INCLUSION OF A SPECIES-SPECIFIC PROBIOTIC OR CALCIUM DIFORMATE IN YOUNG CALVES DIETS: EFFECTS ON GUT MICROBIAL BALANCE, HEALTH STATUS AND GROWTH PERFORMANCE

ABSTRACT Calf morbidity and mortality represent major financial losses for veal and dairy producers, especially intestinal bacterial infections are the primary causes of calf mortality. Thus the preventive intervention is a recommended strategy and as a prior measures probiotics and acidifiers is an alternative to the use of antibiotics. Probiotics play a critical role in the completeness and immune response of the intestinal mucosa while the main mode of action of organic acids is through their bacteriostatic and bactericidal action. The purpose of the present trials was the evaluation of the effects both a species-specific probiotic administration in veal (1st trial) and dairy calves (2nd trial) than an acidifier supplementation in veal calves (3rd trial) on health status, gut microbial balance and growth performance. On veal calves the effects of probiotic and acidifier on slaughter performance and histological survey were further evaluated. The aim of first the study was to evaluate the effects of the administration of a species-specific probiotic supplement to veal calves on performance and microbial parameters in standard rearing conditions. Ninety six male Friesian veal calves (49.31 ± 1.38kg of body weight and 20±5 days of life) were divided at random in two homogeneous groups of 48 animals each from the arrival in the farm and fed either a basal diet (C) or a basal diet plus 1.8x109 CFU/head/day of a probiotic supplement containing Bacillus coagulans, Lactobacillus animalis and Lactobacillus paracasei spp. paracasei in a 35:30:35 ratio (T) for a total of 180 days. Starting from 10th day from arrival and monthly until the end of the trial, individual body weight (BW) was recorded and average daily gain (ADG) was computed. At the same time, on the half of animals per group, faecal samples were collected for faecal score evaluation (FS), Lactobacilli count, Escherichia coli count and Lactobacilli/E.coli ratio, while blood samples were collected for haematological, haematochemical and immunological parameters evaluation. During the whole experimental period daily health status and therapeutic treatments were recorded for General Health Score (GHS) determination. At slaughter 10 animals per group were analysed for macroscopically injuries detection, and gut samples were collected for histological analyses. On each subject carcass weight, dressing percentage, fattening condition, carcass grade (SEUROP classification) and meat pH were recorded. During the trial were considered six times (corresponding to the each month of fattening): from time 0, referred to the 10th day from arrival, to the last considered month (time 6). BW resulted significantly higher (P≤0.05) in C calves than T at the 5th (C=226.76kg vs T=223.27kg) and 6th (C=267.14kg vs T=263.56kg) considered times while ADG was not different between groups. Faecal consistency was, significantly higher in T group at the 4th sampling (P≤0.01) as faecal Lactobacilli content (C=8.37 Log10 vs T=8.49 Log10; P≤0.05). This trend could explain the better GHS observed in supplemented calves during the trial. At different times, some haematological parameters such as basophils, aspartate amino transferase (AST), bilirubin, glucose, urea, bactericidal and complement were higher (P<0.05) in C group, while non-esterificated fatty acids (NEFA) and lactate dehydrogenase (LDH) showed lower values than T (P<0.05). At slaughter no differences were observed for carcass weight, dressing percentage, carcass grade and meat pH. The cecum histological examination revealed a less evident de-epithelialisation and a greater integrity of the epithelium surface in T subjects than C, while in both the ileum than in cecum a numerical increase of the intestinal crypts’ depth was detected. The administration of species-specific probiotic was able to improve the gut microbiological balance and health status with not no evident effects on growth performance. The second study evaluated the effects of the administration of a species-specific multistrain probiotic (Lactobacillus animalis-Lactobacillus paracasei subsp. paracasei-Bacillus coagulans; 30:35:35) on health and performance parameters of newborn Friesian female calves during the first month of life. Twenty-two calves were divided in two groups: control (C) fed with milk replacer and concentrate as a basal diet, and treatment (T), fed C diet plus 1g/head/d of probiotic from the second day after birth to the end of the first month of life. Faecal samples were collected weekly for the count of Lactobacilli and Escherichia coli; blood samples were collected and analyzed weekly. Individual FS was recorded daily, GHS was calculated at the end of the trial, and cell mediate immune response was evaluated by skin test at 7 and 28 days of life. Body weight, biometrical parameters and ADG were recorded weekly while Feed Intake (FI) and Feed Conversion Rate (FCR) were recorded for the overall period. Higher faecal Lactobacilli/E. coli ratio on day 28 of