• Parasites, Hosts and Diseases
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• v.44 no.3
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• pp.187-196
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• 2006

The mammalian trematode Paragonimus westermani is a typical digenetic parasite, which can cause paragonimiasis in humans. Host tissues and blood cells are important sources of nutrients for development, growth and reproduction of P. westermani. In this study, a cDNA clone encoding a 47 kDa hemoglobinase of P. westermani was characterized by sequencing analysis, and its localization was investigated immunohistochemically. The phylogenetic tree prepared based on the hemoglobinase gene showed high homology with hemoglobinases of Fasciola hepatica and Schistosoma spp. Moreover, recombinant P. westermani hemoglobinase degradaded human hemoglobin at acidic pH (from 3.0 to 5.5) and its activity was almost completely inhibited by E-64, a cysteine proteinase inhibitor. Immunohistochemical studies showed that P. westermani hemoglobinase was localized in the epithelium of the adult worm intestine implying that the protein has a specific function. These observations suggest that hemoglobinase may act as a digestive enzyme for acquisition of nutrients from host hemoglobin. Further investigations may provide insights into hemoglobin catabolism in P. westermani.

• Asian-Australasian Journal of Animal Sciences
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• v.17 no.3
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• pp.394-400
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• 2004

To investigate the efficacy of alternatives to antibiotics, the present study was conducted to compare the effects of antibiotic, lactic acid, a blend of commercial essential oils (EOs) and EOs in combination with lactic acid on growth performance and the functional activity of the gut in broiler chickens. A total of 168 broiler chickens were given the basal diet supplemented with 10 ppm colistin (T1), 0.1% lactic acid (T2), 25 ppm EOs (T3), 25 ppm EOs+0.1% lactic acid (T4), 50 ppm EOs (T5) or 50 ppm EOs+0.1% lactic acid (T6) in the period 3 to 35 days of age. As a result, the broiler chickens assigned to T4 group throughout the experimental period had apparently (p<0.05) greater body weight and total gain than these assigned to T1, T2, T3 and T5 groups. However, there was no difference in growth performance among the birds fed the diets supplemented with antibiotic (T1), lactic acid (T2) and EOs (T3 and T5) alone. The weights of digestive organs and the number of lactobacilli and E. coli in the lower ileum were not affected by dietary treatments. Total trypsin activity was significantly (p<0.05) greater in T4 than T1, T2, T3 and T5 groups. Total and specific pancreatic $\alpha$-amylase activities were significantly (p<0.05) enhanced in the broiler chickens fed T4 diet compared with these fed T1, T2 and T3 diets. However, there were no differences in growth performance and digestive enzyme activities including pancreatic trypsin and $\alpha$-amylase between T4 and T6 groups fed the diets supplemented with either low or high EOs levels in combination of lactic acid. In conclusion, a blend of commercial EOs combined with lactic acid showed significant increases in digestive enzyme activities of the pancreas and intestinal mucosa, leading to increase in growth performance.

• Parasites, Hosts and Diseases
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• v.29 no.3
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• pp.293-310
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• 1991

Clonorchis sinensis is a common parasite of man in Korea. Researches on the specific antigens of C. sinensis would be valuable not only because those elucidate the molecular characteristics of this fluke but also because it is applicable to immunodiagnosis. Although many monoclonal antibodies have been used in the field of parasite immunology, few articles on monoclonal antibodies against C. sinensis have been published so far. The aim of this study was to analyse C. sinensis antigens recognized by monoclonal antibodies, and to set up ELISA-inhibition test using C. sinensis specific monoclonal antibodies for improved specificity of immunodiagnostic tests. By fusion between spleen cells of the mice immunized with C. sinensis water-soluble crude adult worm antigens and plasmacytoma cells of mouse origin, 29 hybridoma clones secreting anti-C. sinensis monoclonal antibodies were made, and 8 clones among those were found specific. After cell cloning, isotypes of 6 selected specific monoclonal anti- bodies were determined to be IgGl, IgG2b and IgA. Four exposed antigenic determinants of natural infection were recognized by different specific monoclonal antibodies. By enzyme-immunoelectrotransfer blot, 10 KD, 34 KD antigenic determinants were found to be reacted with CsHyb 0714-20, CsHyb 0605-10 monoclonal antibodies, respectively, The antigenic determinant recognized by CsHyb 0714-20 monoclonal antibody was revealed to be located at the surface and parenchyme of a parasite by indirect immunoauorescent antibody technique, and those reacted with CsHyb 0605-10, CsHyb 0714-25 monoclonal antibodies were found at the parenchyme and intestine. The antigenic determinant reacted with CsHyb 0605-23 monoclonal antibody was found mainly around the uterine eggs. Four antigenic determinants recognized by specific monoclonal antibodies were all found to be present in the early eluted fractions of C. sinensis antigens separated by Sephadex G-200 gel filtration. By conventional ELISA, 75% of clonorchiasis cases were found positive, but 7.1% of normal controls and 37.5% of paragonimiasis cases showed false positives. However, by ELISA-inhibition test using C. sinensis specific monoclonal antibody (CsHyb 0605-23), 77.1% of clonorchiasis cases were found positive, and there were no false positives in normal controls or paragonimiasis cases, indicating 100% specificity. The ELISA- inhibition test using monoclonal antibodies was found to have same sensitivity and definitely high specificity in comparison with conventional ELISA for serodiagnosis of human clonorchiasis.

