Effect of a blend of cinnamaldehyde, eugenol, and Capsicum oleoresin on methane emission and lactation performance of Holstein-Friesian dairy cows.

This study aimed to investigate the effect of administering a standardized blend of cinnamaldehyde, eugenol, and Capsicum oleoresin (CEC) to lactating dairy cattle for 84 d (i.e., 12 wk) on enteric CH4 emission, feed intake, milk yield and composition, and body weight. The experiment involved 56 Holstein-Friesian dairy cows (145 ± 31.1 d in milk at the start of the trial; mean ± standard deviation) in a randomized complete block design. Cows were blocked in pairs according to parity, lactation stage, and current milk yield, and randomly allocated to 1 of the 2 dietary treatments: a diet including 54.5 mg of CEC/kg of DM or a control diet without CEC. Diets were provided as partial mixed rations in feed bins, which automatically recorded individual feed intake. Additional concentrate was fed in the GreenFeed system that was used to measure emissions of CO2, CH4, and H2. Feeding CEC decreased CH4 yield (g/kg DMI) by on average 3.4% over the complete 12-wk period and by on average 3.9% from 6 wk after the start of supplementation onward. Feeding CEC simultaneously increased feed intake and body weight, and tended to increase milk protein content, whereas no negative responses were observed. These results must be further investigated and confirmed in longer-term in vivo experiments.

Haemonchus contortus Adopt Isolate-Specific Life History Strategies to Optimize Fitness and Overcome Obstacles in Their Environment: Experimental Evidence.

Gastrointestinal nematodes (GIN) use flexible life history strategies to maintain their fitness under environmental challenges. Costs incurred by a challenge to one life trait can be recouped by increasing the expression of subsequent life traits throughout their life cycle. Anticipating how parasites respond to the challenge of control interventions is critical for the long-term sustainability of the practice and to further ensure that the parasites withstand favourable adaptive responses. There is currently limited information on whether distinct populations of a GIN species respond to the same environmental challenge in a consistent manner, with similar alterations to their life history strategies or comparable fitness outcomes. This study compared the life history traits and experimental fitness of three distinct Haemonchus contortus isolates exposed to environmental challenges at both the parasitic (i.e., passage through resistant or susceptible sheep) and free-living (i.e., exposure to diverse climatic conditions) life stages. The key findings show that H. contortus maintain their fitness under challenge with isolate-specific alterations to their life history strategies. Further, partial exploration of the H. contortus isolates transcriptomes using cDNA-AFLP methods confirmed disparate expression profiles between them. These results bring fresh insights into our understanding of the non-genetic adaptive processes of GIN that may hinder the efficacy of parasite control strategies.

Garlic-Derived Metabolites Exert Antioxidant Activity, Modulate Gut Microbiota Composition and Limit Citrobacter rodentium Infection in Mice.

The garlic-derived compounds propyl propane thiosulfinate (PTS) and propyl propane thiosulfonate (PTSO) are metabolites with putative health benefits against intestinal inflammation that may be related to their antioxidant activity. However, the underlying mechanisms remain unclear, and whether PTS-PTSO can promote gut health by altering the microbiota and exert protection against enteric pathogens needs further investigation. Here, we explored the antioxidant activity of PTS-PTSO in murine macrophages in vitro, and in an in vivo model of bacterial infection with the bacterial pathogen Citrobacter rodentium. PTS-PTSO attenuated reactive oxygen species in lipopolysaccharide-stimulated macrophages in a nuclear factor erythroid factor 2-related factor 2 (Nrf2)-dependent manner, decreased nitric oxide levels both in macrophages in vitro and in the sera of mice fed PTS-PTSO, and had putatively beneficial effects on the commensal gut microbiota. Importantly, PTS-PTSO decreased faecal C. rodentium counts, concomitant with upregulation of Nrf2-related genes in colon tissue. Thus, PTS-PTSO mediates Nrf2-mediated antioxidant activity and modulates gut microbiota, which may protect the host against C. rodentium colonization. Our results provide further insight into how PTS-PTSO and related bioactive dietary compounds may reduce enteric infections.

