The present study investigated the patterns of microorganisms in an artificial larval diet during Dacus ciliatus (Diptera; Tephritidae) larval development. Microbial population contents in the diet of total heterotrophic bacteria, yeast and molds, coliform and lactobacilli, and their dynamics during development, were monitored. Initially, the microbial composition in diet trays failing to produce viable pupae and in trays successfully producing pupae and adult flies was characterized. The failing diet trays contained large populations of lactobacilli that increased during larval development, and low populations of coliforms. In contrast, the successful diet showed an increasing population of coliforms and a low, or undetected, population of lactobacilli. To study the hypothesis that lactobacilli affect D. ciliatus larval development, we conducted controlled inoculation experiments in which Lactobacillus plantarum was added into fresh diet at the time of egg seeding. L. plantarum inoculated trays showed no production of D. ciliatus. Control trays without lactobacilli inoculation showed variable results. One tray successfully produced viable pupae and adults, and showed a slight and slow increase in the indigenous populations of lactobacilli. The second tray, however, failed to produce pupae and showed a fast increase of the indigenous lactobacilli to very high levels. Monitored pH trends in L. plantarum-inoculated diet showed a sharp pH decrease during the first 4 days of larval development from 5 to less than 4 units, while successful diet, producing viable D. ciliatus pupae and adults, showed a moderate pH drop during most of the larval development period. The paper discusses the possible ecological interactions between D. ciliatus larvae, the microbial content of the diet and the physical properties of the diet. The discussion also points out at the usefulness of this approach in understanding and managing mass production parameters of tephritid fruit flies industrial diets used for Sterile Insect Technique.

The importance of the crop is often underestimated in poultry production. In addition to storing ingested feed, it also can impact nutrient digestion by digesta softening and the initial activity of feed (endogenous and exogenous) and microbial enzymes. The crop represents the first major defence against poultry pathogens and zoonotic organisms with well established adaptive and innate immune function, and a lactobacilli dominated microbiota capable of reducing the passage of these organisms further along the digestive tract. However, the potential to improve bird productivity and health, as well as affect meat and egg safety, are influenced by the nature of the diet, and in particular feed entry and extended presence in the crop. This is required to promote lactobacilli fermentation, the production of lactic acid and other volatile fatty acids, and the lowering of crop pH. Management practices such as meal feeding and the use of lighting programs with extended dark periods encourage crop utilisation. Further, the use of feed additives such as prebiotics and probiotics may enhance crop function, which in turn contributes to well-being of the entire digestive tract. A healthy and functional crop, along with other segments of digestive tract, has increased importance in an era of reduced antibiotic use in poultry feeds.

The neonatal period is often polarised to T helper (Th2) response at the time of birth, predisposing offspring to allergic disorders. Passive immunity through the mother’s milk is critical for immune system development of newborns. Probiotics have been proposed to harmonise Th1/Th2 imbalance in allergic conditions in adults. In the present study, the anti-allergic effects of feeding probiotic Lactobacillus rhamnosus-fermented milk (PFM) either to dams during the suckling period or to their offspring after weaning individually or else in successive periods against ovalbumin (OVA)-induced allergy in newborns was analysed. After allergen sensitisation, physical symptoms of allergy, gut immune response, humoral immune response and cell-mediated response through interleukins were detected. Consumption of PFM by mothers and offspring showed a reduction (P<0·01) in physical allergic symptoms in newborns with an increase (P<0·01) in the numbers of goblet and IgA+ cells in the small intestine. Similarly, considerable (P<0·001) decreases in OVA-specific antibodies (IgE, IgG, IgG1) and ratios of IgE/IgG2a and IgG1/IgG2a in the sera of newborn mice were recorded. A decrease in IL-4 and an increase in interferon-γ levels further confirmed the shift from Th2 to Th1 pathway in PFM-fed mice. It is logical to conclude that the timing of PFM intervention in alleviating allergic symptoms is critical, which was found to be most effective when mothers were fed during the suckling period.

