Ingested versus inhaled limonene in sheep: A pilot study to explore potential different transfer to the mammary gland and effects on milk and Caciotta cheese aroma.

Concentration is a key determinant in the overall positive impact of terpenes on milk and cheese aroma; additionally, route of intake may affect the achievable concentrations of dietary terpenes in milk and cheese. In this study, we explored the possibility that the amount of the monoterpene limonene transferred to sheep milk and its corresponding cheese could differ depending on the route of intake and that the aroma profile of these products could also differ. To this aim, 12 lactating dairy ewes were repeatedly exposed to limonene by the oral or respiratory route during a 48-h test period, according to a 3 × 3 Latin square experimental design. Limonene content was measured in individual and bulk milk samples, in 1-d-old and 15-d-old Caciotta cheese obtained from that milk, in the related whey and curd, and in the air inhaled by the ewes in the respiratory treatment group (to obtain an estimate of the dose actually supplied by this route). Bulk milk and fresh (1-d-old) cheese underwent sensory analysis by ortho-olfactory evaluation. Both intake routes demonstrated transfer of limonene to milk, but the respiratory route transferred limonene with greater efficiency than the oral route. Moreover, according to the protocol used in this study, a short period of respiratory exposure induced a slightly higher limonene content in milk compared with oral exposure. As to the fate of limonene during cheesemaking, an important part of it was lost into the whey, perhaps through volatilization. The differences between milk and cheese tended to dissipate in curd and fresh cheese and disappeared completely after 15 d of ripening. Finally, it was possible to distinguish between the 2 routes of limonene intake using sensory analysis, even though no direct relationship was identified between the different aroma profiles of milks and cheeses from the oral and respiratory groups and their respective limonene contents. Overall, our results expand current knowledge on the biological pathways of terpene transfer from feed to sheep milk and cheese, as well as on the role played by terpenes in the formation of aroma in these products. Our observations may contribute to future development of strategies for external control and better standardization of the presence of odor compounds in milk and cheese from dairy ruminants.

Oral administration of nucleotides in calves: Effects on oxidative status, immune response, and intestinal mucosa development.

The present work aimed to investigate the effects of nucleotide oral administration on oxidative stress biomarkers, immune responses, gut morphology, serum biochemical parameters, and growth performance in calves from birth to 25 d of life. A total of 40 male Holstein Friesian calves were randomly divided in 2 groups. All the calves were born and reared on the same commercial dairy farm. They were fed the same colostrum, milk replacer, and calf starter. Five grams/head of an additive were orally administered with a syringe directly in the mouth to calves of the nucleotide group (NG). The additive contained 74.12 g/100 g of nucleic acids from hydrolyzed yeast, and 75.38% was free nucleotide sodium salt. The other group represented the negative control (CG). At 25 d of life all of the calves were slaughtered. Calves supplemented with nucleotides had a higher final live weight and improved average daily gain, which was associated with better efficiency of nutrient use. Oral nucleotide administration did not affect IgG absorption efficiency; however, NG calves showed greater duodenum villi length and higher crypt depth compared with CG. Oral nucleotide administration increased the activity of antioxidant enzymes (superoxide dismutase, catalase and glutathione peroxidase) and the antioxidant capacity [ferric reducing antioxidant power and 2,2'-azino-bis (3-ethylbenzothiazoline-6-sulphonic acid) scavenging activity] both in plasma and in liver. An enhanced ability of cells to counter reactive oxygen species- and reactive nitrogen species-mediated damage was also observed in peripheral blood mononuclear cells from NG. The findings highlight the effectiveness of oral nucleotide administration, and potentially dietary supplementation of nucleotides, in boosting oxidative and immune status in newborn calves.

Estimation of maximum thermo-hygrometric index thresholds affecting milk production in Italian Brown Swiss cattle.

