• Microbiology and Biotechnology Letters
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• v.16 no.2
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• pp.85-91
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• 1988

Energy efficiency of bacterial cell mass and product formation from cellulose using Ruminococcus albus and Ruminococcus flavefaciens with application of available electron balance were discussed. Values of true growth yield, η$_{max}$ and η$^{max}_{th}$ and maintenance coefficient, m$_{e}$, were estimated us-ing experimental data, and the results were compared with estimates obtained from theoretical ap-proach. Experimental values were similar in magnitude to theoretical values in $Y^{max}_{ATP}$= 10.5 g cells/ mole ATP. Therefore, $Y^{max}_{ATP}$ values of Ruminococcus albus and Ruminocoecus flavefaciens were considered similar to 10.5 g cells/mole ATP.

• Journal of Animal Science and Technology
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• v.51 no.5
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• pp.379-386
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• 2009

The aim of this study was tofind out the effects of supplementation of vitamin A to the diets of high or low amounts of concentrates for ruminants. In the first experiment, ruminal fermentation patterns with the data of pH, VFA production and cellulose disappearance rates in the rumen in vitro were investigated. In the second experiment, enzyme activities, gas production and dry matter degradabilities using cellulolytic bacteria, Ruminococcus flavefaciens were investigated. Ruminal pH was higher in low amounts of concentrates than in high amounts of concentrates as expected, however, no significant differences were found. Cellulose disappearance rates improved in vitamin A addition particularly in early incubation time (before 24h) and also the production of volatile fatty acids increased in vitamin A addition. These trends were more evident in diets containing high amounts of concentrates than in low amounts of concentrates and it may indicate that vitamin A is more required in the diets of high amounts of concentrates. In the second experiment, gas production, enzyme activities and dry matter degradabilities using cellulolytic bacteria, Ruminococcus flavefaciens were not different between vitamin A added and non-added diets. Ruminococcus flavefaciens may not require additional vitamin A for its own growth.

• Korean Journal of Microbiology
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• v.34 no.3
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• pp.91-95
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• 1998

Cellulase producing microorganisms, GPC-1, GPC-2, GNR-1 GNR-2, and GNR-3, were screened from the Rumen fluid of Korean Native Cattle. Isolated GPC-1 and GPC-2 were identified as Ruminococcus sp. according to results of the Gram stain and anaerobic characteristics. Based on morphological and physicochemical identification, the isolate GPC-1 and GPC-2 were identified as strains of Ruminococcus albus and Ruminococcus flavefaciens, respectively. Isolated GNR-1 GNR-2 and GNR-3 were identified as Bacteroides sp., Butyrivibrio sp. and Clostridium sp. according to results of the Gram stain, $H_2S$ producition and spore formation, respectively. Based on morphological and physicochemical identification, the isolate GNR-1 GNR-2 and GNR-3 were identified as strains of Bacteroides succinogenes, Butyrivibrio fibrisolvens and Clostridium cellobioparum, respectively.

• Asian-Australasian Journal of Animal Sciences
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• v.31 no.1
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• pp.54-62
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• 2018

Objective: Due to the threat of global warming, the livestock industry is increasingly interested in exploring how feed additives may reduce anthropogenic greenhouse gas emissions, especially from ruminants. This study investigated the effect of Rhodophyta supplemented bovine diets on in vitro rumen fermentation and rumen microbial diversity. Methods: Cannulated Holstein cows were used as rumen fluid donors. Rumen fluid:buffer (1:2; 15 mL) solution was incubated for up to 72 h in six treatments: a control (timothy hay only), along with substrates containing 5% extracts from five Rhodophyta species (Grateloupia lanceolata [Okamura] Kawaguchi, Hypnea japonica Tanaka, Pterocladia capillacea [Gmelin] Bornet, Chondria crassicaulis Harvey, or Gelidium amansii [Lam.] Lamouroux). Results: Compared with control, Rhodophyta extracts increased cumulative gas production after 24 and 72 h (p = 0.0297 and p = 0.0047). The extracts reduced methane emission at 12 and 24 h (p<0.05). In particular, real-time polymerase chain reaction analysis indicated that at 24 h, ciliate-associated methanogens, Ruminococcus albus and Ruminococcus flavefaciens decreased at 24 h (p = 0.0002, p<0.0001, and p<0.0001), while Fibrobacter succinogenes (F. succinogenes) increased (p = 0.0004). Additionally, Rhodophyta extracts improved acetate concentration at 12 and 24 h (p = 0.0766 and p = 0.0132), as well as acetate/propionate (A/P) ratio at 6 and 12 h (p = 0.0106 and p = 0.0278). Conclusion: Rhodophyta extracts are a viable additive that can improve ruminant growth performance (higher total gas production, lower A/P ratio) and methane abatement (less ciliateassociated methanogens, Ruminococcus albus and Ruminococcus flavefaciens and more F. succinogenes.

