Genetic parameters of differential somatic cell count, milk composition, and cheese-making traits measured and predicted using spectral data in Holstein cows.

Mastitis is one of the most prevalent diseases in dairy cattle and is the cause of considerable economic losses. Alongside somatic cell count (SCC), differential somatic cell count (DSCC) has been recently introduced as a new indicator of intramammary infection. The DSCC is expressed as a count or a proportion (%) of polymorphonuclear neutrophils plus lymphocytes (PMN-LYM) in milk somatic cells. These numbers are complemented to total somatic cell count or to 100 by macrophages (MAC). The aim of this study was to investigate the genetic variation and heritability of DSCC, and its correlation with milk composition, udder health indicators, milk composition, and technological traits in Holstein cattle. Data used in the analysis consisted in single test-day records from 2,488 Holstein cows reared in 36 herds located in northern Italy. Fourier-transform infrared (FTIR) spectroscopy was used to predict missing information for some milk coagulation and cheese-making traits, to increase sample size and improve estimation of the genetic parameters. Bayesian animal models were implemented via Gibbs sampling. Marginal posterior means of the heritability estimates were 0.13 for somatic cell score (SCS); 0.11 for DSCC, MAC proportion, and MAC count; and 0.10 for PMN-LYM count. Posterior means of additive genetic correlations between SCS and milk composition and udder health were low to moderate and unfavorable. All the relevant genetic correlations between the SCC traits considered and the milk traits (composition, coagulation, cheese yield and nutrients recovery) were unfavorable. The SCS showed genetic correlations of -0.30 with the milk protein proportion, -0.56 with the lactose proportion and -0.52 with the casein index. In the case of milk technological traits, SCS showed genetic correlations of 0.38 with curd firming rate (k20), 0.45 with rennet coagulation time estimated using the curd firming over time equation (RCTeq), -0.39 with asymptotic potential curd firmness, -0.26 with maximum curd firmness (CFmax), and of -0.31 with protein recovery in the curd. Differential somatic cell count expressed as proportion was correlated with SCS (0.60) but had only 2 moderate genetic correlations with milk traits: with lactose (-0.32) and CFmax (-0.33). The SCS was highly correlated with the log PMN-LYM count (0.79) and with the log MAC count (0.69). The 2 latter traits were correlated with several milk traits: fat (-0.38 and -0.43 with PMN-LYM and MAC counts, respectively), lactose percentage (-0.40 and -0.46), RCTeq (0.53 and 0.41), tmax (0.38 and 0.48). Log MAC count was correlated with k20 (+0.34), and log PMN-LYM count was correlated with CFmax (-0.26) and weight of water curd as percentage of weight of milk processed (-0.26). The results obtained offer new insights into the relationships between the indicators of udder health and the milk technological traits in Holstein cows.

Could predicting fatty acid profile by mid-infrared reflectance spectroscopy be used as a method to increase the value added by milk production chains?

The aims of this work were (1) to develop prediction equations from mid-infrared spectroscopy (MIRS) to establish a detailed fatty acid (FA) composition of milk; (2) to propose a milk FA index, utilizing MIRS-developed equations, in which the precision of the FA-prediction equations is taken into account to increase the value of milk; and (3) to show application examples. A total of 651 bulk cow milk samples were collected from 245 commercial farms in northwest Italy. The results of the 651 gas chromatography analyses were used to establish (421 samples) and to validate (230 samples) the outcomes of the FA composition prediction that had been obtained by MIRS. A class-based approach, in which the obtained MIRS equations were used, was proposed to define a milk classification. The method provides a numerical index [milk FA index (MFAI)] that allows a premium price to be quantified to increase the value of a favorable FA profile of milk. Ten FA were selected to calculate MFAI, according to their relevance for human health and potential cheese sensory properties, and animal welfare and environmental sustainability were also considered. These factors were selected as dimensions of MFAI. A statistical analysis and expert judgment aggregation were performed on the selected FA by weighting the FA and normalizing the dimensions to reduce redundancy. A class approach was applied, using the precision of the MIRS equations to establish the classes. The median FA concentration of the data set was set as a reference value of class 0. The width, number, and limits of classes above and below the median were calculated using the 95% confidence level of the standard error of prediction, corrected with the bias of each FA. A progressive number and a positive or negative sign were assigned to each FA class above or below the median according to their role in the above mentioned dimensions. The sum of the numbers of each class, associated with its sign for each FA, was used to generate MFAI. The MFAI was applied to dairy farms characterized by different feeding strategies, all of which deliver milk to a commercial dairy plant. The MFAI values ranged from 0.7 to 4.2, and large variations, which depended on the cows' diet and forage quality, were observed for each feeding system. The proposed method has been found to be flexible and adaptable to several contexts on both intensive and extensive dairy farms.

N-linked glycan truncation causes enhanced clearance of plasma-derived von Willebrand factor.

