Development of KISS1 knockout pigs is characterized by hypogonadotropic hypogonadism, normal growth, and reduced skatole.

Kisspeptin is a major regulator of gonadotropin secretion in pigs. Previously, CRISPR/Cas9 knockout of KISS1 was used to develop a mosaic parental line of pigs to generate offspring that would not need castration due to loss of kisspeptin. The current goal was to characterize growth and reproductive development of F1 pigs from this parental line. Body weights, gonadotropin concentrations and gonadal development were measured from birth through development (boars to 220 d of age, n = 42; gilts to 160 d of age, n = 36). Testosterone, skatole, and androstenone were also measured in boars. Blood samples were collected by jugular venipuncture for quantification of serum hormones, gonadal tissues collected for gross morphology and histology, and a fat biopsy collected (boars) for skatole and androstenone analysis. Body weight did not differ with genotype. There were no differences between KISS1+/+ and heterozygote KISS1+/- animals for most parameters measured. Gonadotropin concentrations were reduced in KISS1-/- boars and gilts compared with KISS1+/+ and KISS1+/- animals (P < 0.05). Concentrations of testosterone in serum and both androstenone and skatole in adipose were less in KISS1-/- boars than in KISS1+/+ and KISS1+/- boars (P < 0.05). Hypogonadism was in all KISS1-/- gilts and boars. These data indicate that knocking out KISS1 causes hypogonadotropic hypogonadism but does not negatively affect growth in pigs. Only one KISS1 allele is needed for normal gonadotropin secretion and gonadal development, and accumulation of compounds in adipose leading to boar taint.

Development of a highly reproducible GC-HRMS method for determination of Skatole in pig tissues.

This study presents a sensitive and reproducible mass spectrometry method for quantifying skatole in porcine adipose tissue, muscle, and serum samples applicable for abattoirs and laboratories. Leveraging gas chromatography-high-resolution Orbitrap microscopy and microwave-assisted liquefication of the adipose tissue, the method demonstrates robust performance across key parameters. Impressive linearity (R2) values of 0.9999 and 0.9996 for adipose tissue and serum, respectively. Notably, the method exhibits a low Limit of Detection (LoD) of 0.5 ppb for adipose tissue and 0.9 ppb for serum, with corresponding Limits of Quantification (LoQ) at 1.65 ppb and 3.04 ppb, respectively. The method showed significant reproducibility of 5.9% and repeatability (RSD%) of 8.78% for adipose tissue and 4.08% for serum, with recovery rates of 90% and 87%, respectively. This streamlined method offers promising, effective quantification of boar taint compounds, emphasizing its sensitivity and reproducibility.

Molecular Analysis of Indole and Skatole Decomposition Metabolism in Acinetobacter piscicola p38 Utilizing Biochemical and Omics Approaches.

Indole and skatole (3-methylindole, C9H9N) are common nitrogen-containing heterocyclic pollutants found in waste, wastewater treatment plants, and public restrooms and are the most notorious compounds in animal feces. Biodegradation was considered a feasible method for the removal of indole and skatole, but a comprehensive understanding of the metabolic pathways under both aerobic and anaerobic conditions was lacking, and the functional genes responsible for skatole biodegradation remained a mystery. Through metagenomic and gene cluster functional analysis, Acinetobacter piscicola p38 (NCBI: CP167896), genes 1650 (styrene monooxygenase: ACDW34_08180), and 1687 (styrene monooxygenase: ACDW34_08350) were identified as having the potential to degrade indole and skatole. The heterologous expression results demonstrate that the genes 1650 and 1651 (flavin reductase: ACDW34_08185), when combined, are capable of degrading indole, while the genes 1687 and 1688 (flavin reductase: ACDW34_08355), in combination, can degrade indole as well as skatole. These reactions necessitate the involvement of flavin reductase and NAD(P)H to catalyze the oxygenation process. This work aimed to provide new experimental evidence for the biodegradation of indole and skatole. This study offered new insights into our understanding of skatole degradation. The Acinetobacter_piscicola p38 strain provided an effective bacterial resource for the bioremediation of fecal indole and skatole.

Hepatic Gene Expression and Metabolite Profiles of Androstenone and Skatole Relative to Plasma Estrone Sulfate Levels in Boars.