Effect of dietary magnolia bark extract supplementation in finishing pigs on the oxidative stability of meat.

BACKGROUND: Magnolia bark extract (MBE) is a natural supplement with antioxidant, anti-inflammatory, and antimicrobial activities. Its properties suggest that the dietary supplementation in livestock could improve the quality of products. Therefore, the aim of this study was to investigate, for the first time, the effect of dietary MBE supplementation (0.33 mg/kg) in finishing pigs on the oxidative stability of meat. Oxidative stability is of paramount importance for pork, as it affects storage, retail, and consumer acceptance. For the purpose, the fatty acid profile, cholesterol, fat-soluble vitamins, antioxidant enzymes (catalase, glutathione peroxidase, and superoxide dismutase), non-enzymatic antioxidant capacity (TEAC, FRAP, and Folin-Ciocalteu assays), color stability, and lipid stability of pork were assessed. RESULTS: Concerning carcass characteristics, dietary MBE did not affect cold carcass yield, but reduced (P = 0.040) the chilling weight loss. The meat from pigs fed MBE had a lower (P = 0.031) lightness index than the control meat. No effect on intramuscular fat, cholesterol, and fatty acid profile was observed. Dietary MBE did not affect the content of vitamin E (α-tocopherol and γ-tocopherol) in pork, whereas it reduced (P = 0.021) the retinol content. The catalase activity was 18% higher (P = 0.008) in the meat from pigs fed MBE compared with the control group. The MBE supplementation reduced (P = 0.039) by 30% the thiobarbituric acid reactive substances (TBARS) in raw pork over 6 d of aerobic refrigerated storage. Instead, no effect on lipid oxidation was observed in cooked pork. Last, the meat from pigs fed MBE reduced Fe3+-ascorbate catalyzed lipid oxidation in muscle homogenates, with a lower (P = 0.034) TBARS value than the control group after 60 min of incubation. CONCLUSIONS: Dietary MBE supplementation in finishing pigs delayed the lipid oxidation in raw meat. This effect was combined with an increased catalase concentration. These results suggest that dietary MBE could have implications for improving the shelf-life of pork.

Garlic-Derived Organosulfur Compounds Regulate Metabolic and Immune Pathways in Macrophages and Attenuate Intestinal Inflammation in Mice.

SCOPE: Garlic is a source of bioactive phytonutrients that may have anti-inflammatory or immunomodulatory properties. The mechanism(s) underlying the bioactivity of these compounds and their ability to regulate responses to enteric infections remains unclear. METHODS AND RESULTS: This study investigates if a garlic-derived preparation (PTSO-PTS) containing two organosulfur metabolites, propyl-propane thiosulfonate (PTSO), and propyl-propane thiosulfinate (PTS), regulate inflammatory responses in murine macrophages and intestinal epithelial cells (IEC) in vitro, as well as in a model of enteric parasite-induced inflammation. PTSO-PTS decreases lipopolysaccharide-induced secretion of TNFα, IL-6, and IL-27 in macrophages. RNA-sequencing demonstrates that PTSO-PTS strongly suppresses pathways related to immune and inflammatory signaling. PTSO-PTS induces the expression of a number of genes involved in antioxidant responses in IEC during exposure to antigens from the parasite Trichuris muris. In vivo, PTSO-PTS does not affect T. muris establishment or intestinal T-cell responses but significantly alters cecal transcriptomic responses. Notably, a reduction in T. muris-induced expression of Tnf, Saa2, and Nos2 is observed. CONCLUSION: Garlic-derived organosulfur compounds exert anti-inflammatory effects in macrophages and IEC, and regulate gene expression during intestinal infection. These compounds and related organic molecules may thus hold potential as functional food components to improve gut health in humans and animals.

Effect of different levels of organic zinc supplementation on pork quality.

