Livestock systems aim to improve animal production and welfare. Nowadays, zootechnical systems aim to enhance animal growth and productivity to maximise profit in the shortest time practical. In such systems, beef steers are fed a minimum of roughage and a high amount of concentrate. A high amount of highly fermentable substrates in the diet can lead to imminent rumen dysfunction, induced by an alteration of the microbial population with ruminal inflammation and metabolism disorders (22). Recent studies have demonstrated the direct and indirect positive effects on performance and overall benefits to animal health of the yeast
Ruminants obtain energy from plant compounds through microbial fermentation in the rumen. This process can be potentiated by probiotics. Among probiotics, live yeast cultures have received particular attention from the research community. Yeasts are an important source of products with probiotic activity in livestock systems and provide livestock producers with a replacement for sub-therapeutic antibiotic supplementation, thereby mitigating the potential negative effects of morbidity in order to guarantee the health status of animals as well as maximise their profitability.
The ruminal ecosystem benefits from
In the digestive system of animals which receive yeast supplementation, the addition increases microbial activity by means of the action of its vitamin and enzyme constituents. In particular, several effects induced by yeast supplementation in steers were shown through the evaluation of rumen function, hepatic markers, growth performance, feed intake and digestibility, immunity stimulation and reproductive performance (3, 7). It has been reported that
Physiological or pathological conditions in cattle are shown by different serum protein concentrations. Their evaluation by electrophoresis is a valuable and commonly used laboratory diagnostic tool in veterinary medicine (4). Albumin is the most osmotically active serum protein and is a carrier of many substances. Globulins are a heterogeneous group of serum proteins that includes antibodies and other inflammatory molecules, haemostatic and fibrinolytic proteins, and carriers of lipids, vitamins and hormones. These examples show that serum proteins have multiple functions. The change between physiological and pathological conditions can cause shifts in albumin and globulin concentrations, and their monitoring is an indispensable husbandry tool.
Considering that the acute-phase response is an expression of bulls’ wellness and reflects their inflammation status, and that minimising provocation of the response is crucial for their performance, the aim of this study was to evaluate the effect of diet supplementation with
Sixty clinically healthy, 10-month-old Charolaise bulls with a body weight between 501 and 535 kg were enrolled in this study. The enrolled animals were randomly selected from a farm located in the north-east of Italy. All animals were subjected to a clinical examination before the start of the study and at every time point throughout all experimental periods. The animals were divided into two equal groups (n = 30) and housed in different pens. Thirty bulls were the diet supplementation group (YG), which received daily
The feed ingredients and analytical composition of the diet used are shown in Table 1. The total mixed rations were analysed using near-infrared reflectance spectroscopy (NIRSystem 5000; FOSS Italia, Padova, Italy). A 5g mass corresponding to 5 dL of liquid commercial
Feed ingredients and chemical analysis of total mixed rations provided to the experimental animals
Feed ingredients (kg per day per head) | |
---|---|
BULL 100 11.11* | 0.50 |
Corn gluten feed | 0.60 |
Alfalfa hay | 1.00 |
Corn | 2.00 |
Dry pulp | 1.00 |
Straw | 0.80 |
Corn silage | 5.65 |
Soybean meal | 0.30 |
Total | 11.85 |
Dry matter (%) | 57.17 |
Chemical composition | |
Crude protein (%) | 13.16 |
Ethereal extract (%) | 3.11 |
Fibre (%) | 14.98 |
Ash (%) | 6.07 |
Neutral detergent fibre (%) | 38.93 |
Starch (%) | 32.52 |
Ca (g) | 69.47 |
P (g) | 24.94 |
* – Bull 100 11.11 protein, vitamins and minerals premix: vitamin A (169,000 UI/kg), vitamin D3 (16,900 UI/kg), vitamin E (416 mg/kg), vitamin B1 (42 mg/kg), vitamin B12 (0.22 mg/kg), choline (845 mg/kg), niacinamide (1,793 mg/kg), manganous sulphate (191 mg/kg), manganous oxide (381 mg/kg), zinc chelate of amino acids (5,954 mg/kg), zinc oxide (742 mg/kg), copper sulphate pentahydrate (216 mg/kg), cobalt carbonate (2.2 mg/kg), potassium iodide (14.7 mg/kg), urea (49,500 mg/kg)
All treatments were carried out and housing and animal care conditions provided in accordance with the standards recommended by Directive 2010/63/EU for animal experiments (14).
