Impact of treatments on fecal microbiota and fecal metabolome in symptomatic uncomplicated diverticular disease of the colon: a pilot study.

Symptomatic uncomplicated diverticular disease (SUDD) affects 50% of people having diverticulosis. We performed a pilot study assessing the effect of current treatments on fecal microbiota and metabolome in SUDD. Thirteen consecutive females with SUDD were treated with a 2-week therapeutic trial of 30 g/day fiber supplementation (3 patients), 1.6 g/day of mesalazine (3 patients), 900 billion/day of probiotic mixture VivoMixx® (3 patients), or 800 mg/day of rifaximin (4 patients). Stool samples were collected at entry (T0), at the end of the 2-week therapeutic course (T1), and 30 (T2) and 60 days (T3) after the end of the therapeutic course. Real-time PCR quantified targeted microorganisms. Fecal metabolome patterns were studied by high-resolution proton NMR spectroscopy. At cumulative analysis, symptoms significantly decreased at each time point during follow-up (p less than 0.0001), and only left-lower quadrant pain increased again at T3. The overall bacterial quantity was not altered by the treatments. The amount of Akkermansia muciniphila species was significantly reduced at T1 (p=0.017) and at T2 (p=0.026), while at T3 the reduction was not significant in comparison to enrollment (p=0.090). Fecal molecular profile showed significant changes at T1 and T2, while at T3 it became similar to that of T0. Differences were found for 18 of the quantified molecules (tryptophan, phenylalanine, tyrosine, 4-hydroxyphenylacetate, urocanate, X-6.363, X-5.779, uridylate, galactose, X-4.197, threonine, sarcosine, methionine, 2-oxoisocaproate, 5-aminolevulinate, alanine, leucine, valerate). Metabolome and microbiota changed in patients with SUDD under treatment, confirming a possible role of dysbiosis/dysmetabolome in the pathology.

Biological control of vaginosis to improve reproductive health.

The human vaginal microbiota plays an important role in the maintenance of a woman's health, as well as of her partner's and newborns'. When this predominantly Lactobacillus community is disrupted, decreased in abundance and replaced by different anaerobes, bacterial vaginosis (BV) may occur. BV is associated with ascending infections and obstetrical complications, such as chorioamnionitis and preterm delivery, as well as with urinary tract infections and sexually transmitted infections. In BV the overgrowth of anaerobes produces noxious substances like polyamines and other compounds that trigger the release of pro-inflammatory cytokines interleukin (IL)-1 ß and IL-8. BV can profoundly affect, with different mechanisms, all the phases of a woman's life in relation to reproduction, before pregnancy, during fertilization, through and at the end of pregnancy. BV can directly affect fertility, since an ascending dissemination of the involved species may lead to tubal factor infertility. Moreover, the increased risk of acquiring sexually transmitted diseases contributes to damage to reproductive health. Exogenous strains of lactobacilli have been suggested as a means of re-establishing a normal healthy vaginal flora. Carefully selected probiotic strains can eliminate BV and also exert an antiviral effect, thus reducing viral load and preventing foetal and neonatal infection. The administration of beneficial microorganisms (probiotics) can aid recovery from infection and restore and maintain a healthy vaginal ecosystem, thus improving female health also in relation to reproductive health.

Vaginal microbiota and viral sexually transmitted diseases.

Healthy vaginal microbiota is an important biological barrier to pathogenic microorganisms. When this predominantly Lactobacillus community is disrupted, decreased in abundance and replaced by different anaerobes, bacterial vaginosis (BV) may occur. BV is associated with prevalence and incidence of several sexually transmitted infections. This review provides background on BV, discusses the epidemiologic data to support a role of altered vaginal microbiota for acquisition of sexually transmitted diseases and analyzes mechanisms by which lactobacilli could counteract sexually transmitted viral infections.

Effectiveness of vaginal tablets containing lactobacilli versus pH tablets on vaginal health and inflammatory cytokines: a randomized, double-blind study.

