Distribution of dissolved trace elements in the Laptev Sea affected by the Lena River discharge.

This study presents new data on concentration of dissolved trace elements (DTE) in the Lena River-Laptev Sea mixing zone. Mean concentrations of some dissolved heavy metals in the mixing zone of fresh waters of the Lena River and sea waters of the Laptev Sea on the middle shelf and on the outer shelves are: 0.7± 0.05 µÐœ and 0.5 ± 0.04 µÐœ for Fe, 0.06 ± 0.01 µÐœ and 0.07 ± 0.01 µÐœ for Ni, 0.01 ± 0.003 µÐœ and 0.003 ± 0.002 µÐœ for Zn, 59.2 ± 7.4 nМ and 73.4 ± 12.8 nМ for Cu, respectively. Two major groups of DTE distribution were revealed according to their spatial behavior. The Li, V, As, Rb, Sr, Mo, U concentrations increase towards the outer shelf with increasing salinity. In contrast, mean concentrations of Al, Ti, Mn, Fe, Co decrease with increasing distance from the coast. The identified transport of freshwaters to a distance of 400 km is reflected in the distribution of DTE, which suggests that these elements are able to reach to the Central Arctic Ocean.

Phylogeny and Classification of Yersinia pestis Through the Lens of Strains From the Plague Foci of Commonwealth of Independent States.

The established phylogeny of the etiological agent of plague, Yersinia pestis, is not perfect, as it does not take into account the strains from numerous natural foci of Commonwealth of Independent States (CIS). We have carried out PCR and SNP typing of 359 strains and whole genome sequencing of 51 strains from these plague foci and determined the phylogenetic diversity of the strains circulating here. They belong to 0.ANT3, 0.ANT5, 2.ANT3, 4.ANT branches of antique biovar, 2.MED0, 2.MED1 branches of medieval biovar and to 0.PE2, 0.PE4a. 0.PE4h, 0.PE4t branches. Based on the studies of 178 strains from 23 plague foci of CIS countries, it was determined that the population structure of 2.MED strains is subdivided into Caucasian-Caspian and Central Asian-Chinese branches. In Central-Caucasian high-mountain plague foci in the Russian Federation (RF) the most deeply diverged branch of medieval biovar, 2.MED0, has been found. With the data obtained, the current population structure of Y. pestis species has been refined. New subspecies classification is developed, comprising seven subspecies: pestis, caucasica (0.PE2), angolica (0.PE3), central asiatica (0.PE4), tibetica (0.PE7), ulegeica (0.PE5), and qinghaica (0.PE10).

Yersinia pestis strains of ancient phylogenetic branch 0.ANT are widely spread in the high-mountain plague foci of Kyrgyzstan.

Fifty six Yersinia pestis strains, isolated over the period of more than 50 years in three high-mountain foci of Kyrgyzstan (Tien Shan, Alai, and Talas), have been characterized by means of PCR and single nucleotide polymorphism (SNP) typing methods. Seven of these strains were also characterized by means of whole genome sequencing and genome-wide SNP phylogenetic analysis. It was found that forty two strains belong to 0.ANT2, 0.ANT3 and 0.ANT5 phylogenetic branches. From these, strains of 0.ANT2 and 0.ANT3 branches were earlier detected in China only, whereas 0.ANT5 phylogenetic branch was identified for Y. pestis phylogeny for the first time. According to the results of genome-wide SNP analysis, 0.ANT5 strains are ones of the most closely related to Y. pestis strain responsible for the Justinianic Plague. We have also found out that four of the studied strains belong to the phylogenetic branch 2.MED1, and ten strains from Talas high-mountain focus belong to the phylogenetic branch 0.PE4 (sub-branch 0.PE4t). Established diversity of Y. pestis strains and extensive dissemination of the strains pertaining to the 0.ANT branch confirm the antiquity of the mentioned above plague foci and suggest that strains of the 0.ANT branch, which serve as precursors for all highly virulent Y. pestis strains, had their origin in the Tien Shan mountains.

Sources and behaviour of nitrogen compounds in the shallow groundwater of agricultural areas (Poyang Lake basin, China).

