Browsing by Subject "Rumen microbial community"

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    Comparison of plant cell wall degrading community in the rumen of N’Dama and N’Dama x Jersey crossbred cattle in relation to in vivo and in vitro cell wall degradation
    (2004) Nouala-Fonkou, Simplice; Becker, Klaus
    This thesis presents a unique combination of an in vivo feeding trial, the analysis of the microbial community structure in the rumen, and in vitro fermentation studies, in order to assess the impact of breeds and diets on animal performance in a West African production setting. Pure N?Dama and N?Dama x Jersey crossbred cattle were fed two basal diets, baby corn and groundnut hay, supplemented with graded levels of either conventional concentrate or moringa leaf meal, to compare animal responses in productivity. In this context, Moringa oleifera leaf meal constitutes a locally available, potential alternative to commercial concentrate for cattle production. The cell wall digesting community of N?Dama and its crosses was analysed using phylogenetically based hybridisation probes to account for the contribution of rumen microbes to differences in fermentation patterns and animal response. In vitro fermentation studies were carried out using the same diets and supplementation levels as fed in vivo, to test the accuracy of the in vitro gas production technique in predicting the optimum level of supplementation. The in vivo feeding experiment focussed on the comparison of breed performance with diets relevant for local production conditions. Six N?Dama and six N?Dama x Jersey (crossbred) animals were used in a cross over design. They were fed consecutively three combinations of roughage and supplement, baby corn stover and concentrate (BCS:Co), groundnut hay and concentrate (GNH:Co) and groundnut hay and moringa meal (GNH:Mo), each at 5 levels of supplementation (0, 10, 20, 30 and 40%). Results from this study showed that there was a clear difference in animal response to different feeding regimes between the two breeds. When averaged over all diets organic matter intake (OMI) was higher in crossbred compared to N?Dama (94 and 87.6 g/kg 0.75 d-1, respectively). When analyzed for the diets and averaged over the breeds OMI was higher when animals were fed the baby corn based diet compared to groundnut based diets (95 against 88 g/kg 0.75 d-1). Only when the diet consisted of BCS:Co, and at low levels of supplementation, N?Dama ingested more than crossbred, but the difference was not significant. With GNH:Co crossbred ingested significantly more at levels of supplementation less or equal to 20%. With GNH:Mo crossbred ingested more, whatever the level of supplementation. The optimum level of supplementation in vivo, estimated with the single slope broken line model, was 10% and 20% for both breeds when they were fed BCS:Co and GNH:Co respectively, but 30% for N?Dama and 10% for the crossbreds when animals were fed GNH:Mo. Organic matter digestibility (OMD) was higher in N?Dama (64.6% against 60.7% in crossbreds) when animals were fed BCS:Co and supplementation had no effect on OMD of BCS:Co whatever the breed. When animals were fed groundnut hay as basal diet, OMD was also significantly higher in N?Dama at low levels of supplementation, but the differences became insignificant beyond 10% and 20% of concentrate or moringa, respectively. With GNH:Co OMD showed a quadratic response (p<0.001) with increasing level of supplementation when it was fed to crossbreds and was not affected when it was fed to N?Dama. Increasing levels of moringa meal supplementation increased OMD in both breeds up to a peak at 20 and 30% for N?Dama and crossbred, respectively. Average daily weight gain (ADG) was not affected by the breed, however it was higher on BCS:Co compared to other diets. On BCS:Co ADG increased with the level of supplementation, reaching a peak at 30%, whereas supplementation had no effect on ADG when animals were fed groundnut hay based diets. As N?Dama could take in and digest more of the low quality BCS:Co diet, they were less efficient in feed conversion under this feeding regime (FCE: 14 vs. 9 for the crossbreds). On GNH based diets, however, N?Dama surpassed the crossbreds in feed conversion efficiency with ratios of 11 vs. 13 for GNH:Co and 9 vs. 27 on GNH:Mo. Rumen microbes play the key role for the digestibility of a given feed and thus also for feed intake and finally animal performance. Obviously, the community composition and activity is highly dependent on the diet. With the present set-up, however, with identical external conditions and three different, well defined diets fed to both, N?Dama and crossbred cattle, a comparison of the microbial community structure between breeds could be attempted. The in vivo and in vitro data taken in the other parts of the study allow a sensible interpretation of potential changes in microbial composition. Rumen fluid was collected from three fistulated N?Dama and three crossbred animals adapted to the experimental diets at medium supplementation level. The cell wall degrading community was analyzed using the phylogenetically based 16S rRNA hybridisation probes. The results showed that on BCS:Co diet the Fibrobacter and R. flavefaciens RNA concentrations were higher in rumen fluid of N?Dama compared to crossbred. These concentrations were also significantly affected by the diet, such that they were higher on baby corn stover compared to groundnut hay based diets. The results of the microbial community analysis suggested that the differences between breeds observed in digestibility could be partially explained by the