Browsing by Person "Rodehutscord, Markus"

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The active core microbiota of two high-yielding laying hen breeds fed with different levels of calcium and phosphorus
(2022) Roth, Christoph; Sims, Tanja; Rodehutscord, Markus; Seifert, Jana; Camarinha-Silva, Amélia
The nutrient availability and supplementation of dietary phosphorus (P) and calcium (Ca) in avian feed, especially in laying hens, plays a vital role in phytase degradation and mineral utilization during the laying phase. The required concentration of P and Ca peaks during the laying phase, and the direct interaction between Ca and P concentration shrinks the availability of both supplements in the feed. Our goal was to characterize the active microbiota of the entire gastrointestinal tract (GIT) (crop, gizzard, duodenum, ileum, caeca), including digesta- and mucosa-associated communities of two contrasting high-yielding breeds of laying hens (Lohmann Brown Classic, LB; Lohmann LSL-Classic, LSL) under different P and Ca supplementation levels. Statistical significances were observed for breed, GIT section, Ca, and the interaction of GIT section x breed, P x Ca, Ca x breed and P x Ca x breed (p < 0.05). A core microbiota of five species was detected in more than 97% of all samples. They were represented by an uncl. Lactobacillus (average relative abundance (av. abu.) 12.1%), Lactobacillus helveticus (av. abu. 10.8%), Megamonas funiformis (av. abu. 6.8%), Ligilactobacillus salivarius (av. abu. 4.5%), and an uncl. Fusicatenibacter (av. abu. 1.1%). Our findings indicated that Ca and P supplementation levels 20% below the recommendation have a minor effect on the microbiota compared to the strong impact of the bird’s genetic background. Moreover, a core active microbiota across the GIT of two high-yielding laying hen breeds was revealed for the first time.
Amino acid digestibility and metabolisable energy of spring and winter faba beans grown on two sites and effects of dehulling in caecectomised laying hens
(2021) Siegert, Wolfgang; Ibrahim, Ahmad; Link, Wolfgang; Lux, Guido; Schmidtke, Knut; Hartung, Jens; Nautscher, Nadine; Rodehutscord, Markus
BACKGROUND: The variation in amino acid (AA) digestibility and metabolisable energy (MEN) in four spring and four winter faba bean genotypes differing in vicine/convicine (V/C) concentrations grown on two sites was investigated in caecectomised LSL-Classic laying hens. Effects of dehulling one faba bean genotype were also examined. Diets containing one out of 17 faba bean variants each and a basal diet were fed to ten caecectomised laying hens in a row-column design to achieve five replicates per diet. RESULTS: Ranges and levels of digestibility of the hulled variants differed widely among AA with the lowest and highest range determined for Arg (90–93%) and Cys (−12–65%), respectively. MEN ranged between 10.3 and 12.3 MJ kg−1 dry matter. Lower MEN and digestibility of Cys, Glx, Phe, Pro, Tyr, and Val (P < 0.050) was determined for the winter genotypes grown in Nimtitz compared to the other variants. Digestibility of Ser was lower for the spring than for the winter genotypes (P < 0.050). Negative correlations with AA digestibility were determined for phytate, but not for tannin and V/C concentrations (P < 0.050). Negative correlations between tannin fractions and MEN were weak (P = 0.082–0.099). Dehulling increased MEN by 1.8 MJ kg−1 dry matter and raised the digestibility of Pro, His, and Glx (P < 0.050). CONCLUSIONS: The results indicated that the digestible AA and MEN supply of laying hens was increased by using low phytate faba beans while breeding for low V/C genotypes did not affect AA digestibility or MEN. Dehulling increased MEN and the digestibility of some AA.
Approaches to increase digestibility of Bangladesh ruminant feed resources in order to mitigate enteric methane production
(2024) Bashar, Muhammad Khairul; Rodehutscord, Markus
Multiple analyses have shown that the rising human population, urbanization, and consumer preferences affect the demand for livestock products in developing countries. Concurrently, human population and urbanisation growth are reducing the likelihood of newly cultivated land producing feeds or restoring damaged pastures. Using low-quality roughage more efficiently is an option, but such roughages have drawbacks, including high structural carbohydrate and low nitrogen contents, which lead to poor palatability and nutrient utilisation including emission of methane (CH4) that is harmful to the environment and a loss of energy to the animal. Using commonly accessible legumes and tree foliage could be a possibility to address the need for N and CH4 mitigation due to their high crude protein and secondary compound contents. In order to evaluate the potential of specific combinations of roughages, concentrates, and other feeds, it is necessary to determine the feeding values, in vitro ruminal fermentation, and CH4 production of single feeds, as well as the promising interaction (associative) effects between single feeds when combinded in a ration. This may allow for formulating a better balanced total mixed ration (TMR) for ruminants, which was the main goal of the present doctoral study. Eighteen feedstuffs from Bangladesh were chosen, including roughages such as crop residues (rice straw, urea molasses treated straw and maize stover), silages (Napier silage and Maize silage), common grasses (German grass, Para grass and Napier grass) and leguminous fodder (Ipil-ipil, Glicidia, Alfalfa hay and Moringa tops) and concentrates (crushed wheat, crushed maize, Wheat bran, Kashari bran, and Rice bran). In Study 1, the single feeds were characterized by comprehensive chemical analysis and in vitro production of total gas and methane using the Hohenheim gas test. Based on the results of Study 1, rice straw, German grass, Napier silage, and maize silage were used in Study 2 and incubated in vitro alone or after substitution by Ipil-ipil and Gliricidia at levels of 10%, 20%, 30%, and 40%, respectively. In Study 3, mixtures of roughages and leguminous fodder (rice straw + 40% Ipil-ipil, German grass + 10% Ipil-ipil, Napier silage + 30% Ipil ipil, and maize silage + 30% Ipil-ipil) were combined with single concentrates in order to identify the most promising mixtures in regard to digestibility and methane production. Eventually, 24 TMR were formulated using six for each type of roughage in Study 4. These TMR were comprised of 60% of the same combination of roughage as used in Study 3 and 40% of various combinations of concentrates including TMR-1 (60% rice bran + 10% wheat bran + 30% kashari bran), TMR-2 (30% rice bran + 10% wheat bran + 60% kashari bran), TMR-3 (10% rice bran + 60% wheat bran + 30% kashari bran), TMR-4 (10% rice bran + 30% wheat bran + 60% kashari bran), TMR-5 (60% rice bran + 30% wheat bran + 10 kashari bran), and TMR-6 (30% rice bran + 60% wheat bran + 10% kashari bran) in order to identify suitable TMR for maximising ruminal fermentation and reducing the CH4 production in vitro. In all studies, the total gas production (GP), digestibility of organic matter (dOM), metabolisable energy (ME), and net energy for lactation (NEL) were determined after incubating 200 mg of feed with a rumen fluid-buffer solution for 72 hours. In addition, the CH4 concentration in the produced gas was measured after incubating 120 mg of feed for 24 hours. Measured values of the mixed feed were compared to estimated values, where the estimated values were weighted means of the values of the individual feeds, to determine the associative effects between the feed ingredients (Study 2 and Study 4). In Study 1, the concentration of acid detergent fibre (ADF), neutral detergent fibre (NDF), and acid detergent lignin (ADL) was the lowest in leguminous fodder while the crude protein (CP) concentration was the hightest, followed by the common grasses and the silages. The crop residues showed the lowest CP and the highest cell wall fraction concentrations. The dOM, ME, and NEL of crushed wheat and crushed maize were significantly higher (p<0.05) than those of other feedstuffs. The same differences were found (p<0.05) for CH4 concentration (% of GP) and CH4 production (L CH4/kg dOM). The dOM and ME of German grass and Ipil-ipil were higher (p<0.05), whereas the CH4 concentration and CH4 production were lower compared to that of crop residues and other common grasses. The CH4 production of single feeds decreased with increasing concentrations of CP, ADF, and ADL, whereas it increased with NDF concentration. These findings enabled development of more balanced diets for ruminants with the aim of improved digestibility and reduced CH4 emission while making use of widely available feed resources in Bangladesh. Study 2 aimed to evaluate the in vitro ruminal fermentation, CH4 production, and associative effects between low-quality roughages and CP supplements. The gas production after 24 hours (GP24) and rumen fermentation rate increased (p<0.05) with increasing addition of Ipil-ipil to rice straw. It also resulted in a gradual increase of dOM and ME (p<0.05) of the mixture, reaching maximum values (59.1% and 7.60 MJ ME/kg DM) and maximum reductions in CH4 concentration and CH4/dOM (6.9% and 11.7%) compared to the control with Ipil-ipil addition at 40% of DM. When Ipil-ipil was added at 30% to Napier silage, the same trend was observed for dOM and ME (56.5%, and 7.55 MJ ME/kg DM) and CH4 concentration and CH4/dOM (5.3% and 13.3%). Ipil-ipil did not significantly interact with high quality roughages such as German grass and maize silage. Increasing the addition of Gliricidia to the roughages in Study 2 led to a decrease in GP24, dOM, ME, and CH4 production. The highest levels of the associative effects (p<0.05) were seen when 40% Ipil-ipil was added to rice straw and 30% was added to Napier silage. In conclusion, adding Ipil-ipil to low-quality roughages is superior to Gliricidia and showed promising results, with the ranking as follows: rice straw + 40% Ipil-ipil > Napier silage + 30% Ipil-ipil > maize silage + 30% Ipil-ipil > German gras + 10% Ipil-ipil. Study 3 and Study 4 had the overarching purpose to determine which combination of single concentrates and combinations of concentrates with roughage produced the best mixed feed in terms of rumen fermentation, CH4 reduction potential, and associative effect. The CP and non-stach carbohydrate contents of the Ipil-ipil-based mixed feed was enhanced by adding Wheat bran and Kashari bran as single concentrates, which had a favourable effect on rumen fermentation and gas production but had the opposite effect on CH4 production when compared to the addition of Rice bran. In Study 4, TMR were formulated using roughages and Ipil-ipil and addition of Wheat bran, Kashari bran, and Rice bran in various combinations. The TMR based on German grass and maize silage with specific bran combinations showed significant reductions in in vitro CH4 production, CH4 concentration, and CH4/dOM while maximising GP, dOM, ME, and the associative effects. To conclude, the results of chemical analyses and in vitro fermentation studies showed that specific combinations of roughages, protein feeds, and by-products available in Bangladesh have the potential to formulate rations for cattle that help making livestock production more sustainable. The results may be also relevant for other developing nations. It is suggested to verify the results of the present project by animal trials at local conditions.
