Selecting representative model micro-organisms

Background  

Micro-biological research relies on the use of model organisms that act as representatives of their species or subspecies, these are frequently well-characterized laboratory strains. However, it has often become apparent that the model strain initially chosen does not represent important features of the species. For micro-organisms, the diversity of their genomes is such that even the best possible choice of initial strain for sequencing may not assure that the genome obtained adequately represents the species. To acquire information about a species' genome as efficiently as possible, we require a method to choose strains for analysis on the basis of how well they represent the species.     

Modelling expected trout ranges under current and future water temperature regimes in the Eastern Cape,South Africa

Different values have resulted in conflicts between anglers and conservation lobbies in the management of trout in South Africa. Key to the conflict is the demarcation of boundaries to areas in which brown trout Salmo trutta and rainbow trout Oncorhynchus mykiss currently occur, or are likely to establish following stocking for angling. To provide a longer-term perspective on these areas, we developed models to link salmonid biological thermal thresholds to elevation. These, when applied spatially using a digital elevation model with a probability of occurrence model, provided the basis for estimating potentially available thermal habitat for these two cold water species. Here, we acknowledge that other variables (stocking history; river connectivity) also play a role in understanding trout distributions. Using a simple scenario of an increase in mean daily water temperatures of 2 °C, we demonstrated that both brown and rainbow trout are likely to exhibit considerable range reductions in the future. Because it is possible that these range restrictions will result in an increasing desire to introduce trout into areas above their current distribution limits for the maintenance of angling opportunities, conservation managers should prioritise these areas, with management interventions seeking to understand what will help to limit introductions.     

Bronchoabsorption; a novel bronchoscopic technique to improve biomarker sampling of the airway

Background

Current techniques used to obtain lung samples have significant limitations and do not provide reproducible biomarkers of inflammation. We have developed a novel technique that allows multiple sampling methods from the same area (or multiple areas) of the lung under direct bronchoscopic vision. It allows collection of mucosal lining fluid and bronchial brushing from the same site; biopsy samples may also be taken. The novel technique takes the same time as standard procedures and can be conducted safely.

Methods

Eight healthy smokers aged 40–65 years were included in this study. An absorptive filter paper was applied to the bronchial mucosa under direct vision using standard bronchoscopic techniques. Further samples were obtained from the same site using bronchial brushings. Bronchoalveolar lavage (BAL) was obtained using standard techniques. Chemokine (C-C Motif) Ligand 20 (CCL20), CCL4, CCL5, Chemokine (C-X-C Motif) Ligand 1 (CXCL1), CXCL8, CXCL9, CXCL10, CXCL11, Interleukin 1 beta (IL-1β), IL-6, Vascular endothelial growth factor (VEGF), Matrix metalloproteinase 8 (MMP-8) and MMP-9 were measured in exudate and BAL. mRNA was collected from the bronchial brushings for gene expression analysis.

Results

A greater than 10 fold concentration of all the biomarkers was detected in lung exudate in comparison to BAL. High yield of good quality RNA with RNA integrity numbers (RIN) between 7.6 and 9.3 were extracted from the bronchial brushings. The subset of genes measured were reproducible across the samples and corresponded to the inflammatory markers measured in exudate and BAL.

Conclusions

The bronchoabsorption technique as described offers the ability to sample lung fluid direct from the site of interest without the dilution effects caused by BAL. Using this method we were able to successfully measure the concentrations of biomarkers present in the lungs as well as collect high yield mRNA samples for gene expression analysis from the same site. This technique demonstrates superior sensitivity to standard BAL for the measurement of biomarkers of inflammation. It could replace BAL as the method of choice for these measurements. This method provides a systems biology approach to studying the inflammatory markers of respiratory disease progression.

Trial registration

NHS Health Research Authority (13/LO/0256).     

A universal BMV-based RNA recombination system--how to search for general rules in RNA recombination

At present, there is no doubt that RNA recombination is one of the major factors responsible for the generation of new RNA viruses and retroviruses. Numerous experimental systems have been created to investigate this complex phenomenon. Consequently, specific RNA structural motifs mediating recombination have been identified in several viruses. Unfortunately, up till now a unified model of genetic RNA recombination has not been formulated, mainly due to difficulties with the direct comparison of data obtained for different RNA-based viruses. To solve this problem, we have attempted to construct a universal system in which the recombination activity of various RNA sequences could be tested. To this end, we have used brome mosaic virus, a model (+)RNA virus of plants, for which the structural requirements of RNA recombination are well defined. The effectiveness of the new homomolecular system has been proven in an experiment involving two RNA sequences derived from the hepatitis C virus genome. In addition, comparison of the data obtained with the homomolecular system with those generated earlier using the heteromolecular one has provided new evidence that the mechanisms of homologous and non-homologous recombination are different and depend on the virus' mode of replication.     

