Breed-dependent differences in gut microbiota linked to indole and skatole levels in Hezuo and DLY pigs.

The effects of dietary fibre, lactose and antibiotic on the levels of skatole and indole in faeces and subcutaneous fat in growing pigs

Skatole and indole concentrations in the rumen were compared with the rumen concentrations of ammonia and volatile fatty acids (VFA) in sheep fed fresh white clover (WC; Trifolium repens), perennial ryegrass (PRG; Lolium perenne) or the condensed tannin-containing forage, Lotus corniculatus (LC). Six rumen fistulated Romney wethers were fed the cut forages over two 2-hour meal periods in the morning and afternoon, with rumen contents sampled at intervals after the start of the morning meal. Maximum concentrations of skatole, indole and ammonia were highest when feeding WC (P<0.05). Ammonia accumulates in the rumen when there is rapid protein degradation with unsynchronised energy supply. Condensed tannins in LC slowed protein degradation and consequently maximal ammonia, indole and skatole concentration were lower when feeding this forage (P<0.05). This indicates that skatole and indole formation will be greater when there is high protein degradability. Ammonia, indole and skatole concentrations in sheep fed PRG were intermediate, and not significantly different from either WC or LC. Maximum concentration of straight-chain volatile fatty acids (VFA) were not significantly different for the forages, but branchedchain VFA concentrations tended to be higher for WC (P=0.103) compared with LC. Straight-chain VFAs are a by-product of microbial carbohydrate fermentation in the rumen, but branched-chain VFAs are formed from deamination of amino acids. This suggests that skatole and indole production are not related to carbohydrate digestion in the rumen. In order for meat and milk producers in pastoral systems to ameliorate undesirable flavours and increase demand in certain markets, they will need to consider using alternative forages to reduce protein solubility and degradability.

Effect of non-starch polysaccharides on production and absorption of indolic compounds in entire male pigs

BackgroundPublic complaints concerning odor emissions from intensive livestock and poultry farms continue to grow, as nauseous odorous compounds have adverse impacts on the environment and human health. Itaconic acid is a metabolite from the citric acid cycle of the host and shows volatile odor-reducing effects during animal production operations. However, the specific role of itaconic acid in decreasing intestinal odorous compound production remains unclear. A total of 360 one-day-old chicks were randomly divided into 6 treatment groups: control group (basal diet) and itaconic acid groups (basal diet + 2, 4, 6, 8 and 10 g/kg itaconic acid). The feeding experiment lasted for 42 d.ResultsDietary itaconic acid supplementation linearly and quadratically decreased (P < 0.05) the cecal concentrations of indole and skatole but did not affect (P > 0.05) those of lactic, acetic, propionic and butyric acids. The cecal microbial shift was significant in response to 6 g/kg itaconic acid supplementation, in that the abundances of Firmicutes, Ruminococcus and Clostridium were increased (P < 0.05), while those of Bacteroidetes, Escherichia-Shigella and Bacteroides were decreased (P < 0.05), indicative of increased microbial richness and diversity. Furthermore, a total of 35 significantly (P < 0.05) modified metabolites were obtained by metabolomic analysis. Itaconic acid decreased (P < 0.05) the levels of nicotinic acid, nicotinamide, glucose-6-phosphate, fumatic acid and malic acid and increased (P < 0.05) 5-methoxytroptomine, dodecanoic acid and stearic acid, which are connected with the glycolytic pathway, citrate acid cycle and tryptophan metabolism. Correlation analysis indicated significant correlations between the altered cecal microbiota and metabolites; Firmicutes, Ruminococcus and Clostridium were shown to be negatively correlated with indole and skatole production, while Bacteroidetes, Escherichia-Shigella and Bacteroides were positively correlated with indole and skatole production.ConclusionsItaconic acid decreased cecal indole and skatole levels and altered the microbiome and metabolome in favor of odorous compound reduction. These findings provide new insight into the role of itaconic acid and expand its application potential in broilers.

