Ruminococcin C, a promising antibiotic produced by a human gut symbiont.

A major public health challenge today is the resurgence of microbial infections caused by multidrug-resistant strains. Consequently, novel antimicrobial molecules are actively sought for development. In this context, the human gut microbiome is an under-explored potential trove of valuable natural molecules, such as the ribosomally-synthesized and post-translationally modified peptides (RiPPs). The biological activity of the sactipeptide subclass of RiPPs remains under-characterized. Here, we characterize an antimicrobial sactipeptide, Ruminococcin C1, purified from the caecal contents of rats mono-associated with Ruminococcus gnavus E1, a human symbiont. Its heterologous expression and post-translational maturation involving a specific sactisynthase establish a thioether network, which creates a double-hairpin folding. This original structure confers activity against pathogenic Clostridia and multidrug-resistant strains but no toxicity towards eukaryotic cells. Therefore, the Ruminococcin C1 should be considered as a valuable candidate for drug development and its producer strain R. gnavus E1 as a relevant probiotic for gut health enhancement.

Antidepressants affect gut microbiota and Ruminococcus flavefaciens is able to abolish their effects on depressive-like behavior.

Accumulating evidence demonstrates that the gut microbiota affects brain function and behavior, including depressive behavior. Antidepressants are the main drugs used for treatment of depression. We hypothesized that antidepressant treatment could modify gut microbiota which can partially mediate their antidepressant effects. Mice were chronically treated with one of five antidepressants (fluoxetine, escitalopram, venlafaxine, duloxetine or desipramine), and gut microbiota was analyzed, using 16s rRNA gene sequencing. After characterization of differences in the microbiota, chosen bacterial species were supplemented to vehicle and antidepressant-treated mice, and depressive-like behavior was assessed to determine bacterial effects. RNA-seq analysis was performed to determine effects of bacterial treatment in the brain. Antidepressants reduced richness and increased beta diversity of gut bacteria, compared to controls. At the genus level, antidepressants reduced abundances of Ruminococcus, Adlercreutzia, and an unclassified Alphaproteobacteria. To examine implications of the dysregulated bacteria, we chose one of antidepressants (duloxetine) and investigated if its antidepressive effects can be attenuated by simultaneous treatment with Ruminococcus flavefaciens or Adlercreutzia equolifaciens. Supplementation with R. flavefaciens diminished duloxetine-induced decrease in depressive-like behavior, while A. equolifaciens had no such effect. R. flavefaciens treatment induced changes in cortical gene expression, up-regulating genes involved in mitochondrial oxidative phosphorylation, while down-regulating genes involved in neuronal plasticity. Our results demonstrate that various types of antidepressants alter gut microbiota composition, and further implicate a role for R. flavefaciens in alleviating depressive-like behavior. Moreover, R. flavefaciens affects gene networks in the brain, suggesting a mechanism for microbial regulation of antidepressant treatment efficiency.

Effect of Dietary Chestnut or Quebracho Tannin Supplementation on Microbial Community and Fatty Acid Profile in the Rumen of Dairy Ewes.

Ruminants derived products have a prominent role in diets and economy worldwide; therefore, the capability to control the rumen microbial ecosystem, for ameliorating their quality, is of fundamental importance in the livestock sector. The aim of this study was to evaluate the effect of dietary supplementation with chestnut and quebracho tannins on microbial community and fatty acid profile, in the rumen fluid of dairy ewes. Multivariate analysis of PCR-DGGE profiles of rumen microbial communities showed a correlation among the presence of chestnut or quebracho in the diet, the specific Butyrivibrio group DGGE profiles, the increase in 18:3 cis9, cis12, and cis15; 18:2 cis9 and cis12; 18:2 cis9 and trans11; 18:2 trans11 and cis15; and 18:1 trans11 content, and the decrease in 18:0 concentration. Phylogenetic analysis of DGGE band sequences revealed the presence of bacteria representatives related to the genera Hungatella, Ruminococcus, and Eubacterium and unclassified Lachnospiraceae family members, suggesting that these taxa could be affected by tannins presence in the diets. The results of this study showed that tannins from chestnut and quebracho can reduce the biohydrogenation of unsaturated fatty acids through changes in rumen microbial communities.

