Safety and Immunomodulatory Effects of Three Probiotic Strains Isolated from the Feces of Breast-Fed Infants in Healthy Adults: SETOPROB Study

We previously described the isolation and characterization of three probiotic strains from the feces of exclusively breast-fed newborn infants: Lactobacillus paracasei CNCM I-4034, Bifidobacterium breve CNCM I-4035 and Lactobacillus rhamnosus CNCM I-4036. These strains were shown to adhere to intestinal mucus in vitro, to be sensitive to antibiotics and to resist biliary salts and low pH. In the present study, a multicenter, randomized, double-blind, placebo-controlled trial with 100 healthy volunteers in three Spanish cities was carried out to evaluate the tolerance, safety, gut colonization and immunomodulatory effects of these three probiotics. Volunteers underwent a 15-day washout period, after which they were randomly divided into 5 groups that received daily a placebo, a capsule containing one of the 3 strains or a capsule containing a mixture of two strains for 30 days. The intervention was followed by another 15-day washout period. Patients did not consume fermented milk for the entire duration of the study. Gastrointestinal symptoms, defecation frequency and stool consistency were not altered by probiotic intake. No relevant changes in blood and serum, as well as no adverse events occurred during or after treatment. Probiotic administration slightly modified bacterial populations in the volunteers’ feces. Intestinal persistence occurred in volunteers who received L. rhamnosus CNCM I-4036. Administration of B. breve CNCM I-4035 resulted in a significant increase in fecal secretory IgA content. IL-4 and IL-10 increased, whereas IL-12 decreased in the serum of volunteers treated with any of the three strains. These results demonstrate that the consumption of these three bacterial strains was safe and exerted varying degrees of immunomodulatory effects.

Trial Registration

ClinicalTrials.gov NCT01479543     

Genetic diversity assessment of Tunisian <Emphasis Type="Italic">Mycobacterium bovis</Emphasis> population isolated from cattle

Background

The genetic diversity of M. bovis in Tunisia is still underestimated despite the implementation of an eradication program. The lack of data about spatial distribution of the M. bovis population hinders the control of bovine tuberculosis (bTB) progress. This study represents the largest molecular analysis of M. bovis isolates in Tunisia. It is aimed to upgrade the understanding of bTB epidemiology and the geographical distribution of the infection. Tuberculosis research was performed in cattle (n?=?149) with TB-compatible lesions collected over 5 months from a slaughterhouse located in Sfax, Tunisia.

Results

Ninety-four animals were found to be infected by M. bovis and two others by M. caprae. Spoligotyping revealed twenty-five patterns, SB0120, SB0134, and SB0121 being the most prevalent profiles (36.4%, 11.4%, and 7.2%, respectively). Three new spoligotypes were detected: SB2345, SB2344 and SB2343. MIRU-VNTR analysis classified the isolates in seventy-three profiles and showed a large genotypic variety observed within the main spoligotype which was split into several MIRU-VNTR types: 29 in SB0120 (h?=?0.983), 10 in SB0134 (h?=?0.981) and 7 in SB0121 (h?=?1). Genotyping revealed a common pattern in different geographic regions. It also showed that Sfax, located in southern-Tunisia, represents a high-risk area with an elevated genetic diversity.

Conclusions

Spatial analysis may provide insights into disease transmission, which affects the effectiveness of eradication campaigns in cattle.

     

Sex‐specific density‐dependent secretion of glucocorticoids in lizards: insights from laboratory and field experiments

Negative density feedbacks have been extensively described in animal species and involve both consumptive (i.e. trophic interactions) and non‐consumptive (i.e. social interactions) mechanisms. Glucocorticoids are a major component of the physiological stress response and homeostasis, and therefore make a good candidate for proximate determinants of negative density feedbacks. Here, we combined laboratory and field experiments with enclosed populations to investigate the relationship between density, social stress and plasma corticosterone levels in the common lizard Zootoca vivipara. This species exhibits strong negative density feedbacks that affect females more than males, and its life history is sensitive to experimentally‐induced chronic elevation of corticosterone plasma levels. We found that prolonged crowding in the laboratory can trigger a chronic secretion of corticosterone independent from food restriction. In the field experiments, corticosterone levels of females were not affected by population density. Corticosterone levels of males increased with population density but only during the late activity season in a first field experiment where we manipulated density. They also increased with density during the mating season but only in populations with a female‐biased sex ratio in a second field experiment where we crossed manipulated density and adult sex ratio. Altogether, our results provide limited evidence for a role of basal corticosterone secretion in density feedbacks in this species. Context and density‐dependent effects in males may arise from changes in behavior caused by competition for resources, male–male competition, and mating.     