The purpose of this study was to evaluate the effectiveness of lactobacilli on vaginal health and proinflammatory cytokines. Sixty-seven patients with bacterial vaginosis (BV), 50 with intermediate flora and 42 with normal vaginal flora were enrolled in this double-blind study. The subjects were randomized to receive probiotic lactobacilli vaginal tablets (L. brevis CD2, L. salivarius subsp. salicinius, L. plantarum) or the vaginal pH tablet (active comparator). Cervico-vaginal lavage was collected to measure the concentrations of IL-1ß, TNFα and IL-6 by ELISA. Neutral sphingomyelinase activity was also quantified in both arms before and after treatment. The probiotic vaginal tablet was well tolerated and no side effects were reported. The study demonstrated a cure rate of nearly 80 %; i.e., 32 % of the women could restore normal vaginal flora and 47 % had improved Nugent score, whereas 20 % of the subjects did not clear BV in the first follow-up (after 8 days treatment). The pH tablet containing pH lowering compounds induced resolution of BV and restoration of normal vaginal flora in 74 % and 26 %, respectively. The lactobacilli tablet was found to be better than the pH tablet in preventing BV in healthy subjects. A significant reduction in IL-1ß and IL-6 vaginal cytokines was observed after treatment with lactobacilli, while the active comparator did not have any effect on local proinflammatory cytokines. Vaginal neutral sphingomyelinase activity was not modified in either group. Vaginal tablets containing lactobacilli can cure BV and reduce vaginal inflammatory response.

Effectiveness of Lactobacillus-containing vaginal tablets in the treatment of symptomatic bacterial vaginosis.

The purpose of this study was to determine the effectiveness of Lactobacillus-containing vaginal tablets in the treatment of bacterial vaginosis (BV) and in the restoration of a healthy vaginal flora. Thirty-nine women with BV were enrolled in a double-blind, placebo-controlled clinical trial. Patients received either one Lactobacillus-containing tablet or placebo daily for 7 days. Clinical criteria, vaginal Gram stain scores and symptoms were compared with those at the initial visit and those at completion of therapy and 2 weeks later. After completion of therapy, all of the patients in the Lactobacillus-treated group (n = 18) were free of BV, showing a normal (83%) or intermediate (17%) vaginal flora, as compared with only two patients free of BV with intermediate flora (12%) from among the 16 placebo-treated women (p <0.001). Two weeks after completion of therapy, treatment was successful (score <7) in 61% of Lactobacillus-treated patients as compared with 19% of those in the placebo group (p <0.05). In the treatment group, the total number of symptomatic patients and the intensity of their symptoms, in particular vaginal malodour, were significantly reduced at both follow-up visits. The data indicate that intravaginal administration of exogenous selected strains of lactobacilli can restore a normal vaginal microbiota and be used in treating bacterial vaginosis.

Inhibition of herpes simplex virus type 2 by vaginal lactobacilli.

The protective effect of selected vaginal Lactobacillus strains (L. brevis CD2, L. salivarius FV2, L. plantarum FV9) towards herpes simplex virus type 2 (HSV-2) infection in vitro has been analyzed. Living bacterial cells affect different steps of virus multiplication. The effect on the early phases of virus infection appeared related to the bacterial adhesive potential to the cell membrane while all the strains strongly reduced intracellular events of virus multiplication. The anti HSV-2 activity was not mediated by a virucidal effect. Instead it was exerted through bacterial soluble factors able to down regulate the production of infective virions. In fact HSV-2 yield was significantly reduced in infected cells fed with cell-free supernatants of lactobacilli grown in cell culture medium. Purified lactic acid and H(2)O(2), Lactobacillus metabolites with known antimicrobial activity, produced a dose-dependent virucidal effect. Lactic acid successfully interfered with viral intracellular antigen synthesis and both the virucidal activity and the inhibition of replication were correlated to acidic pH values. L. brevis CD2, the most active strain, does not produce H(2)O(2) and neutralized lactic acid had no effect, thus indicating that factors other than H(2)O(2) and lactic acid could be responsible for the antiviral effect.

Precision measurement of the weak mixing angle in Møller scattering.

