Alterations of the Viable Ileal Microbiota of the Gut Mucosa-Lymph Node Axis in Pigs Fed Phytase and Lactic Acid-Treated Cereals.

The gut-lymph node axis is a critical player in the symbiotic relationship between gut microbiota and the host. However, little is known about the impact of diet-related bacterial shifts in the gut lumen on bacterial translocation into lymph nodes. Here, we (i) characterized changes in the viable microbiota composition along the ileal digesta-mucosa-lymph node axis and (ii) examined the effect of dietary phytase supplementation and lactic acid (LA) soaking of cereals on the bacterial taxonomy along this axis, together with their effect on the mucosal expression of innate immune and barrier function genes in pigs (n = 8/diet). After 18 days on diets, ileal digesta, mucosa, and ileocecal lymph nodes (ICLNs) were collected for RNA isolation and 16S rRNA-based high-resolution community profiling. Bacterial communities were dominated by Lactobacillaceae and Clostridiaceae, with clearly distinguishable profiles at the three sampling sites. Specific bacterial subsampling was indicated by enrichment of the ICLNs with Lactobacillaceae, Lachnospiraceae, Veillonellaceae, and Methanobacteriaceae and less Clostridiaceae, Pasteurellaceae, Helicobacteraceae, and Enterobacteriaceae compared to that of the mucosa. LA treatment of cereals reduced proteolytic taxa in the lumen, including pathobionts like Helicobacteraceae, Campylobacteraceae, and Fusobacteriaceae When combined, phytase- and LA-treated cereals largely increased species richness, while the single treatments reduced Actinobacteria and Bacteroidetes in ICLNs and increased mucosal MUC2 expression. In contrast, phytase reduced mucosal CDH1 expression, indicating altered barrier function with potential effects on bacterial translocation. Overall, both treatments, although often differently, changed the viable microbiome along the digesta-mucosa-lymph node axis in the ileum, probably due to altered substrate availability and microbial-host interactions.IMPORTANCE A host's diet largely determines the gut microbial composition and therefore may influence bacterial translocation into ICLNs. Due to its importance for cell metabolism, the intestinal phosphorus availability, which was modified here by phytase and LA treatment of cereals, affects the intestinal microbiota. Previous studies mainly focused on bacteria in the lumen. The novelty of this work resides mainly in that we report diet-microbe effects along the digesta-mucosa-ICLN axis and linked those effects to mucosal expression of barrier function genes as crucial components for host health. Lymph nodes can serve as reservoir of pathobionts; therefore, present diet-microbiome-host interactions have implications for food safety.

Effect of pre- and post-weaning dietary supplementation with arginine and glutamine on rabbit performance and intestinal health.

BACKGROUND: The purpose of the present study was to assess if the exposure to glutamine (Gln), arginine (Arg) or their combination from pregnancy, through the maternal diet, to a post weaning supplemented diet, can stimulate litter performance, gut development and immune function. To this end does and their litters were fed the same basal diet no supplemented (control C), or supplemented with 0.4% Gln, 0.4% Arg, or 0.4 Gln + 0.4 Arg. Rabbits were weaned at 25 d of age and fed the same experimental diet as their mothers for 10 additional days (35 d of age). Bacterial translocation to mesenteric lymph nodes (MLN) at 6 d of age and intestinal histology, enzymatic activity, phenotypical and functional analysis of intraepithelial lymphocytes (IEL) from the appendix were determined at 6, 25 and 35 d of age. RESULTS: No significant differences on animal performance or mortality rates were observed among dietary treatments. However, kits from rabbit does supplemented with Gln tended (P ≤ 0.10) to reduce the translocation of total number of both aerobic and facultative anaerobic bacteria to the MLN. Also, rabbits fed the Gln supplemented diets maintained intestinal villous height at weaning compared to the non-supplemented diets (P < 0.05). The proportions of CD45+CD4+ and CD45+CD8+ IEL in the appendix were not affected by dietary means. However, in rabbits IEL at weaning dietary Gln significantly upregulated IL-2 and downregulated IL-6 expression. CONCLUSIONS: Despite a lack of effect on performance and mortality the inclusion of 0.4% Gln has a positive effect by maintaining intestinal villous height and modulating the cytokine profile at weaning. The supplementation with Arg or Arg + Gln at the selected doses in this study did not exert positive effects on rabbit intestinal health.

Photodynamic therapy controls of Staphylococcus aureus intradermal infection in mice.

