A human case of Plagiorchis vespertilionis (Digenea: Plagiorchiidae) infection in the Republic of Korea.

Plagiorchis vespertilionis (Digenea: Plagiorchiidae) is generally considered a bat parasite, but here it is reported for the first time in a human. The patient was a 34-yr-old male who lived in a coastal village of Haenam-gun (county), Jeollanam-do (province), Republic of Korea. Only 1 worm, 2.6 mm long and 0.7 mm wide, was recovered after praziquantel treatment and purging with magnesium salts. The fluke was characterized by a large body size, a sucker ratio of 1:1, a straight cirrus organ, a short distance between the ventral sucker and ovary, well-developed vitellaria, a uterus with descending and ascending loops, and fully developed eggs with an average size of 32.5 x 17.5 microm. The patient had habitually eaten the raw flesh of snakehead mullet and gobies that had been caught near his village. The present case represents the first record of a human P. vespertilionis infection.

Radioresistance of Anisakis simplex third-stage larvae and the possible role of superoxide dismutase.

The radioresistance of Anisakis simplex third-stage larvae and the possible role of sublethal radiation on superoxide dismutase (SOD) were investigated. Larvae were isolated from the viscera of the sea eel Anago anago; irradiated with 10, 100, 200, 500, or 1,000 Gy; and then given orally to rats. Worms were recovered at 16 hr postinoculation. Most larvae were found to have invaded the gastric wall, omentum, and abdominal cavity, suggesting that their viability and infectivity were not controlled by irradiation with the doses used. To determine the relationship between SOD activities in parasites and their radiosensitivities, the larvae of A. simplex and the metacercariae of Neodiplostomum seoulense (a radiosensitive control) were irradiated with 0, 30, 100, or 500 Gy, and parasite SOD levels were measured. In nonirradiated A. simplex larvae, the average SOD level was 38.9 U/mg, and this increased to 51.3 U/mg at 500 Gy. However, at all radiation doses applied, SOD activities of N. seoulense metacercariae were significantly (P < 0.05) lower than those of A. simplex larvae. Our results demonstrate that A. simplex third-stage larvae are radioresistant, and suggest that SOD plays a role in this radioresistance.

Suppressed cytokine and immunoglobulin secretions by murine splenic lymphocytes infected in vitro with Toxoplasma gondii tachyzoites.

Mechanisms of host immunosuppression after infection with Toxoplasma gondii are unclear. This study was performed to observe cytokine and immunoglobulin secretions by murine splenic lymphocytes infected in vitro with live, nonreplicating (irradiated) RH tachyzoites on stimulation with concanavalin A (Con A) or lipopolysaccharide (LPS). For lymphocyte cultivation, 3 groups were prepared: coculture with live nonirradiated tachyzoites separated by a transwell (group T), live irradiated tachyzoites without a transwell (group R), and no tachyzoites (group C). Compared with group T, groups R and C, on stimulation with Con A, revealed significantly (P < 0.05) lower levels of interleukin-2 (IL-2) and IFN-gamma, but not IL-10. The levels of IgG1, IgG2a, IgG2b, IgG3, IgA, and IgM were also significantly (P < 0.05) lower in groups R and C than in group T after stimulation with LPS. The results suggest that intracellular infection of murine splenic lymphocytes with T. gondii tachyzoites could impair their capacity to produce cytokine and immunoglobulin secretions.

A trial for a DNA diagnosis of Plasmodium vivax malaria recently reemerging in the Republic of Korea using microtiter plate hybridization assay.

The polymerase chain reaction-based microtiter plate hybridization (PCR-MPH) assay was utilized for a DNA diagnosis of Plasmodium vivax malaria, which has recently reemerged in the Republic of Korea. The subjects were 18 parasite-proven patients and 5 healthy controls. Follow-up blood samples were collected from 4 patients after a standard course of treatment. Polymerase chain reaction and electrophoresis of all the patients' blood showed a prominent band at the 138 base pair area, but not in the controls or after treating the patients. Hybridization of the PCR products with known species-specific probes of the 18S rRNA of various malaria species revealed strong positive reactions against the Plasmodium vivax-specific probe (absorbance 1.30-1.90 at 405 nm) in all of the patients. The absorbance was positively correlated with the degree of blood parasitemia, but with a borderline significance. Sequencing of the probe region of the Korean P. vivax revealed no significant variations from the typical P. vivax. The results show that the PCR-MPH is a highly useful technique for the DNA diagnosis of Korean vivax malaria.