It is known that heat stress affects dairy cow performance in multiple ways: physiological, behavioral, reproductive, and productive. The aim of the present study was to determine if a threshold of temperature-humidity index (THI) exists for multiple milk production traits (milk yield, fat-corrected milk, protein and fat yield and percentage, energy-corrected milk, cheese production, and cheese yield) in Italian Brown Swiss dairy cows from the period 15 d before the day of the Italian Breeders Association test-day sampling. A 10-yr data set (2009-2018) containing 202,776 test-day records of 23,296 Brown Swiss cows was matched with the maximum THI. In all parities considered, no THI thresholds were observed for milk yield in Brown Swiss. In contrast, a THI threshold of 75 was identified for fat-corrected milk. No THI threshold was found for fat percentage, but fat yield showed the highest THI thresholds in cows of first and second parity. Protein yield and cheese production were affected by heat stress with average THI threshold of 74. The THI thresholds identified indicate that the Brown Swiss breed has higher thermal tolerance versus literature values reported for Holstein cows. As THI rises, Brown Swiss cows tend to produce the same volume of milk, but with a decreasing quality with regard to components. Further study is necessary to estimate the genetic component of heat tolerance, in Brown Swiss cattle, considering that the correct estimation of THI thresholds represents the first step to identify components that could be included in selection procedures.

Short communication: Sensory characteristics and volatile organic compound profile of high-moisture mozzarella made by traditional and direct acidification technology.

In the present study, the sensory characteristics and the volatile organic compound (VOC) profiles of high-moisture mozzarella made by different acidification techniques were compared. The cheeses were manufactured at the same dairy by fermentation by autochthonous natural whey starter (traditional backslopping method) and direct acidification with citric acid (the most used industrial technology). Three cheesemaking trials were performed from February to June using raw milk from a single farm. The mozzarella samples were subjected to assessment of the chemical, microbiological, and sensory characteristics and to VOC analysis by headspace solid-phase microextraction coupled to gas chromatography-mass spectrometry. The relevant microbiological differences found between the 2 types of products were reflected in different sensory and VOC profiles. The cheeses were clearly discriminated by the panel, and traditional mozzarella had higher intensity attributes. The most discriminating descriptors were elasticity, overall odor and taste intensity, sour milk and fruity/vegetable odors, sour taste, and aftertaste. Even though some variability was observed among the trials, the VOC profile of traditional product was always much more complex than that obtained by direct acidification. In both products the carbonyl compounds were the most abundant chemical class, accounting for about 50% of the total. In detail, 51 compounds were identified in the entire set of samples, and their contribution to cheese aroma was roughly estimated by calculating the odor activity values on the basis of the odor thresholds available in the literature. The results allowed hypothesizing that only 12 of them could play a primary role. The most important among the odor-active compounds was 3-methyl-butanal that can both derive from metabolism of lactic acid bacteria and yeasts. The results of the study may be very useful in view of European Union PDO labeling of the traditional product, in terms of protecting it from imitations.

Effects of whole linseed supplementation and treatment duration on fatty acid profile and endogenous bioactive compounds of beef muscle.

Diet supplementation with oilseeds is known to improve the fatty acid profile of meat, but few studies have been carried out to determine the time required for the incorporation of a significant quantity of n-3 polyunsaturated fatty acids (PUFA) into meat from steers. Therefore, the present study aimed to assess the effects of linseed supplementation and feeding duration on the fatty acid profile, cholesterol and bioactive compounds of bovine meat. In total, 54 Friesian steers were randomly allocated during the finishing period into six experimental treatments following a 2×3 factorial design. The six treatments consisted of two diets, the control diet (CO) with no supplemental fat and the linseed diet (LS) containing 10% whole linseed, fed 40, 75 or 120 days before slaughter. At the end of each finishing period, steers from the CO and LS groups were slaughtered. After 8 days of ageing chemical analysis, the fatty acid profile, cholesterol content and bioactive compounds were determined from the longissimus thoracis muscle. Including linseed in the diet increased the content of monounsaturated fatty acids, CLA and n-3 PUFA, and reduced the proportion of saturated fatty acids and n-6 PUFA. The percentage of myristic fatty acid increased with the duration of feeding, regardless of diet and a decrease in PUFA and n-6 PUFA was observed in the CO and LS diets, respectively. Furthermore, meat from steers fed linseed showed an increased percentage of n-3 PUFA, linolenic acid, and EPA from 40 to 75 days of feeding, whereas vaccenic acid, CLA 9c,11t, and total CLA increased from 40 and 75 days but declined at 120 days. Beef from the linseed group had a higher content of bioactive substances such as creatine, carnosine and anserine than beef from the control group. The duration of feeding significantly affected the creatine concentrations, with an increase in the LS group from 40 to 75 days of feeding. Feeding linseed did not modify the cholesterol content, on average and the lowest cholesterol content was found in meat after 75 days of linseed administration. This study demonstrates that a short-term diet manipulation is sufficient to improve the nutritional properties of meat, including n-3 PUFA and bioactive compounds.