Bi-objective optimization of nutrient intake and performance of broiler chickens using Gaussian process regression and genetic algorithm
(2023) Ahmadi, Hamed; Rodehutscord, Markus; Siegert, Wolfgang
This study investigated whether quantifying the trade-off between the maxima of two response traits increases the accuracy of diet formulation. To achieve this, average daily weight gain (ADG) and gain:feed ratio (G:F) responses of 7–21-day-old broiler chickens to the dietary supply of three nutrients (intake of digestible glycine equivalents, digestible threonine, and total choline) were modeled using a newly developed hybrid machine learning-based method of Gaussian process regression and genetic algorithm. The dataset comprised 90 data lines. Model-fit-criteria indicated a high model adjustment and no prediction bias of the models. The bi-objective optimization scenarios through Pareto front revealed the trade-off between maximized ADG and maximized G:F and provided information on the needed input of the three nutrients that interact with each other to achieve the trade-off scenarios. The trade-off scenarios followed a nonlinear pattern. This indicated that choosing target values intermediate to maximized ADG and G:F after single-objective optimization is less accurate than feed formulation after quantifying the trade-off. In conclusion, knowledge of the trade-off between maximized ADG and maximized G:F and the needed nutrient inputs will help feed formulators to optimize their feed with a more holistic approach.
Bone ash data in the context of phosphorus and phytase evaluation in poultry
(2021) Künzel, Susanne; Rodehutscord, Markus
Phosphorus (P) is an essential element that is crucial for various metabolic processes in the body of animals and humans. To keep the animals healthy and to obtain food products rich in nutrients, an adequate P supply is indispensable. Plant feedstuffs, the main components of poultry diets, contain P in a form that is only partially available to poultry. For this reason, poultry diets are often supplemented with mineral P. However, global rock phosphate reserves, where mineral P is mined from, are limited. Additionally, excessive P supply should also be avoided because of the environmental impact of P accumulation in the soil. Consequently, P supply not exceeding the requirements of poultry is essential to ensure animal wellbeing and to protect the environment. In order to feed diets with adequate concentrations of P, it is necessary to have suitable approaches for the determination of available P in the animal. The availability of P varies widely between feed components and it is also influenced by feed supplements and other factors. Bone ash analysis is an often-used tool to evaluate the relative bioavailability of P since a high amount of P is stored in the bones. A standard assay for bone ash analyses has never been agreed on. Therefore, many different approaches are described in the literature with an unknown impact on the results of P bioavailability studies. The main objective of the present thesis was to examine the suitability of bone ash data for the evaluation of available P in poultry with emphasis on methodological aspects. Therefore, different studies with broiler chickens and Japanese quail were conducted. The experiments comprised various aspects related to P availability in poultry. The effect of feed supplements in the form of phytase products, myo-inositol and a coccidiostat were evaluated. Furthermore, quantitative genetic analyses were performed. All experiments had in common that tibiotarsus (tibia) or foot ash data or both were used for the examination of the relative bioavailability of P. Based on the data that accrued during the studies described in the four manuscripts of this thesis, comprehensive methodological analyses were performed. The tibia and foot were compared regarding their appropriateness as a trait for the evaluation of the relative bioavailability of P by using bone ash data. The relationship between the two traits was investigated, as well as the relationship between foot or tibia ash and quantitative P measurements. Additionally, P concentration in the ash of both bone fractions was analysed and compared. Results indicated only minor differences between tibia and foot ash data. No clear preference for one of them could be deduced from the data. The left and right feet of broiler chickens were compared in terms of both ash concentration and total ash amount. Significant differences between the two feet of the same animal were detected for both traits. Consequently, not only the choice of the bone fraction but also of the body side should be considered when sampling for bone ash data. Ash data are mostly expressed as a concentration of the dry matter content of the bone. Also possible is the use of the absolute ash amount. The relationship of both ways of expression with traits of quantitative P measurements was analysed by using correlation coefficients and regression analyses. Results showed that the absolute ash amount was at least as suitable as ash concentration but has the advantage that it is easier to determine. Possible selection procedures for animals for bone ash analyses were simulated with data from two of the experiments. Often it is not possible to use all animals involved in an experiment for bone ash analyses. Therefore, the influence of sampling frequency and selection method on the outcome of P availability studies was evaluated. Results indicated that the number and selection method of animals for bone ash data might influence the results. However, it was not possible to recommend a specific selection method based on the obtained results. Estimates of heritability and genetic correlations showed the suitability of bone ash data as a proxy trait for P efficiency breeding of poultry. The absolute amount of bone ash data appeared to be most promising for this purpose. Bone ash data are a very useful and easy to determine trait to estimate the relative bioavailability of P. However, investigations performed in this thesis showed the importance of a careful selection of methods. A standardised assay would be helpful to obtain meaningful and more comparable estimates of relative P bioavailability.
Composition of the ileum microbiota is a mediator between the host genome and phosphorus utilization and other efficiency traits in Japanese quail (Coturnix japonica)
(2022) Haas, Valentin; Vollmar, Solveig; Preuß, Siegfried; Rodehutscord, Markus; Camarinha-Silva, Amélia; Bennewitz, Jörn
Background: Phosphorus is an essential nutrient in all living organisms and, currently, it is the focus of much attention due to its global scarcity, the environmental impact of phosphorus from excreta, and its low digestibility due to its storage in the form of phytates in plants. In poultry, phosphorus utilization is influenced by composition of the ileum microbiota and host genetics. In our study, we analyzed the impact of host genetics on composition of the ileum microbiota and the relationship of the relative abundance of ileal bacterial genera with phosphorus utilization and related quantitative traits in Japanese quail. An F2 cross of 758 quails was genotyped with 4k genome-wide single nucleotide polymorphisms (SNPs) and composition of the ileum microbiota was characterized using target amplicon sequencing. Heritabilities of the relative abundance of bacterial genera were estimated and quantitative trait locus (QTL) linkage mapping for the host was conducted for the heritable genera. Phenotypic and genetic correlations and recursive relationships between bacterial genera and quantitative traits were estimated using structural equation models. A genomic best linear unbiased prediction (GBLUP) and microbial (M)BLUP hologenomic selection approach was applied to assess the feasibility of breeding for improved phosphorus utilization based on the host genome and the heritable part of composition of the ileum microbiota. Results: Among the 59 bacterial genera examined, 24 showed a significant heritability (nominal p ≤ 0.05), ranging from 0.04 to 0.17. For these genera, six genome-wide significant QTL were mapped. Significant recursive effects were found, which support the indirect host genetic effects on the host’s quantitative traits via microbiota composition in the ileum of quail. Cross-validated microbial and genomic prediction accuracies confirmed the strong impact of microbial composition and host genetics on the host’s quantitative traits, as the GBLUP accuracies based on the heritable microbiota-mediated components of the traits were similar to the accuracies of conventional GBLUP based on genome-wide SNPs. Conclusions: Our results revealed a significant effect of host genetics on composition of the ileal microbiota and confirmed that host genetics and composition of the ileum microbiota have an impact on the host’s quantitative traits. This offers the possibility to breed for improved phosphorus utilization based on the host genome and the heritable part of composition of the ileum microbiota.
