Infusion of an internal teat sealant into the mammary gland of the dairy cow at drying off has been claimed to reduce the incidence of clinical mastitis over many months in the subsequent lactation, despite the absence of any ingredient of the sealant remaining for that long. However, these claims have been poorly substantiated, often by lack of identification on when the infection occurred in the period from calving to disease, if the infection was present at post calving sampling and if the pathogen causing the disease was that causing an earlier infection. Moreover, no hypothesis on how any effect on clinical mastitis might occur has been advanced in any of the publications claiming the effect. That the effect might occur is only reported in a minority of publications, and the possibility that this is relatively specific to Gram-negative pathogens is reviewed.

This research communication describes the effect of three automatic cluster removal (ACR) switch-point settings on machine-on time, overmilking duration, and the mean, peak-flow and overmilking vacuums. The objective was to reduce the overmilking and milking time, attaining a lower mean claw vacuum during overmilking. In a randomised experiment, 25 cows were subjected to three ACR switch-point settings at flow rates of 0.840 kg/min, 0.630 kg/min and 0.504 kg/min each for 15 morning-milkings. Pre-milking preparations and cow comfort were observed and evaluated, showing evidence that the switch-point setting of 0.840 kg/ml denoted the least proportion of behavioural problems (1.1% ± 0.6) compared to the settings at 0.640 kg/ml (16.0% ± 2.2) and 0.504 kg/ml (9.4% ± 1.7). A VaDia device (Biocontrol) determined the machine-on time and vacuum levels at the claw, mouthpiece and pulsator chamber. The machine-on time varied from 290 ± 72.8 (mean ± sd) to 289 ± 64.4 and 303 ± 66.3 for the 0.840, 0.630 and 0.504 kg/min settings, respectively. Overmilking was shortened by 29.3% and 26.4% and the claw vacuum during overmilking was reduced by 3.7 kPa and 4.3 kPa when using the 0.840 kg/min switch-point setting, compared to the others. An increased cluster-removal milk flow threshold reduced both overmilking and vacuum level during overmilking, lowering the risk of teat damage. Adapting ACR switch-point settings offers a valuable opportunity to increase parlour efficiency and cow comfort in dairy herds.

This research communication investigated the role and the underlying mechanism of sn-1-acylglycerol-3-phosphate O-acyltransferase 6 (AGPAT6) in acetate-induced mTORC1 signaling activation and milk fat synthesis in dairy cow mammary epithelial cells. The data showed AGPAT6 knockdown significantly decreased acetate-induced phosphorylation of mTORC1 signaling molecules and intracellular triacylglycerol (TAG) content, whereas this inhibition effect was reversed after the addition of 16:0,18:1 phosphatidic acid (PA), suggesting that AGPAT6 could generate PA in response to acetate simulation, that in turn activates mTORC1 signaling. PPARγ is the upstream regulator of AGPAT6 upon acetate stimulation. Luciferase assay with clones containing various deletions and mutation in AGPAT6 promoter showed that there is a RXRα binding sequence located at −96 bp of AGPAT6 promoter. Acetate stimulation significantly increased the interaction between PPARγ and AGPAT6 via this RXRα binding site. Taken together, our data indicated that AGPAT6 could activate mTORC1 signaling by producing PA during acetate-induced milk fat synthesis, and PPARγ acts as a transcription factor to mediate the effect of acetate on AGPAT6 via RXRα.