Blood samples were collected by tail venepuncture using 22 G × 25 mm needles into 10 mL serum vacuum tubes with clot activator (Vacutainer; BD Diagnostics - Preanalytical Solutions, Plymouth, UK). Sampling was carried out in both groups on day 0 (t0) immediately before the supplementation of the experimental diet, and 20 (t1) and 40 (t2) days after its start. The samples were allowed to clot for 2 h at 4°C before centrifugation at 1,350 ×
The concentration of total proteins, albumin, globulin fractions and haptoglobin was assessed in the obtained serum samples. Serum total protein concentration was assessed with a commercial kit by means of an automated ultraviolet spectrophotometer (Slim; SEAC, Florence, Italy) using the biuret method and bovine serum albumin at a concentration of 6.02 g/dL as the standard (Biosystems, Barcelona, Spain).
The protein fraction was assessed by an automated system (Selvet24; Seleo Engineering, Naples, Italy) according to the procedures described by the manufacturer. A total of 25 μL of each serum sample was applied to numbered sample wells in cellulose acetate films. Each holder accommodated up to 24 samples. The films were electrophoresed for 28 min at 180 V. After electrophoresis, the films were immediately fixed using an automated system, stained in red stain acid solution for 10 min, and then dried at 37°C. After being destained in acetic acid and dried completely for 15 min, the films were scanned on a densitometer, electrophoretic curves were plotted and the related quantitative specific protein concentrations calculated using software (SelVet 24; Seleo Engineering). All samples were analysed by the same operator, who determined the lines separating fractions in the densimeter tracing. The major protein fractions were divided into albumin, α1-, α2-, β1-, β2- and γ-globulins from the cathode to the anode, according to the recommendation by the manufacturer (5).
Relative protein concentrations within each fraction were determined as the optical absorbance percentage, then the absolute concentration (g/L) and albumin : globulin ratio (A : G) were calculated using the total protein concentration.
Haptoglobin was assessed with an ELISA kit specific for bovine species (PHASE Haptoglobin assay, Tridelta Development, Maynooth, Republic of Ireland), which had sensitivity of 0.005 mg/mL and intra- and inter-assay coefficients of variation of <7% and <6%, respectively. A microtitre plate reader (EZ Read 400 ELISA; Biochrom, Cambridge, United Kingdom) was used to determine haptoglobin levels. All calibrators and samples were run in duplicate, and samples exhibited parallel displacement to the standard curve for ELISA analysis.
The obtained data were expressed as mean ± standard error of the mean. Data were normally distributed as determined by the Kolmogorov–Smirnov test (P-value > 0.05). Two-way analysis of variance (ANOVA) for repeated measures was applied to determine the influence of feed supplementation and of time (t0, t1 or t2) on the investigated parameters, followed by Bonferroni’s
All investigated parameters were within the physiological range for the species (17). Two-way ANOVA showed a statistically significant effect of yeast supplementation time on serum total protein, albumin, β1-globulins, β2-globulins, γ-globulins and haptoglobin (P-value < 0.001 in all cases). A significant effect of the group was observed on total protein, albumin and γ-globulins (P-value < 0.01 in the three cases) and haptoglobin (P-value < 0.001). In particular, Bonferroni’s
Haptoglobin decreased in both groups at t1 and t2
Mean values ± standard error of the mean of studied parameters with statistical differences related to group, measured in the serum of bulls of the control group (CG) without yeast supplementation and the serum of bulls of the group given 5 g of
