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R C Ko

Publications and source records attributed to R C Ko.

At least 37 records · Page 2Linked to original sources
PubMed1997

Production specific antigens from Trichinella spiralis using a continuous elution method and isoelectric focusing.

Immunodominant antigens of 45-53 kDa (one band per fraction) were obtained from excretory/secretory (E/S) and somatic products of infective-larvae of Trichinella spiralis using a continuous-elution method. They were further resolved by isoelectric focusing into different isoforms (45 kDa: pI4.47, 5.09, 5.47 and 5.86; 47 kDa: pI4.72 and 4.97; 53 kDa: pI4.86, 5.11, 5.44 and 5.78). In immunoblotting, the isoforms of pI 5.09, 5.86, 4.97, 5.44 and 5.78 did not cross-react with antisera against Trichuris suis, Metastrongylus apri, Gnathostoma hispidum and Stephanurus dentatus. Hence, they have the potential to serve as specific antigens for the serodiagnosis of trichinellosis.

Animals↗
PubMed1996

Heat shock response of Trichinella spiralis and T. pseudospiralis.

Heat shock proteins (HSPs) were documented for the first time in both somatic extracts and excretory/secretory (ES) products of the infective-stage larvae of Trichinella spiralis and T. pseudospiralis. Larvae recovered from muscles of infected mice were heat shocked at 37, 40, 43 and 45 degrees C in RPMI 1640 medium containing L(-)[35S]methionine. Somatic extracts and ES products of heat-shocked worms were then analysed by SDS-PAGE, autoradiography and laser densitometry. Prominent bands of HSPs were observed at 43 degrees C which is the optimal heat shock temperature. The major HSPs in somatic extracts of T. spiralis were 20, 47, 50, 70, 80 and 86 kDa. When the temperature was increased from 37 to 43 degrees C, the greatest increase in absorbance was observed in HSPs 70 and 86. In vitro translation of mRNA in a nuclease-treated rabbit reticulocyte lysate system showed an increase in the synthesis of the 80 kDa protein. This suggests that the production of HSP 80 is regulated at the transcriptional level. The major HSPs in the ES products were 11, 45, 53 and 64 kDa. In T. pseudospiralis, the major HSPs in the somatic extracts were 20, 26, 31, 50, 53, 70, 80 and 86 kDa, and in the ES products, 11, 35, 37, 41 and 64 kDa.

Animals↗
PubMed1994

Angiostrongylus cantonensis: characterization of thymidylate synthetase.

Thymidylate synthetase (TS) is the only enzyme that catalyzes the formation of thymidine nucleotides in Angiostrongylus cantonensis. A fraction enriched in TS was obtained from the gravid nematode by gel filtration and affinity chromatography using methotrexate-agarose. TS, which was well separated from dihydrofolate reductase, has a relative molecular mass of 66 kDa. By electrophoresis in sodium dodecyl sulphate gel, a major protein band corresponding to 31 kDa was observed. This band was shown to be TS by comparing the electrophoretic mobility with an enzyme preparation bound with [6-3H]5-fluoro-2'-deoxyuridine 5'-monophosphate (FdUMP). Therefore, the enzyme is composed of two identical or very similar subunits. Velocity studies and product inhibition patterns revealed that the TS reaction undergoes a sequential mechanism in which 2'-deoxyuridine 5'-monophosphate (dUMP) is the first substrate added to the active site and thymidine 5'-monophosphate is the last product released. The apparent Km values for dUMP and 5,10-methylenetetrahydrofolate are 10 and 185 microM, respectively. FdUMP and trimethoprim inhibited the parasite TS competitively with dUMP and the Ki values of 23.5 nM and 852 microM, respectively. Methotrexate was a noncompetitive inhibitor of TS. At 0.2 mM 5,10-methylenetetrafolate, 1 mM methotrexate inhibited the activity by 74%.

Ammonium Sulfate↗
PubMed1994

Changes in host muscles induced by excretory/secretory products of larval Trichinella spiralis and Trichinella pseudospiralis.