Infections caused by Staphylococcus aureus lead to skin infections, as well as soft tissues and bone infections. Given the communal resistance to antibiotics developed by strains of this bacterium, photodynamic therapy emerges as a promising alternative treatment to control and cure infections. Females of the Balb/C mice were infected with 108 CFU of methicillin-resistant S. aureus (MRSA) and divided into four distinct groups: P-L- (negative control group), P+L- (group exposed only to curcumin), P-L+ (group exposed only to LED incidence of 450 nm, 75 mW/cm2, and 54 J/cm2 for 10 min), and P+L+ (group exposed to curcumin followed by 10 min of LED irradiation) (n = 24). The mice were euthanized 48 and 72 h after infection, and biologic materials were collected for analysis of the bacterial load, peripheral blood leukocyte counts, and draining lymph nodes cell counts. The normalization of data was checked and the ANOVA test was applied. The bacterial load in the draining lymph node of P+L+ group was lower when compared to the control groups 72 h post infection (p < 0.0001), indicating that the LED incidence associated with curcumin controls of the staphylococci intradermal infection. The number of the total lymph node cells shows to be lower than control groups in the two availed times (p < 0.01). The histological analysis and the counting of white blood cells did not show differences among cells in the blood and in the tissue of infection. This is the first report showing that photodynamic therapy may be effective against MRSA infection in a murine model of intradermal infection.

Isolation and identification of Caviibacter abscessus from cervical abscesses in a series of pet guinea pigs (Cavia porcellus).

An organism reported in the early literature to be a rare cause of cervical lymphadenitis in guinea pigs, Streptobacillus moniliformis, has been reclassified as Caviibacter abscessus We describe a series of sequential cases of abscesses in guinea pigs that were presented to our clinic from which the only agent isolated was a unique, serum-requiring bacterium. Discrete colonies were not detected in 6.5% CO2 or anaerobically on routine primary isolation media containing up to 5% whole sheep blood, with and without cysteine, vitamin K, and hemin supplementation after 7 days of incubation at 37°C. Based on subsequently determined growth requirements, the organisms were best described as serum-requiring, aerotolerant anaerobes. Colonies were detectable within 24 h at 37°C in an anaerobic atmosphere on a mycoplasma agar-based medium containing 10% pig serum and reached 3 mm in diameter within 3-5 days. Microscopic appearance consisted of small gram-negative rods and coccobacilli with occasional filaments. However, in direct smears from clinical specimens and from weak or dysgonic growth on plates incubated under suboptimal growth conditions (e.g., in 6.5% CO2), irregular rods with occasional small bulbous forms or numerous long wavy filaments were observed. All of the isolates generated unique spectral profiles similar to that of C. abscessus when examined by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Phylogenetic analysis of partial 16S rRNA gene sequences showed that the isolates were identical to each other and shared 99.9% sequence identity with C. abscessus.

Psychological Stress, Immunity, and the Effects on Indigenous Microflora.

Psychological stress is an intrinsic part of life that affects all organs of the body through direct nervous system innervation and the release of neuroendocrine hormones. The field of PsychoNeuroImmunology (PNI) has clearly demonstrated that the physiological response to psychological stressors can dramatically impact the functioning of the immune system, thus identifying one way in which susceptibility to or severity of diseases are exacerbated during stressful periods. This chapter describes research at the interface between the fields of PNI and Microbial Endocrinology to demonstrate that natural barrier defenses, such as those provided by the commensal microflora, can be disrupted by exposure to psychological stressors. These stress effects are evident in the development of the intestinal microflora in animals born from stressful pregnancy conditions, and in older animals with fully developed microbial populations. Moreover, data are presented demonstrating that exposure to different types of stressors results in the translocation of microflora from cutaneous and mucosal surfaces into regional lymph nodes. When considered together, a scenario emerges in which psychological stressors induce a neuroendocrine response that has the potential to directly or indirectly affect commensal microflora populations, the integrity of barrier defenses, and the internalization of microbes. Finally, a hypothesis is put forth in which stressor-induced alterations of the microflora contribute to the observed stressor-induced increases in inflammatory markers in the absence of overt infection.

Effect of probiotic supplementation on bacterial translocation in common bile duct obstruction.