The effect of quercetin on genetic expression of the commensal gut microbes Bifidobacterium catenulatum, Enterococcus caccae and Ruminococcus gauvreauii.

Quercetin is one of the most abundant polyphenols found in fruits and vegetables. The ability of the gut microbiota to metabolize quercetin has been previously documented; however, the effect that quercetin may have on commensal gut microbes remains unclear. In the present study, the effects of quercetin on the commensal gut microbes Ruminococcus gauvreauii, Bifidobacterium catenulatum and Enterococcus caccae were determined through evaluation of growth patterns and cell morphology, and analysis of genetic expression profiles between quercetin treated and non-treated groups using Single Molecule RNA sequencing via Helicos technology. Results of this study revealed that phenotypically, quercetin did not prevent growth of Ruminococcus gauvreauii, mildly suppressed growth of Bifidobacterium catenulatum, and moderately inhibited growth of Enterococcus caccae. Genetic analysis revealed that in response to quercetin, Ruminococcus gauvreauii down regulated genes responsible for protein folding, purine synthesis and metabolism. Bifidobacterium catenulatum increased expression of the ABC transport pathway and decreased metabolic pathways and cell wall synthesis. Enterococcus caccae upregulated genes responsible for energy production and metabolism, and downregulated pathways of stress response, translation and sugar transport. For the first time, the effect of quercetin on the growth and genetic expression of three different commensal gut bacteria was documented. The data provides insight into the interactions between genetic regulation and growth. This is also a unique demonstration of how RNA single molecule sequencing can be used to study the gut microbiota.

Effects of Diets Supplemented with Ensiled Mulberry Leaves and Sun-Dried Mulberry Fruit Pomace on the Ruminal Bacterial and Archaeal Community Composition of Finishing Steers.

This study investigated the effects of ensiled mulberry leaves (EML) and sun-dried mulberry fruit pomace (SMFP) on the ruminal bacterial and archaeal community composition of finishing steers. Corn grain- and cotton meal-based concentrate was partially replaced with EML or SMFP. The diets had similar crude protein (CP), neutral detergent fiber (NDF), and metabolizable energy. Following the feeding trial, the steers were slaughtered and ruminal liquid samples were collected to study the ruminal microbiome. Extraction of DNA, amplification of the V4 region of the 16S rRNA gene, and Illumina MiSeq pyrosequencing were performed for each sample. Following sequence de-noising, chimera checking, and quality trimming, an average of 209,610 sequences were generated per sample. Quantitative real-time PCR was performed to examine the selected bacterial species in the rumen. Our results showed that the predominant phyla were Bacteroidetes (43.90%), Firmicutes (39.06%), Proteobacteria (4.31%), and Tenericutes (2.04%), and the predominant genera included Prevotella (13.82%), Ruminococcus (2.51%), Butyrivibrio (2.38%), and Succiniclasticum (2.26%). Compared to the control group, EML and SMFP groups had a higher abundance of total bacteria (p < 0.001); however, the bacterial community composition was similar among the three groups. At the phylum level, there were no significant differences in Firmicutes (p = 0.7932), Bacteroidetes (p = 0.2330), Tenericutes (p = 0.2811), or Proteobacteria (p = 0.0680) levels among the three groups; however, Fibrobacteres decreased in EML (p = 0.0431). At the genus level, there were no differences in Prevotella (p = 0.4280), Ruminococcus (p = 0.2639), Butyrivibrio (p = 0.4433), or Succiniclasticum (p = 0.0431) levels among the groups. Additionally, the dietary treatments had no significant effects on the archaeal community composition in the rumen. Therefore, EML and SMFP supplementation had no significant effects on the ruminal bacterial or archaeal community composition of finishing steers.

