Short communication: predominance of beta-casein (CSN2) C allele in goat breeds reared in Italy

A protocol for the rapid and simultaneous genotyping of A, C, and 0 'CSN2 alleles in goat was developed by single strand conformational polymorphism polymerase chain reaction (SSCP-PCR) technique. Screening the CSN2 variability in 7 goat breeds reared in Italy validated the genotyping test. The SSCP-PCR technique was also suitable for monitoring CSN2 polymorphism. In particular, the discrimination between CSN2*A and CSN2*C is important because the 2 corresponding protein variants cannot be separated by standard typing techniques. The monitoring of CSN2 variability in the goat breeds indicates the predominance of the C allele. In most breeds, CSN2*C occurred with the highest frequency, except in Saanen where CSN2*A and CSN2*C showed similar frequencies. Variant CSN2*C occurred with a frequency of 0.68 (Camosciata), 0.70 (Jonica), 0.71 (Garganica), 0.82 (Maltese), 0.87 (Cilentana), and 0.97 (Orobica). The alignment among the mature CSN2 sequences of different species suggests that CSN2*A is the ancestral allele compared with CSN2*C. Interestingly, the CSN2*A goat variant showed higher frequencies in selected breeds (Saanen and Camosciata).     

Short communication: the beta-casein (CSN2) silent allele C1 is highly spread in goat breeds

Several single nucleotide polymorphisms have been identified in the goat milk casein genes, most of them modifying the amino acid sequence of the coded protein. At least 9 variants have been found in goat β-CN (CSN2); 6 of them were characterized at the DNA level (A, A1, C, E, 0, and 0′), whereas the other 3 variants were described only at the protein level. The recently identified silent A1 allele is characterized by a C→T transition at the 180th nucleotide of the ninth exon. In the present work, typing results from different breeds (3 Italian, 3 German, and a composite of African breeds for a total of 335 samples) demonstrated that the same mutation is carried by the CSN2*C allele. In addition, the T nucleotide at the 180th nucleotide of the ninth exon was always associated with CSN2*C in all the breeds analyzed. Thus, another silent allele occurs at goat CSN2 and can be named CSN2*C1. The much wider distribution of C1 with respect to the A1 allele indicates that the single nucleotide polymorphisms characterizing the silent mutation originated from CSN2*C. A method for the identification of this allele simultaneously with 5 of the 6 DNA-characterized alleles is also proposed. The mutation involved codifies for the same protein of the C allele; nevertheless, its location in the 3′ untranslated region of the gene might affect the specific casein expression.     

Effect of kappa-casein polymorphism on milk composition in the Orobica goat

The aim of this work was to study the effects of isoelectrofocusing (IEF) milk protein variants on milk composition in the Italian Orobica goat breed, which is characterized by a rather high frequency of the κ-casein (CSN3) B^IEF allele. Significant associations were found between the IEF phenotype and protein and casein percentages. A favorable effect of the CSN3 B^IEF variant was found for both protein and casein percentages, with a codominance trend for the 3 phenotypes: BB > AB > AA. Depending on the selection purpose, emphasis could be given to different κ-casein variants in breeding. The high frequency of B^IEF could be exploited in breeding strategies to improve the protein and casein percentages when cheese making is a selection objective.     

Health implications of milk containing beta-casein with the A2 genetic variant

Milk from dairy cows has long provided a high quality source of protein and selected micronutrients such as calcium to most populations. Recently, a relationship between disease risk and consumption of a specific bovine ss-casein fraction either A1 or A2 genetic variants has been identified. Populations, which consume milk containing high levels of ss-casein A2 variant, have a lower incidence of cardiovascular disease and type 1 diabetes. Furthermore, consumption of milk with the A2 variant may be associated with less severe symptoms of autism and schizophrenia. The mechanism of action focuses on ss-casein A1 and related forms preferentially that are able to produce a bioactive opioid peptide, ss-casomorphin-7 (ss-CM-7) during digestion. Infants may absorb ss-CM-7 due to an immature gastrointestinal tract. Adults, on the other hand, appear to reap the biological activity locally on the intestinal brush boarder. ss-CM-7 can potentially affect numerous opioid receptors in the nervous, endocrine, and immune systems. Whether there is a definite health benefit to milk containing the A2 genetic variant is unknown and requires further investigation.     

