Transfection with steroid-responsive reporter constructs shows glucocorticoid rather than androgen responsiveness in cultured Sertoli cells

It remains unclear why it has proven so difficult to identify androgen target genes in cultured Sertoli cells. Given the lack of useful endogenous reporter genes, we studied the androgen and glucocorticoid responsiveness of these cells by transfection with three different steroid-responsive reporter constructs. The constructs were driven by the tyrosine aminotransferase steroid-responsive region (TAT-GRE4x-Luc), the mouse mammary tumor virus promoter (MMTV-Luc) and the Pem homeobox gene proximal promoter respectively (Pem-Luc). These constructs can be activated either by both the glucocorticoid receptor (GR) and the androgen receptor (AR) (TAT-GRE4x-Luc and MMTV-Luc) or selectively by the AR (Pem-Luc). Despite high transfection efficiency (30-40%) none of the constructs could be activated by treatment of the Sertoli cells with testosterone, 5alpha-dihydrotestosterone or synthetic androgens. Even pretreatment with follicle-stimulating hormone to raise AR levels (from 31 up to 82fmol/mg protein) did not result in androgen responsiveness. In contrast, treatment with dexamethasone markedly stimulated TAT-GRE4x-Luc and MMTV-Luc activity. GR levels reached a value of 172fmol/mg protein in the cultured cells and both AR and GR displayed homogeneous distribution by immunocytochemical evaluation. Androgen responsiveness was restored and glucocorticoid responsiveness was increased by cotransfection with AR or GR expression constructs. Under cotransfection conditions, 1nM of testosterone (a concentration that is some 100 times lower than that estimated to be present in the testis) was sufficient to stimulate the TAT-GRE4x-Luc maximally. Our data indicate that cultured Sertoli cells respond better to glucocorticoids than to androgens and that one of the factors limiting androgen responsiveness is the availability of AR. Other factors limiting the transactivation capacity of the (endogenous) AR, however, cannot be excluded.     

Dynamics of an Oligotrophic Bacterial Aquifer Community during Contact with a Groundwater Plume Contaminated with Benzene, Toluene, Ethylbenzene, and Xylenes: an In Situ Mesocosm Study

An in situ mesocosm system was designed to monitor the in situ dynamics of the microbial community in polluted aquifers. The mesocosm system consists of a permeable membrane pocket filled with aquifer material and placed within a polypropylene holder, which is inserted below groundwater level in a monitoring well. After a specific time period, the microcosm is recovered from the well and its bacterial community is analyzed. Using this system, we examined the effect of benzene, toluene, ethylbenzene, and xylene (BTEX) contamination on the response of an aquifer bacterial community by denaturing gradient gel electrophoresis analysis of PCR-amplified 16S rRNA genes and PCR detection of BTEX degradation genes. Mesocosms were filled with nonsterile or sterile aquifer material derived from an uncontaminated area and positioned in a well located in either the uncontaminated area or a nearby contaminated area. In the contaminated area, the bacterial community in the microcosms rapidly evolved into a stable community identical to that in the adjacent aquifer but different from that in the uncontaminated area. At the contaminated location, bacteria with tmoA- and xylM/xylE1-like BTEX catabolic genotypes colonized the aquifer, while at the uncontaminated location only tmoA-like genotypes were detected. The communities in the mesocosms and in the aquifer adjacent to the wells in the contaminated area consisted mainly of Proteobacteria. At the uncontaminated location, Actinobacteria and Proteobacteria were found. Our results indicate that communities with long-term stability in their structures follow the contamination plume and rapidly colonize downstream areas upon contamination.     

Nickel removal from nickel plating waste water using a biologically active moving-bed sand filter

Efficient removal of dissolved nickel was observed in a biologically active moving-bed `MERESAFIN' sand filter treating rinsing water from an electroless nickel plating plant. Although nickel is fully soluble in this waste water, its passage through the sand filter promoted rapid removal of approximately 1 mg Ni/l. The speciation of Ni in the waste water was modelled; the most probable precipitates forming under the conditions in the filter were predicted using PHREEQC. Analyses of the Ni-containing biosludge using chemical, electron microscopical and X-ray spectroscopic techniques confirmed crystallisation of nickel phosphate as arupite (Ni₃(PO₄)₂.8H₂O), together with hydroxyapatite within the bacterial biofilm on the filter sand grains. Biosorption contributed less than 1% of the overall sequestered nickel. Metabolising bacteria are essential for the process; the definitive role of specific components of the mixed population is undefined but the increase in pH promoted by metabolic activity of some microbial components is likely to promote nickel desolubilisation by others.     

Genomic structure of and genome-wide recombination in the Saccharomyces cerevisiae S288C progenitor isolate EM93

The diploid isolate EM93 is the main ancestor to the widely used Saccharomyces cerevisiae haploid laboratory strain, S288C. In this study, we generate a high-resolution overview of the genetic differences between EM93 and S288C. We show that EM93 is heterozygous for >45,000 polymorphisms, including large sequence polymorphisms, such as deletions and a Saccharomyces paradoxus introgression. We also find that many large sequence polymorphisms (LSPs) are associated with Ty-elements and sub-telomeric regions. We identified 2,965 genetic markers, which we then used to genotype 120 EM93 tetrads. In addition to deducing the structures of all EM93 chromosomes, we estimate that the average EM93 meiosis produces 144 detectable recombination events, consisting of 87 crossover and 31 non-crossover gene conversion events. Of the 50 polymorphisms showing the highest levels of non-crossover gene conversions, only three deviated from parity, all of which were near heterozygous LSPs. We find that non-telomeric heterozygous LSPs significantly reduce meiotic recombination in adjacent intervals, while sub-telomeric LSPs have no discernable effect on recombination. We identified 203 recombination hotspots, relatively few of which are hot for both non-crossover gene conversions and crossovers. Strikingly, we find that recombination hotspots show limited conservation. Some novel hotspots are found adjacent to heterozygous LSPs that eliminate other hotspots, suggesting that hotspots may appear and disappear relatively rapidly.     