Rumen microbial abundance and fermentation profile during severe subacute ruminal acidosis and its modulation by plant derived alkaloids in vitro.

Rumen microbiota have important metabolic functions for the host animal. This study aimed at characterizing changes in rumen microbial abundances and fermentation profiles using a severe subacute ruminal acidosis (SARA) in vitro model, and to evaluate a potential modulatory role of plant derived alkaloids (PDA), containing quaternary benzophenanthridine and protopine alkaloids, of which sanguinarine and chelerythrine were the major bioactive compounds. Induction of severe SARA strongly affected the rumen microbial composition and fermentation variables without suppressing the abundance of total bacteria. Protozoa and fungi were more sensitive to the low ruminal pH condition than bacteria. Induction of severe SARA clearly depressed degradation of fiber (P < 0.001), which came along with a decreased relative abundance of fibrolytic Ruminococcus albus and Fibrobacter succinogenes (P < 0.001). Under severe SARA conditions, the genus Prevotella, Lactobacillus group, Megasphaera elsdenii, and Entodinium spp. (P < 0.001) were more abundant, whereas Ruminobacter amylophilus was less abundant. SARA largely suppressed methane formation (-70%, P < 0.001), although total methanogenic 16S rRNA gene abundance was not affected. According to principal component analysis, Methanobrevibacter spp. correlated to methane concentration. Addition of PDA modulated ruminal fermentation under normal conditions such as enhanced (P < 0.05) concentration of total SCFA, propionate and valerate, and increased (P < 0.05) degradation of crude protein compared with the unsupplemented control diet. Our results indicate strong shifts in the microbial community during severe SARA compared to normal conditions. Supplementation of PDA positively modulates ruminal fermentation under normal ruminal pH conditions.

The effect of fibre source on the numbers of some fibre-degrading bacteria of Arabian camel's (Camelus dromedarius) foregut origin.

The total bacterial community of Fibrobacter succinogenes and Ruminococcus flavefaciens in fibre-enriched culture of the foregut contents of 12 adult feral camels (Camelus dromedaries) fed on native vegetation in Australia was investigated using quantitative PCR. Foregut contents were collected postmortem, pooled and filtered before divided into two fractions. One fraction was used for extraction of DNA, while the other fraction was inoculated straight away into BM 10 contained filter paper (FP), cotton thread (CT) or neutral detergent fibre (NDF) as the sole carbohydrate sources in Hungate tubes. The tubes were incubated anaerobically at 39 °C for 1 week. After a near complete degradation of the FP and CT and extensive turbidity in the NDF, media subculturing was carried out into fresh media tubes. This was repeated twice before genomic DNA was extracted and used for quantification of bacteria. Using an absolute quantification method, the numbers of cells in 1 ml of each sample ranged from 4.07 × 10(6) to 2.73 × 10(9) for total bacteria, 1.34 × 10(3) to 2.17 × 10(5) for F. succinogenes and 5.78 × 10(1) to 3.53 × 10(4) for R. flavefaciens. The mean cell number of F. succinogenes was highest in the FP enrichment medium at approximately 107-fold, whereas for the R. flavefaciens targeted primer, the NDF enrichment media had the highest mean cell number at approximately 4-fold when compared to the rumen content. The data presented here provide evidence of fibre type preference by the two main fibre-degrading bacteria and would help us understand the interaction between fibre type and fibre-degrading microorganisms, which has ramification on camel nutrition at different seasons and environments.

Effects of extracts of Humulus lupulus (hops) and Yucca schidigera applied alone or in combination with monensin on rumen fermentation and microbial populations in vitro.

