Type 2-resistant starch and Lactiplantibacillus plantarum NCIMB 8826 result in additive and interactive effects in diet-induced obese mice.

Little is known about how combining a probiotic with prebiotic dietary fiber affects the ability of either biotic to improve health. We hypothesized that prebiotic, high-amylose maize type 2-resistant starch (RS) together with probiotic Lactiplantibacillus plantarum NCIMB8826 (LP) as a complementary synbiotic results in additive effects on the gut microbiota in diet-induced obese mice and other body sites. Diet-induced obese C57BL/6J male mice were fed a high-fat diet adjusted to contain RS (20% by weight), LP (109 cells every 48 hours), or both (RS+LP) for 6 weeks. As found for mice fed RS, cecal bacterial alpha diversity was significantly reduced in mice given RS+LP compared with those fed LP and high-fat controls. Similarly, both RS+LP and RS also conferred lower quantities of cecal butyrate and serum histidine and higher ileal TLR2 transcript levels and adipose tissue interleukin-6 protein. As found for mice fed LP, RS+LP-fed mice had higher colonic tissue TH17 cytokines, reduced epididymal fat immune and oxidative stress responses, reduced serum carnitine levels, and increased transcript quantities of hepatic carnitine palmitoyl transferase 1α. Notably, compared with RS and LP consumed separately, there were also synergistic increases in colonic glucose and hepatic amino acids as well antagonistic effects of LP on RS-mediated increases in serum adiponectin and urinary toxin levels. Our findings show that it is not possible to fully predict outcomes of synbiotic applications based on findings of the probiotic or the prebiotic tested separately; therefore, studies should be conducted to test new synbiotic formulations.

Viability and In Vitro Gastrointestinal Transit Tolerance of Multispecies Probiotic Combinations Incorporated into Orange Juice and Drinking Water.

Little is known about how combining probiotics affects the storage survival and functional performance of individual probiotics when incorporated into non-dairy drinks. Viability of Lacticaseibacillus rhamnosus GG (LG), Limosilactobacillus reuteri ATCC 55730 (LR), Bifidobacterium animalis subsp. lactis BB-12 (Bb), and Propionibacterium jensenii 702 (PJ), either alone or in multi-species combinations included in orange juice (OJ), were assessed during storage in refrigerated conditions and compared with bottled water (BW). The tolerance of probiotics included in refrigerated OJ to simulated gastrointestinal conditions was also examined. LG and LR viabilities were significantly higher in OJ than in BW (p ≤ 0.001), while the reverse was evident for PJ. Bb maintained high viability in both drinks. LG-PJ in both drinks and Bb-PJ in BW resulted in greater viabilities among the paired combinations compared to their respective monocultures when incorporated separately (p ≤ 0.001). The viability of LG in the LG-Bb-PJ combination improved significantly in BW compared with LG alone (p ≤ 0.001). OJ did not alter bacterial tolerance to simulated gastric juice but diminished tolerance to simulated intestinal juice (SIJ). In all combinations, tolerance of LG and LR to SIJ was improved, whereas tolerance of PJ declined significantly compared with respective monocultures (p ≤ 0.001). In conclusion, probiotic storage stability and gastrointestinal transit tolerance were species-dependent and affected by carrier type and combinations. These effects should be considered when formulating probiotic products.

Maternal probiotic intake attenuates ileal Crh receptor gene expression in maternally separated rat offspring.

Corticotropin-releasing hormone (Crh) and its receptors (Crhr) mediate stress-induced gastrointestinal dysfunctions. Neonatal maternal separation (MS) increased ileal Crhr1 transcript quantities in young rat offspring. Exposure to either MS or adulthood restraint stress increased ileal Crhr1 and Crhr2 transcript quantities only in adult female offspring. Maternal probiotic intervention reversed Crhr overexpression, suggesting a potential early prophylaxis against stress-induced gut dysfunctions.

Growth of Salmonella and Other Foodborne Pathogens on Inoculated Inshell Pistachios during Simulated Delays between Hulling and Drying.

