Efficient Genome-Wide Detection and Cataloging of EMS-Induced Mutations Using Exome Capture and Next-Generation Sequencing

Chemical mutagenesis efficiently generates phenotypic variation in otherwise homogeneous genetic backgrounds, enabling functional analysis of genes. Advances in mutation detection have brought the utility of induced mutant populations on par with those produced by insertional mutagenesis, but systematic cataloguing of mutations would further increase their utility. We examined the suitability of multiplexed global exome capture and sequencing coupled with custom-developed bioinformatics tools to identify mutations in well-characterized mutant populations of rice (Oryza sativa) and wheat (Triticum aestivum). In rice, we identified ∼18,000 induced mutations from 72 independent M2 individuals. Functional evaluation indicated the recovery of potentially deleterious mutations for >2600 genes. We further observed that specific sequence and cytosine methylation patterns surrounding the targeted guanine residues strongly affect their probability to be alkylated by ethyl methanesulfonate. Application of these methods to six independent M2 lines of tetraploid wheat demonstrated that our bioinformatics pipeline is applicable to polyploids. In conclusion, we provide a method for developing large-scale induced mutation resources with relatively small investments that is applicable to resource-poor organisms. Furthermore, our results demonstrate that large libraries of sequenced mutations can be readily generated, providing enhanced opportunities to study gene function and assess the effect of sequence and chromatin context on mutations.     

Isolation of an aboriginal bacterial community capable of utilizing cyanide,thiocyanate, and ammonia from metallurgical plant wastewater

An aboriginal bacterial community capable of degrading cyanide (10 mg/l) and thiocyanate (2 g/l) and eliminating ammonia (120 mg/l) had been isolated from recycled water samples after blast-furnace gas purification of a metallurgical plant wastewater. It was shown that the optimal conditions for this bacterial community were as follows: temperature, 34°C; pH, 8.8–9.0; available organic matter concentration (glucose equivalent), 5 g/l; and dissolved O₂ concentration, 8–10 mg/l. This aboriginal community was formed by the bacteria belonging to the genus Pseudomonas.     

Adipose tissue inflammation and liver fat in patients with highly active antiretroviral therapy-associated lipodystrophy

In this cross-sectional study, we sought to determine whether gene expression of macrophage markers and inflammatory chemokines in lipoatrophic subcutaneous abdominal adipose tissue and liver fat content are increased and interrelated in human immunodeficiency virus (HIV)-1-positive, highly active antiretroviral therapy (HAART)-treated patients with lipodystrophy (HAART+LD+; n = 27) compared with those without (HAART+LD-; n = 13). The study groups were comparable with respect to age, gender, and body mass index. The HAART+LD+ group had twofold more intra-abdominal (P = 0.01) and 1.5-fold less subcutaneous (P = 0.091) fat than the HAART+LD- group. As we have reported previously, liver fat was 10-fold higher in the HAART+LD+ compared with the HAART+LD- group (P = 0.00003). Inflammatory gene expression was increased in HAART-lipodystrophy: CD68 4.5-fold (P = 0.000013), tumor necrosis factor (TNF)-alpha 2-fold (P = 0.0094), chemokine (C-C motif) ligand (CCL) 2 2.5-fold (P = 0.0024), CCL3 7-fold (P = 0.0000017), integrin alphaM (ITGAM) 3-fold (P = 0.00067), epidermal growth factor-like module containing, mucin-like, hormone receptor-like (EMR)1 2.5-fold (P = 0.0038), and a disintegrin and metalloproteinase domain (ADAM)8 3.5-fold (P = 0.00057) higher in the HAART+LD+ compared with the HAART+LD- group. mRNA concentration of CD68 (r = 0.37, P = 0.019), ITGAM (r = 0.35, P = 0.025), CCL2 (r = 0.39, P = 0.012), and CCL3 (r = 0.54, P = 0.0003) correlated with liver fat content. In conclusion, gene expression of markers of macrophage infiltration and adipose tissue inflammation is increased in lipoatrophic subcutaneous abdominal adipose tissue of patients with HAART-associated lipodystrophy compared with those without. CD68, ITGAM, CCL2, and CCL3 expression is significantly associated with accumulation of liver fat.     

MIC and MBC of quinupristin/dalfopristin of erythromycin-resistant MRSA strains isolated from clinical specimens

The MICs and MBCs of quinupristin/dalfopristin were determined for 22 clinical strains MRSA with inducible type of resistance to MLS-B and for 15 of their derivatives with constitutive resistance to MLS-B. For MRSA strains with inducible resistance to MLS-B the obtained results for quinupristin/ dalfopristin were: MIC50 = 0.25, MIC90 = 0.5, MBC50 = 1.0 and MBC90 = 1.0. Mutants of the same strains characterized with the following values for quinopristin/dalfopristin: MIC50 = 0.5, MIC90 = 1.5, MBC50 = 4.0 and MTC90 = 8.0.     