OBJECTIVE: To investigate the effects of probiotics on bacterial translocation in the obstructive common bile duct with comparison to an enteral product containing arginine and glutamine. MATERIAL AND METHOD: In our study, 40 Sprague-Dawley rats each weighing 250-300 g were used. Animals in Group 1 (sham) were laparatomized and fed standard chow supplemented with physiologic saline at daily doses of 2 ml through orogastric tube for 7 days. Common bile ducts of the animals in the other groups were ligated with 3/0 silk sutures. Group 2 (control group) was fed standard chow supplemented with daily doses of 2 ml physiologic saline. Group 3 (probiotic group) was fed standard chow supplemented with a probiotic solution (Acidophilus plus) containing strains of Lactobacillus acidophilus, Bifidobacterium bifidum and Lactobacillus bulgaricus at a daily doses of 2 × 10(9) colony forming units (CFU). Group 4 (formula group) was fed only an enteral solution (Stresson Multi Fiber) containing glutamine, arginine and a medium-chain fatty acid at daily doses of 2 g/kg. At the end of the 7th day, all animals were relaparatomized, and to determine bacterial translocation, aerobic, and anaerobic cultures were obtained from the specimens of mesenteric lymph nodes, intestinal mucosa, and blood samples. Smear cultures prepared from caecum were examined to determine the number of CFU. Finally, for histological examination specimens were excised from terminal ileum, and oxidative damage was assessed in liver tissues. Afterwards all animals were killed. RESULTS: Moderately lesser degrees of bacterial translocation, and mucosal damage were seen in Groups 3, and 4 relative to Group 2 (p < 0.05). In Group 4, any difference was not seen in the number of cecal bacteria relative to baseline values, while in Group 3, significant decrease in cecal colonization was seen. Among all groups, a significant difference between levels of malondialdehyde, and glutathione was not observed. CONCLUSION: At the end of our study, we have concluded that both probiotics, and enteral diets which contain immunomodulators such as glutamine, and arginine alleviate bacterial translocation, and impairment of intestinal mucosa.

Fermentable fibres condition colon microbiota and promote diabetogenesis in NOD mice.

AIMS/HYPOTHESIS: Gut microbiota (GM) and diet both appear to be important in the pathogenesis of type 1 diabetes. Fermentable fibres (FFs), of which there is an ample supply in natural, diabetes-promoting diets, are used by GM as a source of energy. Our aim was to determine whether FFs modify GM and diabetes incidence in the NOD mouse. METHODS: Female NOD mice were weaned to a semisynthetic diet and the effects of FF supplementation on diabetes incidence and insulitis were evaluated. Real-time quantitative PCR was employed to determine the effects imposed to gene transcripts in the colon and lymph nodes. Changes to GM were analysed by next-generation sequencing. RESULTS: NOD mice fed semisynthetic diets free from FFs were largely protected from diabetes while semisynthetic diets supplemented with the FFs pectin and xylan (PX) resulted in higher diabetes incidence. Semisynthetic diet free from FFs altered GM composition significantly; addition of PX changed the composition of the GM towards that found in natural-diet-fed mice and increased production of FF-derived short-chain fatty acid metabolites in the colon. The highly diabetogenic natural diet was associated with expression of proinflammatory and stress-related genes in the colon, while the semisynthetic diet free from FFs promoted Il4, Il22, Tgfß and Foxp3 transcripts in the colon and/or pancreatic lymph node. PX in the same diet counteracted these effects and promoted stress-related IL-18 activation in gut epithelial cells. 16S RNA sequencing revealed each diet to give rise to its particular GM composition, with different Firmicutes to Bacteroidetes ratios, and enrichment of mucin-degrading Ruminococcaceae following diabetes-protective FF-free diet. CONCLUSIONS/INTERPRETATION: FFs condition microbiota, affect colon homeostasis and are important components of natural, diabetes-promoting diets in NOD mice.

Cecum lymph node dendritic cells harbor slow-growing bacteria phenotypically tolerant to antibiotic treatment.

In vivo, antibiotics are often much less efficient than ex vivo and relapses can occur. The reasons for poor in vivo activity are still not completely understood. We have studied the fluoroquinolone antibiotic ciprofloxacin in an animal model for complicated Salmonellosis. High-dose ciprofloxacin treatment efficiently reduced pathogen loads in feces and most organs. However, the cecum draining lymph node (cLN), the gut tissue, and the spleen retained surviving bacteria. In cLN, approximately 10%-20% of the bacteria remained viable. These phenotypically tolerant bacteria lodged mostly within CD103⁺CX3CR1⁻CD11c⁺ dendritic cells, remained genetically susceptible to ciprofloxacin, were sufficient to reinitiate infection after the end of the therapy, and displayed an extremely slow growth rate, as shown by mathematical analysis of infections with mixed inocula and segregative plasmid experiments. The slow growth was sufficient to explain recalcitrance to antibiotics treatment. Therefore, slow-growing antibiotic-tolerant bacteria lodged within dendritic cells can explain poor in vivo antibiotic activity and relapse. Administration of LPS or CpG, known elicitors of innate immune defense, reduced the loads of tolerant bacteria. Thus, manipulating innate immunity may augment the in vivo activity of antibiotics.

