Invited review: Human,cow, and donkey milk comparison: Focus on metabolic effects

Milk is an important food of the daily diet. Many countries include it in their dietary recommendations due to its content in several important nutrients that exert beneficial effects on human health. Human milk is a newborn's first food and plays an important role in the growth, development, and future health of every individual. Cow milk is the type of milk most consumed in the world. However, its relatively high content of saturated fats raises concerns about potential adverse effects on human health, although epidemiological studies have disproved this association. Indeed, dairy consumption appear to be linked to a lower risk of mortality and major cardiovascular disease events. In the last few years many researchers have begun to focus their attention on both the production and quality of cow milk as well as the analysis of milk from other animal species to evaluate their effect on human health. The need to investigate the composition and metabolic effects of milk from other animal species arises from the adverse reactions of individuals in several groups to certain components of cow milk. It has emerged that donkey milk compared with that of other animal species, is the nearest to human milk and an excellent substitute for it. Milk from various animal species shows substantial differences in nutritional composition and distinct metabolic effects. In this review, we discussed the main compositional features and metabolic effects of 3 types of milk: human, cow, and donkey milk.     

Hot topic: using a stearoyl-CoA desaturase transgene to alter milk fatty acid composition

Stearoyl-CoA desaturase enzyme converts specific medium- and long-chain saturated fatty acids to their monounsaturated form. Transgenic goats expressing a bovine beta-lactoglobulin promoter-rat stearoyl-CoA desaturase cDNA construct in mammary gland epithelial cells were produced by pronuclear microinjection. The fatty acid composition of milk from 4 female transgenic founders was analyzed on d 7, 14, and 30 of their first lactation. In 2 animals, the expression of the transgene changed the overall fatty acid composition of the resulting milk fat to a less saturated and more monounsaturated fatty acid profile at d 7 of lactation; however, this effect diminished by d 30. In addition, one animal had an increased proportion of the rumen-derived monounsaturated fatty acid C18:1 trans11 converted by stearoyl-CoA desaturase to the conjugated linoleic acid isomer C18:2 cis9 trans11. Milk that has higher proportions of monounsaturated fatty acids and conjugated linoleic acid may have benefits for human cardiovascular health.     

Expression and bioactivity of recombinant human lysozyme in the milk of transgenic mice

Human milk lysozyme is an important protein for innate immunity, but human breast milk is a fairly poor source for commercial production of this enzyme. Research on the expression of recombinant human lysozyme (rHlys) is therefore potentially valuable to the dairy industry. In this study, 2 different kinds of transgenic mice, PBC-hLY and PBC-sighLY, were generated and used as system models to express rHlys. Six lines of PBC-hLY transgenic mice with human lysozyme genomic DNA-based constructs were generated, and a maximum expression level of rHlys approaching 0.154 mg/mL was achieved. Antibacterial activity of the whey from PBC-hLY female transgenic mice was determined by a turbidimetric assay. Results showed that antibacterial activity of the whey was strongly enhanced, and confirmed that rHlys retained full activity. For rHlys to be secreted efficiently into the milk of transgenic mice, 5 lines of mice were also generated, in which the signal peptide DNA of bovine β-casein was substituted for that of lysozyme in PBC-hLY transgenic mice. Compared with PBC-hLY transgenic mice, both the expression levels of rHlys and the antibacterial activity of the whey were much higher in the PBC-sighLY transgenic mice. The concentration of rHlys in one of these mice amounted to 1.405 mg/mL—3 times higher than the level in human whey. The antibacterial activity of the whey was also 3 times higher than that of human whey. The rHlys from both PBC-hLY and PBC-sighLY transgenic mice had the same antibacterial activity as human milk lysozyme. The effect of the signal peptide and copy numbers of the transgene on expression of rHlys was also evaluated. This work will certainly permit a better understanding of how mammary gland bioreactor systems can be applied to produce rHlys in other mammals, such as cattle.     