Degradation of crude protein and starch of corn and wheat grains in the rumen
(2016) Seifried, Natascha; Rodehutscord, Markus
The major objectives of the present thesis were to characterize the ruminal crude protein (CP) and starch degradation of different genotypes of corn and wheat grains and to predict the effective degradation (ED) of CP and starch with easily measurable characteristics. The in situ method is the standard technique to study the ruminal degradation of feeds in many feed evaluation systems. This technique was originally applied to study forages and it was therefore necessary to clarify methodical details related to the measurements of in situ starch degradation from cereal grains. Two in vitro and one in situ approach were conducted to study the loss of secondary starch particles from bags with different pore sizes used for the in situ incubation of different cereal grains. In the first in vitro study ground wheat was incubated in bags (pore size: 50 µm) over different time spans in a modified rumen simulation technique. Bag residues and fermenter fluids were analyzed for their starch content. In the second in vitro study ground wheat, barley, and corn were incubated with bags of pore sizes of 50, 30 (except corn), 20, and 6 µm. In the in situ study ground wheat, barley, corn, and oats were rumen incubated over different time spans using bags with pore sizes of 50, 20, and 6 µm. The starch content of the grains and bag residues was analyzed enzymatically and the degradation characteristics of starch were calculated for each grain type and pore size. It was shown for the first time that incubating wheat and barley in bags with 50 and 30 µm pore size lead to a substantial amount of secondary starch particle losses during incubation process in vitro. These losses were not detectable when the grains were incubated with bags having pore sizes of 20 and 6 µm. Independent of the bags’ pore size no secondary starch particle losses were found by the incubation of corn. Thus corn can be studied in situ even with bags with 50 µm pore size. Oats showed very high washout losses with all pore sizes tested in the present thesis and therefore none of them is suitable to study the starch degradation measurements of oats. Because of methodical problems of gas accumulation in bags having pore sizes < 50 µm, no recommendations can be provided for the in situ evaluation of wheat and barley. Further research is necessary to solve these problems. In the second and third study of the present thesis ruminal in situ degradation of 20 corn grain genotypes and 20 wheat grain genotypes was measured in three lactating Jersey cows. In both experiments the same techniques were used to characterize the ruminal degradation of CP and starch. Ground grains (2 mm) were rumen incubated in bags (50 µm pore size) over different time spans. Grains and bag residues were analyzed for their CP and starch content. The degradation parameters and the ED were calculated for dry matter (DM), CP, and starch. Gas production (GP) of ground grains (1 mm) was recorded after incubation over different time spans in buffered ruminal fluid and fitted to an exponential equation to determine GP parameters. To predict ED of CP and starch correlations with physical and chemical characteristics and in vitro measurements were evaluated and stepwise multiple linear regression analyses were applied. The in situ parameters (soluble fraction, potential degradable fraction, and degradation rate) varied widely between genotypes of corn and wheat grains. The ED of DM, CP, and starch showed a high variation for corn grain genotypes. Due to the high degradation rates, the ED of wheat grains were similar between genotypes. The GP rate was in good agreement with the in situ values for corn grains, whereas no systematic relationship between both methods was observed for wheat grains. Evaluation of correlation analysis showed significant relationships between calculated ED of CP and several amino acids (AA) for both grain types. This indicates that the protein composition of the grains influences CP degradation in the rumen. Similar relationships were found between the same AA and ED of starch of corn grains which highlights the impact of the protein composition on ruminal starch degradation for this grain type. For both grain types, the ED of starch and CP could be predicted accurately from physical and chemical characteristics alone or in combination with GP measurements. Thus, the equations presented in the present thesis can be used to obtain rapid and cost effective information on ruminal degradation of CP and starch for corn and wheat grains. The results of the present thesis show that there is considerable variation of ruminal CP and starch degradation from different genotypes of corn and also – albeit to a lesser extent – for wheat grains. Differences in ED of starch should be taken into account when formulating rations containing significant amounts of corn and wheat grains. In the case of corn grains differences in ED of CP should also be accounted for.
Dietary phosphorus and calcium in feed affects miRNA profiles and their mRNA targets in jejunum of two strains of laying hens
(2021) Iqbal, Muhammad Arsalan; Ali, Asghar; Hadlich, Frieder; Oster, Michael; Reyer, Henry; Trakooljul, Nares; Sommerfeld, Vera; Rodehutscord, Markus; Wimmers, Klaus; Ponsuksili, Siriluck
Phosphorus (P) and calcium (Ca) are critical for egg production in laying hens. Most of P in plant-based poultry diet is bound as phytic acid and needs to be hydrolysed before absorption. To increase P bioavailability, exogenous phytases or bioavailable rock phosphate is added in feed. There is growing evidence of the importance of miRNAs as the epicentre of intestinal homeostasis and functional properties. Therefore, we demonstrated the expression of miRNA profiles and the corresponding target genes due to the different levels of P (recommended vs. 20% reduction) and/or Ca (recommended vs. 15% reduction) in feed. Jejunal miRNA profiles of Lohmann Selected Leghorn (LSL) and Lohmann Brown (LB) laying hens strains were used (n = 80). A total of 34 and 76 miRNAs were differentially expressed (DE) in the different diet groups within LSL and LB strains respectively. In LSL, the DE miRNAs and their targets were involved in calcium signaling pathway, inositol phosphate metabolism, and mitochondrial dysfunction. Similarly, in LB miRNAs targets were enriched in metabolic pathways such as glutathione metabolism, phosphonate metabolism and vitamin B6 metabolism. Our results suggest that both strains employ different intrinsic strategies to cope with modulated P and Ca supply and maintain mineral homeostasis.