[Investigation of the effects of probiotic bacteria on bacterial translocation that developed during diagnostic laparoscopy: an experimental study]. / Tanisal Amaçla Uygulanan Laparoskopi Sirasinda Olusan Bakteriyel Translokasyona Probiyotik Bakterilerin Etkilerinin Arastirilmasi: Deneysel Bir Çalisma.

Probiotics which are non-pathogenic live microorganisms ingested along with food or as dietary supplements, are thought to be beneficial to the host by supporting the microbial balance in digestive system. Various studies suggest that the effects of probiotics on the intestinal mucosa and immunity are protective against bacterial translocation. We aimed to investigate bacterial translocation related to the amount of CO2 insufflation given during laparoscopy and the effect of probiotic bacteria in an experimental peritonitis model. In this study 60 Wistar rats were used in six groups consisting of 10 rats. Group 1, 3 and 5 consisted of the rats that were fed without probiotics, while the rats in Group 2, 4, and 6 were fed with water containing 5 x 108 cfu/ml probiotic bacteria complex (Bifidobacterium lactis, Lactobacillus bulgaricus, Streptococcus thermophilus) for 15 days. To generate experimental peritonitis, 2 x 107 cfu/ml Escherichia coli ATCC 25922 was inoculated intraperitoneally to all of the rats. Thereafter, laparoscopy was applied in all groups. Application in Group 1 and Group 2 was without CO2; Group 3 and Group 4 with 14 mmHg CO2 insufflation, and Group 5 and Group 6 with 20 mmHg CO2 insufflation. Blood samples were taken in 2nd, 4th, and 6th hours. Mesenteric lymph node, liver and spleen samples were taken at 6th hour when the rats were sacrificed and then these were evaluated microbiologically with qualitative and quantitative methods. Bacterial translocation and bacteremia were found in the rats that were undergone experimental peritonitis during laparoscopy. All positive tissue and blood cultures yielded E.coli. The highest level of bacterial translocation was found to be in mesenteric lymph nodes (in 3/10, 6/10 and 10/10 in groups 1, 3 and 5 fed without probiotics, respectively; in 2/10, 3/10 and 4/10 in groups 2, 4 and 6 fed with probiotics, respectively). The bacterial translocation rates were found to be related to the increased CO2 insufflation. It was found that probiotic bacteria were more effective for decreasing bacterial translocation rates and bacteremia in the groups that were given high CO2 pressure during laparoscopy. It was also found that these results were correlated with bacterial translocation per gram of tissue. As an example, the quantitative bacterial growth values detected in mesenteric lymph node were 5.4 ± 2.9 x 103, 10.6 ± 3.3 x 103 and 21.5 ± 12.4 x 103 cfu/g in groups 1, 3 and 5, fed without probiotics, respectively; and 2.0 ± 1.3 x 103, 3.8 ± 1.9 x 103 and 9.0 ± 3.1 x 103 cfu/g in groups 2, 4 and 6, fed with probiotics, respectively. Our data emphasized that probiotic bacteria may be used as prophylactic agents for the prevention of bacterial translocation during laparoscopy, however comprehensive and clinical studies are needed to support these experimental results.

Effect of organic acids on Salmonella colonization and shedding in weaned piglets in a seeder model.

Piglets (n = 128) weaned at 21 days of age were used in a 35-day seeder model to evaluate the effects of dietary additives differing in active ingredients, chemical, and physical formulation, and dose on Salmonella colonization and shedding and intestinal microbial populations. Treatments were a negative control (basal diet), the positive control (challenged, basal diet), and six treatments similar to the positive control but supplemented with the following active ingredients (dose excluding essential oils or natural extracts): triglycerides with butyric acid (1.30 g kg(-1)); formic and citric acids and essential oils (2.44 g kg(-1)); coated formic, coated sorbic, and benzoic acids (2.70 g kg(-1)); salts of formic, sorbic, acetic, and propionic acids, their free acids, and natural extracts (2.92 g kg(-1)); triglycerides with caproic and caprylic acids and coated oregano oil (1.80 g kg(-1)); and caproic, caprylic, lauric, and lactic acids (1.91 g kg(-1)). On day 6, half the piglets (seeder pigs) in each group were orally challenged with a Salmonella Typhimurium nalidixic acid-resistant strain (4 × 10(9) and 1.2 × 10(9) log CFU per pig in replicate experiments 1 and 2, respectively). Two days later, they were transferred to pens with an equal number of contact pigs. Salmonella shedding was determined 2 days after challenge exposure and then on a weekly basis. On day 34 or 35, piglets were euthanized to sample tonsils, ileocecal lymph nodes, and ileal and cecal digesta contents. The two additives, both containing short-chain fatty acids and one of them also containing benzoic acid and the other one also containing essential oils, and supplemented at more than 2.70 g kg(-1), showed evidence of reducing Salmonella fecal shedding and numbers of coliforms and Salmonella in cecal digesta. However, colonization of tonsils and ileocecal lymph nodes by Salmonella was not affected. Supplementing butyric acid and medium-chain fatty acids at the applied dose failed to inhibit Salmonella contamination in the current experimental setup.