We report on a precision measurement of the parity-violating asymmetry in fixed target electron-electron (Møller) scattering: A(PV) = [-131 +/- 14(stat) +/- 10(syst)] x 10(-9), leading to the determination of the weak mixing angle sin2(thetaW(eff) = 0.2397 +/- 0.0010(stat) +/- 0.0008(syst), evaluated at Q2 = 0.026 GeV2. Combining this result with the measurements of sin2(thetaW(eff) at the Z0 pole, the running of the weak mixing angle is observed with over 6sigma significance. The measurement sets constraints on new physics effects at the TeV scale.

Does hospital work constitute a risk factor for Helicobacter pylori infection?

The aim of this study was to assess whether clinical work constitutes a risk factor for Helicobacter pylori infection among employees in hospitals. The prevalence of H. pylori infection was analysed in 249 individuals employed in a university teaching hospital according to three categories of hospital workers: (A) personnel from gastrointestinal endoscopy units (N=92); (B) personnel from other hospital units with direct patient contact (N=105); and (C) staff from laboratories and other units with no direct patient contact (N=52). Stool samples from each subject were examined with a validated enzyme-linked immunosorbent assay for the presence of H. pylori antigens. A questionnaire inquiring about sociodemographic and occupational characteristics was completed by each participant. The prevalence of H. pylori infection was 37.0% in group A, 35.2% in group B and 19.2% in group C (P<0.05). Among the different healthcare categories, nurses had a significant higher prevalence of H. pylori infection (P<0.01). No significant association was found between the length of employment or exposure to oral and faecal secretions, and H. pylori infection. Hospital work involving direct patient contact seems to constitute a major risk factor for H. pylori infection compared with hospital work not involving direct patient contact.

Observation of parity nonconservation in møller scattering.

We report a measurement of the parity-violating asymmetry in fixed target electron-electron (Møller) scattering: A(PV)=[-175+/-30(stat)+/-20(syst)] x 10(-9). This first direct observation of parity nonconservation in Møller scattering leads to a measurement of the electron's weak charge at low energy Q(e)(W)=-0.053+/-0.011. This is consistent with the standard model expectation at the current level of precision: sin((2)theta(W)(M(Z))((-)MS)=0.2293+/-0.0024(stat)+/-0.0016(syst)+/-0.0006(theory).

Characterization and selection of vaginal Lactobacillus strains for the preparation of vaginal tablets.

AIMS: To characterize and select Lactobacillus strains for properties that would make them a good alternative to the use of antibiotics to treat human vaginal infections. METHODS AND RESULTS: Ten Lactobacillus strains belonging to four different Lactobacillus species were analysed for properties relating to mucosal colonization or microbial antagonism (adhesion to human epithelial cells, hydrogen peroxide production, antimicrobial activity towards Gardnerella vaginalis and Candida albicans and coaggregation with pathogens). The involvement of electrostatic interactions and the influence of bacterial metabolic state in the binding of lactobacilli to the cell surface were also studied. Adherence to epithelial cells varied greatly among the Lactobacillus species and among different strains belonging to the same Lactobacillus species. The reduction in surface negative electric charge promoted the binding of several Lactobacillus strains to the cell membrane whereas lyophilization reduced the adhesion capacity of many isolates. The antimicrobial activity of lactobacilli culture supernatant fluids was not directly related to the production of H2O2. CONCLUSIONS: Three strains (Lactobacillus brevis CD2, Lact. salivarius FV2 and Lact. gasseri MB335) showed optimal properties and were, therefore, selected for the preparation of vaginal tablets. The selected strains adhered to epithelial cells displacing vaginal pathogens; they produced high levels of H2O2, coaggregated with pathogens and inhibited the growth of G. vaginalis. SIGNIFICANCE AND IMPACT OF THE STUDY: The dosage formulation developed in this study appears to be a good candidate for the probiotic prophylaxis and treatment of human vaginal infections.

Technological and biological evaluation of tablets containing different strains of lactobacilli for vaginal administration.