High prevalence and seasonality of cryptosporidiosis in a small rural village occupied predominantly by aged people in the Republic of Korea.

The prevalence of Cryptosporidium parvum infection and its seasonality were studied in a small rural village occupied predominantly by aged people in Hwasun-gun, Chollanam-do, Republic of Korea. Fecal samples were collected monthly from November 1996 to October 1997 and examined for enteric parasites. To detect oocysts of C. parvum, the modified acid-fast stain was applied. To determine effects of climatic factors, local weather reports were collected. The overall oocyst prevalence during 1 year was 57.0% (77 of 135), and was significantly higher (P < 0.0001) in aged people--those aged 50-59 years (80.0%) and those aged 60-69 years (69.0%)--than in younger groups. No significant difference (P > 0.05) was noted between men and boys (53.6%) and women and girls (59.5%), but significant (P < 0.05) household clustering was recognized. People aged > 50 years tended to be more repeatedly positive than people in the younger group. The rate varied monthly from 7.8 to 20.0% (average, 12.6%), with significantly higher (P < 0.05) rates in spring than in summer and winter months. This seasonality had little relation with temperature (r = 0.04), but a weak correlation was found with rainfall (r = 0.4). It is of special interest that cryptosporidiosis is highly endemic among the aged population.

Role of intestinal goblet cells in the expulsion of Gymnophalloides seoi from mice.

Role of intestinal goblet cells (GCs) in the expulsion of Gymnophalloides seoi was studied using 4 strains of mice, ICR, C3H/ HeN, BALB/c, and C57BL/6, after infection with 200 metacercariae isolated from oysters. On day 7 postinfection (PI), significantly higher (P < 0.05) worm recovery rates (WRRs) were observed in ICR (29.5 +/- 12.0%) and C3H/HeN (14.8 +/- 8.2%) than in BALB/c (5.7 +/- 5.3%) and C57BL/6 (0.8 +/- 1.1%) mice. Alteration of the GC mucins was marked in C57BL/6 mice. On day 14 PI, 5.2 +/- 5.2% and 0.6 +/- 0.7% of worms were recovered only from ICR and C3H/HeN mice. When C57BL/6 mice were immunosuppressed with prednisolone, WRR on day 7 PI increased to 11.7 +/- 13.9%, whereas the GC hyperplasia and mucin alteration diminished significantly. The results suggest that expulsion of G. seoi from the intestine is dependent on immune responses of the host, and GCs may be an important effector.

Molecular differentiation of three species of Metagonimus by simple sequence repeat anchored polymerase chain reaction (SSR-PCR) amplification.

Genomic DNAs from 3 species of Metagonimus in the Republic of Korea, M. yokogawai, M. miyatai, and M. takahashii, were amplified by simple sequence repeat anchored polymerase chain reaction. Primers of 3' or 5' termini of the (CA)n repeats were used; 3 kinds of 3' anchored primers (CA)8RG, (CA)4RG, and (CA)8RY, and 2 kinds of 5' anchored primers BDB(CA)7C and DBDA(CA)7. The results showed distinguishable banding patterns among the 3 species of Metagonimus, suggesting that they have different genotypes.

Role of intraepithelial lymphocytes in mucosal immune responses of mice experimentally infected with Cryptosporidium parvum.

In order to investigate the role of intestinal intraepithelial lymphocytes (IELs) in host defense against Cryptosporidium parvum infection, conventionally bred immunocompetent (ImCT) ICR mice and immunosuppressed (ImSP) littermates were infected orally with 10(6) C. parvum oocysts. Then fecal oocyst excretion, the number and location of IELs, and their T lymphocyte subsets were observed on days 4, 7, 10, 13, 16, and 20 postinfection (PI). Uninfected ImCT and ImSP mice were used as controls. The starting point of oocyst excretion was day 4 PI in both ImCT- and ImSP-infected mice. The highest oocyst excretion occurred on day 7 PI in both groups, though the number of oocysts excreted was 3 times greater in ImSP than in ImCT mice. In ImCT mice, IELs greatly increased in number on days 16 and 20 PI (P < 0.05), but the increase was minimal in ImSP mice. IELs changed their location from the basal area to intermediate and apical areas of villous epithelial cells during the early stage of infection. In ImCT-infected mice, IEL phenotypes also changed; whereas CD4+ cells increased temporarily on day 7 PI (P < 0.05), CD8+ cells increased significantly on days 16 and 20 PI (P < 0.05). The results strongly suggest that IELs play a significant role in host defense against C. parvum infection, with helper T cells initiating control of the infection and cytotoxic T cells eliminating the parasites.

