Milk protein synthesis in the mammary gland involves expression of six major milk protein genes whose nutritional regulation remains poorly defined. In this study, the effect of long term under- and over-feeding on the expression of αs1-casein: CSN1S1, αs2-casein: CSN1S2, β-casein: CSN2, κ-casein: CSN3, α-lactalbumin: LALBA and β-lactoglobulin: BLG gene in sheep mammary tissue (MT) was examined. Twenty-four lactating dairy sheep, at 90–98 d in milk, were divided into three groups and fed the same ration, for 60 d, in quantities which met 70% (underfeeding), 100% (control) and 130% (overfeeding) of their energy and crude protein requirements. The results showed a significant reduction on mRNA of CSN1S1, CSN1S2, CSN2 and BLG gene in the MT of underfed sheep compared with the overfed ones and a significant reduction in CSN3 and LALBA gene expression compared with the respective control animals. Significant positive correlations were observed between the mRNA levels of milk proteins’ genes with the milk protein yield and milk yield respectively. In conclusion, the feeding level and consequently the nutrients availability, affected the milk protein yield and milk volume by altering the CSN1S1, CSN1S2, CSN2, CSN3, LALBA and BLG gene expression involved in their metabolic pathways.

Soybean oil (SBO) is rich in polyunsaturated fatty acids (FA) and rumen bypass of SBO can contribute to increase the polyunsaturated FA proportion in milk fat. Citrus pulp (CPP) is a source of antioxidants but there is little information on the effects of CP administration on milk properties. This study was performed to determine the role of rumen microorganisms in the transfer of antioxidants from CPP into milk when cows receive SBO as a source of polyunsaturated FA. Four ruminally fistulated lactating Holstein cows were assigned to a 4 × 4 Latin square design with a 2 × 2 factorial arrangement of treatments: (1) SBO administered in the rumen; (2) SBO infused in the abomasum; (3) SBO + CPP administered in the rumen; and (4) SBO + CPP infused in the abomasum. Product and site of administration had no effect on yield of milk components. Concentrations of total polyphenols and flavonoids, reducing power and production of conjugated diene (CD) hydroperoxides in milk were not affected by products, but infusion in the abomasum compared with administration in the rumen increased production of CD. Milk fat FA profile was not affected by products. However, cows infused in the abomasum compared with those administered in the rumen showed lower proportions of short-chain and monounsaturated FA and higher proportions of polyunsaturated, omega 3 and omega 6 FA in milk fat, which resulted in enhanced health-promoting index of milk. Administration of SBO and CPP (0·2 + 1·0 kg/d) in the rumen or the abomasum resulted in similar milk antioxidant properties, thus suggesting that the rumen microbes have little involvement in the metabolism of antioxidants from CPP.

An experiment was conducted during 6 weeks to evaluate effects of a reduced dietary level of protein-rich concentrates in a moderate dairy production system on cows’ performance, protein efficiency and milk quality including fatty acid profiles. Twenty-three lactating cows (Swiss Fleckvieh) were assigned either to a group receiving on average 2·4 kg/d individually fed concentrates (Prot+, n = 12) or to a group receiving no individually fed concentrates (Prot−, n = 11). All cows had ad-libitum access to a total mixed ration (TMR) mainly based on grass and maize silage, hay and little potatoes and soybean cake. In weeks 4–6 of the experiment, part of the hay was excluded from the TMR, and fed separately in the morning. Individual feed intake and milk yield were recorded during weeks 3 and 6 of the experiment; at the same time feed, faeces and milk samples were collected twice per week for analyses. Data were processed in linear mixed models. Omission of individual concentrates in Prot− was fully compensated by higher roughage intake in terms of dry matter. Crude protein (CP) and net energy intake was almost maintained. Despite a lower apparent CP digestibility in Prot−, the ratio of milk protein to ingested CP was the same in both groups, indicating a higher ruminal utilisation of degraded CP in Prot−. This corresponded with lower milk urea concentrations in Prot−. Milk quality was affected in terms of lower concentrations of linoleic and conjugated linoleic acid in milk fat of Prot−. Concentrations of odd- and branched-chain fatty acids in milk were increased in Prot−. Sequential offer of hay and TMR did not lead to considerable effects in intake, efficiency and milk quality. In conclusion, the results indicate that the efficiency of feed protein utilisation for milk protein is not impaired if concentrates are reduced in a moderate- to low-input dairy production system.

