Influence of hydrological conditions on the <Emphasis Type="Italic">Escherichia coli</Emphasis> population structure in the water of a creek on a rural watershed

Background  

Escherichia coli is a commensal bacterium of the gastro-intestinal tract of human and vertebrate animals, although the aquatic environment could be a secondary habitat. The aim of this study was to investigate the effect of hydrological conditions on the structure of the E. coli population in the water of a creek on a small rural watershed in France composed of pasture and with human occupation.     

Proteomic comparison of the cytosolic proteins of three <Emphasis Type="Italic">Bifidobacterium longum</Emphasis> human isolates and <Emphasis Type="Italic">B. longum</Emphasis> NCC2705

Background  

Bifidobacteria are natural inhabitants of the human gastrointestinal tract. In full-term newborns, these bacteria are acquired from the mother during delivery and rapidly become the predominant organisms in the intestinal microbiota. Bifidobacteria contribute to the establishment of healthy intestinal ecology and can confer health benefits to their host. Consequently, there is growing interest in bifidobacteria, and various strains are currently used as probiotic components in functional food products. However, the probiotic effects have been reported to be strain-specific. There is thus a need to better understand the determinants of the observed benefits provided by these probiotics. Our objective was to compare three human B. longum isolates with the sequenced model strain B. longum NCC2705 at the chromosome and proteome levels.     

Resveratrol suppresses body mass gain in a seasonal non-human primate model of obesity

Background  

Resveratrol, a natural polyphenolic compound, was shown to protect rodents against high-fat-diet induced diabesity by boosting energy metabolism. To the best of our knowledge, no data is yet available on the effects of resveratrol in non-human primates. Six non-human heterotherm primates (grey mouse lemurs, Microcebus murinus) were studied during four weeks of dietary supplementation with resveratrol (200 mg/kg/day) during their winter body-mass gain period. Body mass, spontaneous energy intake, resting metabolic rate, spontaneous locomotor activity and daily variations in body temperature were measured. In addition, the plasma levels of several gut hormones involved in satiety control were evaluated.     

Adaptation and response of Bifidobacterium animalis subsp. lactis to bile: a proteomic and physiological approach

Bile salts are natural detergents that facilitate the digestion and absorption of the hydrophobic components of the diet. However, their amphiphilic nature makes them very inhibitory for bacteria and strongly influences bacterial survival in the gastrointestinal tract. Adaptation to and tolerance of bile stress is therefore crucial for the persistence of bacteria in the human colonic niche. Bifidobacterium animalis subsp. lactis, a probiotic bacterium with documented health benefits, is applied largely in fermented dairy products. In this study, the effect of bile salts on proteomes of B. animalis subsp. lactis IPLA 4549 and its bile-resistant derivative B. animalis subsp. lactis 4549dOx was analyzed, leading to the identification of proteins which may represent the targets of bile salt response and adaptation in B. animalis subsp. lactis. The comparison of the wild-type and the bile-resistant strain responses allowed us to hypothesize about the resistance mechanisms acquired by the derivative resistant strain and about the bile salt response in B. animalis subsp. lactis. In addition, significant differences in the levels of metabolic end products of the bifid shunt and in the redox status of the cells were also detected, which correlate with some differences observed between the proteomes. These results indicate that adaptation and response to bile in B. animalis subsp. lactis involve several physiological mechanisms that are jointly dedicated to reduce the deleterious impact of bile on the cell's physiology.     

In situ monitoring of the nascent Pseudomonas fluorescens biofilm response to variations in the dissolved organic carbon level in low-nutrient water by attenuated total reflectance-Fourier transform infrared spectroscopy

