Trabecular bone strains around a dental implant and associated micromotions—A micro-CT-based three-dimensional finite element study

The first objective of this computational study was to assess the strain magnitude and distribution within the three-dimensional (3D) trabecular bone structure around an osseointegrated dental implant loaded axially. The second objective was to investigate the relative micromotions between the implant and the surrounding bone. The work hypothesis adopted was that these virtual measurements would be a useful indicator of bone adaptation (resorption, homeostasis, formation).In order to reach these objectives, a μCT-based finite element model of an oral implant implanted into a Berkshire pig mandible was developed along with a robust software methodology. The finite element mesh of the 3D trabecular bone architecture was generated from the segmentation of μCT scans. The implant was meshed independently from its CAD file obtained from the manufacturer. The meshes of the implant and the bone sample were registered together in an integrated software environment. A series of non-linear contact finite element (FE) analyses considering an axial load applied to the top of the implant in combination with three sets of mechanical properties for the trabecular bone tissue was devised. Complex strain distribution patterns are reported and discussed. It was found that considering the Young’s modulus of the trabecular bone tissue to be 5, 10 and 15 GPa resulted in maximum peri-implant bone microstrains of about 3000, 2100 and 1400. These results indicate that, for the three sets of mechanical properties considered, the magnitude of maximum strain lies within an homeostatic range known to be sufficient to maintain/form bone. The corresponding micro-motions of the implant with respect to the bone microstructure were shown to be sufficiently low to prevent fibrous tissue formation and to favour long-term osseointegration.     

Sleep and circadian phase in a ship's crew

Numerous factors influence the increased health risks of seamen. This study investigated sleep (by actigraphy) and the adaptation of the internal clock in watch-keeping crew compared to day workers, as possible contributory factors. Fourteen watch keepers, 4 h on, 8 h off (0800-1200/2000-2400 h, 1200-1600/2400-0400 h, 1600-2000/0400-0800 h) (fixed schedule, n = 6; rotating by delay weekly, n = 8), and 12 day workers participated during a voyage from the United Kingdom to Antarctica. They kept daily sleep diaries and wore wrist monitors for continuous recording of activity. Sleep parameters were derived from activity using the manufacturer's software and analyzed by repeated-measures ANOVA using SAS 8.2. Sequential urine samples were collected for 48 h weekly for 6-sulphatoxymelatonin measurement as an index of circadian rhythm timing. Individuals working watches of 1200-1600/2400-0400 h and 1600-2000/0400-0800 h had 2 sleeps daily, analyzed separately as main sleep (longest) and 2nd sleep. Main sleep duration was shorter in watch keepers than in day workers (p < 0.0001). Objective sleep quality was significantly compromised in rotaters compared to both day workers and fixed watch keepers, the most striking comparisons being sleep efficiency (percentage desired sleep time spent sleeping) main sleep (p < 0.0001) and sleep fragmentation (an index of restlessness) main sleep (p < 0.0001). The 2nd sleep was substantially less efficient than was the main sleep (p < 0.0001) for all watch keepers. There were few significant differences in sleep between the different watches in rotating watch keepers. Circadian timing remained constant in day workers. Timing of the 6-sulphatoxymelatonin rhythm was later for the watch of 1200-1600/2400-0400 h than for all others (1200-1600/2400-0400 h, 5.90 +/- 0.85 h; 1600-2000/0400-0800 h, 1.5 +/- 0.64 h; 0800-1200/ 2000-2400 h, 2.72 +/- 0.76 h; days, 2.09 +/- 0.68 h [decimal hours, mean +/- SEM]: ANOVA, p < 0.01). This study identifies weekly changes in watch time as a cause of poor sleep in watch keepers. The most likely mechanism is the inability of the internal clock to adapt rapidly to abrupt changes in schedule.     

Managing and mining protein crystallization data

The crystallization of macromolecules remains a major bottleneck in structural biology. The routine screening of more than one thousand crystallization conditions and subsequent optimization by fine screening presents a challenge to conventional laboratory notebook keeping. In addition, the development of high-throughput robotic crystallization and imaging systems presents a pressing need for low-cost laboratory information management system (LIMS). Here we describe CLIMS2, a crystallization LIMS that features a simple, user-friendly graphical interface, allowing the storage, management, retrieval and mining of crystallization data. The CLIMS2 executable and documentation is freely available at http://clims.med.monash.edu.au.     

A specific secretion system mediates PPE41 transport in pathogenic mycobacteria

Mycobacterial genomes contain two unique gene families, the so-called PE and PPE gene families, which are highly expanded in the pathogenic members of this genus. Here we report that one of the PPE proteins, i.e. PPE41, is secreted by pathogenic mycobacteria, both in culture and in infected macrophages. As PPE41 lacks a signal sequence a dedicated secretion system must be involved. A single gene was identified in Mycobacterium marinum that showed strongly reduced PPE41 secretion. This gene was located in a gene cluster whose predicted proteins encode components of an ESAT-6-like secretion system. This cluster, designated ESX-5, is conserved in various pathogenic mycobacteria, but not in the saprophytic species Mycobacterium smegmatis. Therefore, different regions of this cluster were introduced in M. smegmatis. Only introduction of the complete ESX-5 locus resulted in efficient secretion of heterologously expressed PPE41. This PPE secretion system is also involved in the virulence of pathogenic mycobacteria, as the ESX-5 mutant of M. marinum was affected in spreading to uninfected macrophages.     