Essentials von Willebrands factor (VWF) glycosylation plays a key role in modulating in vivo clearance. VWF glycoforms were used to examine the role of specific glycan moieties in regulating clearance. Reduction in sialylation resulted in enhanced VWF clearance through asialoglycoprotein receptor. Progressive VWF N-linked glycan trimming resulted in increased macrophage-mediated clearance. Click to hear Dr Denis discuss clearance of von Willebrand factor in a free presentation from the ISTH Academy SUMMARY: Background Enhanced von Willebrand factor (VWF) clearance is important in the etiology of both type 1 and type 2 von Willebrand disease (VWD). In addition, previous studies have demonstrated that VWF glycans play a key role in regulating in vivo clearance. However, the molecular mechanisms underlying VWF clearance remain poorly understood. Objective To define the molecular mechanisms through which VWF N-linked glycan structures influence in vivo clearance. Methods By use of a series of exoglycosidases, different plasma-derived VWF (pd-VWF) glycoforms were generated. In vivo clearance of these glycoforms was then assessed in VWF-/- mice in the presence or absence of inhibitors of asialoglycoprotein receptor (ASGPR), or following clodronate-induced macrophage depletion. Results Reduced amounts of N-linked and O-linked sialylation resulted in enhanced pd-VWF clearance modulated via ASGPR. In addition to this role of terminal sialylation, we further observed that progressive N-linked glycan trimming also resulted in markedly enhanced VWF clearance. Furthermore, these additional N-linked glycan effects on clearance were ASGPR-independent, and instead involved enhanced macrophage clearance that was mediated, at least in part, through LDL receptor-related protein 1. Conclusion The carbohydrate determinants expressed on VWF regulate susceptibility to proteolysis by ADAMTS-13. In addition, our findings now further demonstrate that non-sialic acid carbohydrate determinants expressed on VWF also play an unexpectedly important role in modulating in vivo clearance through both hepatic ASGPR-dependent and macrophage-dependent pathways. In addition, these data further support the hypothesis that variation in VWF glycosylation may be important in the pathophysiology underlying type 1C VWD.

von Willebrand factor arginine 1205 substitution results in accelerated macrophage-dependent clearance in vivo.

BACKGROUND: Enhanced von Willebrand factor (VWF) clearance is important in the etiology of type 1 and type 2 von Willebrand disease (VWD). More than 20 different VWF point mutations have already been reported in patients with enhanced clearance. These include the VWD-Vicenza variant, which is characterized by an Arg1205His substitution in the VWF D3 domain. Critically, however, the molecular mechanisms through which single amino acid substitutions in VWF result in enhanced clearance of this complex multimeric glycoprotein have not been defined. OBJECTIVES: In this study, we have investigated the biological basis underlying the enhanced clearance of the VWF-R1205H variant. METHODS: Using VWF(-/-) mice, in vivo clearance rates were determined for a series of full-length and truncated recombinant VWF variants. In addition, the role of macrophages in modulating enhanced VWD-Vicenza clearance was investigated using clodronate liposome administration. RESULTS: Our findings demonstrate that substitutions of R1205 with histidine, cysteine or serine all result in markedly reduced survival of full-length recombinant VWF. Importantly, D'A3 fragments containing these same R1205 substitutions also demonstrated significantly enhanced clearance. In contrast to the reduced in vivo survival observed with R1205H, clearance of R1204H was not enhanced. Recent studies have demonstrated that hepatic and splenic macrophages play key roles in regulating VWF clearance. Importantly, macrophage-depletion also served to markedly attenuate the enhanced clearance phenotypes associated with VWF-R1205H, VWF-R1205S and VWF-R1205C. CONCLUSIONS: Collectively, these novel findings demonstrate a specific and critical role for the R1205 residue in modulating macrophage-mediated clearance of VWF in vivo.

The O-linked glycans of human von Willebrand factor modulate its interaction with ADAMTS-13.

BACKGROUND: O-linked glycans (OLGs) are clustered on either side of the von Willebrand factor (VWF) A1 domain and modulate its interaction with platelets; however, their influence on the VWF interaction with ADAMTS-13 is unknown. OBJECTIVES: To assess the role of the OLGs in VWF susceptibility to ADAMTS-13 proteolysis, which would help to explain their specific distribution. METHODS: OLG sites were mutated individually and as clusters on either and both sides of the A1 domain, and expressed in HEK293T cells. First, their proteolysis by ADAMTS-13 was assayed in the presence of urea. Next, a parallel-flow chamber was used to analyze VWF-mediated platelet capture on collagen in the presence and absence of ADAMTS-13 under a shear stress of 1500 s(-1) . The decrease in platelet capture in the presence ADAMTS-13 was used as a measure of VWF proteolysis. RESULTS: Initially, we found that, under denaturing conditions, the C-terminal S1486A and Cluster 2 and double cluster (DC) variants were less susceptible to ADAMTS-13 proteolysis than wild-type VWF. Next, we showed that addition of ADAMTS-13 diminished VWF-mediated platelet capture on collagen under flow; surprisingly, this was more pronounced with the S1486A, Cluster 2 and DC variants than with wild-type VWF, indicating that these are proteolyzed more rapidly under shear flow. CONCLUSIONS: OLGs provide rigidity to peptide backbones, and our findings suggest that OLG in the A1-A2 linker region regulates VWF conformational changes under shear. Importantly, the impact of OLGs on ADAMTS-13 cleavage under shear stress is the opposite of that under denaturing conditions, highlighting the non-physiologic nature of in vitro cleavage assays.

