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ANIMAL PRODUCTION |
* Department of Animal Science,
and
Department of Poultry Science, Texas A&M University, 2471 TAMU, College Station 77843
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Abstract |
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 Top Abstract INTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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Key Words: Ascophyllum nodosum • exercise • immune • lamb • seaweed • temperature
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INTRODUCTION |
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TopAbstractINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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,b), increase cell mediated immune function (Allen et al., 2001b; Saker et al., 2001), increase weight gain (Turner et al., 2002), increase marbling (Allen et al., 2001a), increase shelf life of meat (Montgomery et al., 2001), and to possess antibacterial characteristics (Allen et al., 2001b). Other species of seaweed have also been observed to increase feed intake, increase carcass weight, and decrease digestive tract fill (Al-Shorepy et al., 2001). All of these findings suggest that ANOD may have beneficial effects for livestock subjected to handling and transport.
Handling and transporting livestock are known to be stressful. Numerous environmental factors contribute to the stress. One response to stress is an increase in core body temperature (Ingram et al., 2002). In lambs, evaporative cooling through panting is the major heat loss mechanism. There is also a correlation between respiratory frequency and heat loss by evaporation (Hales and Brown, 1974). When environmental temperatures exceed 25°C, lambs increase evaporative heat loss via increased respiration and sweating (Degen and Shkolnik, 1978). In the goat, progressive dehydration led to suppressed sweating and increased panting (Baker, 1989). A panting animal does not lose salt, and therefore electrolyte concentrations increase as water is lost from the body. Increased environmental temperatures also decrease red blood cell count, white blood cell count, and hematocrit in sheep (da Silva et al., 1992) and increase cortisol concentrations (Lowe et al., 2002).
The objective of this study was to determine if ANOD could be useful in moderating the detrimental effect of exercise and transport on lambs and to estimate the amount of ANOD needed to alleviate these detrimental effects.
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MATERIALS AND METHODS |
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TopAbstractINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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General
Forty-four Rambouillet-Suffolk crossbred lambs averaging 26.4 kg were used in this study. Lambs were randomly assigned to receive A. nodosum as a ground meal (Tasco 14, Acardian Agritech, Dartmouth, Nova Scotia, Canada) at the rate of 0 (control), 0.5, 1, and 2% of DMI/d, as estimated by using the NRC (1985) recommendations. Treatments were equivalent to 0, 0.25, 0.5, and 1 g of ANOD·kg of BW–1·d–1. Feeding more than 2% ANOD was reported to decrease feed intake in cattle (V. Allen, Texas Tech Univ., Lubbock, personal communication).
Lambs in this study were kept on pasture and fed a 16% CP (DM basis) pelleted ration on an ad libitum basis. Twice daily, beginning on d 0 (first day of ANOD) and continuing until d 14, lambs were brought from their pasture to an adjoining set of working pens. They then were given 3 gelatin capsules (size 14, Torpac, Fairfield, NJ) containing the appropriate amount of ANOD or their standard pelleted ration using a balling gun. After receiving the capsules, the lambs were released back into the pasture.
Lambs were weighed at the beginning of the 2-wk ANOD supplementation and at the end of supplementation on d 14 to determine their growth rate. An initial vaccination against ovalbumin was given to each lamb 6 wk before the onset of supplementation (Krawczel et al., 2007). On d 11, all lambs were injected i.m. in each of their hind leg muscles with 0.5 mg of ovalbumin suspended in 0.25 mL of saline to test antibody production. Blood samples (10 mL) were taken via jugular venipuncture on d 11, 15, and 18.
An indirect ELISA modified from Ameiss et al. (2004) was performed on all plasma samples using 96-well plates coated with 5 µg of ovalbumin per well. All plates were read for absorbance at 450 nm. Anti-sheep IgG or donkey anti-sheep IgM (Bethyl Laboratories, Montgomery, TX) were used as the secondary antibodies in this study. Plasma samples were initially diluted to a concentration of 1 µL of plasma in 319 µL of PBS for IgG and 2 µL of plasma in 318 µL of PBS for IgM. Five serial dilutions of each diluted sample and positive control (8 µL in 1.272 mL) were made to generate a response curve. Positive and negative control samples were also run on each plate. The initial dilutions were based on the IgG and IgM titers of the positive controls, which were obtained from the greatest responder of 3 ewes vaccinated with ovalbumin every 3 wk, for 4 mo. The negative control was plasma from an unvaccinated sheep at a dilution of 1 µL in 10.24 mL for IgG and IgM. Using the response curve, variation between IgG plates was corrected by dividing the response at 1:2,550 µL by the response of the positive control at 1:10,240 µL. Variations between IgM plates were corrected by dividing the response at 1:1,272 µL by the response of the positive control at 1:2,560 µL. All dilutions were duplicated, and the mean of the corrected duplicates was used for analysis of treatment effects.