• Journal of Animal Science and Technology
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• v.53 no.3
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• pp.227-235
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• 2011

A total of 96 crossbred weanling barrows aged 21 days were randomly allocated to 32 pens of a new nursery to investigate the effects of antibiotics, phytogenics, and probiotics on intestinal growth and development. The animals were fed a set of three-phase basal diets containing 0.3% zinc oxide (CON) or the basal diets supplemented with 353 ppm of a combination of tiamulin, neomycin, chlortetracycline, and oxytetracycline (ANTI), 75 ppm triterpenoid saponin plus 150 ppm mixed saccharides (HERB; Sacchapin$^{(R)}$), or $1{\times}10^7$ brewer's yeasts plus $8{\times}10^7$ spores of each of Bacillus licheniformis and Bacillus subtilis per kilogram feed (PROBIO; Yeasture Plus 2B$^{(R)}$) for five weeks. Thirty-two pigs representing as many pens were slaughtered at the end of the feeding trial, after which morphological measures and digestive enzyme activities of intestinal mucosa were determined. Weight gain and gain:feed of the pigs were not affected by the dietary treatments (TRT) during the overall feeding trial. Total intestinal length was greater in PROBIO than in ANTI (P<0.05). Wet mucosa weight of the duodenum was not affected by TRT. However, jejunal mucosa weight was greater in PROBIO than in any other group sum of mucosa weights of the duodenum and jejunum was greater (P<0.05) in PROBIO than in ANTI and HERB. The height and width of duodenal villus were not affected by TRT, but crypt depth decreased (P<0.05) in response to HERB and PROBIO vs CON. Specific activities of alkaline phosphatase, sucrase, maltase, lactase, and leucine aminopeptidase in the duodenum and jejunum were not changed by TRT. In conclusion, results suggest that the present dietary treatments have no effects on growth performance of weanling pigs and that of PROBIO enhances intestinal growth and development under a clean experimental setting.

• Microbiology and Biotechnology Letters
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• v.36 no.4
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• pp.299-306
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• 2008

Human pancreatic lipase is a digestive enzyme which is synthesized in pancreas, secreted into small intestine, and there hydrolyze the fat in food. Pancreatic lipase protein composes of catalytic domain and colipase-binding domain. In this research, the gene segments corresponding to total protein, catalytic domain, and co lipase-binding domain were cloned by PCR method, inserted into an expression vector, and then used to transform Escherichia coli BL21 (DE3). The recombinant proteins produced were purified and injected intramuscularly three times into laying hens. The egg yolk antibodies (IgY) were obtained from the egg yolks and tested for their antibody titer. Among three IgY, the IgY against colipase-binding domain showed the highest antibody titer. All three IgY had inhibitory effects on the porcine pancreatic lipase. Among them, the IgY against colipase-binding domain showed the highest inhibition effects. The fat diet with corn oil and IgY was administrated to the experimental rats and their blood compositions were examined with time course. The triglyceride concentration of treated rats was decrease meaningfully when compared with those of control rats. This suggested that the IgY against colipase-binding domain antigen inhibited pancreatic lipase in vivo.

• Journal of Life Science
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• v.19 no.9
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• pp.1226-1231
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• 2009

Enzyme substitution with recombinant phenylalanine ammonia-lyase (PAL, EC 4.3.1.5) is currently being explored for treatment of phenylketonuria (PKU), an autosomal recessive genetic disorder with mutations of the gene encoding phenylalanine-4-hydroxylase (EC 1.14.16.1). However, oral administration of PAL is limited because of proteolytic digestion in the gastrointestinal tract. The aim of this study was to determine the biochemical properties of PAL and delinate the susceptibility of wild-type PAL to pancreatic proteolysis by exploring several mutants, and to develop therapeutic drugs with PAL for PKU. The specific activity of PAL was assayed and its optimal pH, temperature stability, and intestinal protease susceptibility were investigated. Its $V_{max}$ values for phenylalanine and tyrosine were 1.77 and $0.47{\mu}mol$/ min/mg protein, respectively, and its $K_m$ values were $4.77{\times}10^{-4}$ and $4.37{\times}10^{-4}\;M$, respectively. PAL showed an optimal pH at 8.5, corresponding to the average pH range of the small intestine. It showed no loss of activity at $-80^{\circ}C$ for 5 months and possessed 93.4% of its activity under $4^{\circ}C$ for 4 wks. PAL was susceptible to chymotrypsin digestion and, to a lesser extent, to trypsin, elastase, carboxypeptidase A, and B. The trypsin and chymotrypsin cleaving sites were mutated to investigate protection from pancreatic digestion and the specific activities of these mutants were evaluated. The six mutants displayed low specific activities compared to the wild-type, suggesting that the primary trypsin and chymotrypsin cleaving sites may be essential for catalytic reaction. The PAL mutants could therefore be applied as a pretreatment modality without susceptibility to proteolytic attack, however, additional modification for enhancing enzymatic activity is needed to reduce the Phe levels effectively.