Drinking water quality management requires early warning tools which enable water supply companies to detect quickly and to forecast degradation of the microbial quality of drinking water during its transport throughout distribution systems. This study evaluated the feasibility of assessing, in real time, drinking water biostability by monitoring in situ the evolution of the attenuated total reflectance-Fourier transform infrared (ATR-FTIR) fingerprint of a nascent reference biofilm exposed to water being tested. For this purpose, the responses of nascent Pseudomonas fluorescens biofilms to variations in the dissolved organic carbon (DOC) level in tap water were monitored in situ and in real time by ATR-FTIR spectroscopy. Nascent P. fluorescens biofilms consisting of a monolayer of bacteria were formed on the germanium crystal of an ATR flowthrough cell by pumping bacterial suspensions in Luria-Bertani (LB) medium through the cell. Then they were exposed to a continuous flow of dechlorinated sterile tap water supplemented with appropriate amounts of sterile LB medium to obtain DOC concentrations ranging from 1.5 to 11.8 mg/liter. The time evolution of infrared bands related to proteins, polysaccharides, and nucleic acids clearly showed that changes in the DOC concentration resulted in changes in the nascent biofilm ATR-FTIR fingerprint within 2 h after exposure of the biofilm to the water being tested. The initial bacterial attachment, biofilm detachment, and regrowth kinetics determined from changes in the areas of bands associated with proteins and polysaccharides were directly dependent on the DOC level. Furthermore, they were consistent with bacterial adhesion or growth kinetic models and extracellular polymeric substance overproduction or starvation-dependent detachment mechanisms.     

Poly(I:C) and lipopolysaccharide innate sensing functions of circulating human myeloid dendritic cells are affected in vivo in hepatitis C virus-infected patients

The role of peripheral dendritic cells (DCs) in hepatitis C virus (HCV) infection is unclear. To determine if persistent infection exerts an inhibitory pressure on HCV-specific innate responses, we analyzed DC function in blood through quantification of cell-associated HCV RNA levels in conjunction with multiparametric flow cytometry analysis of pathogen recognition receptor-induced cytokine expression. Independently of the serum viral load, fluorescence-activated cell sorter-purified total DCs had a wide range of cell-associated HCV genomic RNA copy numbers (mean log(10), 5.0 per 10(6) cells; range, 4.3 to 5.8). Here we report that for viremic patients with high viral loads in their total DCs, the myeloid DC (MDC) subset displayed impaired expression of interleukin-12 (IL-12) and tumor necrosis factor alpha (TNF-alpha) but normal IL-6 or chemokine CCL3 expression in response to poly(I:C) and lipopolysaccharide (LPS). IL-6-expressing cells from this subgroup of viremic patients demonstrated a significant increase (sixfold more) in TNF-alpha(-) IL-12(-) cell frequency compared to healthy donors (mean, 38.8% versus 6.5%; P < 0.0001), indicating a functional defect in a subpopulation of cytokine-producing MDCs ( approximately 6% of MDCs). Attenuation of poly(I:C) and LPS innate sensing was HCV RNA density dependent and did not correlate with viremia or deficits in circulating MDC frequencies in HCV-infected patients. Monocytes from these patients were functionally intact, responding normally on a per-cell basis following stimulation, independent of cell-associated HCV RNA levels. Taken together, these data indicate that detection of HCV genomic RNA in DCs and loss of function in the danger signal responsiveness of a small proportion of DCs in vivo are interrelated rather than independent phenomena.     

Comparative analysis of expressed genes from cacao meristems infected by Moniliophthora perniciosa

BACKGROUND AND AIMS: Witches' broom disease is caused by the hemibiotrophic basidiomycete Moniliophthora perniciosa, and is one of the most important diseases of cacao in the western hemisphere. Because very little is known about the global process of such disease development, expressed sequence tags (ESTs) were used to identify genes expressed during the Theobroma cacao-Moniliophthora perniciosa interaction. METHODS: Two cDNA libraries corresponding to the resistant (RT) and susceptible (SP) cacao-M. perniciosa interactions were constructed from total RNA, using the DB SMART Creator cDNA library kit (Clontech). Clones were randomly selected, sequenced from the 5' end and analysed using bioinformatics tools including in silico analysis of the differential gene expression. KEY RESULTS: A total of 6884 ESTs were generated from the RT and SP cDNA libraries. These ESTs were composed of 2585 singlets and 341 contigs for a total of 2926 non-redundant sequences. The redundancy of the libraries was low and their specificity high when compared with the few other cacao libraries already published. Sequence analysis allowed the assignment of a putative functional category for 54 % of sequences, whereas approx. 22 % of sequences corresponded to unknown function and approx. 24 % of sequences did not show any significant similarity with other proteins present in the database. Despite the similar overall distribution of the sequences in functional categories between the two libraries, qualitative differences were observed. Genes involved during the defence response to pathogen infection or in programmed cell death were identified, such as pathogenesis related-proteins, trypsin inhibitor or oxalate oxidase, and some of them showed an in silico differential expression between the resistant and the susceptible interactions. CONCLUSIONS: As far as is known this is the first EST resource from the cacao-M. perniciosa interaction and it is believed that it will provide a significant contribution to the understanding of the molecular mechanisms of the resistance and susceptibility of cacao to M. perniciosa, to develop strategies to control witches' broom, and as a source of polymorphism for molecular marker development and marker-assisted selection.     

