Dietary butyrate and valerate glycerides impact diarrhea severity and immune response of weaned piglets under ETEC F4-ETEC F18 coinfection conditions.

Enterotoxigenic Escherichia coli (ETEC) causes post-weaning diarrhea in piglets, significantly impacting animal welfare and production efficiency. The two primary ETEC pathotypes associated with post-weaning diarrhea are ETEC F4 and ETEC F18. During the post-weaning period, piglets may be exposed to both ETEC F4 and ETEC F18. However, the effects of coinfection by both strains have not been studied. Short chain fatty acid feed additives, such as butyrate and valerate, are being investigated for their potential to improve animal performance and disease resistance. Therefore, this pilot experiment aimed to test the effects of butyrate glycerides or valerate glycerides on growth performance, diarrhea incidence, and immune responses of piglets under ETEC F4-ETEC F18 coinfection conditions. Twenty piglets were individually housed and assigned to one of the three dietary treatments immediately at weaning (21 to 24 d of age). The dietary treatments included control (basal diet formulation), control supplemented with 0.1% butyrate glycerides or 0.1% valerate glycerides. After a 7-d adaptation, all pigs were inoculated with ETEC F4 and ETEC F18 (0.5 × 109 CFU/1.5 mL dose for each strain) on three consecutive days. Pigs and feeders were weighed throughout the trial to measure growth performance. Fecal cultures were monitored for hemolytic coliforms, and blood samples were collected for whole blood and serum analysis. Pigs fed valerate glycerides tended (P = 0.095) to have higher final body weight compared with control. The overall severity of diarrhea was significantly (P < 0.05) lower in both treatment groups than control. Pigs fed valerate glycerides tended (P = 0.061) to have lower neutrophils and had significantly (P < 0.05) lower serum TNF-α on day 4 post-inoculation. This pilot experiment established an appropriate experimental dose for an ETEC F4-ETEC F18 coinfection disease model in weaned piglets. Results also suggest that butyrate glycerides and valerate glycerides alleviated diarrhea and regulated immune responses in piglets coinfected with ETEC F4 and ETEC F18.

Piglets suffer from post-weaning diarrhea associated with Enterotoxigenic Escherichia coli (ETEC) F4 and F18, two prevalent strains on swine farms globally. Short chain fatty acids (SCFAs), such as butyrate and valerate, are natural, organic compounds that could potentially promote intestinal health when used as dietary supplements. During the post-weaning period, piglets are vulnerable to simultaneous infection by ETEC F4 and F18. Therefore, this experiment aimed to develop an experimental disease model for coinfection with ETEC F4 and F18, employing a dose of 0.5 × 109 CFU/1.5 mL of each strain, administered over three consecutive days. In addition, the experiment evaluated treatment diets supplemented with 0.1% butyrate or valerate glycerides compared with the control diet. Results from this experiment revealed that the inoculation dose incited infection and diarrhea in piglets, implying its suitability for use in a disease challenge model. Moreover, the results indicated that the inclusion of butyrate and valerate glycerides to pig's diet reduced the severity of diarrhea. Furthermore, pigs fed SCFA glycerides exhibited lowered levels of inflammatory blood markers. In conclusion, the experimental dose induced diarrhea in piglets, and dietary supplementation of butyrate and valerate glycerides alleviated the severity of diarrhea while augmenting inflammatory status.

Dietary supplementation of botanical blends enhanced performance and disease resistance of weaned pigs experimentally infected with enterotoxigenic Escherichia coli F18.

