INTRODUCTION:

The prevalence of hydroelectrolyte and acid-base disorders is common in variable bovine diseases; therefore, fluid therapy has become an indispensable part of the buiatric routine (^{RIBEIRO FILHO et al., 2013}). In this way, accurate evaluation of dehydration and acid-base disorders is necessary for optimal hydroelectrolyte replacement.

In bovine medicine, fluid therapy is most commonly administered by intravenous and oro-ruminal routs (^{RIBEIRO FILHO et al., 2013}). Due to the large volumes administered, professional supervision is required during fluid therapy, and the final cost of administration via the parenteral route is sometimes unviable (^{RIBEIRO FILHO et al., 2011}). Consequently, this route of fluid therapy is only used under critical scenarios, such as hypovolemic shock.

Enteral fluid therapy, like the oro-ruminal route, allows a large volume of electrolyte fluid to be administered directly into the rumen in cases of mild and moderate dehydration (^{ROUSSEL, 2014}), and is used careful in cases with motility disorder. Although, this technique is simple, when performed over several days, successive tube passages increase the risk of laryngeal and esophagus lesions. In addition, large volumes of fluid can cause some discomfort to the animal when administered quickly into the rumen.

An alternative method that has been used and studied in Brazil for more than 10 years, is enteral fluid therapy with continuous flow, using a naso-gastric or naso-ruminal tube with a small diameter. This technique has been used successfully in horses (^{AVANZA et al., 2009}; ^{RIBEIRO FILHO et al., 2012}; ^{RIBEIRO FILHO et al., 2015}); owing to the small gastric capacity of this species, several doses or slow and continuous administration is required throughout the day.

In goats (^{ATOJI-HENRIQUE et al., 2012}), calves (^{RIBEIRO FILHO et al., 2017}), and bubaline calves (^{ERMITA et al., 2016}) continuous flow via the naso-ruminal route has been tested and promising results have been reported. Some studies have already been performed in adult cattle (^{RIBEIRO FILHO et al., 2009}; ^{RIBEIRO FILHO et al., 2011}; ^{RIBEIRO FILHO et al., 2013}); however, many questions remain unanswered, including osmolarity, the correct electrolyte composition, energy precursor, and infusion rate for these animals. This is because this technique is not widespread in this species, and animals need to remain in an individual stall during therapy.

One important characteristic of enteral fluid therapy is the possibility of creating new solutions with altered compositions that meet the needs of the animal and increase the therapeutic efficiency (^{RIBEIRO FILHO, 2011}). When used in continuous flow, the deleterious effects described above are minimized, besides not preventing the animal from walking and laying inside the stall and feeding during fluid therapy.

According to ^{CONSTABLE (2003}), solutions for enteral use should contain sodium, potassium, chloride, calcium, phosphate, and a glycemic precursor, such as propionate. In addition, ^{RIBEIRO FILHO et al. (2014}) stated that by maintaining volemia and electrolyte balance, the solution should not cause any adverse events.

The objective of this study was to evaluate the effects of enteral fluid therapy administered as continuous flow by the naso-ruminal route with three enteral electrolyte solutions, containing different energy precursors, on the physiological biomarkers and the hemogram of clinically healthy cattle.

MATERIALS AND METHODS:

This experiment included six healthy Holstein heifers from the flock of the Experimental Research and Extension Unit in Dairy Cattle of the Federal University of Viçosa (Unidade Experimental de Pesquisa e Extensão em Gado de Leite da Universidade Federal de Viçosa), aged 16 and 18 months and with a mean body weight of 300kg. Animals were adapted to the experimental environment for 10 days before the beginning of the study, during which they were restricted in an individual stall in a Tie Stall system. Animals were fed with corn silage as a balanced ration and water was provided ad libitum.

