of Asian Taenia saginata with special reference to its extrahepatic viscerotropism.

Asian Taenia saginata, tentatively called Taenia saginata taiwanensis, has been described to be infected in its metacestode stage only in the liver of intermediate host animals. Experimentally, however, we found that the metacestodes of the Asian Taenia saginata are also infected in other viscera than the liver of pigs (Landrace-Duroc-Hampshire) 4 days to 4 months postinoculation (PI). Viscerotropism of cysticercosis was apparent because a majority (70.7%) of the non-calcified cysticerci were found in the livers while a minority were found in extrahepatic organs such as the omentum (19.2%), lungs (8.1%) and serosa of colon (2.0%). When experimentally infected to cattle, Asian T. saginata cysticerci were also observed calcified in the livers. On the other hand, classical Taenia saginata metacestodes infected the muscles and viscera of the Holstein-Friesian cattle whereas no infection was observed in experimental pigs. Extrahepatic metacestodes of Asian T. saginata, which were obtained from an experimental pig were confirmed to be infective to a male volunteer. This extrahepatic viscerotropism of Asian T. saginata metacestodes in experimental pigs explains well the transmission modes of Asian T. saginata among people considering the eating habits.

[Antigenic localities in the tissues of Metagonimus yokogawai in the period of growth].

In order to observe the antigenic localization in the tissues of Metagonimus yokogawai in growth stages, immunogoldlabeling method was applied to using serum of the cat which infected with isolated metacercariae from Plecoglossus altivelis. The sectioned worm tissues from each growth stages were embedded in Lowicryl HM 20 medium, stained with infected serum IgG and protein A gold complex (particle size: 12 nm) and observed by electron microscopy. In the worm tissues of all experimental groups, the gold particles were specifically concentrated on the tegumental syncytium and cytoplasm of the tegumental cell as well as the secretory granules in the parenchymal tissue. In the 16th and 20th week grown worm tissues, the gold particles were specifically concentrated on the vesicles in the tegumental syncytium and cytoplasm of the tegumental cell. The gold particles were specifically concentrated on the caecal epithelia of the 4th, 8th and 12th week growth groups but slightly concentrated on those of the 16th and 20th week.