Meat quality of farmed red deer fed a balanced diet: effects of supplementation with copper bolus on different muscles.

Supplementation with copper (Cu) improves deer antler characteristics, but it could modify meat quality and increase its Cu content to levels potentially harmful for humans. Here, we studied the effects of Cu bolus supplementation by means on quality and composition of sternocephalicus (ST) and rectus abdominis (RA) muscles (n=13 for each one) from yearling male red deer fed with a balanced diet. Each intraruminal bolus, containing 3.4 g of Cu, was administered orally in the treatment group to compare with the control group. Meat traits studied were pH at 24 h postmortem (pH24), colour, chemical composition, cholesterol content, fatty acid (FA) composition, amino acid (AA) profile and mineral content. In addition, the effect of Cu supplementation on mineral composition of liver and serum (at 0 and 90 days of treatment) was analysed. No interactions between Cu supplementation and muscle were observed for any trait. Supplementation with Cu increased the protein content of meat (P<0.01). However, Cu content of meat, liver and serum was not modified by supplementation. In fact, Cu content of meat (1.20 and 1.34 mg/kg for Cu supplemented and control deer, respectively) was much lower in both groups than 5 mg/kg of fresh weight allowed legally for food of animal origin. However, bolus of Cu tended to increase the meat content of zinc and significantly increased (P<0.05) the hepatic contents of sodium and lead. Muscles studied had different composition and characteristics. The RA muscle had significantly higher protein content (P<0.001), monounsaturated FA content (P<0.05) and essential/non-essential AA ratio (P<0.01) but lower pH24 (P<0.01) and polyunsaturated FA content (P=0.001) than the ST muscle. In addition, RA muscle had 14.4% less cholesterol (P=0.001) than ST muscle. Also, mineral profile differed between muscles with higher content of iron, significantly higher (P<0.001) content of zinc and lower content of calcium, magnesium and phosphorus (P<0.05) for ST muscle compared with RA. Therefore, supplementation with Cu modified deer meat characteristics, but it did not increase its concentration to toxic levels, making it a safe practice from this perspective. Despite the lower content of polyunsaturated FA, quality was better for RA than for ST muscle based on its higher content of protein with more essential/non-essential AA ratio and lower pH24 and cholesterol content.

Glucocorticoid effects on sheep peripheral blood mononuclear cell proliferation and cytokine production under in vitro hyperthermia.

The present experiment aimed at understanding the effects of cortisol levels on sheep peripheral blood mononuclear cell (PBMC) proliferation and cytokine production during hyperthermia. To mimic stress related to the exposition of high ambient temperatures, PBMC were cultured at 43°C for 12 h, and subsequently at 39°C for additional 12 h. Cells in normothermia were cultured at 39°C for 24 h. Phytohemagglutinin-stimulated PBMC were cultured with different cortisol levels: 0 ng/mL; 100 ng/mL, representing the physiological cortisol concentration simulating stress condition (Cort100); and 1,000 ng/mL, representing the hyperactivated hypothalamic-pituitary-adrenal axis (Cort1000). Phytohemagglutinin-stimulated PBMC with 0 ng/mL of cortisol concentration represented the positive control, whereas nonstimulated PBMC without cortisol represented the negative control (NC). The free cell supernatants were collected for the determination of IL-6, IL-1ß, and IL-10 by ELISA. Bromodeoxyuridine assay was performed on cells to determine cell proliferation. Exposition to hyperthermia negatively affected cell proliferation, IL-6, IL-1ß, and IL-10 concentrations in cell supernatants. The interaction of hyperthermia and cortisol level affected both cell proliferation and IL-10 production. Both PBMC proliferation and IL-10 production in positive control, Cort100, and Cort1000 decreased at 43°C as compared with 39°C NC. On average, the Cort100 treatment displayed higher concentrations of IL-6 than NC. The present experiment demonstrated that the action of cortisol concentration simulating stress condition on cell proliferation and cytokine production was a permissive/stimulatory action during normothermia, whereas it was a suppressive action during hyperthermia. These data confirmed that cortisol concentration simulating stress condition could have a role in the immune system of sheep via mediating cellular homeostasis in the condition of hyperthermia. The negative effects of hyperthermia on sheep immune responses were apparent when performing an immunological challenge.

Survey on basal blood plasma catecholamine concentrations in Martina Franca donkey (Equus asinus).