Acanthoparyphium tyosenense: the discovery of human infection and identification of its source.

Acanthoparyphium tyosenense Yamaguti, 1939 (Digenea: Echinostomatidae), was originally reported as an avian intestinal parasite; here, its presence is reported in 10 humans in the Republic of Korea. The patients were 9 adults aged 35-66 yr (males and females) and a young girl aged 7 yr residing in 2 coastal villages in Puan-gun, Chollabuk-do. The worms were recovered after treatment with praziquantel and purgation with magnesium salts. A total of 158 specimens (1-107 specimens/individual) was collected, together with varying numbers of other intestinal flukes. The patients had eaten various kinds of brackish water mollusks caught in an estuary near their villages. Five bivalves and a gastropod species suspected as sources of human infection were collected and examined. Two bivalves (Mactra veneriformis and Solen grandis) and the gastropod (Neverita bicolor) were found to be infected with the metacercariae of A. tyosenense; adult flukes were confirmed after the experimental infection of chicks. The results show that A. tyosenense infects humans and that brackish water mollusks are the source of human infection.

Genetic difference in susceptibility and fatality of three strains of mice experimentally infected with Neodiplostomum seoulense.

The genetic influence on host susceptibility to Neodiplostomum seoulense infection and fatality of the host was studied in 3 inbred strains of mice (BALB/c [H-2d], C3H/He [H-2k], and C57BL/6 [H-2b]). The survival of the mice, worm expulsion kinetics, worm size, number of eggs produced per day (EPD), and number of uterine eggs were observed from day 1 to day 40 postinfection (PI) with 100 or 200 metacercariae per mouse. Infection with N. seoulense was highly lethal to all 3 strains of mice, but the lethality was dose-dependent and varied according to the genetic backgrounds of the mice. The C3H/He mice exhibited the highest mortality, the lowest worm burdens and EPD, and the quickest expulsion of worms. It is suggested that different genetic backgrounds of mice appear to affect the host's capacity to expel N. seoulense and the fatality of the hosts themselves.

Stictodora lari (Digenea: Heterophyidae): the discovery of the first human infections.

Stictodora lari Yamaguti, 1939 (Digenea: Heterophyidae) was originally reported as an avian intestinal parasite and is here reported in humans. The patients were adults, aged 46-69 (2 men and 4 women), who resided in coastal villages in Sachon-gun, Kyongsangnam-do, or Shinan-gun, Chollanam-do. in the Republic of Korea. The worms were recovered after praziquantel treatment and purgation with magnesium salts. A total of 15 S. lari specimens (1-10 per individual) was collected, together with 5 other species of intestinal flukes. The flukes were morphologically characterized by a small body size (0.70-0.86 mm long and 0.27-0.36 mm wide), a ventrogenital sac, and a gonotyl armed with 70-80 spines in the form of a comma or reversed comma lying along their lateral margin. The patients had eaten the raw flesh of mullets and gobies that had been caught in an estuary near their villages. To the best of our knowledge, this is the first record of a human S. lari infection.

Susceptibility of various species of animals and strains of mice to Gymnophalloides seoi infection and the effects of immunosuppression in C3H/HeN mice.

Susceptibility to Gymnophalloides seoi infection was studied in 8 species of animals, including 7 strains of mice; the effects of immunosuppression on susceptibility were examined in C3H/HeN mice. One hundred metacercariae of G. seoi isolated from naturally infected oysters were orally administered to each animal. Worm recovery rate (WRR), worm dimensions, and the number of uterine eggs were obtained at day 3 and day 7 postinfection (PI). Average WRR from gerbils, hamsters, and cats at day 7 PI was 28.0%, 14.2%, and 10.9%, respectively, the former 2 figures of which were significantly higher than the rate of 0.0-4.0% from Sprague-Dawley rats, dogs, ducks, guinea pigs, and chicks. In the case of mice, average WRR at day 7 PI was 12.4% (KK strain), 11.8% (C3H/HeN), 9.6% (ICR), 6.4% (BALB/c), and 6.3% (ddY), respectively; the first 3 figures were significantly higher than the rates from other strains, which were 1.8% (A) and 0% (C57BL/6). At day 3 PI, WRR was much higher in all strains except C57BL/6. Worm maturation was the highest in C3H/HeN mice. Immunosuppression of C3H/HeN mice by injecting prednisolone for 7, 14, or 21 days prior to infection increased WRR at day 7 PI to 27.8%, 33.8%, or 67.5%, respectively. The results show that gerbils, hamsters, cats, and KK, C3H/HeN, ICR. BALB/c, and ddY mice are laboratory hosts that are fairly susceptible to G. seoi infection. In C3H/HeN mice, susceptibility was markedly enhanced by immunosuppression.