Lactobacilli are indigenous microbes of the stomach of rodents, with much lower numbers being present in mice fed a purified diet than in those fed a non-purified diet. We postulated that gastric emptying (GE) is responsible for the different colonisation levels of lactobacilli and tested this hypothesis in the present study. BALB/cCr Slc mice were fed either a non-purified diet or a purified diet for 2 weeks. The number of gastric tissue-associated lactobacilli was lower in mice fed the purified diet than in those fed the non-purified diet. GE, estimated by measuring the food recovered from the stomach, was higher in mice fed the purified diet than in those fed the non-purified diet and correlated negatively with the number of lactobacilli. Mice fed the non-purified diet exhibited lower GE rates even when lactobacilli were eliminated by ampicillin administration through the drinking-water, suggesting that GE is the cause but not the consequence of different Lactobacillus colonisation levels. The plasma concentrations of acylated ghrelin, a gastric hormone that promotes GE, were higher in mice fed the purified diet than in those fed the non-purified diet. There was a negative correlation between GE and the number of lactobacilli in mice fed the non-purified diet, the purified diet, and the purified diet supplemented with sugarbeet fibre (200 g/kg diet) or carboxymethyl cellulose (40 g/kg diet). We propose that a higher GE rate contributes, at least in part, to lower gastric colonisation levels of lactobacilli in mice fed a purified diet.

Thylakoid membranes derived from green leaf chloroplasts affect appetite-regulating hormones, suppress food intake, reduce blood lipids and lead to a decreased body weight in animals and human subjects. Thylakoids also decrease the intestinal in vitro uptake of methyl-glucose in the rat. The aim of this study was to investigate the effect of dietary thylakoids on the gut microbiota composition, mainly the taxa of lactobacilli and bifidobacteria, in rats fed either a thylakoid-enriched diet or a control diet for 10 d. At the same time, a glucose-tolerance test in the same rats was also performed. Food intake was significantly decreased in the thylakoid-fed rats compared with the control-fed rats over the 10-d study. An oral glucose tolerance test after 10 d of thylakoid- or control-food intake resulted in significantly reduced plasma insulin levels in the thylakoid-fed rats compared with the control-fed rats, while no difference was observed for blood glucose levels. Analysis of gut bacteria showed a significant increase of lactobacilli on the ileal mucosa, specifically Lactobacillus reuteri, in the rats fed the thylakoid diet compared with rats fed the control diet, while faecal lactobacilli decreased. No difference in bifidobacteria between the thylakoid and control groups was found. Analyses with terminal restriction fragment length polymorphism and principal component analysis of faeces demonstrated different microbial populations in the thylakoid- and control-fed animals. These findings indicate that thylakoids modulate the gut microbial composition, which might be important for the regulation of body weight and energy metabolism.

Probiotics are live microorganisms that, when ingested in adequate amounts, provide health benefits to the host. The strains most frequently used as probiotics include lactic acid bacteria and bifidobacteria, which are isolated from traditional fermented products and the gut, faeces and breast milk of human subjects. The identification of microorganisms is the first step in the selection of potential probiotics. The present techniques, including genetic fingerprinting, gene sequencing, oligonucleotide probes and specific primer selection, discriminate closely related bacteria with varying degrees of success. Additional molecular methods, such as denaturing gradient gel electrophoresis/temperature gradient gel electrophoresis and fluorescence in situ hybridisation, are employed to identify and characterise probiotics. The ability to examine fully sequenced genomes has accelerated the application of genetic approaches to elucidate the functional roles of probiotics. One of the best-demonstrated clinical benefits of probiotics is the prevention and treatment of acute and antibiotic-associated diarrhoea; however, there is mounting evidence for a potential role for probiotics in the treatment of allergies and intestinal, liver and metabolic diseases. These positive effects are generally attributed to the ability of probiotics to regulate intestinal permeability, normalise host intestinal microbiota, improve gut immune barrier function and equilibrate the balance between pro-inflammatory and anti-inflammatory cytokines. However, the positive effects of probiotics are not always substantiated by findings from properly conducted clinical trials. Notably, even when the results from randomised, placebo-controlled trials support the beneficial effects of a particular probiotic for a specific indication, the benefits are generally not translatable to other probiotic formulations.