life (3.73 Log UFC/g vs. 2.02 Log UFC/g; P<0.05) and improved faecal consistency at 6, 25 and 27d of life were found in T group. Final body weight (48.92 Kg vs 46.92 Kg; P<0.05) and hearth girth (81.16 cm vs 78.49 cm; P<0.05) were significantly higher in T group. Concentrate FI (%DM) was significantly higher in T group overall the trial period, while ADG, FCR and milk replacer FI (%DM) were not influenced by probiotic administration. GHS showed increased mean value, but non significantly, in T group (T=20.45 vs C=18.91). Increased haemoglobin (10.04g/dl vs 8.60g/dl) and hematocrit (26.68% vs 22.17%) plasma content at 8d in T group with lower eosinophils percentage (0.05% vs 0.22%) were found at 8d in T group, while basophiles content was increased at 28d in species-specific probiotc-fed animals than C (0.21% vs 0.16%, respectively; P<0.05). The administration of a species-specific probiotic compound during the first month of life of newborn calves improved gut microflora, increased performance and some biometric parameters. The last trial involving 36 male Friesian calves was conducted to determine the effect of calcium diformate administration and its inclusion level on growth performance, intestinal balance, metabolism, health status, and calcium and formate organ content. The calves (44.55kg±5.51kg BW, 25±10 days old), were divided in three homogeneous groups of twelve subjects each: C fed with a basal diet, T1 fed with a basal diet plus 1%/milk replacer of calcium diformate, and T3 fed a basal diet plus 3%/milk replacer of calcium diformate for a total of 84 days of supplementation. During the experimental period daily FI, AD and FCR were determined. At 0, 42 and 84 days of trial individual blood samples were collected in order to detect haematological and biochemical parameters, Ca and FO content. At the same time faecal samples were collected for Lactobacilli, total coliforms and E.coli count along with faecal pH. Weekly BW and FS were recorded, while the health status was constantly monitored and the cause of deaths analyzed by necropsy. At the 91st day of trial the calves were slaughtered: any presence of ulcers were detected, individual carcass weight and dressing percentage were determined, and kidney and liver weight recorded. On seven animals per group liver, kidney, muscle and fat samples were collected and total Ca and formate were evaluated. The administration of calcium diformate decreased dry matter intake from either milk powder, concentrate and both milk powder and concentrate together with increasing level of calcium diformate in the diet (C=1419,05g/h/d; T1=1217,78g/h/d; T3=1190,73g/h/d; P≤0.01). As a result, final body weight in treated animals was lower than control calves (C=121,00kg; T1=112,17kg; T3=108,74kg; P≤0.01). Mean ADG in 3%/milk replacer CaFO calves was decreased if compared to C subjects (C=0.87kg/h/d vs T3=0.73kg/h/d; P<0.01), while a similar feed conversion rate was detected between the experimental groups. Faecal score was decreased in T3 animals (P≤0.05) than control while no differences were found for faecal pH, E.coli or Lactobacilli. Higher total coliforms count was evidenced in T1 animals than C (7.45 Log10 vs T1=2.75Log10 respectively; P≤0.05). At slaughter no difference was observed for dressing percentage, while carcass weight resulted higher in C compared to T3 (C=67.49kg vs T3=58.87; P≤0.05). Organ weight and organ formate and calcium content were similar between the experimental groups. Red blood cells count (RBC), haemoglobin (HB) and hematocrit (HT) resulted higher (P≤0.05) in C and T3 than T1, while control calves showed higher platelets (P≤0.05) value than T1, and greater (P≤0.05) triglycerides than T3. Mean cell haemoglobin (MCH) value resulted higher (P≤0.05) in T3 subjects than the other groups. Increased formate blood content in 3% of calcium difromate/milk replacer animals did not show any differences for organ formate content thus giving no accumulation. Immune response was not affected by the treatment and necropsies on dead animals suggest no interaction among treatment and death itself. The histological examination revealed altered structural aspects of the intestinal mucosa in all the experimental corresponding with no detectable differences between control T1 and T3 groups. Epithelial detachment in the intestinal villi associated with mucous secretion excess and some diffuse mixed cellular infiltration sub mucosa and lamina propria were highlighted. The sub mucosal Brunner glands were in normal physiological conditions in C, but showed empty and enlarged lumen in treated animals. While not showing toxic effects in veal calves, the administration of calcium diformate is not useful in promoting the growth performance and gut balance. In conclusion, the species-specific probiotic administration in veal and dairy calves can improve the gut microbial balance and consequently the general health status, although no effect on growth performance, while the calcium diformate supplementation, even if not showing toxic effects in veal calves, isn’t helpful to increase the intestinal microbial balance and the growth performance.