Effects of a reduction of dietary levels of calcium and phosphorus on performance, bone minerals and mineral excretion of turkey breeder hens in the rearing and laying period
(2023) Gickel, Julia Maria; Rodehutscord, Markus
Phosphorus (P) is an essential mineral in feed for livestock and has finite resources all over the world. The aim of this study was to obtain an idea about the reduction potential of P in the diets of turkey breeders. As the metabolism of P is interlinked with the metabolism of calcium (Ca), Ca was also examined. Therefore, the requirements of P and Ca were studied using a factorial approach. As data about the requirements of these minerals in turkey breeders is limited, the present study mostly used data from other poultry species. Thus, this study can be viewed as an approach to building new resilient data for turkey breeder hens. The results of the factorial approach were embedded in a feeding program for turkey breeder hens during rearing and laying with two different treatments. One group was fed a standard feed (practical diet used before the trial), while the other group was fed a Ca/P reduced feed. In total, four trials were conducted: trials I and III focused on the rearing period while trials II and IV focused on the laying period. All trials were observed independently but the hens from trial I were used also in trial II afterwards for studying long-term effects. In each trial, body weight and feed intake were measured throughout the trial period. Bone mineralization was studied in trial I, including analyses of bone ash as well as Ca and P in the bone ash from the tibia of fallen and culled animals. The egg components were studied in eggs from trial II, including analyses of the percentages of albumen, egg yolk, and egg shell; Ca in albumen; P in albumen; Ca in egg yolk; P in egg yolk; water in egg shell; Ca in egg shell; and P in egg shell. In trials II and IV, laying performance, egg weights, number (and causes) of culled eggs, fertility, hatchability, as well as body weight and fitness of the hatchlings were additionally observed. At the end of each trial, the concentrations of dry matter, Ca, and P in manure were analyzed and a nutrient balance was calculated to classify the results. Body weight development exhibited significant differences between the treatments in eight out of 30 weeks in trial I (five weeks had a higher mean weight in the group with standard feed; three weeks had a higher mean weight in the group with Ca/P reduced feed); three out of 28 weeks in trial II (two weeks had a higher mean weight in group with Ca/P reduced feed; one week had a higher mean weight in the group with standard feed); four out of 30 weeks in trial III (higher mean weight in the group with Ca/P reduced feed); and two weeks out of 28 in trial IV (higher mean weight in the group with Ca/P reduced feed). The feed intake data also fluctuated, with three out of 30 weeks exhibiting a significant difference in trial I (higher mean feed intake in the group with Ca/P reduced feed) and one week with a significant difference in trial II (higher mean feed intake in the group with standard feed). The observations throughout all of the trials revealed the tendency for a higher mean feed intake in the groups with Ca/P reduced feed. A significant difference concerning bone mineralization was not observed. Moreover, the results of laying performance, fertility, and hatchability exhibited no differences between the treatments. Regarding the egg weight in trial II, there were eight weeks with a significant difference between the treatments (seven weeks with a higher mean egg weight in the group with standard feed; one week with a higher mean egg weight in the group with Ca/P reduced feed). In trial IV, only one week exhibited a significant difference between the treatments (higher mean egg weight in the group with Ca/P reduced feed). Regarding the weight of hatchlings in trial II, three weeks had a significant difference between the groups (higher mean weight in the group with standard feed). In trial IV, no significant difference was observed for any week all throughout the trial period. The analyses of egg components revealed significant differences between the treatments according to the percentages of albumen and egg yolk and the calculated value of total Ca in the egg (including the shell). In trials I and III, significant differences existed between the treatments in the concentration of Ca in manure, with a lower level in the group with Ca/P reduced feed. This study concluded that a reduction in the dietary levels of Ca from 1.00–1.10% to 0.56–0.80% and of P from 0.48–0.61% to 0.35–0.50% (av. P) or 0.70–0.80% to 0.50–0.60% (total P) in rearing and a reduction of the dietary levels of Ca from 2.90% to 2.80% or 2.60% and of P from 0.36% to 0.30% or 0.24% (av. P) or from 0.65% to 0.50% (total P) in laying in the feed of turkey breeder hens are possible and did not result in disadvantages. As the present study also compared the results with required recommendations and target levels from breeding companies, it was also able to conclude that these levels are obsolete and should be adjusted downwards.
Effects of a transient lack of dietary mineral phosphorus on renal gene expression and plasma metabolites in two high-yielding laying hen strains
(2025) Qasir, Hiba; Reyer, Henry; Oster, Michael; Ponsuksili, Siriluck; Trakooljul, Nares; Sommerfeld, Vera; Rodehutscord, Markus; Wimmers, Klaus
Background: There is an emerging body of evidence that current poultry feed is formulated in excess for phosphorus (P), which results in unnecessarily high P excretions. Sustainable concepts for agricultural P flows should trigger animal-intrinsic mechanisms for efficient P utilization. In the current study, Lohmann Brown (LB) and Lohmann Selected Leghorn (LSL) laying hens were fed either a high P diet (P+) with 1 g/kg mineral P supplement or a low P diet (P-) with 0 g/kg mineral P supplement for a period of 4 weeks prior to sampling. Before and after onset of laying, i.e., at 19 and 24 weeks of life, kidney and plasma samples were collected to investigate the endogenous P utilization in response to restricted dietary P, laying hen strain, and sexual maturation. Results: Plasma analyses of minerals and metabolites confirmed the response to a low P diet, which was characterized by a significant reduction in plasma P levels at week 19 in both strains. The plasma calcium (Ca) levels were tightly regulated throughout the entire experimental period. Notably, there was a numerical trend of increased plasma calcitriol levels in P- fed birds of both strains compared to the P + group, which might have mediated a substantial role regarding the adaptive responses to low P supply. At week 19, RNA sequencing of kidney identified 1,114 and 556 differentially expressed genes (DEGs) unique to the LB and LSL strains, respectively. The number of DEGs declined with increasing maturity of the hens culminating in 90 and 146 DEGs for LB and LSL strains at week 24. Analyses revealed an enrichment of pathways related to energy metabolism and cell cycle, particularly at week 19 in both strains. The diet-specific expression of target genes involved in P homeostasis highlighted transcripts related to active ( SLC34A1 , SLC20A2 ) and passive mineral transport ( CLDN14 , CLDN16 ), Ca utilization ( STC1 , CALB1 ), and acid-base balance ( CA2 , SLC4A1 ). Conclusions: Results suggest that both laying hen strains adapted to the lack of mineral P supplements and achieved a physiological Ca: P-ratio in body compartments through endogenous regulation as evidenced via the endocrine profile.
Effects of calcium level and source, formic acid, and phytase on phytate degradation and the microbiota in the digestive tract of broiler chickens
(2021) Krieg, Jochen; Borda-Molina, Daniel; Siegert, Wolfgang; Sommerfeld, Vera; Chi, Yung Ping; Taheri, Hamid Reza; Feuerstein, Dieter; Camarinha-Silva, Amélia; Rodehutscord, Markus
Background: Diet acidification, dietary calcium (Ca) level, and phytase supplementation are known influences on the microbial community in the digestive tract and on phosphorus (P) utilization of broiler chickens. Effects of dietary factors and microbiota on P utilization may be linked because microorganisms produce enzymes that release P from phytate (InsP6), the main source of P in plant feedstuffs. This study aimed to detect linkages between microbiota and InsP6 degradation by acidifying diets (i.e., replacing Ca carbonate (CaCO3) by Ca formate or adding formic acid to CaCO3-containing diets), varying Ca levels, and supplementing phytase in a three-factorial design. We investigated i) the microbial community and pH in the digestive tract, ii) prececal (pc) P and Ca digestibility, and iii) InsP6 degradation. Results: All factors under investigation influenced digesta pH and the microbiota composition. Predicted functionality and relative abundance of microorganisms indicated that diets influenced the potential contribution of the microbiota on InsP degradation. Values of InsP6 degradation and relative abundance of the strains Lactobacillus johnsonii and Lactobacillus reuteri were correlated. Phytase supplementation increased pc InsP6 disappearance, with differences between Ca levels, and influenced concentrations of lower inositol phosphate isomers in the digestive tract. Formic acid supplementation increased pc InsP6 degradation to myo-inositol. Replacing CaCO3 by Ca-formate and the high level of these Ca sources reduced pc InsP6 disappearance, except when the combination of CaCO3 + formic acid was used. Supplementing phytase to CaCO3 + formic acid led to the highest InsP6 disappearance (52%) in the crop and increased myo-inositol concentration in the ileum digesta. Supplementing phytase leveled the effect of high Ca content on pc InsP6 disappearance. Conclusions: The results point towards a contribution of changing microbial community on InsP6 degradation in the crop and up to the terminal ileum. This is indicated by relationships between InsP6 degradation and relative abundance of phosphatase-producing strains. Functional predictions supported influences of microbiota on InsP6 degradation. The extent of such effects remains to be clarified. InsP6 degradation may also be influenced by variation of pH caused by dietary concentration and solubility of the Ca in the feed.
Effects of dietary phosphorus and myo-inositol supplementation on NaPi-IIb and TRPV6 protein expression in duodenal apical membranes of laying hens from two strains
(2026) Shomina, Nataliia; Sommerfeld, Vera; Hanauska, Anna; Oster, Michael; Rodehutscord, Markus; Huber, Korinna
Phosphorus (P) and calcium (Ca) absorption in the intestine is mediated by apical brush border membrane (BBM) transporters, including the sodium-dependent phosphate (Pi) transporter NaPi-IIb and the Ca²⁺-selective channel TRPV6. Both are highly expressed in the duodenum and exhibit dietary adaptability; yet little is known about how this adaptability varies with strain and age in laying hens. The present study examined the effects of dietary mineral P renunciation and myo-inositol (MI) supplementation on NaPi-IIb and TRPV6 protein expression in the duodenal BBM of Lohmann Brown-Classic (LB) and Lohmann LSL-Classic (LSL) hens. Two independent feeding trials were conducted: hens received diets either with or without mineral P supplementation (wk 15 - 19 and 20 – 24), or with graded MI levels (0 - 3 g/kg; wk 26 – 30). At the end of each period, hens were euthanized and protein expression of NaPi-IIb and TRPV6 in duodenal BBM was studied by western blotting. Statistical correlation with additional traits of mineral metabolism was analyzed. An immunoreactive NaPi-IIb band was detected at ∼ 45 kDa; therefore, all results reported here refer to this NaPi-IIb fragment. Mineral P renunciation did not affect NaPi-IIb fragment or TRPV6 expression in either hen strain. In LSL hens NaPi-IIb fragment expression increased from wk 19 to wk 24, whereas in LB hens it remained unchanged. NaPi-IIb fragment expression was positively associated with duodenal phosphatase activity and plasma estradiol. TRPV6 expression tended to reduce in LB hens from wk 19 to wk 24, but remained stable in LSL hens. TRPV6 expression was positively associated with duodeno-jejunal P content. MI supplementation upregulated NaPi-IIb fragment expression in LB, but downregulated it in LSL hens with high dietary MI levels, without affecting TRPV6. These findings demonstrate strain-dependent regulatory patterns of duodenal expression of NaPi-IIb fragment and TRPV6 in response to physiological stage and MI supply, indicating that mineral feeding strategies may benefit from genotype-specific consideration, whereas the mechanisms underlying MI-related effects require further clarification.
