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This research was co-funded by the European Fund of Regional Development of the European Union within the framework of the FEDER operating program of Catalunya 2014-2020 (project COMRDI16-1-0033) and managed by ACCIÓ. Nutritional strategies to improve gut health of broilers are under research. This study investigated the effect of dietary supplementation with sodium butyrate protected by sodium salts of medium-chain fatty acids as a feed additive on broiler gut health. The first experiment was conducted to evaluate the effect of supplementing at 0.5, 1, and 2 kg/t in broilers housed under optimal conditions. Supplementation at 0.5 and 1 kg/t maintained goblet cell counts at 10 days of age (p ≤ 0.05), and supplementation at 1 kg/t decreased intraepithelial lymphocyte counts compared to 2 kg/t at 39 days (p ≤ 0.10). Abdominal fat pad levels of lauric and myristic acids were gradually increased by supplement dose (p ≤ 0.05). In the second experiment, the feed additive at 1 kg/t was evaluated in coccidiosis-challenged broilers. Experimental treatments were as follows: non-challenged, control-challenged, and supplemented-challenged treatments. Coccidiosis negatively impact performance and modify histomorphometry and microbiota (p ≤ 0.05). The feed additive increased crypt depth at 7 days post-inoculation and goblet cell count at 14 days post-inoculation (p ≤ 0.05). Further, supplementation interacted with the microbiota modification led by the coccidiosis (p ≤ 0.05). These results suggest that this feed additive could be a useful strategy to reinforce the gut barrier, especially for birds under coccidiosis-challenge treatments.

The relation between nutrition and intestinal health is a subject with an increasing interest in research, as nutritionists need knowledge about how formulation affects different parameters in the gastrointestinal tract (GIT). That is why 4 trials were conducted to evaluate the effect of nutrient concentration and a feed additive (sodium butyrate protected with sodium salts of palm fatty acid distillates (PSB, Gustor N'RGY produced by Norel S.A., Spain, dosed at 1 kg/t), on performance, diet digestibility, intestinal morphology, volatile fatty acid concentration (VFA) in the GIT and intestinal microbiota of broiler chickens, when fed diets with different energy and amino acids concentration. Control diets, C, with the recommended metabolizable energy (ME) and ideal amino acid (AA) composition; Reduction 1, R1, C – 60 kcal ME and – 2.3% AA and Reduction 2, R2, C – 120 kcal ME and – 4.6% AA) based on different feed ingredients (Corn Soy [CS] and Wheat Barley Soy (WBS) were formulated. All trials lasted 42 d. In trials 2 and 4, the nutrient dilution decreased performance of the animals. In all trials, PSB improved animal performance (growth or FCR), despite the different situations. In trials 1 and 4, animals receiving R1 diets and PSB showed similar performance to those receiving C diets without PSB. PSB improved Gross Energy metabolizability (69.94 vs. 72.55; P: 0.02). Nutrient concentration affected histology results in T2 (ileum) and T3 (jejunum); PSB showed effects in T2 (jejunum, ileum) and in T3 (jejunum). In T1, PSB affected VFA in duodenum, jejunum, and ileum, changing the profile depending on diet nutrient concentration. PSB altered microbiology in caecum of animals in T2. It can be concluded that the dilution of ME and AA concentration of the diet impairs animal performance, influences intestinal microbiota and affects intestinal histology. PSB improves animal performance, increases gross energy metabolizability, steers intestinal microbiota and alters VFA concentrations in the intestine. The addition of PSB may help the animal to counteract the negative effects of diluted diets.

Butyric acid has received great attention as a feed additive to maintain or increase the gut integrity and health of broiler chickens. Particularly, the protection of butyrate is under research to allow slow intestinal release of butyric acid and to promote its beneficial effects throughout the intestine. This study evaluated in vivo the intestinal release of butyric acid from sodium butyrate protected by salts of medium-chain fatty acid in broilers. Brilliant blue was used as an inert marker, so it was included in the feed additive that broilers ingested for two days. The gastrointestinal tract was then colored in blue from jejunum and backward. Considering the digesta color of the broilers non-supplemented as blank, it allowed quantification of the amount of brilliant blue, and consequently, butyric acid delivered in the intestine from the protected feed additive. Few traces of butyric acid were released in the duodenum and proximal jejunum, whereas the major amount (45.9%) was delivered in the distal ileum (p < 0.001). These results suggest that this in vivo approach allows for evaluation of the intestinal delivery of butyric acid supplemented as protected sodium butyrate by medium-chain fatty acids, showing a gradual intestinal release of butyric acid in broiler chickens.