Walking Study
All lambs had temperature data recorders (Thermochrons, Maxim/Dallas Semiconductor Corp., Sunnyvale, CA) placed in one of their ear canals to measure ear canal temperature on d 14, 1 h before the walking study. Each data logger was placed into the toe of an infant-sized cotton sock. The sock was then filled with cotton and placed in the ear canal, with the toe as far inside the ear canal as possible. The ear was then taped shut to seal it and to ensure the sock did not fall out during the studies. Loggers were set to record the temperature every 5 min. At the end of the study, all lambs had their rectal temperature taken using a digital thermometer to determine the difference between the ear canal and rectal temperature. Tympanic temperature is generally 2°C lower than rectal temperature (Goodwin, 1998).
On d 14, at 1300, all lambs were relocated to an enclosed pen (9 x 9 m) formed from a section of their home pasture. The lambs were walked in a circular motion for 30 min around a 3 x 3 m rectangular structure within the pen. The lambs were walked at a brisk pace until their respiration rate increased and they began panting. The pace was then slowed and maintained for the remainder of the 30 min. Increased respiration rates occur during hyperthermia (Hales and Brown, 1974; Entin et al., 1998, 1999; Friend et al., 1998). The lambs were then allowed to rest for 30 min to allow their respiration rates to return to normal, after which another 30-min walk was repeated.
Ten milliliters of blood was collected from all lambs via jugular venipuncture (BD brand plasma separation tubes with lithium heparin and polymer separator gel, VWR, West Chester, PA) immediately before the first walk and immediately after the second walk. Plasma samples were analyzed in duplicate for cortisol concentration using commercially available RIA kits (Diagnostic Protocols Corp., Los Angeles, CA). Any duplicates that differed by more than 12% were reassayed. The intra- and interassay CV for the cortisol assay were less than 9%.
Transport Study
Lambs were transported on d 15 over local roads near College Station, TX, for 12 h (730 to 1930) during September in a goose-neck trailer. Treatments were distributed into 3 groups of 15 lambs among three 2.1 x 2.3-m compartments within the trailer. A lamb that was not part of the study but was in the same flock was used so each compartment contained 15 lambs. Temperature-humidity data recorders (HOBO-H8, Onset Computers, Pocasset, MA) were mounted within the trailer at 1 m from the front and rear of the trailer and at a height 0.5 m above the deck. The lambs were weighed immediately before transport and before being offered feed and water after transport.
The thermal heat index was used to assess periods of heat stress during transport. This index is a combination of the temperature and the humidity, and is calculated using the following formula: thermal heat index = (dry-bulb temperature, °C) + (0.36 x dew point temperature, °C) + 41.2. In cattle, a thermal heat index above 72 indicates mild heat stress, above 80 indicates medium heat stress, and above 90 indicates severe heat stress (Pennington and Van Devender, 2004).
Temperature recorders were allowed to remain in the ears of the lambs after the walking study so they could be used to monitor ear canal temperature during transport the next day. Minimum, maximum, mean, and range of the ear canal temperature were calculated for each treatment. Each individual’s minimum temperature was subtracted from their maximal temperature to obtain an ear canal temperature range during transport. Ten milliliters of blood was collected from all lambs via jugular venipuncture at 4-h intervals throughout transport and 3 d posttransport. The only restraint needed for blood sampling consisted of 1 person briefly holding the lamb and raising its head so the neck was exposed. The 0 h and d 18 samples were obtained in the holding pens, whereas the 4, 8, and 12 h samples were collected within the trailer.
Plasma was frozen until analyzed for cortisol, and the 0 and 12 h samples were also analyzed for aldosterone using commercially available RIA kits (Diagnostic Protocols Corp.). Any duplicates that differed by more than 12% were reassayed. The intra- and interassay CV for the cortisol and aldosterone assays were less than 9%. To determine general health and hydration of the lambs, a portion of the plasma samples taken at 0 and 12 h of transport were used for determination of electrolyte concentrations (sodium, chloride, and potassium) and plasma chemistry profiles by the Texas Veterinary and Medical Diagnostic Laboratory (College Station, TX). The plasma chemistry profile consisted of: albumin:globulin ratio, aspartate amino-transferase, albumin, blood urea nitrogen, calcium, creatinine, gamma glutamyltransferase, globulins, glucose, magnesium, phosphorus, total plasma protein, and total bilirubin. Plasma chemistry profiles are an accepted method of documenting the overall health of an animal (Kumar et al., 2003). Components of a chemistry panel can indicate problems such as organ dysfunction, dehydration, metabolic state, and muscle damage.