Botanicals exhibit promising impacts on intestinal health, immune-regulation, and growth promotion in weaned pigs. However, these benefits may vary depending on major active components in the final feed additive products. Therefore, this study aimed to investigate two types of botanical blends (BB) that were comprised of 0.3% capsicum oleoresin and 12% garlic extracts from different sources on performance, diarrhea, and health of weaned piglets experimentally infected with a pathogenic Escherichia coli F18. Sixty weanling pigs (7.17 ± 0.97 kg body weight (BW)) blocked by weight and gender were assigned to one of five dietary treatments: negative control (NC), positive control (PC), or dietary supplementation with 100 mg/kg of BB1, 50 mg/kg or 100 mg/kg of BB2. This study lasted 28 d with 7 d before and 21 d after the first E. coli inoculation (day 0). All pigs, except negative control, were orally inoculated with 1010 cfu E. coli F18/3-mL dose for 3 consecutive days. Blood samples were collected periodically to analyze systemic immunity. Intestinal tissues and mucosa were collected on days 5 and 21 PI for analyzing histology and gene expression. All data, except for frequency of diarrhea, were analyzed by ANOVA using the PROC MIXED of SAS. The Chi-square test was used for analyzing frequency of diarrhea. Escherichia coli infection reduced (P < 0.05) growth rate and feed intake and increased (P < 0.05) frequency of diarrhea of weaned pigs throughout the experiment. Supplementation of 100 mg/kg BB1 or BB2 alleviated (P < 0.05) frequency of diarrhea of E. coli challenged pigs during the entire experiment. Escherichia coli infection also enhanced (P < 0.05) serum TNF-α and haptoglobin concentrations on day 4 post-inoculation (PI) but reduced (P < 0.05) duodenal villi height and area on day 5 PI, while pigs supplemented with 100 mg/kg BB1 or BB2 had lower (P < 0.05) serum TNF-α than pigs in PC on day 4 PI. Pigs fed with 100 mg/kg BB2 had higher (P < 0.05) jejunal villi height than pigs in PC on day 5 PI. Pigs fed with 100 mg/kg BB2 had reduced (P < 0.05) gene expression of IL1B, PTGS2, and TNFA in ileal mucosa than pigs in PC on day 21 PI. In conclusion, dietary supplementation of botanical blends at 100 mg/kg could enhance disease resistance of weaned pigs infected with E. coli F18 by enhancing intestinal morphology and regulating local and systemic immunity of pigs.

This experiment aimed to investigate two botanical blends consisting of 0.3% capsicum oleoresin and 12% garlic extracts on performance, diarrhea, and health of weaned piglets experimentally infected with a pathogenic Escherichia coli F18. The two botanical blends have the same formulation, except that different garlic oils were used. A total of 60 weaned pigs were randomly allotted to one of five experimental treatments: 1) a complex control diet without E. coli F18 challenge; 2) control diet with E. coli F18 challenge; 3) supplementing 100 mg/kg of botanical blend type 1 to pigs challenged with E. coli F18; 4) and 5) supplementing 50 or 100 mg/kg of botanical blend type 2 to pigs challenged with E. coli F18. The experiment lasted 28 d with 7 d adaptation and 21 d after the first F18 E. coli inoculation. Results of this experiment demonstrate that supplementation of 100 mg/kg of botanical blend enhanced disease resistance and tended to improve growth of weaned pigs, regardless of garlic oil variety. An improved intestinal morphology and reduced systemic inflammation was also observed in pigs supplemented with 100 mg/kg of botanical blends. In conclusion, supplementation of 100 mg/kg of botanical blends could reduce diarrhea of E. coli infected pigs and modify local or systemic immunity of pigs.

Supplementation of oligosaccharide-based polymer enhanced growth and disease resistance of weaned pigs by modulating intestinal integrity and systemic immunity.