Testicular steroids can alter the activity and expression of enzymes within the liver and may influence the metabolism of skatole and androstenone, which are responsible for boar taint. Plasma levels of estrone sulfate (E1S) are indicative of the steroidogenic capacity of the boar and are variable between animals of similar live weights at slaughter. This study aimed to characterize the relationship between steroidogenic capacity and the metabolism of boar taint compounds by relating plasma E1S levels at slaughter weight to the expression levels of genes regulating the metabolism of androstenone and skatole, along with their respective metabolite profiles. RT-qPCR was used to evaluate gene expression in the liver. Hepatocytes were also isolated and treated with androstenone or skatole, with metabolite levels in the incubation media quantified by high-performance liquid chromatography. Plasma E1S levels ranged from 2.2-108.5 ng/mL and were positively correlated with overall skatole metabolism (p = 0.038), the production of metabolites 3-methyloxindole (p = 0.026) and 3-hydroxy-3-methyloxindole (p = 0.036), and expression levels of key genes involved in skatole metabolism, specifically CYP2C33 (p = 0.0042), CYP2C49 (p = 0.022), and CYB5R1 (p = 0.017). There was no association between androstenone metabolism and plasma E1S concentrations; however, there was evidence of possible co-regulation amongst genes involved in the metabolism of androstenone, skatole, and estrogens. These findings indicate that steroidogenic capacity is related to the rate of skatole, but not androstenone metabolism, in slaughter-weight boars.

The Effect of the Boar Taint Masking Strategy (Adding Dried Origanum vulgare or Allium sativum) on Sensory Characteristics.

With increasing efforts to ban surgical castration, it is important to find ways to mask the level of boar taint in meat. The aim of this study was to test the possibility of masking boar taint or skatole levels by adding dried Origanum vulgare or Allium sativum and to evaluate consumer sensory preferences towards the skatole concentration in different carcass parts (longissimus lumborum; semimembranosus; neck chop and subcutaneous fat) and the masking strategy (addition of Allium sativum or Origanum vulgare). In the first experiment, the effect of the masking strategy was evaluated at three different skatole concentrations (0.069, 0.269 and 0.463 µg/g). The results showed that the samples with low and medium skatole levels were significantly different between the control group and the groups treated with Origanum vulgare or Allium sativum. In both cases, the addition of Allium sativum and Origanum vulgare had a positive effect on the parameters of abnormal odour and pleasantness of odour (p < 0.05). According to the results of the second experiment, meat samples from leaner parts, such as the neck chop semimembranosus and longissimus lumborum, not treated with Allium sativum and Origanum vulgare for masking, were significantly (p < 0.05) worse in terms of the occurrence of boar taint or abnormal odour than the masked samples. No significant differences were found between the two masking methods.

Relationships between conformational and vibrational features of tryptophan characteristic Raman markers.

Tryptophan (Trp) residue provides characteristic vibrational markers to the middle wavenumber spectral region of the Raman spectra recorded from peptides and proteins. In this report, we were particularly interested in eight Trp Raman markers, referred to as Wi (i = 1,…,8). All responsible for pronounced Raman lines, these markers originate from indole moiety, a bicyclic conjugated segment involved in the Trp structure. Numerous investigations have previously attempted to relate the variations observed in the spectral features of these markers to the environmental changes of Trp residues. To emphasize the most important points we can mention (i) the variations in the Raman profile of W4 (∼1360 cm-1) and W5 (∼1340 cm-1), frequently observed as a doublet with variable intensity ratio. These two markers were thought to result from a Fermi-resonance effect between certain planar and nonplanar modes; (ii) the changes observed in the wavenumbers and relative intensities of W4, W7 (∼880 cm-1) and W8 (∼760 cm-1) were supposed to be related to the accessibility of Trp to surrounding water molecules; and (iii) the wavenumber fluctuations of W3 (∼1550 cm-1), taken as a Trp side chain orientational marker. However, some ambiguities still exist regarding the interpretation of these markers, needing further clarification. Herein, upon a joint experimental and theoretical analysis based on a multiconformational approach, attention was paid to the relationships between structural and vibrational features of three indole-containing compounds with increasing structural complexity, i.e., skatole (3-methylindole), tryptophan, and tripeptide Gly-Trp-Gly. This study clearly shows that the existing assignments given to certain Trp Raman markers should be reconsidered, especially those based on the Fermi-resonance origin of W4-W5 (∼1360-1340 cm-1) doublet, as well as the purely environmental dependence of W7 and W8 markers.

