Detection of monochlorobenzene metabolizing bacteria under anoxic conditions by DNA-stable isotope probing

Cultivation-independent analyses were applied to study the structural diversity of the bacterial community which developed in groundwater inoculated microcosms actively metabolizing monochlorobenzene (MCB) under anaerobic conditions. Addition of ¹³C-labelled MCB demonstrated that the community produced ¹³CO₂ as a metabolite at slightly increasing rates over a period of 1,051 days while no ¹³C-methane evolved. Genetic profiles of partial 16S rRNA genes generated with the single-strand conformation polymorphism (SSCP) technique by PCR from directly extracted total DNA revealed that, despite the long incubation period, six replicate microcosms were characterized by almost the same microbial members. Nine distinguishable contributors to the SSCP-profiles were characterized by DNA sequencing, revealing the presence of different members from the phyla Proteobacteria, Fibrobacteres and from the candidate division OD1. DNA-stable isotope probing (SIP) was applied to distinguish the actual MCB metabolizing bacteria from the other community members. This study reveals for the first time the structural diversity of an anaerobic MCB metabolizing bacterial community. However, it also demonstrates the limitations of SIP to detect bacteria slowly metabolizing carbon sources under anaerobic conditions.     

Testing the emergence of New Caledonia: fig wasp mutualism as a case study and a review of evidence

While geologists suggest that New Caledonian main island (Grande Terre) was submerged until ca 37 Ma, biologists are struck by the presence of supposedly Gondwanan groups on the island. Among these groups are the Oreosycea fig trees (Ficus, Moraceae) and their Dolichoris pollinators (Hymenoptera, Agaonidae). These partners are distributed in the Paleotropics and Australasia, suggesting that their presence on New Caledonia could result from Gondwanan vicariance. To test this hypothesis, we obtained mitochondrial and nuclear markers (5.3 kb) from 28 species of Dolichoris, used all available sequences for Oreosycea, and conducted phylogenetic and dating analyses with several calibration strategies. All our analyses ruled out a vicariance scenario suggesting instead that New Caledonian colonization by Dolichoris and Oreosycea involved dispersal across islands from Sundaland ca 45.9-32.0 Ma. Our results show that successful long-distance dispersal of obligate mutualists may happen further suggesting that presence of intimate mutualisms on isolated islands should not be used as a priori evidence for vicariance. Comparing our results to a review of all the published age estimates for New Caledonian plant and animal taxa, we showed that support for a vicariant origin of the island biota is still lacking. Finally, as demonstrating a causal relationship between geology and biology requires independent evidence, we argue that a priori assumptions about vicariance or dispersal should not be used to constrain chronograms. This circular reasoning could lead to under or overestimation of age estimates.     

GABA neurons of the VTA drive conditioned place aversion

Salient but aversive stimuli inhibit the majority of dopamine (DA) neurons in the ventral tegmental area (VTA) and cause conditioned place aversion (CPA). The cellular mechanism underlying DA neuron inhibition has not been investigated and the causal link to behavior remains elusive. Here, we show that GABA neurons of the VTA inhibit DA neurons through neurotransmission at GABA(A) receptors. We also observe that GABA neurons increase their firing in response to a footshock and provide evidence that driving GABA neurons with optogenetic effectors is sufficient to affect behavior. Taken together, our data demonstrate that synaptic inhibition of DA neurons drives place aversion.     

Spatial patterns and GIS habitat modelling of fish in two French Mediterranean coastal areas

The spring and summer distribution of adults and larval fish stages of Sparids and Scorpaenids was studied in two sites in the western Mediterranean. Fish adults and larvae of those two taxa were identified and sorted according to their life stages in order to study their distribution and develop suitable habitat maps. Study areas were located on French coastal waters, the “Côte Bleue” Marine Park (CBMP) has an east-west orientation with substratum dominated by Posidonia beds and the Marine Reserve of Cerbère-Banyuls (MRCB) runs north–south and is dominated by rocky substrates. Generalised Additive Models (GAM) combined with Geographic Information System (GIS), were used to model the suitable habitats for fish larvae and adults. During spring months, waters exhibit low Sea Surface Temperatures (SST), low Photosynthetically Active Radiation (PAR), high values of Sea Surface Chlorophyll-a concentration (Chl-a), and mostly negative values of Sea Level Anomaly (SLA) indicating anticyclonic eddies. During summer months, waters have higher values of SST, lower values of Chl-a and positive values of SLA indicating cyclonic eddies. The results revealed different environmental responses in the distribution of fish adults and larvae in the CBMP and MRCB. Suitable habitats for adult were mainly dependent on the substrate types (Posidonia meadows and sand) and they were found close to the coast, whereas fish larvae were dependent on environmental cues (Chl-a, SLA, SST) with a sparse spatial distribution.     

Rainbow trout gonadal masculinization induced by inhibition of estrogen synthesis is more physiological than masculinization induced by androgen supplementation

The present study was designed to obtain new insights into fish gonadal sex differentiation by comparing the effects of two different masculinizing treatments on some candidate gene expression profiles. Masculinization was induced in rainbow trout, Oncorhynchus mykiss, genetic all-female populations using either an active fish androgen (11betaAnd, 11beta-hydroxyandrostenedione) or an aromatase inhibitor (ATD, 1,4,6-androstatriene-3,17-dione). The expression profiles of 100 candidate genes were obtained by real-time RT-PCR, and 46 profiles displayed a significant differential expression between control populations (males and females) and ATD/11betaAnd-treated populations. These expression profiles were grouped in four temporally correlated expression clusters. Among the common responses shared by the two masculinizing treatments, the inhibition of some early female differentiating genes (cyp19a1, foxl2a, fst, and fshb) appears to be crucial for effective masculinization, suggesting that these genes act together via a short regulation loop to maintain high sex-specific ovarian expression of cyp19a1. This simultaneous down-regulation of female-specific genes could be triggered by some testicular genes, such as dmrt1, nr0b1 (also known as dax1), and pdgfra, which are quickly up-regulated by the two masculinizing treatments. In contrast to 11betaAnd, ATD quickly restored the expression levels of steroidogenesis related genes (cyp11b2.1, cyp11b2.2, hsd3b1, cyp17a, star, and nr5a1) and some Sertoli cell markers (sox9a2 and amh) to the expression levels observed during control testicular differentiation. This demonstrates that these genes are probably not needed for active masculinization and that the inhibition of endogenous estrogen synthesis produces a much more complete and specific testicular pattern of gene expression than that observed following androgen-induced masculinization.     