Short communication: Simultaneous identification of five kappa-casein (CSN3) alleles in domestic goat by polymerase chain reaction-single strand conformation polymorphism

Until now, a total of nine polymorphic sites corresponding to six different alleles have been described at the κ-casein (CSN3) locus in the domestic goat (Capra hircus). A protocol for the rapid and simultaneous genotyping of five goat CSN3 alleles by using polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) technique was developed. Moreover, the developed test was validated by screening the CSN3 variability in four Italian breeds, Garganica, Jonica, Maltese, and Camosciata. Seven different patterns were readily identifiable. These corresponded to five known alleles and two newly identified variants. The G/A substitution at nucleotide position 471, which is not identifiable at the protein level but was found to be very frequent in the typed breeds, is easily detectable by the protocol developed. The PCR-SSCP analysis is a powerful tool for the genetic study of CSN3 variability in domestic goats, allowing both the simultaneous identification of different alleles, and the detection of new variants.     

Short communication: characterization of a new genetic variant in the caprine kappa-casein gene

A new polymorphism has been identified in the goat kappa-casein gene by evaluating genomic DNA from the Montefalcone breed in Italy. The polymorphic site consists of a single nucleotide substitution A to G at position 242 of the exon 4 and produces an amino acid substitution Asp/Gly. A polymerase chain reaction-restriction fragment length polymorphism protocol for rapid genotyping of the variant has been developed, using the HaeIII enzyme. Animals from Italian, Spanish, and French breeds have been analyzed to investigate the occurrence of the allele in other populations. The allele appears to be exclusive to the Montefalcone breed.     

山羊奶与牛奶和人奶营养成分的比较

对山羊奶、牛奶和人奶中蛋白质、脂肪、维生素、矿物质等主要营养成分进行了比较,分析了3种奶中主要营养成分的差别。通过比较发现,山羊奶在总体营养成分方面优于牛奶,更接近人奶。但山羊奶存在铁、叶酸和维生素B12含量较低以及羊奶膻味的问题,应在营养上对这方面加以重视。     

Evaluation of the allergenicity of goat milk, cow milk, and their lactosera in a guinea pig model

The object of this study was to determine the allergenicity of goat milk (GM) and cow milk (CM) and that of their respective lactosera (GML and CML), by in vivo and in vitro assays. Two systemic tests for anaphylaxis were carried out in guinea pigs, the animals being sensitized orally with the 2 types of milk and lactosera. Sera were taken from the orbital sinus of the experimental animals at 0 and 22 d of the experiment to perform the serological study and the passive cutaneous anaphylaxis test. For the latter, the guinea pigs were sensitized passively with antibodies against the 4 antigen solutions. Enzyme-linked immunosorbent assay and Western blot were used to determine the specific antibodies of the isotypes immunoglobulin G1 and immunoglobulin G(Fc) developed against the same 4 antigen solutions. From these anaphylaxis and antibody-production tests, it was concluded that GM is hypoallergenic when compared with CM. The lactosera produced more closely grouped results, with values always below those of the corresponding milk. None of the proteins in the 4 immunizing solutions were identified as being their main allergen. These results show the hypoallergenicity of GM versus CM, and also that both casein and lactoserum proteins may be responsible for allergy in each case. To analyze the possibility of producing an innocuous food for those allergic to milk proteins, it would be of interest to identify the epitope(s) responsible for such allergenicity.     