Efficacy of ciprofloxacin-gentamicin combination therapy in murine bubonic plague.

Potential benefits of combination antibiotic therapy for the treatment of plague have never been evaluated. We compared the efficacy of a ciprofloxacin (CIN) and gentamicin (GEN) combination therapy with that of each antibiotic administered alone (i) against Yersinia pestis in vitro and (ii) in a mouse model of bubonic plague in which animals were intravenously injected with antibiotics for five days, starting at two different times after infection (44 h and 56 h). In vitro, the CIN+GEN combination was synergistic at 0.5x the individual drugs' MICs and indifferent at 1x- or 2x MIC. In vivo, the survival rate for mice treated with CIN+GEN was similar to that observed with CIN alone and slightly higher than that observed for GEN alone 100, 100 and 85%, respectively when treatment was started 44 h post challenge. 100% of survivors were recorded in the CIN+GEN group vs 86 and 83% in the CIN and GEN groups, respectively when treatment was delayed to 56 h post-challenge. However, these differences were not statistically significant. Five days after the end of treatment, Y. pestis were observed in lymph nodes draining the inoculation site (but not in the spleen) in surviving mice in each of the three groups. The median lymph node log(10) CFU recovered from persistently infected lymph nodes was significantly higher with GEN than with CIN (5.8 vs. 3.2, p = 0.04) or CIN+GEN (5.8 vs. 2.8, p = 0.01). Taken as the whole, our data show that CIN+GEN combination is as effective as CIN alone but, regimens containing CIN are more effective to eradicate Y. pestis from the draining lymph node than the recommended GEN monotherapy. Moreover, draining lymph nodes may serve as a reservoir for the continued release of Y. pestis into the blood - even after five days of intravenous antibiotic treatment.

ß-Glucan plus ascorbic acid in neonatal calves modulates immune functions with and without Salmonella enterica serovar Dublin.

To determine if ß-glucan plus ascorbic acid affects adherence and pathogenicity of Salmonella Dublin and innate immune response in neonatal calves, 20 calves were fed control or supplemented diets (ß-glucan, 0.9 g/d, plus ascorbic acid, 500 mg/d) until d 23. On d 21, 5 calves per treatment received 2.4 × 10(8)CFU of S. Dublin orally. S. Dublin spread through intestinal tissues into mesenteric lymph nodes (MLN), spleen, and lung tissues within 48 h. All supplemented calves had less mRNA expression of IL-1 receptor antagonist in liver. Leukocyte cell surface markers changed in lung cells, but not in blood, MLN, or spleen. CD14 in lungs was greatest for calves receiving supplement and challenge, but CD18 in lungs was greater for challenged than control calves. Lung DEC205 was greatest for challenged calves with and without supplement compared to controls, but more lung cells expressed CD14 for all treated groups compared to controls. These data show that S. Dublin briefly inhabited the intestinal tract, moving quickly to spleen, MLN, and lung tissues. Lung tissue was modulated by S. Dublin, but supplement alone increased CD14 expressing cells. The supplement appears not to attenuate invasiness but modified some lung cell populations by 48h.

Pretreatment with arginine preserves intestinal barrier integrity and reduces bacterial translocation in mice.