This study investigated the effect of two supplementation levels of zinc glycinate (ZnGly) on performance, carcass characteristics, and meat quality of growing-finishing pigs. Thirty pigs (bodyweight: 61 ± 4.0 kg) were assigned to three treatments and fed ad libitum for 56 days a diet supplemented with 0 (control), 45 (Zn45), or 100 mg/kg (Zn100) of ZnGly. The highest ZnGly supplementation lowered the average daily gain (P = 0.031); while, cold carcass weight did not differ between treatments. Both ZnGly levels reduced carcass chill loss (P < 0.001). Micromineral content, color stability, and fatty acid profile of meat were not altered by ZnGly. Superoxide dismutase activity was lowered by Zn45 compared to control (P = 0.007); while, catalase activity was enhanced by Zn100 (P = 0.003). Although ZnGly supplementation did not influence lipid oxidation in raw meat and in meat homogenates incubated with pro-oxidant catalysts, Zn45 limited lipid oxidation in cooked meat (P = 0.037). Our results demonstrated that supplementing pigs with 45 mg/kg of ZnGly could improve the oxidative stability of pork subjected to strong pro-oxidant conditions, but this effect needs to be further elucidated.

Potential of a mixture of eugenol and garlic tincture to improve performance and intestinal health in broilers under necrotic enteritis challenge.

Plant extracts (PE) are gaining increased attention as potential alternatives to in-feed antimicrobials (AM) due to their known antimicrobial activities. This study was conducted to examine the potential of PE, a microencapsulated product composed of eugenol and garlic tincture as an alternative to AM-agent on performance and intestinal health in broilers under necrotic enteritis (NE) challenge. A total of 960 day-old mixed-sex Cobb 500 chicks were randomly distributed to 48-floor pens with 6 treatments replicated 8 times with 20 birds each. The 6 treatments were as follows: UC, unchallenged control; CC, challenged control; PE, challenged group plus PE; AM, challenged group plus AM; FAP, challenged group plus a full dose of AM with PE; HAP, challenged group plus a half dose of AM with PE in starter, grower and finisher phases. Birds in the challenged groups were inoculated with Eimeria spp. on d 9 and Clostridium perfringens on d 14. The body weight gain (BWG), feed intake (FI), feed conversion ratio (FCR), and livability of birds were compromised, and intestinal lesions and mortality were increased (P < 0.05) by NE challenge, illustrating a successful clinical NE challenge. Birds fed AM had higher BWG and FI, and lower FCR, mortality, and intestinal lesions compared to the CC group (P < 0.05). Birds fed PE had improved FCR (P < 0.05) and livability (5.8%) in an overall period compared to the CC group. On d 16, PE supplementation reduced ileal lesion scores in only male birds (P < 0.05). Birds fed PE had decreased Eimeria maxima and Eimeria acervulina oocyst counts in caecal content (P < 0.05). Birds fed PE had decreased Escherichia brunetti and total oocyst counts in caecal content, and E. acervulina oocyst counts in ileal content in only female birds (P < 0.05). On d 35, PE supplementation reduced variation of BW in both male and female birds and increased yellowness (b∗ value, 14.4%) in the thigh. These findings suggest the potential of PE supplementation in diets to improve the performance and intestinal health of birds under clinical NE as indicated by improved FCR, livability, uniformity, reduced ileal lesions, oocyst counts and increased skin yellowness. However, the protective effect of PE may not be apparent in the presence of AM in the feed.

The phytonutrient cinnamaldehyde limits intestinal inflammation and enteric parasite infection.

