The infestation rates and abundances of human infecting metacercariae (Clonorchis sinensis, Metagonismus spp., Centrocestus armatus, Echinostoma hortense, Echinochasmus japonicus, Clinostomum complanatum) in freshwater fish were investigated at the three river areas- Taewhagang (river), Hyongsangang(river), Nakdonggang(river) -in Kyongsang-do and at four streams-Yonpungchon, Munsanchon, Kyonganchon, and Konjiamchon-in Kyonggi-do, Korea in 1994-1995. The fish caught at Taewhagang were heavily infested with metacercariae of Clonorchis sinensis and Centrocestus armatus. At Hyongsangang, Zacco platypus and Z. temmincki were moderately infested with metacercariae of C. armatus.
Chomanpo, at the basin of Nakdonggang, was still endemic for C. sinensis. In the fish caught at four streams of Kyonggi-do, metacercariae of C. sinensis exhibited the highest infestation rate and intensity out of 6 species of metacercariae. The infestation intensity of C. sinensis metacercariae in fish flesh was markedly different according to each division of flesh. The cause of this difference was conjectured as a result of larval behavior. The metacercariae of C. armatus were found in almost all parts, except scales and fins, of fish. The infestation rates and intensities of C. sinensis and C. armatus metacercariae in Taewhagang greatly increased as compared with those of previous reports. Rhinogobius brunneus and Acanthorhodeus macropterus are newly recorded intermediate hosts of Echinostoma hortense. The reason of large differences from previous data was discussed and the standard method of metacercaria examination was proposed.
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A nationwide survey was performed to know the geographical distribution of Gymnophalloides seoi (Digenea: Gymnophallidae) metacercariae in Korea, by examining the infection status of locally produced oysters, Crassostrea gigas. A total of 24 coastal areas (myons) of 14 guns ( = counties) in Kyonggi-do, Chollabuk-do, Chollanam-do, Kyongsangnam-do, Kyongsangbuk-do, or Kangwon-do, where natural oysters are produced but G. seoi has never been reported, and 13 areas (myons) of Shinan-gun, Chollanam-do, nearby the known endemic area, were surveyed. Oysters from nonendemic areas were free from G. seoi infection, except Byonsan- myon of Buan-gun, Chollabuk-do, where one of 50 oysters examined was infected with 15 metacercariae of G.
seoi. In Shinan-gun, oysters from 10 areas including Aphae-myon ( = township) and Anjwa-myon were infected with the metacercariae, with the infection rate ranging from 1.7% to 100% by areas. The intensity of infection was the highest in Aphae-myon. 785.9 metacercariae per oyster. The results indicate that high prevalence of G. seoi is confined to Shinan-gun, but low grade prevalence is also present in adjacent areas such as Buan-gun, Chollabuk-do.
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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.
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For the detection of Cryptosporidium oocysts, fecal samples were collected from 201 calves which showed diarrhea. Among the 201 samples, 29 samples (14.4%) were positive for Cryptosporidium spp. by the DMSO- modified acid-fast stain (MAFS), 23 samples (11.4%) were positive by commercial kit (Meridian Diagnostics, Cincinnati, Ohio) and 23 by the indirect immunofluorescence antibody (IFA) assay employing the monoclonal antibody (mAb C6). When tested by both IFA and MAFS, 20 fecal samples were positive for Cryptosporidium oocysts whereas 169 fecal samples were negative. If the MAFS is considered a standard method for oocyst detection, the IFA showed 69% of sensitivity and 98% of specificity. When tested by both IFA and commercial kit, 22 fecal samples were positive for Cryptosporidium oocysts while 177 samples were negative. One sample tested by IFA was found to be false negative, when compared with the results by commercial kit.
The sensitivity of IFA was calculated as high as 96%; the specificity as 99% and the predictive value was also 99%. In the present study, IFA employing the mAb C6 revealed that 23 samples (11.4%) were positive among the 201 calves showing diarrhea. Of 23 IFA positive samples, 4 samples (5%) showed cryptosporidial oocysts more than 10(5) OPG. Therefore, it is concluded that the calves showing cryptosporidial oocysts more than 10(5) OPG in the feces were highly associated with clinical cryptosporidiosis.
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Twelve isolates of Acanthamoeba spp. assigned to either A.
castellanii or A. polyphaga, and type strains of A.
culbertsoni, A. healyi, A. palestinensis, and A. astronyxis were examined by restriction fragment length polymorphism (RFLP) of a conserved region of small subunit ribosomal RNA gene (ssu rDNA) amplified by polymerase chain reaction (PCR). The PCR products of the isolates measured approximately 910-930 bp, except for that of A. astronyxis which was extraordinarily long, approximately 1,170 bp.
Average of estimated sequence divergence of the amplified DNA among the isolates assigned to A. castellaii was 9.8% whereas that among the isolates assigned to A. polyphaga 9.6%. The maximum intraspecific sequence divergence among the isolates assigned to A. castellanii was observed between the Chang and Ma strains (17.3%) while that among the isolates assigned to A. polyphaga was observed between KA/S3 and KA/S7 strains (16.1%). The both maximum sequence divergences were much greater than the minimum interspecific sequence divergence between A. castellanii and A. polyphaga (2.6%) which appeared between the Castellani (or CCAP 1501/2 g) and KA/S3 strains. The PCR-RFLP patterns of A.
culbertsoni, A. healyi, A. palestinensis, and A. astronyxis were quite diverse from one another and from those of isolates assigned to either A. castellanii or A. polyphaga.
It is suggested that taxonomic validity of the isolates assigned to either A. castellanii or A. polyphaga should be reevaluated.
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, and
Naegleria fowleri
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Antigenic domain of major surface protein (p30) of Toxoplasma gondii was analyzed after polymerase chain reaction (PCR) of its gene fragments. Hydrophilic or hydrophobic moiety of amino acid sequences were expressed as glutathione S-transferase (GST) fusion proteins. Fragments of p30 gene were as follows: T37, total p30 open reading frame (ORF); S28, total ORF excluding N-terminal signal sequence and C- terminal hydrophobic sequence: A19, N-terminal 2/3 parts of S28; P19, C- terminal 2/3 of S28; X9, N-terminal 1/3 part of S28; Y10, middle 1/3 of S28; and Z9, C-terminal 1/3 of S28, respectively. Primer of each fragment was synthesized to include clamp sequence of EcoR I restriction site. PCR amplified DNA was inserted into GST (26 kDa) expression vector, pGEX-4T-1 to transform into Escherichia coli (JM105 strain). GST fusion proteins were expressed with IPTG induction as 63, 54, 45, 45, 35, 36, and 35 kDa proteins measured by SDS-PAGE. Each fusion protein was confirmed with GST detection kit. Western blot analysis with the serum of a toxoplasmosis patient revealed antigenicity in proteins expressed by T37, S28, and A19 but not those by P18, X9, Y10, and Z9. Antigenicity of p30 seems to be located either in N-terminal 1/3 part in the presence of middle 1/3 part or in the oligopeptides between margins of the first and second 1/3 parts.
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