A crossover 6×3 design was adopted (six animals × three treatments) and all animals received all treatments. To avoid an overlap effect, there was a 7-day interval between treatments. The composition of the electrolyte solutions was as follows: Solution with calcium propionate solution (SEPCa), 4g of NaCl^a, 0.5g of KCl^a, 0.3g of MgCl₂ ^aand 10g de calcium propionate^b in 1000mL of water (measured osmolarity: 299mOsm/L); glycerol solution (SEGly), 4g of NaCl^a, 0.5g of KCl^a, 0.3g of MgCl₂ ^a, 1g of calcium acetate^a, and 10mL of glycerol^c in 1000mL of water (measured osmolarity: 287mOsm/L); and a solution with propylene glycol, 4g of NaCl, 0.5g of KCl, 0.3g of MgCl₂, 1g of calcium acetate, and 15mL of propylene glycol in 1000mL of water (measured osmolarity: 378mOsm/L).

A naso-ruminal tube with a small diameter (4mm diameter and 1.8m length) was used to administer fluid. The tube was attached to the halter and connected to a gallon with a 20L capacity with a spiral hydration system^d, set 1.5m above the head of the animal. The continuous flow rate was 15mL/kg/h, over 12 hours, and animals did not receive food or water during the hydration period. After the hydration period, the animals were in water and food fasting for 12 hours, for a 24-hour experimental cycle. Samples were collected and evaluated at the following times: T0h, just before the beginning of the experiment; T3h, 3 hours after the beginning of hydration; T6h, 6 hours after the beginning of hydration; T9h, 9 hours after the beginning of hydration; T12h, 12 hours after the beginning of hydration and the end of hydration period; and T24h, 12 hours after the end of hydration (observation period).

The following physiological parameters were analyzed: heart rate (HR, bpm), respiratory rate (RR, mpm), rectal temperature (RT, °C), ruminal movements (RM in 3 minutes), abdominal girth (AG, cm), and fecal humidity (FH%). Blood samples were collected via jugular venipuncture using a vacuum system^e in K₂EDTA tubes, and blood counts were determined in an automatic blood counter device^f. White blood cell counts were obtained by blood scrubbing as recommended by ^{STOCKHAM & SCOTT (2011}). Abdominal girth was calculated using a measuring tape across the flank of the bovine. Feces were collected directly from the rectal ampulla and were immediately weighed and dried in a drying oven at 60°C for several days. The feces were weighed until a constant weight was achieved. Humidity was determined using the following equation: [(initial weight - final weight) × 100/initial weight].

Results were analyzed descriptively to obtain the mean and the standard deviation. The normality of the data was evaluated with the Shapiro-Wilk test and the sphericity of the variances was determined with the Mauchly test. When the pre-requisites of normality and sphericity were achieved, analysis of variance (ANOVA) based on the factorial design of repetitive measures was used to evaluate the main effects across time, treatments, and in the time^*treatment effect. Multiple comparisons were compared using the Tukey post hoc test. When it was not possible to use ANOVA, the Friedman test with the Wilcoxon post hoc test associated with a Bonferroni correction was used. All analyses were performed with the statistical package SPSS 20 (IBMD, Statistic). Significance was considered when P<0.05.

RESULTS AND DISCUSSION:

Physiological variables have been used to evaluate the welfare and adaptation of cattle under variable edaphoclimatic and management conditions (^{DINIZ et al., 2017}). In this study, a decrease (P<0.05) in the heart rate of animals in the SEPCa group was observed during the hydration phase, and the respiratory rate remained the same (P>0.05) at baseline in the three groups. Notably, under all situations, these biomarkers remained within the reference values suggested by ^{FEITOSA (2004}). In this way, the data observed in the present study (Table 1), confirmed that enteral fluid therapy administered as continuous flow under the proposed conditions (solution type and fluid therapy technique), does not cause stress to the animals, demonstrating that the use of naso-ruminal tubes and the administration of these solutions did not cause discomfort to animals. This was clearly observed during the hydration period when the heifers were mostly laying down, quiet, and ruminating, as previously described by ^{RIBEIRO FILHO et al. (2011}, ²⁰¹³).

