1. Introduction
Feeding cattle high-grain (HG) diets is commonly implemented in the dairy and beef cattle production industry to enhance the energy intake for milk or accelerate daily gains, respectively. Grains are rich in starch and less voluminous and are thereby a better source of metabolizable energy than forages for cattle diets in many parts of the world. However, feeding large amounts of grain is known to influence chewing behavior, which could affect animal health due to the increased risk of gut disorders. More specifically, HG diets have been reported to impair rumination and total chewing times, which are essential physiologic processes in ruminants [
1,
2,
3,
4] often used as indicators of cattle welfare and health [
5]. Additionally, the lying behavior of cattle is an important indicator of their comfort and welfare [
6]. For instance, Haley et al. [
7] demonstrated that lying time is closely related to comfort and health changes. Cattle ruminate more while lying down than when standing. However, feeding them HG diets may change the lying behavior of cows [
2], with longer lying times and shorter rumination times usually reflecting distress and discomfort [
8].
The feed-sorting behavior in cattle may also be affected by a change in dietary composition [
9,
10,
11]. For example, cows are known to select diet fractions during eating, sorting for shorter particles in the ration (concentrate) and refusing longer particles (forages) [
12,
13,
14]. Another report by Greter and DeVries [
15] demonstrated that cows fed with a 54% grain diet sorted against long particles and tended to sort in favor of short particles. Nonetheless, this feeding behavior may contribute to further impaired chewing activity and salivation due to the reduced intake of dietary physically effective NDF (peNDF) and could consequently affect animal health or gut function. Specifically, peNDF in the diet is important because it stimulates chewing activity, greater salivary buffer secretion, and the regulation of ruminal pH [
16]. Therefore, diet composition plays an important role in feed sorting, with the time invested in eating and ruminating positively correlated with the intake of peNDF [
17]. In this regard, diets with more fiber are associated with more meals per day and reduced eating rates [
18], which positively modulates rumen fermentation.
The essential physiologic role of chewing in cattle is based on its contribution to stimulating salivary secretion [
19,
20]. Salivary buffers help stabilize the ruminal pH [
21] because salivary buffers such as bicarbonate and phosphate represent important components for ruminal proton removal in the rumen [
22]. Furthermore, salivary secretions contain different proteins such as mucins, lysozymes, and immunoglobulins [
23], which contribute to health and gut function [
24]. Therefore, an increase in mastication and salivary flow can enhance the rumen acid–base balance and ultimately improve health [
5,
25]. In this context, phytogenic compounds, such as thymol and thyme oil, have shown a potential to modulate the salivary secretions in cattle. Similarly, menthol has been reported to stimulate chewing and increase salivation in non-ruminants [
26,
27].
The dietary shift to HG feeds is known as the time with the greatest risk for cattle health due to the major adaptive changes occurring in the host during this interval. However, there is limited research on the effects of the duration of the HG feeding challenge after the dietary change on salivary secretions [
28] as well as the chewing activity and lying behavior. In addition, there is a paucity of information regarding the effect of the supplementation of feed with phytogenic compounds on salivary composition and production [
29], chewing activity, or the eating behavior of cows. Thus, there is a need to fill or strengthen these research gaps in the scientific literature. Therefore, the aims of this study were to evaluate the effect of the duration of an HG feeding challenge on the chewing activity, eating and lying behavior, and salivary composition and production in dairy cows without or with phytogenic feed supplementation. Our hypothesis stated that the advanced duration of the HG challenge would exacerbate the negative effects on rumination, the eating and lying behavior, and the salivary production and composition. We also hypothesized that the phytogenic supplementation would alleviate the decrease in the chewing activity as well as improve the feed sorting and salivary properties.