• Asian-Australasian Journal of Animal Sciences
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• v.12 no.5
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• pp.708-714
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• 1999

Fermentation of legume fodder tree leaves by rumen microorganisms was evaluated. The substrates were sun-dried, ground leaves. Gas and volatile fatty acid (VFAs) production were estimated. Using gas production as an index of fermentation at 12 h, the leaves tested ranked as follows; Chamaecytisus palmensis>Gliricidia sepium>Sebania sesban>Tephrosia bracteolate>Leucaena pallida>Vernonia amygdalina>Acacia sieberiana>Sesbania goetzei>Acacia angustissima. Using VFA production, the ranking was a follows; G. sepium>S. sesban>S. goetzei>L. pallida>C. palmensis/V. amygdalina>T. bracteolate> A. sieberiana>A. angustissima. Absolute gas or VFA production rates, were also used to rank the leaves. Extracts (70% acetone) of A. angustissima inhibited the growth of Ruminococcus albus 8, R. flavefaciens FD-1, Prevotella ruminicola D3ID and Streptococcus bovis JBI while the trowth of Selenomonas ruminantium D was depressed when 0.6 ml exracts were added. C. palmensis water extracts enhanced cellulose hydrolysis by R. flavefaciens FD-1. All extracts reduced celluloysis by R. albus 8. R. flavefaciens FD-1 hydrolyzed more (p<0.001) cellulose than R. albus 8.

• Asian-Australasian Journal of Animal Sciences
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• v.14 no.6
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• pp.880-884
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• 2001

Rumen microbiology research has undergone several evolutionary steps: the isolation and nutritional characterization of readily cultivated microbes; followed by the cloning and sequence analysis of individual genes relevant to key digestive processes; through to the use of small subunit ribosomal RNA (SSU rRNA) sequences for a cultivation-independent examination of microbial diversity. Our knowledge of rumen microbiology has expanded as a result, but the translation of this information into productive alterations of ruminal function has been rather limited. For instance, the cloning and characterization of cellulase genes in Escherichia coli has yielded some valuable information about this complex enzyme system in ruminal bacteria. SSU rRNA analyses have also confirmed that a considerable amount of the microbial diversity in the rumen is not represented in existing culture collections. However, we still have little idea of whether the key, and potentially rate-limiting, gene products and (or) microbial interactions have been identified. Technologies allowing high throughput nucleotide and protein sequence analysis have led to the emergence of two new fields of investigation, genomics and proteomics. Both disciplines can be further subdivided into functional and comparative lines of investigation. The massive accumulation of microbial DNA and protein sequence data, including complete genome sequences, is revolutionizing the way we examine microbial physiology and diversity. We describe here some examples of our use of genomics- and proteomics-based methods, to analyze the cellulase system of Ruminococcus flavefaciens FD-1 and explore the genome of Ruminococcus albus 8. At Illinois, we are using bacterial artificial chromosome (BAC) vectors to create libraries containing large (>75 kbases), contiguous segments of DNA from R. flavefaciens FD-1. Considering that every bacterium is not a candidate for whole genome sequencing, BAC libraries offer an attractive, alternative method to perform physical and functional analyses of a bacterium's genome. Our first plan is to use these BAC clones to determine whether or not cellulases and accessory genes in R. flavefaciens exist in clusters of orthologous genes (COGs). Proteomics is also being used to complement the BAC library/DNA sequencing approach. Proteins differentially expressed in response to carbon source are being identified by 2-D SDS-PAGE, followed by in-gel-digests and peptide mass mapping by MALDI-TOF Mass Spectrometry, as well as peptide sequencing by Edman degradation. At Ohio State, we have used a combination of functional proteomics, mutational analysis and differential display RT-PCR to obtain evidence suggesting that in addition to a cellulosome-like mechanism, R. albus 8 possesses other mechanisms for adhesion to plant surfaces. Genome walking on either side of these differentially expressed transcripts has also resulted in two interesting observations: i) a relatively large number of genes with no matches in the current databases and; ii) the identification of genes with a high level of sequence identity to those identified, until now, in the archaebacteria. Genomics and proteomics will also accelerate our understanding of microbial interactions, and allow a greater degree of in situ analyses in the future. The challenge is to utilize genomics and proteomics to improve our fundamental understanding of microbial physiology, diversity and ecology, and overcome constraints to ruminal function.

