Improving the antiprotozoal effect of saponins in the rumen by combination with glycosidase inhibiting iminosugars or by modification of their chemical structure.

The antiprotozoal effect of saponins is transitory, as when saponins are deglycosylated to sapogenins by rumen microorganisms they become inactive. We hypothesised that the combination of saponins with glycosidase-inhibiting iminosugars might potentially increase the effectiveness of saponins over time by preventing their deglycosylation in the rumen. Alternatively, modifying the structure of the saponins by substituting the sugar moiety with other small polar residues might maintain their activity as the sugar substitute would not be enzymatically cleaved. The aim of this in vitro study was to evaluate the acute antiprotozoal effect and the stability of this effect over a 24 h incubation period using ivy saponins, a stevia extract rich in iminosugars, ivy saponins with stevia extract, and a chemically modified ivy saponin, hederagenin bis-succinate (HBS). The effects on fermentation parameters and rumen bacterial communities were also studied. Ivy saponins with stevia and HBS had a greater antiprotozoal effect than ivy saponins, and this effect was maintained after 24 h of incubation (P<0.001). The combination of ivy and stevia extracts was more effective in shifting the fermentation pattern towards higher propionate (+39%) and lower butyrate (-32%) and lower ammonia concentration (-64%) than the extracts incubated separately. HBS caused a decrease in butyrate (-45%) and an increase in propionate (+43%) molar proportions. However, the decrease in ammonia concentration (-42%) observed in the presence of HBS was less than that caused by ivy saponins, either alone or with stevia. Whereas HBS and stevia impacted on bacterial population in terms of community structure, only HBS had an effect in terms of biodiversity (P<0.05). It was concluded that ivy saponins with stevia and the modified saponin HBS had a strong antiprotozoal effect, although they differed in their effects on fermentation parameters and bacteria communities. Ivy saponins combined with an iminosugar-rich stevia extract and/or HBS should be evaluated to determine their antiprotozoal effect in vivo.

In vitro screening of natural feed additives from crustaceans, diatoms, seaweeds and plant extracts to manipulate rumen fermentation.

BACKGROUND: Eight natural products from animal, unicellular algae, brown seaweed and plant origins were chosen according to their theoretical antimicrobial activity: Diatomaceous earths (DE), insoluble chitosan (ICHI), soluble chitosan (CHI), seaweed meal (SWM), Ascophyllum nodosum (ASC), Laminaria digitata (LAM), neem oil (NOIL) and an ivy fruit extract rich in saponins (IVY). Dose-response incubations were conducted to determine their effect on rumen fermentation pattern and gas production, while their anti-protozoal activity was tested using (14) C-labelled bacteria. RESULTS: DE, SWM, NOIL and ICHI had very small effects on rumen function when used at inclusion rate up to 2 g L(-1) . ASC had anti-protozoal effects (up to -23%) promoting a decrease in gas production and methanogenesis (-15%). LAM increased VFA production (+7%) and shifted from butyrate to acetate. CHI also shifted fermentation towards propionate production and lower methane (-23%) and protozoal activity (-56%). IVY decreased protozoal activity (-39%) and ammonia concentration (-56%), as well as increased feed fermentation (+11% VFA concentration) and shifted from acetate to propionate production. CONCLUSIONS: ASC, LAM, CHI and IVY showed promising potential in vitro as feed additives to improve rumen function, thus more research is needed to investigate their mode of action in the rumen microbial ecosystem. © 2015 Society of Chemical Industry.

Effects and mode of action of chitosan and ivy fruit saponins on the microbiome, fermentation and methanogenesis in the rumen simulation technique.

This study investigates the effects of supplementing a control diet (CON) with chitosan (CHI) or ivy fruit saponins (IVY) as natural feed additives. Both additives had similar abilities to decrease rumen methanogenesis (-42% and -40%, respectively) using different mechanisms: due to its antimicrobial and nutritional properties CHI promoted a shift in the fermentation pattern towards propionate production which explained about two thirds of the decrease in methanogenesis. This shift was achieved by a simplification of the structure in the bacterial community and a substitution of fibrolytic (Firmicutes and Fibrobacteres) by amylolytic bacteria (Bacteroidetes and Proteobacteria) which led to greater amylase activity, lactate and microbial protein yield with no detrimental effect on feed digestibility. Contrarily, IVY had negligible nutritional properties promoting minor changes in the fermentation pattern and on the bacterial community. Instead, IVY modified the structure of the methanogen community and decreased its diversity. This specific antimicrobial effect of IVY against methanogens was considered its main antimethanogenic mechanism. IVY had however a negative impact on microbial protein synthesis. Therefore, CHI and IVY should be further investigated in vivo to determine the optimum doses which maintain low methanogenesis but prevent negative effects on the rumen fermentation and animal metabolism.