Enhanced HPLC Method for Boar Taint Quantification.

Boar taint is an unpleasant odour found in the carcasses of entire male pigs, resulting from androstenone and skatole accumulation during pubertal development, and impacting pork quality. This study proposes the validation of an adapted chromatographic method for quantifying skatole and androstenone in the pigs' liquid fat using fluorescence detection. A good chromatographic separation was achieved, with skatole (SKA) and androstenone (AND) elution at 4.4 and 9.9 min., respectively. An external calibration method was applied, with calibration curves correlation coefficient of 0.9999 for both analytes. Detection limit values were 1.53 and 16.02 ng/g for SKA and AND, respectively. SKA recovery was 99.72±2.34 % (2.34 % RSD) and 102.84±1.62 % (1.57 % RSD) for AND. Results showed good precision values (repeatability <2.46 % RSD for SKA, <6.85 % RSD for AND; intermediate precision <2.87 % RSD for SKA, <6.98 % RSD for AND). The method's robustness was tested and the values were within the reference ranges. The validation results proved that the adaptation of an existing method resulted in good assessments of robustness, reliability and accuracy.

Effects of dietary selenized glucose on intestinal microbiota and tryptophan metabolism in rats: Assessing skatole reduction potential.

3-Methylindole (Skatole), a degradation product of tryptophan produced by intestinal microbial activity, significantly contributes to odor nuisance. Its adverse effects on animal welfare, human health, and environmental pollution have been noted. However, it is still unclear whether the intestinal microbiota mediates the impact of selenium (Se) on skatole production and what the underlying mechanisms remain elusive. A selenized glucose (SeGlu) derivative is a novel organic selenium compound. In this study, a diverse range of dietary SeGlu-treated levels, including SeGlu-deficient (CK), SeGlu-adequate (0.15 mg Se per L), and SeGlu-supranutritional (0.4 mg Se per L) conditions, were used to investigate the complex interaction of SeGlu on intestinal microbiome and serum metabolome changes in male Sprague-Dawley (SD) rats. The study showed that SeGlu supplementation enhanced the antioxidant ability in rats, significantly manifested in the increases of the activity of catalase (CAT) and glutathione peroxidase (GSH-Px), while no change in the level of malonaldehyde (MDA). Metagenomic sequencing analysis verified that the SeGlu treatment group significantly increased the abundance of beneficial microorganisms such as Clostridium, Ruminococcus, Faecalibacterium, Lactobacillus, and Alloprevotella while reducing the abundance of opportunistic pathogens such as Bacteroides and Alistipes significantly. Further metabolomic analysis revealed phenylalanine, tyrosine, and tryptophan biosynthesis changes in the SeGlu treatment group. Notably, the biosynthesis of indole, a critical pathway, was affected by SeGlu treatment, with several crucial enzymes implicated. Correlation analysis demonstrated strong associations between specific bacterial species - Treponema, Bacteroides, and Ruminococcus, and changes in indole and derivative concentrations. Moreover, the efficacy of SeGlu-treated fecal microbiota was confirmed through fecal microbiota transplantation, leading to a decrease in the concentration of skatole in rats. Collectively, the analysis of microbiota and metabolome response to diverse SeGlu levels suggests that SeGlu is a promising dietary additive in modulating intestinal microbiota and reducing odor nuisance in the livestock and poultry industry.

A Preliminary Evaluation of Sex and Dietary Field Pea Effects on Sensory Characteristics of Dry-Cured Loins.