Several factors can alter body temperature, including exercise, environmental temperature, digestion, water intake, disease, and time of day (circadian or nictemeral variation), which can cause variation of 1.5°C in 24 hours (^{FEITOSA, 2004}). During the fluid therapy period between T0h and T3h, the rectal temperature of animals in the SEPCa group, was not correlated with the technique used or experimental time; this variation was probably caused by the circadian variation and, at all time points observed, the temperature remained inside the reference interval for the species (^{REECE, 2015}). Stability in rectal temperature was also observed by ^{RIBEIRO FILHO et al. (2015}) and ^{ERMITA et al. (2016}).

The ruminal movements were not affected over time with either treatment, as described by ^{ATOJI-HENRIQUE et al. (2012}). Effects of enteral fluid therapy on gastrointestinal tract motility seem to be associated with the dose and solution used, as described by ^{RIBEIRO FILHO et al. (2012}) who used enteral fluid therapy in healthy horses and observed an increase in intestinal motility. Although, using the same dose in the present study (15mL/kg/h), solution used by authors contained different concentrations of electrolytes and other energy sources. This suggested that in addition to physiological differences between species, the solution used could be a major factor for the effects observed in the intestinal tract.

Abdominal girth has been utilized in studies investigating enteral fluid therapy to verify the degree of abdominal distention, as some energy sources can sustain ruminal microbiota fermentation. This can result in gas production and accumulation inside the gut, causing discomfort or conditions such as a bloat. This kind of change was not observed in the present study, as there was no significant change in abdominal girth (P>0.05) across the whole fluid therapy period under all treatments, corroborating the findings of ^{RIBEIRO FILHO et al. (2013}). The choice of energy precursor directly affects these findings, because calcium propionate, glycerol, and propylene glycol can be oxidized by the ruminal microbiota or absorbed in the integral form, not over-stimulating gas production.

There was no variation in fecal humidity (P>0.05) over time or between treatments. This result confirmed that the electrolyte solutions proposed were readily absorbed by the intestinal tract with minimal losses through feces. Osmolarity is an important characteristic for the efficacy of an electrolyte solution, as hypo or isotonic solutions are absorbed quickly and in large amounts, as suggested by ^{AVANZA et al. (2009}). Hypertonic enteral solutions (osmolarity>320mOsm/L) could have decreased volemic effects, remaining longer in the intestinal lumen, increasing the risk of osmotic diarrhea; however, this was not observed in the present study, even for the SEProp group with 378mOsm/L, which was well tolerated by the animals.

The concentrations of red blood cells and hemoglobin (Table 2) decreased significantly after 6 hours of fluid therapy (T6h) with all treatments, and remained low over 12 hours of fluid therapy (T12h) in the SEGly and SEProp groups, returning to basal levels at 24 hours (T24h). These results confirmed that the solutions were well absorbed by the gastrointestinal tract and that the osmolarity of SEProp did not limit its use. The expanse capacity of the solutions (hemodilution) is demonstrated by the decreased hematocrit concentration (P<0.05) observed following 3 hours of fluid therapy (T3h) in all groups. After being absorbed in the rumen and gut, the solutions dilute the solid components of the blood and proteins, explaining the results of this study, as described by ^{RIBEIRO FILHO et al. (2011}) and ^{RIBEIRO FILHO et al. (2013)}. These findings showed that enteral fluid therapy as continuous flow is an effective alternative to intravenous fluid therapy for adult cattle. All other parameters of the erythrogram remained unchanged and within the reference intervals for this species.

There were no changes in the leucogram profile throughout the experimental period or between treatments (Table 3). This result clearly shows that intubation with a naso-ruminal tube for 12 hours, and all experimental management, had no significant excitatory or stressful effects on the animals, which would result in relative polycythemia following the release of adrenaline (^{THRALL et al., 2012}).

CONCLUSION:

Enteral fluid therapy given as a continuous flow via the naso-ruminal route is well-tolerated by animals with minimal effects on welfare, even when administered for 12 hours, and is indicated as an alternative route for parenteral maintenance fluid therapy. The three electrolyte solutions proposed here are able to significantly expand blood volume.