• Current Research on Agriculture and Life Sciences
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• v.2
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• pp.115-121
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• 1984

Some investigations on chronic mastitis in dairy cattle in Taegu-Kyungpook Provinces from the beginning of October, 1984 till the end of August, 1985 were conducted with the particular regard to the causative agents and their drug susceptibility. Milk samples from 83 isolated cases of chronic mastitis cattle were investigated bacteriologically and the causative organisms recovered were examined for their antibiotic susceptibility by using disc diffusion susceptibility technique against the major antibiotics of current veterinary use. Major causative agents involved in chronic mastitis in Taegu-Kyungpook Provinces were in order of prevalence Staphylococcus spp. (48.2 %), Escherichia coli (18.1 %), Candida spp. (10.8 %) and Corynebacterium spp. (8.4 %), Streptococcus agalactiae (3.6 %), Bacillus cereus (3.6 %) and Pseudomonas aeruginosa (2.4 %) were found to be one of the minor agents. The majority of staphylococcal isolates and E. coli were highly resistant to the most antibiotics tested. The percentages of staphylococcal cultures resistant to penicillin, methicillin, lincomycin, novobiocin, ampicillin and tetracycline were 87.2 %, 78.7 %, 68.1 %, 61.7% and 57.4 %, respectively, while the majority of them were susceptible to gentamicin(78.7 %), cephalothin(76.6 %) and chloramphenicol (74.5%). E. coli isolates were found to be highly resistant to streptomycin, cephalothin, tetracycline and ampicillin while the majority of them were susceptible to colistin (83.3 %), gentamicin (77.8 %) and chloramphenicol (66.7 %). Corynebacterium spp. were susceptible to ampicillin, chloramphenicol, erythromycin, gentamicin, oleandomycin and tetracycline although they showed resistance to novobiocin and penicillin. Two cultures of Pseudomonas aeruginosa recovered from mastitis milk were highly resistant to the antibiotics employed in the present study.

• Applied Biological Chemistry
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• v.22 no.4
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• pp.191-197
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• 1979

${\beta}-Tyrosinase$ was purified and crystallized from cells of Escherichia intermedia A-21 grown in a medium supplemented with 0.2% L-tyrosine. Molecular weight of its subunit, Km value and absorption spectra were determined. Crystallization methods were also studied to eliminate any unnecessary procedures. The results obtained were as follows: 1. The purification procedure included ammonium sulfate fractionation, dialysis against potassium phosphate buffer, pH 6.0 and pH 7.0, and DEAE-Sephadex column chromatography. In the column chromatography, 11 mg of protein was applied per ml of DEAE-Sephadex for efficiency. 2. Steps of protamine sulfate treatment and Sephadex G-150 gel filtration could be eliminated for this enzyme from the known procedures. 3. The purified enzyme was dissolved in 0.01M potassium phosphate buffer containing 2-mercaptoethanol, with a concentration of 20mg/ml. Crystalline enzyme, which appears as hexagonal rods, was obtained by adding solid fine powdered ammonium sulfate to the enzyme solution. 4. Absorption maxima of the enzyme appeared at 340 and 430nm when associated with pyridoxal phosphate. 5. Km value of the enzyme for L-tyrosine was $2.31{\times}10^{-4}M$ and the molecular weight of its subunit was determined by SDS-polyacrylamide electrophoresis to be approximately 50,000.

• Current Research on Agriculture and Life Sciences
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• v.4
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• pp.132-140
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• 1986