Two of the main issues related to cured meat products are castration to avoid boar taint and the hefty reliance on soybean meal to feed pigs. However, data on the effects of immunocastration in pigs and alternative crop protein feeds on the sensory traits and consumers' acceptance of dry-cured loin are still limited. A preliminary study was conducted on the effect of sex type (surgical castrated male pigs and immunocastrated male and female pigs, at approximately 140 kg in weight and 7 months of age) and animal diet (pea vs. soya) on dry-cured loins. The study involved a sensory evaluation of six treatments, with a 3 × 2 factorial design, conducted by trained panellists and untrained consumers in Spain (n = 126) and Portugal (n = 80). The consumers were also checked for their skatole sensitivity using a pure substance. The results showed that the pea-based diet was significantly different (p < 0.001) from the soy-based diet, as determined by a trained panel of surgically castrated male pigs, although the immunocastration treatments were not split. Dry-cured loin from immunocastrated male pigs fed with peas was considered tenderer and juicier (p < 0.05) than those fed soya-based diets. The untrained consumer panel scored higher on the sensory traits (flavour, juiciness, and overall liking) from the loin samples of surgically castrated male pigs fed a pea diet. All consumers scored similarly in boar taint detection. This study demonstrates the applicability of the pea-based diet for the feeding of pigs destined for cured meat and highlights immunocastration as a tool that does not compromise the skatole detection score.

Magnolol-driven microbiota modulation elicits changes in tryptophan metabolism resulting in reduced skatole formation in pigs.

Skatole of gut origin has garnered significant attention as a malodorous pollutant due to its escalating emissions, recalcitrance to biodegradation and harm to animal and human health. Magnolol is a health-promoting polyphenol with potential to considerably mitigate the skatole production in the intestines. To investigate the impact of magnolol and its underlying mechanism on the skatole formation, in vivo and in vitro experiments were conducted in pigs. Our results revealed that skatole concentrations in the cecum, colon, and faeces decreased by 58.24% (P = 0.088), 44.98% (P < 0.05) and 43.52% (P < 0.05), respectively, following magnolol supplementation. Magnolol supplementation significantly decreased the abundance of Lachnospira, Faecalibacterium, Paramuribaculum, Faecalimonas, Desulfovibrio, Bariatricus, and Mogibacterium within the colon (P < 0.05). Moreover, a strong positive correlation (P < 0.05) between skatole concentration and Desulfovibrio abundance was observed. Subsequent in silico studies showed that magnolol could dock well with indolepyruvate decarboxylase (IPDC) within Desulfovibrio. Further in vitro investigation unveiled that magnolol addition led to less indole-3-pyruvate diverted towards the oxidative skatole pathway by the potential docking of magnolol towards IPDC, thereby diminishing the conversion of substrate into skatole. Our findings offer novel targets and strategies for mitigating skatole emission from the source.

Mulberry leaf supplementation inhibits skatole deposition by regulating gut microbiota and upregulating liver cytochrome P450 1A1 expression in finishing pigs.

Skatole, a strong fecal odor substance, is generated through microbial degradation of tryptophan in the animal hindgut. It easily accumulates in adipose tissue and affects meat quality. In this study, the effect of mulberry leaf supplementation on skatole in finishing pigs was studied. In a 35-day trial, 20 finishing pigs (barrows and gilts) were fed with a basal diet or basal diet with 6% mulberry leaves. Growth performance of the pigs (n = 10) was automatically recorded by a performance-testing feeder system and 8 pigs in each treatment were slaughtered and sampled for the remaining tests. Skatole and short-chain fatty acids were detected using HPLC and gas chromatography, respectively. Fecal microbiota were analyzed using 16S rRNA gene sequencing. The metabolomics analysis of feces and serum was performed with UHPLC-MS/MS. The major cytochrome P450 (CYP) enzymes that catalyze skatole degradation in the liver were tested by using RT-PCR and Western blot. Effects of major bioactive compounds in mulberry leaves on the CYP genes were verified in the hepatic cell line HepG2 in an in vitro test (n = 3). In finishing pigs, mulberry leaf supplementation had no significant effect on the average daily gain, average daily feed intake, and feed conversion ratio (P > 0.05), but reduced skatole levels in feces, serum, and backfat (P < 0.05), and increased acetic acid levels in feces (P = 0.027). Mulberry leaf supplementation decreased the relative abundance of the skatole-producing bacteria Megasphaera and Olsenella (P < 0.05). Indole-3-acetic acid, the intermediate that is essential for skatole production, was significantly reduced in feces by mulberry leaf supplementation (P < 0.05) and was positively correlated with skatole content in feces (P = 0.004). In pigs treated with mulberry leaves, liver CYP1A1 expression was increased (P < 0.05) and was negatively correlated with skatole content in backfat (P = 0.045). The in vitro test demonstrated that mulberry leaf polyphenols and polysaccharides could directly stimulate CYP1A1 expression in hepatic cells. These findings suggest that mulberry leaf supplementation reduces skatole production and deposition in finishing pigs by regulating the gut microbiota and promoting skatole degradation in liver.