Serum parameters | Group | t0 | t1 | t2 |
---|---|---|---|---|
Total protein (g/L) | CG | 73.07 ± 0.45 | 72.40 ± 0.63 | 74.08 ± 0.40*** |
YG | 71.47± 0.53 | 71.26 ± 0.51 | 76.91 ± 0.47*** | |
Albumin (g/L) | CG | 34.46 ± 0.42 | 31.75 ± 0.34** | 33.41 ± 0.41** |
YG | 33.84 ± 0.44 | 33.49 ± 0.34** | 35.20 ± 0.40** | |
α1-globulins (g/L) | CG | 3.68 ± 0.12 | 3.35 ± 0.09 | 3.65 ± 0.25 |
YG | 3.48 ± 0.08 | 3.19 ± 0.11 | 3.48 ± 0.08 | |
α2-globulins (g/L) | CG | 8.52 ± 0.16 | 8.77 ± 0.12 | 8.47 ± 0.28 |
YG | 8.66 ± 0.16 | 8.20 ± 0.18 | 9.09 ± 0.16 | |
β1-globulins (g/L) | CG | 9.03 ± 0.38 | 8.14 ± 0.11 | 9.16 ± 0.21 |
YG | 8.84 ± 0.25 | 8.54 ± 0.23 | 9.82 ± 0.19 | |
β2-globulins (g/L) | CG | 9.78 ± 0.29 | 9.51 ± 0.28 | 9.77 ± 0.24 |
YG | 9.21 ± 0.24 | 8.79 ± 0.21 | 10.15 ± 0.22 | |
γ-globulins (g/L) | CG | 8.95 ± 0.51 | 10.89 ± 0.47** | 10.95 ± 0.37 |
YG | 8.54 ± 0.34 | 9.05 ± 0.32** | 10.17 ± 0.36 | |
A: G ratio | CG | 0.87 ± 0.02 | 0.79 ± 0.02*** | 0.80 ± 0.01 |
YG | 0.88 ± 0.01 | 0.89 ± 0.01*** | 0.83 ± 0.01 | |
Haptoglobin (mg/L) | CG | 1.28 ± 0.07 | 0.75 ± 0.08 | 0.74 ± 0.06 |
YG | 1.28 ± 0.09 | 0.49 ± 0.05* | 0.47 ± 0.04* |
* – P < 0.05;
** – P < 0.01;
*** – P < 0.001 (in Bonferroni’s
One of the principal parameters used for assessing animal nutritional status is total protein. In our study, higher serum total protein values were found in the YG compared to the CG after 40 days of yeast supplementation. The proteins comprising the total are sources of amino acids for muscle protein synthesis and they can be related to muscle mass increase (3). A higher serum concentration of total protein was found in the CG at t2 relative to t1, in the YG at t2 compared to t1 and in the YG at t2 than at t0, and this related to the well-known lower serum total protein concentration in young animals than in adults due to muscle mass increase with age (10). The increased concentration of serum total protein in the YG at the end of the experimental period may indicate the improved nutrient status of the bulls and may have been induced by yeast-supplemented feed intake. In agreement with our results, Adeyemi
An increase of α2- and β2-globulins was found in the YG, while β1- and γ-globulins increased in both the YG and CG at the end of the experimental period. These findings suggest an activation of the acute-phase response in both groups (1). Burdick Sanchez
It is well documented that some changes in diet could result in rumen pH changes, alterations to the microbial ecosystem and the accumulation of large amounts of endotoxin in the organism, and that it is linked to increased peak concentrations of acute-phase proteins, such as haptoglobin. Haptoglobin is one of the most important α2-globulins. It is synthesised by hepatocytes in negligible concentrations in healthy animals and in high amounts in unhealthy ones, and may increase more than 100-fold during the inflammatory and acute-phase response in cattle (28). For this reason, the serum haptoglobin value has been exploited as a useful marker for disease conditions, infections, inflammations, and stressor effects (9). Serum haptoglobin concentration was significantly affected by time of sampling in steers studied by Shen
It is well established that the use of yeast supplementation in feed has positive effects on the physiology of bulls. Yeast supplementation induced an increase in the serum levels of total protein and albumin as indices of higher body mass gain and a reduced acute-phase response, in association with an increase of the a2-globulins after 40 days of yeast supplementation. This aspect together with a delayed increase of γ-globulins with respect to the control group and a decrease in serum haptoglobin concentration starting from 20 days of yeast supplementation indicates the reductive effect of