BACKGROUND: There is a great demand for antibiotic alternatives to maintain animal health and productivity. The objective of this experiment was to determine the efficacy of dietary supplementation of a blood group A6 type 1 antigen oligosaccharides-based polymer (Coligo) on growth performance, diarrhea severity, intestinal health, and systemic immunity of weaned pigs experimentally infected with an enterotoxigenic Escherichia coli (ETEC), when compared with antibiotics. RESULTS: Pigs in antibiotic carbadox or Coligo treatment groups had greater (P < 0.05) body weight on d 5 or d 11 post-inoculation (PI) than pigs in the control group, respectively. Supplementation of antibiotics or Coligo enhanced (P < 0.05) feed efficiency from d 0 to 5 PI and reduced (P < 0.05) frequency of diarrhea throughout the experiment, compared with pigs in the control group. Supplementation of antibiotics reduced (P < 0.05) fecal ß-hemolytic coliforms on d 2, 5, and 8 PI. Pigs in antibiotics or Coligo groups had reduced (P < 0.05) neutrophil counts and serum haptoglobin concentration compared to pigs in the control group on d 2 and 5 PI. Pigs in Coligo had reduced (P < 0.05) total coliforms in mesenteric lymph nodes on d 5 and 11 PI, whereas pigs in antibiotics or Coligo groups had reduced (P < 0.05) total coliforms in spleen on d 11 PI compared with pigs in the control group. On d 5 PI, pigs in the Coligo group had greater (P < 0.05) gene expression of ZO1 in jejunal mucosa, but less (P < 0.05) mRNA expression of IL1B, IL6, and TNF in ileal mucosa, in comparison with pigs in the control group. Supplementation of antibiotics enhanced (P < 0.05) the gene expression of OCLN in jejunal mucosa but decreased (P < 0.05) IL1B and IL6 gene expression in ileal mucosa, compared with the control. On d 11 PI, supplementation of antibiotics or Coligo up-regulated (P < 0.05) gene expression of CLDN1 in jejunal mucosa, but Coligo reduced (P < 0.05) IL6 gene expression in ileal mucosa compared to pigs in the control group. CONCLUSIONS: Supplementation of Coligo improved growth performance, alleviated diarrhea severity, and enhanced gut health in weaned pigs infected with ETEC F18 in a manner similar to in-feed antibiotics.

Mint Oils: In Vitro Ability to Perform Anti-Inflammatory, Antioxidant, and Antimicrobial Activities and to Enhance Intestinal Barrier Integrity.

The objectives of the study were to test the biological activities of peppermint and spearmint oils via (i) measuring in vitro anti-inflammatory effects with porcine alveolar macrophages (PAMs), (ii) determining the barrier integrity of IPEC-J2 by analyzing transepithelial electrical resistance (TEER), (iii) testing their antioxidant activities, and (iv) investigating the antimicrobial activity against enterotoxigenic Escherichia coli (ETEC) F18+. Briefly, (i) macrophages were seeded at 106 cells/mL and treated (24 h) with mint oils and lipopolysaccharide (LPS). The treatments were 2 (0 or 1 µg/mL of LPS) × 5 (0, 25, 50, 100, 200 µg/mL of mint oils). The supernatants were collected for TNF-α and IL-1ß measurement by ELISA; (ii) IPEC-J2 cells were seeded at 5 × 105 cells/mL and treated with mint oils (0, 25, 50, 100, and 200 µg/mL). TEER (Ωcm2) was measured at 0, 24, 48, and 72 h; (iii) the antioxidant activity was assessed (0, 1, 50, 100, 200, 500, and 600 mg/mL) using the 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging and reducing power assays; (iv) overnight-grown ETEC F18+ were quantified (CFU/mL) after supplementing with peppermint and spearmint oils (0, 1.44, 2.87, 5.75, 11.50, and 23.00 mg/mL). All data were analyzed using the MIXED procedure. Both mint oils significantly inhibited (p < 0.05) IL-1ß and TNF-α secretion from LPS-stimulated PAMs. Mint oil treatments did not affect TEER in IPEC-J2. Spearmint and peppermint oils exhibited (p < 0.05) strong antioxidant activities in DPPH and reducing power assays. Both mint oils also dose-dependently inhibited (p < 0.05) the growth of ETEC F18+ in vitro. The results of the study indicated that both mint oils are great candidate feed additives due to their in vitro anti-inflammatory, antioxidant, and antimicrobial effects. Further research is needed to evaluate their efficacy in vivo.

Trace amounts of antibiotic exacerbated diarrhea and systemic inflammation of weaned pigs infected with a pathogenic Escherichia coli.