BACKGROUND: ß-Acids in hops (Humulus lupulus) and saponins in yucca (Yucca schidigera) have been found to possess antimicrobial properties similar to that of monensin and could be an alternative to in-feed antibiotics. The effects of monensin (MON) and ethanol extracts of hops (HE) and Y. schidigera (YE) alone and in combination with MON were assessed on ruminal microbial composition and fermentation in vitro of a barley-based diet. RESULTS: All treatments decreased (P < 0.05) CH4 production (per unit of dry matter), microbial protein (mg), and NH3 -N accumulation. All treatments reduced (P < 0.01) the acetate:propionate (A:P) ratio and molar proportions of butyrate, but increased (P < 0.01) those of propionate, whereas those of acetate decreased (P < 0.001) with addition of MON (10 µg mL(-1)) and combined with HE or YE. Methane produced per unit of true digested dry matter decreased (P < 0.001) with all treatments except YE. Monensin reduced (P < 0.001) proportions of 16S rRNA copies of Ruminococcus flavefaciens, but increased (P < 0.01) those of Selenomonas ruminantium. Hops extract alone or combined with MON reduced (P < 0.01) proportions of R. flavefaciens but combined with MON tended (P < 0.1) to increase those of S. ruminantium. Yucca extract combined with MON increased (P < 0.01) the proportions of R. flavefaciens and S. ruminantium. All treatments except MON (2.5 µg mL(-1)) reduced (P < 0.01) the relative abundance of methanogens. CONCLUSION: Hops extract and YE altered rumen microbes and fermentation in a manner similar to MON with many responses being additive when applied in combination.

Effects of quillaja and yucca saponins on communities and select populations of rumen bacteria and archaea, and fermentation in vitro.

AIMS: The objective of this study was to comprehensively evaluate quillaja (QSP) and yucca saponin (YSP) products with respect to their effects on diversity of rumen bacteria and archaea, abundance of selected microbes, and feed degradability and fermentation. METHODS AND RESULTS: Both QSP and YSP at doses 0-0.6 g l(-1) tended to increase degradability of feed substrate in in vitro rumen cultures, but to different extents. Neither one of the saponins affected the concentrations of ammonia, total volatile fatty acids, or molar proportion of acetate. However, QSP increased molar proportion of propionate and decreased that of butyrate, whereas YSP tended to decrease that of butyrate. As determined by qPCR, QSP and YSP did not affect the abundance of total bacteria or Ruminococcus albus. The QSP did not affect the abundances of Fibrobacter succinogenes or genus Prevotella, but tended to decrease that of Ruminococcus flavefaciens, whereas YSP significantly increased the abundance of R. flavefaciens and Prevotella, and numerically increased that of F. succinogenes. Both saponins increased archaeal abundance, although to small magnitudes (0.3-0.4 log). The protozoal populations were decreased significantly by QSP, but not by YSP. Based on DGGE and T-RFLP analysis, both saponins altered the bacterial community and species organization, but less so the archaeal community. CONCLUSIONS: This study demonstrated that saponins, although not effective in mitigating methane emission, may improve feed utilization at low doses, and modulate ruminal microbial communities in a dose-dependent manner. SIGNIFICANCE AND IMPACT OF THE STUDY: The results of this study suggest that saponins at low doses may directly stimulate the growth of some rumen bacteria including cellulolytic bacteria, thus improving digestibility of feeds, independent of their defaunation activity. In contrast, saponins at high doses modulate rumen fermentation characteristically similar to defaunation.

Effect of disodium fumarate on microbial abundance, ruminal fermentation and methane emission in goats under different forage: concentrate ratios.