Effect of different feeding strategies in intensive dairy farming systems on milk fatty acid profiles, and implications on feeding costs in Italy.

The aim of this work was to characterize the fatty acid (FA) profile of milk from intensive dairy farming systems in the Po Plain (Italy) to estimate the costs of the adopted feeding strategies and to simulate the effect of supplementary premiums on the basis of milk FA composition on milk income. Twenty dairy farms with 5 different feeding strategies were studied: 3 corn silage-based systems in which cows were supplemented with a great proportion (CCH), a medium proportion (CCM), or without commercial concentrate mix (CC0), and 2 systems in which part of corn silage was replaced with grass or legume silage (HF) or with fresh herbage (G), cut and fed indoors. Bulk milk was sampled and lactating cow performance, feeding strategies and forage characteristics were recorded through a survey, 3 times during a year. The milk FA supplementary premium was calculated considering C18:3n-3 and saturated FA (SFA) concentrations, and ratio of total cis C18:1 isomers to C16:0. The CCH, CCM, and CC0 systems bought most of their dairy cow feeds off farm, which allowed them to increase milk production to 35,000 L/yr per hectare. Their low dry matter and crude protein self-sufficiency led to higher feeding costs per liter of milk (from €0.158 to €0.184), and highest income over feed cost was achieved only for milk yield performance greater than 10,000 kg/cow per year. The use of homegrown forages in HF and G increased dry matter and crude protein self-sufficiency and reduced the feeding costs per liter of milk from 9 to 22%, compared with the other studied systems, making HF and G feeding economically competitive, even for a lower milk yield per cow. The studied systems highlighted a remarkable variation in FA profiles. The concentrations of C16:0 and SFA were the highest in CCH (31.53 and 67.84 g/100g of FA) and G (31.23 and 68.45 g/100g of FA), because of the larger proportion of commercial concentrate mix in the cow diet. The concentrations of C16:0 and SFA were the lowest in CCM (27.86 and 63.10 g/100g of FA), because of low roughage-to-concentrate ratio in the cow diet, which is known to favor milk fat depression, affecting particularly these FA. The calculated supplementary premium was the highest in the CCM system, based on milk FA profiles from those herds. The HF diet was rich in forages and resulted in greater concentration of C18:3n-3 in milk (0.57 g/100g of FA) than the other systems and thus led to an increase in milk FA supplementary premium. Milk from G and HF milk had the lowest ratio of Σn-6:Σn-3 FA compared with milk from the systems based on higher corn silage proportion in the cow diet (3.71, and 3.25, respectively, vs. 4.58 to 4.78), with the lower ratios being closer to recommendation for human nutrition.

Effect of Lactobacillus buchneri LN4637 and Lactobacillus buchneri LN40177 on the aerobic stability, fermentation products, and microbial populations of corn silage under farm conditions.

This study determined the efficacy of the use of 2 commercial inoculants containing Lactobacillus buchneri alone or in combination with homofermentative lactic acid bacteria in improving aerobic stability of corn silage stored in commercial farm silos in northern Italy. In the first survey, samples were collected from 10 farms that did not inoculate their silages and from 10 farms that applied a Pioneer 11A44 inoculant (L. buchneri strain LN4637; Pioneer Hi-Bred International, Des Moines, IA). In the second survey, corn silage samples were collected from 11 farms that did not inoculate their silages and from 11 farms that applied a Pioneer 11CFT inoculant (L. buchneri strain LN40177; Pioneer Hi-Bred International). Inoculants were applied directly through self-propelled forage harvesters, at the recommended rate of 1 g/t of fresh forage, to achieve a final application rate of 1.0 × 10(5) cfu/g of L. buchneri. One corn bunker silo, which had been open for at least 10 d, was examined in detail on each farm. The silages inoculated with L. buchneri had lower concentrations of lactic acid, a lower lactic-to-acetic acid ratio, a lower yeast count, and higher aerobic stability compared with the untreated silages. Unexpectedly, concentrations of acetic acid and 1,2-propanediol, 2 hallmarks of L. buchneri activity, did not differ between treatments and were only numerically higher in the inoculated silages compared with untreated ones, in both surveys. Aerobic stability, on average, was 107 and 121 h in the inoculated silages and 64 and 74 h in the untreated silages, for surveys 1 and 2, respectively, and decreased exponentially as the yeast count in the silage at the time of sampling increased, regardless of treatment. Inoculation with L. buchneri proved to be effective in reducing the yeast count to <2 log cfu/g of silage in 16 of 21 of the studied farm silages, confirming the ability of this inoculum to enhance the aerobic stability of corn silages in farm bunker silos.