The aim of this study was to investigate the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of organic acid salts against six field isolates of Streptococcus suis. The three products evaluated were sodium salt of coconut fatty acids distillate (DIC) alone and two combinations with sodium butyrate (NaBut): DIC70:30, being 70% of NaBut protected with 30% of DIC; and DIC50:50, being 50% of NaBut protected with 50% of DIC. Antimicrobial susceptibility testing was performed to estimate the MIC values for each product and strain by the broth microdilution method at pH 6.0. MBC values were also determined by sub-culturing supernatant from wells without evident bacterial growth. The values of MIC50/MBC50 were calculated as the concentration which inhibited/killed 50% of the isolates tested. The MIC50 showed DIC as the most effective (8 ppm) against S. suis followed by DIC50:50 (32 ppm) and DIC70:30 (64 ppm). The MBC50 demonstrated a similar trend, DIC being the most effective (16 ppm) followed by DIC50:50 (64 ppm) and DIC70:30 (64 ppm). It is well known that butyric acid is a short-chain fatty acid which has strong antimicrobial activity against Gram-negative bacteria. In contrast, coconut fatty acids distillate is a medium-chain fatty acid source (MCFA) rich in lauric acid which has strong antimicrobial activity against Gram-positive bacteria. Both products are generally available as salts to facilitate their application in feed. In this study, the results showed that DIC was the most effective against the Gram-positive bacteria tested, followed by DIC50:50 and DIC70:30, the sodium butyrate-based products. As expected, a higher concentration of MCFA in the tested product was associated with a higher inhibitory and bactericidal activity. Further studies would be required to better understand these interactions as well as in vivo studies to demonstrate the effects on microbial populations.

Essential oils (EOs) have different mechanisms, most of them targeting the bacterial wall. This fact can explain differences in the effectivity of EOs between Gram-positive and Gram-negative bacteria. Therefore, combining certain EOs can broaden their individual spectrum of efficacy due to potential synergistic effects. This trial aimed to test the in vitro antibacterial activity of an EO combination (oregano and clove oils) against a collection of relevant bacterial pathogens in swine production. The Gram-negative bacterial species chosen were Salmonella enterica ssp. enterica, Escherichia coli and Brachyspira hyodysenteriae and the Gram-positive bacterial species were Clostridium perfringens and Streptococcus suis. In addition, Lactobacillus fermentum was included to compare the susceptibility between this beneficial intestinal bacteria and the pathogens tested. The broth microdilution method at pH 6 and the subculturing from wells without bacterial growth were used to determine the minimum concentration of active principle necessary to inhibit (MIC) or kill (MBC) the 50% and 90% (MIC50/90/MBC50/90) of the population of every bacteria. The results showed that the lowest MIC50/90 were obtained for B. hyodysenteriae (37.5/75 ppm) while for S. enterica ssp. enterica (150/300 ppm), C. perfringens (150/150 ppm), E. coli (300/300 ppm) and S. suis (150/300 ppm) results were similar. Regarding the MBC50/90; B. hyodysenteriae (18.8/75 ppm) was the most susceptible pathogen, again, compared to S. enterica ssp. enterica (300/300 ppm), C. perfringens (150/150 ppm), E. coli (300/300 ppm) and S. suis (150/300 ppm). In contrast, the highest bacteriostatic/bactericidal concentrations were obtained against L. fermentum (MIC50/90 600/1,200 ppm and MBC50/90 600/2,400 ppm). These results suggest that the bacterial category (Gram-positive or Gram-negative) did not have an influence on the MIC and MBC. It can also be concluded that B. hyodysenteriae is the most susceptible enteropathogen to this EO blend. However, the in vivo effect of this combination of EOs must be further studied.