The ability of land plants to synthesize glucuronoxylans predates the evolution of tracheophytes

Glucuronoxylans with a backbone of 1,4-linked β-D-xylosyl residues are ubiquitous in the secondary walls of gymnosperms and angiosperms. Xylans have been reported to be present in hornwort cell walls, but their structures have not been determined. In contrast, the presence of xylans in the cell walls of mosses and liverworts remains a subject of debate. Here we present data that unequivocally establishes that the cell walls of leafy tissue and axillary hair cells of the moss Physcomitrella patens contain a glucuronoxylan that is structurally similar to glucuronoxylans in the secondary cell walls of vascular plants. Some of the 1,4-linked β-D-xylopyranosyl residues in the backbone of this glucuronoxylan bear an α-D-glucosyluronic acid (GlcpA) sidechain at O-2. In contrast, the lycopodiophyte Selaginella kraussiana synthesizes a glucuronoxylan substituted with 4-O-Me-α-D-GlcpA sidechains, as do many hardwood species. The monilophyte Equisetum hyemale produces a glucuronoxylan with both 4-O-Me-α-D-GlcpA and α-D-GlcpA sidechains, as does Arabidopsis. The seedless plant glucuronoxylans contain no discernible amounts of the reducing-end sequence that is characteristic of gymnosperm and eudicot xylans. Phylogenetic studies showed that the P. patens genome contains genes with high sequence similarity to Arabidopsis CAZy family GT8, GT43 and GT47 glycosyltransferases that are likely involved in xylan synthesis. We conclude that mosses synthesize glucuronoxylan that is structurally similar to the glucuronoxylans present in the secondary cell walls of lycopodiophytes, monilophytes, and many seed-bearing plants, and that several of the glycosyltransferases required for glucuronoxylan synthesis evolved before the evolution of tracheophytes.     

Predicted warming and browning affect timing and magnitude of plankton phenological events in lakes: a mesocosm study

1. Aquatic ecosystems in Northern Europe are expected to face increases in temperature and water colour (TB) in future. While effects of these factors have been studied separately, it is unknown whether and how a combination of them might affect phenological events and trophic interactions. 2. In a mesocosm study, we combined both factors to create conditions expected to arise during the coming century. We focused on quantifying effects on timing and magnitude of plankton spring phenological events and identifying possible mismatches between resources (phytoplankton) and consumers (zooplankton). 3. We found that the increases in TB had important effects on timing and abundance of different plankton groups. While increased temperature led to an earlier peak in phytoplankton and zooplankton and a change in the relative timing of different zooplankton groups, increased water colour reduced chlorophyll‐a concentrations. 4. Increased TB together benefitted cladocerans and calanoid copepods and led to stronger top‐down control of algae by zooplankton. There was no sign of a mismatch between primary producers and grazers as reported from other studies. 5. Our results point towards an earlier onset of plankton spring growth in shallow lakes in future with a stronger top‐down control of phytoplankton by zooplankton grazers.     

Effects of brown and turbid water on piscivore–prey fish interactions along a visibility gradient

1. Environmental changes such as eutrophication and increasing inputs of humic matter (brownification) may have strong effects on predator–prey interactions in lakes through a reduction in the visual conditions affecting foraging behaviour of visually oriented predators. 2. In this experiment, we studied the effects of visual range (25–200 cm) in combination with optically deteriorating treatments (algae, clay or brown humic water) on predator–prey interactions between pike (Esox lucius) and roach (Rutilus rutilus). We measured effects on reaction distance and strike distance for pike and escape distance for roach, when pike individuals were exposed to free‐swimming roach as well as to roach held in a glass cylinder. 3. We found that reaction distance decreased with decreasing visual range caused by increasing levels of algae, clay or humic matter. The effect of reaction distance was stronger in turbid water (clay, algae) than in the brown water treatment. 4. Strike distance was neither affected by visual range nor by optical treatment, but we found shorter strike distances when pike attacked roach using visual cues only (roach held in a cylinder) compared to when pike could use multiple senses (free‐swimming roach). Escape distance for roach was longer in turbid than in brown water treatments. 5. Changes in environmental drivers, such as eutrophication and brownification, affecting the optical climate should thus have consequences for the strength of predator–prey interactions through changes in piscivore foraging efficiency and prey escape behaviour. This in turn may affect lake ecosystems through higher‐order interactions.     

Six <Emphasis Type="Italic">Medicago truncatula</Emphasis> Dicer-like protein genes are expressed in plant cells and upregulated in nodules

Key message

Here we report the existence of six putative Dicer-like genes in the Medicago truncatula genome. They are ubiquitously expressed throughout the plant and significantly induced in root nodules.

Abstract

Over the past decade, small noncoding RNAs (sncRNA) have emerged as widespread and important regulatory molecules influencing both the structure and expression of plant genomes. One of the key factors involved in sncRNA biogenesis in plants is a group of RNase III-type nucleases known as Dicer-like (DCL) proteins. Based on functional analysis of DCL proteins identified in Arabidopsis thaliana, four types of DCLs were distinguished (DCL1-4). DCL1 mainly produces 21 nt miRNAs. The products generated by DCL2, DCL3, and DCL4 belong to various classes of siRNAs that are 22, 24 and 21 nt in length, respectively. M. truncatula is a model legume plant closely related to many economically important cultivable species. By screening the recent M. truncatula genome assembly, we were able to identify three new DCL genes in addition to the MtDCL1-3 genes that had been earlier characterized. The newly found genes include MtDCL4 and two MtDCL2 homologs. We showed that all six M. truncatula DCL genes are expressed in plant cells. The first of the identified MtDCL2 paralogs encodes a truncated version of the DCL2 protein, while the second undergoes substantial and specific upregulation in the root nodules. Additionally, we identified an alternative splicing variant of MtDCL1 mRNA, similar to the one found in Arabidopsis. Our results indicate that DCL genes are differently activated during Medicago symbiosis with nitrogen fixing bacteria and upon pathogen infection. In addition, we hypothesize that the alternative splicing variant of MtDCL1 mRNA may be involved in tissue-specific regulation of the DCL1 level.