The main objective of this experiment was to evaluate the effects of two milking intervals (8 and 16 h) on milk constituents (fat, protein, lactose, dry matter, and log10 SCC) of nineteen Istrian × Awassi × East-Friesian crossbred ewes in different milk fractions (0–25, 25–50, 50–75 and 75–100%) during the course of milking and in machine stripping (MS) milk. Furthermore, we sought to determine the effect of the two milking intervals on milking characteristics (average milk flow rate, peak milk flow rate, machine-on time, total milk yield, and milk production rate) and whether each milk constituent within each milking interval is best described by a linear, quadratic, or cubic function. Average milk flow rate and milk yield per milking decreased in the 8 h milking interval compared to the 16 h milking interval (P < 0.05). Peak milk flow rate, machine-on time, and milk production rate were not different between the two milking intervals. Overall, milk fat content, dry matter content, and log10 SCC increased in the 8 h milking interval compared to the 16 h milking interval (P < 0.05). Milk protein content did not change through the main milk fractions at either milking interval. Milk lactose content did not change through the milk fractions at the 8 h milking interval, whereas it decreased in the 75–100% and stripping milk fractions at the 16 h milking interval (P < 0.05). The 0–25% and stripping milk fractions contained the highest log10 SCC compared to all other milk fractions (P < 0.05). Changes of milk fat and dry matter content throughout milking were best described by quadratic functions, whereas milk protein content, milk lactose content, and log10 SCC were best described by different functions depending on the milking interval. These results demonstrate that milking interval influenced all milk constituents in various milk fractions during the course of ewe milking. Moreover, milking characteristics such as average milk flow and total milk yield, and the appropriate mathematical function to characterize milk constituents throughout a milking, were affected by milking interval.

Streptococcus agalactiae (S. agalactiae) infection is a significant cause of mastitis, resulting in loss of cellular homeostasis and tissue damage. Autophagy plays an essential function in cell survival, defense, and the preservation of cellular homeostasis, and is often part of the response to pathogenic challenge. However, the effect of autophagy induced by S. agalactiae in bovine mammary epithelial cells (bMECs) is mainly unknown. So in this study, an intracellular S. agalactiae infection model was established. Through evaluating the autophagy-related indicators, we observed that after S. agalactiae infection, a significant quantity of LC3-I was converted to LC3-II, p62 was degraded, and levels of Beclin1 and Bcl2 increased significantly in bMECs, indicating that S. agalactiae induced autophagy. The increase in levels of LAMP2 and LysoTracker Deep Red fluorescent spots indicated that lysosomes had participated in the degradation of autophagic contents. After autophagy was activated by rapamycin (Rapa), the amount of p-Akt and p-mTOR decreased significantly, whilst the amount of intracellular S. agalactiae increased significantly. Whereas the autophagy was inhibited by 3-methyladenine (3MA), the number of intracellular pathogens decreased. In conclusion, the results demonstrated that S. agalactiae could induce autophagy through PI3K/Akt/mTOR pathway and utilize autophagy to survive in bMECs.

Teat dipping is widely used in dairy cattle, especially to protect against contagious mastitis. Here we determine the effect of the device called ‘Electrical Teat Dipping’ (ETD), which was developed by combining teat dipping application and electrical field stimulation technique on teats. For this purpose, the front teats of 100 Holstein breed milking cows were evaluated in two groups, with ETD being applied once to the left front teat of these cows, and conventional teat dipping (CTD) being applied once to the right front teat, both after milking. Ultrasonographic measurements of the teats were made before milking and after teat dipping. We found that the width of the teat canal (1.88 ± 0.07 mm) in the teat using ETD was narrower after the application compared to those with CTD (2.28 ± 0.05 mm). Based on our findings, we conclude that the effects of ETD on the teat are very positive and can potentially be used as a new approach in the preventative control of mastitis in cows.

The main objective of this research communication was to explore the extent of milkability changes caused by the incidence of clinical mastitis. Our second objective was to investigate if the milkability of cows shortly before mastitis incidence significantly differed in comparison with healthy cows. Milk yield, milking time, average milk flow, partial milk flows and the occurrence of bimodal milk flows were monitored during the first 120 d in milk for all cows that calved on the farm during the experimental period (n = 127). A veterinarian diagnosed 27 cows with clinical mastitis, while the remaining healthy cows served as a control group. The period surrounding the mastitis was monitored in a timeline from 2 weeks before to 4 weeks after the diagnosis. We did not observe any significant differences in milkability between the healthy cows and the cows in the pre-mastitis period, which suggests that monitoring these might not be useful for early mastitis detection. Milk yield and milk flow were significantly decreased in the week of mastitis diagnosis compared to the pre-mastitis period. Milking time was not affected in the week of diagnosis, but was significantly increased in the following week, because milk yield returned to pre-mastitis values, but average milk flow was still significantly decreased. Milk yield was normalized to pre-mastitis values as the first of monitored parameters. Milk flow returned to pre-mastitis values in the fourth week after mastitis diagnosis. Milking time was the only parameter that was significantly affected up to a month after diagnosis. Our results showed that milkability changes caused by mastitis are significant and should not be ignored. The ability to adjust milking settings for cows diagnosed with mastitis could become a useful tool for improving mastitis treatment.