The influence of different pasture-based feeding systems on fatty acids, organic acids and volatile organic flavour compounds in yoghurt was studied. Pasture is the main source of nutrients for dairy cows in many parts of the world, including southeast Australia. Milk and milk products produced in these systems are known to contain a number of compounds with positive effects on human health. In the current study, 260 cows were fed supplementary grain and forage according to one of 3 different systems; Control (a traditional pasture based diet offered to the cows during milking and in paddock), PMR1 (a partial mixed ration which contained the same supplement as Control but was offered to the cows as a partial mixed ration on a feedpad), PMR 2 (a differently formulated partial mixed ration compared to Control and PMR1 which was offered to the cows on a feedpad). Most of the yoghurt fatty acids were influenced by feeding systems; however, those effects were minor on organic acids. The differences in feeding systems did not lead to the formation of different volatile organic flavour compounds in yoghurt. Yet, it did influence the relative abundance of these components.

In addition to the main components of the somatotrophic axis (GH/GHR/IGF-I/IGF-IR), great importance in the control of growth and development is also attached to the Janus kinase 2 (JAK2) pathway. Induced by the GH/GHR complex, JAK2 activates signal transducer and activator of transcription 5 (STAT5), and in consequence, may be involved in the regulation of expression of insulin-like growth factor I (IGF-I) in the mammary gland. Silent mutation (rs110298451) has been identified within exon 20 using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). A total of 904 individuals of four dairy or dual-purpose breeds (Polish Holstein-Friesian, Montbeliarde, Simmental and Jersey) were genotyped. A genotypic imbalance in the populations was observed. In the case of dual-purpose breeds (Montbeliarde and Simmental), the frequencies of both alleles were almost equal. In contrary, the JAK2G allele was predominant in the Polish Holstein-Friesian breed while JAK2A allele in Jersey. A pronounced relationship between JAK2/RsaI polymorphism and milk production traits was found where, irrespective of breed and lactation order, the GG genotype was significantly associated with higher milk, protein and fat yields, as compared to the AA genotype. Heterozygous individuals were generally characterised by intermediate values of the analysed milk traits. It can be argued that the JAK2 gene polymorphism is a potential marker for milk production traits. However, due to the fact that rs110298451 SNP does not directly affect amino acid sequence, other association studies involving missense mutation should also be performed.

Objective of the study was to record, by means of ultrasonographic examination, changes occurring during lactogenesis in the udder of healthy ewes and of ewes with pregnancy toxaemia. The work was carried out in 28 ewes, 16 with pregnancy toxaemia (group A) and 12 healthy controls (group B). B-mode and Doppler ultrasonographic examination of the udder of ewes was performed. During the last month of pregnancy, grey-scale intensity values of mammary parenchyma in group A were significantly greater than in group B (P = 0·007), as was also the progressive increase in grey-scale intensity values in both groups (P < 0·001). Blood mammary input was significantly greater in ewes of group B than in ewes of group A (P < 0·05), as was also the progressive increase in blood input in both groups (P < 0·001). Further, differences between the two groups were identified in pulsatility index (P = 0·007) and in mean blood velocity (P = 0·036), but only during the last fortnight of pregnancy. After lambing, grey-scale values decreased sharply compared to those in pregnancy (P < 0·01), whilst blood input, pulsatility index and mean blood velocity continued the same trend as at the last stage of pregnancy, with differences between the two groups still prevalent (P < 0·05). There was a reverse correlation between grey-scale intensity values and milk quantities (P < 0·035) and a correlation between blood input and milk quantities (P < 0·07). The progressive increase in the diameter of the external pudendal artery was significant (P < 0·001), but no significant differences were evident between the two groups (P > 0·35). Differences between group A and group B in all other haemodynamic parameters studied were not significant, neither throughout the last month of pregnancy (P > 0·25), nor during the first week of lactation (P > 0·06). However, their progressive changes during the last month of pregnancy were significant (P < 0·02).