The present study was undertaken to determine the prebiotic efficacy of gum arabic upon consumption by man for up to 4 weeks and, if any, to establish the dose–effect relationship. Human healthy volunteers consumed various daily doses (5, 10, 20, 40 g) of gum arabic (EmulGold®) in water for up to 4 weeks. Daily consumption of water was taken as the negative control and that of 10 g inulin as the positive control. At 0, 1, 2 and 4 weeks quantification of bacterial numbers in stool samples was performed via real time-PCR techniques and questionnaires were filled in to account for potential drawbacks. The genera of Bifidobacteria and Lactobacilli were taken as potentially beneficial bacteria and those of Bacteroides, Clostridium difficile and Enterococci as potentially non-beneficial, this distinction was dependent on the issue of these numbers being or becoming out of balance in the host. Compared with the negative control the numbers of Bifidobacteria and Lactobacilli 4 weeks after consumption were significantly higher for gum arabic: the optimal dose being around 10 g. Moreover, at this dose the numbers of Bifidobacteria, Lactobacilli and Bacteroides were significantly higher for gum arabic than for inulin. No significant drawback was encountered during the study. It is concluded that gum arabic establishes prebiotic efficacy, at least as good as inulin. The optimal daily dose was found to be 10 g.

This work was aimed at decreasing the glycaemic index (GI) of white wheat bread. Breads made with wheat flour (WF) or wholemeal flour (WMF) and fermented with baker's yeast had similar values of resistant starch (RS; 1·4–1·7 %, starch basis). Sourdough Lactobacillus plantarum P1 and Lactobacillus brevis P2 favoured the highest formation of RS (approximately 5 %) when fermented with WF and WMF. The mixture (1:1) of WF and WMF (WF/WMF) was selected. The effect of dietary fibres, chemical or sourdough acidification on the hydrolysis index (HI) of WF/WMF bread was determined. Among fibres, only the addition of oat fibre (5 %) decreased the HI to 90·84 %. Lactic acid determined the lowest HI, and the effect was related to the decrease of pH. For the same decrease of pH, breads fermented with L. plantarum P1 and L. brevis P2 (sourdough WF/WMF) showed values of HI lower than chemical acidification. The glucose response and GI of WF bread or sourdough WF/WMF bread enriched with oat fibre was determined by using fifteen healthy volunteers. Anhydrous glucose was used as reference. The area under the glucose response curve and the value of GI (72 %) of WF bread were significantly (P < 0·05) higher than sourdough WF/WMF bread enriched with oat fibre (GI = 53·7 %). The decrease of GI of the sourdough WF/WMF bread may be due to both fibre content and decreased pH. Compared to WMF bread, sourdough WF/WMF bread, enriched with oat fibre, had higher specific volume, better cell crumb structure and more appreciated acidulous smell, taste and aroma.

Milk fermented with mixed dahi cultures NCDC167, Lactococcus lactis ssp diacetylactis NCDC60 and two probiotic strains; Lactobacillus acidophilus NCDC14 and Lb. casei NCDC19 were evaluated after fermentation (14 h) and during 8 d storage at 7 °C. The β-galactosidase activity was found to increase after fermentation leading to the hydrolysis of lactose and production of glucose, galactose and oligosaccharides; that subsequently decreased during storage. The viable counts of lactococci and lactobacilli decreased during storage yet remained >106 cfu/ml after storage. The results of present study indicate that all the selected cultures have ability to produce oligosaccharides (prebiotics) due to transgalactosidal and lactose hydrolysis activities of β-galactosidase. The cultures developed an active synbiotic formula by maintaining sufficient probiotic viable counts to exert health benefits to the consumers.