Predictability of Painful Stimulation Modulates the Somatosensory-Evoked Potential in the Rat

Somatosensory-evoked potentials (SEPs) are used in humans and animals to increase knowledge about nociception and pain. Since the SEP in humans increases when noxious stimuli are administered unpredictably, predictability potentially influences the SEP in animals as well. To assess the effect of predictability on the SEP in animals, classical fear conditioning was applied to compare SEPs between rats receiving SEP-evoking electrical stimuli either predictably or unpredictably. As in humans, the rat’s SEP increased when SEP-evoking stimuli were administered unpredictably. These data support the hypothesis that the predictability of noxious stimuli plays a distinctive role in the processing of these stimuli in animals. The influence of predictability should be considered when studying nociception and pain in animals. Additionally, this finding suggests that animals confronted with (un)predictable noxious stimuli can be used to investigate the mechanisms underlying the influence of predictability on central processing of noxious stimuli.     

Phenotypic evidence for hidden biodiversity in the <Emphasis Type="Italic">Merodon aureus</Emphasis> group (Diptera,Syrphidae) on the Balkan Peninsula: conservation implication

Cryptic species and phenotypic divergent units provided useful information about hidden biodiversity in the Merodon genus (Diptera, Syrphidae) on the Balkan Peninsula. Discrimination three cryptic species within both the M. aureus (M. aureus A, M. aureus B and M. aureus C) and M. cinereus (M. cinereus A, M. cinereus B and M. cinereus C) complexes was done by study of subtle wing variation (wing size and shape) using a geometric morphometric approach. Observed interspecific differentiation is generally in agreement with a previous study using molecular markers (allozyme loci, COI mtDNA). A considerable evolutionary and phenotypic intraspecies diversity of the M. aureus A, M. aureus B, M. aureus C, M. cinereus A, M. cinereus B, M. cinereus C and M. funestus species from the Balkan Peninsula has important implications for diagnosing biodiversity, including endemic and cryptic species. Observed phenotypic divergent units within the species might be considered as an evolutionary potential of the M. aureus group and used for defining conservation priorities. This study has contributed to the recognition of the value of wing traits in order to decipher the hidden diversity and evolutionary diversification.     

A widespread plant-fungal-bacterial symbiosis promotes plant biodiversity,plant nutrition and seedling recruitment

Highly diverse microbial assemblages colonize plant roots. It is still poorly understood whether different members of this root microbiome act synergistically by supplying different services (for example, different limiting nutrients) to plants and plant communities. In order to test this, we manipulated the presence of two widespread plant root symbionts, arbuscular mycorrhizal fungi and nitrogen-fixing rhizobia bacteria in model grassland communities established in axenic microcosms. Here, we demonstrate that both symbionts complement each other resulting in increased plant diversity, enhanced seedling recruitment and improved nutrient acquisition compared with a single symbiont situation. Legume seedlings obtained up to 15-fold higher productivity if they formed an association with both symbionts, opposed to productivity they reached with only one symbiont. Our results reveal the importance of functional diversity of symbionts and demonstrate that different members of the root microbiome can complement each other in acquiring different limiting nutrients and in driving important ecosystem functions.     

Integrative approach revealed contrasting pattern of spatial structuring within urban and rural biotypes of Culex pipiens

Population structure of pests is an important issue when designing management strategies to optimize control measures. In this study, we investigated a spatial pattern of genetic and phenotypic variation within seven urban and within six rural populations of Culex pipiens from Vojvodina Province (Serbia) incorporating landscape genetic methods (using allozyme data) and wing size and shape (using geometric morphometric approach). Comparing rural samples, no strong genetic groupings of individuals were detected. Nevertheless, traditional approaches where individuals are pre‐assigned to populations, including F statistics and amova (analysis of molecular variance), revealed low, but significant genetic differentiation among samples. Similarly, phenotypic data (wing size and shape) indicated some level of heterogeneity among rural samples. Contrary to genetic homogeneity found within rural biotype, the individual‐based structuring characterized urban biotype. Geneland revealed the presence of two genetic clusters within urban group which is in concordance with F_ST and amova results. These results showed that sample from Novi Sad (NS) is a distinct genetic unit, which has been likely resulted in intensive insecticide use over several decades. Furthermore, phenotypic differentiation supported the existence of spatial structuring. Therefore, complementary use of molecular markers and phenotypic traits may be a powerful tool for revealing hidden spatial diversity within Cx. pipiens.     

Genetic structure of Suillus luteus populations in heavy metal polluted and nonpolluted habitats

The genetic structure of populations of the ectomycorrhizal basidiomycete Suillus luteus in heavy metal polluted and nonpolluted areas was studied. Sporocarps were collected at nine different locations and genotyped at four microsatellite loci. Six of the sampling sites were severely contaminated with heavy metals and were dominated by heavy metal-tolerant individuals. Considerable genetic diversity was found within the geographical subpopulations, but no reduction of the genetic diversity, current or historic, was observed in subpopulations inhabiting polluted soils. The genetic differentiation between the geographical subpopulations was low, and no evidence for clustering of subpopulations from polluted soils vs. subpopulations from nonpolluted soils was found. These results indicate that heavy metal pollution has a limited effect on the genetic structure of S. luteus populations, and this may be due to the high frequency of sexual reproduction and extensive gene flow in S. luteus, which allows rapid evolution of the tolerance trait while maintaining high levels of genetic diversity.