The frequency of the occurrence of genes ermA, ermB, ermC and msrA/B among methicillin-resistant Staphylococcus aureus strains resistant to erythromycin

The group of 50 clinical MRSA strains resistant to MLS-B was examined for the presence of ermA, ermB, ermC, msrA/B genes by using PCR. Gene ermA was found in 43 strains (86%). 20 of ermA strains demonstrated inducible whereas 23 constitutive type of expression. The gene ermC was present in 15 of examined MRSA strains (30%). The expression of the gene was inducible in the case of 9 and constitutive in the case of 6 of the strains. The msrA/B gene was present in the case of 5 strains (10%). The ermB gene was not detected among the investigated strains.     

Does Human–Animal Similarity Lower the Need to Affirm Humans’ Superiority Relative to Animals? A Social Psychological Viewpoint

This research integrates different social psychological theories to test whether human–animal similarity promotes affiliation with animals and lowers the need to affirm humans’ superiority relative to animals. On the basis of theories of intergroup relations, terror management theory, and work conducted in the field of human–animal relations, we expected that higher human–animal similarity would decrease the need to affirm humans’ superiority relative to animals, by triggering a greater sense of shared social identity. Two correlational studies (ns = 187 and 191) tested a mediation model whereby perceived human–animal similarity was expected to predict a lower need to differentiate the perceived status of humans from animals through a process of social identification with animals. Mediated regressions provided support for these associations. A repeated-measures experiment (n = 176) replicated these findings by systematically comparing objectively (phylogenetically) more vs. less similar animals and assessing perceived status and identification. Results are discussed in light of theories of intergroup relations, terror management theory, as well as recent advances in the field of social psychology and self and identity processes.     

The application of succession analysis in combination with extremely high-frequency irradiation to the selective isolation of actinomycetes from soil

A new method employing succession analysis and extremely high-frequency (EHF) irradiation is proposed for the selective isolation of actinomycetes from soil. Total actinomycetes were efficiently isolated from soil suspensions irradiated in the wavelength band 4.6-5.8 mm on the 14th and 45th days of succession initiated by soil wetting and from soil suspensions irradiated in the wavelength band 8-11.5 mm on the 7th day of succession. The rare actinomycete genera Actinomadura, Micromonospora, Nonomuraea, Microbispora, Amycolatopsis, Pseudonocardia, Saccharothrix, Streptosporangium, Actinosynnema, Nocardioides, and Saccharopolyspora were isolated by either of the two approaches (succession analysis and EHF irradiation); however, the range of isolated rare actinomycetes was considerably wider when the combination of the two approaches was used. For instance, actinomycetes of the rare genera Actinocorallia, Promicromonospora, Actinoplanes, and Kibdelosporangium were isolated only when EHF irradiation was employed at the early stages of succession.     

A keystone-like grain in hornet comb cells: its nature and physical properties

In the roof of cells built by the Oriental hornet, Vespa orientalis (Vespinae, Hymenoptera), there is one or more grains differing in their composition and shape from the surrounding structures. These grains have a diameter of 100-200 microm and protrude slightly from the roof inward. They contain primarily a large percentage of Ti but also Fe and occasionally Si. The mineral makeup in the grains is still uncertain, but it seems that the minerals belong to the group called perovskite. These grains in the hornet's comb may originate from the soil around the nest, that is, comprised of ceramic matter, but it is also possible that they are secreted from a gland within the frons plate of the hornet because the fungus-like secretion in this gland contains elements similar to those in the roof of the comb cells. We conjecture that the grain in question serves as a reference point for the hornets that build the cell walls in the direction of gravity. The cement whereby the building hornets conjoin the various building materials, whether organic (cellulose and the like) or inorganic (sand grains or tiny stones), is a saliva that is secreted from their oral gland, which upon release hardens into a polymeric glue. The ceramic grain embedded within this polymeric matrix is endowed with different physical properties and can therefore respond differently to the tension or pressure of the surrounding walls.     

Mitochondria structural reorganization during mouse embryonic stem cell derivation

Mouse embryonic stem (ES) cells are widely used in developmental biology and transgenic research. Despite numerous studies, ultrastructural reorganization of inner cell mass (ICM) cells during in vitro culture has not yet been described in detail. Here, we for the first time performed comparative morphological and morphometric analyses of three ES cell lines during their derivation in vitro. We compared morphological characteristics of blastocyst ICM cells at 3.5 and 4.5 days post coitum on feeder cells (day 6, passage 0) with those of ES cells at different passages (day 19, passage 2; day 25, passage 4; and passage 15). At passage 0, there were 23–36% of ES-like cells with various values of the medium cross-sectional area and nucleocytoplasmic parameters, 55% of fibroblast-like (probably trophoblast derivatives), and ~?19% of dying cells. ES-like cells at passage 0 contained autolysosomes and enlarged mitochondria with reduced numerical density per cell. There were three types of mitochondria that differed in matrix density and cristae width. For the first time, we revealed cells that had two and sometimes three morphologically distinct mitochondria types in the cytoplasm. At passage 2, there were mostly ES cells with a high nucleocytoplasmic ratio and a cytoplasm depleted of organelles. At passage 4, ES cell morphology and morphometric parameters were mostly stable with little heterogeneity. According to our data, cellular structures of ICM cells undergo destabilization during derivation of an ES cell line with subsequent reorganization into the structures typical for ES cells. On the basis of ultrastructural analysis of mitochondria, we believe that the functional activity of these organelles changes during early stages of ES cell formation from the ICM.