Intramammary infection (IMI), comprises a group of costly diseases affecting dairy animals worldwide. Many dairy parlours are equipped with on-line computerised data acquisition systems designed to detect IMI. However, the data collected is related to the cow level, therefore the contribution of infected glands to the recorded parameters may be over estimated. The present study aimed at evaluating the influence of single gland IMI by different bacteria specieson the cow's overall milk quality. A total of 130 cows were tested 239 times; 79 cows were tested once and the others were examined 2–8 times. All of the analysed data refer to the number of tests performed, taking into account the repeated testing of the same cows. Of the cows tested ~50% were free of infection in all 4 glands and the others were infected in one gland with different coagulase negative staphylococci (CNS), Streptococcus dysgalactiae, or were post infected with Escherichia coli (PIEc), i.e., free of bacterial infection at the time of sampling but 1–2 months after clinical infection by E. coli. Overall, infection with bacteria had significant effects on somatic cell count (SCC) and lactose concentration. Examining each bacterium reveals that the major influence on those parameters was the sharp decrease in lactose in the PIEc and curd firmness in PIEc and Strep. Individual gland milk production decreased ~20% in Strep. dysgalactiae- and ~50% in PIEc-infected glands with respect to glands with no bacterial findings. Significant differences were found in lactose, SCC, rennet clotting time and curd firmness in the milk of infected glands and among those, these parameters were significantly higher in Strep. dysgalactiae and PIEc than in CNS infected cows. The current results using quarter-milking reinforces the importance of accurate IMI detection in relation to economic and welfare factors, and moreover, emphasises the need for technical sensing and constant reporting to the farmer about changes in the milk quality of every animal.

The biofilm-associated protein (Bap) of Staphylococcus aureus is a high molecular weight cell-wall-anchored protein involved in biofilm formation, first described in bovine mastitis strains from Spain. So far, studies regarding Bap were mainly based on the Spanish strain V329 and its mutants, but no information on the genetic variability of bap-positive Staph. aureus strains is yet available in the literature. The present study investigated the molecular characteristics of 8 bap-positive Staph. aureus strains from subclinical bovine mastitis, isolated in 5 herds; somatic cell counts (SCC) of milk samples were also registered. Strains were characterised using MLST, SPA typing and microarray and the results were compared with V329. All isolates from this study and V329 were assigned to ST126, t605, but some molecular differences were observed. Only herd A and B strains harboured the genes for β-lactams resistance; the leukocidin D/E gene, a type I site-specific deoxyribonuclease subunit, 3rd locus gene and serin-protease A and B were carried by all strains, but not by V329, while serin-protease E was absent in V329 and in another isolate. Four isolates and V329 harboured the fibronectin-binding protein B gene. SCC showed the highest value in the milk sample affected by the only strain carrying all the virulence factors considered. Potential large variability of virulence was evidenced among V329 and all bap-positive Staph. aureus strains considered: the carriage of fnb could enhance the accumulation of biofilm, but the lack of lukD/E and splA, B or E might decrease the invasiveness of strain.

Few studies have described the relationship between genotypic and phenotypic methods for detecting penicillin resistance in Staphylococcus aureus isolated from bovine intramammary infection (IMI). Six phenotypic methods for penicillinase detection were compared with a genotypic method testing the presence of the β-lactamase gene blaZ in Staph. aureus (n = 150) isolated from bovine IMI. Highest sensitivities and specificities were observed for disk diffusion (DD) (93 and 97·4%), minimum inhibitory concentration (MIC) (90·3 and 97·4%), Cefinase™ (85·9 and 97·4%) and Diatabs™ (85·7 and 98·7%). The estimated cut-off points estimated in the present study can be considered close to the ones indicated by CLSI (2013). The molecular detection of blaZ gene is the only method that may indicate the real or potential capacity of producing β-lactamase in Staph. aureus. Considering that from a clinical standpoint a false negative result from a phenotypic test is the most unfavourable situation, a combination of standard DD with Diatabs™ or Cefinase™ should be performed by routine mastitis laboratories to minimise false negative results.