This study investigated the effects of disodium fumarate (DF) on methane emission, ruminal fermentation and microbial abundance in goats under different forage (F) : concentrate (C) ratios and fed according to maintenance requirements. Four ruminally fistulated, castrated male goats were used in a 4 × 4 Latin square design with a 2 × 2 factorial arrangement of treatments and the main factors being the F : C ratios (41 : 59 or 58 : 42) and DF supplementation (0 or 10 g/day). DF reduced methane production (P < 0.05) on average by 11.9%, irrespective of the F : C ratio. The concentrations of total volatile fatty acids, acetate and propionate were greater in the rumen of goats supplemented with DF (P < 0.05), whereas the abundance of methanogens was lower (P < 0.05). In high-forage diets, the abundance of Selenomonas ruminantium, a fumarate-reducing bacterium, was greater in the rumen of goats supplemented with DF. The abundance of fungi, protozoa, Ruminococus flavefaciens and Fibrobacter succinogenes were not affected by the addition of DF. Variable F : C ratios affected the abundance of methanogens, fungi and R. flavefaciens (P < 0.05), but did not affect methane emission. The result implied that DF had a beneficial effect on the in vivo rumen fermentation of the goats fed diets with different F : C ratios and that this effect were not a direct action on anaerobic fungi, protozoa and fibrolytic bacteria, the generally recognized fiber-degrading and hydrogen-producing microorganisms, but due to the stimulation of fumarate-reducing bacteria and the depression of methanogens.

Exposure to Al2O3 nanoparticles changes the fatty acid profile of the anaerobe Ruminococcus flavefaciens.

One of the main mechanisms of nanoparticle toxicity is known to be the generation of reactive oxygen species (ROS) which primarily damage cell membranes. However, very limited data on membrane effects in anaerobic environments (where ROS could not be the cause of membrane damage) are available. In the following study, rumen anaerobe Ruminococcus flavefaciens 007C was used as a bacterial model to assess the potential effects of Al(2)O(3) and TiO(2) nanoparticles on membranes in an anaerobic environment. Fatty acid profiles of cultures after exposure to Al(2)O(3) or TiO(2) nanoparticles were analyzed and compared with the profiles of non-exposed cultures or cultures exposed to bulk materials. Analysis revealed dose-effect changes in membrane composition exclusively when cells were exposed to Al(2)O(3) nanoparticles in a concentration range of 3-5 g/L, but were not present in cultures exposed to bulk material. On the other hand, the tested concentrations of nano-TiO(2) did not significantly affect the membrane profile of the exposed bacterium. The results suggest the possibility that Al(2)O(3) induces changes in bacterial membranes by direct physical interaction, which was supported by TEM image analysis.

Impact of polydextrose on the faecal microbiota: a double-blind, crossover, placebo-controlled feeding study in healthy human subjects.

In this placebo-controlled, double-blind, crossover human feeding study, the effects of polydextrose (PDX; 8 g/d) on the colonic microbial composition, immune parameters, bowel habits and quality of life were investigated. PDX is a complex glucose oligomer used as a sugar replacer. The main goal of the present study was to identify the microbial groups affected by PDX fermentation in the colon. PDX was shown to significantly increase the known butyrate producer Ruminococcus intestinalis and bacteria of the Clostridium clusters I, II and IV. Of the other microbial groups investigated, decreases in the faecal Lactobacillus-Enterococcus group were demonstrated. Denaturing gel gradient electrophoresis analysis showed that bacterial profiles between PDX and placebo treatments were significantly different. PDX was shown to be slowly degraded in the colon, and the fermentation significantly reduced the genotoxicity of the faecal water. PDX also affected bowel habits of the subjects, as less abdominal discomfort was recorded and there was a trend for less hard and more formed stools during PDX consumption. Furthermore, reduced snacking was observed upon PDX consumption. This study demonstrated the impact of PDX on the colonic microbiota and showed some potential for reducing the risk factors that may be associated with colon cancer initiation.

Long-term defaunation increases the abundance of cellulolytic ruminococci and methanogens but does not affect the bacterial and methanogen diversity in the rumen of sheep.