Use of human lysozyme transgenic goat milk in cheese making: effects on lactic acid bacteria performance

Genetically engineered goats expressing elevated levels of the antimicrobial enzyme lysozyme in their milk were developed to improve udder health, product shelf life, and consumer well-being. The purpose of this study was to evaluate the effect of lysozyme on the development of lactic acid bacteria (LAB) throughout the cheese-making process. Raw and pasteurized milk from 7 lysozyme transgenic goats and 7 breed-, age-, and parity-matched nontransgenic controls was transformed into cheeses by using industry methods, and their microbiological load was evaluated. The numbers of colony-forming units of LAB were determined for raw and pasteurized goat milk, whey, and curd at d 2 and at d 6 or 7 of production. Selective plating media were used to enumerate lactococcal species separately from total LAB. Although differences in the mean number of colony-forming units between transgenic and control samples in raw milk, whey, and cheese curd were non-significant for both total LAB and lactococcal species from d 2 of production, a significant decrease was observed in both types of LAB among d 6 transgenic raw milk cheese samples. In pasteurized milk trials, a significant decrease in LAB was observed only in the raw milk of transgenic animals. These results indicate that lysozyme transgenic goat milk is not detrimental to LAB growth during the cheese-making process.     

Detection of recombinant human lactoferrin and lysozyme produced in a bitransgenic cow

Lactoferrin and lysozyme are 2 glycoproteins with great antimicrobial activity, being part of the nonspecific defensive system of human milk, though their use in commercial products is difficult because human milk is a limited source. Therefore, many investigations have been carried out to produce those proteins in biological systems, such as bacteria, yeasts, or plants. Mammals seem to be more suitable as expression systems for human proteins, however, especially for those that are glycosylated. In the present study, we developed a bicistronic commercial vector containing a goat β-casein promoter and an internal ribosome entry site fragment between the human lactoferrin and human lysozyme genes to allow the introduction of both genes into bovine adult fibroblasts in a single transfection. Embryos were obtained by somatic cell nuclear transfer, and, after 6 transferences to recipients, 3 pregnancies and 1 viable bitransgenic calf were obtained. The presence of the vector was confirmed by fluorescent in situ hybridization of skin cells. At 13 mo of life and after artificial induction of lactation, both recombinant proteins were found in the colostrum and milk of the bitransgenic calf. Human lactoferrin concentration in the colostrum was 0.0098 mg/mL and that in milk was 0.011 mg/mL; human lysozyme concentration in the colostrum was 0.0022 mg/mL and that in milk was 0.0024 mg/mL. The molar concentration of both human proteins revealed no differences in protein production of the internal ribosome entry site upstream and downstream protein. The enzymatic activity of lysozyme in the transgenic milk was comparable to that of human milk, being 6 and 10 times higher than that of bovine lysozyme present in milk. This work represents an important step to obtain multiple proteins or enhance single protein production by using animal pharming and fewer regulatory and antibiotic-resistant foreign sequences, allowing the design of humanized milk with added biological value for newborn nutrition and development. Transgenic animals can offer a unique opportunity to the dairy industry, providing starting materials suitable to develop specific products with high added value.     

Impacts of Milk Fraud on Food Safety and Nutrition with Special Emphasis on Developing Countries

Milk in its natural form has a high food value, since it is comprised of a wide variety of nutrients which are essential for proper growth and maintenance of the human body. In recent decades, there has been an upsurge in milk consumption worldwide, especially in developing countries, and it is now forming a significant part of the diet for a high proportion of the global population. As a result of the increased demand, in addition to the growth in competition in the dairy market and the increasing complexity of the supply chain, some unscrupulous producers are indulging in milk fraud. This malpractice has become a common problem in the developing countries, which lack strict vigilance by food safety authorities. Milk is often subjected to fraud (by means of adulteration) for financial gain, but it can also be adulterated due to ill‐informed attempts to improve hygiene conditions. Water is the most common adulterant used, which decreases the nutritional value of milk. If the water is contaminated, for example, with chemicals or pathogens, this poses a serious health risk for consumers. To the diluted milk, inferior cheaper materials may be added such as reconstituted milk powder, urea, and cane sugar, even more hazardous chemicals including melamine, formalin, caustic soda, and detergents. These additions have the potential to cause serious health‐related problems. This review aims to investigate the impacts of milk fraud on nutrition and food safety, and it points out the potential adverse human health effects associated with the consumption of adulterated milk.     