Effects of different dietary energy and essential amino acid levels on growing turkeys under organic farming conditions
(2025) Kirn, Anna Isabella; Rodehutscord, Markus
An adequate supply of dietary apparent metabolizable energy (AMEN) and essential amino acids (EAA) is crucial for turkeys to reach their genetic growth potential. However, optimizing diets in organic turkey production is challenging due to regulatory restrictions under EU Regulation 2018/848, which often lead to increased crude protein levels, potentially compromising animal health and environmental sustainability. To address this, targeted feeding strategies can be applied, leveraging compensatory growth through an initial qualitative feed restriction followed by a re-alimentation phase. Previous research showed that a consistent 10% reduction in AMEN combined with an initial EAA reduction of up to 20% (relative to commercial breeder guidelines), gradually restored to 90-100% during refeeding, effectively triggered compensatory growth, resulting in full catch-up growth. Nevertheless, as nutrient intake also affects welfare indicators such as plumage condition and injury prevalence, a balance between growth and welfare is essential. The present thesis aimed to evaluate whether dietary EAA levels could be further reduced by up to 30%, maintaining a constant 10% AMEN reduction, in male and female turkeys of two genotypes (slow-growing Auburn and fast-growing B.U.T. 6) across various housing systems (indoor, indoor with silage, and mobile housing with free-range). The study focused on evaluating growth, slaughter performance, and welfare indicators. Additionally, the potential of a newly introduced scoring system for wing feather structure and depigmentation was examined as a sensitive indicator of dietary EAA supply status. In the first study, 600 male day-old chicks from two genotypes (B.U.T. 6 and Auburn) were allocated to three feeding strategies differing in EAA levels across five 4-week phases: F1 (80/90/90/90/90%), F2 (80/80/90/100/90%), and F3 (70/80/90/100/90%). After rearing in indoor housing until week 8, the turkeys were transferred to one of three housing systems (indoor, indoor with silage, and mobile with free-range; weeks 9–20). Results showed that up to 30% initial EAA reduction followed by gradual re-alimentation triggered compensatory growth without affecting final body weight, total feed intake, or valuable cuts. No genotype by feeding interaction was observed. However, compensatory feed intake increased the feed conversion ratio and nutrient intake per kilogram of weight gain. While there was no housing by feeding interaction, housing significantly affected performance. Mobile housing with free-range and silage supplementation improved the feed conversion ratio and reduced abdominal fat. Additionally, mobile housing increased breast meat yield. Notably, a significant genotype by housing interaction suggested that B.U.T. 6 may demonstrate superior adaptability to the nutritional and environmental variability of the housing systems compared to Auburn. Building on this, the second study tested a similar approach in 432 female turkeys across three feeding strategies (F1: 80/90/90/90%, F2: 80/80/90/90%, F3: 70/80/90/90%) over four four-week phases. After being reared indoors until week 8, birds were assigned to indoor or mobile housing with free-range access (weeks 9-16). While a 20% EAA reduction maintained growth and slaughter performance, a 30% reduction led to significant decreases in final body weight, slaughter weight, and dressing percentage. A significant genotype by feeding interaction suggested that B.U.T. 6 was more sensitive to the 30% reduction than Auburn. Mobile housing improved breast meat yield and reduced abdominal fat but increased feed conversion ratio, likely due to winter fattening conditions. The third study examined the effects of dietary AMEN and EAA reductions on injuries, plumage damage, and the potential of a newly introduced scoring system for wing feather structure and depigmentation as an indicator of dietary EAA supply status. A total of 216 non-beak-trimmed female turkeys from the second study, housed indoors and subjected to the same feeding strategies, were evaluated. The most restrictive feeding strategy (30% EAA reduction) increased the prevalence of skin injuries, plumage damage, alterations in feather structure, and depigmentation. In contrast, the 20% EAA reduction preserved welfare and performance outcomes. Importantly, wing feather structure and pigmentation showed graded responses to varying EAA levels, supporting their potential as sensitive indicators of dietary EAA status in turkeys. In conclusion, this thesis demonstrated that reducing EAA levels by 20%, followed by a re-alimentation phase, alongside a 10% AMEN reduction, can be successfully applied in organic turkey production. This approach maintained final growth performance, slaughter performance, and welfare in male and female Auburn and B.U.T. 6 turkeys by effectively leveraging compensatory growth. Wing feather structure and pigmentation scoring proved to be a sensitive indicator of EAA supply status. External feed sources such as silage and free-range foraging contributed nutritionally, while mobile housing enhanced breast muscle yield and reduced abdominal fat. Overall, turkey growth and performance under organic conditions resulted from complex interactions between endogenous factors (sex, genotype, age) and exogenous factors (housing, environment, feeding), highlighting the need for integrated and adaptive management approaches.
Effects of feeding diets without mineral P supplement on intestinal phytate degradation, blood concentrations of Ca and P, and excretion of Ca and P in two laying hen strains before and after onset of laying activity
(2024) Sommerfeld, Vera; Bennewitz, Jörn; Camarinha-Silva, Amélia; Feger, Martina; Föller, Michael; Huber, Korinna; Oster, Michael; Ponsuksili, Siriluck; Schmucker, Sonja; Seifert, Jana; Stefanski, Volker; Wimmers, Klaus; Rodehutscord, Markus
The objective of this study was to characterize intestinal phytate degradation and mineral utilization by 2 laying hen strains before and after the onset of egg laying using diets without or with a mineral phosphorus (P) supplement. One offspring of 10 roosters per strain (Lohmann Brown-classic [LB] and Lohmann LSL-classic [LSL]) was sacrificed before (wk 19) and after (wk 24) the onset of egg-laying activity and following 4 wk placement in a metabolic unit. Diets were corn-soybean meal-based and without supplemented P (P-) or with 1 g/kg supplemented P (P+) from monocalcium phosphate. In wk 19 and 24, the blood plasma and digesta of duodenum+jejunum and distal ileum were collected. The concentration of P in blood plasma was higher in hens fed P+ than P- (P < 0.001). In duodenum + jejunum and ileum content, the concentrations of InsP6, Ins(1,2,4,5,6)P5 and Ins(1,2,3,4,5)P5 were lower in P- than in P+ (P ≤ 0.009). In duodenum+jejunum, the concentrations of InsP6, Ins(1,2,4,5,6)P5 and Ins(1,2,3,4,5)P5 were lower in wk 24 than 19 and lower in LSL than LB hens (P < 0.001). The concentration of myo-inositol (MI) in duodenum + jejunum content was lower in wk 19 than 24 (P < 0.001). Following a 4-d total excreta collection, the retained amount of P was higher in P+ than P- (P < 0.001). Phosphorus retention was lower in LB hens fed P- than in other treatments (P × strain: P = 0.039). In the jejunal tissue, some genes related to intracellular InsP metabolism were higher expressed in LB than LSL hens. The renunciation of mineral P increased endogenous phytate degradation, but more P was retained with supplemented P. Differences in endogenous phytate degradation between the periods before and after the onset of egg laying might be attributed to different Ca concentrations in intestinal digesta caused by different Ca needs in both periods.