OBJECTIVE: To evaluate the effects of arginine on intestinal barrier integrity and bacterial translocation (BT) in mice undergoing intestinal obstruction. METHODS: Mice were divided into 3 groups, treated for 7 d before surgical intervention with isocaloric and isoprotein diets. The ARG group received a diet containing 2% arginine, the IO (intestinal obstruction) and Sham groups, standard chow diet. On the eighth day of treatment, all animals received diethylenetriamine pentaacetic acid (DTPA) solution labeled with 99mTechnetium (99mTc-DTPA) by gavage for intestinal permeability analysis. After 90 min, the animals were anesthetized and the terminal ileum ligated. The Sham group only underwent laparotomy. After 4, 8, and 18 h, blood was collected for radioactivity determination. Samples of ileum were collected 18 h after surgery for histological analysis. In another set of animals, BT was evaluated. After 7 d of treatment, all animals received 10(8) CFU/mL of 99mTc-E.coli by gavage; 90 min later they were submitted to the surgical procedure described above. BT was determined by the uptake of 99mTc-E.coli in blood, mesenteric lymph nodes, liver, spleen, and lungs, assessed 18 h after the surgery. RESULTS: The intestinal permeability and BT were higher in the IO group when compared with the Sham group (P < 0.05). Arginine supplementation reduced intestinal permeability and BT to physiologic levels. Histological analysis showed mucosal ileum preservation in animals treated with arginine. CONCLUSION: Arginine was able to preserve barrier integrity, thus reducing BT.

The preliminary experimental and clinical study of the relationship between the pigment gallstone and intestinal mucosal barrier.

AIMS: To investigate the relations between the formation of pigment gallstone and the function of the intestinal mucosal barrier, as well as the underlying mechanism. METHODS: Eighty guinea pigs were randomly divided into three groups in which they were respectively given normal diet, gallstone-causing diet, and gallstone-formation diet with a supplementary intestinal mucosal protection compound known as glutamine. The model of pigment gallstone was established after 8 weeks of dietary administration. Indices about the function of the intestinal mucosal barrier and bacterial translocation were measured. Clinical cases were divided into three groups: control, cholesterol gallstone, and pigment gallstone, where the levels of plasma diamine oxidase (DAO), plasma endotoxin and the excretion rates of technetium 99m-diethylene triamine pentaacetic acid (99mTC-DTPA) in the urine of each group were measured. RESULTS: In the pigment gallstone group, the level of plasma DAO and endotoxin, the excretory ratio of lactulose and mannitol in urine, the bacterial translocation ratio in the celiac lymph nodes and the activities of beta-glucuronidase increased comparing to the control group. The gallstone-formation rate for the intestinal mucosal protection group (GLN) decreased, and other indices, except the activity of beta-glucuronidase, were all lower than that of gallstone-formation group. In the clinical experiment, the levels of plasma DAO and endotoxin, as well as the excretory rate of 99mTC-DTPA in urine were higher in the patients with gallstones than that in the control group. CONCLUSIONS: The formation of pigment gallstone was related to the abnormal function of the intestinal mucosal barrier. The abnormality in the function of the intestinal mucosal barrier probably induced the formation of gallstone by a bacterial translocation mechanism.

Enteral supplementation enriched with glutamine, fiber, and oligosaccharide prevents gut translocation in a bacterial overgrowth model.

BACKGROUND: Normal gut flora plays an important role in the intestinal mucosal barrier function under various critical conditions. The flora may alter after severe insults, such as trauma and shock. Enteral nutrition should preserve the gut environment; however, full support is usually difficult for severely ill patients because of impaired gastrointestinal motility. Currently, we have commercial enteral supplementation product enriched with glutamine, dietary fiber, and oligosaccharide (GFO) in Japan. This study examines the hypothesis that the enteral supplementation ameliorates gut injury induced by a bacterial overgrowth model, even in small volumes and quantities. MATERIALS: Balb/c mice received antibiotics (4 mg/mL of streptomycin) in their drinking water for 4 days to kill the normal gut flora after which they were orally inoculated with a streptomycin-resistant strain of Escherichia coli, known as E. coli C-25. The mice that were administered bacterial monoassociation received 0.5 mL of GFO twice daily (GFO group) or 10% of glucose solution (GLU group). Unsupplemented drinking water was used for control animals (control) whose gut flora was normal. The mice were killed and their mesenteric lymph nodes complex was harvested and processed to test gut bacterial translocation. The cecal population levels of bacteria and ileum histology were also examined. RESULTS: The incidence and magnitude of gut translocation to the lymph nodes complex in the GLU group were significantly higher than those in the control (p < 0.01). Treatment with GFO prevented the gut translocation although animals in the GFO group had same level of the cecal bacterial population. Histologic findings in the ileum were not different between the GLU and GFO. CONCLUSION: GFOs supplement prevented gut translocation for bacterial overgrowth even in small volumes and quantities. The intestinal histologic findings could not explain the protective mechanisms of GFO. Further studies may be needed to elucidate the benefit of the partial enteral nutrition.

Dietary glutamine and oral antibiotics each improve indexes of gut barrier function in rat short bowel syndrome.