Phytonutrients such as cinnamaldehyde (CA) have been studied for their effects on metabolic diseases, but their influence on mucosal inflammation and immunity to enteric infection are not well documented. Here, we show that consumption of CA in mice significantly down-regulates transcriptional pathways connected to inflammation in the small intestine, and alters T-cell populations in mesenteric lymph nodes. During infection with the enteric helminth Heligomosomoides polygyrus, CA treatment attenuated infection-induced changes in biological pathways connected to cell cycle and mitotic activity, and tended to reduce worm burdens. Mechanistically, CA did not appear to exert activity through a prebiotic effect, as CA treatment did not significantly change the composition of the gut microbiota. Instead, in vitro experiments showed that CA directly induced xenobiotic metabolizing pathways in intestinal epithelial cells and suppressed endotoxin-induced inflammatory responses in macrophages. Collectively, our results show that CA down-regulates inflammatory pathways in the intestinal mucosa and can limit the pathological response to enteric infection. These properties appear to be largely independent of the gut microbiota, and instead connected to the ability of CA to induce antioxidant pathways in intestinal cells. Our results encourage further investigation into the use of CA and related phytonutrients as functional food components to promote intestinal health in humans and animals.

Effect of raw and encapsulated policosanol on lipid profiles, blood biochemistry, activity, energy expenditure and macronutrient metabolism of adult cats.

OBJECTIVES: The objective of this study was to verify the safety of policosanol supplementation for domestic cats. The effects of raw and encapsulated policosanol were compared with positive (L-carnitine) and negative (no supplementation) controls on outcomes of complete blood count, serum biochemistry, energy expenditure, respiratory quotient and physical activity in healthy young adult cats. METHODS: The study was a replicated 4 × 4 complete Latin square design. Eight cats (four castrated males, four spayed females; mean age 3.0 ± 1.0 years; mean weight 4.36 ± 1.08 kg; mean body condition score 5.4 ± 1.4) were blocked by sex and body weight then randomized to treatment groups: raw policosanol (10 mg/kg body weight), encapsulated policosanol (50 mg/kg body weight), L-carnitine (200 mg/kg body weight) or no supplementation. Treatments were supplemented to a basal diet for 28 days with a 1-week washout between periods. Food was distributed equally between two offerings to ensure complete supplement consumption (first offering) and measure consumption time (second offering). Blood collection (lipid profile, complete blood count, serum biochemistry) and indirect calorimetry (energy expenditure, respiratory quotient) were conducted at days 0, 14 and 28 of each period. Activity monitors were worn 7 days prior to indirect calorimetry and blood collection. Data were analyzed using a repeated measures mixed model (SAS, v.9.4). RESULTS: Food intake and body weight were similar among treatments. There was no effect of treatment on lipid profile, serum biochemistry, activity, energy expenditure or respiratory quotient (P >0.05); however, time to consume a second meal was greatest in cats fed raw policosanol (P <0.05). CONCLUSIONS AND RELEVANCE: These data suggest that policosanol is safe for feline consumption. Further studies with cats demonstrating cardiometabolic risk factors are warranted to confirm whether policosanol therapy is an efficacious treatment for hyperlipidemia and obesity.

Genetic × environment variation in sheep lines bred for divergent resistance to strongyle infection.

Drug-resistant parasites threaten livestock production. Breeding more resistant hosts could be a sustainable control strategy. Environmental variation linked to animal management practices or to parasite species turnover across farms may however alter the expression of genetic potential. We created sheep lines with high or low resistance to Haemonchus contortus and achieved significant divergence on both phenotypic and genetic scales. We exposed both lines to chronic stress or to the infection by another parasite Trichostrongylus colubriformis, to test for genotype-by-environment and genotype-by-parasite species interactions respectively. Between-line divergence remained significant following chronic stress exposure although between-family variation was found. Significant genotype-by-parasite interaction was found although H. contortus-resistant lambs remained more resistant against T. colubriformis. Growth curves were not altered by the selection process although resistant lambs were lighter after the second round of divergence, before any infection took place. Breeding for resistance is a sustainable strategy but allowance needs to be made for environmental perturbations and worm species.

A Microencapsulated Mixture of Eugenol and Garlic Tincture Supplementation Mitigates the Effect of Necrotic Enteritis on Intestinal Integrity and Increases Goblet Cells in Broilers.