Sow milk fats not only provide energy but also essential nutrients for piglets. Thus, feeding strategies must be aligned with fat composition, especially triacylglycerols (TAGs) and their isomers. The triacylglycerol (TAG) profiles of sow milk fats from five typical breeds (Landrace × Large White, Landrace, Large White, Duroc, Pietrain) and two lactation stages (colostrum and milk) were systematically studied. A total of 45 major TAG species were identified using ultra-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry. The most abundant TAG was oleic acid-palmitic acid-linoleic acid (O-P-L) (13.92% and 12.03% in colostrum and milk, respectively), which was not significantly different in colostrum among all breeds. TAG composition of sow milk was affected mainly by the lactation stage rather than sow breed. Furthermore, TAG compositions of sow milk fats were similar to those of human milk fats, but significant differences were observed between commercial piglet formulas and sow milk. Therefore, the results will contribute to the optimization of piglet formulas to improve the growth and wellness of piglets, as well as potentially providing a basis for food usage as a new source of nutrients for human infants in future.

The objective of this research communication was to produce low potassium milk in which other electrolyte changes and changes in taste were minimized. To reduce potassium concentrations, several studies have reported batch methods of directly mixing milk or formula with sodium polystyrene sulfonate, which can exchange cations such as potassium for sodium. However, they also reported increases in sodium content, decreases in calcium and magnesium content, and changes in taste, because sodium polystyrene sulfonate exchanged other substances such as calcium and magnesium for sodium. In the present study, a method of dialyzing whole cow's milk using both sodium polystyrene sulfonate and a small amount of water through cellophane membranes was developed. A batch method for comparison was also performed. Each milk sample was evaluated biochemically and analyzed for taste and aroma in a sensory analysis. We showed that the potassium concentration in the dialyzed milk was reduced to 38% of that in unreacted milk. It was also shown that changes in sodium (increased) as well as calcium and magnesium (decreased) in the dialyzed milk were less than half of those in the batch method milk. Sensory analysis showed that minimal changes occurred in the taste of the dialyzed milk.

This research communication reports concentrations of two sialic acids (SA), N-acetylneuraminic (Neu5Ac) and N-glycolylneuraminic (Neu5Gc), in fresh milk from different cow breeds throughout lactation. According to published studies, the two SA types found in animal-derived products have diverse and conflicting effects on human health, but SA content is not routinely analysed in individual milk cows samples. We measured the content of Neu5Ac and Neu5Gc in milk from Holstein Friesian (HO), Simmental (SM), Simmental × Holstein crossbred (SM×HO), and Podolica (POD) cows at 60 and 120 d following calving. HO, SM and SM×HO were reared in an intensive production while POD were raised in an extensive system. Results showed that total Neu5Ac was overall thirty times more abundant than Neu5Gc, and their concentrations were higher at 120 d than at 60 d (P < 0.001). Neu5Gc values were greater in HO, SM, and SM × HO than in POD (P < 0.001), while HO had a higher Neu5Ac value than the other three breeds (P < 0.001). These findings shed light on the differences in SA content among cow breeds and lay the groundwork for future research to select animals that produce milk with desirable characteristics for human health.

We wished to determine if Mycoplasma bovis infection can negatively impact milk quality and production in Holstein dairy cows. For this Research Communication, milk samples (271) from Holstein cows from 3 herds were screened for M. bovis by real-time PCR. Positive (n = 21) and negative animals (n = 21) were matched by herd, age, lactations and days in milk (DIM). Pairs were evaluated in 7 stages of lactation: D1–50, D51–100, D101–150, D151–200, D201–250, D251–300, and D ≥ 301. A mixed model was used to assess the effect of groups (M. bovis+ × M.bovis−), time (lactation) and groups × time interaction. Cows positive for M. bovis had lower average milk production per day and high somatic cells count (SCC).