Genetic Correlations between Boar Taint Compound Concentrations in Fat of Purebred Boars and Production and Ham Quality Traits in Crossbred Heavy Pigs.

Selecting pigs with reduced ability to accumulate boar taint (BT) compounds in their tissues is an alternative to male surgical castration. As the majority of slaughter pigs are crossbred, before selecting against BT in purebreds, it is essential to consider possible impacts on commercial traits in crossbreds. This study estimated the genetic correlations between BT compound levels measured in 1115 purebred pigs and carcass and ham quality traits collected in 26,577 crossbred Italian heavy pigs. Genetic correlations were estimated in bivariate Bayesian analyses including one BT trait and one production or ham quality trait at a time. Heritability of androstenone, skatole, and indole was 0.41, 0.49, and 0.37, respectively. A moderate negative correlation between skatole and carcass yield (-0.40), and between all BT compounds and backfat (from -0.26 to -0.55) was observed. Conversely, positive correlations (from 0.11 to 0.54) were found between skatole and ham fat thickness traits. Correlations between BT compounds and iodine number ranged from -0.07 (for androstenone) to -0.64 (for skatole), whereas those with PUFA ranged from -0.13 (for indole) to -0.33 (for skatole). Hence, reducing BT could decrease ham fat thickness and increase unsaturated fatty acids, with potential negative impacts on product quality.

Genomic Prediction and Genome-Wide Association Study for Boar Taint Compounds.

With a perspective future ban on surgical castration in Europe, selecting pigs with reduced ability to accumulate boar taint (BT) compounds (androstenone, indole, skatole) in their tissues seems a promising strategy. BT compound concentrations were quantified in the adipose tissue of 1075 boars genotyped at 29,844 SNPs. Traditional and SNP-based breeding values were estimated using pedigree-based BLUP (PBLUP) and genomic BLUP (GBLUP), respectively. Heritabilities for BT compounds were moderate (0.30-0.52). The accuracies of GBLUP and PBLUP were significantly different for androstenone (0.58 and 0.36, respectively), but comparable for indole and skatole (~0.43 and ~0.47, respectively). Several SNP windows, each explaining a small percentage of the variance of BT compound concentrations, were identified in a genome-wide association study (GWAS). A total of 18 candidate genes previously associated with BT (MX1), reproduction traits (TCF21, NME5, PTGFR, KCNQ1, UMODL1), and fat metabolism (CTSD, SYT8, TNNI2, CD81, EGR1, GIPC2, MIGA1, NEGR1, CCSER1, MTMR2, LPL, ERFE) were identified in the post-GWAS analysis. The large number of genes related to fat metabolism might be explained by the relationship between sexual steroid levels and fat deposition and be partially ascribed to the pig line investigated, which is selected for ham quality and not for lean growth.

Change in fat skatole and indole content in lambs switched from a concentrate-based diet indoors to alfalfa grazing for various durations before slaughter.

Meat from lambs fattened on alfalfa is at risk of excessive pastoral flavours due to high levels of in-fat volatile indolic compounds (especially, skatole). Skatole has also been identified as a potential marker of interest for authenticating pasture-fed lamb meat. Here, we investigated the change in skatole and indole concentrations in kidney fat from lambs switched from an indoor-fed concentrate based diet to outdoor alfalfa grazing for various durations (0, 21, 42, 63 days) before slaughter. The study used a total of 219 lambs over 3 consecutive years. Kidney-fat skatole and indole concentrations increased from as early as 21 days on alfalfa, and then reached a plateau. Similarly, the proportion of lambs that had a kidney fat-skatole concentration above 0.15 µg/g liquid fat, a value that has been established as a sensory rejection threshold for pork, increased significantly from as early as 21 days on alfalfa and then reached a plateau. This value was reached or exceeded in a significant proportion of lambs fattened on alfalfa pastures (45.1%). However, skatole was not detected in kidney fat from 20 out of 164 alfalfa-fattened lambs (i.e., 12.2%) but was detected in 15 out of 55 concentrate-fed lambs (i.e., 27.3%). We thus conclude that while skatole content in kidney fat can inform on dietary changes made shortly before slaughter, it does not have the discrimination power needed to reliably authenticate pasture-fed lamb meat, let alone duration of finishing on pasture.