After transport and weighing, the lambs were simultaneously released into a 9 x 9-m pen made from part of their home pasture and containing feed and water troughs. Feed was offered in the same round feeder from which the lambs fed daily, and 2 additional 4-m troughs were filled with feed (16% CP, pelleted ration) to allow all lambs room to eat. The water trough was 1 m long with a float valve for automatic refilling. The latency to eat, drink, and lay down was recorded by observers for all lambs.
Statistical Analysis
Mean antibody response, plasma chemistry, electrolytes, aldosterone, and cortisol were analyzed using a split-plot, repeated measures design. The GLM procedure (SAS Inst. Inc., Cary, NC) was used, with treatment, time, treatment x time interaction, and individual lambs nested within treatment as the factors. Lamb nested within treatment was the error term used to test the treatment effects. Average daily gain, ear canal temperature (minimum, maximum, range, and mean during hot periods), BW loss, and behavioral data were analyzed using PROC GLM of SAS. Posthoc contrasts were performed to determine any linear or quadratic dose effects.
Because comparisons of individual dosages were required to identify the optimum dosage (to determine whether 2 dosages caused similar or different responses), the 4 dosages were evaluated using the GLM procedure, and the means were evaluated using the LSD procedure (Bartle et al., 1992). Differences for pairwise comparisons were considered significant at P < 0.05, unless otherwise stated.
Lambs were released as 1 group containing all treatments during behavioral data collection; however, the subjects were considered to be independent because lambs do not have an established social hierarchy and the feed and water were in close proximity, limiting any reluctance of separating from the flock to eat, drink, or lay down.
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RESULTS |
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TopAbstractINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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). On d 15, and to a lesser extent on d 18 (4 and 7 d after the vaccination), there was a linear dose-dependent suppression (P < 0.002) of the antibody response. Only the control group exhibited the expected immune response (Abbas et al., 2000). A similar linear dose-dependent effect also occurred in the IgM response on d 11, 15, and 18 (P < 0.02; Figure 2).
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, ear canal temperature ranged from 2 to 3°C below rectal temperature. The average ear canal temperature at the end of the walking study was 38.4°C, whereas the average rectal temperature was 41.2°C.
Although no treatment x time interaction (P = 0.40) was observed, a treatment (P = 0.02) effect on cortisol concentration was observed before and after forced walking. Before walking, the control group had (P = 0.05) greater cortisol concentrations than the 2% ANOD group (Table 1), with the 0.5 (P = 0.14) and 1% (P = 0.10) ANOD groups being intermediate following a dose-dependent linear trend (P = 0.03). The same linear trend (P = 0.04) occurred postwalk.
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During transport, no interaction of time x treatment (P = 0.70) was observed for cortisol; however, the control group had greater (P = 0.01) cortisol concentrations than all other groups in a linear dose-dependent manner (P = 0.03). Differences during transport were only observed at 4 and 8 h of transport when the control group had greater concentrations of cortisol (0.91 and 1.24 µg/dL) than the 2% ANOD group (0.61 and 0.90 µg/dL, P = 0.05, 0.10, respectively). Before transport a quadratic response (P = 0.03) was observed, and 4 h (P = 0.01) into transport there was a linear trend for cortisol concentration with the control group having the greatest concentrations.
An interaction of treatment x time was observed for concentrations of aldosterone (P = 0.01). Greater levels of supplementation with ANOD lowered pretransport concentrations of aldosterone in a linear pattern (P = 0.03, Table 2). After transport, all lambs had similar concentrations (P > 0.65) of aldosterone compared with controls, except the 1% ANOD group, which was greater (P = 0.05) in aldosterone concentrations compared with all other groups, resulting in a quadratic response (P = 0.03).
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). A linear trend (P = 0.01) was observed in plasma protein, albumin, phosphorus, glucose, blood urea nitrogen, magnesium, and calcium. As rate of ANOD increased above the 0.5% rate, differences among treatments were observed (P < 0.05).
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). All lambs increased from pretransport glucose concentrations during transport except the 2% ANOD group. The 1 and 2% ANOD groups also decreased from pretransport blood urea nitrogen concentrations as a result of transport. Globulins, creatinine, total biliubin, creatine kinase, the albumin:globulin ratio, and gamma glutamyltransferase were not influenced by treatments and therefore are not shown in Table 3.