This research communication aims to characterize the prevalence, molecular characterization and antimicrobial resistance profiling of Streptococcus agalactiae isolated from clinical mastitis in China. A total of 140 Strep. agalactiae isolates were identified from 12 out of 201 farms in 6 provinces, overall herd prevalence was 18.6% and the MLST analysis showed clonal complexes (CC) 103 and CC 67 were present in these herds with CC 103 predominant, accounting for 97.9%. Isolates were mostly sensitive to the tested antimicrobials: penicillin, ceftiofur, amoxi/clav, cefquinome, and vancomycin (100%), followed by cefalexin (97.9%), oxacillin (96.4%), enrofloxacin (95.7%), erythromycin (89.3%), and clindamycin (88.6%). Only 19.3 and 0.7% of isolates were sensitive to tetracycline and daptomycin, respectively, and sequence type (ST) 103 was most resistant to antimicrobials. In conclusion, CC 103 was the predominant subgroup of bovine mastitis Strep. agalactiae in China, and most antimicrobials apart from tetracycline and daptomycin were effective.

This study evaluated the impact of three distinct diets; perennial ryegrass (GRS), perennial ryegrass/white clover (CLV) and total mixed ration (TMR), on the sensory properties and volatile profile of whole milk powder (WMP). The samples were evaluated using a hedonic sensory acceptance test (n = 99 consumers) and by optimised descriptive profiling (ODP) using trained assessors (n = 33). Volatile profiling was achieved by gas chromatography mass spectrometry using three different extraction techniques; headspace solid phase micro-extraction, thermal desorption and high capacity sorptive extraction. Significant differences were evident in both sensory perception and the volatile profiles of the WMP based on the diet, with WMP from GRS and CLV more similar than WMP from TMR. Consumers scored WMP from CLV diets highest for overall acceptability, flavour and quality, and WMP from TMR diets highest for cooked flavour and aftertaste. ODP analysis found that WMP from TMR diets had greater caramelised flavour, sweet aroma and sweet taste, and that WMP from GRS diets had greater cooked aroma and cooked flavour, with WMP derived from CLV diets having greater scores for liking of colour and creamy aroma. Sixty four VOCs were identified, twenty six were found to vary significantly based on diet and seventeen of these were derived from fatty acids; lactones, alcohols, aldehydes, ketones and esters. The abundance of δ-decalactone and δ-dodecalactone was very high in WMP derived from CLV and GRS diets as was γ-dodecalactone derived from a TMR diet. These lactones appeared to influence sweet, creamy, and caramelised attributes in the resultant WMP samples. The differences in these VOC derived from lipids due to diet are probably further exacerbated by the thermal treatments used in WMP manufacture.

The low levels of toxicity and cytoprotective effect attributed to Achyrocline satureioides (Lam.) DC, a medicinal plant native to South America, are of interest for bovine mastitis therapy. This research paper reports the hypothesis that a nanoemulsion of macela extract (Achyrocline satureioides) exerts protective effects on bovine mammary alveolar cells -T (MAC-T) and increases the permeation of flavonoid compounds through mammary epithelium. Extract-loaded nanoemulsions (2.5 mg/ml) (NE-ML) (n = 4) were prepared using high-pressure homogenization with varying concentrations of flaxseed oil and Tween 80. Permeation and retention of free and nanoencapsulated quercetin, 3-O-methylquercetin and luteolin were performed on mammary glandular epithelium using Franz diffusion cells. The cell viability was evaluated on mammary epithelial cells (MAC-T lineage) using the MTT method (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) after exposure to loaded and blank nanoemulsions (NE-ML and NE-BL). Necrotic or apoptotic cell death was evaluated by flow cytometry after exposure to nanoemulsions (NE-ML and NE-BL). Subsequently, the cell death was assessed by previously treating MAC-T cells with NE-ML for 23 h, followed by exposure to H2O2 (2 mM) for 1 h. Higher permeation of quercetin and 3-O-methylquercetin in NE-ML was found compared to that of free extract with a final permeated amount of 50.7 ± 3.2 and 111.2 ± 0.6 μg/cm2 compared to 35.0 ± 0.6 and 48.9 ± 1.2, respectively. For NE-BL, the IC50 was at least 1.3% (v/v), while for the NE-ML, it was at least 2.6% (v/v). After exposure to NE-ML (5 and 1.2%, v/v), the percentage of apoptotic cells was reduced (±30%). For the H2O2 assay, the percentage of cells in necrosis was reduced by 40% after exposure to NE-ML1% (v/v) + H2O2 2 mM. The protective effects and increased permeation of macela nanoemulsion make this a promising new candidate for bovine mastitis therapy.