Milk oligosaccharides: A review

Milk oligosaccharides (OSs) confer unique health benefits to the neonate. Although human digestive enzymes cannot degrade these sugars, they support specific commensal microbes and act as decoys to prevent the adhesion of pathogenic micro‐organisms to gastrointestinal cells. The limited availability of human milk oligosaccharides (HMOs) impedes research into these molecules and their potential applications in functional food formulations. Recent studies show that complex OSs with fucose and N‐acetyl neuraminic acid (key structural elements of HMO bioactivity) also exist in caprine milk, suggesting a potential source of bioactive milk OSs suitable as a functional food ingredient.     

Milk biologically active components as nutraceuticals: review

Milk contains components that provide critical nutritive elements, immunological protection, and biologically active substances to both neonates and adults. Milk proteins are currently the main source of a range of biologically active peptides. Concentrates of these peptides are potential health-enhancing nutraceuticals for food and pharmaceutical applications. Several bioactive peptides may be used as nutraceuticals, for example, in the treatment of diarrhea, hypertension, thrombosis, dental diseases, as well as mineral malabsorption, and immunodeficiency. Minor whey proteins, such as lactoferrin, lactoperoxidase, lysozyme, and immunoglobulins, are considered antimicrobial proteins. Milk also contains some natural bioactive substances. These include oligosaccharides, fucosylated oligosaccharides, hormones, growth factors, mucin, gangliosides, and endogenous peptides, which are present in milk at secretion. Most of the claimed physiological properties of milk bioactive components have been carried out in vitro or in animal model systems, and these hypothesized properties remain to be proven in humans. Whether these milk bioactive components will replace drugs entirely in the immediate future is still unclear, but the increasing appreciation of "drug foods" or nutraceuticals plays a complementary rather than a substitutional role to the synthetic pharmacological drugs.     

Effect of rabbit kappa-casein expression on the properties of milk from transgenic mice

Transgenic mice were produced carrying the coding region of the rabbit kappa-casein gene linked to the upstream region of the rabbit whey acidic protein gene. Mice from the highest-expressing line produced 2.5 mg rabbit kappa-casein/ml in their milk. The foreign protein was associated with the casein micelles and altered micelle size, though in the high-expressing line rabbit kappa-casein also segregated into the whey fraction obtained after centrifuging the milk samples. Milk from transgenic mice had the same overall protein content as that from non-transgenic mice, except for the transgene product. However, litters fed with this transgenic mouse milk grew less well than litters given milk from non-transgenic mice. This reduction in growth was not related to changes in mammary gland structure or mammary cell morphology. Preliminary results indicated that milk from the transgenic mice had a higher viscosity.     

Hygienic and health characteristics of donkey milk during a follow-up study

For its characteristics, donkey milk has been proposed as an alternative to goat or artificial milk to feed allergic infants. Therefore, it is important to increase our knowledge on health and immunological characteristics of donkey milk. Ten donkeys, bred as companion animals, were enrolled in this study and sampled once a month, for eight months. Milk (10 ml) was collected from each half udder for somatic cell count (SCC), bacteriological analysis and total bacteria count (TBC). The major pathogens were tested for antimicrobial susceptibility, and Staphylococcus aureus isolates were further genotyped by nanoarray analysis. Whey lysozyme and NAGase (NAG) activities were also assessed. Overall, 101 half-udder milk samples were taken. They showed very low values of TBC (<250 cfu/ml) and SCC (<50 000 cells/ml) and a minor prevalence of pathogens: Staph. aureus was isolated only from 5 milk samples (3 animals), Streptococcus equi from 2 samples and Str. equisimilis from a single sample. All the isolates were sensitive to all antibiotic classes used in veterinary medicine. None of the Staph. aureus isolates were shown to harbour genes coding for any enterotoxin, toxic-shock syndrome toxin or antibiotic resistance. Lysozyme levels were always very high (4000-5000 U/ml), while NAG values were mostly low (<50 U/ml), out of the last part of lactation. The results of this study confirmed the low prevalence of intramammary infections in donkey and the absence of food-borne pathogens, suggesting that donkey milk could be a safe food, if the mammary gland is healthy and the animals are milked in proper hygienic conditions.     