During harvest, pistachios are hulled, separated in water into floater and sinker streams (in large part on the basis of nut density), and then dried before storage. Higher prevalence and levels of Salmonella were previously observed in floater pistachios, but contributing factors are unclear. To examine the behavior of pathogens on hulled pistachios during simulated drying delays, floater and sinker pistachios collected from commercial processors were inoculated at 1 or 3 log CFU/g with cocktails of Salmonella and in some cases Escherichia coli O157:H7 or Listeria monocytogenes and incubated for up to 30 h at 37°C and 90% relative humidity. Populations were measured by plating onto tryptic soy agar and appropriate selective agars. In most cases, no significant growth (P > 0.05) of Salmonella was observed in the first 3 h after inoculation in hulled floaters and sinkers. Growth of Salmonella was greater on floater pistachios than on corresponding sinkers and on floater pistachios with ≥25% hull adhering to the shell surface than on corresponding floaters with <25% adhering hull. Maximum Salmonella populations (2 to 7 log CFU/g) were ∼2-log higher on floaters than on corresponding sinkers. The growth of E. coli O157:H7 and Salmonella on hulled pistachios was similar, but a longer lag time (approximately 11 h) and significantly lower maximum populations (4 versus 5 to 6 log CFU/g; P < 0.05) were predicted for L. monocytogenes. Significant growth of pathogens on hulled pistachios is possible when delays between hulling and drying are longer than 3 h, and pathogen growth is enhanced in the presence of adhering hull material.

Growth of Salmonella on Inoculated Inhull Pistachios during Postharvest Handling.

Salmonella has been isolated from dried pistachios in both postharvest and retail surveys. The source of Salmonella in pistachios is unknown, but introduction is possible at points during production, harvest, and postharvest activities. To examine the behavior of Salmonella on pistachios during simulated postharvest conditions, early-, mid-, and late-season inhull pistachios were collected from two commercial processors over five different harvests. Pistachios were inoculated with cocktails of nalidixic acid- or rifampin-resistant Salmonella at 0.64 to 1.59 log CFU/g (low) or 2.73 to 3.27 or 4.29 to 4.31 log CFU/g (high) and were incubated for up to 30 h under commercially relevant conditions (23, 35, or 37°C and 50 or 90% relative humidity [RH]). Populations of Salmonella were measured by plating onto tryptic soy agar and CHROMagar Salmonella with added nalidixic acid or rifampin. Individual growth curves at the same temperature and RH differed significantly among different lots of pistachios. Except for a single late-season lot in which no significant growth was observed, Salmonella multiplied under all storage conditions. In the first 3 h after inoculation, insignificant (most cases) to small (0.41 to 0.67 log CFU/g) but significant ( P < 0.05) mean increases in Salmonella populations were measured; the mean predicted time to achieve maximum populations (5 to 8 log CFU/g) was 16 ± 4 h. In paired samples, longer lag phases, lower growth rates, and lower maximum increases were observed with inoculated inhull pistachios incubated at 23°C and 50% RH compared with 35 or 37°C and 90% RH. Similar growth curves were observed at the low and high inoculum levels; throughout the 30 h of incubation, Salmonella populations were consistently ∼1 to 2 log CFU/g lower on pistachios inoculated at the low inoculum level. Managing the time between harvesting and hulling will reduce the potential for growth of Salmonella on pistachios during postharvest handling.

Lactobacillus plantarum bacteriocin is associated with intestinal and systemic improvements in diet-induced obese mice and maintains epithelial barrier integrity in vitro.