Cell-permeable peptides induce dose- and length-dependent cytotoxic effects

We have explored the threshold of tolerance of three unrelated cell types to treatments with potential cytoprotective peptides bound to Tat_48-57 and Antp_43-58 cell-permeable peptide carriers. Both Tat_48-57 and Antp_43-58 are well known for their good efficacy at crossing membranes of different cell types, their overall low toxicity, and their absence of leakage once internalised. Here, we show that concentrations of up to 100 μM of Tat_48-57 were essentially harmless in all cells tested, whereas Antp_43-58 was significantly more toxic. Moreover, all peptides bound to Tat_48-57 and Antp_43-58 triggered significant and length-dependent cytotoxicity when used at concentrations above 10 μM in all but one cell types (208F rat fibroblasts), irrespective of the sequence of the cargo. Absence of cytotoxicity in 208F fibroblasts correlated with poor intracellular peptide uptake, as monitored by confocal laser scanning fluorescence microscopy. Our data further suggest that the onset of cytotoxicity correlates with the activation of two intracellular stress signalling pathways, namely those involving JNK, and to a lesser extent p38 mitogen-activated protein kinases. These responses are of particular concern for cells that are especially sensitive to the activation of stress kinases. Collectively, these results indicate that in order to avoid unwanted and unspecific cytotoxicity, effector molecules bound to Tat_48-57 should be designed with the shortest possible sequence and the highest possible affinity for their binding partners or targets, so that concentrations below 10 μM can be successfully applied to cells without harm. Considering that cytotoxicity associated to Tat_48-57- and Antp_43-58 bound peptide conjugates was not restricted to a particular type of cells, our data provide a general framework for the design of cell-penetrating peptides that may apply to broader uses of intracellular peptide and drug delivery.     

Age-related changes in microarchitecture and mineralization of cancellous bone in the porcine mandibular condyle

The mandibular condyle is considered a good model for developing cancellous bone because of its rapid growth and high rate of remodeling. The aim of the present study was to analyze the simultaneous changes in microarchitecture and mineralization of cancellous bone during development in a three-dimensional fashion. Eight mandibular condyles of pigs aged 8 weeks prepartum to 108 weeks postpartum were scanned using microCT with an isotropic spatial resolution of 10 microm. The number of trabeculae decreased during development, whereas both the trabecular thickness and the distance between the trabeculae increased. The bone surface to volume ratio decreased during development, possibly limiting the amount of (re)modeling. Both the mean degree of mineralization and intratrabecular differences in mineralization between the surfaces and cores of trabecular elements increased during development. The trabecular surfaces were more highly mineralized in the older condyles compared to the younger ones. Together with the observed decrease in the relative size of trabecular surface, this finding suggests a decrease in (re)modeling activity during development. In accordance with the general growth and development of the pig, it was concluded that most developmental changes in cancellous bone occur until the age of 40 weeks postpartum.     

FliK regulates flagellar hook length as an internal ruler

The mechanism of length control of the flagellar hook is under debate between two theories. One claims that the FliK directly measures the hook length as a molecular ruler, while the other claims that the cytoplasmic substructure measures the amount of hook subunits to determine the hook length. Both agree that the FliK C-terminal domain catalyses the substrate-specificity switch to terminate hook elongation. In this study, we systematically created fliK mutants with deletions and insertions at various sites within the FliK N-terminal domain and analysed their effects on the final hook length. Insertions of peptide fragments from the Yersinia YscP into FliK gave rise to hooks with defined lengths, which was proportional to the molecular size of the FliK-YscP chimeras. Among fliK deletion mutants, only those with small truncations in three specific sites of FliK produced hooks of a defined, shortened length. For the majority of deletion mutants, FliK was secreted, but hook length was not controlled. On the other hand, for some deletion mutants FliK was not secreted, but the hook length was controlled, indicating that FliK secretion is not necessary for hook-length control. We conclude that FliK regulates hook length as an internal molecular ruler.