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© 2009 Plant Management Network. Composition of Horse Diets on Cool-Season Grass Pastures using Microhistological Analysis Jesse I. Morrison, Graduate Research Assistant, and S. Ray Smith, Associate Professor, Department of Plant and Soil Sciences, University of Kentucky, Lexington, KY 40546; Glen E. Aiken, USDA-ARS Forage-Animal Production Research Unit (FAPRU), Lexington, KY 40546; and Laurie M. Lawrence, Professor, Department Animal and Food Sciences, Lexington, KY 40546 Corresponding author: S. Ray Smith. raysmith1@uky.edu Morrison, J. I., Smith, S. R., Aiken, G. E., and Lawrence, L. M. 2009. Composition of horse diets on cool-season grass pastures using microhistological analysis. Online. Forage and Grazinglands doi:10.1094/FG-2009-0325-01-RS. Abstract Grazing patterns and diet composition can be difficult to determine with horses, but are important when pastures contain species that have the potential to cause animal toxicity. The objective of this study was to determine the composition of domesticated horse diets when grazing mixed cool-season pastures using microhistology of fecal samples. Samples of tall fescue, Kentucky bluegrass, and orchardgrass were evaluated for microscopically unique characteristics from plant fragments in fecal material. Grazing studies were conducted in cool-season grass pastures in November 2006 and April 2007 in Lexington, KY. Eight mature Thoroughbred mares were placed in individual paddocks of varying botanical compositions and grazed for six days. Fecal samples from manure piles were taken from each paddock to determine diet composition. There was a high correlation between tall fescue and orchardgrass in the pasture and in the diet (0.90 and 0.80, respectively), but no correlation with bluegrass. For each percent increase of tall fescue or orchardgrass in the pasture, there was a corresponding increase of 0.44% and 0.42%, respectively, in the diet. In conclusion, microhistological analysis of plant fecal fragments showed that horses consume tall fescue and orchardgrass in similar proportions to those found in their pastures. Introduction Tall fescue toxicity in late-term broodmares can be detrimental to both the mare and foal, and is a major concern for the horse industry as a whole (7,12,13,14). In the United States, an estimated 700,000 horses are managed on pastures containing varying amounts of tall fescue (2). Since tall fescue is endemic and thrives in the transition zone climate, it is difficult and costly to completely eliminate from existing pastures (16). Techniques that help horse farm owners and managers know what their animals are consuming when grazing mixed-species pastures could be useful for making management decisions for pastures containing tall fescue. Several techniques are available to evaluate diet composition of grazing animals, but the standard method is to analyze ingested plant material from esophageal fistulas (9). This invasive technique is not acceptable for equine research, but microhistological analysis of plant fecal fragments provides a useful non-invasive technique. There has been extensive research to validate this technique (3,4,5,6,15) for use with a variety of animal species in varying climates and ecosystems. This technique identifies plant tissues microscopically and performs frequency counts of fragments for each species contained in samples of feces (6). Microhistological analysis of plant fecal fragments is a valid predictor of diet composition using the assumptions developed by Sparks and Malechek (15): (i) the fragments of plants are randomly distributed on microscope slides; (ii) fragments from different species are the same average size; and (iii) dry weight bulk densities are the same for the different species. This technique was validated by Morrison et al. (10) for horses consuming cool-season grass species (tall fescue, orchardgrass, and Kentucky bluegrass). The objective of this study was to determine the composition of domesticated horse diets when grazing mixed cool-season pastures using microhistology of fecal samples. Determining Diet Composition of Horses Grazing Mixed Cool-season Pastures This study was conducted from 3 to 10 November 2006 and 23 April to 1 May 2007, at the University of Kentucky’s Maine Chance Farm. These time periods were chosen because they correspond to ideal growing conditions for the cool-season grass species of interest. Tall fescue, orchardgrass, and Kentucky bluegrass were all in a vegetative growth stage (15 to 25 cm) during this experiment, with the exception of some bluegrass seedheads in late April 2007. All methods of pasture sampling, manure collection, slide preparation, and analysis were identical across dates. A 4.0-ha pasture, with predominantly Maury silt loam soil, was pre-selected for use and managed similar to Thoroughbred horse farm pastures in the region (Table 1). Table 1. Management procedures during this study, at the University of Kentucky’s Maine Chance Farm, Lexington, KY.