composition of the cell wall degrading community. Parallel to the in vivo experiment, in vitro fermentation studies were undertaken to evaluate the predictability of the in vivo response to supplementation by the in vitro data. Rumen fluid from 3 N?Dama and 3 crossbred donor animals was used for 24 hour in vitro fermentations. The donor animals were fed consecutively the same three diets used in vivo (BCS:Co, GNH:Co and GNH:Mo) at 20% level of supplementation. Each of these inocula was incubated with in vitro substrates consisting of all the combinations tested in vivo (i.e. 3 diets, 5 levels of supplementation) plus supplement alone. This design should allow to analyze for both, the impact of donor breed as well as that of the donor diet and to conclude which factors may be varied while maintaining predictability. The breed of the donor animals did not significantly affect 24 hour gas production, but short chain fatty acid concentration was higher with rumen fluid from crossbreds when donors were fed BCS:Co and GNH:Mo. Moringa meal as supplement to donor animals changed the fermentation pattern of all the substrates, such that gas production and SCFA increased substantially in groundnut hay based substrates, whereas gas production of BCS:Co substrates decreased and SCFA did not substantially change. In vitro digestibility was higher with rumen fluid from N?Dama whatever the diet of donor animals and the substrate incubated. GNH:Mo as donor diet also increased IVTD of all the substrates. Even though there was no clear response in vivo, this indicates a general stimulation of microbial activity in the rumen and renders moringa leaf meal a promising supplement. Averaged over all data there was a positive correlation (r2=0.53 p<0.001) between IVTD and in vivo OMD. This correlation was much stronger when calculated for a specific diet (e.g. r2=0.90 p<0.001 for BCS:Co, averaged over the breeds). Analyzing the data for the individual breeds affected correlations only to a minor degree. Thus, when testing a supplementation strategy in vitro, it should be important that donor animals are fed the same components (roughage and supplement) that will be combined at different levels in vitro, whilst the breed of donor animals may be of second importance. This work provides conclusive evidence that in vitro incubations may be used to design supplementation strategies, thus reducing the need for in vivo experiments. Moringa leaf meal is a promising local resource to substitute for conventional concentrate. Differences in productivity between breeds could be correlated to (and may be partially manifested through) a divergent community structure of rumen microbes. That, in turn, indicates that animals of different breeds might have a ?genetic background? that favours the establishment of a certain community, even if the animals are kept under identical conditions. This relationship should be investigated by more advanced molecular techniques.
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    Studies on the composition of the ruminal microbial community using grass silage and corn silage
    (2016) Lengowski, Melanie; Rodehutscord, Markus
    Grass silage and corn silage are the most commonly used silages for feeding dairy cows and fattening bulls. Due to their different chemical composition these silages have diverse effects on the ruminal microbial community and, therefore, on the fermentation process in the rumen. The major objective of this thesis was to evaluate the effect of grass silage and corn silage on the ruminal microbial community composition. The focus was on the incubation of silages without using concentrates. Furthermore, diurnal changes of the ruminal microbial community were investigated. In the first study (Manuscript 1), effects of incubation of grass silage and corn silage on the ruminal microbial community were investigated using an established rumen simulation technique (Rusitec). Furthermore, diurnal changes and changes during the first 48 hours of incubation (adaption phase) were observed. A significant decrease of the complete ruminal microbial community in the fermenter liquids, on species and group level, was observed. During the adaption phase, the silage source lost relevance because in the silage-containing fermenters as well as in the blank fermenters decreasing numbers were observed with the exception of Clostridium aminophilum. For this species, after 48 hours higher numbers were found as compared to the inoculum. As has already been described in the literature, in the current thesis changes in microbial abundance were lesser in feed residues than in fermenter liquids. The abundance of protozoa in feed residues decreased during the first 48 hours while for Prevotella bryantii and C. aminophilum higher numbers were found after 24 and 48 hours as compared to the inoculum. The adaption in the Rusitec had not yet been fully completed after 48 hours, since for several species a different abundance was found on day 13 of incubation (Period 2). The provision of fresh substrate at the beginning of Period 2 led to an increase of almost all species and groups within the following two to four hours, with the exception of the protozoa and methanogens. The reduction of the protozoal population and methanogens could have been the result of migration from fermenter liquids to new feed particles and also the fact that some species of methanogens are associated to protozoa. Likewise, sampling of the feed residues after 24 and 48 hours was too late since the microorganisms could have been already detached. However, sampling of the feed residues at shorter intervals is