4. Discussion
This study aimed to evaluate the effect of the duration of an HG feeding challenge on chewing, eating, and lying behaviors as well as the salivary production and composition in cows with or without a phytogenic feed supplement. In agreement with our hypothesis, there was a reduction in the rumination time not only at the start of the HG feeding regimen, but also due to advanced duration on the HG diet. Consequently, the reduction in the rumination time likely contributed to the lower feed ensalivation observed after 4 weeks of the HG feeding regimen. Furthermore, the total chewing time decreased because of the duration on the HG diet. Chewing activity is essential for adequate rumen function because it stimulates salivation and normal rumen fermentation [
16]. It has also been reported that a greater chewing time improves feed digestion by exposing nutrients and increasing feed surface area, thereby facilitating the activity of microbial enzymes [
41]. Therefore, our findings clearly indicate that the duration on an HG diet exacerbates the negative effects on the chewing activity in cattle. Our observations for rumination time were lower than the values reported by Ben Meir et al. [
42] for lactating cows consuming diets with similar levels of concentrate. These contrasting findings may be because of the higher DMI intake in lactating cows used by Ben Meir et al. [
42], with intakes twice as high as the dry cows utilized in our study, resulting in more ruminal digesta available for rumination. Although there are limited data regarding the effects of phytogenic supplementation on rumination, the research conducted by our group showed that a blend of essential oils increased the rumination time during the first 2 weeks of an HG feeding challenge compared to a control TMR [
43]. In another experiment, Castillo-Lopez et al. [
29] demonstrated in a short-term trial that thymol supplementation tended to increase chews/min. However, in the present study, this effect was not observed, suggesting that thymol may exert only a temporary stimulating effect on chewing activity.
Our findings also indicate that the DMI increased from the first week of the HG feeding regimen. These results support reports from Dann et al. [
44] for diets containing similar levels of starch. The increase in the DMI might have occurred because of the small particle size of the HG diet, which allows for a greater feed intake due to the decreased gut fill. In addition, the greater feed consumption may be explained by the improved feed acceptability with the inclusion of concentrate in the diets. Our findings showed a greater meal size and eating rate due to the advanced duration on the HG diet, which is a factor that increases the risk of ruminal acidosis, because this results in the accumulation of volatile fatty acids in the rumen. The latter observations explain the greater feed boli measured after 4 weeks on the HG. In general, the number of meals per day observed in this study were lower compared to previous studies in lactating cows [
18,
33,
42]. This may be because the nutrient utilization and energy metabolism are slower in dry cows compared to lactating animals. With regard to the effect of the TRT on the DMI, reports suggest that individual phytogenic compounds may influence feed intake [
29]. In this trial, the TRT tended to reduce meal sizes in week 2 on the HG diet compared to the CON. This may be explained by the increased intake of long feed particles and peNDF for the TRT in that week, which contributed to gut fill. The increased preference for long feed particles possibly was due to olfactory and gustatory stimulation of TRT. This is a beneficial effect because of the role of fibrous feed ingredients in the regulation of ruminal pH, particularly when there is a need to modulate fermentation due to a low ruminal pH, as reported in the companion paper [
30]. Furthermore, the findings from the correlation analyses in this study agree with our expectations and showed that a greater amount of saliva per gram of the feed that cows consume contributes to an increased ruminal pH by neutralizing the acids produced in the rumen. The positive association between the feed ensalivation and the ratio of acetate to propionate agrees with the simultaneous change in the feed ensalivation and acetate production due to a change in the proportion of the concentrate in the diets. For example, forage-based rations are associated with greater feed ensalivation and an increased acetate production.
The majority of the studies on feed sorting behavior have evaluated feed management or the fiber of the diet [
3,
25,
45], but the influence of the duration of the HG challenge or phytogenic supplementation on feed sorting remains yet to be elucidated. Interestingly, the present study showed that the advanced duration on the HG diet decreased the cows’ preference for the short feed particles. This may reflect a response of the cows to counteract the negative effects of low fiber diets on rumen pH, because short feed particles are rich in readily fermentable starch that increase ruminal fermentation and acidification.