Short bowel syndrome (SBS) is associated with gut barrier dysfunction. We examined effects of dietary glutamine (GLN) or oral antibiotics (ABX) on indexes of gut barrier function in a rat model of SBS. Adult rats underwent a 60% distal small bowel + proximal colonic resection (RX) or bowel transection (TX; control). Rats were pair fed diets with or without l-GLN for 20 days after operation. Oral ABX (neomycin, metronidazole, and polymyxin B) were given in some RX rats fed control diet. Stool secretory immunoglobulin A (sIgA) was measured serially. On day 21, mesenteric lymph nodes (MLN) were cultured for gram-negative bacteria. IgA-positive plasma cells in jejunum, stool levels of flagellin- and lipopolysaccharide (LPS)-specific sIgA, and serum total, anti-flagellin- and anti-LPS IgG levels were determined. RX caused gram-negative bacterial translocation to MLN, increased serum total and anti-LPS IgG and increased stool total sIgA. After RX, dietary GLN tended to blunt bacterial translocation to MLN (-29%, P = NS) and significantly decreased anti-LPS IgG levels in serum, increased both stool and jejunal mucosal sIgA and increased stool anti-LPS-specific IgA. Oral ABX eliminated RX-induced bacterial translocation, significantly decreased total and anti-LPS IgG levels in serum, significantly decreased stool total IgA and increased stool LPS-specific IgA. Partial small bowel-colonic resection in rats is associated with gram-negative bacterial translocation from the gut and a concomitant adaptive immune response to LPS. These indexes of gut barrier dysfunction are ameliorated or blunted by administration of dietary GLN or oral ABX, respectively. Dietary GLN upregulates small bowel sIgA in this model.

[Relationship between pigment gallstone and intestinal barrier function: experiment with guinea pigs and clinical observations].

OBJECTIVE: To investigate the relationship between the intestinal barrier function and pigment gallstone formation. METHODS: Ninety Guinea pigs were divided randomly into 3 groups: normal control (CON) group receiving normal forage, pigment gallstone (PS) group receiving pigment gallstone-forming forage, and intestinal mucosa protection group receiving pigment gallstone-forming forage with supplemental glutamine intestinal (GLN), a mucosa protector. The guinea pigs were observed for 8 weeks, the gallstone-forming rate, plasma diamine oxidase ( DAO), serum endotoxin, proportionality of urine lactulose/mannitol, and biliary beta-glucuronidase were detected. PCR was used to detect the bacteria in abdominal lymph node taking 16SrRNA as the target gene common in most bacteria. 32 gallstone patients, 16 with cholesterol gallstone and 16 with pigmental gallstone, and 27 patients with non-gastroenterological diseases, as controls, underwent detection of the plasma DAO and serum endotoxin. Another 109 gallstone patients, 31 with cholesterol gallstone and 78 with pigmental gallstone, and 21 patients with nongastroenterological diseases, as controls, underwent detection of urine technetium-labeled diethylenetriamine-pentaacetate (99mTc-DTPA). RESULTS: The gallstone-forming rate of the guinea pigs of the GLN group was 44.4% was, significantly lower than that of the PS group (73.9%, P < 0.05). The plasma DAO, serum endotoxin levels, proportionality of urine lactulose/mannitol, and activity of biliary beta-glucuronidase of the PS group were all significantly higher than those of the CON group (P < 0.05 or P < 0.01). The plasma endotoxin level of the pigmental GLN group was significantly lower than that of the PS group (P < 0.01). The positive rate of bacteria in abdominal lymph node of the PS group was 80%, significantly higher than those of the CON and GLN groups (30% and 45% respectively, P < 0.01 and P < 0.05). The level of plasma DAO and endotoxin of the pigmental gallstone patients were significantly higher than those of the controls (P < 0.05 and P < 0.01). The urine 99mTc-DTPA excretion rate of gallstone patients was 11.4%, significantly higher than that of the controls (4.7%, P < 0.01). CONCLUSION: Intestinal barrier function is correlated with pigment gallstone forming. Intestinal barrier function disorder may promote pigment gallstone formation through bacteria translocation, endotoxemia, and increase of biliary beta-glucuronidase.

Assessment of different formulations of oral Mycobacterium bovis Bacille Calmette-Guérin (BCG) vaccine in rodent models for immunogenicity and protection against aerosol challenge with M. bovis.