The experiment was conducted to investigate the effects of trace amounts of antibiotic on growth performance, diarrhea, systemic immunity, and intestinal health of weaned pigs experimentally infected with an enterotoxigenic Escherichia coli. Weaned pigs (n = 34, 6.88 ± 1.03 kg body weight [BW]) were individually housed in disease containment rooms and randomly allotted to one of the three dietary treatments: nursery basal diet (CON) and two additional diets supplemented with 0.5 or 50 mg/kg carbadox to the nursery basal diet (TRA or REC), respectively. The experiment lasted 18 d with 7 d before and 11 d after the first E. coli inoculation. The E. coli F18 inoculum was orally provided to all pigs with a dose of 1010 colony-forming unit (CFU)/3 mL for three consecutive days. Fecal and blood samples were collected on day 0 before inoculation and days 2, 5, 8, and 11 postinoculation (PI) to test the percentage of ß-hemolytic coliforms in total coliforms and complete blood cell count, respectively. Sixteen pigs were euthanized on day 5 PI, whereas the remaining pigs were euthanized at the end of the experiment to collect the jejunal and ileal mucosa and mesenteric lymph node for gene expression and bacterial translocation, respectively. Pigs in REC had greater (P < 0.05) final BW and lower (P < 0.05) overall frequency of diarrhea compared with pigs in the CON and TRA groups. Pigs in TRA had the lowest (P < 0.05) average daily gain and feed efficiency from day 0 to 5 PI, highest (P < 0.05) percentage of ß-hemolytic coliforms in fecal samples on days 2 and 5 PI, and greatest (P < 0.05) bacterial colonies in mesenteric lymph nodes on day 11 PI compared with pigs in the CON and REC groups. Pigs in TRA had the greatest (P < 0.05) neutrophils on day 5 PI and higher (P < 0.05) white blood cell counts and lymphocytes than other groups on day 11 PI. Pigs in TRA had the greatest (P < 0.05) serum C-reactive protein on days 2 and 5 PI and serum tumor necrosis factor-α on day 5 PI, compared with pigs in the CON and REC groups. Pigs fed REC had increased (P < 0.05) mRNA expression of zona occludens-1 (ZO-1) and occludin (OCDN) and reduced (P < 0.05) interleukin-1 beta (IL1B), interleukin-6 (IL6), and tumor necrosis factor-alpha (TNFA) in ileal mucosa on day 5 PI, compared with the CON, whereas TRA upregulated (P < 0.05) mRNA expression of IL1B, IL6, and cyclooxygenase-2 (COX2) in the ileal mucosa on day 11 PI, compared with the REC. In conclusion, trace amounts of antibiotic may exacerbate the detrimental effects of E. coli infection on pig performance by increasing diarrhea and systemic inflammation of weanling pigs.

Bacillus subtilis: a potential growth promoter in weaned pigs in comparison to carbadox.