The experiment was conducted to investigate the in vitro effects of inulin and soya bean oligosaccharide (SBO) on the metabolism of L-tryptophan (L-try) to skatole production, and the intestinal microbiota in broilers. Treatments were as follows: caecal microbiota control (Cc), Cc+inulin, Cc+SBO, rectal microbiota control (Rc), Rc+inulin and Rc+SBO. Microbial suspensions were anaerobically incubated at 38°C for 24hr. The results showed that concentrations of skatole and acetic acid were significantly lower in caecal microbiota fermentation broth (MFB) than those in rectal MFB (p<.05). Addition of inulin or SBO significantly decreased the concentrations of indole and skatole and rate of L-try degradation (p<.05). Inulin groups had lower indole than SBO groups (p<.05). PCR-DGGE analysis revealed that addition of inulin or SBO decreased the microbiota richness (p<.05), but no significant differences in Shannon index (p>.05). Four distinct bands were detected in inulin and SBO groups, which were related to two of Bacteroides, one of Firmicutes and Bifidobacteria. Six bands were detected only in control groups, which represented uncultured Rikenellaceae, Roseburia, Escherichia/Shigella dysenteriae, Bacteroides uniformis (T), Parabacteroides distasonis and Enterobacter aerogenes. Populations of Lactobacilli, Bifidobacteria and total bacteria in inulin groups were higher than those in control groups (p<.05). For SBO groups, only population of total bacteria increased (p<.05). However, there were no significant differences in Escherichia coli population among treatments (p>.05). These results suggest that reduced concentrations of skatole and indole in the presence of inulin and SBO may be caused by decrease in L-try degradation rate, which were caused by change in microbial ecosystem and pH value. Uncultured B.uniformis (T) and E.aerogenes may be responsible for degradation of L-try to skatole.

The gut microbiota plays vital roles in metabolizing nutrient, maintaining the intestinal epithelial barrier but also in modulating immunity. Host genetics and the pig breed are implicated in shaping gut microbiota. Tibetan pig is a unique native Chinese breed and has evolved to manifest a strong disease resistance. However, the immunity and microbiota of growing Tibetan (TP) pigs were still rarely understood. The jejunal immunity phenotype and microbial composition of TP and Duroc × (Landrace × Yorkshire) (DLY) pigs were explored through immunohistochemistry and 16S rRNA sequencing. Higher scores of clusters of differentiation 4 (CD4+) and Toll-like receptor 9 (TLR9) were observed in TP pigs than those of DLY pigs (p < 0.05), as were Interleukin 10 (IL-10) and zonular occludens 1 (ZO-1) (p < 0.01). Similar levels of bacterial richness and diversity were found in the jejunal microbiota of the TP and DLY pigs. However, the TP pigs showed a significantly different microbiome compared to DLY pigs at the genus level (ANOSIM; p < 0.05). Pseudomonas, Stenotrophomonas, Phenylobacterium, and Sandaracinobacter were enriched in DLY pigs (p < 0.05), while the Lactobacillus and Solibacillus had higher abundances in TP pigs than DLY pigs (p < 0.05). Tibetan pigs have “healthier” intestinal microbial communities than DLY pigs. Close relationships were found between jejunal immune performance and the differential bacteria, Lactobacillus can enhance porcine jejunal immunity, while Stenotrophomonas will have a negative impact on porcine gut immunity.

Skatole and indole concentration and the odour of fat from lambs that had grazed perennial ryegrass/white clover pasture or Lotus corniculatus

BACKGROUND: An excessive accumulation of skatole in pigs is a major contributor to boar taint. Intestinal skatole concentrations may vary among different pig segments and breeds. The objective of this study was to evaluate the spatial variation of intestinal skatole production and the microbial community in local Jinhua and exotic Landrace pigs.RESULTS: For both breeds, skatole concentration increased towards the rectum and segment had a significant effect on skatole (P &lt; 0.001). The caecal skatole concentration in Landrace was significantly lower than proximal, distal colonic and rectal skatole levels (P &lt; 0.05). Compared with Jinhua pigs, the rectal skatole and proximal and distal colonic indole levels were significantly higher in Landrace pigs (P &lt; 0.05). Volatile fatty acids were significantly affected by breed except for isobutyrate and isovalerate (P &lt; 0.05), with higher butyrate level and lower pH in Landrace pigs (P &lt; 0.05). There were no significant differences in mitosis and apoptosis for the two breeds. The denaturing gradient gel electrophoresis profile showed differences between breeds and segments within one breed, though some bands were common to all samples.CONCLUSION: The higher skatole levels in Landrace pigs may be not associated with higher intestinal apoptosis. The results suggested that pig genotype plays a role in the establishment of the host‐specific microbiota and that the variations in fermentation patterns are more likely to explain differences in intestinal skatole production. Copyright © 2009 Society of Chemical Industry