Microbiome-metabolomics analysis reveals abatement effects of itaconic acid on odorous compound production in Arbor Acre broilers.

BACKGROUND: Public 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. RESULTS: Dietary 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. CONCLUSIONS: Itaconic 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.

Outdoor Finishing of Intact Male Portuguese Alentejano Pigs on a Sustainable High-Fiber Diet: Impacts on Blood, Growth, Carcass, Meat Quality and Boar Taint Compounds.

This trial evaluated the effect of sex on the blood, growth, carcass, meat quality, and boar taint compounds in male Alentejano (AL) pigs (n = 30). From ~40 to 130 kg LW, castrated (C) and intact pigs (I and IExp groups) were fed commercial diets ad libitum. Between ~130 and 160 kg (slaughter), C and I pigs continued on commercial diets, while IExp were fed an experimental diet containing locally produced pulses and by-products aimed at reducing boar taint. At ~160 kg, blood urea levels were higher in IExp than C pigs, triacylglycerols were lower in both intact groups, and cortisol was lower in IExp. IExp pigs exhibited faster growth, improved feed conversion ratio, carcass higher commercial yield and leaner meat than C pigs. The loin intramuscular fat in intact pigs was lower, less saturated and more polyunsaturated, while total collagen was higher. Fat androstenone content was higher in intact pigs and skatole content was similar across treatments, although they were below threshold values for consumer detection. Finally, although boar taint compounds were low in intact AL pigs raised outdoors, adding pulses and by-products to the experimental diet did not result in a reduction in fat skatole content compared to pigs fed the commercial diet.

Toxicological evaluation of a nonlethal riot control combinational formulation upon dermal application using animal models.

Numerous adverse effects on human health have been reported in epidemiological studies of oleoresin capsicum (OC) and other riot control agents (RCAs). Importantly, the daunting risk of such RCAs can be neutralized by optimizing the desired concentration of such agents for mob dispersal. Hence, a nonlethal riot control combinational formulation (NCF) was prepared for dispersing rioters without imparting fatal outcomes. However, for desired utilization of NCF, it is essential to recognize its extent of potential toxicity. Therefore, the current investigation evaluated the dermal toxicity of NCF using experimental animals in compliance with the OECD guidelines. Additionally, few essential metal ions were analyzed and found non -significantly different in the test rats as compared to control rats. Moreover, abnormal dermal morphology and lesions ultrastructural tissue defects were not noticed as evinced by different studies like ultrasonography, histology, and scanning electron microscopy (SEM) respectively. Further, Doppler ultrasonography exhibited non-significantly different blood flow velocity in both groups, whereas miles test demonstrated a significantly increased Evans blue concentration in test rats compared to the control rats, which might be due to an initial increase in blood flow via an instant action of the NCF at the cutaneous sensory nerve endings. However, our results demonstrated NCF can produce initial skin irritating and sensitizing effects in guinea pigs and rabbits without the antecedence of acute toxicity (≤2000 mg/kg) in Wistar rats.

Effects of fermented feed on growth performance, immune organ indices, serum biochemical parameters, cecal odorous compound production, and the microbiota community in broilers.