This study was conducted to examine the effects of a plant extract mixture, a microencapsulated product composed of eugenol and garlic tincture (PE), on intestinal health in broilers under necrotic enteritis (NE) challenge. A total of 960 d-old mixed-sex Cobb 500 chicks were randomly distributed to 48-floor pens housing 20 birds per pen. Six treatments were applied: UC, unchallenged control; CC, challenged control; PE, challenged group plus PE; AM, challenged group plus antimicrobial (AM); FAP, challenged group plus a full dose of AM with PE; HAP, challenged group plus a half dose of AM with PE in starter, grower and finisher phases. Birds in the challenged groups were inoculated with Eimeria spp. on d 9 and Clostridiumperfringens on d 14. On d 16, the CC group had increased serum fluorescein isothiocyanate dextran (FITC-d), reduced villus surface area, goblet cell number, upregulated CLDN1, JAM2 genes and reduced microbial diversity compared to the UC group (p < 0.05). Birds fed PE had reduced FITC-d, increased goblet cell number and Bifidobacterium compared to the CC group (p < 0.05). Birds fed PE had reduced CLDN5 expression in male birds, and Bacteroides spp. in female birds than CC group (p < 0.05). These findings suggest that PE supplementation mitigates the effect of NE by improving the intestinal health of birds.

Integrative biology defines novel biomarkers of resistance to strongylid infection in horses.

The widespread failure of anthelmintic drugs against nematodes of veterinary interest requires novel control strategies. Selective treatment of the most susceptible individuals could reduce drug selection pressure but requires appropriate biomarkers of the intrinsic susceptibility potential. To date, this has been missing in livestock species. Here, we selected Welsh ponies with divergent intrinsic susceptibility (measured by their egg excretion levels) to cyathostomin infection and found that their divergence was sustained across a 10-year time window. Using this unique set of individuals, we monitored variations in their blood cell populations, plasma metabolites and faecal microbiota over a grazing season to isolate core differences between their respective responses under worm-free or natural infection conditions. Our analyses identified the concomitant rise in plasma phenylalanine level and faecal Prevotella abundance and the reduction in circulating monocyte counts as biomarkers of the need for drug treatment (egg excretion above 200 eggs/g). This biological signal was replicated in other independent populations. We also unravelled an immunometabolic network encompassing plasma beta-hydroxybutyrate level, short-chain fatty acid producing bacteria and circulating neutrophils that forms the discriminant baseline between susceptible and resistant individuals. Altogether our observations open new perspectives on the susceptibility of equids to strongylid infection and leave scope for both new biomarkers of infection and nutritional intervention.

Feed Concentrate Palatability in Welsh Ponies: Acceptance and Preference of Flavors.

In horses, it is well established that nutrients and the palatability of feed material (odor and taste) play an important role in diet selection. For example, high-fiber feed taste is not well accepted by horses. Consequently, manufacturers have begun to supplement horse feed with flavors to mask feed bitterness, to overcome feed neophobia and to encourage water drinking. However, only few studies have been performed to evaluate the acceptance and preference of flavors in horses. The aim of this study was to evaluate the acceptance and preference of flavors supplemented on top of concentrate offered to ponies. Thirty-three female Welsh ponies aged between four to 13 years were enrolled in the experiment. Ponies were offered 4 flavored concentrates and one control with no flavor. The flavored concentrates were anise, caramel, raspberry and apple. The inclusion rate of the flavors was 300 g/t on an as-is basis. During the adaptation period (one week), the ponies were gently guided to each bucket containing the flavored concentrate (200 g) during 10 sec/bucket for olfaction only. During the test period, ponies were allowed to freely choose flavored concentrates for 2 minutes. The flavors and the position of the buckets in front of the ponies were randomized. Each period was video-recorded and number of chews were counted during test period. The concentrate intake, eating time, and animal behavior were recorded during the test period. The apple concentrate was consumed the most at 116 g/2-min offering, whereas the raspberry and control concentrates were consumed the least, at 85.31 and 90.80 g/2-min offering, respectively. Apple flavor was preferred over caramel, raspberry and anise as indicated by higher consumption rate (g/sec) (chi-squared=16.68, df=4, P<0.05). There was no effects on chews, smell or headshaking time per sec between treatments. In conclusion, the ponies accepted a wide range of flavors with a preference for apple over raspberry flavored concentrate.

