Cross Inoculation of Rumen Fluid to Improve Dry Matter Disappearance and Its Effect on Bacterial Composition Using an in vitro Batch Culture Model

Abstract

Environmental pressures of ruminant production could be reduced by improving digestive efficiency. Previous in vivo attempts to manipulate the rumen microbial community have largely been unsuccessful probably due to the influencing effect of the host. Using an in vitro consecutive batch culture technique, the aim of this study was to determine whether manipulation was possible once the bacterial community was uncoupled from the host. Two cross inoculation experiments were performed. Rumen fluid was collected at time of slaughter from 11 Holstein-Friesian steers from the same herd for Experiment 1, and in Experiment 2 were collected from 11 Charolais cross steers sired by the same bull and raised on a forage only diet on the same farm from birth. The two fluids that differed most in their in vitro dry matter disappearance (IVDMD; "Good," "Bad") were selected for their respective experiment. The fluids were also mixed (1:1, "Mix") and used to inoculate the model. In Experiment 1, the mixed rumen fluid resulted in an IVDMD midway between that of the two rumen fluids from which it was made for the first 24 h batch culture (34, 29, 20 g per 100 g DM for the Good, Mix, and Bad, respectively, P < 0.001) which was reflected in fermentation parameters recorded. No effect of cross inoculation was seen for Experiment 2, where the Mix performed most similarly to the Bad. In both experiments, IVDMD increased with consecutive culturing as the microbial population adapted to the in vitro conditions and differences between the fluids were lost. The improved performance with each consecutive batch culture was associated with reduced bacterial diversity. Increases in the genus Pseudobutyrivibrio were identified, which may be, at least in part, responsible for the improved digestive efficiency observed, whilst Prevotella declined by 50% over the study period. It is likely that along with host factors, there are individual factors within each community that prevent other microbes from establishing. Whilst we were unable to manipulate the bacterial community, uncoupling the microbiota from the host resulted in changes in the community, becoming less diverse with time, likely due to environmental heterogeneity, and more efficient at digesting DM.

EFFECT OF NITRO COMPOUNDS ON in vitro RUMINAL METHANE, CARBON DIOXIDE, HYDROGEN AND DRY MATTER DEGRADABILITY

“…An effect of the consecutive inoculation was observed for the control tubes and those supplemented with NPA, where DMD decreases from batch A to C, following the same pattern as TG, showing the lowest digestibility for all tubes during the B series. These results differ from those of McDermott et al (2020), who indicated that digestibility of dry matter on ruminal cultures tends to increase with consecutive culturing, due to a microbial adaptation under in vitro conditions, associated with reduced diversity of the microbiome. Nevertheless, Lin et al (2019) demonstrated that adaptation to in vitro conditions may take a long time, and that the transfer-to-transfer process has an impact on fermentative parameters, including gas produced, pH, and short-chain fatty acids produced, which may cause variability in dry matter degradability of our experiment.…”

Effect of Ethyl Nitroacetate, Ethyl 2-Nitropropionate and 3-Nitropropionic Acid on Ruminal Fermentation Characteristics and Dry Matter Degradability under <em>In Vitro</em> Conditions

“…On the other side, an effect of the consecutive inoculation was observed for the control tubes and those supplemented with NPA, where DMD decreases from series A to C, following the same tendency that TG, showing the lowest digestibility for all tubes during the B series. These results differ from others (McDermott et al, 2020), who indicate that the digestibility of dry matter on ruminal cultures tends to increase with consecutive culturing, due to a microbial adaptation under in vitro conditions, associated with reduced diversity of the microbial ecosystem. Nevertheless, Lin et al, (2019) demonstrate that the adaptation to in vitro conditions may take a long time, and the transfer-to-transfer process has an impact on fermentative parameters, including gas produced, pH, and short chain fatty acids SCFA produced, which may cause variability in dry matter degradability of our experiment.…”

In vitro Inoculation of Fresh or Frozen Rumen Fluid Distinguishes Contrasting Microbial Communities and Fermentation Induced by Increasing Forage to Concentrate Ratio

“…Fresh rumen fluid is commonly employed as INO for in vitro fermentation. Increasing evidences show that frozen rumen fluid can be also used as INO. Frozen rumen fluid as INO can easily preserve rumen microorganisms, and allows multiple experiments to be carried out on the same fluids to reduce interexperimental variation (8,9). Although strong correlations of DM degradation have been observed between fresh and frozen INO (10), evidence also indicates that frozen INO can alter the fermentation pattern with decreased VFA production and shift of fermentation to favor propionate production (11).…”