Simple SummaryThe elimination of boar taint by a method other than surgical castration without anaesthesia is currently one of the main topics in pig research. Boar taint occurs in meat from some entire male pigs and is undesirable for sensitive consumers. Boar taint is mainly caused by skatole. Skatole is produced by the breakdown of proteins by intestinal bacteria and can be stored in meat and reduce its sensory quality (taste and odour). Boar taint can be reduced by a diet high in easily fermentable saccharides, such as Jerusalem artichoke (Helianthus tuberosus L.). These saccharides change the bacterial colonisation in the intestines and thus reduce the production of skatole. The aim of this study was to evaluate the effects of different levels of Jerusalem artichoke on performance, carcass composition and skatole and indole levels in adipose tissue and on microbiota in faecal samples. In the present study, Jerusalem artichoke had no negative effect on the growth performance or carcass value in male pigs. Moreover, Jerusalem artichoke led to decreased skatole levels in the adipose tissue, probably due to the decreased level of proteolytic bacteria, which cause a higher rate of skatole production in the gastrointestinal tract. It seems that a dietary concentration of 8.1% of Jerusalem artichoke fed 13 days before slaughter is a sufficient dose for decreasing the skatole levels to those of castrated males, and this approach could be an alternative to the surgical castration of male pigs.Jerusalem artichoke contains inulin polysaccharide, which has prebiotic effects and influences the microbiota of the digestive tract. The addition of Jerusalem artichoke in boar diets may decrease the content of skatole and indole, which are the main constituents of boar taint, and may also negatively affect the taste and odor. The objective of this study was to evaluate the effects of different levels of Helianthus tuberosus L. (H. tuberosus) in feed mixtures on performance, carcass composition, the levels of microbiota in faecal samples, and the concentrations of skatole and indole in adipose tissue. The study was performed with 47 crossbred entire male pigs of the Large White sire × (Large White dame × Landrace) genotype fed a basal diet with 0%, 4.1%, 8.1% or 12.2% H. tuberosus for 13 days before slaughter. Significant differences in daily weight gain and daily feed intake were found (p = 0.045), with the values being lower in the group with the highest level of H. tuberosus. In addition, increasing levels of H. tuberosus decreased the concentration of skatole in the adipose tissue (p = 0.003). The highest level of H. tuberosus decreased the level of Escherichia coli (p ≤ 0.001) in the faeces. The enterococcal count increased (p = 0.029) in groups with a diet that included 4.1% and 8.1% H. tuberosus. There was also a significant correlation between the concentration of H. tuberosus and the concentration of E. coli (p < 0.001; −0.64) and the skatole levels in the adipose tissue (p = 0.001; –0.46). Moreover, there was also a positive correlation between the concentration of E. coli and the skatole levels in the adipose tissue (p = 0.023; 0.33). In conclusion, feeding pigs with H. tuberosus leads to decreased levels of skatole in the adipose tissue. According to the results of our study, a diet with 8.1% H. tuberosus is sufficient for decreasing skatole levels, which could be due to the decreased levels of pathogenic bacteria in the intestines.

Skatole and indole concentrations in Longissimus dorsi and fat samples of pigs

ObjectiveThe objective of this study was to investigate the in vitro fermentation profiles of different soybean oligosaccharides (SBOs) and their effects on skatole production and cecal microbiota of broilers.MethodsFive SBOs with varying main component contents were fermented using an in vitro batch incubation inoculated with broiler cecal microbiota. Gas production was recorded automatically, skatole, indole and short-chain fatty acids (SCFAs) were determined using high-performance liquid chromatography, and microbial changes were analyzed using 16S DNA gene sequencing.ResultsThe addition of SBOs increased (p<0.05) gas production, suggesting bacterial growth-stimulating activities. In addition, the concentrations of indole were significantly (p<0.05) decreased after SBO supplementation, and SBO III, with higher sucrose and stachyose contents, decreased (p<0.05) the skatole level. Our results also revealed that the fermentation of SBOs by cecal microbiota produced (p<0.05) SCFAs, which were dominated by propionic acid, butyrate acid and lactic acid compared to the control. In addition, SBO III increased (p<0.05) the abundance of Firmicutes and Subdoligranulum and decreased that of Bacteroides.ConclusionThese results suggest that SBOs with higher sucrose and stachyose contents are promising prebiotics in modulating gut microbiota and reducing odor emission in broilers.