Excretory/secretory (ES) products obtained by in vitro culture of infective-stage larvae of Trichinella spiralis and T. pseudospiralis were injected intramuscularly at various intervals into mice. Mini-osmotic pumps containing T. spiralis ES products were also implanted subcutaneously and intraperitoneally into rats. The introduction of ES materials into muscles elicited extensive lesions which included dissolution of myofibres, mobilization of mononuclear and polymorphonuclear leucocytes, angiogenesis, hypertrophy of myonuclei, myotube formation, mitosis, muscle bundles becoming rounded and separated from each other, disappearance of Z, I and A bands of sarcomeres, increase in endoplasmic reticulum and Golgi complexes, decrease in glycogen and relocation of mitochondria. These are considered as degenerative/regenerative changes of muscles to injury. Immunodominant epitopes of specific 45-53 kDa glycoproteins in ES antigens of T. spiralis could not be detected in hypertrophic nuclei of injected muscles by using polyclonal and monoclonal antibodies and immunocytochemical methods. ES products of T. spiralis failed to stimulate unsensitized lymphocytes in the lymphocyte transformation test. Infective-stage larvae of T. spiralis released from muscles were found capable of forming nurse cells after injection subcutaneously into rats. It is postulated that the invasion of muscles by trichinellids elicits two independent events, i.e. a general degenerative/regenerative response of muscles and a specific change in genomic expression of myonuclei. The two events are probably mediated by different effector molecules.

Animals↗
PubMed1993

Pathways of pyrimidine nucleotide biosynthesis in gravid Angiostrongylus cantonensis.

The incorporation of radioactive precursors into pyrimidine nucleotides via de novo and salvage pathways was measured in gravid Angiostrongylus cantonensis by HPLC and thin-layer chromatography. 14C-labelled orotate, uridine, uracil and deoxyuridine were traced to UMP, UDP, UTP, UDP-glucose, dTMP, CMP, CDP and CTP. 3H-labelled cytidine was also incorporated into both uracil and cytosine nucleotides in a ratio of 2:1. Cytosine was a major end-product for all the precursors. Cytosine nucleotides were probably formed from UTP by the action of CTP synthetase whose activity in crude cell-free extract was 31.5 +/- 4.9 pmol min-1 (mg protein)-1. It was dependent on glutamine, ATP and GTP and was inhibited by CTP. The total amount of pyrimidine nucleotides formed from uridine was 3 times of that from uracil. The presence of uracil in the metabolism of uridine indicates that UMP is formed by uracil phosphoribosyltransferase as well as by uridine kinase. UMP is a key intermediate for cytidylate and thymidylate biosynthesis in the gravid worms.

Angiostrongylus cantonensis↗
PubMed1992

Precursors of pyrimidine nucleotide biosynthesis for gravid Angiostrongylus cantonensis (Nematoda: Metastrongyloidea).

Gravid Angiostrongylus cantonensis can utilize radiolabelled bicarbonate, orotate, uracil, uridine and cytidine but not cytosine, thymine and thymidine for the synthesis of RNA and DNA. In cell-free extracts of the worm, a phosphoribosyltransferase was shown to convert orotate to OMP and uracil to UMP. A similar reaction was not observed with cytosine and thymine. Uridine was readily phosphorylated by a kinase but a similar reaction for thymidine and deoxyuridine was not found. Cytidine could be phosphorylated by a kinase or be deaminated by a deaminase to uridine. No deaminase for cytosine was detected. There was also no phosphotransferase activity for pyrimidine nucleosides in the cytosolic or membrane fractions. Pyrimidine nucleosides were, in general, converted to the bases by a phosphorylase reaction but only uracil and thymine could form nucleosides in the reverse reaction. The activity of thymidylate synthetase was also measured. These results indicate that the nematode synthesizes pyrimidine nucleotides by de novo synthesis and by utilization of uridine and uracil and that cytosine and thymine nucleotides are formed mainly through UMP. The thymidylate synthetase reaction appears to be vital for the growth of the parasite.

Angiostrongylus↗
PubMed1992

Specificity of affinity-purified Trichinella spiralis antigens.