Protozoa are commensal eukaryotes in the rumen of herbivores. Protozoa are large producers of hydrogen, which is utilized by methanogenic archaea to produce methane, a greenhouse gas. The removal of protozoa from the rumen (defaunation) decreases methanogenesis, but also negatively affects fiber digestion, which is the main function of the rumen. The aim of this study was to examine the effect of long-term defaunation on the structure of the microbiota and particularly methanogenic archaea and fibrolytic bacteria to better understand the microbial mechanisms responsible for the decrease in methanogenesis and fibrolysis. The trial was conducted in 5 adult sheep subjected successively to long-term defaunation (2 yr), refaunation (12 wk), and short-term defaunation (10 wk). Methanogens were enumerated by quantitative PCR targeting the rrs (16S ribosomal RNA subunit) and mcrA (methyl coenzyme-M reductase) genes. The rrs gene was used to quantify the 3 major culturable rumen cellulolytic bacterial species (i.e., Fibrobacter succinogenes, Ruminococcus albus, and Ruminococcus flavefaciens) and total bacteria. Bacterial and methanogen diversity was also examined by PCR-DGGE (PCR-denaturing gradient gel electrophoresis) analysis targeting the rrs and mcrA genes, respectively. Total rumen bacterial density estimated as rrs copies per gram of DM of rumen content increased in response to long- and short-term defaunation (+1 log, P < 0.001), but without noticeable shifts in diversity. Defaunation increased the rrs copies per gram of DM of rumen content of R. albus and R. flavefaciens (+2 log, P < 0 0.001), but did not affect that of F. succinogenes. Despite a 20% reduction in methane emission in the 2 defaunated periods, the mcrA and rrs copies of methanogens per gram of DM of rumen content increased (+1 log, P < 0.001) in the absence of protozoa, whereas the diversity of the dominant methanogenic community was not modified. This study shows no major difference between long- and short-term defaunation in abundance and diversity of bacteria and archaea. It also provides evidence that monitoring the abundance and diversity of methanogens is not sufficient to comprehend the microbial mechanisms leading to a reduction in methane emissions by ruminants. This study also reports for the first time in sheep a selective effect of defaunation on the abundance of cellulolytic bacterial species.

Phylotypes related to Ruminococcus bromii are abundant in the large bowel of humans and increase in response to a diet high in resistant starch.

To further understand how diets containing high levels of fibre protect against colorectal cancer, we examined the effects of diets high in nonstarch polysaccharides (NSP) or high in NSP plus resistant starch (RS) on the composition of the faecal microbial community in 46 healthy adults in a randomized crossover intervention study. Changes in bacterial populations were examined using denaturing gradient gel electrophoresis (DGGE) of 16S rRNA gene fragments. Bacterial profiles demonstrated changes in response to the consumption of both RS and NSP diets [analysis of similarities (ANOSIM): R=0.341-0.507, P<0.01]. A number of different DGGE bands with increased intensity in response to dietary intervention were attributed to as-yet uncultivated bacteria closely related to Ruminococcus bromii. A real-time PCR assay specific to the R. bromii group was applied to faecal samples from the dietary study and this group was found to comprise a significant proportion of the total community when individuals consumed their normal diets (4.4+/-2.6% of total 16S rRNA gene abundance) and numbers increased significantly (+/-67%, P<0.05) with the RS, but not the NSP, dietary intervention. This study indicates that R. bromii-related bacteria are abundant in humans and may be significant in the fermentation of complex carbohydrates in the large bowel.

Ruminococcus gauvreauii sp. nov., a glycopeptide-resistant species isolated from a human faecal specimen.

A novel strictly anaerobic, vancomycin-resistant, Gram-positive coccus (strain CCRI-16,110(T)) was isolated from a human faecal specimen. This strain was characterized using morphological, biochemical and molecular taxonomic methods. The organism was unable to hydrolyse aesculin and failed to produce acid from cellobiose, d-lactose and alpha-raffinose. Acetic acid was the sole product of glucose fermentation by the organism. On the basis of 16S rRNA and tuf gene sequence comparison, strain CCRI-16,110(T) was most closely related to species of the genus Ruminococcus and formed a hitherto unknown sublineage within the Clostridium coccoides rRNA cluster of organisms (cluster XIVa). Based on phenotypic and phylogenetic evidence, a novel species, Ruminococcus gauvreauii sp. nov., is proposed. The type strain is CCRI-16,110(T) (=NML 060141(T) =CCUG 54,292(T) =JCM 14987(T)).

vanD and vanG-like gene clusters in a Ruminococcus species isolated from human bowel flora.