This research paper assessed textural, microstructural, sensory and colour properties of set-yoghurts produced using milk from two indigenous cattle types, Thamankaduwa White (TW) and Lankan cattle (LC) compared to two generic cattle breeds, Friesian and Jersey. Instrumental texture profile (firmness, adhesiveness, cohesiveness and springiness), colour space (L* a* b*) and scanning electron micrographs of set-yoghurts during 21 d of storage (4 ± 1°C) were evaluated. Sensory quality attributes were evaluated with 40 untrained panellists using a five-point hedonic scale. Set-yoghurts prepared using indigenous cow milk showed higher (P < 0.05) firmness, cohesiveness and apparent viscosity values compared to those prepared using generic cow milk. As revealed by micrographs, set-yoghurts made from TW milk had lesser and smaller void spaces and a dense protein gel network than gels made from LC and the two generic breeds. The gel network made from Friesian milk showed a comparatively larger porous gel structure and thinner protein strands resulting in a weaker gel than other milk gels. The highest lightness (L*) and yellowness (b*) were observed from set-yoghurt produced from Friesian and LC milk, respectively. Set-yoghurts from TW milk had the highest (P < 0.05) sensory scores for all sensory attributes. The lowest sensory acceptance was recorded in set-yoghurt made from Friesian milk. Thus, milk from TW and LC is likely to be suitable in producing set-yoghurts with superior textural, microstructural and sensory properties, compared to milk from Jersey and Friesian. Our results suggest the merits of using indigenous cow milk in producing set-yoghurts and, thereby, prioritizing the preservation of the genetic pool of these indigenous breeds.

Bovine κ-casein glycomacropeptide (GMP) is a sialic acid containing glycopeptide, which is considered as a health promoting compound found in cheese whey. The study described in this research communication was undertaken to determine whether GMP with undetectable level of contaminating protein or phenylalanine can be isolated from bovine whey fraction using batch anion exchange technique with chitin as an adsorbent. A soluble whey fraction (SWF) prepared from 1 g whey protein isolate (WPI) was mixed with a slurry of 1 g chitin, and the mixture was incubated at pH 3.0. After incubation, the mixture was filtered, and the residue obtained (containing chitin-GMP complex) was washed with water and eluted stepwise with 0.5 M NaCl and 2.0 M NaCl. Most of GMP (corresponding to 75.8% of total sialic acid recovered) was eluted with 0.5 M NaCl. The recovered GMP accounted for 5.4% dry weight of WPI (or 18.9% dry weight of SWF). Amino acid analysis showed that there was no detectable level of contaminating amino acids including phenylalanine, histidine, arginine and tyrosine in the GMP fraction. It was concluded that the batch anion exchange method with chitin developed in this study can be used for the isolation of high purity GMP from bovine SWF.

In this research communication the genetic diversity of Pseudomonas fluorescens (n = 67) and Pseudomonas putida (n = 44) isolated from refrigerated raw milk from bulk tank trucks were verified. The relationship between the genetic profile of the isolates and their lipoproteolytic potential was evaluated using skim milk agar and tributyrin agar (21°C/72 h). The lipoproteolytic potential (low or high), evaluated by the diameter of the halos (cm), was correlated with the number of milk producing properties that contributed to each sample (one sample = one bulk tank truck; 8−80 producers/sample) and the distance between the dairy properties and the processing plant (21−370 km). P. fluorescens was confirmed in all samples, while P. putida in 60% samples. For both species, two clusters (I and II) were observed, and the first one showed lower genotypic diversity and the presence of isolates with 100% similarity. P. fluorescens isolates presenting at least 70% similarity were 83.9% in Cluster I (n = 31) and 44.4% in Cluster II. In both clusters (I and II) observed in the P. fluorescens dendrogram, the occurrence of high proteolytic and lipolytic potential were equivalent. The higher the number of farms per milk sample, the greater the lipoproteolytic intensity of P. fluorescens isolates. In relation to P. putida isolates, 74% presented at least 50% similarity in Cluster I (n = 27) and only 35% in Cluster II (n = 17). The occurrence of high proteolysis linked to P. putida was proportional between both Clusters, but the occurrence of high lipolysis was greater in Cluster II. No significant association was detected between P. putida isolates and the variables studied. The results indicate the circulation of P. putida and P. fluorescens with 100% similarity in different milk producing regions. The level of genetic diversity was related only to the lipolytic capacity of P. putida.