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.

Experimental infection of murine splenic lymphocytes and granulocytes with Toxoplasma gondii RH tachyzoites.

Toxoplasma gondii, an intracellular protozoan infecting many kinds of eukaryotic cells, has been used to experimentally infect macrophages, epithelial cells, fibroblasts, and various cancer cells, but rarely T and B lymphocytes or granulocytes. The present study was performed to determine the susceptibility of murine (BALB/c or CBA) splenic T and B lymphocytes, and granulocytes to infection with T. gondii RH tachyzoites. The ultrastructure of the infected host cells was observed by TEM, and the degree of intracellular parasite proliferation was quantified using 3H-uracil uptake assay. At 24 hrs post-culture, the host cell cytoplasm was found to contain 1 or 2, or a maximum of 7-8 tachyzoites. Infected T lymphocytes demonstrated a peripherally displaced nucleus, a parasitophorous vacuole enveloping the parasite, and an increased number of mitochondria. In B lymphocytes infected with tachyzoites, RER was not well developed compared to uninfected B lymphocytes. Uninfected granulocytes contained many electron-dense granules, but T. gondii-infected granulocytes demonstrated a decreased number of granules. Based on the 3H-uracil uptake assay, the susceptibility of T and B lymphocytes, and granulocytes, to infection with T. gondii tachyzoites was fairly high irrespective of cell type and strain of mouse. This strongly suggests deterioration in the functioning of infected host immune cells.

An epidemiological survey of Cryptosporidium parvum infection in randomly selected inhabitants of Seoul and Chollanam-do.

An epidemiological survey was performed to know the status of Cryptosporidium sp. infection among the people in Seoul and Chollanam-do in 1992. One village of Chollanam-do (Hwasun-gun) which showed the highest oocyst positive rate was re-surveyed in 1995 for human infection and for cattle also. The subjected areas consisted of 8 urban villages ( = dongs) of Seoul and 4 urban ( = dongs) and 7 rural ( = myons) villages of Chollanam-do. A total of 3,146 fecal samples was collected randomly, and smears were made from formalin-ether sediments. They were examined for Cryptosporidium oocysts by modified acid fast staining. The overall oocyst positive rate was 7.9% (248/3,146), but the rate was remarkably different between Seoul and Chollanam-do, 0.5% (4/853) and 10.6% (244/2,293), respectively. The average size of oocysts was 4.8 +/- 0.5 by 4.2 +/- 0.5 microns, compatible with C. parvum. In Chollanam-do, rural villages showed significantly higher rate (14.0%) than urban villages (3.7%). Especially the people in Iyang-myon. Hwasun-gun, a typical rural village, revealed a very high rate of 40.0% (74/185). Adults aged 51-70 years revealed the highest positive rate among all age groups. At the re-survey of the same village of Hwasun-gun in 1995, 44 (35.2%) of 125 villagers and 14 (93.3%) of 15 cattle examined were positive for C. parvum oocysts. The results suggest that C. parvum is highly prevalent in rural areas of Chollanam-do, and an important source or mode of infection seems to be contaminated water or contact with the feces of infected cattle.

Recovery of Pseudoterranova decipiens (Anisakidae) larvae from codfish of the Antarctic Ocean.

Anisakid larvae were found from the liver and stomach of the codfish (Notothenia neglecta) caught from the Antarctic Ocean, South Pole, where the Polar Research Center of The Korea Ocean Research and Development Institute is operating. Through observations by scanning electron (SEM) and light microscopy (LM), they were identified as the third stage larvae of Pseudoterranova decipiens, a potential agent of human codworm anisakiasis. They measured 25.0-28.5 mm in length, and 0.6-0.8 mm in width. The SEM revealed a prominent boring tooth, three lips, and excretory pore at the anterior end, and a small but prominent mucron at the posterior end. From whole mounts and cross sections of the larvae the ventriculus, intestinal cecum and a Renette cell were found characteristically at the same plane. Raw eating of the codfish caught from the Antarctic Ocean near the South Pole should be avoided so as to prevent human codworm anisakiasis in this area.