• Korean Journal of Organic Agriculture
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• v.20 no.3
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• pp.327-343
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• 2012

In order to develop a high cellulolytic direct-fed microorganism (DFM) for ruminant productivity improvement, this study isolated cellulolytic bacteria from the rumen of Holstein dairy cows, and compared their cellulolytic abilities via DM degradability, gas production and cellulolytic enzyme activities. Twenty six bacteria were isolated from colonies grown in Dehority's artificial (DA) medium with 2% agar and cultured in DA medium containing filter paper at $39^{\circ}C$ for 24h. 16s rDNA gene sequencing of four strains from isolated bacteria showed that H8, H20 and H25 strains identified as Ruminococcus flavefaciens, and H23 strain identified as Fibrobacter succinogenes. H20 strain had higher degradability of filter paper compared with others during the incubation. H8 (R. flavefaciens), H20 (R. flavefaciens), H23 (F. succinogenes), H25 (R. flavefaciens) and RF (R. flavefaciens sijpesteijn, ATCC 19208) were cultured in DA medium with filter paper as a single carbon source for 0, 1, 2, 3, 4 and 6 days without shaking at $39^{\circ}C$, respectively. Dry matter degradability rates of H20, H23 and H25 were relatively higher than those of H8 and RF since 2 d incubation. The cumulative gas production of isolated cellulolytic bacteria increased with incubation time. At every incubation time, the gas production was highest in H20 strain. The activities of carboxymethylcellulase (CMCase) and Avicelase in the culture supernatant were significantly higher in H20 strain compared with others at every incubation time (p<0.05). Therefore, although further researches are required, the present results suggest that H20 strain could be a candidate of DFM in animal feed due to high cellulolytic ability.

• Asian-Australasian Journal of Animal Sciences
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• v.14 no.6
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• pp.797-802
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• 2001

Two strains of rumen fungi (Piromyces rhizinflata B157, Orpinomyces joyonii SG4) and three strains of rumen cellulolytic bacteria (Ruminococcus albus B199, Ruminococcus flavefaciens FD1 and Fibrobacter succinogenes S85) were used as mono-cultures or combinationally arranged as co- and sequential-cultures to assess the relative contributions and interactions between rumen fungi and cellulolytic bacteria on rice straw degradation. The rates of dry matter degradation of co-cultures were similar to those of corresponding bacterial mono-cultures. Compared to corresponding sequential-cultures, the degradation of rice straw was reduced in all co-cultures (P<0.01). Regardless of the microbial species, the cellulolytic bacteria seemed to inhibit the degradation of rice straw by rumen fungi. The high efficiency of fungal cellulolysis seems to affect bacterial degradation rates.

• Asian-Australasian Journal of Animal Sciences
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• v.20 no.2
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• pp.200-207
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• 2007

In vitro rumen incubation studies were conducted to determine effects of initial pH on bacterial attachment and fiber digestion. Ruminal fluid pH was adjusted to 5.7, 6.2 and 6.7, and three major fibrolytic bacteria attached to rice straw in the mixed culture were quantified with real-time PCR. The numbers of attached and unattached Fibrobacter succinogenes, Ruminococcus flavefaciens and Ruminocococcus albus were lower (p<0.05) at initial pH of 5.7 without significant difference between those at higher initial pH. Lowering incubation media pH to 5.7 also increased bacterial numbers detached from substrate regardless of bacterial species. Dry matter digestibility, gas accumulation and total VFA production were pH-dependent. Unlike bacterial attachment, maintaining an initial pH of 6.7 increased digestion over initial pH of 6.2. After 48 h in vitro rumen fermentation, average increases in DM digestion, gas accumulation, and total VFA production at initial pH of 6.2 and 6.7 were 2.8 and 4.4, 2.0 and 3.0, and 1.2 and 1.6 times those at initial pH of 5.7, respectively. The lag time to reach above 2% DM digestibility at low initial pH was taken more times (8 h) than at high and middle initial pH (4 h). Current data clearly indicate that ruminal pH is one of the important determinants of fiber digestion, which is modulated via the effect on bacterial attachment to fiber substrates.

• Asian-Australasian Journal of Animal Sciences
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• v.21 no.6
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• pp.818-823
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• 2008

The interaction of fibre degrading microbes and methanogens was studied using two forages, lucerne (Medicago sativa) hay and maize (Zea mays) hay, as substrate and 2-bromoethanesulphonic acid (BES) as an additive in an in vitro gas production test. Gas and methane production (ml/g dry matter) were significantly higher (p<0.05) on lucerne as compared to maize hay. Inclusion of BES in the incubation medium significantly suppressed methane emission irrespective of substrate. The population density of total bacteria, fungi, Ruminococcus flavefaciens and Fibrobacter succinogenes was higher, whereas that of methanogens was lower with maize hay as compared to lucerne as substrate. BES suppressed methanogen population by 7 fold on lucerene and by 8.5 fold on maize at 24 h incubation as estimated by real time-PCR. This suppression was accompanied by almost complete (>98% of control) inhibition of methanogenesis. The proportion of acetate decreased, whereas that of propionate increased significantly by inclusion of BES, resulting in narrowing of acetate to propionate ratio. In vitro true digestibility (IVTD) of lucerne was significantly higher as compared to maize but BES inclusion did not affect IVTD.