Mycobacterium avium ssp. paratuberculosis in foods: current evidence and potential consequences

Mycobacterium avium ssp . paratuberculosis (MAP), the cause of Johne's disease in cattle, sheep and goats, may have a role in Crohn's disease in humans. Animals with Johne's disease shed viable MAP in their milk and faeces. The organism is also widely disseminated in the blood and tissues of infected animals. Consequently, transmission to humans via consumption of animal-derived foods is a distinct possibility. Milk, other dairy products, beef and water have been identified as possible food vehicles of transmission. To date, viable MAP has been cultured from raw cows', sheep and goats' milk, retail pasteurized cows' milk, and some retail cheeses in several countries during recent studies. MAP has not been isolated from retail beef to date, although limited testing has been carried out. The public health consequences, if any, of low numbers of viable MAP being periodically consumed by susceptible individuals are uncertain. An association between MAP and Crohn's disease is not proven, but neither can it be discounted on the basis of current evidence. A precautionary approach is therefore warranted in relation to the existence of MAP in food, and action is needed to reduce the prevalence of Johne's disease in the cattle population worldwide, in order to minimize public exposure to this potential human pathogen.     

Lactoferrin research,technology and applications

Lactoferrin is an iron-binding glycoprotein present in milk as well as other exocrine secretions and neutrophil granules in mammals. Lactoferrin is considered to be an important host defense molecule and has a diverse range of physiological functions such as antimicrobial/antiviral activities, immunomodulatory activity, and antioxidant activity. During the past decade, it has become evident that oral administration of lactoferrin exerts several beneficial effects on the health of humans and animals, including anti-infective, anticancer, and anti-inflammatory effects. This has enlarged the application potential of lactoferrin as a food additive. The technology of producing bovine lactoferrin on a factory scale was established over 20 years ago. Bovine lactoferrin is purified by cation-exchange chromatography from bovine skim milk or whey, and is commercially available from several suppliers. Recombinant human lactoferrin is produced by Aspergillus niger, transgenic cows, and rice, and its efficacy is being evaluated. In this article, we review basic research and technological aspects of the application of lactoferrin.     

Practical Food Safety Interventions for Dairy Production

Consumers are increasingly concerned about the safety of their food and uncertain about food production practices. Potential threats to human health related to dairy products and dairy farming include errors in pasteurization, consumption of raw milk products, contamination of milk products by emerging heat-resistant pathogens, emergence of antimicrobial resistance in zoonotic pathogens, chemical adulteration of milk, transmission of zoonotic pathogens to humans through animal contact, and foodborne disease related to cull dairy cows. Most dairy farmers feel responsible for the safety of milk and beef that originate on their farms, but linkage between farm production practices and the quality of processed products have been weak. The safety of dairy products can be enhanced by adoption of a number of management practices. Sources of microbial contamination of milk must be minimized by adoption of hygienic standards that can be easily evaluated. Uniform adoption of milking practices that reduce microbial contamination of milk should be emphasized. The diagnosis of salmonellosis or listeriosis on a dairy farm should be regarded as an indication that other potentially infected animals may be present in the herd. Coliform counts on bulk tank milk should be routinely performed as an indicator of fecal contamination. A reduction in the national regulatory limit for somatic cells in bulk tank milk should be considered based on potential enhancements in milk safety. Dairy farmers must take responsibility for the market cattle leaving their farms. The inappropriate or prophylactic use of antimicrobial agents must be minimized to ensure that antimicrobial resistance does not develop in animal pathogens. Consumers can have confidence in food safety programs on dairy farms that promote awareness and accountability for the products that are produced.     