In this study, we developed and optimized a growth media by evaluating various nitrogen sources for the cultivation of Lactobacillus bulgaricus, a probiotic and an important dairy starter culture. We modified the composition of deMan, Rogosa and Sharpe (MRS) culture media and substituted the nitrogen content with alternative nitrogen sources X-Seed KAT, X-Seed Carbo Max and X-Seed Nucleo Max in various blends of 5 g/l and 10 g/l respectively. Results showed that bacterial growth was significantly higher when the nitrogen source blend KCMax (10/10) was used. The optical density (OD610 nm) of the Lactobacillus bulgaricus strains were higher (1.34 and 1.79) in the KCMax (10/10) medium than in the MRS medium (0.89 and 1.42) (P < 0.05). There was no significant difference in the bacterial counts for both the MRS medium and the KCMax (10/10) medium, and all bacterial counts were estimated at 8 log CFU/ml. The buffering capacity of KCMax (10/10) was also tested and supplemented with l-histidine and was significantly different (P < 0.05) than that of the MRS control medium. Calcium supplemented in the KCMax (10/10) also served as a cryoprotectant for the cells during freezing and freeze-drying. Bacterial counts of the recovered calcium-treated freeze-dried cells were statistically significant (P < 0.05). We hypothesized that alternative nitrogen sources such as selected yeast extracts from the X-Seed brand of complex nitrogen sources could efficiently support the viability of Lb. bulgaricus. Our results thus suggested the growth of Lb. bulgaricus was efficiently supported by the X-Seed KAT, X-Seed Nucleo Max and X-Seed Carbo Max nitrogen sources. Consequently, these alternative nitrogen sources could potentially be recommended for dairy starter culture fermentations.

We investigated the use of near-infrared spectroscopy (NIR) for measuring water-holding capacity (WHC) in fermented milk. Increased WHC ensures improved texture and decreased syneresis in fermented dairy products and also improves cheese yield. NIR combined with partial least-squares-discriminant analysis (PLS-DA) was found to be a promising rapid and non-invasive method with no pretreatment of the samples for prediction of WHC in fermented milk samples. Analysis of the chemical bonds in the region 10 700–4500 cm−1 (935–2200 nm) of the electromagnetic spectrum was able to distinguish between samples with high vs. low WHC. This technique was successfully used to screen different strains of lactic acid bacteria for their ability to provide fermented milk with increased WHC, which is of great importance for use in various dairy products.

Lactoperoxidase (LPO) is a glycosylated antimicrobial protein present in milk with a molecular mass of 78 kDa. LPO is included in many biological processes and is well-known to have biocidal actions, acting as an active antibiotic and antiviral agent. The wide spectrum biocidal activity of LPO is mediated via a definite inhibitory system named lactoperoxidase system which plays a potent role in the innate immune response. With the current advancement in nanotechnology, nanoformulations can be developed for stabilizing and potentiating the activity of LPO for several applications. In the research described in this Research Communication, fresh LPO purified from bovine mammary gland secretions was used for nanoparticle synthesis using a simple thermal process at different pH and temperatures. The round-shaped nanoparticles (average size 229 nm) were successfully synthesized at pH 7.0 and a temperature of 75°C. These nanoparticles were tested against four different bacterial species namely S. flexineri, P. aeruginosa, S. aureus, and E. coli. The prepared nanoparticles exhibited strong inhibition of the growth against all four bacterial species as stated by their MIC and ZOI values. These results may help in increasing the efficiency of lactoperoxidase system and will assist in identifying novel avenues to enhance the stability and antimicrobial function of LPO in drug discovery and industrial processes.