Organic selenium supplement partially alleviated diquat-induced oxidative insults and hepatic metabolic stress in nursery pigs.

The study investigated antioxidant effects of Se on resilience to diquat-induced oxidative stress in nursery pigs. Thirty-five weaned pigs were individually housed and randomly assigned to one of the five treatments. Pigs were (1) fed a basal diet and intraperitoneally injected with sterile saline (negative control), (2) fed the basal diet and injected with diquat solution (positive control, PC), or fed the basal diet supplemented with 0·3 mg Se/kg as (3) sodium selenite (SS), (4) soyabean protein-chelated Se (SC) or (5) selenised yeast (SY) and intraperitoneally injected with diquat. Pigs were fed the experimental diets for 17 d and injected with diquat at 10 mg/kg body weight or saline on the 11th day of the study (day 0 post-injection (PI)). Diquat exposure induced acute stress and innate immune activation (P < 0·05) at 6 h PI and compromised (P < 0·05) plasma glutathione peroxidase activity on day 2 PI, which was accompanied by an increase in plasma malondialdehyde at 6 h and day 2 PI (P < 0·10). Organic Se, particularly SY, enhanced (P < 0·05) endogenous antioxidant activity in various aspects compared with the PC group. The growth rate and feed intake from day 0 to day 7 PI were significantly lower in the PC, SS and SC groups than the NC group (P < 0·05). Untargeted metabolomics analysis revealed that twenty-two hepatic metabolites (false discovery rate < 0·15) associated with lipid and cellular antioxidant metabolism were altered by diquat. SY restored hepatic metabolic profiles in some but not all samples.

Standardized Plant Extract Alleviates the Negative Effects of FMD Vaccination on Animal Performance.

The present study was conducted to assess the efficacy of a plant extract (PE) on growth performance and immune status in foot and mouth disease (FMD)-vaccinated growing pigs. A total of 120 crossed ((Landrace × Yorkshire) × Duroc) growing pigs with an average initial body weight (BW) of 24.66 ± 2.34 kg and an average age of 70 days were randomized into three groups (10 pens; 4 pigs per pen per treatment) as follows: a nonvaccinated negative control group (NV), a FMD vaccinated group (OV), and a third group received a 0.0125% PE supplement after vaccination (PV), in a 6-week trial. The PV group receiving PE supplementation increased (p < 0.05) the BW compared with the OV group, and average daily gain (ADG) during days 1-14, overall and gain-to-feed ratio (G: F) in days 1-14, and dry matter (DM) digestibility at week 6 were higher (p < 0.05) in the PV compared with the OV group. A significant increase (p < 0.05) in haptoglobin concentration was observed in the OV group compared with the NV group at 25 days postvaccination. The inhibition percentage of antibodies against FMD in the sera reached above 50% in the PV group 5 days earlier than in the OV group. The findings suggest that the inclusion of PE in the diet promoted the performance of vaccinated growing pigs.

Dietary phytonutrients and animal health: regulation of immune function during gastrointestinal infections.