Histoplasmosis in a Brazilian center: clinical forms and laboratory tests

Histoplasmosis, caused by the dimorphic fungus Histoplasma capsulatum, is endemic in many regions of the Americas, Asia and Africa. It has a wide spectrum of clinical manifestations, from asymptomatic infection to severe disseminated disease. A retrospective study was carried out to describe the clinical forms and assess the clinical significance of the laboratory diagnostic tests of patients with histoplasmosis during the period of July 1987 to December 2003 at Instituto de Pesquisa Clínica Evandro Chagas/ FIOCRUZ, RJ, Brazil. Seventy-four patients were included. Forty-nine percent of the cases (n = 36) occurred in HIV positive patients who presented with disseminated disease. The remaining 38 cases were classified in different clinical forms. Histoplasma capsulatum was isolated from 69.5% of the clinical specimens sent to culture. Immunodiffusion and immunoblot were positive in 72.6% and 100% of the performed tests, respectively. Histopathologic findings suggestive of H. capsulatum were found in 63.2% of the performed exams. Serology had a lower proportion of positivity amongst AIDS patients, when compared with HIV negative patients (X2 = 6.65; p lower than 0.008). Statistical differences between AIDS and non-AIDS patients were not observed with culture and histopathology. The specific role of each test varies according to the clinical form. Physicians need to know the value and limitations of the available diagnostic tests, but before that, they have to think about histoplasmosis and consider this clinical entity in their differential diagnosis.     

Structural rearrangement of model membranes by the peptide antibiotic NK-2

We have developed a novel α-helical peptide antibiotic termed NK-2. It efficiently kills bacteria, but not human cells, by membrane destruction. This selectivity could be attributed to the different membrane lipid compositions of the target cells. To understand the mechanisms of selectivity and membrane destruction, we investigated the influence of NK-2 on the supramolecular aggregate structure, the phase transition behavior, the acyl chain fluidity, and the surface charges of phospholipids representative for the bacterial and the human cell cytoplasmic membranes. The cationic NK-2 binds to anionic phosphatidylglycerol liposomes, causing a thinning of the membrane and an increase in the phase transition temperature. However, this interaction is not solely of electrostatic but also of hydrophobic nature, indicated by an overcompensation of the Zeta potential. Whereas NK-2 has no effect on phosphatidylcholine liposomes, it enhances the fluidity of phosphatidylethanolamine acyl chains and lowers the phase transition enthalpy of the gel to liquid cristalline transition. The most dramatic effect, however, was observed for the lamellar/inverted hexagonal transition of phosphatidylethanolamine which was reduced by more than 10 °C. Thus, NK-2 promotes a negative membrane curvature which can lead to the collapse of the phosphatidylethanolamine-rich bacterial cytoplasmic membrane.     

Phylogenetic and molecular clock inferences of cyanobacterial strains within Rivulariaceae from distant environments

Heterocyst-forming cyanobacteria are important players at both evolutionary and ecological scales, but to date it has been difficult to establish their phylogenetic affiliations. We present data from a phylogenetic and molecular clock analysis of heterocystous cyanobacteria within the family Rivulariaceae, including the genera Calothrix, Rivularia, Gloeotrichia and Tolypothrix. The strains were isolated from distant geographic regions including fresh and brackish water bodies, microbial mats from beach rock, microbialites, pebble beaches, plus PCC strains 7103 and 7504. Phylogenetic inferences (distance, likelihood and Bayesian) suggested the monophyly of genera Calothrix and Rivularia. Molecular clock estimates indicate that Calothrix and Rivularia originated ～1500 million years ago (MYA) ago and species date back to 400-300 MYA while Tolypothrix and Gloeotrichia are younger genera (600-400 MYA).     