The aim of this study was to explore the effects of dietary fermented feed addition on growth performance, immune organ indices, serum biochemical parameters, cecal odorous compound production, and the bacterial community in broilers. A total of 480 broiler chicks (1-day-old) were randomly assigned to 6 groups, including a basal diet (control group), a basal diet supplemented with 10, 15, 20, and 25% dried fermented feed, and 10% wet fermented feed. Each group contained 8 replicates of 10 chicks each. The results showed that fermentation increased (P < 0.05) the total acid level and the number of Lactobacillus, Yeast, and Bacillus. The 15% dried fermented feed group had an increased (P < 0.05) body weight (BW) than the control, while the 25% dried fermented feed group had the lowest (P < 0.05) BW on 42 d. Compared to the control group, the feed intake (FI) was increased (P < 0.05) in the 10, 15% dried and 10% wet fermented feed groups from 22 to 42 d and from 1 to 42 d. No significant difference (P > 0.05) was observed in feed conversion ratio (FCR) among all groups. Supplementation with fermented feed increased (P < 0.05) the bursa of Fabricius index but not (P > 0.05) the thymus and spleen indices. Compared with the control, the broilers fed fermented feed had increased (P < 0.05) serum total protein, albumin, globulin, IgA, IgG, IgM, lysozyme, complement 3, and complement 4 levels. The cecal concentrations of acetic acid, propionic acid, butyric acid, and lactic acid were increased and the pH values were decreased in the fermented feed groups (P < 0.05). Among the groups, the 15% dried fermented feed group showed the lowest concentrations of skatole and indole in the cecum (P < 0.05). The composition of the cecal microbiota was characterized, in which an increased abundance of Ruminococcaceae, Lactobacillaceae, and unclassified Clostridiales and a decreased abundance of Rikenellaceae, Lachnospiraceae, and Bacteroidaceae were found in the fermented feed groups. Taken together, dietary fermented feed supplementation can improve growth performance, immune organ development, and capacity and decrease cecal odorous compound production, which may be related to the regulation of microbial composition.

Increasing sustainability in pork production by using high inclusion levels of co-products distillers dried grains with solubles, wheat middling and canola meal doesn't affect pig growth performance and meat quality but reduces boar taint.

OBJECTIVE: The present study is to examine the effect of high inclusion of co-products in pig diets (referred to as an alternative diet) during the finishing stage on pig growth performance, meat quality and boar taint compounds. METHODS: Growing pigs were fed an alternative diet made with distillers dried grains with solubles (25%), canola meal (20%), and wheat middling (15%) or a control diet based on barley and soybean meal to investigate the impact of co-products on pig performance and meat quality. Sixteen female and sixteen entire male Duroc×(Large White×Landrace) pigs (22.6±2.07 kg, body weight±standard error) were equally allocated to the diets. RESULTS: Pigs fed the alternative diet had a lower feed intake; however, growth rate and feed conversion efficiency were unaffected by diet. A diet by sex interaction was found for gain:feed whereby males fed the alternative diet had the best feed conversion (p<0.01). Pork from pigs fed the alternative diet had lower a* and Chroma and protein % (p<0.05), while other meat quality characteristics were unaffected. The alternative diet reduced backfat skatole levels (p<0.001). CONCLUSION: A diet containing high inclusion levels of co-products can be fed to pigs during the finishing stage without detrimental effects on pig performance or meat quality and with the potential to enhance pork flavour. This finding suggests a solution to increase the sustainable development of pig production.

Natural Product Skatole Ameliorates Lipotoxicity-Induced Multiple Hepatic Damage under Hyperlipidemic Conditions in Hepatocytes.

Skatole (3-methylindole, 3MI) is a natural-origin compound derived from plants, insects, and microbial metabolites in human intestines. Skatole has an anti-lipid peroxidation effect and is a biomarker for several diseases. However, its effect on hepatocyte lipid metabolism and lipotoxicity has not been elucidated. Hepatic lipotoxicity is induced by excess saturated free fatty acids in hyperlipidemia, which directly damages the hepatocytes. Lipotoxicity is involved in several metabolic diseases and hepatocytes, particularly affecting nonalcoholic fatty liver disease (NAFLD) progression. NAFLD is caused by the accumulation of fat by excessive free fatty acids (FFAs) in the blood and is accompanied by hepatic damage, such as endoplasmic reticulum (ER) stress, abnormal glucose and insulin metabolism, oxidative stress, and lipoapoptosis with lipid accumulation. Hepatic lipotoxicity causes multiple hepatic damages in NAFLD and has a directly effect on the progression from NAFLD to nonalcoholic steatohepatitis (NASH). This study confirmed that the natural compound skatole improves various damages to hepatocytes caused by lipotoxicity in hyperlipidemic conditions. To induce lipotoxicity, we exposed HepG2, SNU-449, and Huh7 cells to palmitic acid, a saturated fatty acid, and confirmed the protective effect of skatole. Skatole inhibited fat accumulation in the hepatocytes, reduced ER and oxidative stress, and recovered insulin resistance and glucose uptake. Importantly, skatole reduced lipoapoptosis by regulating caspase activity. In conclusion, skatole ameliorated multiple types of hepatocyte damage induced by lipotoxicity in the presence of excess free fatty acids.