We investigated the Lactobacillus plantarum bacteriocin plantaricin EF (PlnEF) system for its contributions to L. plantarum mediated benefits in a mouse model of diet-induced obesity. C57BL/6J mice on a high-fat diet (HFD) were administered a rifampicin resistant mutant of L. plantarum NCMIB8826 (NICMB8826-R) or an isogenic ΔplnEFI mutant strain, LM0419, every 48 h for nine weeks. Mice fed wild-type L. plantarum, but not LM0419, reduced their consumption of the HFD starting three weeks into the study and exhibited an overall 10% reduction in weight gain. The responses were independent of glucose homeostasis, as both NCMIB8826-R and LM0419 fed mice had improved oral glucose tolerance compared to sham controls. Although bacteriocins have antibacterial properties, the ileal, cecal, and fecal microbiota and cecocolic metabolomes were unchanged between mice fed either wild-type L. plantarum or the ΔplnEFI mutant. Instead, only mice fed NCMIB8826-R showed an increased production of ZO-1 in ileal tissues. To verify a potential role for the plantaricin EF system in supporting intestinal epithelial function, synthesized PlnEF peptides were applied to Caco-2 cell monolayers challenged with TNF-α and IFN-γ. The combination of PlnE and PlnF were required to prevent sustained cytokine-induced losses to Caco-2 cell para- and transcellular permeability and elevated IL-8 levels. In conclusion, this study shows that probiotic L. plantarum ameliorates the effects of obesogenic diets through a mechanism that involves the plantaricin EF system and likely includes L. plantarum - induced fortification of the intestinal epithelium.

Supplementation of Lactobacillus plantarum Improves Markers of Metabolic Dysfunction Induced by a High Fat Diet.

Obesity is a prevalent chronic condition in many developed and developing nations that raises the risk for developing heart disease, stroke, and diabetes. Previous studies have shown that consuming particular probiotic strains of Lactobacillus is associated with improvement in the obese and diabetic phenotype; however, the mechanisms of these beneficial effects are not well understood. In this study, C57BL/6J male mice were fed a lard-based high fat diet for 15 weeks with Lactobacillus plantarum supplementation NCIMB8826 (Lp) between weeks 10 and 15 ( n = 10 per group). Systemic metabolic effects of supplementation were analyzed by NMR metabolomics, protein expression assays, gene transcript quantification, and 16S rRNA marker gene sequencing. Body and organ weights were not significantly different with Lp supplementation, and no microbiota community structure changes were observed in the cecum; however, L. plantarum numbers were increased in the treatment group according to culture-based and 16S rRNA gene quantification. Significant differences in metabolite and protein concentrations (serum, liver, and colon), gene expression (ileum and adipose), and cytokines (colon) were observed between groups with increases in the gene expression of tight junction proteins in the ileum and cecum and improvement of some markers of glucose homeostasis in blood and tissue with Lp supplementation. These results indicate Lp supplementation impacts systemic metabolism and immune signaling before phenotypic changes and without large-scale changes to the microbiome. This study supports the notion that Lp is a beneficial probiotic, even in the context of a high fat diet.

Microbiota, metabolome, and immune alterations in obese mice fed a high-fat diet containing type 2 resistant starch.

SCOPE: We examined the intestinal and systemic responses to incorporating a type 2 resistant starch (RS) into a high fat diet fed to obese mice. METHODS AND RESULTS: Diet-induced obese, C57BL/6J male mice were fed an HF diet without or with 20% (by weight) high-amylose maize resistant starch (HF-RS) for 6 weeks. Serum adiponectin levels were higher with RS consumption, but there were no differences in weight gain and adiposity. With HF-RS, the expression levels of ileal TLR2 and Reg3g and cecal occludin, TLR2, TLR4, NOD1 and NOD2 were induced; whereas colonic concentrations of the inflammatory cytokine IL-17A declined. The intestinal, serum, liver, and urinary metabolomes were also altered. HF-RS resulted in lower amino acid concentrations, including lower serum branched chain amino acids, and increased quantities of urinary di/trimethylamine, 3-indoxylsulfate, and phenylacetylglycine. Corresponding to these changes were enrichments in Bacteroidetes (S24-7 family) and certain Firmicutes taxa (Lactobacillales and Erysipelotrichaceae) with the HF-RS diet. Parabacteroides and S24-7 positively associated with cecal maltose concentrations. These taxa and Erysipelotrichaceae, Allobaculum, and Bifidobacterium were directly correlated with uremic metabolites. CONCLUSION: Consumption of RS modified the intestinal microbiota, stimulated intestinal immunity and endocrine-responses, and modified systemic metabolomes in obese mice consuming an otherwise obesogenic diet.

Modeling the risk of salmonellosis from consumption of pistachios produced and consumed in the United States.