A STUDY ON THE OCCURRENCE OF AFLATOXIN IN RAW MILK DUE TO FEEDS

Milk and milk products are the most important group of food which carries aflatoxin to the people. Therefore, the existence of aflatoxin M₁ (AFM₁) in milk is a potential risk for public health. AFM₁ existence in milk and milk products was found in studies performed in Turkey. In this study, aflatoxin contamination was detected in 48 raw milk and 48 feed samples. Milk and feed samples, in which the positive sample rate was determined as 24%, included different levels of aflatoxin. Altogether, 20 raw milk samples (41.67%) and 15 feed samples (31.25%) contained over the level of legal limits established by the Turkish Food Codex (TFC) and the European Communities (EC) regulation in positive samples. According to these results, the milk and feed samples collected from this area constitute a potential risk for public health. The most effective way of controlling aflatoxin M₁ in milk is to reduce the contamination of aflatoxin in raw milk and feed samples. Both the producers and the consumers, as well as the government, have important obligations on this subject.     

Short communication: effects of increased expression of alpha-lactalbumin in transgenic mice on milk yield and pup growth

Lactose synthase (a complex of beta1,4-galactosyltransferase and alpha-lactalbumin) forms lactose in the Golgi complex of mammary epithelial cells. To determine whether alpha-lactalbumin is a limiting component in this complex, transgenic mice that expressed bovine alpha-lactalbumin were studied. Transgenic mice produced 0.5 to 1.5 mg/ml of bovine alpha-lactalbumin in their milk, 5- to 15-fold more alpha-lactalbumin than in milk of control mice. Transgenic and control mice produced milk with the same concentrations of lactose, cream, and total solids, and showed similar mammary gland growth, morphology, and histology. Milk from transgenic mice had 0.6% less protein than milk from control mice (P < 0.05). The in vitro lactose synthase activity in mammary gland homogenates from alpha-lactalbumin transgenic mice was increased (P < 0.05), demonstrating that bovine alpha-lactalbumin could interact with murine beta1,4-galactosyltransferase. Pups reared by lactating transgenic mice showed a 4% increase in growth on d 10 of lactation, suggesting that milk production was increased (P = 0.06). Milk volume, estimated using the weigh-suckle-weigh technique, tended to be higher (although not significantly) in transgenic mice (P = 0.11). These results suggest that augmenting alpha-lactalbumin expression in the dam increases the growth of suckling offspring.     

Effects of animal selection on milk composition and processability

One goal of animal breeding is to increase the economic output through increased production, improved milk quality, and cow health. The objective of this study was to evaluate genetic progress in relation to milk composition, processability, and yield as a correlated response to selection for the Swedish breeding objective. Dairy cows with high genetic merit, classified as elite dairy cows, of the Swedish Red and Swedish Holstein breeds were used. Milk samples were collected on the farm level in winter and summer from a research herd at Nötcenter Viken, a bovine research farm in Sweden. Comparisons were made with milk from a Swedish Red herd, a Swedish Holstein herd, and a Swedish dairy processor in the same geographical area. Protein, lipid, and carbohydrate profiles as well as minerals were analyzed, and technological properties, including rennet-induced gelation characteristics, lipid oxidation, total antioxidant capacity, and fat globule size, were determined. Higher yields were found for elite cows for components of the protein, lipid, and carbohydrate profiles as well as for minerals, implying genetic progress in relation to milk yield; however, the content of some milk components (e.g., lipid and whey protein contents) had decreased on average. Milk from the elite cows had good gelation characteristics, but was more susceptible to lipid autooxidation and had a lower total antioxidant capacity. These results demonstrate that milk composition and processing characteristics could be used to adjust breeding practices to optimize the quality and stability of milk and dairy products.     