A study of beta-casein tertiary structure by intramolecular crosslinking and mass spectrometry

The objective of this study was to obtain experimental evidence to extend the discussion on the 3-D structure of beta-casein (beta-CN). The approach involved the preparation of homobifunctional crosslinkers, bis(sulfosuccinimidyl) derivatives of dicarboxylic acids of several lengths, which specifically react with primary amines of lysinyl residues or the N-terminal in the protein. The intramolecular crosslinks formed were determined by enzymatic digestion and by matrix-assisted laser desorption and ionization time-of-flight mass spectrometry combined with comparison against the theoretical digestion patterns. This procedure allowed the measurement of distances between the crosslinked residues. Ten different masses arising from 8 different specific intramolecular crosslinks were identified. Of these, 5 crosslinks were in good agreement with a published model (Kumosinski et al., 1993). Two other crosslinks each connected 2 residues that are much closer together, according to the model, than the maximum length of the crosslink. However, one of the crosslinks apparently connected 2 residues that are predicted by the model to be 16.7 A farther apart than the crosslink's stretched length. This disparity might be explained by structural flexibility. The structure expressed by the model is probably one of several energetically favorable conformations of the beta-CN molecule, whose structure is best described as rheomorphic rather than either a fixed structure or a random coil.     

掺假羊乳及其制品中牛乳的检测技术研究进展

羊乳具有营养价值高、蛋白质组成更接近人乳、脂肪球直径小及致敏性低等优点,更利于人体消化吸收,受到消费者和乳品企业的青睐.近年来我国羊乳产业发展迅速且潜力巨大,但由于受羊乳产量和养殖规模的限制,羊乳价格昂贵,市场中存在羊乳及其制品掺假牛乳的现象,且掺假手段多样,难以辨别.为了保证消费者的健康和权益,保障羊乳市场良性发展,...     

Short communication: Fluoroquinolone susceptibility of Staphylococcus aureus strains isolated from caprine clinical mastitis in southeast Spain

The antibiotic susceptibility of 32 strains of Staphylococcus aureus isolated from the mastitic milk of dairy goats was evaluated. The antibiotics tested were 3 fluoroquinolones that have been developed especially for use in veterinary medicine: danofloxacin, marbofloxacin, and orbifloxacin. Minimum inhibitory concentration (MIC) tests were performed according to the microdilution broth method. The MIC₉₀ (concentration at which 90% inhibition is achieved) values obtained were 0.5, 1, and 1 mg/L for danofloxacin, marbofloxacin, and orbifloxacin, respectively. Danofloxacin was the most active fluoroquinolone tested against Staph. aureus strains isolated from milk; however, specific testing is required before using these drugs as therapy for the control of clinical mammary infections in goats.     

Short communication: Phenotypic characterization of total antioxidant activity of buffalo,goat, and sheep milk

Free radicals are reactive and unstable waste molecules produced by cells, responsible of damages and alteration on DNA, proteins, and fat. The daily intake of antioxidant compounds, acting against free radicals and their detrimental effects, is essential for human health. Milk contains several compounds with antioxidant activity, and the sum of their reducing potential blocking free radicals development is defined as total antioxidant activity (TAA). This novel trait has been described in literature both in individual and bulk cow milk, but there are no reports from other dairy species. Therefore, the present study aimed to investigate phenotypic variation of TAA in individual samples of buffalo (n = 105), goat (n = 112), and sheep (n = 198) milk. Total antioxidant activity was measured through a reference spectrophotometric method, and expressed as millimoles per liter of Trolox equivalents (TE). The greatest TAA was observed in sheep milk, averaging 7.78 mmol/L of TE and showing also the broadest phenotypic variation expressed as coefficient of variation (13.98%). Significantly lower TAA values were observed for buffalo (7.35 mmol/L of TE) and goat (6.80 mmol/L of TE) milk, with coefficients of variation of 8.18 and 8.47%, respectively. Total antioxidant activity exhibited weak correlations with milk yield and chemical composition. Phenotypic values of TAA presented in this study will be used to assess the ability of mid-infrared spectroscopy to predict this new trait and thus to collect data at the population level.     

Short communication: Identification of two polymorphisms in the goat lipoprotein lipase gene and their association with milk production traits

Lipoprotein lipase (LPL) is a glycoprotein that plays a central role in plasma triglyceride metabolism by hydrolyzing triglyceride-rich chylomicrons and very low density lipoproteins. The activity of milk LPL has been shown to differ among several goat breeds, suggesting the existence of a genetic polymorphism influencing the functional properties of this enzyme. We have characterized the complete coding sequence of the goat LPL gene in 18 individuals belonging to 3 breeds. The coding region of the goat LPL cDNA was 1,437 bp long and encoded a protein of 478 amino acids. Moreover, we have identified 2 single nucleotide polymorphisms (SNP) including a G50C missense mutation, which involved a Ser→Thr amino acid replacement at position 17 of the signal peptide, and a C2094T substitution in the 3′ untranslated region. A univariate mixed model was used to evaluate the association between LPL genotypes and milk production and composition in 130 Murciano-Granadina goats. The G50C SNP was suggestively associated with milk fat content and tended to affect the milk dry weight basis. The C2094T SNP was not associated with any of the measured traits.     