An affinity-purified fraction (APF) was obtained by passing crude somatic antigens of Trichinella spiralis muscle larvae through an Affi-Gel 10 column coupled with anti-Trichuris suis IgG. The fraction contained seven antigens with molecular weights ranging from 28 to 55 kDa. When tested with antiserum against other common nematodes of pigs from China, the APF was found to be markedly more specific than S3 antigens (prepared by a combination of cell fractionation and differential centrifugation according to Despommier and Lacetti, 1981) and fractions produced by Sephacryl S-200 gel filtration (F1 to F12). When the APF was used in an indirect IgG-enzyme-linked immunosorbent assay (IgG-ELISA) to screen serum samples from 2000 pigs imported from China, a positive rate of 7.5% was obtained. Similar screenings using the crude somatic antigens F1 and S3 gave a large number of cross-reactions and false positive reactions. Positive rates of 48%, 39% and 59.5% respectively were obtained for the three antigens.

Animals↗
PubMed1992

Purification of larval Taenia solium antigens by gel filtration.

Crude larval Taenia solium extracts were fractionated by Sephacryl S-200 gel filtration into four fractions (W1-W4). The sensitivities of the fractions to rabbit and pig antiserum against Taenia solium were tested by double immunodiffusion, immunoelectrophoresis, and ELISA. Fraction W2 which was highly sensitive to antisera was shown by immunoblotting to contain antigen B (95 and 105 kDa). The four fractions were shown to contain antigenic determinants common with pig serum proteins and crude extracts of other Platyhelminthes (especially Taenia hydatigena). Fraction W2 has the potential to be used as a serodiagnostic antigen.

Animals↗
PubMed1992

Enhanced chemiluminescent enzyme immunoassay for the detection of trichinellosis antibodies in pigs.

A modified enhanced chemiluminescent enzyme assay (ECIA) was developed for mass screening of pigs for trichinellosis antibodies in abattoirs. Using Bionectics beads as solid support, the assay time could be reduced to 45 min. Optimal conditions for washing, blocking, incubation, concentration of serum, antigens and conjugates as well as timing of film exposure were determined. The sensitivity and specificity of the assay were found to be comparable to those of the triple antibody-IgG ELISA. The assay was tested in an abattoir and its efficacy was found to be satisfactory. However, the major disadvantage of the assay is the high cost of magnetic beads.

Animals↗
PubMed1992

Trichinella spiralis: specificity of ES antigens from pre-encysted larvae.

Excretory/secretory (ES) antigens were obtained by culturing pre-encysted Trichinella spiralis larvae which were recovered from muscles of experimentally infected mice 14-15 days postinfection. Analyses of these antigens (PEL ES) with immunoblotting, SDS-PAGE and Triple Antibody ELISA showed that they yielded a low sensitivity and specificity when tested with antisera against the common nematodes of Chinese pigs. As compared to ES antigens from encysted larvae, PEL ES also contained more low molecular mass proteins.

Animals↗
PubMed1991

Isolation of specific antigens from Trichinella spiralis by the rotating horizontal ampholine column method.

By preparative isoelectric focusing in a rotating ampholine column, specific antigens with molecular weights of 45, 47 and 53 kDa were successfully isolated from crude somatic extracts of Trichinella spiralis muscle larvae. These antigens, with pI 5.5, 4.3 and 4.4, were found to possess the same epitope found in specific antigens in the excretory/secretory products that were recovered by in vitro culture of muscle larvae. When the antigens obtained by the two methods were compared using enzyme-linked immunosorbent assays, they showed similar specificity and sensitivity.

Ampholyte Mixtures↗
PubMed1991

Cross-reactions between crude antigens of larval Taenia solium (Cysticercus cellulosae) and other helminths of pigs.

A crude antigen extract of larval Taenia solium was shown by immunodiffusion (ID) and immunoelectrophoresis (IEP) to cross-react with rabbit antisera against pig serum proteins and larval T. hydatigena, and by enzyme-linked immunosorbent assay (ELISA) with antisera against pig serum proteins, Fasciolopsis buski, larval T. hydatigena, hydatid cyst, Hymenolepis diminuta and Dipylidium caninum. Immunoblotting demonstrated that the crude antigens extract contained epitopes of pig serum proteins of 48 and 66 kDa. The crude extract also contained a subunit of antigen B (95 kDa) which was also found in T. hydatigena and H. diminuta. Immunoperoxidase and indirect immunofluorescence studies showed that cross-reacting antigens were distributed mainly on the tegument of T. solium.

Animals↗
PubMed1991

Trichinella spiralis: antigenic epitopes from the stichocytes detected in the hypertrophic nuclei and cytoplasm of the parasitized muscle fibre (nurse cell) of the host.