The colonization history of Juniperus brevifolia (Cupressaceae) in the Azores Islands

Background

A central aim of island biogeography is to understand the colonization history of insular species using current distributions, fossil records and genetic diversity. Here, we analyze five plastid DNA regions of the endangered Juniperus brevifolia, which is endemic to the Azores archipelago.

Methodology/Principal Findings

The phylogeny of the section Juniperus and the phylogeographic analyses of J. brevifolia based on the coalescence theory of allele (plastid) diversity suggest that: (1) a single introduction event likely occurred from Europe; (2) genetic diversification and inter-island dispersal postdated the emergence of the oldest island (Santa Maria, 8.12 Ma); (3) the genetic differentiation found in populations on the islands with higher age and smaller distance to the continent is significantly higher than that on the younger, more remote ones; (4) the high number of haplotypes observed (16), and the widespread distribution of the most frequent and ancestral ones across the archipelago, are indicating early diversification, demographic expansion, and recurrent dispersal. In contrast, restriction of six of the seven derived haplotypes to single islands is construed as reflecting significant isolation time prior to colonization.

Conclusions/Significance

Our phylogeographic reconstruction points to the sequence of island emergence as the key factor to explain the distribution of plastid DNA variation. The reproductive traits of this juniper species (anemophily, ornithochory, multi-seeded cones), together with its broad ecological range, appear to be largely responsible for recurrent inter-island colonization of ancestral haplotypes. In contrast, certain delay in colonization of new haplotypes may reflect intraspecific habitat competition on islands where this juniper was already present.     

TLR5-dependent immunogenicity of a recombinant fusion protein containing an immunodominant epitope of malarial circumsporozoite protein and the FliC flagellin of Salmonella Typhimurium

Recently, we described the improved immunogenicity of new malaria vaccine candidates based on the expression of fusion proteins containing immunodominant epitopes of merozoites and Salmonella enterica serovar Typhimurium flagellin (FliC) protein as an innate immune agonist. Here, we tested whether a similar strategy, based on an immunodominant B-cell epitope from malaria sporozoites, could also generate immunogenic fusion polypeptides. A recombinant His6-tagged FliC protein containing the C-terminal repeat regions of the VK210 variant of Plasmodium vivax circumsporozoite (CS) protein was constructed. This recombinant protein was successfully expressed in Escherichia coli as soluble protein and was purified by affinity to Ni-agarose beads followed by ion exchange chromatography. A monoclonal antibody specific for the CS protein of P. vivax sporozoites (VK210) was able to recognise the purified protein. C57BL/6 mice subcutaneously immunised with the recombinant fusion protein in the absence of any conventional adjuvant developed protein-specific systemic antibody responses. However, in mice genetically deficient in expression of TLR5, this immune response was extremely low. These results extend our previous observations concerning the immunogenicity of these recombinant fusion proteins and provide evidence that the main mechanism responsible for this immune activation involves interactions with TLR5, which has not previously been demonstrated for any recombinant FliC fusion protein.     

Land, language, and loci: mtDNA in Native Americans and the genetic history of Peru

Despite a long history of complex societies and despite extensive present-day linguistic and ethnic diversity, relatively few populations in Peru have been sampled for population genetic investigations. In order to address questions about the relationships between South American populations and about the extent of correlation between genetic distance, language, and geography in the region, mitochondrial DNA (mtDNA) hypervariable region I sequences and mtDNA haplogroup markers were examined in 33 individuals from the state of Ancash, Peru. These sequences were compared to those from 19 American Indian populations using diversity estimates, AMOVA tests, mismatch distributions, a multidimensional scaling plot, and regressions. The results show correlations between genetics, linguistics, and geographical affinities, with stronger correlations between genetics and language. Additionally, the results suggest a pattern of differential gene flow and drift in western vs. eastern South America, supporting previous mtDNA and Y chromosome investigations.