Nitrogen contamination of natural water is a typical problem for various territories throughout the world. One of the regions exposed to nitrogen pollution is located in the Poyang Lake basin. As a result of agricultural activity and dense population, the shallow groundwater of this area is characterised by a high concentration of nitrogen compounds, primarily NO3-, with the concentration varying from 0.1mg/L to 206mg/L. Locally, high ammonium content occurs in the shallow groundwater with low reduction potential Eh (<100mV). However, in general, the shallow groundwater of the Poyang Lake basin has Eh>100mV. To identify sources of nitrogen species and the factors that determine their behaviour, the dual stable isotope approach (δ15N and δ18О) and physical-chemical modelling were applied. Actual data were collected by sampling shallow groundwater from domestic water supply wells around the lake. The δ18О values from -4.1‰ to 13.9‰ with an average value of 5.3 permille indicate a significant influence of nitrification on nitrogen balance. The enrichment of nitrate with the 15N isotope indicates that manure and domestic sewage are the principal sources of nitrogen compounds. Inorganic nitrogen speciation and thermodynamic calculations demonstrate the high stability of nitrate in the studied groundwater. Computer simulation and field observations indicate the reducing conditions formed under joint effects of anthropogenic factors and appropriate natural conditions, such as the low-level topography in which decreased water exchange rate can occur. The simulation also demonstrates the growth in pH of the groundwater as a consequence of fertilisation, which, in turn, conduced to the clay mineral formation at lower concentrations of aqueous clay-forming components than the ones under the natural conditions.

Antisense noncoding RNA promoter regulates the timing of de novo methylation of an imprinting control region.

Epigenetic marks at cis acting imprinting control regions (ICRs) regulate parent of origin-specific expression of multiple genes in imprinted gene clusters. Epigenetic marks are acquired during gametogenesis and maintained faithfully thereafter. However, the mechanism by which differential epigenetic marks are established and maintained at ICRs is currently unclear. By using Kcnq1 ICR as a model system, we have investigated the functional role of genetic signatures in the acquisition and maintenance of epigenetic marks. Kcnq1 ICR is methylated on the maternal chromosome but remains unmethylated on the paternal chromosome. Here, we show that a paternal allele of Kcnq1 ICR lacking the Kcnq1ot1 promoter remains unmethylated during spermatogenesis; however, it becomes methylated specifically during pre-implantation development. Analysis of the chromatin structure at the paternal ICR in spermatogenic cells and in E13.5 embryonic tissues revealed that the ICRs of both wild type and mutant mice are enriched with H3K4me2 in spermatiogenic cells of the testicular compartment, but the mutant ICR lost H3K4me2 specifically in epididymal sperm and an increase in repressive marks was observed in embryonic tissues. Interestingly, we also detected a decrease in nucleosomal histone levels at the mutant ICR in comparison to the wild-type ICR in epididymal sperm. Taken together, these observations suggest that the Kcnq1ot1 promoter plays a critical role in establishing an epigenetic memory in the male germline by ensuring that the paternal allele remains in an unmethylated state during pre-implantation development.

Kcnq1ot1 noncoding RNA mediates transcriptional gene silencing by interacting with Dnmt1.

A long noncoding RNA, Kcnq1ot1, regulates the expression of both ubiquitously and tissue-specific imprinted genes within the Kcnq1 domain. However, the functional sequences of the Kcnq1ot1 RNA that mediate lineage-specific imprinting are unknown. Here, we have generated a knockout mouse with a deletion encompassing an 890-bp silencing domain (Delta890) downstream of the Kcnq1ot1 promoter. Maternal transmission of the Delta890 allele has no effect on imprinting, whereas paternal inheritance of the deletion leads to selective relaxation of the imprinting of ubiquitously imprinted genes to a variable extent in a tissue-specific manner. Interestingly, the deletion affects DNA methylation at somatically acquired differentially methylated regions (DMRs), but does not affect the histone modifications of the ubiquitously imprinted genes. Importantly, we found that Kcnq1ot1 recruits Dnmt1 to somatic DMRs by interacting with Dnmt1, and that this interaction was significantly reduced in the Delta890 mice. Thus, the ubiquitous and placental-specific imprinting of genes within the Kcnq1 domain might be mediated by distinct mechanisms, and Kcnq1ot1 RNA might mediate the silencing of ubiquitously imprinted genes by maintaining allele-specific methylation through its interactions with Dnmt1.

Diagnostic microarrays in hematologic oncology: applications of high- and low-density arrays.

Microarrays have become important tools for high-throughput analysis of gene expression, chromosome aberrations, and gene mutations in cancer cells. In addition to high-density experimental microarrays, low-density, gel-based biochip technology represents a versatile platform for translation of research into clinical practice. Gel-based microarrays (biochips) consist of nanoliter gel drops on a hydrophobic surface with different immobilized biopolymers (primarily nucleic acids and proteins). Because of the high immobilization capacity of the gel, such biochips have a high probe concentration and high levels of fluorescence signals after hybridization, which allow the use of simple, portable detection systems. The notable accuracy of the analysis is reached as a result of the high level of discrimination between positive and negative gel-bound probes. Different applications of biochips in the field of hematologic oncology include analysis of chromosomal translocations in leukemias, diagnostics of T-cell lymphomas, and pharmacogenetics.

Differences in Chlamydia trachomatis serovar E growth rate in polarized endometrial and endocervical epithelial cells grown in three-dimensional culture.