BACKGROUND: Catecholamines are among the most frequently investigated parameters for studying sympathoadrenal activity in response to stress conditions. OBJECTIVES: To evaluate basal plasma concentrations of catecholamines (adrenaline, noradrenaline and dopamine) in healthy donkeys. STUDY DESIGN: Cross-sectional study. METHODS: Catecholamine concentrations from 440 Martina Franca donkeys were determined: 269 females and 171 entire males, aged from 4 months to 24 years. Animals were subdivided into four age categories: under 12 months old (64 males and 54 females), from 13 to 36 months (56 males and 75 females), from 37 to 120 months (49 males and 80 females) and over 120 months (24 males and 38 females). Indwelling jugular catheters were inserted at least 12 h before drawing blood. The data set was subjected to analysis of variance considering age, sex and the two-way interaction between them as independent variables. Pearson's correlation coefficients between the three catecholamines were evaluated. RESULTS: Confidence intervals (CI) for noradrenaline concentration ranged between 239.98 and 255.07 ng/L (mean 247.52 ng/L), for adrenaline between 129.27 and 137.90 ng/L (mean 133.59 ng/L), dopamine concentrations between 149.62 and 160.80 ng/L (mean 155.21 ng/L) and noradrenaline/adrenaline ratio between 1.91 and 2.05 (mean 1.98). Catecholamine plasma concentrations were not influenced by sex. Donkeys older than 37 months had lower adrenaline and noradrenaline plasma concentrations (P<0.001) and higher noradrenaline/adrenaline ratios (P<0.01) than younger animals. MAIN LIMITATIONS: Indwelling catheters and blood drawing procedures may have influenced catecholamine levels. CONCLUSIONS: Catecholamine concentrations were established within a large group of healthy Martina Franca donkeys.

Martina Franca donkey meat quality: Influence of slaughter age and suckling technique.

This work aimed to evaluate the effect of suckling technique and slaughter age on Martina Franca donkey meat quality. Twenty Martina Franca male foals were involved in the trial. Foals naturally assumed colostrum within 4h from birth. Afterwards, 10 foals were partially artificially suckled (AS), and 10 foals were naturally suckled (NS). All the foals were weaned at 180d, then housed indoors and fed the same diet. Ten donkeys were slaughtered at 12months and the other 10 at the age of 18months. Samples of Longissimus thoracis et lumborum (LTL) were taken from each foal for chemical analysis, then rheological parameters, oxidative profile, colorimetric parameters and fatty acid profile were assessed. Older donkeys (18months) fed with natural milk presented the highest intramuscular fat (IMF) and meat protein content. From a dietary view point, IMF acid composition showed a more favourable profile in meat from artificially-reared donkeys compared to naturally-suckled ones.

Effects of two different packaging materials on veal calf meat quality and shelf life.

The aim of the present work was to evaluate the effects of 2 different packaging films on chemical, physical, and microbiological variables in veal calf meat. Slices from the right half carcass were sampled from each calf: 8 were packaged with Cryovac film (25 µm thick) and 8 were packaged with Weegal film (45 µm thick). In both packaging types, the same gas mixture was used. The samples were analyzed at these postpackaging times: 0, 2, 4, 6, 8, 10, 12, and 14 d. Chromatic patterns, water holding capacity (WHC), cooking loss, drip loss, protein oxidation, and hydroperoxide concentration were influenced by the packaging type (P < 0.001). Moreover, during the postpackaging time the WHC decreased but oxidative reactions increased (P < 0.001). The film that showed better characteristics in packaging of veal calf meat was the Weegal, characterized by decreased gas permeability due to its ability to reduce and delay chemical and physical alterative processes.

Slaughtering age effect on carcass traits and meat quality of italian heavy draught horse foals.

