Experiments with artificial nests provide evidence for ant community stratification and nest site limitation in a tropical forest

Ants are dominant in tropical forests and many species nest in hollow cavities. The manner in which species are vertically stratified in these complex habitats is not known, with lack of nest sites being proposed to limit ant populations. Here, we assess ant community stratification and nest site limitation in a lowland rainforest in New Guinea using experimental addition of artificial bamboo nests of two cavity sizes (small: ~12 mm large: ~32 mm diameter) placed at ground level, in the understorey, and in the canopy. We also conducted a pilot experiment to test the utility of nest translocation. Nests were checked for occupancy after 10 weeks and half of the occupied nests were then translocated between forest plots, while keeping same vertical position. Occupancy of small nests was much higher in the understorey and canopy than at ground level (~75% vs. ~25%). Translocation was successful, as a majority of nests was inhabited by the same species before and after translocation and there was no impact of translocation to a different plot compared to the control, except for a reduction in colony size at ground level. Our experiment demonstrates a vertical stratification in community composition of ants nesting in hollow dead cavities and shows that these ants are more nest site limited in the higher strata than at ground level. Use of small artificial cavities has great potential for future experimental studies, especially for those focused on arboreal ants, as occupancy is high and translocation does not negatively affect their colony size. Abstract in Tok Pisin is available with online material.     

Validation of a Laboratory Method for Evaluating Dynamic Properties of Reconstructed Equine Racetrack Surfaces

Background

Racetrack surface is a risk factor for racehorse injuries and fatalities. Current research indicates that race surface mechanical properties may be influenced by material composition, moisture content, temperature, and maintenance. Race surface mechanical testing in a controlled laboratory setting would allow for objective evaluation of dynamic properties of surface and factors that affect surface behavior.

Objective

To develop a method for reconstruction of race surfaces in the laboratory and validate the method by comparison with racetrack measurements of dynamic surface properties.

Methods

Track-testing device (TTD) impact tests were conducted to simulate equine hoof impact on dirt and synthetic race surfaces; tests were performed both in situ (racetrack) and using laboratory reconstructions of harvested surface materials. Clegg Hammer in situ measurements were used to guide surface reconstruction in the laboratory. Dynamic surface properties were compared between in situ and laboratory settings. Relationships between racetrack TTD and Clegg Hammer measurements were analyzed using stepwise multiple linear regression.

Results

Most dynamic surface property setting differences (racetrack-laboratory) were small relative to surface material type differences (dirt-synthetic). Clegg Hammer measurements were more strongly correlated with TTD measurements on the synthetic surface than the dirt surface. On the dirt surface, Clegg Hammer decelerations were negatively correlated with TTD forces.

Conclusions

Laboratory reconstruction of racetrack surfaces guided by Clegg Hammer measurements yielded TTD impact measurements similar to in situ values. The negative correlation between TTD and Clegg Hammer measurements confirms the importance of instrument mass when drawing conclusions from testing results. Lighter impact devices may be less appropriate for assessing dynamic surface properties compared to testing equipment designed to simulate hoof impact (TTD).

Potential Relevance

Dynamic impact properties of race surfaces can be evaluated in a laboratory setting, allowing for further study of factors affecting surface behavior under controlled conditions.     

Botulinum neurotoxin type D enables cytosolic delivery of enzymatically active cargo proteins to neurones via unfolded translocation intermediates

Multi-domain bacterial protein toxins are being explored as potential carriers for targeted delivery of biomolecules. Previous approaches employing isolated receptor binding subunits disallow entry into the cytosol. Strategies in which catalytic domains are replaced with cargo molecules are presumably inefficient due to co-operation of domains during the endosomal translocation step. Here, we characterize a novel transport vehicle in which cargo proteins are attached to the amino terminus of the full-length botulinum neurotoxin type D (BoNT/D). The intrinsic enzymatic activity of the neurotoxin allowed quantification of the efficacy of cargo delivery to the cytosol. Dihydrofolate reductase and BoNT type A (BoNT/A) light chain (LC) were efficiently conveyed into the cytosol, whereas attachment of firefly luciferase or green fluorescent protein drastically reduced the toxicity. Luciferase and BoNT/A LC retained their catalytic activity as evidenced by luciferin conversion or SNAP-25 hydrolysis in the cytosol of synaptosomes, respectively. Conformationally stabilized dihydrofolate reductase as cargo considerably decreased the toxicity indicative for the requirement of partial unfolding of cargo protein and catalytic domain as prerequisite for efficient translocation across the endosomal membrane. Thus, enzymatically inactive clostridial neurotoxins may serve as effective, safe carriers for delivering proteins in functionally active form to the cytosol of neurones.     

Within brown-fat cells, UCP1-mediated fatty acid-induced uncoupling is independent of fatty acid metabolism

In the present investigation, we have utilized the availability of UCP1(-/-) mice to examine a wide range of previously proposed lipid activators of Uncoupling Protein 1 (UCP1) in its native environment, i.e. in the brown-fat cells. A non-metabolizable fatty acid analogue, beta,beta cent-methyl-substituted hexadecane alpha,omega-dicarboxylic acid (Medica-16) is a potent UCP1 (re)activator in brown-fat cells, despite its bipolar structure. All-trans-retinoic acid activates UCP1 within cells, whereas beta-carotene only does so after metabolism. The UCP1-dependent effects of fatty acids are positively correlated with their chain length. Medium-chain fatty acids are potent UCP1 activators in cells, despite their lack of protonophoric properties in mitochondrial membranes. Thus, neither the ability to be metabolized nor an innate uncoupling/protonophoric ability is a necessary property of UCP1 activators within brown-fat cells.     