The risk of salmonellosis from consumption of pistachios produced and consumed in the U.S. was assessed through quantitative microbial risk assessment. Data on Salmonella prevalence and concentration on pistachios, nut crop volume, storage times and temperatures during processing and handling, and reductions during storage or from roasting were derived from laboratory experiments, published literature, and industry expert opinion. Uncertainty was analyzed via what-if scenarios for Salmonella prevalence, concentration, storage reduction, treatment variability, portion of crop treated, and increased consumption. The estimated U.S. incidence of salmonellosis when 100% of pistachios were exposed to a 4 ± 0 log reduction treatment averaged 1.4 cases per billion servings, or <1 case/year, without considering Salmonella decline during storage. Including Salmonella decline during storage reduced the salmonellosis estimates approximately 10-fold. The predicted arithmetic mean number of cases associated with individual 500,000-kg storage silos, contaminated at the highest observed levels, ranged from 5 to 530 when the product was consumed untreated, but was reduced to below 1 case per silo when a 4 ± 0 log reduction treatment was applied. Assuming a uniform 4-log reduction treatment is applied to 100% of the crop and there is no decline of Salmonella during storage, the assessment indicates the following: 10-fold increases in either Salmonella prevalence or concentration, 2-fold increases in both prevalence and concentration, or consumption of >0.05% of untreated product volume yield an arithmetic mean risk of >1 case/year.

Altered nociceptive, endocrine, and dorsal horn neuron responses in rats following a neonatal immune challenge.

The neonatal period is characterized by significant plasticity where the immune, endocrine, and nociceptive systems undergo fine-tuning and maturation. Painful experiences during this period can result in long-term alterations in the neurocircuitry underlying nociception, including increased sensitivity to mechanical or thermal stimuli. Less is known about the impact of neonatal exposure to mild inflammatory stimuli, such as lipopolysaccharide (LPS), on subsequent inflammatory pain responses. Here we examine the impact of neonatal LPS exposure on inflammatory pain sensitivity and HPA axis activity during the first three postnatal weeks. Wistar rats were injected with LPS (0.05mg/kg IP, Salmonella enteritidis) or saline on postnatal days (PNDs) 3 and 5 and later subjected to the formalin test at PNDs 7, 13, and 22. One hour after formalin injection, blood was collected to assess corticosterone responses. Transverse spinal cord slices were also prepared for whole-cell patch clamp recording from lumbar superficial dorsal horn neurons (SDH). Brains were obtained at PND 22 and the hypothalamus was isolated to measure glucocorticoid (GR) and mineralocorticoid receptor (MR) transcript expression using qRT-PCR. Behavioural analyses indicate that at PND 7, no significant differences were observed between saline- or LPS-challenged rats. At PND 13, LPS-challenged rats exhibited enhanced licking (p<.01), and at PND 22, increased flinching in response to formalin injection (p<.05). LPS-challenged rats also displayed increased plasma corticosterone at PND 7 and PND 22 (p<.001) but not at PND 13 following formalin administration. Furthermore, at PND 22 neonatal LPS exposure induced decreased levels of GR mRNA and increased levels of MR mRNA in the hypothalamus. The intrinsic properties of SDH neurons were similar at PND 7 and PND 13. However, at PND 22, ipsilateral SDH neurons in LPS-challenged rats had a lower input resistance compared to their saline-challenged counterparts (p<.05). These data suggest neonatal LPS exposure produces developmentally regulated changes in formalin-induced behavioural responses, corticosterone levels, and dorsal horn neuron properties following noxious stimulation later in life. These findings highlight the importance of immune activation during the neonatal period in shaping pain sensitivity later in life. This programming involves both spinal cord neurons and the HPA axis.

Effect of maternal probiotic intervention on HPA axis, immunity and gut microbiota in a rat model of irritable bowel syndrome.

