Growth and morphologic response of rumen methanogenic archaea and bacteria to cashew nut shell liquid and its alkylphenol components.

The growth and morphology of rumen methanogenic archaea (15 strains of 10 species in 5 genera, including 7 strains newly isolated in the present study) and bacteria (14 species in 12 genera) were investigated using unsupplemented in vitro pure cultures and cultures supplemented with cashew nut shell liquid (CNSL) and its phenolic compound components, anti-methanogenic agents for ruminant animals. Growth of most of the methanogens tested was inhibited by CNSL and alkylphenols at different concentrations ranging from 1.56 to 12.5 µg/ml. Of the alkylphenols tested, anacardic acid exhibited the most potent growth inhibition. Three gram-negative bacterial species involved in propionate production were resistant to CNSL and alkylphenols (>50 µg/ml). All the methanogens and bacteria that were sensitive to CNSL and alkylphenols exhibited altered morphology; disruption of the cell surface was notable, possibly due to surfactant activity of the tested materials. Cells division was inhibited in some organisms, with cell elongation and unclear septum formation observed. These results indicate that CNSL and alkylphenols, particularly anacardic acid, inhibit both rumen bacteria and methanogens in a selective manner, which could help mitigate rumen methane generation.

Feeding cashew nut shell liquid decreases methane production from feces by altering fecal bacterial and archaeal communities in Thai local ruminants.

The effect of feeding cashew nut shell liquid (CNSL) on fecal fermentation products and microbiota was investigated in Thai native cattle and swamp buffaloes. Four of each animal were fed rice straw and concentrate diet with control pellets without CNSL for 4 weeks, followed by the same diet with pellets containing CNSL for another 4 weeks, so that CNSL was administered at a level of 4 g/100 kg body weight. Feces were collected the last 2 days in each feeding period. CNSL alkyl phenols were recovered from feces (16%-28%) in a similar proportion to those in the diet, indicating that most functional anacardic acid was not selectively removed throughout the digestive tract. In vitro production of gas from feces, particularly methane, decreased with CNSL feeding. The proportion of acetate in feces decreased with CNSL feeding, whereas that of propionate increased, without affecting total short-chain fatty acid concentration. CNSL feeding changed fecal microbial community, particularly in swamp buffaloes, which exhibited decreases in the frequencies of Treponema, unclassified Ruminococcaceae, and Methanomassiliicoccaceae. These results suggest that CNSL feeding alters not only rumen fermentation but also hindgut fermentation via modulation of the microbial community, thereby potentially attenuating methane emission from the feces of ruminant animals.

Microbial community structure of the bovine rumen as affected by feeding cashew nut shell liquid, a methane-inhibiting and propionate-enhancing agent.

The effect of cashew nut shell liquid (CNSL) feeding on bacterial and archaeal community of the bovine rumen was investigated by analyzing clone libraries targeting 16S rRNA genes, methyl-coenzyme reductase A-encoding genes (mcrA), and their respective transcripts. Rumen samples were collected from three non-lactating cows fed on a hay and concentrate diet with or without CNSL supplementation. DNA and complementary DNA (cDNA) libraries were generated for investigating rumen microbial communities. MiSeq analysis also was performed to understand more comprehensively the changes in the microbial community structures. Following CNSL supplementation, the number of operational taxonomical unit (OTU) and diversity indices of bacterial and archaeal community were decreased. Bacterial OTUs belonging to Proteobacteria, including Succinivibrio, occurred at a higher frequency with CNSL feeding, especially in cDNA libraries. The methanogenic archaeal community became dominated by Methanomicrobium. A bacterial community shift also was observed in the MiSeq data, indicating that CNSL increased the proportion of Succinivibrio and other genera known to be involved in propionate production. Methanogenic archaeal community shifts to increase Methanoplanus and to decrease Methanobrevibacter also were observed. Together, these results imply the occurrence of significant changes in rumen communities, not only for bacteria but also for methanogens, following CNSL feeding.

Rumen responses to dietary supplementation with cashew nut shell liquid and its cessation in sheep.

Rumen responses to cashew nut shell liquid (CNSL) were evaluated in a feeding study. Four wethers were fed a hay and concentrate diet for 4 weeks (pre-CNSL period), and then fed the same diet supplemented with low and high levels of CNSL for 2 weeks each (L-CNSL and H-CNSL periods respectively). The diet was then reverted to the unsupplemented control diet for another 2 weeks (post-CNSL period). Rumen parameters were monitored in each feeding period. CNSL, regardless of the two levels tested, did not show any adverse effects on total short chain fatty acid concentration and dry matter digestibility in the rumen. Propionate proportion increased in the H-CNSL period, while methane production potential, acetate and butyrate proportions, viscosity, foam formation and its stability, and ammonia concentration decreased. Values of these parameters returned to those in the unsupplemented control period after cessation of CNSL supplementation. Clone library analysis of 16S rRNA genes revealed the following shifts in the H-CNSL period. For bacteria, Firmicutes was frequently detected, while Bacteroidetes and Spirochetes were not. For archaea, Methanobrevibacter wolinii was predominant. These results indicate that CNSL could be a methane inhibitor and propionate enhancer by altering the rumen microbial community.

Effect of cashew nut shell liquid on metabolic hydrogen flow on bovine rumen fermentation.

Effect of cashew nut shell liquid (CNSL), a methane inhibitor, on bovine rumen fermentation was investigated through analysis of the metabolic hydrogen flow estimated from concentrations of short-chain fatty acids (SCFA) and methane. Three cows were fed a concentrate and hay diet without or with a CNSL-containing pellet. Two trials were conducted using CNSL pellets blended with only silica (trial 1) or with several other ingredients (trial 2). Methane production was measured in a respiration chamber system, and energy balance and nutrient digestibility were monitored. The estimated flow of metabolic hydrogen demonstrated that a part of metabolic hydrogen was used for hydrogen gas production, and a large amount of it flowed into production of methane and SCFA in both trial 1 and 2, when CNSL was administered to the bovine rumen. The results obtained by regression analyses showed that the effect of CNSL supply on methane reduction was coupled with a significant (P < 0.01) decrease of acetate and a significant (P < 0.01) increase of propionate and hydrogen gas. These findings reveal that CNSL is able to reduce methane and acetate production, and to increase hydrogen gas and propionate production in vivo.