The effect of high levels of dietary chicory roots (25%) and intracecal exogenous butyrate infusion on skatole formation and gut microbiota was investigated in order to clarify the mechanisms underlying the known reducing effect of chicory roots on skatole production in entire male pigs. A Latin square design with 3 treatments (control, chicory, and butyrate), 3 periods, and 6 animals was carried out. Chicory roots showed the lowest numerical levels of skatole in both feces and plasma and butyrate infusion the highest. In the chicory group, an increased abundance of the skatole-producing bacterium Olsenella scatoligenes compared to the control group (P = 0.06), and a numerically higher relative abundance of Olsenella than for the control and butyrate groups, was observed. Regarding butyrate-producing bacteria, the chicory group had lower abundance of Roseburia but a numerically higher abundance of Megasphaera than the control group. Lower species richness was found in the chicory group than in the butyrate group. Moreover, beta diversity revealed that the chicory group formed a distinct cluster, whereas the control and butyrate groups clustered more closely to each other. The current data indicated that the skatole-reducing effect of chicory roots is neither via inhibition of cell apoptosis by butyrate nor via suppression of skatole-producing bacteria in the pig hindgut. Thus, the mode of action is most likely through increased microbial activity with a corresponding high incorporation of amino acids into bacterial biomass, and thereby suppressed conversion of tryptophan into skatole, as indicated in the literature.IMPORTANCE Castration is practiced to avoid the development of boar taint, which negatively affects the taste and odor of pork, and undesirable aggressive behavior. Due to animal welfare issues, alternatives to surgical castration are sought, though. Boar taint is a result of high concentrations of skatole and androstenone in back fat. Skatole is produced by microbial fermentation in the large intestine, and therefore, its production can be influenced by manipulation of the microbiota. Highly fermentable dietary fiber reduces skatole production. However, various theories have been proposed to explain the mode of action. In order to search for other alternatives, more efficient or less expensive, to reduce skatole via feeding, it is important to elucidate the mechanism behind the observed effect of highly fermentable dietary fiber on skatole. Our results indicate that highly fermentable dietary fiber does not affect skatole production by reducing the number of skatole-producing bacteria or stimulating butyrate production in the large intestine.

Condensed tannin in the form of a grape seed extract (GSE) was dosed to weaned wether lambs fed white clover (WC) or perennial ryegrass (PRG) over a 9‐week period to determine whether the ‘pastoral’ flavour and odour of meat could be altered. The concentrations of the pastoral flavour compounds indole and skatole were determined in the rumen fluid, blood plasma and intermuscular fat. The odour and flavour of fat and meat from the slaughtered lambs was assessed by a trained panel. The rumen fluid and blood plasma concentrations of indole and skatole were higher in those lambs fed WC compared to PRG (P &lt; 0.05) and the overall meat flavour intensity was greater when feeding WC (P &lt; 0.01). The observed concentration of indole and skatole in the fat between WC and PRG feeding treatments was not statistically different. Power analysis indicated that increasing the number of lambs per treatment group from 20 to 65 would result in a higher fat skatole concentration (P &lt; 0.05) being detected in lambs fed WC compared to PRG. Dosing with GSE gave a small reduction in skatole concentration in the rumen fluid and reduced plasma concentration of indole and skatole (P &lt; 0.001). Odour and flavour scores of the fat and meat samples were not particularly high however, dosing with GSE lowered the overall and sweet odour and the sheepy, camphor, faecal and barnyard flavour (P &lt; 0.05). Although the plasma concentration of indole and skatole suggests that GSE reduced indole and skatole formation, the intermittent supply of the GSE to the rumen environment was not sufficient to reduce their concentration in the fat. Hence, the small difference in the scores for pastoral odour and flavour attributes associated with GSE treatment may arise from other unknown factors. From a primary investigation, there was no difference in the concentration of indole and skatole in fat samples collected from carcasses before and after chilling. Further investigations into meat pastoral flavour are warranted through feeding condensed tannin‐containing forages. Copyright © 2007 Society of Chemical Industry

The effect of cereal type and enzyme supplementation on carcass characteristics, volatile fatty acids and intestinal microflora and boar taint in entire male pigs

Boars play a critical role in pig production. Numerous studies have reported important effects of the gut microbiota on pig production traits. However, whether the gut microbiota is associated with reproduction traits in boars remains largely unknown. Understanding the gut microbial composition and its influencing factors in large-scale boar populations is an essential first step to investigate this association. In this study, shotgun metagenomic sequencing was performed on fecal samples of 1,651 commercial boars from three breeds raised in three pig farms to uncover their gut microbial structures. We observed significant differences in boar gut microbial compositions across three breeds, even when raised in the same farm. Permutational multivariate analysis of variance (PERMANOVA) within-farm breeds and with-age stages found that the effect size of each factor on boar gut microbial composition varied across farms and age stages. Breeds accounted for 2% ~ 9% of the variance of boar gut microbial compositions in different farms. We then identified gut microbial taxa enriched in each boar breed using MaAsLin2. Lactic acid and butyrate-producing taxa, such as Lactobacillus amylovorus and Faecalibacterium prausnitzii, were enriched in Duroc boars; Akkermansia muciniphila and Lactobacillus reuteri showed the enrichment in Landrace boars, accompanied by increased relative abundance of Enterobacteriaceae members. Meanwhile, the species from Bacteroides, Prevotella, and Treponema had higher abundances in the gut of Large White pigs than in the other two pig breeds. We also identified bacterial species enriched in each of the three age stages. These breed and age-associated microbial enrichment patterns might reflect the combined effects of long-term genetic selection of pig breeds, age, and differences in feeding diets. The results of this study provide important insights for further investigating the effects of gut microbiota on boar reproductive traits and for developing strategies to modulate the gut microbiota to improve boar health and production performance.