Bacillus subtilis (DSM17299) significantly reduces Salmonella in broilers.

Salmonella continues to be a major public health burden worldwide. Poultry are known to be one of the main reservoirs for this zoonotic pathogen. It has previously been shown that a single dose of Bacillus subtilis reduces fecal shedding of Salmonella enterica serovar Enteritidis, whereas no effect on long-term colonization of the cecum has been observed. Here we report experiments that were undertaken to test the efficacy of a conventional diet supplemented with a probiotic (B. subtilis DSM17299) on 1) Salmonella colonization in the intestinal tract of broiler chickens, and 2) fecal shedding of Salmonella under production-like conditions. The trial birds fed the B. subtilis diet showed a significant 58% reduction in Salmonella-positive drag swabs compared with control birds, which had 100% presence of Salmonella. Feeding B. subtilis significantly reduced the average Salmonella load of cecum samples of the chickens, by 3 log units. This reduction in Salmonella colonization might not only positively affect broilers on the live production side by reducing the risk of infection between birds, but could also aid on the processing side by decreasing the amount of Salmonella entering the facility and improving food safety. Furthermore, numerical, but not statistically significant, improvements in feed conversion rate and BW gain at d 42 were observed in the B. subtilis-treated group compared with control birds.

Characterization of Streptococcus thermophilus lytic bacteriophages from mozzarella cheese plants.

Phage infection still represents the main cause of fermentation failure during the mozzarella cheese manufacturing, where Streptococcus thermophilus is widely employed as starter culture. Thereby, the success of commercial lactic starter cultures is closely related to the use of strains with low susceptibility to phage attack. The characterization of lytic S. thermophilus bacteriophages is an important step for the selection and use of starter cultures. The aim of this study was to characterize 26 bacteriophages isolated from mozzarella cheese plants in terms of their host range, DNA restriction profile, DNA packaging mechanism, and the variable region VR2 of the antireceptor gene. The DNA restriction analysis was carried out by using the restriction enzymes EcoRV, PstI, and HindIII. The bacteriophages were distinguished into two main groups of S. thermophilus phages (cos- and pac-type) using a multiplex PCR method based on the amplification of conserved regions in the genes coding for the major structural proteins. All the phages belonged to the cos-type group except one, phage 1042, which gave a PCR fragment distinctive of pac-type group. Furthermore, DNA sequencing of the variable region VR2 of the antireceptor gene allowed to classify the phages and examine the correlation between typing profile and host range. Finally, bacterial strains used in this study were investigated for the presence of temperate phages by induction with mitomycin C and only S. thermophilus CHCC2070 was shown to be lysogenic.

Dose-response study of probiotic bacteria Bifidobacterium animalis subsp lactis BB-12 and Lactobacillus paracasei subsp paracasei CRL-341 in healthy young adults.

OBJECTIVE: This study was performed to investigate the dose-response effects of supplementation with Bifidobacterium animalis subsp lactis (BB-12) and Lactobacillus paracasei subsp paracasei (CRL-431) on blood lipids, recovery from feces and bowel habits. Changes of the fecal microflora was analyzed in the 10(10) CFU/day probiotic and placebo group. DESIGN: The study was designed as a randomized, placebo-controlled, double-blinded, parallel dose-response study. SUBJECTS: Healthy young adults (18-40 years) were recruited by advertising in local newspapers. Of the 75 persons enrolled, 71 (46 women, 25 men, mean age 25.6 years (range 18-40 years)) completed the study. INTERVENTION: The volunteers were randomly assigned into five groups receiving either placebo or a mixture of the two probiotics in the concentration of 10(8), 10(9), 10(10) or 10(11) CFU/day in 2 weeks run-in period, 3 weeks intervention and 2 weeks wash-out. Diary reporting bowel habits and well being (abdominal bloating, flatulence and headache) was kept for all 7 weeks and blood lipids, fecal recovery of BB-12 and CRL-431, as well as fecal microflora was tested before, immediately and 2 weeks after intervention. RESULTS: The fecal recovery of BB-12 increased significantly (P < 0.001) with increasing dose. In the group receiving 10(11) CFU/day BB-12 was recovered from 13 out of 15 volunteers. CRL-431 was not recovered in any of the fecal samples. Supplementation with probiotics did not change the fecal bacterial composition. A significant linear increase in fecal consistency (looser stool) with increasing probiotic dose (P = 0.018) was observed. No overall dose-response effect was found on the blood lipids. High doses of probiotics were well tolerated. CONCLUSION: A dose-related recovery of BB-12 from feces was observed.

Prevalence of Helicobacter pylori vacuolating cytotoxin and its allelic mosaicism as a predictive marker for Iranian dyspeptic patients.