Eighty one products from 36 kinds of vitamin and mineral feed supplement collected during August, 1984 to February, 1985 were examined for microbiological contamination. In addition, 83 strains of coliform isolated from the samples were tested for the resistance to 8 kinds of antimicrobial drugs and distribution of R plasmid. General bacteria were detected in all of samples tested. Bacterial population was varied from less than 10 per gram of the sample to 1,400,000 per gram and 34 (42%) of 81 samples were contaminated with 100 to 1,000 cells per gram. Coliform isolation, which was more frequent in samples with larger number of general bacteria, was possible in 14 (17.3%) out of 81 samples tested and 6 (33.3%) out of 18 companies were coliform positive in their products. Forty one (49.4%) out of 83 coliform isolates were fecal coliform. The frequency of resistant strains was the highest to sulfadimethoxine (Sa) with 92.8% and followed by streptomycin (Sm, 67.5%), tetracycline (Tc, 50.6%), kanamycin (Km, 26.5%), chloramphenicol (Cm, 18.1%) and ampicillin (Am, 15.7%). No strain was resistant to nalidixic acid (Na) and gentamicin (Gm). The resistance frequency of fecal coliform strains were higher compare to non-fecal coliform strains. There were minimum inhibitory concentration (MIC) of $3,200{\mu}g/m{\ell}$ or higher in 7 strains to Am, 3 to Sm and 3 to Km, and 70 strains had MIC of $1,600{\mu}g/m{\ell}$ of higher to Sa while Tc had MICs from $1.6{\mu}g/m{\ell}$ to $400{\mu}g/m{\ell}$. All strains had MICs of $6.3{\mu}g/m{\ell}$ of lower to Na and $3.1{\mu}g/m{\ell}$ of lower to Gm. Seventy nine (95.2%) of 83 strains were resistant to one or more drugs tested. The most frequent resistance patterns were SaSm (14.5%) and followed by SaSmTc(12%), SaSmTcKm(8.4%) SaTc (8.4%) and SaSmKm (7.2%) ; total 19 different patterns were noted. Thirty two (40.5%) of 79 resistant strains were transferred all of a part of their resistance to Escherichia coli ML 1410. The frequency of transferable resistance was high in Am (100%) and Cm (80%) while low in Tc (38.1%), Sa (18.2%), Sm (17.9%) and Km (4.5%).

• Applied Biological Chemistry
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• v.22 no.4
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• pp.198-209
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• 1979

L-Tyrosine, 2-chloro-L-tyrosine, 2-bromo-L-tyrosine, and 2-iodo-L-tyrosine were synthesized by ${\beta}-tyrosinase$ obtained from cells of Escherichia intermedia A-21, through the reversal of the ${\alpha},{\beta}-elimination$ reaction, and their molecular structures were analyzed by element analysis, NMR spectroscopy, mass spectrometry and IR spectroscopy. Rates of synthesis and hydrolysis of halogenated tyrosines by ${\beta}-tyrosinase$, inhibition of the enzyme activity by halogenated phenols, and effects of addition of m-bromophenol on the synthesis of 2-bromotyrosine were determined. The results obtained were as follows: 1) In the synthesis of halogenated tyrosines, the yield of 2-chlorotyrosine from m-chlorophenol were approximately 15 per cent, that of 2-bromotyrosine from m-bromophenol 13.8 per cent, and that of 2-iodotyrosine from m-iodophenol 9.8 per cent. 2) Rate of synthesis of halogenated tyrosines by ${\beta}-tyrosinase$ was slower than that of tyrosine and the rates were decreased in the order of chlorine, bromine and iodine, that is, by increasing the atomic radius. Relative rate of 2-chlorotyrosine synthesis was determined to be 28.2, that of 2-bromotyrosine to be 8.13, and that of 2-iodotyrosine to be 0.98, respectively, against 100 of tyrosine. However 3-iodotyrosine was not synthesized by the enzyme. 3) The relative rate of 2-chlorotyrosine hydrolysis by ${\beta}-tyrosinase$ was 70.7, that of 2-bromotyrosine was 39.0, and that of 2-iodotyrosine was 12.6 against 100 of tyrosine, respectively. The rate of hydrolysis appeared to be decreased in the order of chlorine, bromine and iodine, that is, by increasing the atomic radius or by decreasing the electronegativity. But 3-iodotyrosine was not hydrolyzed by the enzyme. 4) The activity of ${\beta}-tyrosinase$ was inhibited by phenol markedly. Of the halogenated phenols, o-, or m-chlorophenol and o-bromophenol gave marked inhibition on the enzyme action, however inhibition by iodophenol was not strong. Plotting by Lineweaver-Burk method, a mixed-type inhibition by m-chlorophenol was observed and its Ki value was found to be $5.46{\times}10^{-4}M$. 5) During the synthesizing reaction of 2-bromotyrosine by the enzyme, sequential addition of substrate which was m-bromophenol with time intervals and in a small amount resulted in better yield of the product. 6) The halogenated tyrosines which were produced by ${\beta}-tyrosinase$ from pyruvate, ammonia and m-halogenated phenols were analysed to determine their molecular structures by element analysis, NMR spectroscopy, mass spectrometry, and IR spectroscopy. The result indicated that they were 2-chloro-L-tyrosine, 2-bromo-L-tyrosine, and 2-iodo-L-tyrosine, respectively.