Despite the limited use of antibiotics as growth promoters (AGPs) in a number of countries, many others find a useful tool in them. However, many producers in the later regions are transitioning to AGP-free production for commercial reasons. The aim of this trial was to determine whether a blend of essential oils (EO), like oregano or clove, could reach similar production/health parameters compared to an AGP treatment used in broiler production. A total of 1,200 one-day-old chicks were distributed in 2 treatments within 12 floor pens (n = 6). The treatments were: T1-basal feed + AGP (Enramycin 10ppm); T2-basal feed + EO (1kg/t). The trial lasted 35d and performance parameters were recorded weekly. On day 21 and 35, one bird per pen was euthanized for intestinal and carcass analysis. Data were analyzed by one-way ANOVA using the GLM procedure of SAS 9.0. There were no significant differences in the performance parameters at 35d. A trend was observed of higher BW (P = 0.062) at 28d for EO birds (1,708g) compared to AGP birds (1,671g), which relates to another trend (P = 0.071) of increased daily gain from 21-28d (685g and 653g for EO and AGP treatments respectively). A relevant finding was a significantly (P = 0.034) higher mortality in AGP birds from 0-28d (4.83%) compared to EO birds (3.00%). In the post-mortem analysis no differences were observed in intestinal segments in relative and total length. However, animals from T2 showed a trend (P = 0.077) to have heavier gizzards at 21d, which could indicate an effect of EO on their development, while T1 animals had a significantly heavier cecum at 35d (P = 0.026). No relevant findings were observed in carcass weights and yield. These results suggest that this EO blend could be a valid alternative to AGPs, although further trials under commercial conditions should be carried out to confirm its potential.

This study evaluated the effect of two feed lauric acid (LA) based additives on faecal microbiota of piglets. A total of 256 piglets were distributed (from 25 to 66d of age) in 4 treatments: CON-, diet without additives nor antibiotics; CON+, diet with Doxycycline; DIC, CON- diet plus sodium salt of coconut fatty acids distillate at 3kg/t and 1kg/t in starter and grower diets; MON, CON- diet plus monolaurine at 2.1kg/t and 0.7kg/t in starter and grower diets. The dosage of both products was made at the same level of LA (0.96kg LA/t starter and 0.32kg LA/t grower). At the end of the study, the 16S rDNA sequencing was performed. The result showed a drastic decrease in the abundance of the dominating genus Prevotella within the MON group compensated by other genera increase, mostly Selenomonadales. Streptococcus, Treponema, and unclassified Erysipelotrichaceae, Dialister or Bacillus increased their respective abundances. Treponema was more pronounced in the DIC, however, all other mentioned genera did not show a similar increase in abundance. Dialister (a marker for disease in humans) showed significantly lower abundance in DIC. In the bacterial fingerprints, a trend for clustering according to treatment was visible. Six of eight CON- samples formed a separate cluster. Six of eight samples of the DIC formed a cluster that shared very low similarity to the CON-. Four of eight CON+ samples formed a separate cluster. MON samples were interspersed among all samples and showed no distinct clustering. Even though quantitative microbial comparisons were only anecdotal, observations may indicate a more “healthier” microbiota in the DIC than was present in the MON. DIC also showed a much closer similarity of bacterial fingerprints for the pooled samples in the qualitative analysis indicating that DIC had a higher potential to modify the faecal (hindgut) microbiota than MON.