Within the pasture, eight 0.25-ha paddock subdivisions were assigned using temporary electric fencing. Paddock orientation was chosen to create a range of tall fescue percentages across paddocks and to insure equal availability of herbage mass between paddocks. The study was repeated in April 2007 to provide data during another growing season. Each paddock was evaluated for botanical composition 5 November and 26 April with hand-separated herbage harvested within a 0.6/m² quadrat, made of 3.8-cm (OD) PVC. The PVC qradrat was randomly thrown 10 times within each paddock and individual sample areas were photographed and harvested to a 5-cm stubble within the quadrat area. Eight mature open Thoroughbred mares were turned out into the 4-ha pasture on 3 November 2006 and 23 April 2007, but excluded from the eight 0.25-ha paddocks. After a 2- to 3-day adjustment period, each mare was pastured in an individual paddock, where they were kept for 6 days (5 to 10 November and 26 April to 1 May). On 3 November 2006, tall fescue tiller samples were taken from each paddock to test for endophyte infection rate. Endophyte infection measurements were not repeated in April 2007 since endophyte infection rate would not be expected to change over winter. Fresh plant samples collected for pasture botanical composition were dried in a forced-air oven at approximately 70°C. They were then ground through a 2-mm screen in a Thomas Wiley mill, and then reground through a 1-mm screen in a Foss Cyclotec 1093 Sample Mill. Each sample was analyzed using Near-Infrared Spectrometry (NIRS) with a Foss 6500 autosampler and ISIscan software for crude protein, acid detergent fiber, neutral detergent fiber, and relative feed value. Fecal samples were taken by collecting and bulking the freshest manure piles in each paddock (~2 kg per paddock) on each of days 4 to 6 (8 to 10 November and 29 April to 1 May) to allow sufficient time for passage. Only fecal material collected on day 5 was used for this research. These samples were stored separately by paddock and dried in a forced-air oven at 55°C for 48 h. Dried fecal samples were ground through a 1-mm screen with a Wiley mill for subsequent analysis. Pigments were cleared from sample material by mixing a 1-g subsample in 20 ml of household liquid bleach (6% NaClO) in a 100-ml specimen cup for 6 min. Cups were shaken occasionally to blend contents, and gas buildup was allowed to vent after each blending. Samples were then rinsed with water over an 80-mesh screen, and allowed to thoroughly air dry. Six microscope slides were prepared per sample (paddock) and analyzed as described by Sparks and Malechek (15) using Johnson’s (8) plastic mounting medium. The identifiable cellular structures used for identification included: stomata and their density patterns, size of bulliform cells, shape and frequency of prickle hairs, cell wall construction, shape and thickness, and cuticle size (1,11,15). The slides were evaluated for percent species composition, which was then compared to pasture species composition measurements. Determining Effect of Tall Fescue Pasture Composition on Diet Composition Since paddock was the experimental unit in this study correlation and regression analysis procedures were used to determine the relationship between the percentage by weight of tall fescue and other available grasses in the pasture and the corresponding ratio that was actually ingested by the animals. Standard errors for paddock botanical composition between fall and spring sampling dates were determined using PROC GLM in SAS 9.1 (SAS Institute Inc., Cary, NC). Paddock Botanical Composition Paddocks were selected within the overall pasture to reflect the wide range in tall fescue percentage and to evaluate how diet composition was associated with pasture composition. The percent botanical composition of tall fescue in paddocks on 5 November 2006 ranged from 16% to 71%, and on 26 April 2007 from 13% to 75%. Percent composition of bluegrass (Fig. 2), orchardgrass (Fig. 3), and weeds varied greatly. Clover populations in paddocks were low, likely due to the chemical application regimen used for broadleaf weed control. Percentages of bare soil also were low.
