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WALCZAK-WoJCIAK, M., KLUPCZYnSKI, J., MICInSKI, J., HOsEK, M.: Associations between microsatellite polymorphism within the macrophage expressed lysozyme (mLys) gene and milk Indices Properties in Polish Black-and-White Cows. Acta univ. agric. et silvic. Mendel. Brun., 2005, LIII, No. 2, pp. 99-106 The experiment was performed in the years 1998–2000 on two farms located near to Vistula Lowlands. The experimental materials were comprised of 52 daughters of bull Paran, among which 21 possessed the mLys-mic 7 allele and 31 – the mLys-mic 3 allele. The serum and whey bacteriolytic activity of the lysozyme, concentrations of selected mineral elements, technological properties of milk at successive the first lactation stages (30, 100, 150 and 200 days), somatic cell count and bacterial count were investigated in this study. The results obtained indicate that Lys-mic polymorphism has low suitability as a marker for milk production capacity in cows. The lysozyme gene, treated as a mastitis resistance factor, showed no effects on somatic cell count and bacterial count. Further investigations, conducted not only during the first lactation, but also during the next lactation in a herd threatened by a variety of pathogenic factors, would be carried to validate this Lys-mic gene polymorphism on mastitis resistance. alleles, lysozyme, mastitis, somatic cells, bacterial cells, lactation Rocnik LIII 10 Cislo 2, 2005 99 The genetic aspects of differentiation of the bacteriolytic activity of the bovine lysozyme have been already studied in Norway and Poland. At the beginning statistical detection methods were employed to confirm the presence of a hypothetical gene for high activity, denoted by the symbol LZM+. Three breeding bulls (one Red-and-White in Norway, two Blackand-White in Poland), independently segregating with high and normal capability to lyse bacterial cells, have been identified. The structure and organization and the bovine lysozyme gene have been described. A microsatellite fragment has been detected recently within the immuno-relevant lysozyme gene (mLys-mic) exhibiting expression in both macrophages and mammary gland tissue (Weikard et al., 1996). One of the polymorphic microsatellite variants, denoted by the symbol mLys-mic 7, showed an analogy to the phenotypically identified high-activity allele LZM+. Moreover, three mutations located in introns 2 and 3 have been identified. One of them, found at the position 8603 bp, also shows full compatibility with the hypothetical LZM+ allele. There100 M. Walczak-Wojciak, J. Klupczynski, J. Micinski, M. Hosek fore, DNA molecular markers enable the identification of genotypes determining differentiated phenotypic effect and, in consequence, directional mating in cattle herds, resulting in a threefold higher capability of bacterial pathogen destruction in the progeny (Steinhoff et al., 1994; Pareek et al., 2003; Prusinowska et al., 2003). The objective of the present study was to determine the associations between the occurrence of the mLysmic 7 allele, and concentrations of mineral elements and physicochemical properties of milk at successive lactation stages in primiparous cows. The activity of the serum and whey lysozymes, and the somatic cell and bacteria counts in milk, as dependent upon the activity of the Lys-mic 7 and Lys-mic 3 alleles, were also determined. MATERIAL AND METhoDS The experiment was performed in the years 1998– 2000, on Black-and-White cows kept under similar conditions on two farms in the Vistula Lowlands (Zulawy Wislane). The cows were free from tuberculosis, brucellosis and bovine leukemia. Both herds were characterized by comparable productivity. The cows were kept indoor and fed farm-made roughage, i.e. green forage (grass-alfalfa mixtures) in summer, and maize, grass and pulp silage supplemented with hay in winter. The cows were also given feed concentrate composed of ground grain, soybean meal and cake, supplemented with premixes. The experimental materials were comprised of daughters of bull Paran (51681 1 – 8). This bull was characterized by genetically differentiated bacteriolytic activity of the serum lysozyme, and was identified (Walawski et al., 1997) as a carrier of the allele for high lysozyme activity (LZM+/LZMo genotype) and a mLys-mic 7/ mLys-mic 3 heterozygote (Pareek et al., 1998). His semen (100 doses) was used for cow insemination on both farms. Both the number of heifers and bulls produced, and the presence of the sire allele, were purely accidental. Among 52 Paran’s daughters, one group (21 cows) possessed the sire mLys-mic 7 allele for high bacteriolytic activity, and the other (31 cows) – the mLys-mic 3 allele for normal activity, were identified according to Weikard et al. (1996). The heifers were successively introduced into the production herd and kept under the same environmental conditions. Tab. I presents the indices analyzed and methods used for their determination. Samples for analyses were taken at successive lactation stages, i.e. 30 days (stage I), 100 days (stage II), 150 days (stage III) and 200 days (stage IV). The serum and whey lysozyme activity was expressed as a percentage of lysis of Micrococcus lysodeicticus, according to the formula: % lysis = (EZ – EC) x 100% / EZ , where: Ez = extinction before incubation; Ec = extinction after 10 min. incubation. I: Indices analyzed and methods used for their determination Index Unit of measure Determination/analysis Authors Calcium g/l Spectophotometry Whiteside and Miner (1984) Phosphorus g/l Colorimetry Rutkowska (1981) Sodium g/l Photometry Rutkowska (1981) Potassium g/l Photometry Rutkowska (1981) Chlorides g/l Mohr method Budslawski (1973) Density oLd lacto-densitometry Manual Active acidity pH “PICCOLO –PLUS” pH–meter PN-68/A-86122 Clotting sec. Rennet method Budslawski (1973) Thermostability ml ethanol per 10 ml milk Ethanol method Polish Standard PN-68/A-86122 Somatic cell count 103 /ml “FOSSOMATIC” apparatus Manual Bacterial count 103 /ml “BACTOSCAN” apparatus Manual Lysozyme activity % lysis Micrococcus lysodeicticus Turbidimetric method Smolelis and Hartsell acc. to Metzger (Ślopek 1970) Associations between microsatellite polymorphism within the macrophage expressed lysozyme 101 The results were analyzed statistically (STATISTICA 6.0), by calculating arithmetic means (x) and standard deviations (Sd). The effects of mLys–mic polymorphism and lactation stage on productivity parameters, milk composition and properties, were verified by a two-factor analysis of variance in a nonorthogonal design and the Duncan test. In the case of somatic cell count and bacterial count the analysis was also performed for logarithmized values.

acta universitatis agriculturae et silviculturae mendelianae brunensis

Associations between microsatellite polymorphism within the macrophage expressed lysozyme (mLys) gene and milk Indices Properties in Polish Black-and-White Cows