The composition of dietary macronutrients (proteins, carbohydrates, and fibers) and micronutrients (vitamins, phytochemicals) can markedly influence the development of immune responses to enteric infection. This has important implications for livestock production, where a significant challenge exists to ensure healthy and productive animals in an era of increasing drug resistance and concerns about the sector's environmental footprint. Nutritional intervention may ultimately be a sustainable method to prevent disease and improve efficiency of livestock enterprises, and it is now well established that certain phytonutrients can significantly improve animal performance during challenge with infectious pathogens. However, many questions remain unanswered concerning the complex interplay between diet, immunity, and infection. In this review, we examine the role of phytonutrients in regulating immune and inflammatory responses during enteric bacterial and parasitic infections in livestock, with a specific focus on some increasingly well-studied phytochemical classes-polyphenols (especially proanthocyanidins), essential oil components (cinnamaldehyde, eugenol, and carvacrol), and curcumin. Despite the contrasting chemical structures of these molecules, they appear to induce a number of similar immunological responses. These include promotion of mucosal antibody and antimicrobial peptide production, coupled with a strong suppression of inflammatory cytokines and reactive oxygen species. Although there have been some recent advances in our understanding of the mechanisms underlying their bioactivity, how these phytonutrients modulate immune responses in the intestine remains mostly unknown. We discuss the complex inter-relationships between metabolism of dietary phytonutrients, the gut microbiota, and the mucosal immune system, and propose that an increased understanding of the basic immunological mechanisms involved will allow the rational development of novel dietary additives to promote intestinal health in farmed animals.

Accurate quantification of metal-glycinates-sulphate complexes and free metals in feed by capillary electrophoresis inductively coupled plasma mass spectrometry.

Traceability of metal-glycinate-sulphate complexes (Metal-GLY) in feed requires specific analysis to differentiate complexes from inorganic forms. A previously described method focused on the quantification of Metal-GLY at one single concentration but not on the quantification of free metal ion forms. The objective of this work was to extend the method to quantify both Metal-GLY and free metal ion forms of various metals at low inclusion levels. A 50/50 w/w mix of corn flour and soybean meal was used as feed. Copper-glycinate(Cu-GLY), Manganese-glycinate (Mn-GLY) and Zinc-glycinate (Zn-GLY) complexes (provided by Pancosma SA) were used for in-feed inclusions. The feed metal background concentrations and species repartitions were assessed. Cu-GLY was spiked on feed at levels matching 5, 15 and 45 mg/kg, corresponding to metal concentrations of 1.2, 3.6 and 10.8 mg/kg. Mn-GLY and Zn-GLY were spiked at 15, 45 and 100 mg/kg, corresponding to 3.3, 9.9, 22 mg/kg Mn and 3.9, 11.7, 26mg/kg Zn, respectively. The water soluble fraction of un-supplemented feed contained 0.06 mg/kg Cu, 0.05 mg/kg Mn and 0.12 mg/kg Zn, with 69.5% of Cu, 33.2% of Mn and 24.3% of Zn being present under free metal ions but 30.4% of Cu being present under Cu-GLY, 66.82% of Mn and 75.7% of Zn being present under Mn-GLY and Zn-GLY, respectively. The supplemented feeds at the 3 tested doses, from the lowest to the highest inclusion levels, contained in total respectively: 1.1, 3.05 and 9.06 mg/kg Cu; 2.99, 8.9 and 18.2 mg/kg Mn; 3.72, 10.9 and 23.4 mg/kg Zn. The M-GLY species recovered by analysis within the different supplemented feeds ranged from 76.26 to 89.32% for Cu-GLY, form 94.5 to 98.51% for Mn-GLY and from 76.05 to 98.96% for Zn-GLY. These results showed that CE-ICP-MS technique can be used to quantify low doses and to measure metal-species repartition between Metal-GLY and free metal ions, when included in feeds. For the first time, this study highlighted that the raw materials used contain Metal-GLY compounds. This raises the question of the occurrence of these compounds within the different raw materials used in feed production that could dramatically affect the way to supplement minerals in animal feed.

The transcription factor NHR-8: A new target to increase ivermectin efficacy in nematodes.