Aminotransferase (AT) activity against 18 amino acids was studied in ten strains of three species of Lactobacillus. A method for permeabilisation of cells was developed using toluene and ethanol combined with mechanical treatment. It was found that the AT activities in the washed permeabilised cells (W-PC) corresponded well to that in cell-free extracts (CFE). The AT specificity pattern was species as well as strain dependant. Strains of Lb. helveticus had high specificity for aromatic amino acids (ArAA) and lower activity against branched-chain amino acids (BcAA) and Asp, while strains of Lb. paracasei subsp. paracasei degraded BcAA and Asp, but had a lower and variable specificity against ArAA. One of the Lb. paracasei strains was characterised by having very high AT activity against all three BcAA (Ile, Leu, Val) compared with any of the other Lb. paracasei strains tested. Strains of Lb. danicus, which is a newly discovered Lactobacillus species isolated from cheese, had up to about 20 times higher AT activity against Leu than Lb. paracasei and Lb. helveticus. The permeabilised cells of Lb. danicus had also considerably higher AT activity against ArAA than Lb. paracasei and Lb. helveticus strains, and also higher AT activity against Asp. All Lactobacillus strains tested had AT activity against Met, but at a much lower rate than against other amino acids. Results of this study also demonstrated a chemical reaction between α-ketoglutaric acid and Asp that was catalysed by pyridoxal-5-phosphate without any AT present.

The aim was to select potentially probiotic lactobacilli from 88 strains isolated from unpasteurized milk and cheese products, and to incorporate these bacteria in a viable state into a soft cheese, without changing its quality. The survival of these bacteria was assessed in acidic and bile conditions, after freezing at −80 °C. Four strains from unpasteurized Camembert – two Lactobacillus plantarum strains and two Lb. paracasei/casei strains – were identified and typed by PCR and PFGE and were found to display potentially probiotic characteristics in addition to resistance to low pH and bile. These characteristics were resistance to lysozyme, adhesion to CACO-2 cells, antimicrobial effects against common foodborne pathogens (Listeria monocytogenes, Staphylococcus aureus, Salmonella spp., Escherichia coli), innocuity following the ingestion of high doses by mice and appropriate antibiotic susceptibility profiles. The potential of Lb. plantarum strain UCMA 3037 for incorporation into a soft cheese (Pont-l'Eveque registered designation of origin (RDO)) was investigated. This strain grew well and survived in sufficient numbers (more than 107 cfu/g throughout the shelf-life of the product) in the cheese. This strain did not change the quality score of the product until the best before date (75 days after manufacture). Thus, unpasteurized Camembert is a natural source of potentially probiotic lactobacilli, which could be used as an additive in the development of potentially probiotic soft cheeses. Further work is required to demonstrate the persistence and efficacy of these strains in the human host upon ingestion.

In the present study we report for the first time the presence of S-layer proteins in Lactobacillus kefir and Lactobacillus parakefir isolated from kefir grains. Soluble whole-cell protein profile obtained either by mechanical disruption (X-press) or by a combined treatment with lysozyme and SDS on whole cells, showed a significant band of apparent molecular mass of 66–71 kDa as measured by SDS–PAGE. The intensity of this band was considerably reduced when cells were treated with 5 M-LiCl. The above mentioned proteins were recovered in the LiCl extracts. After dialysis and concentration, the proteins extracted were able to reassemble in a regular array. Negative staining of these protein preparations were analysed by transmission electron microscopy and a paracrystalline arrangement was seen. Thin sections of bacteria analysed by transmission electron micrographs showed an outermost layer over the bacterial cell wall, that was lost after the LiCl treatment. The production of this surface structure under different culture conditions was also evaluated. Finally, the relationship between the presence of S-layer proteins and surface properties (e.g. adhesion to Caco-2 cells, autoaggregation, and hemagglutination) was investigated.