Effects of monensin and tannin extract supplementation on methane production and other criteria of rumen fermentation in vitro and in long-term studies with sheep
(2013) Wischer, Gerald; Rodehutscord, Markus
Ruminants increasingly attract public concern due to their methane release and contribution to the greenhouse effect. One strategy to reduce the release of methane is to modify microbial fermentation in the rumen by the use of feed additives such as monensin and tannin extracts. However, other characteristics of fermentation including the synthesis of microbial protein may also be affected. The aim of the present studies was to provide a comprehensive evaluation of the effects of monensin and tannin extracts on ruminal fermentation and methane production. The ionophore monensin is known to increase feed efficiency in ruminants. Although the use of silages is common practice in cattle feeding, the effects of monensin on the fermentation of silages in the rumen and microbial protein synthesis are lacking. Monensin has often been described to have indirect effects on methane production resulting from its effects on feed intake, protozoa and Gram-positive bacteria. It has rarely been studied whether monensin can reduce methane production without adverse effects on other criteria of rumen fermentation. The first objective therefore was to investigate the effects of different dosages of monensin on methane production and microbial protein synthesis when supplemented to different silages in two in vitro systems (Study 1). In Experiment 1 of Study 1, 15 g of oven-dried grass silage alone or combined with a concentrate was incubated in a rumen simulation (Rusitec) over a period of 13 d to examine the effects of monensin supplementation (2 or 4 mg/d, n = 4) on the production of total gas, methane, volatile fatty acids (VFA), degradation of nutrients and microbial protein synthesis. In Experiment 2 of Study 1, different dosages of monensin (0.5, 1, 2, 6 and 10 µg) were supplemented to syringes containing 120 mg of grass silage alone, grass silage combined with concentrates, or maize silage alone. After 24 h of incubation the effects of monensin on total gas, methane and VFA production were determined. In Experiment 1 monensin inclusion to grass silage and grass silage combined with concentrate resulted in a decreased total gas, methane and acetate production, while propionate production was increased. Along with a decreased degradation of crude protein, ammonia concentration in the system was reduced. While microbial protein originating from solid associated microbes decreased with monensin inclusion, microbial protein from liquid associated microbes was increased, resulting in an increase in total microbial protein synthesis. In Experiment 2, different dosages of monensin reduced methane production in grass silage (17%), grass silage combined with concentrate (10%) and maize silage (13%) without adverse effects on total gas production. Based on these two in vitro experiments it was concluded that monensin is able to reduce methane production without a major decrease in total gas and VFA production and degradation of organic matter. Although microbial fractions were differently affected, the total microbial protein synthesis was increased upon monensin supplementation. Tannins are secondary plant compounds that are known to complex with feed and microbial proteins. Several products from this heterogeneous group have shown potential to affect rumen fermentation in vivo and, even more, in vitro, but are often accompanied by negative effects on digestibility, feed intake and microbial protein synthesis. In Study 2 of the present work, ten tannin extracts (chestnut, mimosa, myrabolan, quebracho, sumach, tara, valonea, oak, cocoa and grape seed) and four monomers of rapeseed tannin (pelargonidin, catechin, cyanidin and sinapinic acid) were screened in grass silage based diets in successive runs using the Hohenheim Gas Test. The objective was to determine the optimal dosage of each tannin extract to cause a maximal methane reduction without negative effects on total gas production. Whereas the supplementation of pelargonidin and cyanidin to grass silage did not reduce methane production; catechin and sinapinic acid reduced methane production without affecting total gas production. Except tara extract, all tannin extracts reduced methane production by 8 to 28% without adverse effects on total gas production. Based on these results, chestnut, grape seed, myrabolan, sumach and valonea extract were investigated in a second step in a Rusitec to determine their effects on degradation of nutrients, VFA and ammonia production, and particularly on microbial protein synthesis. All tannin extracts were supplemented at similar dosages of 1.5 g to 15 g of grass silage. The supplementation of chestnut resulted in the greatest decrease in methane production (63%), followed by valonea (35%), grape seed (23%), sumach (18%), and myrabolan (7%; not significantly different from the control). While chestnut extract reduced acetate production by 19%, supplementation with grape seed or myrabolan extract increased acetate production; however, degradation of fibre fractions was reduced in all tannin treatments. Degradation of dry and organic matter was reduced by all tannin extracts, but there were no differences between tannin treatments. Crude protein degradation and ammonia production were also reduced by tannin extract supplementation. Microbial protein synthesis and its efficiency were not affected by tannin supplementation, which indicates that a reduction in methane production due to tannin extract supplementation is possible without negatively affecting microbial protein synthesis. Chestnut and valonea extract had the greatest potential in reducing methane production without negative effects on rumen fermentation of grass silage and microbial protein synthesis. Therefore, these tannin extracts were investigated for their long-term effects in sheep (Study 3). In Experiment 1 of Study 3, sheep receiving the control, chestnut or valonea treatment (each n = 4) were fed 842 g/d of hay (fresh weight). The animals on the control treatment also received 464 g/d of concentrate, and animals on the tannin treatments received the same amount of concentrate but were also fed 20 g of the respective tannin extract. Following initiation of tannin feeding, methane release from sheep was measured in 23.5 h intervals in respiration chambers on day 1, 8, 15, 29, 57, 85, 113, 148, and 190. In three balances periods faeces and urine were collected for 6 and 3 days, respectively. Effects on nutrient digestibility, nitrogen and energy metabolism were evaluated, with microbial protein synthesis estimated from the urinary excretion of purine derivatives. Based on the results of Experiment 1, a second experiment was conducted four month after the start of Experiment 1. Experiment 2 had the same study design and data collected, but the dosage of tannin extracts was doubled compared to Experiment 1 (0.9 vs. 1.7 g tannin extract/kg body weight) and the duration was shorter (85 days). Hay and concentrates used in both experiments were also evaluated using the Hohenheim Gas for their effects on total gas and methane production. In both experiments, methane release was not significantly reduced by tannin extract supplementation when analysed over the whole experimental period. In Experiment 1 the supplementation of chestnut extract on day 190 resulted in a reduced methane release. In both experiments, on day 1 a numeric reduction in methane release for the tannin treatments was observed, with a greater reduction recorded for the higher dosage used in Experiment 2. This trend disappeared by day 57. In the third balance period of Experiment 1, digestibility of dry and organic matter was reduced by tannin supplementation. The digestibility of crude protein was reduced in both experiments, whereas the digestibility of fibre fractions was not influenced. In both experiments a long-lasting shift in nitrogen excretion from urine to faeces was observed, which occurred to a greater extent in Experiment 2. The urinary excretion of purine derivatives was not significantly affected by tannin supplementation, indicating that the microbial protein synthesis was not altered in either experiment. The in vitro methane production was reduced for concentrates containing tannin extracts, but it was not significantly affected when concentrates were incubated with hay. It is concluded that monensin added to different silages caused a decrease in methane production without affecting total gas production but with an increased microbial protein synthesis. Nine of the ten considered tannin extracts and two tannin monomers decreased methane production without affecting total gas production. The Rusitec study confirmed the great potential of chestnut and valonea extract to reduce methane production without negative effects on microbial protein synthesis. However, neither chestnut nor valonea extract reduced the methane release in sheep when fed over a longer period of time. It is assumed, that rumen microbes adapted to the tannin dosages in terms of methane release but not nitrogen metabolism, as there were long-lasting effects on nitrogen excretion. The shift in nitrogen excretion can have a positive effect on the environment due to the reduced potential of ammonia emission from the urine. Both in vitro systems used in the present studies showed effects of tannin extracts that were considerably different from those observed in sheep. The monomers investigated in the present study are the basic units of condensed tannins, whereas the tannin extracts selected in vitro only contain hydrolysable tannins. It is possible that monomers of chestnut and valonea extract may reduce methane production, whereas higher dosages of these tannin extracts cause negative effects on feed intake, digestibility and microbial protein synthesis. Further investigations should focus systematically on the transfer of in vitro studies to estimate in vivo responses. Therefore, a parallel implementation of different in vitro and respiration studies would be of great value.