Monoclonal antibodies raised against antigens present in the excretions/secretions (E/S) of larval Trichinella spiralis, polyclonal antibodies raised against E/S and antisera from rabbits and pigs infected with T. spiralis were used in conjunction with immunocytochemical techniques to detect antigens in sections of muscle from mice that had been infected with T. spiralis for 15 or 30 days. The antibodies recognized epitopes in the stichocytes, on the surface of the cuticle, in the lumen of the oesophagus and in the lumen of the intestine of encysted larvae. Monoclonal antibodies 7C2C5 and 1H7 and the polyclonal antibodies recognized epitopes in the cavity occupied by the larva, in the cytoplasm of the nurse cell, and in the hypertrophic nuclei of the nurse cell, but did not recognize material in the smaller nuclei of the nurse cell, in the cyst wall or in the surrounding muscle. Monoclonals 3B2E6 and 1D11G8B2, which recognized epitopes in the stichocytes and on the surface of the cuticle of the larvae, gave negative results with the cytoplasm and nuclei of the nurse cell. A polyclonal antibody raised against Trichuris suis recognized epitopes in the muscle and hypodermis of encysted T. spiralis but gave negative results with material in the nurse cell and nurse cell nuclei. The possibility that the antigen detected in the cytoplasm and nuclei of the nurse cell is produced by the stichocytes of the nematode and that it is controlling genes of the altered muscle fibre, either directly or indirectly, is discussed.

Animals↗
PubMed1991

Current status of food-borne parasitic zoonoses in Hong Kong.

Although there have been no recent epidemiological studies on parasitic zoonoses in Hong Kong, the following diseases are known to occur locally: toxoplasmosis, clonorchiasis, fasciolopsiasis, cysticercosis, hydatidosis, sparganosis, trichinellosis, angiostrongyliasis, gnathostomiasis and trichostrongylosis. Most of them are due to the consumption of infected animals imported from China. Due to the general unawareness of parasitic diseases by physicians and health authorities, the importance of zoonoses has been under recognized.

Animals↗
PubMed1990

Lipids of Angiostrongylus cantonensis (Nematoda: Metastrongyloidea): a comparison between young adults and gravid worms.

1. All major classes of lipids were found in the young adults in brain (22 days post-infection) and gravid Angiostrongylus cantonensis in lung of rats (34 days post-infection) comprising approximately 60% of phospholipids, 30% of neutral lipids and the rest, glycolipids. 2. The relative composition of phospholipids were quite similar between worms from the two different habitats, with phosphatidylcholine predominating. The glycolipid profiles were also similar. 3. More neutral lipids in the worms from brain existed as cholesterol and cholesterol esters than those from the lung. More than 20% of the fatty acids in these lipids of the brain were found as C10-C14 acids while oleic acid was the main component in the lung worm.

Aging↗
PubMed1988

Comparison between standard ELISA and dot-ELISA for serodiagnosis of human trichinosis.

The dot enzyme-linked immunosorbent assay (dot-ELISA) provided rapid and accurate detection of human trichinosis. Comparable sensitivity and specificity of the standard ELISA and dot-ELISA were obtained when sera of 18 confirmed cases of trichinosis were screened. Cross-reactivity was assayed with sera from 31 Chinese subjects with other parasitic infections. Only 2 cross-reactions occurred in the dot-ELISA, compared with 7 in the standard ELISA.

Antigens, Helminth↗
PubMed1987

Four documented cases of eosinophilic meningoencephalitis due to Angiostrongylus cantonensis in Hong Kong.

4 cases of eosinophilic meningoencephalitis in Hong Kong are described. The major clinical features of the patients, who were 2-60 years old, were low grade fever, headache, mild meningeal signs, right facial palsy or hemiplegia. Eosinophilia in the peripheral blood and eosinophilic pleocytosis were prominent. Computerized tomography scans of brain showed a small area of attenuation with surrounding hypodense area; the lesion was resolved 1-2 months after admission to hospital. Electroencephalograms revealed abnormally slow dysrhythmia. Sections of a nematode observed in the brain of a patient who died were identified as those of young adult Angiostrongylus cantonensis. High ELISA titres against the crude antigens of this nematode were also noted in the serum of 3 patients. The disease is probably under-recognized in Hong Kong.

Adult↗