Ten strains of lactobacilli were evaluated for the administration of viable microorganisms to restore the normal indigenous flora in the treatment of urogenital tract infections (UTI) in women. As the strains considered are facultative anaerobes, optimization of the production process was particularly critical to preserve bacterial viability. The microorganisms were formulated in single- and double-layer vaginal tablets. The two layers were characterized by different release properties: one is an effervescent composition that ensures a rapid and complete distribution of the active ingredient over the whole vaginal surface; while the second is a sustained release composition capable of releasing the lactobacilli over a longer period of time. Three different retarding polymers were tested, and all the formulations and tablets were evaluated in terms of technological processability, bacterial viability and stability, and cell adhesion properties of the microorganisms. From the results obtained, three out of ten strains appear particularly suitable for their application in the treatment of UTI. A larger batch of tablets made with a mixture of the three strains was then evaluated, confirming the feasibility of their industrial production and a good bacterial viability in the final dosage form.

Synthesis and anti-rhinovirus activity of 2-styrylchromones.

2-Styrylchromones were synthesized as vinylogues of 2-phenylchromones (flavones), a broad class of anti-rhinovirus compounds. The antiviral activity of 2-styrylchromones and 3-hydroxy-1-(2-hydroxyphenyl)-5-phenyl-2,4-pentadien-1-ones, which are intermediates in the synthesis, was evaluated against two selected serotypes of human rhinovirus, 1B and 14, by a plaque reduction assay in HeLa cell cultures. All of the compounds interfered with HRV 1B replication, with the exception of 3-hydroxy-1-(2-hydroxyphenyl)-5-(4-methoxyphenyl)-2,4-pentadien-1-one. The majority of derivatives were also found to be effective against serotype 14, often with a higher potency.

Synthesis and biological evaluation of 2-methoxy- and 2-methylthio-6-[2'-alkylamino)ethyl]-4(3H)-pyrimidinones with anti-rubella virus activity.

The synthesis of a new family of antiviral compounds, 2-methoxy-, and 2-methylthio-6-[(2'-alkylamino)ethyl]-4(3H)-pyrimidinones, has been accomplished. The activity of these agents against positive strand (rubella virus and sindbis virus) and negative strand (vesicular stomatitis virus) RNA viruses is reported. Some of these compounds are efficient and selective inhibitors of rubella virus.

Antiviral effect of hyperthermic treatment in rhinovirus infection.

Human rhinoviruses (HRV) are recognized as the major etiologic agents for the common cold. Starting from the observation that local hyperthermic treatment is beneficial in patients with natural and experimental common colds, we have studied the effect of brief hyperthermic treatment (HT) on HRV replication in HeLa cells. We report that a 20-min HT at 45 degrees C is effective in suppressing HRV multiplication by more than 90% when applied at specific stages of the virus replication cycle. Synthesis of virus proteins is not affected by HT, indicating that the target for treatment is a posttranslational event. The antiviral effect is a transient cell-mediated event and is associated with the synthesis of the 70-kDa heat shock protein hsp70. Unlike poliovirus, rhinovirus infection does not inhibit the expression of hsp70 induced by heat. The possibility that hsp70 could play a role in the control of rhinovirus replication is suggested by the fact that a different class of HSP inducers, the cyclopentenone prostaglandins PGA1 and delta 12-PGJ2, were also effective in inhibiting HRV replication in HeLa cells. Inhibition of hsp70 expression by actinomycin D prevented the antiviral activity of prostaglandins in HRV-infected cells. These results indicate that the beneficial effect of respiratory hyperthermia may be mediated by the induction of a cytoprotective heat shock response in rhinovirus-infected cells.

Anti-picornavirus activity of synthetic flavon-3-yl esters.