Effects of myo-inositol supplementation in the diet on myo-inositol concentrations in the intestine, blood, eggs, and excreta of laying hens
(2025) Sommerfeld, Vera; Hanauska, Anna; Huber, Korinna; Bennewitz, Jörn; Camarinha-Silva, Amélia; Feger, Martina; Föller, Michael; Oster, Michael; Ponsuksili, Siriluck; Schmucker, Sonja; Seifert, Jana; Stefanski, Volker; Wimmers, Klaus; Rodehutscord, Markus
The objectives of this study were to investigate whether an increased dietary myo-inositol (MI) supply translates into changes in MI concentrations and endogenous mucosal phosphatase activities in the intestine of laying hens and whether different laying hen strains respond differently to MI supplementation. The diets were corn–soybean meal-based and supplemented without (MI0) or with 1 (MI1), 2 (MI2), or 3 (MI3) g MI/kg feed. Ten hens per strain (Lohmann Brown-classic (LB) and Lohmann LSL-classic (LSL)) and diet were sacrificed at the age of 30 wk following a 4-wk stay in a metabolic unit. The blood plasma, digesta of the duodenum+jejunum and distal ileum, mucosa of the duodenum, and eggs were collected at wk 30. The concentration of MI in the blood plasma was increased by MI supplementation (P < 0.001); however, that of MI3 did not further increase compared with MI2. The concentration of MI in the duodenum+jejunum and ileum increased steadily (P < 0.001). The MI concentration in the duodenum+jejunum was higher in LB than in LSL hens (P = 0.017). The MI concentration in egg yolk was increased by MI supplementation (P < 0.001) and was higher in LB than in LSL hens (P = 0.015). Strain or diet did not affect mucosal phosphatase activity. Myo-inositol flow at the terminal ileum and postileal disappearance increased with each increment in MI supplementation (P < 0.001) and was higher in LB than in LSL hens (P ≤ 0.041). Regression analysis indicated that, on average, 84% of supplemented MI was retained in the body or metabolized and excreted in a different form. Based on the measured MI concentrations in the blood and eggs, dietary MI was not completely absorbed in the small intestine and, to a different extent, in the two laying hen strains. A higher dietary MI supply was followed by higher intestinal absorption or metabolism by microorganisms. The fate of supplemented MI and its relevance to birds warrant further research.
Effects of Prosopis juliflora pods on sheep performance and carcass traits, and their methane mitigation potential as assessed in vitro
(2025) Tadesse, Assefa; Titze, Natascha; Rodehutscord, Markus; Melesse, Aberra
This study aimed to assess the effect of partial substitution of concentrate mix (CM) with Prosopis juliflora pod (PJP) on growth performance and carcass traits in sheep, and its potential to reduce methane (CH4) production in vitro. A total of 25 yearling rams with an initial body weight of 15.8±1.53 kg was randomly assigned to five treatment diets. The diets were a control diet (JP0) and JP0 replaced with PJP at the level of 5% (JP5), 10% (JP10), 15% (JP15), and 20% (JP20). Data were collected on feed intake, body weight, and carcass characteristics. Methane production was determined from 24 h in vitro gas production. The substitution of CM with PJP did not affect feed intake, body weight, weight gain, feed efficiency, and carcass traits (p>0.05). The in vitro CH4 production was reduced (p<0.001) in diets containing increased levels of PJP. The metabolizable energy for JP10, JP15 and JP20 diets was 11.9, 11.6 and 11.5 MJ/kg DM, respectively (p>0.05). In conclusion, replacing CM with PJP did not negatively affect growth performance and carcass characteristics and could be used as potential supplement to mitigate methane emissions. Further in vivo studies involving respiration chambers are recommended to investigate the CH4 reduction potential of PJP.
Endogenous mucosal phosphatases characterization in duodenum brush border membrane of laying hens
(2025) Hanauska, Anna; Sommerfeld, Vera; Schollenberger, Margit; Huber, Korinna; Rodehutscord, Markus
Chicken mucosal phosphatases can partially degrade phytate contained in the feed. Little is known about the characteristics and degradation products of such mucosal phosphatases and the effects of age and genetic strain of the chicken. The objective of this study was to characterize endogenous mucosal phosphatases of two laying hen strains fed diets with or without mineral phosphorus (P) before and after the onset of egg laying. Hens of the strains Lohmann Brown-classic (LB) and Lohmann LSL-classic (LSL) were sacrificed in weeks 19 and 24 of age after 4 weeks of feeding one of two diets with (P+) or without (P-) mineral P supplement. Mucosa of the duodenum was collected, and the brush border membrane (BBM) of enterocytes was enriched and used for phosphatase activity determination. Additionally, the BBM was used in a modified three-step in vitro assay to study the InsP6 degradation products. The results of both in vitro assays were not significantly affected by hen strain and diet. The activity of mucosal phosphatase in 19-week-old hens was, on average, 0.8 µmol Pi/g BBM protein/min lower than in 24-week-old hens (P < 0.002). Consistently, the InsP6 concentration in the incubation residue was significantly higher in 19-week-old hens than in 24-week-old hens (P < 0.001). In the incubation residue, the concentrations of Ins(1,2,3,4,5)P5, Ins(1,2,3,4,6)P5, and Ins(1,2,3,4)P4 were significantly lower (P ≤ 0.002), and those of InsP3 and InsP2 were significantly higher (P ≤ 0.027) when BBM of 24-week-old hens was used compared to 19-week-old hens. The InsP6 degradation products suggest the activity primarily of a 6- and secondarily of a 5-phytase in the duodenal mucosa. The consistent results from both in vitro assays provide a comprehensive characterization of these enzymes. Under the conditions of this study, small intestine calcium concentration appeared to influence mucosal enzyme activity more than dietary mineral P supplementation.
Entwicklung und ernährungsphysiologische Bewertung mikrobieller Hybrid-Phytasen
(2023) Metten, Alexander; Rodehutscord, Markus
To degrade the organic phosphate storage in the best possible way, it is necessary to increase phytase efficiency in vivo. Both a better understanding of the influencing factors limiting phytate degradation in vivo and a continuous improvement of the biochemical properties of phytases to be best adapted to the conditions in the digestive tract of non-ruminants will help to achieve this. Therefore, the main objective of this work was the generation of a large number of sequentially unique hybrid phytases by directed recombination of known phytase genes with the goal to achieve improved biochemical properties compared to the wild-type phytases used. The focus of this work was the biochemical and nutritional evaluation of the newly generated hybrid phytases with respect to their suitability as feed supplements. All hybrid phytases examined showed more efficient InsP6 degradation at pH 3.0 than at pH 5.5, although the phytase activity supplemented was the same at both pH values. While InsP6 was dephosphorylated to InsP1-2 in many cases at pH 3.0, accumulation of the Ins(1,2,5,6)P4 isomer occurred at pH 5.5. In an in vitro model simulating the digestive tract of broilers, hybrid phytases with high sequential homology to the E. coli and C. braakii phytase showed high accumulation of InsP4 isomers. Interestingly, these phytases preferentially formed the Ins(1,2,5,6)P4 isomer. In contrast, other hybrid phytases were able to degrade all InsP4 isomers and in some cases high InsP2 concentrations were observed. Another in vitro experiment with a complex feed matrix consisting of soybean meal, rapeseed meal, and wheat with a high mineral content, illustrated the negative influence of certain feed-related factors on phytase efficiency. InsP6 present in the feed was significantly less degraded by all phytases used compared to a corn and soy-based feed matrix with a low mineral content. While a hybrid phytase was able to completely dephosphorylate the InsP6 of the corn and soy-based feed matrix down to the InsP3 isomer resulting in high InsP2 concentrations, the InsP6 were still detectable in the in vitro model with the more complex feed matrix and high mineral content, despite identical reaction conditions. In a final feeding trial with broilers, one of the hybrid phytases was supplemented at two doses each (500 and 1500 FTU/kg) to evaluate its suitability as a feed supplement. Also, a commercial phytase was included in the study design at the same doses setting the benchmark for phytase efficiency. A low phosphorus experimental feed based on corn and soybean meal was used. The supplementation of the used hybrid phytase resulted in a dose-dependent increase in broiler performance data such as daily weight gain, feed intake and significantly improved feed efficiency compared to the basal ration without enzyme supplementation. In addition, foot ash content was increased by 21.6% at a dose of 1500 FTU/kg phytase, indicating significantly improved bone mineralization due to the released InsP6 phosphate. By analyzing InsP6 concentration and its degradation products in different segments of the digestive tract, efficient InsP6 degradation was observed. In contrast to the in vitro experiments, no accumulation of InsP3-4 isomers could be detected in crop, gizzard or small intestine. In addition to a high exogenous phytase activity, this result also suggests a high endogenous phytase as well as phosphatase activity in the digestive tract of broilers. It can be assumed that the absence of monocalcium phosphate in the experimental rations may have induced the expression of endogenous phytases and phosphatases. This assumption is confirmed by the high precaecal InsP6 degradation, which was 63.5% in the basal ration without phytase supplementation. Nevertheless, the used hybrid phytase significantly increased the precaecal InsP6 degradation to 76.3%. The high phytase efficiency was also reflected in the measured precaecal phosphorus digestibility, which was increased by 6.8% compared to the basal ration. The commercial phytase used showed comparable improvement in broiler performance data to the non optimized hybrid phytase. This project demonstrated the development of a variety of sequentially unique hybrid phytases by recombination of known phytase genes, which exceeded the biochemical properties of the wild-type phytases in some relevant aspects. Some of the phytases showed very efficient phytate degradation when simulating the digestive tract of broilers in vitro. Also, the suitability of the tested hybrid phytase as feed additives was demonstrated by the increased performance data of broilers. The higher performance data of the broilers could be attributed to efficient phytate degradation. To achieve maximum InsP6 degradation in vivo, the feed-related and animal-related factors on phytase efficiency need to be better understood.