Effect of Nutritive Value on Diet Composition Forage quality analysis (Table 2) showed that the three desirable grasses were of higher quality in the spring. Each species contained higher crude protein (CP), lower acid detergent fiber (ADF) and neutral detergent fiber (NDF), and higher overall relative feed values (RFV). Forage quality was likely not a factor in diet composition because of the small amount of difference between species quality values. Further studies should involve more thorough quality analysis as well as analysis of forage carbohydrate concentration since these plant characteristics may affect horse selectivity. Table 2. Forage quality values from grab samples taken from study pasture before the initiation of grazing by horses (3 November 2006 and 23 April 2007), Maine Chance Farm, Lexington, KY.
Effect of Endophyte Infection on Diet Composition Paddocks 1, 2, 4, and 5 occupied large areas of previously seeded Jesup endophyte-free and Jesup Max-Q tall fescue cultivars, while other paddocks contained tall fescue infected with the wild type fescue endophyte. Paddocks 1, 2, 4, and 5 were all less than 5% infected with the wild-type endophyte, while wild-type endophyte infection rates for other paddocks were: Paddock 3, 8%; Paddock 6, 100%; Paddock 7, 83%; and Paddock 8, 41%. There was no apparent negative effect of wild-type endophyte infection level on consumption of tall fescue. In November horses in paddock 6 (18% tall fescue, 100% infected) and paddock 7 (16% tall fescue, 83% infected) consumed 23 and 22% of their diet in tall fescue while horses in paddock 3 (23% tall fescue, 8% infected) consumed 20% tall fescue. Since these data are very limited further research should be conducted to conclusively determine the effect of endophyte infection on tall fescue consumption by horses. Pasture Botanical Composition and Diet Composition Relationship There was strong correlation (r = 0.90; P < 0.0001) between botanical composition in the paddock and fecal composition of the tall fescue component. There also was a strong correlation between orchardgrass measurements (r = 0.80; P < 0.0002). Bluegrass measurements were not correlated (r = 0.52), most likely because of a narrow range in botanical composition of this species within paddocks. Bluegrass botanical composition in paddocks ranged from 5 to 35% (fall samples) and 5 to 45% (spring samples), while fecal compositions ranged from 22 to 35% and 15 to 30%, respectively. There was also no significant relationship between paddock botanical composition and fecal composition of "other" component. Figure 4 shows a scatter plot of tall fescue fecal composition against tall fescue botanical composition by paddock across both seasons. There was a strong relationship (r² = 0.81) between the percentage of tall fescue in the paddock and the corresponding percentage in the diet. With each percent increase in tall fescue composition by weight in the paddock, there was a corresponding 0.43% increase in diet composition. A quadratic analysis model was designed to test the tall fescue botanical composition and diet composition for curvilinear tendencies; however, the quadratic term was not significant.
These results suggest that horses do not avoid or select against tall fescue. At low botanical composition percentages, horses in this experiment consumed a slightly higher percentage of their diet in tall fescue than what was available; while at higher percentages (> 35%) they selected slightly lower amounts. Therefore, horses did not necessarily select against tall fescue when grazing these mixed species pastures. The practical application of this research is that decisions concerned the potential for fescue toxicity for pregnant mares can be made based on the percentage of tall fescue present in a specific pasture. It is noteworthy that this study was conducted during late fall and early spring when vegetative growth was more succulent and palatable than summer growth. Though not measured here, ergovaline levels in endophyte-infected tall fescue on area farms were lower than normal during fall 2006 and spring 2007, and much lower than would be expected during the summer (data not shown). Therefore, more succulent forage and lower levels of ergovaline may have contributed to the higher rates of tall fescue consumption. In other words, horses may show a different preference for tall fescue when the plant material is more mature. Figure 5 shows a scatter plot of orchardgrass fecal composition against orchardgrass botanical composition by paddock across both seasons. There was a strong linear relationship (r² = 0.57) between the percentage of orchardgrass in the paddock by weight and the corresponding percentage in the diet. With each percent increase in orchardgrass composition in the paddock, there was a corresponding 0.42% increase in diet composition.