Short communication: growth characteristics of Streptococcus uberis in UHT-treated milk

Streptococcus uberis is an important environmental pathogen associated with bovine mastitis as well as with high total bacterial numbers in bulk tank milk. This study was conducted to determine whether S. uberis reproduction is likely to contribute to high bacterial numbers in bulk tank milk. Four S. uberis raw milk isolates were individually inoculated into UHT-treated milk and incubated at 4.4 or 7 degrees C for up to 5 d to simulate appropriate cooling; at 10 degrees C for 5 d to simulate marginally inadequate cooling; at 21 or 25 degrees C for 7 h to simulate ambient temperatures; or at 32 degrees C for 7 h to simulate elevated temperature conditions. None of the S. uberis isolates grew at either 4.4 or 7 degrees C. Streptococcus uberis growth at 10 degrees C appeared to be ribotype-specific. Although ribotype 116-520-S-1 isolates did not grow at 10 degrees C, ribotype 116-520-S-2 isolate numbers increased up to 3.5 log10 cfu/mL within 5 d. Generation times were calculated as 2.7 +/- 0.1 h, 2.1 +/- 0.1 h, and 1.0 +/- 0.1 h for 116-520-S-1 isolates and 1.8 +/- 0.4 h, 1.3 +/- 0.3 h, and 0.8 +/- 0.1 h for 116-520-S-2 isolates at 21, 25, and 32 degrees C, respectively. Our results suggest that high numbers of S. uberis in bulk tank milk are more likely to reflect high numbers of S. uberis shed by mastitic cows, rather than multiplication of these organisms under cooling conditions required for production of Grade A milk.     

Beneficial effect of goat milk on nutritive utilization of iron and copper in malabsorption syndrome

The search for diets that improve the digestive and metabolic use of iron and copper in malabsorption syndrome led us to study goat milk for particular nutritional characteristics and compare it with cow milk, which is usually supplied. We studied the metabolism of iron and copper in transected rats (control) and in resected rats (resection of 50% of the distal small intestine). The diets used were the standard diet recommended by the American Institute of Nutrition and diets based on goat or cow milk. Intestinal resection reduced the apparent digestibility coefficient (ADC) of iron in the three diets tested. In the transected and resected rats, the ADC of iron was highest with the goat milk diet, followed by the standard diet and lowest with the cow milk diet. The ADC of copper was not affected by intestinal resection in the animals fed the goat milk diet, and was higher than that in the two groups of animals fed the other diets. Intestinal resection reduced the ADC of copper with the standard diet and the cow milk diet. When both groups of animals were fed the goat milk diet, the deposit of iron in the organs was greater than with those fed the cow milk diet and similar to that in those animals given the standard diet. The copper content in the kidneys was lower in the resected than in the transected animals, except in the case of those fed the goat milk diet, in which it was similar to that of the control (transected) rats. This study shows the beneficial effect of goat milk, with respect to cow milk, on the metabolism of iron and copper in control rats, especially those with malabsorption syndrome.     

牛羊乳酪蛋白制备及电泳分析比较

采采用新鲜和复原的牛羊乳为原料,等电点沉淀,通过洗涤、干燥等步骤分别制得牛乳和羊乳酪蛋白,用凯氏定氮法对其进行定量检测,应用十二烷基硫酸钠聚丙烯酰胺凝胶电泳（SDS—PAGE）分析比较牛羊乳酪蛋白组分差异。结果表明：牛乳酪蛋白得率为86．75％,羊乳酪蛋白得率为90．55％,高含量的酪蛋白主要集中在电泳图谱的中分子量组,可分为ds,CN、as2-CN、β-CN和K—CN,牛羊乳as2-CN分子量羊乳大于牛乳,牛乳a-CN含量比较多,羊乳β—CN含量比较多,鲜乳与复原乳全蛋白主要组分在电泳图谱中除酪蛋白差别外,上端的高分子量组清蛋白区羊乳IgG重链的分子量比牛乳小。应用蛋白质电泳分析技术可以区分羊乳和牛乳的蛋白质组分,等电点沉淀法制备酪蛋白的方法简单,易于操作。     