Distribution of phytoplankton pigments in nine European estuaries and implications for an estuarine typology

Phytoplankton pigments were studied by LiquidChromatography (HPLC) in nine West Europeanestuaries. Three estuaries, i.e. the Rhine,Scheldt and the Gironde were sampled four timesto cover the different seasons, whereas theother six estuaries were sampled once. Pigmentdistributions in estuaries reflect bothriverine inputs as well as autochthonousblooms. Fucoxanthin was the most commonaccessory photosynthetic pigment showing thatDiatoms were the most common group in thestudied estuaries and were particularlydominant during autumn and winter. In the veryturbid Gironde estuary, degradation processeswere predominant between salinities 1 and 20,while Diatoms, Dinoflagellates and Cryptophytesbloomed above 20 salinity during spring andsummer. This contrasted with the highlyeutrophic but less turbid Scheldt, wherephytoplanktonic blooms occurred at lowsalinities close to the city of Antwerp. In theScheldt, we observed both a tenfold fluctuationof phytoplankton biomass and a fluctuatingpigment diversity index. In contrast,chlorophyll a was always low in theGironde, but we observed large variations ofpigment diversity among samplings duringdifferent seasons. Distribution of pheopigmentsshowed that the maximum turbidity zone (MTZ)was a highly reactive region for heterotrophicphytoplankton degradation. The Scheldt and theThames were the most anthropogenic influencedestuaries contrasting with the Gironde estuarythat has a less urbanised watershed. Anestuarine typology is proposed based on threeclusters emerging from a correspondenceanalysis of pigment variables and variablescharacterising the anthropogenic impact andphysical forcing.     

Contributions of the response regulators Ssk1p and Skn7p in the pseudohyphal development, stress adaptation, and drug sensitivity of the opportunistic yeast Candida lusitaniae

We recently characterized the histidine kinase receptor genes of Candida lusitaniae. For the present study, we have further investigated the role of SSK1 and SKN7, encoding response regulators. The results of functional analysis of mutants indicated that Ssk1p is involved in osmotolerance and pseudohyphal development, whereas Skn7p appears crucial for oxidative stress adaptation.     

Insight into the role of HOG pathway components Ssk2p, Pbs2p, and Hog1p in the opportunistic yeast Candida lusitaniae

In the present study, we have investigated the role of SSK2, PBS2, and HOG1, encoding modules of the high-osmolarity-glycerol mitogen-activated protein kinase pathway in Candida lusitaniae. Functional analysis of mutants indicated that Ssk2p, Pbs2p, and Hog1p are involved in osmotolerance, drug sensitivity, and heavy metal tolerance but not in oxidant stress adaptation.     

Glycerol induces reuterin production and decreases Escherichia coli population in an in vitro model of colonic fermentation with immobilized human feces

Lactobacillus reuteri ATCC 55730 is a probiotic strain that produces, in the presence of glycerol, reuterin, a broad-spectrum antimicrobial substance. This strain has been shown to prevent intestinal infections in vivo; however, its mechanisms of action, and more specifically whether reuterin production occurs within the intestinal tract, are not known. In this study, the effects of L. reuteri ATCC 55730 on intestinal microbiota and its capacity to secrete reuterin from glycerol in a novel in vitro colonic fermentation model were tested. Two reactors were inoculated with adult immobilized fecal microbiota and the effects of daily addition of L. reuteri into one of the reactors (c.10(8) CFU mL(-1)) without or with glycerol were tested on major bacterial populations and compared with addition of glycerol or reuterin alone. The addition of glycerol alone or with L. reuteri increased numbers of the Lactobacillus-Enterococcus group and decreased Escherichia coli. The addition of reuterin significantly and selectively decreased E. coli without affecting other bacterial populations. The observed decrease in E. coli concentration during the addition of glycerol (in presence or absence of L. reuteri) could be due to in situ reuterin production because 1,3-propanediol, a typical product of glycerol fermentation, was detected during the addition of glycerol.     

Single-nucleotide primer extension assay for detection and sequence typing of "Dehalococcoides" spp

A single-nucleotide primer extension (SNuPE) assay in combination with taxon-specific 16S rRNA gene PCR analysis was developed for the detection and typing of populations of the genus "Dehalococcoides". The specificity of the assay was evaluated with 16S rRNA gene sequences obtained from an isolate and an environmental sample representing two Dehalococcoides subgroups, i.e., the Cornell and the Pinellas subgroups. Only one sequence type, belonging to the Pinellas subgroup, was detected in a Bitterfeld-Wolfen region aquifer containing chlorinated ethenes as the main contaminants. The three-primer hybridization assay thus provided a fast and easy-to-implement method for confirming the specificity of taxon-specific PCR and allowed rapid additional taxonomic classification into subgroups. This study demonstrates the great potential of SNuPE as a novel approach for rapid parallel detection of microorganisms and typing of different nucleic acid signature sequences from environmental samples.