A vancomycin-resistant, anaerobic, gram-positive coccus containing the vanD and vanG-like genes (strain CCRI-16110) was isolated from a human fecal specimen during a hospital surveillance program to detect carriers of vancomycin-resistant enterococci. Comparison of the 16S rRNA gene sequence of strain CCRI-16110 with databases revealed a potentially novel Ruminococcus species that was most similar (<94% identity) to Clostridium and Ruminococcus species. Strain CCRI-16110 was highly resistant to vancomycin and teicoplanin (MICs of >256 microg/ml). The complete DNA sequence of the vanD cluster was most similar (98.2% identity) to that of Enterococcus faecium BM4339, containing the vanD1 allele. An intD gene with 99% identity with that of this E. faecium strain was found to be associated with the vanD gene cluster of this novel anaerobic bacterium. Strain CCRI-16110 also harbors genes encoding putative VanS(G), VanG, and VanT(G) proteins displaying 56, 73.6, and 55% amino acid sequence identity, respectively, compared to the corresponding proteins encoded by the vanG1 and vanG2 operons of Enterococcus faecalis BM4518 and N03-0233. This study reports for the first time an anaerobic bacterium containing the vanD gene cluster. This strain also harbors a partial vanG-like gene cluster. The presence of vanD- and vanG-containing anaerobic bacteria in the human bowel flora suggests that these bacteria may serve as a reservoir for the vanD and vanG vancomycin resistance genes.

[Liver abscess and infective endocarditis cases caused by Ruminococcus productus]. / Ruminococcus productus'un etken oldugu karaciger apsesi ve enfektif endokardit olgulari.

The genus Ruminococcus which are anaerobe Gram positive cocci, previously classified as Peptostreptococcus, may colonize the upper respiratory tract, gastrointestinal tract, vagina and skin of humans and animals. In this report a case of liver abscess and a case of infective endocarditis caused by Ruminoccocus productus, which is very rarely encountered in the clinical practice were presented. The first case was a 32 years old male who was admitted to the hospital in 2002, with the complaints of fever lasting for 20 days and pain while breathing. The abdominal ultrasonography revealed the presence of a liver abscess, and the drainage material from the abscess yielded Ruminococcus productus, identified in BACTEC 9200 (Becton Dickinson, Sparks, Md) anaerobe system. As the isolate was found to be sensitive to penicilin, the empirical gentamicin and ampicillin/sulbactam therapy was continued. The second case was a 25 years old male who was admitted to the hospital in 2005, with the signs of fever lasting for 3-4 months, chills, bone and joint pains. As multiple vegetations were detected in echocardiography, blood cultures were collected and empirical therapy with ceftriaxone and gentamicin was initiated with the preliminary diagnosis of infective endocarditis. Bacteria which were isolated from blood cultures by BACTEC 9200 system have been identified as R. productus. As this strain was also sensitive to penicillin, the empirical therapy was changed to penicilin and gentamicin. These two cases indicated that R. productus should be considered in complicated infections even if it is a rarely isolated species from the clinical samples.

Effects of difructose anhydride III (DFA III) administration on rat intestinal microbiota.