This study aimed to describe chronological patterns of changes of various candidate blood components in milk during the acute phase of a mammary immune response in detail. Eight dairy cows were challenged with Escherichia coli lipopolysaccharide in one udder quarter. Milk from challenged and control quarters and blood samples were taken before, and 1 and 2 h after challenge and then every 15 min until 5 h after challenge. The SCC, serum albumin, immunoglobulin (Ig)G1, IgG2, lactate dehydrogenase (LDH), and L-lactate in milk and blood, and α-lactalbumin in blood were analysed. All selected parameters in milk increased in challenged quarters but did not increase in control quarters. Milk IgG1, IgG2, serum albumin, and LDH were already significantly increased at 2 h after challenge whereas a significant increase of SCC was detectable at 2·75 h and L-lactate was increased at 2·25 h after challenge. In blood L-lactate was increased at 3·75 h after challenge, however, other factors in blood did not change significantly within the 5 h of experiment. In conclusion, the increase of blood components in milk during inflammation follows two different patterns: There is a rapid increase for IgG1, IgG2, or LDH, before the increase of SCC, and their concentrations reach a plateau within 3 h. On the other hand, SCC and L-lactate show a slower but consistent increase not reaching a plateau within 5 h after LPS challenge. L-lactate increases to higher concentrations in milk than in blood. This clearly shows that the increase of blood components follows different patterns and is therefore a controlled and compound-specific process and not exclusively an unspecific type of leakage.

While the Complement protein system in human milk is well characterised, there is little information on its presence and activity in bovine milk. Complement forms part of the innate immune system, hence the importance of its contribution during milk ingestion to the overall defences of the neonate. A bactericidal sequestration assay, featuring a Complement sensitive strain, Escherichia coli 0111, originally used to characterise Complement activity in human milk was successfully applied to freshly drawn bovine milk samples, thus, providing an opportunity to compare Complement activities in both human and bovine milks. Although not identical in response, the levels of Complement activity in bovine milk were found to be closely comparable with that of human milk. Differential counts of Esch. coli 0111 after 2 h incubation were 6·20 and 6·06 log CFU/ml, for raw bovine and human milks, respectively – the lower value representing a stronger Complement response. Exposing bovine milk to a range of thermal treatments e.g. 42, 45, 65, 72, 85 or 95 °C for 10 min, progressively inhibited Complement activity by increasing temperature, thus confirming the heat labile nature of this immune protein system. Low level Complement activity was found, however, in 65 and 72 °C heat treated samples and in retailed pasteurised milk which highlights the outer limit to which high temperature, short time (HTST) industrial thermal processes should be applied if retention of activity is a priority. Concentration of Complement in the fat phase was evident following cream separation, and this was also reflected in the further loss of activity recorded in low fat variants of retailed pasteurised milk. Laboratory-based churning of the cream during simulated buttermaking generated an aqueous (buttermilk) phase with higher levels of Complement activity than the fat phase, thus pointing to a likely association with the milk fat globule membrane (MFGM) layer.

Fatty acids (FAs), tocopherols and retinoids were analysed in raw milk and cheese from six commercial sheep flocks monitored from early lactation in winter to late lactation in summer. In winter, animals received concentrate and forage indoors; in early spring, animals grazed part-time on cultivated or natural valley grasslands; and from mid spring on, animals were kept outdoors constantly on mountain natural pastures. Mountain grazing in late lactation significantly increased the amount of healthy desirable unsaturated FAs such as C18:1t11 (VA), C18:2c9t11 (RA), C18:2t11c13, C18:3c9c12c15 (ALA) and C20:5c5c8c11c14c17 (EPA), and those of α-tocopherol and α-tocotrienol of milk and cheese. Stepwise discriminant analysis was applied to classify cheese samples according to seasonal feeding management. The multivariate approach was able to discriminate beyond doubt mountain cheeses from those of indoor feeding and part-time valley grazing.

The aim of this study was to assess the effect of ovine bulk tank somatic cell count (BTSCC) on composition, proteose-peptone (p-p) content and casein fractions as indicating parameters for proteolysis and coagulating properties of milk. A total of 97 samples of bulk tank milk from Manchega breed ewe flocks were grouped according to somatic cell count (SCC) into four classes: fewer than 500 000 cells/ml, from 500 000 to 10 00 000 cells/ml, from 10 00 000 to 15 00 000 and more than 15 00 000 cells/ml. The casein : protein ratio and lactose content decreased with BTSCC. Proteolysis increased with BTSCC, causing a drop in β-casein and an increase in the γ-caseins from a concentration of 500 000 cells/ml. Regarding coagulation behaviour, the rennet clotting time (RCT) and firming time (k20) rose from 10 00 000–15 00 000 cells/ml of milk. The results showed that the impairment of milk quality and milk ability to make cheese as affected by intramammary infection (IMI) can be inferred from the bulk tank milk of flocks with poor udder health.