The effects of a probiotic yeast on the bacterial diversity and population structure in the rumen of cattle.

It has been suggested that the ability of live yeast to improve milk yield and weight gain in cattle is because the yeast stimulates bacterial activity within the rumen. However it remains unclear if this is a general stimulation of all species or a specific stimulation of certain species. Here we characterised the change in the bacterial population within the rumen of cattle fed supplemental live yeast. Three cannulated lactating cows received a daily ration (24 kg/d) of corn silage (61% of DM), concentrates (30% of DM), dehydrated alfalfa (9% of DM) and a minerals and vitamins mix (1% of DM). The effect of yeast (BIOSAF SC 47, Lesaffre Feed Additives, France; 0.5 or 5 g/d) was compared to a control (no additive) in a 3 × 3 Latin square design. The variation in the rumen bacterial community between treatments was assessed using Serial Analysis of V1 Ribosomal Sequence Tag (SARST-V1) and 454 pyrosequencing based on analysis of the 16S rRNA gene. Compared to the control diet supplementation of probiotic yeast maintained a healthy fermentation in the rumen of lactating cattle (higher VFA concentration [high yeast dose only], higher rumen pH, and lower Eh and lactate). These improvements were accompanied with a shift in the main fibrolytic group (Fibrobacter and Ruminococcus) and lactate utilising bacteria (Megasphaera and Selenomonas). In addition we have shown that the analysis of short V1 region of 16s rRNA gene (50-60 bp) could give as much phylogenetic information as a longer read (454 pyrosequencing of 250 bp). This study also highlights the difficulty of drawing conclusions on composition and diversity of complex microbiota because of the variation caused by the use of different methods (sequencing technology and/or analysis).

Changes in the bacterial population of the caecum and stomach of the rabbit in response to addition of dietary caprylic acid.

The effect of caprylic acid, either in its pure form, or as Akomed R, on the microbial community of the stomach and caecum of farmed rabbits was investigated. This fatty acid, which is often added to the diet of farmed rabbits to reduce mortality rates was shown to reduce the number of coliforms isolated from both the stomach and the caecum. Moreover, it led to a reduction in the total number of anaerobic bacteria isolated from the caecum, but not for those isolated from the stomach. Its mode of action remains unclear, but here it is shown by use of both DGGE and TRFLP analysis that these changes are not confined to one specific group of bacteria, but rather affects a number of species.

Regeneration of cryoresistance of in vitro rumen ciliate cultures.

The purpose of this study was to investigate factors affecting mechanical- and cryo-resistance of the rumen ciliates Entodinium caudatum (E.c.), Entodinium furca monolobum (E.f.m.), Entodinium simplex (E.s.), Diplodinium denticulatum (two clones, D.d.01 and D.d.02), Diploplastron affine (D.a.) and Epidinium ecaudatum forma caudatum (E.e.c.) after long-term in vitro cultivation. Following prolonged in vitro cultivation (more than six months), the ciliates were very sensitive to both centrifugation and 5% (v/v) dimethylsulphoxide, with motility decreased to: 39 and 23% for E.c., 66 and 32% for E.f.m., 46 and 27% for D.d. 01, 64 and 41% for D.a., and 44 and 28% for E.e.c., respectively. Thus, cryopreservation was unsuccessful. The effect of supplementing the ciliate growth medium with rumen fluid, glycine-betaine, proline, myo-inositol, linoleic acid, Sel-Plex or insulin, together with the effect of the source of rumen fluid on ciliate resistance to centrifugation, dimethylsulphoxide and freezing was also tested. The omission of rumen fluid from the growth medium resulted in the loss of cryoresistance after one-month cultivation. Supplementing the growth environment with a combination of glycine-betaine, proline, linoleic acid, Sel-Plex, insulin plus improved quality rumen fluid significantly enhanced survival of the ciliates after the freezing-thawing procedure (from 1 to 33% survival in un-supplemented vs. supplemented for E.c., P<0.01; 4-40% E.f.m., P<0.01; 0-17% D.d., P<0.05; 5-7% D.a. and 4-36% E.e.c., P<0.01).