The effects of difructose anhydride III (di-D-fructofuranose-1,2':2,3'-dianhydride; DFA III) administration (3% DFA III for 4 weeks) on rat intestinal microbiota were examined using denaturing gradient gel electrophoresis (DGGE). According to DGGE profiles, the number of bacteria related to Bacteroides acidofaciens and uncultured bacteria within the Clostridium lituseburense group decreased, while that of bacteria related to Bacteroides vulgatus, Bacteroides uniformis and Ruminococcus productus increased in DFA III-fed rat cecum. In the cecal contents of DFA III-fed rats, a lowering of pH and an increase in short chain fatty acids (SCFAs), especially acetic acid, were observed. The DFA III-assimilating bacterium, Ruminococcus sp. M-1, was isolated from the cecal contents of DFA III-fed rats. The strain had 98% similarity with R. productus ATCC 27340T (L76595), and mainly produced acetic acid. These results confirmed that the bacteria harmful to host health were not increased by DFA III administration. Moreover, DFA III stimulated the growth of Ruminococcus sp. M-1 producing acetic acid, which may alter the intestinal microbiota towards a healthier composition. It is expected that DFA III would be a new candidate as a prebiotic.

Effects of difructose anhydride III (DFA III) administration on bile acids and growth of DFA III-assimilating bacterium Ruminococcus productus on rat intestine.

The growth of DFA III-assimilating bacteria in the intestines of rats fed 3% DFA III for 2 weeks was examined. Sixty-four percent of the DFA III intake had been assimilated on day 3 of ingestion, and almost all of the DFA III was assimilated at the end of the experiment. The DFA III-assimilating bacterium, Ruminococcus productus, in DFA III-fed rats was in the stationary state of 10(8)-10(9) cells/g dry feces within a week from 10(6) cells/g dry feces on day 1 of DFA III ingestion. The number of R. productus cells was associated with the amount of DFA III excreted in the feces. The acetic acid produced from DFA III by R. productus lowered the cecal pH to 5.8. In control-fed rats and DFA III-fed rats, 94% of secondary bile acids and 94% of primary bile acids, respectively, were accounted for in the total bile acids analyzed. DFA III ingestion increased the ratio of primary bile acids and changed the composition of fecal bile acids. In conclusion, R. productus assimilated DFA III, produced short chain fatty acids, and the cecal pH was lowered. The acidification of rat intestine perhaps inhibited secondary bile acid formation and decreased the ratio of secondary bile acids. Therefore, it is expected that DFA III may prevent colorectal cancer and be a new prebiotic candidate.

Use of community genome arrays (CGAs) to assess the effects of Acacia angustissima on rumen ecology.

This research developed a community genome array (CGA) to assess the effects of Acacia angustissima on rumen microbiology. A. angustissima produces non-protein amino acids as well as tannins, which may be toxic to animals, and CGA was used to assess the effects of this plant on the ecology of the rumen. CGAs were developed using a 7.5 cmx2.5 cm nylon membrane format that included up to 96 bacterial genomes. It was possible to separately hybridize large numbers of membranes at once using this mini-membrane format. Pair-wise cross-hybridization experiments were conducted to determine the degree of cross-hybridization between strains; cross-hybridization occurred between strains of the same species, but little cross-reactivity was observed among different species. CGAs were successfully used to survey the microbial communities of animals consuming an A. angustissima containing diet but quantification was not precise. To properly quantify and validate the CGA, Fibrobacter and Ruminococcus populations were independently assessed using 16S rDNA probes to extracted rRNA. The CGA detected an increase in these populations as acacia increased in the diet, which was confirmed by rRNA analysis. There was a great deal of variation among strains of the same species in how they responded to A. angustissima. However, in general Selenomonas strains tended to be resistant to the tannins in the acacia while Butyrivibrio fibrisolvens was sensitive. On the other hand some species, like streptococci, varied. Streptococcus bovis-like strains were sensitive to an increase in acacia in the diet while Streptococcus gallolyticus-like strains were resistant. Strep. gallolyticus has independently been shown to be resistant to tannins. It is concluded that there is significant variation in tannin resistance between strains of the same species. This implies that there are specific molecular mechanisms at play that are independent of the phylogenetic position of the organism.