Evaluation of fresh and preserved sheep faeces as an inoculum source in in vitro gas production assays
(2025) Rippstein, Lena; Rodehutscord, Markus
In order to meet the animals’ requirements of energy and nutrients, knowledge of the feed value of individual feed components is essential. In this context, information on ruminal degradability of feeds is crucial for formulating rations for ruminants. Information of this kind can be obtained using in vitro methods, such as the Hohenheim gas test (HGT). This method allows for the estimation of the organic matter digestibility and the energy value, as well as the protein value of ruminant feeds when applying the extended HGT (eHGT). In vitro methods provide a cost-efficient, rapid, and standardisable alternative to in situ and in vivo approaches, while contributing to reducing animal burden and the number of experimental animals. However, the HGT currently depends on using rumen fluid, which is commonly obtained from rumen-cannulated animals. Due to ethical concerns related to animal welfare and practical considerations, there is growing interest in replacing rumen-cannulated animals for the in vitro feed evaluation in the HGT and eHGT systems. However, the use of individual enzymes or enzyme mixtures for in vitro evaluation of feeds has not yet proven suitable for adequately representing the complex microbial activity of a rumen fluid inoculum (RI). In contrast, several findings in the literature indicate the potential of faeces as an alternative inoculum source to rumen fluid. However, to date, this approach has not been established in the routine analysis of ruminant feeds. The overarching aim of the present thesis was to systematically evaluate the suitability of sheep faeces as an alternative inoculum source to rumen fluid in the HGT. To this end, in vitro gas production (GP) was compared between faecal inoculum (FI) and RI. Additionally, the potential of using preserved faeces as an alternative inoculum source to fresh faeces, as well as the application of FI in the eHGT system, was evaluated. To compare faeces and rumen fluid as inoculum sources in the HGT and eHGT, RI was prepared according to the standard procedure using rumen fluid obtained from two rumen-cannulated lactating dairy cows and FI was prepared from rectally collected faeces of three adult wether sheep. The objective of Manuscript 1 was to examine whether FI and RI generally follow similar GP kinetics and to assess whether feed-specific variation could be observed. Furthermore, there was considerable interest in determining whether FI-GP and RI-GP are related to each other, as this could provide the basis for the future applicability of FI. A total of 90 currently relevant ruminant feeds from various categories and differing nutrient compositions were incubated in vitro with both FI and RI for 72 h, with multiple readings in the HGT. By using FI, lower GP kinetics were observed across all feed categories compared to RI. On average of all feeds, the potential GP was 9 mL/200 mg dry matter (DM) lower and the GP rate was 3.1%/h lower with FI than RI. Additionally, a lag phase of 1.51 h was estimated with FI, whereas no lag phase was observed for RI. The results indicate an overall lower fermentation activity of FI compared to RI. Despite these differences, the GP kinetic curves of the two inocula exhibited a very similar progression. Moreover, strong linear relationships were found between RI-GP at 24 h, the common incubation time of RI in the HGT, and FI-GP at both 24 h (Slope = 1.02, R² = 0.97) and 48 h (Slope = 1.1, R² = 0.97). Additionally, within the scope of this thesis, linear regression analyses were conducted based on a combined dataset from Manuscript 1 and previous studies. By using data from more than 400 different feeds in these analyses, the strong linear relationships between RI-GP at 24 h and FI-GP at 24 h (Slope = 0.98, R² = 0.93) as well as 48 h (Slope = 1.02, R² = 0.96) were confirmed. Dividing the dataset into the feed categories roughages and concentrates for the calculation of separate regression equations did not provide a clear advantage over using a single equation for all feeds. Despite the lower GP observed with FI, a consistent relationship was evident between the GP of both inocula across the different feeds, enabling a reliable estimation of RI-GP from FI-GP in the HGT. The use of preserved instead of fresh faeces would allow for a centralised housing of donor sheep, thereby reducing the number of animals required and improving standardisation. The aim of Manuscript 2 was to investigate the effect of differently preserved sheep faeces on the in vitro GP of nine different feeds and the microbiome in the HGT, intending to maintain a high level of microbial activity during incubations. Seven different freezing and freeze-drying treatments were applied. On average across all feeds, the potential GP of the frozen treatments (61 mL/200 mg DM) was comparable to that of the fresh faeces (62 mL/200 mg DM), whereas the freeze-dried treatments accounted for only 71-85% of the fresh faecal value. The results were confirmed by metaproteome analyses, as the microbiomes of the fresh and frozen treatments were significantly different from that of the freeze-dried treatments based on the relative abundance of the core proteins (p < 0.001). This demonstrated that stress factors associated with the freeze-drying process significantly impaired the microbiome, consequently affecting fermentation activity and GP. By contrast, the freezing process appeared more gentle on the microbiome, preserving a high microbial activity. Furthermore, strong relationships were found between RI-GP at 24 h and GP of the frozen treatments at 48 h of incubation (Slope = 1.27, R² = 0.96). Additionally, the effect of storage on freeze-dried and frozen treatments was investigated, revealing a considerable negative impact on GP and its relationship with RI-GP for both treatments. This limits the high potential for estimating RI-GP, which was particularly demonstrated with frozen faeces, and therefore requires further research. Manuscript 3 aimed to investigate the potential suitability of FI in the eHGT for estimating the protein value of ruminant feeds. The eHGT is used to estimate ruminally undegradable crude protein (RUP) and microbial crude protein. Ammonia-nitrogen (NH3-N) is a key parameter in this context, as it is released during microbial crude protein degradation and provides a nitrogen source for the microbes. FI and RI were therefore compared based on NH3-N and calculated microbially bound nitrogen (mN) following in vitro incubation of six different feeds for 8, 24, and 48 h. The NH3 N content was 17 and 23% lower with FI than with RI after 24 and 48 h, respectively. With RI, mN values decreased over the incubation time for most feeds, whereas with FI, mN initially increased before declining at later incubation times. This suggests that crude protein degradation and microbial binding of nitrogen occur more slowly and to a lesser extent with FI. However, both inocula demonstrated a comparable response to an additional energy source and showed strong linear relationships for NH3-N, particularly after 24 h (Slope = 1.39, R² = 0.98), indicating similar microbial mechanisms in faeces and rumen fluid. The RUP was also estimated for both inocula in this thesis, and the results showed an inconsistent ratio between FI and RI incubations across the six feeds. Similarly, an inconsistent ratio between the two inocula was observed for the mN data. Therefore, further studies involving a larger number of feeds, as well as the testing of mathematical approaches, are necessary to better evaluate the suitability of FI for estimating the protein value of ruminant feeds with the eHGT. In conclusion, the use of sheep faeces as an inoculum source for the in vitro analysis of ruminant feeds can be considered suitable for replacing rumen fluid and thus rumen-cannulated animals in the HGT. By reliably predicting the RI-GP from FI-GP, the predicted RI-GP can be used in the official and validated equations to estimate organic matter digestibility and metabolisable energy of ruminant feeds. The methodological approach applied in this thesis, including sheep feeding, faeces collection, and inoculum preparation, appeared appropriate in achieving a high and consistent microbial activity in the FI. Furthermore, a high potential was demonstrated for using preserved, particularly frozen, sheep faeces for application in the HGT, as well as the use of FI for estimating the protein value of feeds in the eHGT. However, further investigations are required for the two application fields to assess the suitability of FI comprehensively.