OBJECTIVE: To examine whether maternal probiotic intervention influences the alterations in the brain-immune-gut axis induced by neonatal maternal separation (MS) and/or restraint stress in adulthood (AS) in Wistar rats. DESIGN: Dams had free access to drinking water supplemented with Bifidobacterium animalis subsp lactis BB-12® (3 × 10(9) CFU/mL) and Propionibacterium jensenii 702 (8.0 × 10(8) CFU/mL) from 10 days before conception until postnatal day (PND) 22 (weaning day), or to control ad lib water. Offspring were subjected to MS from PND 2 to 14 or left undisturbed. From PND 83 to 85, animals underwent 30 min/day AS, or were left undisturbed as controls. On PND 24 and 86, blood samples were collected for corticosterone, ACTH and IgA measurement. Colonic contents were analysed for the composition of microflora and luminal IgA levels. RESULTS: Exposure to MS significantly increased ACTH levels and neonatal fecal counts of aerobic and anaerobic bacteria, E. coli, enterococci and clostridia, but reduced plasma IgA levels compared with non-MS animals. Animals exposed to AS exhibited significantly increased ACTH and corticosterone levels, decreased aerobic bacteria and bifidobacteria, and increased Bacteroides and E. coli counts compared to non-AS animals. MS coupled with AS induced significantly decreased anaerobes and clostridia compared with the non-stress adult controls. Maternal probiotic intervention significantly increased neonatal corticosterone levels which persisted until at least week 12 in females only, and also resulted in elevated adult ACTH levels and altered neonatal microflora comparable to that of MS. However, it improved plasma IgA responses, increased enterococci and clostridia in MS adults, increased luminal IgA levels, and restored anaerobes, bifidobacteria and E. coli to normal in adults. CONCLUSION: Maternal probiotic intervention induced activation of neonatal stress pathways and an imbalance in gut microflora. Importantly however, it improved the immune environment of stressed animals and protected, in part, against stress-induced disturbances in adult gut microflora.

Disgust elevates core body temperature and up-regulates certain oral immune markers.

Recent findings suggest that disgust can activate particular aspects of the immune system. In this study we examine whether disgust can also elevate core body temperature (BT), a further feature of an immune response to disease. In addition, we also examined whether food based disgust--a core eliciting stimulus--may be a more potent immune stimulus than non-food based disgust. Healthy males were randomly assigned to view one of four sets of images--food disgust, non-food disgust, food control and negative emotion control. Measures of BT, salivary immune and related markers, and self-report data, were collected before, and at two time points after image viewing. Disgust elevated BT relative to the negative emotion control condition, as did food images. Different mechanisms appeared to account for these effects on BT, with higher initial levels of Tumor Necrosis Factor alpha (TNF-a) and disgust, predictive of BT increases in the disgust conditions. Disgust also increased TNF-a, and albumin levels, relative to the control conditions. Type of disgust exerted little effect. These findings further support the idea that disgust impacts upon immune function, and that disgust serves primarily a disease avoidance function.

The effect of disgust on oral immune function.

Disgust motivates avoidance of pathogen sources, but whether its role in disease avoidance extends into activating the immune system is unexplored. This was tested here by comparing oral immune markers before and after a disgust induction, relative to neutral and negative induction control groups. The disgust group, but not controls, revealed an oral inflammatory response, with increased salivary tumor necrotizing factor alpha and albumin, as well as a down-regulation of immunoglobulin A (SIgA) secretion. It has been hypothesized that disgust evolved in animals to clear toxins from the oral cavity by gaping and increased salivary flow. Our data suggest down-regulated SIgA secretion may be a vestige of this response so as to conserve protein, while the inflammatory reaction may reflect an adaptive response to disease threat, selectively triggered by disgust. The broader implications of these data for a discrete neuro-gut-immune axis are examined.

A scale for measuring hygiene behavior: development, reliability and validity.

BACKGROUND: There is currently no general self-report measure for assessing hygiene behavior. This article details the development and testing of such a measure. METHODS: In studies 1 to 4, a total of 855 participants were used for scale and subscale development and for reliability and validity testing. The latter involved establishing the relationships between self-reported hygiene behavior and existing measures, hand hygiene behavior, illness rates, and a physiological marker of immune function. In study 5, a total of 507 participants were used to assess the psychometric properties of the final revised version of the scale. RESULTS: The final 23-item scale comprised 5 subscales: general, household, food-related, handwashing technique, and personal hygiene. Studies 1 to 4 confirmed the scale's reliability and validity, and study 5 confirmed the scale's 5-factor structure. CONCLUSIONS: The scale is potentially suitable for multiple uses, in various settings, and for experimental and correlational approaches.