title: Comparison of three 15N methods to correct for microbial contamination when assessing in situ protein degradability of fresh forages.
abstract: The use of stable (15)N as a marker to determine microbial contamination in nylon bag incubation residues to estimate protein degradability was investigated. Three methods using (15)N were compared: (15)N-labeled forage (dilution method, LF), (15)N enrichment of rumen solids-associated bacteria (SAB), and (15)N enrichment of rumen liquid-associated bacteria (LAB). Herbage from forages differing in protein and fiber contents (early-cut Italian ryegrass, late-cut Italian ryegrass, and red clover) were freeze-dried and ground and then incubated in situ in the rumen of 3 steers for 3, 6, 12, 24, and 48 h using the nylon bag technique. The (15)N-labeled forages were obtained by fertilizing the plots where herbage was grown with (15)NH4 (15)NO3. Unlabeled forages (obtained from plots fertilized with NH4NO3) were incubated at the same time that ((15)NH4)2SO4 was continuously infused into the rumen of the steers, and then pellets of labeled SAB and LAB were isolated by differential centrifugation of samples of ruminal contents. The proportion of bacterial N in the incubation residues increased from 0.09 and 0.45 g bacterial N/g total N at 3 h of incubation to 0.37 and 0.85 g bacterial N/g total N at 48 h of incubation for early-cut and late-cut ryegrass, respectively. There were differences (P < 0.001) between uncorrected N degradability values and those corrected for microbial contamination with all of the methods. Apparent N degradability of the low-N, high-fiber forage (late-cut ryegrass) was 0.51, whereas the corrected values were 0.85, 0.84, and 0.77 for the LF, SAB, and LAB methods, respectively. With early-cut ryegrass and red clover, the differences between uncorrected and corrected values ranged between 6% and 13%, with small differences among the labeling methods. Generally, methods using labeled forage or labeled SAB and LAB provided similar corrected degradability values. The accuracy in estimating the extent of degradation of protein in the rumen from in situ disappearance curves is improved when values are corrected for microbial contamination of the bag residue.
authors.affiliation: Département Sciences Agronomiques, Animal Science Unit, Gembloux Agro Bio-Tech, Université de Liège, Passage Deportes 2, B-5030 Gembloux, Belgium Ecole Nationale de Médecine Vétérinaire, Département des Productions Animales, Sidi Thabet, Tunis, Tunisia.
authors.forename: M
authors.initials: M
authors.lastname: Kamoun
authors.affiliation: Ecole Supérieure d'Agriculture de Mograne, 1121 Mograne-Zaghouan, Tunisia.
authors.forename: H
authors.initials: H
authors.lastname: Ammar
authors.affiliation: Département Sciences Agronomiques, Animal Science Unit, Gembloux Agro Bio-Tech, Université de Liège, Passage Deportes 2, B-5030 Gembloux, Belgium.
authors.forename: A
authors.initials: A
authors.lastname: Théwis
authors.affiliation: Département Sciences Agronomiques, Animal Science Unit, Gembloux Agro Bio-Tech, Université de Liège, Passage Deportes 2, B-5030 Gembloux, Belgium.
authors.forename: Y
authors.initials: Y
authors.lastname: Beckers
authors.affiliation: Centre for Nutrition Modelling, Department of Animal and Poultry Science, University of Guelph, Guelp, ON N1G 2W1, Canada.
authors.forename: J
authors.initials: J
authors.lastname: France
authors.affiliation: Instituto de Ganadería de Montaña (CSIC-Universidad de León), Departamento de Produccion Animal, Universidad de León, 24071 León, Spain s.lopez@unileon.es.
authors.forename: S
authors.initials: S
authors.lastname: López
mesh_terms: D000821:Animal Feed / Q000032:analysis*; D000818:Animals; D001419:Bacteria / Q000378:metabolism; D015415:Biomarkers; D001826:Body Fluids; D004043:Dietary Fiber; D005516:Food Microbiology; D008129:Lolium / Q000737:chemistry / Q000382:microbiology*; D009584:Nitrogen / Q000378:metabolism*; D009587:Nitrogen Isotopes; D059748:Proteolysis; D012417:Rumen; D013997:Time Factors; D029921:Trifolium / Q000737:chemistry / Q000382:microbiology*
chemical_list: D015415:Biomarkers; D004043:Dietary Fiber; D009587:Nitrogen Isotopes; D009584:Nitrogen
keywords: 15N; forage; microbial contamination; nylon bag; protein degradability; rumen