Release of ethane from immobilised plant cell protoplasts in response to chemical treatment

In isolated protoplasts of Vicia faba L. (cv. Weißkernige Hangdown) which were immobilised in a cross-linked alginate matrix, ethane production and ribulose bisphosphate carboxylase (RuBP carboxylase) aetivity were determined after treatment of the alginate strips with various chemical substances.
The stimulation of ethane formation and the inhibition of RuBP carboxylase activity by the toxic materials might provide the basis for a simple, rapid and sensitive system for the determination of chemicals in the environment.     

A method to generate multilocus barcodes of pinned insect specimens using MiSeq

For molecular insect identification, amplicon sequencing methods are recommended because they offer a cost‐effective approach for targeting small sets of informative genes from multiple samples. In this context, high‐throughput multilocus amplicon sequencing has been achieved using the MiSeq Illumina sequencing platform. However, this approach generates short gene fragments of <500 bp, which then have to be overlapped using bioinformatics to achieve longer sequence lengths. This increases the risk of generating chimeric sequences or leads to the formation of incomplete loci. Here, we propose a modified nested amplicon sequencing method for targeting multiple loci from pinned insect specimens using the MiSeq Illumina platform. The modification exists in using a three‐step nested PCR approach targeting near full‐length loci in the initial PCR and subsequently amplifying short fragments of between 300 and 350 bp for high‐throughput sequencing using Illumina chemistry. Using this method, we generated 407 sequences of three loci from 86% of all the specimens sequenced. Out of 103 pinned bee specimens of replicated species, 71% passed the 95% sequence similarity threshold between species replicates. This method worked best for pinned specimens aged between 0 and 5 years, with a limit of 10 years for pinned and 14 years for ethanol‐preserved specimens. Hence, our method overcomes some of the challenges of amplicon sequencing using short read next generation sequencing and improves the possibility of creating high‐quality multilocus barcodes from insect collections.     

Divergence in coloration and ecological speciation in the Anolis marmoratus species complex

Adaptive divergence in coloration is expected to produce reproductive isolation in species that use colourful signals in mate choice and species recognition. Indeed, many adaptive radiations are characterized by differentiation in colourful signals, suggesting that divergent selection acting on coloration may be an important component of speciation. Populations in the Anolis marmoratus species complex from the Caribbean island of Guadeloupe display striking divergence in the colour and pattern of adult males that occurs over small geographic distances, suggesting strong divergent selection. Here we test the hypothesis that divergence in coloration results in reduced gene flow among populations. We quantify variation in adult male coloration across a habitat gradient between mesic and xeric habitats, use a multilocus coalescent approach to infer historical demographic parameters of divergence, and examine gene flow and population structure using microsatellite variation. We find that colour variation evolved without geographic isolation and in the face of gene flow, consistent with strong divergent selection and that both ecological and sexual selection are implicated. However, we find no significant differentiation at microsatellite loci across populations, suggesting little reproductive isolation and high levels of contemporary gene exchange. Strong divergent selection on loci affecting coloration probably maintains clinal phenotypic variation despite high gene flow at neutral loci, supporting the notion of a porous genome in which adaptive portions of the genome remain fixed whereas neutral portions are homogenized by gene flow and recombination. We discuss the impact of these findings for studies of colour evolution and ecological speciation.     

Detection of dental plaque and its potential pathogenicity using quantitative light‐induced fluorescence

Quantitative light‐induced fluorescence (QLF) technology can detect some dental plaque as red fluorescence. This in vivo study aimed to identify the microbial characteristics of red fluorescent (RF) dental plaque using 16S rRNA gene sequencing and evaluate the correlations between RF plaque and the clinical symptoms of dental diseases. Paired supragingival plaque samples collected from each 10 subjects and consisted of RF and non‐RF dental plaques as observed by QLF technology using a 405 nm blue light source for excitation. The characteristics of the bacterial communities in the RF and non‐RF plaque samples were compared by sequencing analysis. An increase in microbial diversity was observed in RF plaque compared with the non‐RF plaque. There were significant differences in the community compositions between the 2 types of dental plaque. Periodontopathic bacteria were significantly more abundant in the RF plaque than non‐RF plaque. The fluorescence intensity of RF plaque was significantly related to the proportion of the periodontopathic bacterial community and the presence of gingival inflammation. In conclusion, the plaque red fluorescence is associated with changes in the microbial composition and enrichment of periodontopathic pathogens, which suggests that RF plaque detected by QLF technology could be used as a risk indicator for gingival inflammation.      