免疫学检测羊乳中掺入牛乳成分

由于营养、价格和原料等原因，市场上在羊乳中掺入牛乳的现象常有发生。针对这种乳源性掺假陆续产生了多种检测方法，本文主要综述了以ELISA为主的免疫学快速检测方法。     

Short communication: Effect of the addition of Bifidobacterium monocultures on the physical,chemical, and sensory characteristics of fermented goat milk

The aim of the study was to use 3 monocultures of Bifidobacterium (Bifidobacterium animalis ssp. lactis AD600, Bifidobacterium animalis ssp. lactis BB-12, and Bifidobacterium longum AD50) in fermented goat milk to assess the microbial, physicochemical, rheological, and sensory quality of beverages during a 3-wk storage period at 5°C. The results indicated that selected bifidobacteria may be used for production of fermented goat milk because they comply with the minimum standards specified by the Food and Agriculture Organization of the United Nations and the World Health Organization during the entire period of storage. However, goat milk fermented by Bif. longum AD50 had less than 10⁶ cfu/g after 21 d of storage. The acidity, acetaldehyde content, viscosity, and hardness of fermented goat milk beverages depended on the strain and the storage period. Sensory properties were similar and acceptable, with a tendency for the quality to be reduced with an extended storage time. Depending on the monoculture of bifidobacteria used to manufacture fermented goat milk, the product had a different pH value. Titratable acidity in all fermented goat milk increased significantly along with the time of storage. Our study has shown that monocultures of bifidobacteria had a significant effect on the content of acetaldehyde, but the lowest effect over the entire storage period was observed in goat milk fermented by Bif. animalis ssp. lactis BB-12. This sample also had the lowest viscosity values compared with other samples and the best organoleptic properties during a 3-wk storage period.     

Dependence of the interfacial behavior of beta-casein on phosphoserine residues

The role of the phosphoserine residues on the dynamical and structural properties of beta-casein was studied by molecular dynamics of the protein in water/lipid interfacial regions. The initial protein structure adopted in the modeling was that proposed for bovine beta-casein A2, where the five phosphoserine residues, originally present in its primary structure, were partially or totally substituted by serine residues. The simulations revealed a dependence of the interfacial behavior of beta-casein on the phosphorylation grade. When only partially dephosphorylated, the protein showed a similar behavior as that observed for the original beta-casein reported in previous work. During dynamics, the protein migrated from the aqueous environment towards the lipid medium, and remained attached to the interface separating both media. Quite different was the dynamics of the totally dephosphorylated beta-casein, that did not perceive the interface and immersed incessantly into lipid medium. The results suggest that the phosphoserine residues appear to be, in fact, intrinsically related to the mechanisms of beta-casein emulsion stabilization.     

Short communication: Temperature sensibility of Prototheca blaschkeae strains isolated from bovine mastitic milk

Dairy cow mastitis associated with microalgae of the genus Prototheca has been reported worldwide. This alga is extremely resistant to most antimicrobials commonly used in mastitis therapy. In milk processing, different thermal treatments are generally efficient at inactivating and eliminating microorganisms. Until recently, no reports on Prototheca blaschkeae susceptibility to heat treatment have been described. Thus, considering the potential zoonotic risk that Prototheca may represent, the objective of this study was to test the susceptibility of P. blaschkeae field isolates retrieved from bovine mastitis to different temperature/time ratios that are generally used in the milk processing industry: 62°C/15 min and 30 min; 70°C/20 s, 15 min, and 30 min; 75°C/20 s; 90°C/1 s; and 100°C/1 s. The results showed a growth reduction of all isolates after the heat treatments, but only at 100°C was a total growth inhibition observed.