High endemicity of Metagonimus yokogawai infection among residents of Samchok-shi, Kangwon-do.

A small-scale epidemiological survey was undertaken during 1997-1998 on the residents along the Osib-chon (Stream), Samchok-shi (City), Kangwon-do (Province), to evaluate the status of Metagonimus yokogawai infection. A total of 165 fecal samples was collected and examined by cellophane thick smear and formalin-ether sedimentation techniques. The egg positive rate of M. yokogawai was 29.7%, showing a remarkable difference between males (46.6%) and females (16.3%). To obtain the adult flukes of M. yokogawai, 11 egg positive persons were treated with praziquantel and purged with magnesium sulfate. A total of 242,119 adult flukes (average 22,010 per person, 367-119,650 in range) was collected from diarrheic stools, all of which were identified as M. yokogawai. The results show that M. yokogawai is still highly endemic in this area.

Surface ultrastructure of Metagonimus miyatai metacercariae and adults.

A scanning electron microscopic study was performed to observe surface ultrastructures of excysted metacercariae and adults of Metagonimus miyatai. Metacercariae were collected from the scale of the pale chub (Zacco platypus), and adult flukes were harvested 1-4 weeks after infection to rats. In excysted metacercariae, the oral sucker was devoid of tegumental spines and had type I and type II sensory papillae. Anteriorly to the ventral sucker, spines were dense and digitated into 5-7 points, whereas near the posterior end of the body spines were sparse and digitated into 2-3 points. In one-week adults, 7 type II sensory papillae were arranged around the lip of the oral sucker, and at inner side of the lip one pair of small and two pairs of large type 1 sensory papillae were seen on each side. The distribution of tegumental spines was similar to that of metacercariae, but they were more differentiated with 9-11 pointed tips. In two- to four-week old adults, the surface ultrastructure was nearly the same as in one-week old adults, however, sperms were frequently seen entering into the Laurer's canal. Conclusively, the surface ultrastructure of M. miyatai was generally similar to that of M. yokogawai, however, differentiation of tegumental spines and distribution of sensory papillae around the oral sucker were different between the two species, which may be of taxonomic significance.

Surface ultrastructure of Metagonimus takahashii metacercariae and adults.

A scanning electron microscopic study was performed on the surface ultrastructure of metacercariae and adults of Metagonimus takahashii. Metacercariae were collected from the scale of crucian carp (Carassius auratus), and adult flukes were harvested 1-4 weeks after infection to rats. In excysted metacercariae, the oral sucker had type I (numerous) and type II (seven in total) sensory papillae. Tegumental spines were dense and digitated into 5-7 points on the surface anterior to the ventral sucker, but became sparse and less digitated posteriorly toward the end of the body. In adults, seven type II sensory papillae were characteristically arranged around the lip of the oral sucker, and on the inner side of the lip four small and two large type I sensory papillae were symmetrically seen on each side (12 in total). Tegumental spines on anterior two-thirds of the body, were digitated with 9-12 tips ventrally and 8-13 tips dorsally. Sperms entering into the Laurer's canal were observed. The results show that the surface ultrastructure of M. takahashii is generally similar to those of M. yokogawai and M. miyatai except for the digitation of tegumental spines.

Growth and development of Gymnophalloides seoi in immunocompetent and immunosuppressed C3H/HeN mice.

The growth and development of Gymnophalloides seoi were studied in C3H/HeN mice and effects of immunosuppression of the host on the worm development were observed. Two hundred metacercariae of G. seoi were orally administered to each mouse, and worms were recovered on days 1, 3, 5, 7, 14 and 21 post-infection (PI). The worm recovery rate was significantly higher in immunosuppressed (ImSP) mice than in immunocompetent (ImCT) mice except on days 1 and 3 PI. The worms attained sexual maturity by day 3 PI with eggs in the uterus, and worm dimensions and the number of uterine eggs continuously increased until day 14 PI in ImSP mice. Worms recovered from ImSP mice were significantly larger in size than those from ImCT mice on days 1 and 3 PI, and the number of uterine eggs was significantly larger in ImSP mice on days 5 and 7 PI. Genital organs such as the ovary, testes, and vitellaria, that were already developed in the metacercarial stage, grew a little in size until day 14 PI. The results show that the C3H/HeN mouse is, though not excellent, a suitable laboratory host for G. seoi.