These results indicate that, similar to tall fescue, horses did not avoid or select against orchardgrass. At low botanical composition percentages, horses consumed a higher percentage of their diet in orchardgrass than was available; while at higher percentages (> 30%) they selected slightly lower amounts in relation to what was available. Levels of orchardgrass in the diet may have reached a plateau at approximately 35%. These data are of great importance to horse owners concerned about what pasture grasses are best suited for their animals. It is also useful to producers trying to minimize consumption of tall fescue, because orchardgrass can be used as an acceptable replacement forage grass for endophyte-infected tall fescue. In conclusion, this research shows that diet composition of tall fescue and orchardgrass are closely related to botanical composition in the pasture. Acknowledgment This research was funded in part by a Specific Cooperative Agreement (58-6440-7-135) with the USDA-ARS, Forage Animal Production Research Unit, Lexington, KY. Literature Cited 1. Aiken, G. E. 1989. Plant and animal responses to a complex grass-legume mixture under different grazing intensities. PhD diss. Univ. of Florida. Gainesville, FL. 2. Blodgett, D. J. 2001. Fescue toxicosis. Vet. Clin. N. Am.-Equine. 17:567-577. 3. Fracker, S. B., and Brischle, H. A. 1944. Measuring the local distribution of ribes. J. Ecol. 25:283-304. 4. Havstad, K. M., and Donart, G. B. 1978. The microhistological technique: testing two central assumptions in South-central New Mexico. J. Range Manage. 31:469-470. 5. Holechek, J. L., and Gross, B. D. 1982. Evaluation of different calculation procedures for microhistological analysis. J. Range Manage. 35:721-723. 6. Holechek, J. L., Gross, B. D., Dabo, S. M., and Stephenson, T. 1982. Effects of sample preparation, growth stage, and observer on microhistological analysis of herbivore diets. J. Wildl. Manage. 46:502-505. 7. Hoveland, C. S. 1993. Importance and economic significance of the Acremonium endophytes to performance of animals and grass plants. Agric. Ecosys. Enviro. 44:3-12. 8. Johnson, M. K., Wofford, H., and Pearson, H. A. 1983. Microhistological techniques for food habits analyses. Res. Paper SO-199, December. USDA/USFS, New Orleans, LA. 9. Laycock, W. A., Buchanan, H., and Kraegen, W. C. 1972. Three methods for determining diet, utilization and trampling damage on sheep ranges. J. Range. Manage. 25:352-356. 10. Morrison, J. I., Smith, S. R., and Aiken, G. E. 2007. Manure and a microscope: The keys to keeping Kentucky's Thoroughbreds kicking. Proc. of the Amer. Forage and Grassland Counc. Ann. Meeting, 23-26 June 2007. State College, PA. Amer. Forage and Grassland Counc., Chicago, IL. 11. Paolo, V. S., Cid, M. S., Brizuela, M. A., and Ferri, C. M. 2005. Microhistological estimation of grass leaf blade percentages in pastures and diets. Rangeland Ecol. Manage. 58:207-214. 12. Poppenga, R. 2003. An overview of fetotoxic agents and their possible role as agents in mare reproductive loss syndrome. Proc. 1st Workshop on Mare Reproductive Loss Syndrome, Lexington, KY. 2002. KY Ag. Exp. Stn., Lexington, KY. 4:44-47. 13. Schlafer, D. H. 2003. Placental toxicology: Recognized placental toxicants in veterinary medicine and consideration of a likely role of a placental toxicant in mare reproductive loss syndrome. Proc. 1st Workshop on Mare Reproductive Loss Syndrome, Lexington, KY. 2002. KY Ag. Exp. Stn., Lexington, KY. 4:48-50. 14. Schultz, C. L., and Bush, L. P. 2003. The potential role of ergot alkaloids in mare reproductive loss syndrome. Proc. 1st Workshop on Mare Reproductive Loss Syndrome, Lexington, KY. 2002. KY Ag. Exp. Stn., Lexington, KY. 5:60-63. 15. Sparks, D. R., and Malechek, J. C. 1968. Estimating percentage dry weight in diets using a microscopic technique. J. Range Manage. 21:264-265. 16. Zhuang, J., Marchant, M. A., Schardl, C. L., and Butler, C. M. 2005. Economic analysis of replacing endophyte-infected tall fescue pastures. Agron. J. 97:711-716. |
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