ATR kinase activity regulates the intranuclear translocation of ATR and RPA following ionizing radiation

Upon damage of DNA in eukaryotic cells, several repair and checkpoint proteins undergo a dramatic intranuclear relocalization, translocating to nuclear foci thought to represent sites of DNA damage and repair. Examples of such proteins include the checkpoint kinase ATR (ATM and Rad3-related) as well as replication protein A (RPA), a single-stranded DNA binding protein required in DNA replication and repair. Here, we used a microscopy-based approach to investigate whether the damage-induced translocation of RPA is an active process regulated by ATR. Our data show that in undamaged cells, ATR and RPA are uniformly distributed in the nucleus or localized to promyelocytic leukemia protein (PML) nuclear bodies. In cells treated with ionizing radiation, both ATR and RPA translocate to punctate, abundant nuclear foci where they continue to colocalize. Surprisingly, an ATR mutant that lacks kinase activity fails to relocalize in response to DNA damage. Furthermore, this kinase-inactive mutant blocks the translocation of RPA in a cell cycle-dependent manner. These observations demonstrate that the kinase activity of ATR is essential for the irradiation-induced release of ATR and RPA from PML bodies and translocation of ATR and RPA to potential sites of DNA damage.     

Impaired activity of volume-sensitive Cl- channel is involved in cisplatin resistance of cancer cells

The platinum-based drug cisplatin is a widely used anticancer drug which acts by causing the induction of apoptosis. However, resistance to the drug is a major problem. In this study we show that the KCP-4 human epidermoid cancer cell line, which serves as a model of acquired resistance to cisplatin, has virtually no volume-sensitive, outwardly rectifying (VSOR) chloride channel activity. The VSOR chloride channel's molecular identity has not yet been determined, and semi-quantitative RT-PCR experiments in this study suggested that the channel corresponds to none of three candidate genes. However, because it is known that the channel current plays an essential role in apoptosis, we hypothesized that lack of the current contributes to cisplatin resistance in these cells and that its restoration would reduce resistance. To test this hypothesis, we attempted to restore VSOR chloride current in KCP-4 cells. It was found that treatment with trichostatin A (TSA), a histone deacetylase inhibitor, caused VSOR chloride channel function to be partially restored. Treatment of the cells with both TSA and cisplatin resulted in an increase in caspase-3 activity at 24 h and a decrease in cell viability at 48 h. These effects were blocked by simultaneous treatment of the cells with a VSOR chloride channel blocker. These results indicate that restoration of the channel's functional expression by TSA treatment leads to a decrease in the cisplatin resistance of KCP-4 cells. We thus conclude that impaired activity of the VSOR chloride channel is involved in the cisplatin resistance of KCP-4 cancer cells.     

Evolution of sociality by natural selection on variances in reproductive fitness: evidence from a social bee

Background  

The Central Limit Theorem (CLT) is a statistical principle that states that as the number of repeated samples from any population increase, the variance among sample means will decrease and means will become more normally distributed. It has been conjectured that the CLT has the potential to provide benefits for group living in some animals via greater predictability in food acquisition, if the number of foraging bouts increases with group size. The potential existence of benefits for group living derived from a purely statistical principle is highly intriguing and it has implications for the origins of sociality.     

DNA barcoding of euryglossine bees and the description of new species of Euhesma Michener (Hymenoptera,Colletidae, Euryglossinae)

This paper launches an open access DNA barcoding project “AUSBS” under the Barcoding of Life Datasystems (BOLD). The aims of the project are to help scientists who lack the necessary morphological knowledge to identify known species using molecular markers, to aid native bee specialists with the recognition of species groups that morphologically are difficult to define, and, eventually, to assist with the recognition of new species among known species. Using integrative taxonomy, i.e. morphological comparison to type specimens in Australian museum collections combined with phylogenetic analysis of a fragment of the mitochondrial DNA cytochrome c oxidase subunit I (mtCOI) gene sequences led to the recognition of four new species of Euhesma Michener (Hymenoptera: Colletidae: Euryglossini) collected during intensive surveys in remote Australian conservation areas, which are described. The new species are Euhesma micans, Euhesma lyngouriae, and Euhesma aulaca in a species group associated with Eremophila flowers, and Euhesma albamala in the walkeriana species group.