Bacteriocins are natural compounds used as food biopreservatives instead of chemical preservatives. Bifidobacterium animalis subsp. lactis (Bifid. lactis) was shown to produce a bacteriocin-like inhibitory substance (BLIS) able to inhibit the growth of Listeria monocytogenes selected as an indicator microorganism. To enhance this production by the strain Bifid. lactis BL 04, skim milk (SM) was used as a fermentation medium either in the presence or in the absence of yeast extract, Tween 80 or inulin as stimulating additives, and the results in terms of bacterial growth and BLIS production were compared with those obtained in a traditional high cost complex medium such as Man, Rogosa and Sharpe (MRS). To this purpose, all the cultivations were carried out in flasks at 200 rpm under anaerobic conditions ensured by a nitrogen flowrate of 1·0 L/min for 48 h, and BLIS production was quantified by means of a modified agar diffusion assay at low values of both temperature and concentration of List. monocytogenes. Although all these ingredients were shown to exert positive influence on BLIS production in both media, yeast extract and SM were by far the best ingredient and the best medium, respectively, allowing for a BLIS production at the late exponential phase of 2000 AU/ml.

β-Galactosidases from Kluyveromyces lactis and Kluyveromyces marxianus isolated from artisanal ewes’ milk cheeses, were used to transgalactosylate lactose from cheese whey permeate (WP). The content of galactooligosaccharides (GOS) obtained by transgalactosylation was comparable with that formed using pure lactose as substrate. In order to obtain a mixture with higher prebiotic oligosaccharide content, isomerisation of the transgalactosylated WP was carried out using sodium aluminate as catalyst. The transgalactosylated mixtures at 6 h of reaction contained amounts of prebiotic carbohydrates (tagatose, lactulose, GOS and oligosaccharides derived from lactulose, OsLu) close to 50 g/100 g of total carbohydrates for all the strains tested, corresponding to 322 g prebiotics/kg whey permeate. Thus, the suitability of this methodology to produce mixtures of dietary non-digestible carbohydrates with prebiotic properties from WP has been demonstrated, which is interesting for the food industry since it increases the value and the applicability of this by-product from cheese manufacture.

Iranian White cheese was manufactured from ultrafiltered cows’ milk using different concentrations of salt consisting of 1, 2·5, 4% and salt free. Chemical composition, proteolysis, counts for lactic acid bacteria and sensory evaluation were examined during 90 d of ripening. It was found that the use of different salt concentrations significantly influenced all chemical composition, proteolysis, total number of lactic acid bacteria and sensory characteristics of the cheeses. Increasing the salt concentrations caused a proportional decrease in proteolysis determined by both urea-PAGE of caseins and RP-HPLC of peptides. With increased salt concentration, total number of lactic acid bacteria decreased. Cheeses with 1 and 2·5% salt were suitable and acceptable in odour and flavour that may be due to the proportional level of proteolysis products. In conclusion, reducing salt concentration from 4 to 2·5 and 1% had no ineligible effect on the quality and acceptability of the cheese.

In this work, we studied the influence of the type of coagulant enzyme and the curd scalding temperature on the proteolysis and residual coagulant and plasmin activities of a cooked cheese, Reggianito, in the interest of reducing ripening time. A two-factor experimental design was applied in two levels: type of coagulant enzyme, bovine chymosin or camel chymosin, and curd scalding temperature, 50 or 56 °C. The experimental treatments were applied in Reggianito cheese making experiments, and the samples were ripened for 90 d at 12 °C. Scalding temperature influenced residual coagulant activity; the cheeses cooked at 50 °C had significantly higher activity than those treated at 56 °C. In contrast, scalding temperature did not modify plasmin activity. Proteolysis was primarily affected by curd cooking temperature because chymosin-mediated hydrolysis of αs1 casein was slower in cheeses treated at 56 °C. Additionally, the nitrogen content in the cheese soluble fractions was consistently lower in the cheeses scalded at 56 °C than those cooked at 50 °C. A significant influence of the type of coagulant enzyme was observed, especially in the nitrogen fractions and peptide profiles, which demonstrated that camel chymosin was slightly less proteolytic; however, these differences were lower than those caused by the scalding temperature.