Persistently stalled replication forks inhibit nucleotide excision repair in trans by sequestering Replication protein A

Rev3, the catalytic subunit of DNA polymerase ζ, is essential for translesion synthesis of cytotoxic DNA photolesions, whereas the Rev1 protein plays a noncatalytic role in translesion synthesis. Here, we reveal that mammalian Rev3^−/− and Rev1^−/− cell lines additionally display a nucleotide excision repair (NER) defect, specifically during S phase. This defect is correlated with the normal recruitment but protracted persistence at DNA damage sites of factors involved in an early stage of NER, while repair synthesis is affected. Remarkably, the NER defect becomes apparent only at 2 h post-irradiation indicating that Rev3 affects repair synthesis only indirectly, rather than performing an enzymatic role in NER. We provide evidence that the NER defect is caused by scarceness of Replication protein A (Rpa) available to NER, resulting from its sequestration at stalled replication forks. Also the induction of replicative stress using hydroxyurea precludes the accumulation of Rpa at photolesion sites, both in Rev3^−/− and in wild-type cells. These data support a model in which the limited Rpa pool coordinates replicative stress and NER, resulting in increased cytotoxicity of ultraviolet light when replicative stress exceeds a threshold.     

Exploring the prevalence of ten polyomaviruses and two herpes viruses in breast cancer

Several different viruses have been proposed to play a role in breast carcinogenesis. The aim of this study was to investigate the prevalence of a subset of viruses in breast cancer tissue. We investigated the prevalence of 12 DNA viruses: EBV and CMV from the Herpesviridae family and SV40, BKV, JCV, MCV, WUV, KIV, LPV, HPyV6, HPyV7, and TSV from the Polyomaviridae family in 54 fresh frozen breast tumour specimens. Relevant clinical data and basic lifestyle data were available for all patients. The tissue samples were DNA extracted and real-time PCR assays were used for viral detection.The highest prevalence, 10% (5/54), was found for EBV. MCV, HPyV6, and HPyV7 were detected in single patient samples (2% each), while WUV, KIV, JCV, BKV, LPV, SV40, TSV and CMV were not detected in the 54 breast cancer specimens analysed here. Further investigations are needed to elucidate the potential role of viruses, and particularly EBV, in breast carcinogenesis.     

Kainate receptors and RNA editing in cholinergic neurons

Parasympathetic ganglia are considered simple relay systems that have cholinergic input and output, with modulation occurring centrally. Greater complexity is suggested, however, by our showing here that avian ciliary ganglion (CG) neurons also express a different excitatory receptor type--ionotropic glutamate receptors of the kainate subtype (KARs). This is the first report of glutamate receptor expression in the CG and KAR expression in any cholinergic neuron. We show that KARs form functional channels on CG neurons. KARs localize to CG neuron axons and somata as well as axons and terminals of pre-synaptic inputs to the CG. Glutamate transporters are expressed on Schwann cells that surround synapses on neuronal somata, and may provide a local source of glutamate. CG neurons express multiple KAR subunit mRNAs (GluR5, GluR7, and KA1), and their relative levels change dramatically during axon outgrowth and synaptic differentiation. The developmental role for KARs may depend upon their calcium permeability, a property regulated by mRNA editing. We show GluR5 editing increases predominantly at the time CG axons contact peripheral targets. Our data suggest that glutamatergic signaling may function as a local circuit mechanism to modulate excitability and calcium signaling during synapse formation and maturation in the CG in vivo.     

Metabolic profiling, metabolomic and metabonomic procedures for NMR spectroscopy of urine, plasma, serum and tissue extracts

Metabolic profiling, metabolomic and metabonomic studies mainly involve the multicomponent analysis of biological fluids, tissue and cell extracts using NMR spectroscopy and/or mass spectrometry (MS). We summarize the main NMR spectroscopic applications in modern metabolic research, and provide detailed protocols for biofluid (urine, serum/plasma) and tissue sample collection and preparation, including the extraction of polar and lipophilic metabolites from tissues. 1H NMR spectroscopic techniques such as standard 1D spectroscopy, relaxation-edited, diffusion-edited and 2D J-resolved pulse sequences are widely used at the analysis stage to monitor different groups of metabolites and are described here. They are often followed by more detailed statistical analysis or additional 2D NMR analysis for biomarker discovery. The standard acquisition time per sample is 4-5 min for a simple 1D spectrum, and both preparation and analysis can be automated to allow application to high-throughput screening for clinical diagnostic and toxicological studies, as well as molecular phenotyping and functional genomics.     

Oritavancin exhibits dual mode of action to inhibit cell-wall biosynthesis in Staphylococcus aureus

Solid-state NMR measurements performed on intact whole cells of Staphylococcus aureus labeled selectively in vivo have established that des-N-methylleucyl oritavancin (which has antimicrobial activity) binds to the cell-wall peptidoglycan, even though removal of the terminal N-methylleucyl residue destroys the d-Ala-d-Ala binding pocket. By contrast, the des-N-methylleucyl form of vancomycin (which has no antimicrobial activity) does not bind to the cell wall. Solid-state NMR has also determined that oritavancin and vancomycin are comparable inhibitors of transglycosylation, but that oritavancin is a more potent inhibitor of transpeptidation. This combination of effects on cell-wall binding and biosynthesis is interpreted in terms of a recent proposal that oritavancin-like glycopeptides have two cell-wall binding sites: the well-known peptidoglycan d-Ala-d-Ala pentapeptide stem terminus and the pentaglycyl bridging segment. The resulting dual mode of action provides a structural framework for coordinated cell-wall assembly that accounts for the enhanced potency of oritavancin and oritavancin-like analogues against vancomycin-resistant organisms.