The in vitro antiviral activity against picornaviruses (rhinovirus serotype 1B and 14, and poliovirus type 2) of new synthetic 3-hydroxyflavones, 3-acetoxyflavones, and substituted cinnamic and benzoic acid flavon-3-yl esters was evaluated. The maximum non-toxic concentration of compounds was determined in a human cell line (HeLa) suitable for the replication of the three viruses. Their antiviral potency was measured by a plaque reduction assay. Generally, rhinoviruses exhibited a higher sensitivity to the new flavonoids than poliovirus. Flavones, with sterically small substituents in position 3, showed good activity against both rhinoviruses tested. However, the introduction of bulky substituents in the same position resulted in analogues with a higher toxicity and often with a lower efficacy.

Antiviral activity of natural and semisynthetic polysaccharides on the early steps of rubella virus infection.

The natural and semisynthetic carbohydrates scleroglucan, locust bean gum, tamarind gum (glyloid) and its three sulphate derivatives (GP4311, GP4327 and GP4324), glycogen and its two sulphate derivatives (GP4427 and GP4435), alginic acid and dextran sulphate, were investigated for their inhibitory effect on rubella virus (RV) infection of Vero cells. The neutral polymer scleroglucan and two highly negatively charged compounds, glyloid sulphate 4324 and dextran sulphate, had the highest inhibitory effect on RV antigen synthesis. The antiviral properties of active molecules appears to be dependent on the shape of the macromolecule and/or on the electric charge, while saccharide units play a minor role. The results indicated that polysaccharides blocked a step in virus replication subsequent to virus attachment, such as internalization and/or uncoating. Confirmation that the inhibitory activity of the compounds was directed at the early steps of RV multiplication, was that none of the polysaccharides had any effect on infection initiated by transfection of cells with RVRNA.

Pathway of rubella virus infectious entry into Vero cells.

The mechanism and the kinetics of rubella virus (RV) penetration into Vero cells were studied. By using pronase or acid treatment to inactivate virus which had adsorbed to cell membrane but had not been internalized, it was found that a period of 7 h was required in order for all of the adsorbed virus to enter the host cells. Lysosomotropic agents (monensin, methylamine, ammonium chloride and chloroquine) were used to study the mechanism by which RV penetrates host cells. Virus replication was inhibited if treatment of cells with these compounds was performed for at least 9 h after infection. However, if extracellular adsorbed virions were eliminated by acid treatment following removal of the lysosomotropic compounds, RV replication was completely inhibited by treatment with these drugs for any time period after adsorption. This indicated that the prolonged period of treatment with these compounds necessary to inhibit virus replication is due to the slow rate of RV internalization. None of the compounds had any effect on infection initiated by transfection of RV RNA, confirming that these drugs were exerting their inhibitory activity at penetration. The inhibition of RV replication by lysosomotropic compounds indicates that RV penetrates host cells by the endosomal pathway.

Inhibition of poliovirus replication by prostaglandins A and J in human cells.

Cyclopentenone prostaglandins (PGs) inhibit the replication of a wide variety of enveloped DNA and RNA viruses. The antiviral activity is associated with alterations in the synthesis, maturation, and intracellular translocation of viral proteins. In the present report, we describe the effects of cyclopentenone PGs PGA1 and delta 12-PGJ2 on poliovirus (PV) replication in HeLa cells. Both PGs were found to inhibit PV replication dose dependently. Virus yield was significantly reduced at nontoxic concentrations, which did not suppress RNA or protein synthesis in uninfected or PV-infected cells. Both the pattern of PV proteins synthesized and the kinetics of viral protein synthesis and degradation appeared to be similar in PGA1-treated cells and control cells. Antiviral PGs have been shown to selectively inhibit virus protein synthesis during the replication of several viruses, including vesicular stomatitis virus (VSV), and this effect has been recently associated with the induction of a 70-kDa heat shock protein (HSP70). PGA1 and delta 12-PGJ2 were found to induce HSP70 synthesis in uninfected or VSV-infected HeLa cells. PV infection was found to inhibit PG-induced HSP70 synthesis in these cells, suggesting that the lack of ability of cyclopentenone PGs to block PV protein synthesis could be related to an impaired heat shock response in PV-infected cells. The finding that PV protein synthesis was not inhibited by PGs suggests that cyclopentenone PGs could interfere with a late event in the virus replication cycle, such as protein assembly and maturation of PV virions.