The study was conducted to investigate the efficacy of a probiotic Bacillus subtilis strain on growth performance, diarrhea, systemic immunity, and intestinal health of weaned pigs experimentally infected with an enterotoxigenic Escherichia coli and to compare the efficacy of B. subtilis with that of carbadox. Weaned pigs (n = 48, 6.17 ± 0.36 kg body weight [BW]) were individually housed in disease containment rooms and randomly allotted to one of four dietary treatments: negative control (NC, control diet without E. coli challenge), positive control (PC, control diet with E. coli challenge), and supplementation of 50 mg/kg of carbadox (antibiotic growth promotor [AGP]) or 2.56 × 109 CFU/kg of B. subtilis probiotics (PRO). The experiment lasted for 28 d with 7 d before and 21 d after the first E. coli inoculation. Fecal and blood samples were collected on days 0, 3, 7, 14, and 21 post inoculation (PI) to analyze ß-hemolytic coliforms and complete blood cell count, respectively. Diarrhea score was recorded daily for each pig to calculate the frequency of diarrhea. All pigs were euthanized at day 21 PI to collect jejunal and ileal mucosa for gene expression analysis. Pigs in AGP had greater (P < 0.05) BW on days 7, 14, and 21 PI than pigs in PC and PRO groups. Supplementation of PRO enhanced pigs' BW on day 21 PI compared with the PC. Escherichia coli F18 challenge reduced (P < 0.05) average daily gain (ADG) and feed efficiency from day 0 to 21 PI, while supplementation of carbadox or PRO enhanced ADG and feed efficiency in E. coli F18-challenged pigs from day 0 to 21 PI. Pigs in AGP and PRO groups had reduced (P < 0.05) frequency of diarrhea throughout the experiment and fecal ß-hemolytic coliforms on day 7 PI than pigs in the PC. Pigs in PRO had greater (P < 0.05) gene expression of CLDN1 in jejunal mucosa than pigs in the PC. Supplementation of carbadox or PRO reduced (P < 0.05) the gene expression of IL6 and PTGS2 in ileal mucosa of E. coli-infected pigs compared with pigs in the PC. Pigs in the PRO group had lower (P < 0.05) white blood cell number and neutrophil count, and serum haptoglobin concentration on day 7 PI, and less (P < 0.05) monocyte count on day 14 PI, compared with PC. In conclusion, supplementation of probiotic B. subtilis could enhance disease resistance and promote the growth performance of weaned pigs under disease challenge conditions. The potential mechanisms include but not limited to enhanced gut barrier integrity and local and systemic immune responses of weaned pigs.

Correction: Kovanda, L., et al. In Vitro Antimicrobial Activities of Organic Acids and Their Derivatives on Several Species of Gram-Negative and Gram-Positive Bacteria. Molecules 2019, 24, 3770.

The authors wish to make the following corrections to this paper published in Molecules [...].

In Vitro Antimicrobial Activities of Organic Acids and Their Derivatives on Several Species of Gram-Negative and Gram-Positive Bacteria.

The objective of this study was to determine the in vitro antimicrobial activity of several organic acids and their derivatives against Gram-positive (G+) and Gram-negative (G-) bacteria. Butyric acid, valeric acid, monopropionin, monobutyrin, monovalerin, monolaurin, sodium formate, and ProPhorce-a mixture of sodium formate and formic acid (40:60 w/v)-were tested at 8 to 16 concentrations from 10 to 50,000 mg/L. The tested bacteria included G- bacteria (Escherichia coli, Salmonella enterica Typhimurium, and Campylobacter jejuni) and G+ bacteria (Enterococcus faecalis, Clostridium perfringens, Streptococcus pneumoniae, and Streptococcus suis). Antimicrobial activity was expressed as minimum inhibitory concentration (MIC) of tested compounds that prevented growth of tested bacteria in treated culture broth. The MICs of butyric acid, valeric acid, and ProPhorce varied among bacterial strains with the lowest MIC of 500-1000 mg/L on two strains of Campylobacter. Sodium formate at highest tested concentrations (20,000 mg/L) did not inhibit the growth of Escherichia coli, Salmonella Typhimurium, and Enterococcus faecalis, but sodium formate inhibited the growth of other tested bacteria with MIC values from 2000 to 18,800 mg/L. The MIC values of monovalerin, monolaurin, and monobutyrin ranged from 2500 to 15,000 mg/L in the majority of bacterial strains. Monopropionin did not inhibit the growth of all tested bacteria, with the exception that the MIC of monopropionin was 11,300 mg/L on Clostridia perfringens. Monolaurin strongly inhibited G+ bacteria, with the MIC value of 10 mg/L against Streptococcus pneumoniae. The MIC tests indicated that organic acids and their derivatives exhibit promising antimicrobial effects in vitro against G- and G+ bacteria that are resistant to antimicrobial drugs. The acid forms had stronger in vitro antimicrobial activities than ester forms, except that the medium chain fatty acid ester monolaurin exhibited strong inhibitory effects on G+ bacteria.