Helicobacter pylori infects the majority of the population in the developing countries. However, the rate of gastrointestinal complications such as peptic ulcers and gastric malignancies has no parallel with the infection. In order to determine whether cytotoxin (vacA) and its allelic polymorphism can serve as screening markers for such a population, H. pylori strains were isolated from one hundred and thirty two dyspeptic patients. H. pylori genomic DNA was extracted and underwent PCR-amplification for the cytotoxin alleles. Genotyping of the signal sequence region of the vacA gene identified 68% (70 out of 103) of patients with non ulcer dyspepsia (NUD) and 79% (23 out of 29) of the patients with peptic ulcer disease (PUD) possessing the s1 genotype. S1 strains were significantly more prevalent among patients with PUD as compared to the NUD (p < 0.05). In regard to the middle region, 55% of the patient isolates belonged to the m2 genotype with no correlation to disease. The s1m2 genotype was the most prevalent among all patients and significantly correlated with the PUD group (p < 0.05).

Gastric mucosal cytokine responses in Helicobacter pylori-infected patients with gastritis and peptic ulcers. Association with inflammatory parameters and bacteria load.

Helicobacter pylori is an important pathogen in gastroduodenal inflammation and ulceration. Several mechanisms have been proposed to explain its role. We studied the cytokine production patterns in situ in gastric mucosal biopsies from H. pylori-positive and H. pylori-negative patients with dyspepsia. Immunohistochemistry with monoclonal antibodies was used. The study showed enhanced expression of interleukin (IL) -8, IL-10 and interferon-gamma (IFN-gamma) in H. pylori infection and a significant association was found between these cytokines and the following parameters: bacteria load, chronic inflammation and activity. These parameters were significantly correlated with the cell markers CD19 and CD56. The study indicates a dual effect of H. pylori on the Th1 response, i.e. a stimulation of the response verified by increased IFN-gamma and a feed-back verified by an increase of the counterinflammatory IL-10, which may dampen the inflammatory and cytotoxic effect of the Th1 response. Furthermore, the study confirms the connection between increase of IL-8 and inflammatory activity in gastric mucosa in H. pylori infection.

[Prevalence of Chlamydia among young men. A screening among men liable for military service and coming before the military board]. / Forekomsten af klamydia blandt unge maend. En screeningsundersøgelse blandt maend fremmødt på session.

INTRODUCTION: Chlamydia is the most common sexually transmitted bacterial infection in Denmark. Each year 13-14,000 cases are diagnosed; of these about 3000 occur in men. In order to describe the prevalence of chlamydia among young men, screening for chlamydia was conducted in those liable for military service and coming before the medical board. MATERIALS AND METHODS: Of 2500 men coming before the medical boards in North Jutland State County, Arhus State County, and Copenhagen County during the winter of 1996-97, 1345 men aged 17-32 (median 18 years) were entered in the study. The participants sent a urine sample and a filled-in questionnaire to Statens Serum Institut. The urine samples were analysed by an in-house PCR and the test results were sent to the participant; if permission had been granted, the positive test results were further sent to the general practitioner, along with a second questionnaire. RESULTS: Chlamydia was found in 4.8% (65/1345) of all participants and in 6.9% of those sexually active. In North Jutland State County 7.1 and 9.8%, respectively, had chlamydia, whereas the corresponding figures for Arhus State County and Copenhagen County were about 4% and 6%. Two-thirds of the patients had no symptoms of urethritis. DISCUSSION: The results of this study indicate that a considerable reservoir of unrecognised chlamydia must exist in young men. The prevalence of chlamydia was higher in North Jutland State County than in the other two districts. One explanation could be that there is more focus on sexually transmitted infections and contact tracing in the largest cities than in the rest of the country.

Purification and enzymatic activity of an NADH-fumarate reductase and other mitochondrial activities of Leishmania parasites.

A 65 kD membrane-associated NADH-fumarate reductase subunit, which has a molecular weight similar to that of one of the enzyme subunits from bacteria, was purified from Leishmania donovani promastigotes. NADH-fumarate reductase and other mitochondrial enzymatic activities of L. major and L. donovani promastigotes and amastigotes were investigated. The presence of NADH-fumarate reductase was demonstrated in digitonin-permeabilized L. major promastigotes and mitochondria of L. major and L. donovani promastigotes and amastigotes. The activity of solubilized NADH-fumarate reductase was measured in L. major and L. donovani promastigotes. Succinate exhibited a clear concentration-dependent inhibitory effect on fumarate reductase, whereas fumarate also exhibited a clear concentration-dependent inhibitory effect on succinate dehydrogenase. The data indicate that fumarate reductase is an obligatory component of the respiratory chain of the parasite. Since the enzyme is an important component in the intermediate metabolism in the Leishmania parasite and is absent in mammalian cells, it could be a potential target for antileishmanial drugs.

Helicobacter heilmannii-associated primary gastric low-grade MALT lymphoma: complete remission after curing the infection.