The present work describes the effect of slaughtering age on horse carcass traits and on meat quality. Eighteen male Italian heavy draught horse (IHDH) breed foals were employed in the study. Soon after foaling they were randomly subdivided in 3 groups according to 3 age at slaughtering classes: 6 months old, 11 months old and 18 months old. Live weight, hot carcass weight and dressing percentage of each animal were recorded. After slaughtering, meat samples were collected from Longissimus Dorsi muscle between 13th and 18th thoracic vertebra of each animal and then analyzed. The right half carcass of each animal was then divided in cuts. Each one was subdivided into lean, fat and bones. Then, the classification of the lean meat in first and second quality cuts was performed according to the butchers' customs. Older animals were characterized by a lower incidence of first quality cuts (p<0.01) on carcass. Younger animals showed greater content in protein (p<0.01). Fatty acid profile showed an increasing trend of PUFA connected to the increasing of slaughtering age (p<0.05). The unsaturation index of intramuscular fatty acids was not affected by slaughtering age, confirming that horse meat, if compared to beef, is more suitable from a nutritional point of view. Season influenced reproduction, birth as well as production aspects of this species. The different slaughtering age could represent the way to produce meat of IHDH foals during the entire year without change in the qualitative standard expected by consumers.

Application of Wood's model to lactation curve of Italian Heavy Draft horse mares.

The aim of this work was to study, using Wood's model, the lactation curve in Italian Heavy Draft horse (IHDH) mares, both for milk yield and composition. Interest in mare's milk has grown for use in human nutrition, as a substitute for human and cow milk for premature newborns and allergic children. For this study, 7 IHDH mares were used. Milk yield was evaluated from d 15 to 180 postpartum (every 15 d). Wood's lactation model was used to describe milk yield, fat, protein, and lactose lactation curves in the first 180 d in milk (DIM), with a prediction to 210 DIM. Mean values recorded across the considered lactation period for milk yield and constituents were 16.11 kg of milk/d, 11.38 g of fat/kg, 15.54 g of protein/kg, and 69.72 g of lactose/kg. The lactation curve showed a peak at 69 DIM, with a persistence index of 6.26. Fat (g/kg) and protein (g/kg) lactation curves decreased during the lactation period; instead, the lactose (g/kg) curve increased from the first to the last DIM. The coefficient of determination values indicated a good model fit by Wood's model application to IHDH mare lactation. The research showed good aptitude of a heavy horse breed, such as IHDH, for milk production.

Colour Changes in Meat of Foals as Affected by Slaughtering Age and Post-thawing Time.

The aim of the present work was to investigate how colour changes of foal meat can vary after thawing out in relation to the slaughtering age of the horses and to the post-thawing time. Eighteen Italian Heavy Draught Horse (IHDH) foals were used for the trial. They were subdivided in three groups according to their slaughtering age (6, 11 and 18 months). Two different surfaces were investigated for each sample: a fresh cut surface (daily renewed cutting surface: DRCS), and not-renewed cutting surface (NRCS). The redness of both investigated surfaces increased with slaughtering age (p<0.01). Moreover, this parameter decreased during post-thawing time (p<0.01) only on the NRCS, probably due to the myoglobin oxidation processes. Colour is an important visual cue denoting perceived quality by consumers. So, by a chromatic perspective the thawed meat of IHDH foals slaughtered at 6 and 11 months proved to be that which best meets the market requirements.

Podolian beef production on pasture and in confinement.

From February to August 2007 the effect of rearing system (confined (C) v. free ranging (FR)) and season (spring v. summer) was evaluated on behaviour, immune and blood parameters and beef production (experiment 1). From February to August 2008, the effect of rearing system was only evaluated on beef production (experiment 2). A total of 12 Podolian young bulls were used each year. They were slaughtered at 18 and 15 months of age in 2007 and 2008, respectively. Herbage mass and pasture composition were monitored during the 2 years. Pasture availability, in terms of herbage mass and composition, as well as its seasonal development, was similar in the 2 years. In the first experiment, FR animals spent more time walking (P < 0.05), feeding (P < 0.001) and standing (P < 0.01) and showed a lower number of agonistic (P < 0.05) and non-agonistic social interaction than C animals (P < 0.01). Significant lower concentrations of serum urea nitrogen (P < 0.001) and creatinine (P < 0.10) in FR animals indicated a lower protein nutritional status due to inadequate protein availability at pasture. As a consequence, average daily gains (P < 0.05), slaughter weight (P < 0.05) and body condition score (P < 0.01) were lower in grazing animals as compared with C bulls. Cellular immune responsiveness was higher in FR animals (P < 0.05). Similarly, antibody titre to keyhole limpet hemocyanin was higher in FR bulls at the 2nd and 3rd month after antigen injection (P < 0.05), whereas it tended to be higher at the 4th month (P < 0.10). In both experiments, grazing negatively affected meat colour in terms of lightness. Eighteen-month-old bulls also showed lower final weight, weight gain and body conditions when kept outdoor: a possible consequence of nutrient deficits, as suggested by the metabolic status of FR animals. The same animals, however, benefited from FR in terms of natural behaviour expression and immune responsiveness. When the experiment was replicated the subsequent year, on animals slaughtered at 15 months of age, no differences between the performances of FR and C animals were detected. The earlier slaughter age system was also proportionally less dependent on external inputs as grazing was not extended to the dry season when herbage mass availability was lower.