not possible in the Rusitec. No effect of silage was observed on the abundance of total bacteria, methanogens and Selenomonas ruminantium. Methanogens and S. ruminantium do not use the substrate itself but rather fermentation end products of other ruminal microorganisms. Although no effect of silage was observed on the abundance of methanogens, more methane was produced during the incubation of grass silage than corn silage. This is in accordance with results described in literature that an abundance of methanogens is not associated with the production of methane. The corn silage promoted the abundance of protozoa and Ruminobacter aminophilus whereby the latter could only be detected during the first 48 hours of incubation. Quantification was not possible because of the generation of unspecific products during real-time qPCR. Fibrobacter succinogenes was also promoted by corn silage even though this is one of the most active cellulolytic species in the rumen. It could be possible that this species had an advantage in the degradation of the cell wall structures of C4 plants (which corn accounts to) as compared to other cellulolytic species. The grass silage promotes the abundance of the cellulolytic Ruminococcus albus as well as P. bryantii and C. aminophilum. The last two species use amino acids and proteins as energy sources thereby the latter belongs to hyper ammonia producing bacteria which could be the reason for the higher ammonia concentration in fermenter liquids during the incubation of grass silage compared to corn silage. In the second study (Manuscript 2), the effects of supplementing corn silage with different nitrogen sources on ruminal microbial community composition and ruminal microbial crude protein synthesis was investigated. Higher efficiency of microbial crude protein synthesis with corn silage has been reported from in vivo studies, while in vitro contradictory results were found. It was proposed that the different nitrogen content of the silages, and especially the lack of rumino-hepatic circulation in vitro, could be the reason for the lower efficiency of microbial crude protein synthesis when using corn silage. Different microbial species prefer different nitrogen sources as already observed in pure cultures and partially also in mixed cultures. Thus, in the second study, corn silage was supplemented with urea, pea protein, pea peptone or mixed free amino acids. With the supplementation of peptone and urea to corn silage, the highest efficiency of microbial crude protein synthesis was observed followed by amino acids and protein supplementation. But none of the used nitrogen sources allowed corn silage to achieve the level of microbial crude protein synthesis observed for grass silage. The protozoal population was negatively correlated with the efficiency of microbial crude protein synthesis which could have been a result of the higher energy requirement of protozoa compared to bacteria. Furthermore, protozoa engulfs and ingests bacteria and uses it as the main energy and protein source which could also have a negative effect on the efficiency of microbial crude protein synthesis. The effects of different nitrogen sources on the abundance of microbial groups and species not always were in accordance with reports in the literature. This could be attributed to the use of different substrate and pure cultures. In the third study (Manuscript 3), the effect of grass silage and corn silage on the ruminal microbial community composition in vivo was investigated. Three lactating Jersey cows fitted with a permanent cannula were fed with rations based on grass silage or corn silage. The rations contained the same amount of concentrate. In order to adjust the nitrogen content of the rations the corn silage based ration was supplemented with urea. The corn silage based ration promoted the abundance of total bacteria in the solid fraction obtained from the rumen, as well as the protozoal population in the solid and liquid fraction. It could be possible that the higher content of readily fermentable carbohydrates, mainly starch, of the corn silage in combination with the urea could have had a positive effect on both groups. Furthermore, a higher abundance of F. succinogenes was observed in animals fed with the corn silage-based ration. As expected, the cellulolytic bacteria R. albus and R. flavefaciens were enhanced by the grass silage-based ration as well as P. bryantii and R. amylophilus. The latter are known to use starch and maltose as energy source and to use their proteolytic activity only to achieve access to protein-coated starch. It is assumed that the combination of grass silage and concentrate could have had an influence on this species. For S. ruminantium, a feeding effect on the abundance was only observed in the liquid fraction while in the solid fraction no effect was found. There was also no observable effect of ration on the abundance of C. aminophilum, RCC and the methanogens. Changes of the ruminal microbial community over time were observed. But no consistent increase of all species and groups after the phase with the highest dry matter intake of fresh feed became obvious. In conclusion, the results of the current thesis showed that grass and corn silages differently affected the composition of the ruminal microbial community in the liquid and solid fractions of the rumen content. Furthermore, significant diurnal changes of the ruminal microbial community could be observed in vitro as well as in vivo. These results highlighted the need of repeated sampling during the day and suggest the consideration of both ruminal compartments, liquid and solid.