BACKGROUND & AIMS: Cure of Helicobacter pylori infection may lead to complete remission of associated low-grade mucosa-associated lymphoid tissue (MALT) lymphoma in stage EI. This study investigated whether Helicobacter heilmannii infection-associated primary gastric MALT lymphoma will regress after cure of the infection. METHODS: H. heilmannii-induced gastritis was diagnosed histologically, by a new specific immunoglobulin G enzyme-linked immunosorbent assay, and with 16S ribosomal RNA amplification and sequencing in 5 consecutive patients with primary gastric MALT lymphoma clinical stage EI. Patients received 40 mg omeprazole and 750 mg amoxicillin 3 times per day for 14 days. Polymerase chain reaction (PCR) was used to detect rearrangement of immunoglobulin heavy-chain genes before treatment and during follow-up. RESULTS: Five patients (3 men, 2 women; mean age, 65 years; range, 42-79 years) were studied. H. pylori was not detected by culture, histology, serology, or PCR. Treatment resulted in the cure of H. heilmannii infection in each case and complete histological and endoscopic remission of the tumors. Three of 5 patients showed monoclonal B cells before treatment, 2 of whom remained PCR positive. Within a median follow-up period of 24 months, no relapse of the lymphoma or reinfection with H. heilmannii occurred. CONCLUSIONS: These data suggest that gastric MALT lymphoma may arise in patients with H. heilmannii infection. Cure of this infection may lead to complete remission of the MALT lymphoma.

[Pseudolymphoma and ventricular maltoma in patients with chronic gastritis, ulcer and Helicobacter pylori infection]. / Pseudolymfom og ventrikelmaltom hos patienter med kronisk gastritis, ulcus og Helicobacter pylori-infektion.

Among 128 patients with malignant B-lymphoproliferative disorders, 19 patients had long lasting dyspepsia and gastroscopy showed chronic active gastritis or gastric ulcer. PCR analysis for TCR and IgH clonality in biopsies showed local involvement of the malignant lymphocyte clone in four patients out of eight indicating presence of these cells in the inflammatory infiltrate. Weak B-cell clonality was found in four patients. A close relationship was seen between lymphocytic clonality and immune response to H. pylori Cag A, and all patients had parietal cell antibodies. Thus, the malignant clone may participate in the local inflammatory reaction, and continued local stimulation by H. pylori as well as parietal cell antigens may lead both to autoimmunity as well as a clonal development of lymphocytes.

Treatment of H. pylori infected mice with antioxidant astaxanthin reduces gastric inflammation, bacterial load and modulates cytokine release by splenocytes.

Helicobacter pylori is a gram-negative bacterium affecting about half of the world population, causing chronic gastritis type B dominated by activated phagocytes. In some patients the disease evolves into gastric ulcer, duodenal ulcer, gastric cancer or MALT lymphoma. The pathogenesis is in part caused by the immunological response. In mouse models and in human disease, the mucosal immune response is characterized by activated phagocytes. Mucosal T-lymphocytes are producing IFN-gamma thus increasing mucosal inflammation and mucosal damage. A low dietary intake of antioxidants such as carotenoids and vitamin C may be an important factor for acquisition of H. pylori by humans. Dietary antioxidants may also affect both acquisition of the infection and the bacterial load of H. pylori infected mice. Antioxidants, including carotenoids, have anti-inflammatory effects. The aim of the present study was to investigate whether dietary antoxidant induced modulation of H. pylori in mice affected the cytokines produced by H. pylori specific T-cells. We found that treatment of H. pylori infected mice with an algal cell extract containing the antioxidant astaxanthin reduces bacterial load and gastric inflammation. These changes are associated with a shift of the T-lymphocyte response from a predominant Th1-response dominated by IFN-gamma to a Th1/Th2-response with IFN-gamma and IL-4. To our knowledge, a switch from a Th1-response to a mixed Th1/Th2-response during an ongoing infection has not been reported previously.

Production and isolation of guinea pig IgE antibody.

Immunoglobulins of the IgE and IgG classes have been causally associated with hypersensitivity reactions in man and in numerous animal species including mice, rats and guinea pigs. The use of the guinea pig as an animal model for both pulmonary and dermal hypersensitivity reactions, and the recent recognition of the importance of IgE antibodies in both early- and late-onset hypersensitivity responses, has heightened interest in production, separation, and isolation of this immunoglobulin class from the guinea pig. IgE antibodies were produced by treatment of strain 13 guinea pigs with cyclophosphamide followed by injection with S. aureus enterotoxin. Serum was obtained and the globulin fraction isolated by addition of caprylic acid then ammonium sulfate. Immunoglobulins were separated into classes using fast protein liquid chromatography (FPLC) employing a Mono Q column and a linear gradient of 0.01-0.3 M Na,K phosphate buffer, pH 7.5 (buffer B). IgG eluted in two major peaks. IgG2 was not retained on the column and emerged with the starting buffer; IgG1 was eluted with 15-20% buffer B. IgE, detected as heat labile homocytotropic antibody, was found in the fraction eluting with 30-35% buffer B. The elution profile of the guinea pig immunoglobulins was predicted from the pattern obtained with immunoglobulin classes from other species. This chromatographic procedure enabled rapid isolation of immunoglobulin classes from guinea pig sera and effectively separated IgG1 from IgE, the two classes associated with hypersensitivity reactions.