Physicochemical properties of meat of Italian Heavy Draft horses slaughtered at the age of eleven months.

To investigate rheological and chemical characteristics of the meat from Italian Heavy Draft horse, 24 foals (12 males and 12 females) were weaned at 6 mo, reared and fed in the same way, and slaughtered at 11 mo of age. The results obtained showed that there were no significant differences between the sexes but that the muscle type is a significant variation source. The rectus femoris muscle was lighter, and the biceps femoris had a lower a* index than longissimus dorsii, rectus femoris, and semimembranosus muscle. The most tender muscle was the semitendinosus, and the toughest even after cooking was the biceps femoris. The male animals had a greater protein and lipid percentage. The acidic composition of the intramuscular fat showed a greater presence of MUFA (P < 0.05) in the females and of PUFA (P < 0.01) in the males. Colorimetry analysis of the subcutaneous and perirenal fat from these animals indicated the unfavorable yellow color seen in adult animals had not yet been acquired. The meat produced had low redness due to the low myoglobin content and high lightness. Besides, the low collagen content and its high solubility indicates an appreciable tenderness. The high level of unsaturation of the intramuscular fat resulted in a high ratio of unsaturated to saturated fat, making horse meat favorable from a health point of view.

Some qualitative and chromatic aspects of thawed buffalo (Bubalus bubalis) meat.

After thawing, the meat of beef calves (Italian Frisian breed) and buffalo calves (Mediterranean breed) slaughtered at 4, 8 and 12 months of age was examined. Both the pH and the thawing loss confirmed that the meat of buffalo calves is more suitable for preservation by freezing. With increased age and time of exposure to air the lightness of the non-renewed surface was reduced. The lightness of the fresh cut surface remained stable in the various thawing phases though it was less in the older animals. The a(∗) index increased with animal age but decreased during the 4 days post-thawing. The fresh cut surface of buffalo meat from calves slaughtered at 4 and 8 months was not darker than beef slaughtered at the same age. On the contrary at 12 months of age, the buffalo meat had a lower redness index than beef and a higher haematin concentration.

Influence of free-stall flooring on comfort and hygiene of dairy cows during warm climatic conditions.

An evaluation of behavioral and hygienic conditions was carried out with 4 materials used as free-stall flooring for dairy cows: polyethylene vinyl acetate (EVA) and polypropylene vinyl acetate (PVA) mats, wood shavings, and solid manure. The free-stall type selected by cows was evaluated in response to changes in environmental temperature and humidity. Two tests were used: 1) a preference test, in which 8 cows were housed in a pen with 32 free stalls and 4 types of flooring; and 2) an aversion test, in which 32 cows were placed in 4 pens, each with 8 free stalls. The free stalls in each pen had a single type of bedding material. These tests showed that the comfort of dairy cows was predominantly influenced by environmental conditions. The preference test for lying showed that cows preferred free-stall floors with EVA mats over those with PVA mats, wood shavings, and solid manure (332.4 +/- 24.0 vs. 130.8 +/- 6.2, 160.9 +/- 23.7, and 102.6 +/- 23.2 min/d, respectively), but under conditions of heat stress, with a temperature-humidity index > 80, they chose wood shavings and solid manure lying areas. These results were confirmed by the aversion test. In all experimental and environmental conditions, the PVA mats were the least suitable. The mats contaminated with organic manure and the free stalls bedded with wood shavings and organic solids did not differ in either the coliform load on the lying surfaces (EVA mats: 290 +/- 25; PVA mats: 306 +/- 33; wood shavings: 290 +/- 39; and solid manure: 305 +/- 23 log(10) cfu/mL) or the total bacterial count in the raw milk (EVA mats: 232 +/- 22; PVA mats: 233 + 24; wood shavings: 221 +/- 24; and solid manure: 220 +/- 25 log(10) cfu/mL). These results demonstrate that the comfort of dairy cows housed in barns with free stalls as resting areas does not depend only on the material used, but also on the value of the material in microenvironmental conditions.