In vitro studies of obligate intracellular chlamydia biology and pathogenesis are highly dependent on the use of experimental models and growth conditions that mimic the mucosal architecture and environment these pathogens encounter during natural infections. In this study, the growth of Chlamydia trachomatis genital serovar E was monitored in mouse fibroblast McCoy cells and compared to more relevant host human epithelial endometrium-derived HEC-1B and cervix-derived HeLa cells, seeded and polarized on collagen-coated microcarrier beads, using a three-dimensional culture system. Microscopy analysis of these cell lines prior to infection revealed morphological differences reminiscent of their in vivo architecture. Upon infection, early chlamydial inclusion distribution was uniform in McCoy cells but patchy in both epithelial cell lines. Although no difference in chlamydial attachment to or entry into the two genital epithelial cell lines was noted, active bacterial genome replication and transcription, as well as initial transformation of elementary bodies to reticulate bodies, were detected earlier in HEC-1B than in HeLa cells, suggesting a faster growth, which led to higher progeny counts and titers in HEC-1B cells upon completion of the developmental cycle. Chlamydial development in the less relevant McCoy cells was very similar to that in HeLa cells, although higher progeny counts were obtained. In conclusion, this three-dimensional bead culture system represents an improved model for harvesting large quantities of infectious chlamydia progeny from their more natural polarized epithelial host cells.

Characterization of estrogen-responsive epithelial cell lines and their infectivity by genital Chlamydia trachomatis.

Chlamydial attachment and infectivity in vitro and ascending disease and sequelae in vivo have been reported to be enhanced/modulated by estrogen. Endometrial carcinoma cell lines Ishikawa and HEC-1B and the breast cancer lines MCF-7 and HCC-1806 were examined for Chlamydia trachomatis E infectivity. Estrogen receptor (ER) presence was confirmed by Western blot and qRT-PCR analyses. FACS analysis was used to determine the percent of plasma membrane-localized ERs (mERs), and their activity was tested by estrogen binding and competitive estrogen antagonists assays. Chlamydiae grew in all cell lines with HEC (90%) >> MCF-7 (57%)>Ishikawa (51%) >> HCC-1806 (20%). The cell line ER isoform composition was re-defined as: ERalpha + ERbeta + for MCF-7, HCC-1806 and Ishikawa; and ERbeta only for HEC-1B. HeLa cells were also tested and found to express ERbeta, but not ERalpha. A small percentage of both ERs were surface-exposed and functionally active. The endometrium-predominant ERbeta isoform was found in all cell lines, including those most representative of the common sites of C. trachomatis infection. Thus, the role of chlamydial attachment/infectivity will now be analyzed in ERbeta+and-isogenic HEC-1B cells.

Primary cultures of female swine genital epithelial cells in vitro: a new approach for the study of hormonal modulation of Chlamydia infection.

Previous studies have demonstrated that female reproductive hormones influence chlamydial infection both in vivo and in vitro. Due to the reduced availability of human genital tissues for research purposes, an alternative hormone-responsive model system was sought to study chlamydial pathogenesis. Mature female swine eliminated from breeding programs were selected as the animals of choice because of the similarity of a sexually transmitted disease syndrome and sequelae in swine to a disease syndrome and sequelae found in humans, because of the near identity of a natural infectious chlamydial isolate from swine to Chlamydia trachomatis serovar D from humans, and because a pig's epithelial cell physiology and the mean length of its estrous cycle are similar to those in humans. Epithelial cells from the cervix, uterus, and horns of the uterus were isolated, cultivated in vitro in Dulbecco's minimum essential medium-Hanks' F-12 (DMEM-F-12) medium with and without exogenous hormone supplementation, and analyzed for Chlamydia suis S-45 infectivity. The distribution of chlamydial inclusions in swine epithelial cells was uneven and was influenced by the genital tract site and hormone status. This study confirmed that, like primary human endometrial epithelial cells, estrogen-dominant swine epithelial cells are more susceptible to chlamydial infection than are progesterone-dominant cells. Further, the more differentiated luminal epithelial cells were more susceptible to infection than were glandular epithelial cells. Interestingly, chlamydial growth in mature luminal epithelia was morphologically more active than in glandular epithelia, where persistent chlamydial forms predominated. Attempts to reprogram epithelial cell physiology and thereby susceptibility to chlamydial infection by reverse-stage, exogenous hormonal supplementation were unsuccessful. Freshly isolated primary pig epithelial cells frozen at -80 degrees C in DMEM-F-12 medium with 10% dimethyl sulfoxide for several weeks can, after thawing, reform characteristic polarized monolayers in 3 to 5 days. Thus, primary swine genital epithelia cultured ex vivo appear to be an excellent cell model for dissecting the hormonal modulation of several aspects of chlamydial pathogenesis and infection.