Our hypothesis also stated that the lying time would increase with the duration of the HG feeding regimen. Several experiments have demonstrated that there is a close relationship between the standing and lying time and laminitis in cows. For example, reduced lying times and abnormal standing times seem to be indicators of the development of laminitis [
46]. In this study, we found that there was an increase in the number of lying bouts on either the right or left side from the start of the HG feeding regimen, and these increased values were maintained throughout the HG feeding challenge. Greater lying bouts may reflect animal discomfort and may be due to the effect of the acidogenic diets that lead to damage in the lamina of the foot [
47]. Furthermore, Fukasawa et al. [
48] reported similar results, describing a tendency to increase lying bouts when implementing high concentrate feeding compared to forage feeding. The higher nutritive value of the diet has been suggested to influence lying time or lying bouts as well, with increasing lying times associated with a greater body condition score [
6]. In this study, there was an increase in the body weight of cows (69 kg), which could have contributed to the greater lying bouts throughout the HG feeding.
Our results show that laterality of the lying behavior followed a similar pattern as reports from Tucker et al. [
49], with a fairly even distribution between the left or right lying times. Previous research has demonstrated that cows preferably ruminate while lying down [
17,
50], which coincides with our findings. It is possible that as the cows experienced increased discomfort as a consequence of intensive rumen fermentation, they preferably ruminated while lying on the left side. These results are supported by pioneering findings by Bailey and Balch [
51] and Albright [
52], who suggested that lying on the left side is a strategic position for cows to increase rumination efficiency, because this position may facilitate the regurgitation process of the digesta due to the improved alignment of the esophagus with the rumen contents. In addition, the TRT group increased its propensity to lie on the left compared to CON, especially in the second week on the HG diet. Nonetheless, the exact association between the TRT supplementation and the lying side of cows remains to be elucidated.
Another aim of this study was to evaluate the changes in salivary composition and dynamics. In agreement with our hypothesis and with previous studies [
28,
35], the change to an HG diet had a negative impact on feed ensalivation. Beauchemin et al. [
35] also suggested that eating time may influence salivary secretion in cows, which supports our findings showing that a lower eating time was associated with a lower feed ensalivation in week 4 on the HG diet. Moreover, salivary secretion is not influenced by rumination alone. For instance, it has been demonstrated that when the eating rate increases, there is a reduction in the daily salivation of cows [
35]. This supports our findings showing that when the cows consumed the HG rations at a faster pace, the feed ensalivation was lower. With regard to the effect of phytogenic compounds on salivation, the increased salivation rate via individual phytogenic compounds previously reported [
37] in a short-term trial was not confirmed in the present study. These contrasting findings may be because the stimulus for the salivation flow of individual phytogenic compounds could decrease when combined with other compounds. It is also possible that differences in the effects of these compounds are related to their distinct modes of action, with some substances being active in the oral cavity [
53], while others influence salivary secretion through the olfactory stimulation of the nervous system [
54,
55].
Salivary physico-chemical properties play important roles in gut function and health. In this trial, we observed that salivary lysozyme activity increased at the start of the HG feeding regimen. The salivary lysozyme is known to act as an antimicrobial bioactive component [
56,
57]. Thus, this observation may reflect a host response to counteract a potential outgrowth of pathogens due to the drastic diet shift [
57]. Additionally, we found that salivary pH increased with the advanced time on the HG diet, which may be a host response for ruminal pH regulation, given the role of saliva for proton removal and ruminal pH balance. On the other hand, we found a reduction in salivary phosphate during the first 2 weeks on the HG, an observation that is highly relevant because of the role of this salivary buffer in the regulation of ruminal pH. The latter effect may exacerbate the reduction in ruminal pH commonly observed when cattle are switched from forage to HG rations. Another finding was the tendency for TRT to increase the salivary buffer capacity in the third week of the HG feeding regimen, and to increase the salivary pH in the fourth week compared to CON; these changes may have contributed to the greater ruminal pH for TRT reported in those weeks in the companion paper [
30]. However, there was no increment in bicarbonate or phosphate due to TRT. These findings indicate that the salivary pH or buffer capacity are also influenced by factors other than salivary major buffers [
19,
51]. Although the mechanism by which TRT supplementation influences buffer capacity or pH is not clear at the moment, TRT possibly influenced the profile of specific proteins in saliva [
58] increasing salivary pH, a topic that deserves further investigation. Another potential explanation to the change in the salivary buffer capacity and pH is the hydration status, as reported in other studies [
59], indicating that hydration status affects these salivary variables in other animal species.