Bovine tuberculosis (bTB) caused by infection with Mycobacterium bovis is causing considerable economic loss to farmers and Government in the United Kingdom as its incidence is increasing. Efforts to control bTB in the UK are hampered by the infection in Eurasian badgers (Meles meles) that represent a wildlife reservoir and source of recurrent M. bovis exposure to cattle. Vaccination of badgers with the human TB vaccine, M. bovis Bacille Calmette-Guérin (BCG), in oral bait represents a possible disease control tool and holds the best prospect for reaching badger populations over a wide geographical area. Using mouse and guinea pig models, we evaluated the immunogenicity and protective efficacy, respectively, of candidate badger oral vaccines based on formulation of BCG in lipid matrix, alginate beads, or a novel microcapsular hybrid of both lipid and alginate. Two different oral doses of BCG were evaluated in each formulation for their protective efficacy in guinea pigs, while a single dose was evaluated in mice. In mice, significant immune responses (based on lymphocyte proliferation and expression of IFN-gamma) were only seen with the lipid matrix and the lipid in alginate microcapsular formulation, corresponding to the isolation of viable BCG from alimentary tract lymph nodes. In guinea pigs, only BCG formulated in lipid matrix conferred protection to the spleen and lungs following aerosol route challenge with M. bovis. Protection was seen with delivery doses in the range 10(6)-10(7) CFU, although this was more consistent in the spleen at the higher dose. No protection in terms of organ CFU was seen with BCG administered in alginate beads or in lipid in alginate microcapsules, although 10(7) in the latter formulation conferred protection in terms of increasing body weight after challenge and a smaller lung to body weight ratio at necropsy. These results highlight the potential for lipid, rather than alginate, -based vaccine formulations as suitable delivery vehicles for an oral BCG vaccine in badgers.

Probiotics and blueberry attenuate the severity of dextran sulfate sodium (DSS)-induced colitis.

We studied the anti-inflammatory properties of probiotic strains and blueberry in a colitis model. The disease activity index (DAI) was significantly lower on days 9 and 10 in all groups compared to the colitis control. Myeloperoxidase (MPO) and bacterial translocation to the liver and to the mesenteric lymph nodes (MLN) decreased significantly in all groups compared to colitis control. Cecal Enterobacteriaceae count decreased significantly in blueberry with and without probiotics compared to the other groups. Lactobacillus plantarum reisolated from the cecal content in the presence of blueberry, contrary to Lactobacillus fermentum. Colonic MDA decreased significantly in all groups, except the L. fermentum group, compared to the colitis control. The cecal concentration of acetic, propionic, and butyricbutyric acid was significantly higher in the L. plantarum group, while the L. fermentum group yielded the highest concentration of lactic acid compared with all other groups. Lactobacillus plantarum DSM 15313, Lactobacillus fermentum 35D, and blueberry alone and in combination improve the DAI, reduce bacterial translocation, and reduce inflammation.

Combination of allopurinol and hyperbaric oxygen therapy: a new treatment in experimental acute necrotizing pancreatitis?

AIM: To investigate the individual and combined effects of allopurinol and hyperbaric oxygen (HBO) therapy on biochemical and histopathological changes, oxidative stress, and bacterial translocation (BT) in the experimental rat acute pancreatitis (AP). METHODS: Eighty-five Sprague-Dawley rats were included in the study. Fifteen of the eighty-five rats were used as controls (sham, Group I). AP was induced via intraductal taurocholate infusion in the remaining seventy rats. Rats that survived to induction of acute necrotizing pancreatitis were randomized into four groups. Group II received saline, Group III allopurinol, Group IV allopurinol plus HBO and Group V HBO alone. Serum amylase levels, oxidative stress parameters, BT and histopathologic scores were determined. RESULTS: Serum amylase levels were lower in Groups III, IV and V compared to Group II (974 +/- 110, 384 +/- 40, 851 +/- 56, and 1664 +/- 234 U/L, respectively, P < 0.05, for all). Combining the two treatment options revealed significantly lower median [25-75 percentiles] histopathological scores when compared to individual administrations (13 [12.5-15] in allopurinol group, 9.5 [7-11.75] in HBO group, and 6 [4.5-7.5] in combined group, P < 0.01). Oxidative stress markers were significantly better in all treatment groups compared to the controls. Bacterial translocation into the pancreas and mesenteric lymph nodes was lower in Groups III, IV and V compared to Group II (54%, 23%, 50% vs 100% for translocation to pancreas, and 62%, 46%, 58% vs 100% for translocation to mesenteric lymph nodes, respectively, P < 0.05 for all). CONCLUSION: The present study confirms the benefit of HBO and allopurinol treatment when administered separately in experimental rat AP. Combination of these treatment options appears to prevent progression of pancreatic injury parameters more effectively.