Resistance to the anthelmintic macrocyclic lactone ivermectin (IVM) has a great impact on the control of parasitic nematodes. The mechanisms by which nematodes adapt to IVM remain to be deciphered. We have identified NHR-8, a nuclear hormone receptor involved in the xenobiotic response in Caenorhabditis elegans, as a new regulator of tolerance to IVM. Loss-of-function nhr-8(ok186) C. elegans mutants subjected to larval development assays and electropharyngeogram measurements, displayed hypersensitivity to IVM, and silencing of nhr-8 in IVM-resistant worms increased IVM efficacy. In addition, compared to wild-type worms, nhr-8 mutants under IVM selection pressure failed to acquire tolerance to the drug. In addition, IVM-hypersensitive nhr-8(ok186) worms displayed low transcript levels of several genes from the xenobiotic detoxification network and a concomitant low Pgp-mediated drug efflux activity. Interestingly, some pgp and cyp genes known to impact IVM tolerance in many nematode species, were down regulated in nhr-8 mutants and inversely upregulated in IVM-resistant worms. Moreover, pgp-6 overexpression in nhr-8(ok186) C. elegans increased tolerance to IVM. Importantly, NHR-8 function was rescued in nhr-8(ok186) C. elegans with the homolog of the parasitic nematode Haemonchus contortus, and silencing of Hco-nhr-8 by RNAi on L2 H. contortus larvae increased IVM susceptibility in both susceptible and resistant H. contortus isolates. Thus, our data show that NHR-8 controls the tolerance and development of resistance to IVM in C. elegans and the molecular basis for this relates to the NHR-8-mediated upregulation of IVM detoxification genes. Since our results show that Hco-nhr-8 functions similarly to Cel-nhr-8, this study helps to better understand mechanisms underlying failure in drug efficacy and open perspectives in finding new compounds with NHR-8 antagonist activity to potentiate IVM efficacy.

100 Questions in Livestock Helminthology Research.

An elicitation exercise was conducted to collect and identify pressing questions concerning the study of helminths in livestock, to help guide research priorities. Questions were invited from the research community in an inclusive way. Of 385 questions submitted, 100 were chosen by online vote, with priority given to open questions in important areas that are specific enough to permit investigation within a focused project or programme of research. The final list of questions was divided into ten themes. We present the questions and set them briefly in the context of the current state of knowledge. Although subjective, the results provide a snapshot of current concerns and perceived priorities in the field of livestock helminthology, and we hope that they will stimulate ongoing or new research efforts.

Deciphering the molecular determinants of cholinergic anthelmintic sensitivity in nematodes: When novel functional validation approaches highlight major differences between the model Caenorhabditis elegans and parasitic species.

Cholinergic agonists such as levamisole and pyrantel are widely used as anthelmintics to treat parasitic nematode infestations. These drugs elicit spastic paralysis by activating acetylcholine receptors (AChRs) expressed in nematode body wall muscles. In the model nematode Caenorhabditis elegans, genetic screens led to the identification of five genes encoding levamisole-sensitive-AChR (L-AChR) subunits: unc-38, unc-63, unc-29, lev-1 and lev-8. These subunits form a functional L-AChR when heterologously expressed in Xenopus laevis oocytes. Here we show that the majority of parasitic species that are sensitive to levamisole lack a gene orthologous to C. elegans lev-8. This raises important questions concerning the properties of the native receptor that constitutes the target for cholinergic anthelmintics. We demonstrate that the closely related ACR-8 subunit from phylogenetically distant animal and plant parasitic nematode species functionally substitutes for LEV-8 in the C. elegans L-AChR when expressed in Xenopus oocytes. The importance of ACR-8 in parasitic nematode sensitivity to cholinergic anthelmintics is reinforced by a 'model hopping' approach in which we demonstrate the ability of ACR-8 from the hematophagous parasitic nematode Haemonchus contortus to fully restore levamisole sensitivity, and to confer high sensitivity to pyrantel, when expressed in the body wall muscle of C. elegans lev-8 null mutants. The critical role of acr-8 to in vivo drug sensitivity is substantiated by the successful demonstration of RNAi gene silencing for Hco-acr-8 which reduced the sensitivity of H. contortus larvae to levamisole. Intriguingly, the pyrantel sensitivity remained unchanged thus providing new evidence for distinct modes of action of these important anthelmintics in parasitic species versus C. elegans. More broadly, this highlights the limits of C. elegans as a predictive model to decipher cholinergic agonist targets from parasitic nematode species and provides key molecular insight to inform the discovery of next generation anthelmintic compounds.