An interaction of treatment x time was detected for sodium (P = 0.06), potassium (P = 0.05), sodium:potassium ratio (P = 0.07), and chloride concentrations (P = 0.06). Supplementation with ANOD caused no differences (P > 0.10) in the pretransport concentrations of sodium, potassium, or chloride. However, the sodium:potassium ratio in blood taken pretransport was lower in lambs fed 1 (P = 0.03) and 2% (P = 0.06) ANOD than the control group. Posttransport, the 1 (P < 0.05) and 2% (P < 0.05) ANOD groups had lower sodium and chloride concentrations than the controls (Table 4). A linear response (P < 0.01) was observed in sodium, chloride, and potassium concentrations after transport.
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No effect of treatment was observed on the amount of BW lost in response to transport (P = 0.17). No differences were noted in latency to eat, drink, or lie down posttransport (P > 0.50). All lambs went to feed immediately after being released after transport, and time to lie down posttransport was approximately 1.5 h for all lambs. Latency to drink demonstrated a very slight linear (P = 0.18) response with the control, 0.5, 1, and 2% ANOD groups drinking 823, 747, 595, and 589 s after transport.
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DISCUSSION |
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TopAbstractINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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,b; Saker et al., 2001) did not have precise control over the amount of ANOD consumed. An ANOD extract was sprayed on the grass the animals were grazing, mixed in the feed (Allen et al., 2001a,b), or fed as a mineral supplement (V. Allen, personal communication). In those studies, it was assumed that the animals ate ANOD at the 2% rate. In the current study lambs received ANOD in a bolus. This enabled a more accurate investigation of effects of different supplementation rates than previous studies. This study confirmed that the 2% rate of supplementation suggested in other studies (Allen et al., 2001a,b) was the appropriate rate to feed lambs to lower ear canal temperature during heat stress by 0.3°C.
Supplementation did not affect growth during the 2-wk feeding period, which is contrary to swine fed ANOD for 2 wk (Turner et al., 2002). Feeding ANOD suppressed antibody production in response to the booster vaccination in a dose-dependent manner. It is important to note that this decreased antibody production was not only seen in the postboost response for IgG and IgM, but IgM was already lower in the 2% ANOD group before the booster. This was indicative of decreased IgM production before the booster. Decreased IgM production implies that it may not be productive to vaccinate animals that are being supplemented with ANOD. Vaccinating prior to supplementation with ANOD may be advisable. The actual impact on health and the dynamics of the antibody suppression observed in this study merit further study.
Although it has been reported that ANOD decreased basal body temperature during hot summer weather (Allen et al., 2001b), that effect was not observed in this study in response to forced walking. This was not unexpected as the amount of heat produced internally during exercise can cause marked increases in body temperature, especially during hot weather. These increases are often difficult for the body to dissipate through thermoregulation. The ability of ANOD to moderate ear canal temperature during hot temperatures was observed in this study during transport and is consistent with previous research in cattle (Allen et al., 2001a,b).
Cortisol concentrations in the 1 and 2% ANOD groups were lower compared with the control group prior to the walking and transport studies. It was not clear whether decreases in cortisol were due to decreased perception of stress or to a decrease in the function of the hypothalamic-pituitary adrenal axis. The adrenal gland releases cortisol in response to stress and also secretes aldosterone to maintain electrolyte balance. Lowered cortisol is indicative of lower stress, but additional measurements are needed to confirm that stress was actually reduced. The initial difference in cortisol concentration between the 2% ANOD and the control group, with the control group having greater concentrations, was maintained throughout walking and transport. The decrease in cortisol concentration in response to walking was likely due to depletion of adrenal reserves and not lowered stress because the lambs were walked to the point of showing distress as indicated by panting, increased body temperature, and decreased vigor in walking. Further research is needed to determine if supplementation with ANOD decreases adrenal function to the detriment or benefit of the animal.
Supplementation of ANOD at the 1 and 2% rate also appeared to affect the adrenal gland’s control of water balance. Before transport began, the 1 and 2% ANOD groups had lower concentrations of aldosterone, indicating that they were excreting sodium and retaining potassium (Dickson, 1993). Low aldosterone concentration before transport may have allowed the 2% ANOD group to maintain pretransport plasma electrolyte concentrations; their electrolyte concentrations did not change significantly as a result of transport. The 2% ANOD group could have already been primed for excreting excess sodium, possibly counteracting the increase usually seen in electrolyte concentrations due to hemoconcentration from dehydration. This hypothesis was supported by a trend for decreases in sodium, calcium, phosphorus, and magnesium concentrations in the 1 and 2% ANOD groups, whereas these plasma constituents increased in the control lambs after transportation. Aldosterone concentrations after transport were similar among all treatments, possibly indicating that the control group was compensating for their altered electrolyte concentrations by lowering aldosterone concentrations.
1 Corresponding author: t-friend{at}tamu.edu
Received for publication August 20, 2005. Accepted for publication August 17, 2006.
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