Effect of feed additives based on lauric acid (LA) on performance and faecal score in piglets. A total of 192 piglets were distributed in 24 pens from 25 to 66d of age (42d feeding period). The study was divided into two feeding phases; starter (25 to 38d) and grower period (39 to 66d). Treatments were: CON-, diet without additives nor antibiotics; CON+, diet with Doxycycline (0.25g/kg and 0.13g/kg in starter and grower diets); DIC, CON- diet plus sodium salt of coconut fatty acids distillate (DICOSAN) at 3kg/t and 1kg/tn in starter and grower diets; MON, CON- diet plus monolaurine at 2.1kg/t and 0.7kg/tn in starter and grower diets. The dosage of both products was made at same level of LA (0.96kg LA/t starter and 0.32kg LA/t grower). All parameters were analyzed using a one-way ANOVA via SPSS, with pen as the experimental unit. At 38d, CON+ and DIC piglets had significantly lower FCR than CON- (1.534a, 1.353b, 1.380b and 1.526ab, for CON-, CON+, DIC and MON, P = 0.021). At 66d, CON+ tended to have lower FCR than CON- (1.613x, 1.541y, 1.560xy and 1.605xy, for CON-, CON+, DIC and MON, P = 0.054). For the overall period, DIC and CON+ piglets had significantly lower FCR than CON- (1.603a, 1.514b, 1.537b and 1.589ab, for CON-, CON+, DIC and MON, P = 0.003), there was also a tendency to lower daily weight gain in MON treatment when compared with CON+ (443xy, 468y, 449xy and 440x g/d, for CON-, CON+, DIC and MON, P = 0.079). Finally, the faecal score showed higher values for DIC and CON+, meaning more consistent faeces (3.71a, 3.93b, 3.84b and 3.82ab, for CON-, CON+, DIC and MON, P = 0.001). The results demonstrated that DICOSAN, administered in feed at the same dosage of lauric acid, achieved better results than monolaurine and CON- and without significant differences with CON+.

The aim of this trial was to evaluate the effect of prenatal and postnatal inclusion of a combination of essential oils (EO) on performance parameters and diarrheas after 2 weeks post-weaning. The trial followed a 2x3 factorial design; 2 sow diets: basal or with EO at 375 g/t x 3 piglet diets: basal diet, medicated with 200 ppm of colistin or treated with 375 g/t of EO. A total of 168 weaned piglets were randomly distributed in the 6 groups: 3 from basal sows (CC, CA and CT) and 3 from EO sows (TC, TA and TT), being the animals from the TT treatment the ones imprinted. The incidence of diarrhea was recorded twice a day and calculated using the percentage of animals with diarrhea over the total period of observations carried out. The animals were weighed at 14 days post-weaning. Average daily gain (ADG), average daily feed intake (ADFI) and feed conversion ratio (FCR) were calculated. Data were analyzed by two-way ANOVA using GLM procedure of SAS. Animals from the TT group had significantly lower incidence of diarrheas from 0 to 14 days post weaning. Diarrhea was recorded on 4.7b% of the observations (TT group), being significantly lower (P ≤ 0.001) than the animals from CC (10.8a%), CA (13.7a%), CT (10.7a%), TC (14.4a%) and TA (12.9a%). Performance parameters indicated that there were non-significant differences in BW, ADFI, ADG with numerically lower FCR (kg/kg) 1.14 in TT group (CC 1.23, CA 1.33, CT 1.27, TC 1.22, TA 1.23). Despite the non-significant differences in performance parameters, in this study the effect of imprinting with essential oils reduced diarrhea incidence.

Background Accurate screening of new alternative antimicrobial compounds is essential for their use to control pathogens in swine production due to the replacement of antibiotics and zinc oxide. Most in vitro studies have separately reported the antimicrobial activity of organic acids and essential oils (EOs) using diverse methods for susceptibility testing. In addition, in vitro outcomes can help in the selection of the suitable antimicrobial compound and effective combinations of these compounds in the control of pathogens of interest in pork production. Therefore, the aim of this study is to determinate the antibacterial activity of six organic acids and six EOs against Escherichia coli, Salmonella spp. and Clostridium perfringens isolates, some of them multi-resistant to antibiotics, from swine origin. The synergistic effects between the products with higher activity for each bacteria were also calculated. Results All products tested showed activity against at least one bacterial species, except for black pepper EO. The results showed that formic acid with the shortest chain length was the most effective against E. coli and Salmonella spp., while the sodium salt of coconut fatty acid distillates with long chain acids was the most effective against C. perfringens. The susceptibility of isolates tested to EOs was similar, a result that demonstrates a similar activity of these products against phylogenetically unrelated pathogens. In addition, an additive effect was shown for carvacrol-oregano EO for E. coli, formic acid-carvacrol and formic acid-thymol for Salmonella spp. and carvacrol-cinamaldehyde for C. perfringens. Conclusions The susceptibility of isolates to EOs was similar, a result that demonstrates a similar activity of these products against phylogenetically unrelated pathogens in contrast to organic acids. In addition, an additive effect was shown for several combinations of these compounds.