Probiotic microorganisms, especially lactic acid bacteria, are effective in the treatment of infectious diarrhoeal diseases and experimental colitis. Although the mechanisms by which these organisms exert their anti-inflammatory effects are largely unknown, immunomodulating effects are suggested. The objective of this study was to examine the effect of a 5-week oral administration of Lactobacillus rhamnosus subspecies GG (Lb. GG) on the cellular immune response to intestinal microorganisms in ten healthy volunteers. Peripheral blood cells (PB) were stimulated with either ‘self’ or ‘non-self’ preparations of faecal samples and isolated Bacteroides fragilis group-organisms (Bfg) or Escherichia coli (Esch. coli), and pro- and anti-inflammatory cytokines (IL-10, IL-4, IL-6, IFN-γ, TNF-α) were measured in the culture supernatant. CD4+ T-lymphocyte activation was determined by measurement of intracellular ATP following lysis of the cells. The activational response of CD4+ T-lymphocytes towards isolated and heat-inactivated intestinal organisms was increased after the probiotic treatment. Additionally, TNF-α, IL-6 and in part IFN-γ cytokine secretion by PB cells following stimulation with whole stool preparations and single members of the flora was significantly decreased, whereas the IL-10 and in part IL-4 cytokine secretion was increased at the end of the study. In contrast, the activational response of CD4+ T-lymphocytes following stimulation with whole ‘non-self’ intestinal flora was higher than by ‘self’ intestinal flora, but both responses showed a trend towards a reduction at the end of the study. This study documents a direct effect by Lb. GG on the cellular immune system of healthy volunteers and offers a promising tool to investigate systemic immunomodulation due to oral administration of probiotic microorganisms.

Curiosity about probiotic bacteria is high. This is not surprising given the rapid growth of the natural products market in the USA, which was 22% in 1995 (New Hope Natural Media, Boulder, CO). Although probiotic cultures are only one component of that market, this trend shows that consumers in the USA are increasingly taking a pro-active stance towards their health and are purchasing products not only to eliminate what is perceived as dietary negatives but to increase the levels of dietary positives (Wrick, 1995). This shift in consumer attitude bodes well for the development of the American probiotic market. This article describes the composition, promotion and labelling of probiotic-containing products, discusses attributes of commercial probiotic bacteria, and estimates the size of the probiotic market in the USA. This market information was obtained from several published (Anon, 1996a,b)and unpublished but commercially available (SPINS Information Services, San Francisco, CA and New Hope Natural Media, Boulder, CO) sources.

The effect of feeding live Lactobacillus reuteri cells containing active bile salt hydrolase (BSH) on plasma cholesterol levels was studied in pigs. During an experiment lasting 13 weeks, twenty pigs were fed on a high-fat, high-cholesterol, low-fibre diet for the first 10 weeks, and a regular pig diet for the last 3 weeks. One group of animals received, twice daily, 11·25 (SD 0·16) log10 colony forming units of the potential probiotic bacteria for 4 weeks (from week 3 until week 7). From week 8 onwards, the treated group was again fed on the same diet as the control group without additions. The total faecal Lactobacillus counts were only significantly higher in the treated pigs during the first 2 weeks of L. reuteri feeding. Based on limited data, it was suggested that the administered Lactobacillus species had caused a temporary shift within the indigenous Lactobacillus population rather than permanently colonizing the intestinal tract. The probiotic feeding brought about significant lowering (P ≤ 0·05) of total and LDL-cholesterol concentrations in the treated pigs compared with the control pigs, while no change in HDL-cholesterol concentration was observed. The data for faecal output of neutral sterols and bile salts were highly variable between the animals of each group, yet they indicated an increased output in the treated pigs. Although the blood cholesterol levels went up in both groups during the 3 weeks following the Lactobacillus administration period, significantly lower serum total and LDL-cholesterol levels were observed in the treated pigs. During the final 3 weeks of normalization to the regular diet, cholesterol concentrations significantly decreased in both animal groups and the differences in total and LDL-cholesterol concentrations between the groups largely disappeared.