Technical note: A noninvasive method for measuring mammary apoptosis and epithelial cell activation in dairy animals using microparticles extracted from milk

Milk production from dairy animals has been described in terms of 3 processes: the increase in secretory cell numbers in late pregnancy and early lactation, secretion rate of milk per cell, and the decline in cell numbers as lactation progresses. This latter process is thought to be determined by the level of programmed cell death (apoptosis) found in the animal. Until now, apoptosis has been measured by taking udder biopsies, using magnetic resonance imaging scans, or using animals postmortem. This paper describes an alternative, noninvasive method for estimating apoptosis by measuring microparticles in milk samples. Microparticles are the product of several processes in dairy animals, including apoptosis. Milk samples from 12 Holstein cows, at or past peak lactation, were collected at 5 monthly samplings. The samples (n = 57) were used to measure the number of microparticles and calculate microparticle density for 4 metrics: annexin V positive and merocyanine 540 dye positive, for both and total particles, in both whole milk (WM) and spun milk. Various measures of milk production were also recorded for the 12 cows, including daily milk yield, fat and protein percentage in the milk, somatic cell count, and the days in milk when the samples were taken. A high correlation was found between the 4 WM microparticle densities and days in milk (0.46 to 0.64), and a moderate correlation between WM microparticle densities and daily milk yield (−0.33 to −0.44). No significant relationships were found involving spun milk samples, somatic cell count, or fat and protein percentage. General linear model analyses revealed differences between cows for both level of microparticle density and its rate of change in late lactation. Persistency of lactation was also found to be correlated with the WM microparticle traits (−0.65 to −0.32). As apoptosis is likely to be the major contributor to microparticle numbers in late lactation, this work found a noninvasive method for estimating apoptosis that gave promising results. Further investigation is required to find out the factors affecting microparticle production and how it changes throughout lactation.     

4 种哺乳动物乳中低聚糖的定性和定量分析研究进展

低聚糖是乳中的一类生物活性成分,具有益生元、抗病毒、抗炎及免疫调节等功能,可作为一种新型食品组分添加至特殊医学用途配方食品或功能性食品中,近年来受到了人们的广泛关注。哺乳动物乳是低聚糖的主要来源,前人围绕哺乳动物的初乳及成熟乳中的低聚糖种类及含量开展了相关研究,但是不同物种间低聚糖种类和含量的对比还鲜有报道。本文对4 种哺乳动物乳（山羊、牛、猪、骆驼）中的低聚糖种类和含量进行了总结及对比,旨在为动物乳源低聚糖的应用提供参考。     

Transgenic Over-Expression of Bovine α-Lactalbumin and Human Insulin-Like Growth Factor-I in Porcine Mammary Gland

The first week postpartum is the period of greatest loss for US swine producers, with most morbidity and mortality attributed to malnutrition and scours. Despite the benefit to be gained by improving lactation performance, little progress has been made in this area through conventional means. Transgenic technology provides an important tool for addressing the problem of low milk production and its detrimental impact on swine production. Transgenic swine over-expressing bovine α-lactalbumin (α-LA) and human IGF-I were developed. α-Lactalbumin was selected for its role in lactose synthesis and regulating milk volume. IGF-I regulates mammary development and piglet intestinal development. The IGF-I construct consisted of the IGF-I gene inserted directly behind the α-LA signal peptide coding sequence to allow secretion of IGF-I into milk. Outcomes assessed were milk composition and yield and piglet growth and intestinal development. First parity α-LA gilts (n = 8) had higher milk lactose content in early lactation and 20 to 50% greater milk yield on d 3, 6, and 9 of lactation than nontransgenic gilts (n = 10). Weight gain of piglets suckling α-LA gilts was greater from d 7 and 21 postpartum than control piglets. Colostral IGF-I was ∼10-fold higher in first-parity IGF-I transgenic gilts (n = 5) than non-transgenic gilts (n = 4), and elevated milk IGF-I is maintained throughout lactation at ∼0.6 mg/L. Milk yield tended to be greater in IGF-I transgenic swine in early lactation. Thus, transgenic over-expression of milk proteins and growth factors may provide a means to improve swine lactation performance.