Biodiversity management of fens and fen meadows by grazing,cutting and burning

Question: Can the biodiversity of fens in Europe and North America be maintained through the use of grazing (especially cattle grazing), fire, and/or cutting? Location: European and North American fens. Methods: This paper is a review of the literature on the effects of grazing, fire and cutting on fens, to explore the relationship between management and biodiversity in fens. Results: A reduction of cattle grazing, mowing and burning in fens has led to a reduction in biodiversity in fens. The vegetation of abandoned fens shifts to trees and shrubs after 10–15 years, which shade the smaller and rarer species of these wetlands. While careful use of fire is used to manage fens in North America, it is not widely used in European fens, perhaps because the peat of drained fens may catch fire. Cattle grazing cannot be considered a natural disturbance in North America, since cattle did not evolve on that continent. In Europe, cattle do not generally graze in unaltered fens, but they do use slightly drained fen meadows. Conclusions: Three approaches have been used to control the dominance of tall woody and herbaceous species in abandoned fens, including the re‐introduction of cattle, mowing, and burning. Overgrazing results in a permanent reduction in biodiversity, therefore cattle re‐introduction must be approached cautiously. In Europe, but not in North America, mowing has been an important management tool, and mowing has been successful in maintaining species richness, particularly in fens that have been mowed annually for centuries. Fire has been the most common and successful management tool in North America although it is not effective in removing shrubs that have become large. Because the problems and solutions are similar, the literature of both European and North American fen management can be analyzed to better assess the management of fens on both continents. Many management questions require further study and these are listed in the paper.     

Detection of nucleotide binding to Na,K-ATPase in an aqueous membrane suspension by 13C cross-polarization magic-angle spinning NMR spectroscopy

Binding of uniformly (13)C labelled ATP to Na,K-ATPase was studied by (13)C cross-polarization magic-angle spinning (CP-MAS) NMR. In the presence of 30 mM Na(+) , and with sample- and time-averaging, NMR spectra obtained at 4 degrees C exhibited several resonances for the bound nucleotide. Chemical shifts suggested that site-specific changes in the micro-environment or conformation of the nucleotide occurred in the high affinity binding site. These experiments permit further studies of nucleotide dynamics, structure and binding under physiologically relevant conditions.     

Specific Chaperones for the Type VII Protein Secretion Pathway

Mycobacteria use the dedicated type VII protein secretion systems ESX-1 and ESX-5 to secrete virulence factors across their highly hydrophobic cell envelope. The substrates of these systems include the large mycobacterial PE and PPE protein families, which are named after their characteristic Pro-Glu and Pro-Pro-Glu motifs. Pathogenic mycobacteria secrete large numbers of PE/PPE proteins via the major export pathway, ESX-5. In addition, a few PE/PPE proteins have been shown to be exported by ESX-1. It is not known how ESX-1 and ESX-5 recognize their cognate PE/PPE substrates. In this work, we investigated the function of the cytosolic protein EspG(5), which is essential for ESX-5-mediated secretion in Mycobacterium marinum, but for which the role in secretion is not known. By performing protein co-purifications, we show that EspG(5) interacts with several PPE proteins and a PE/PPE complex that is secreted by ESX-5, but not with the unrelated ESX-5 substrate EsxN or with PE/PPE proteins secreted by ESX-1. Conversely, the ESX-1 paralogue EspG(1) interacted with a PE/PPE couple secreted by ESX-1, but not with PE/PPE substrates of ESX-5. Furthermore, structural analysis of the complex formed by EspG(5) and PE/PPE indicates that these proteins interact in a 1:1:1 ratio. In conclusion, our study shows that EspG(5) and EspG(1) interact specifically with PE/PPE proteins that are secreted via their own ESX systems and suggests that EspG proteins are specific chaperones for the type VII pathway.     

Complete genome sequence of Mycobacterium phlei type strain RIVM601174

Mycobacterium phlei is a rapidly growing nontuberculous Mycobacterium species that is typically nonpathogenic, with few reported cases of human disease. Here we report the whole genome sequence of M. phlei type strain RIVM601174.     

Disruption of the ESX-5 system of Mycobacterium tuberculosis causes loss of PPE protein secretion, reduction of cell wall integrity and strong attenuation

The chromosome of Mycobacterium tuberculosis encodes five type VII secretion systems (ESX-1-ESX-5). While the role of the ESX-1 and ESX-3 systems in M. tuberculosis has been elucidated, predictions for the function of the ESX-5 system came from data obtained in Mycobacterium marinum, where it transports PPE and PE_PGRS proteins and modulates innate immune responses. To define the role of the ESX-5 system in M. tuberculosis, in this study, we have constructed five M. tuberculosis H37Rv ESX-5 knockout/deletion mutants, inactivating eccA(5), eccD(5), rv1794 and esxM genes or the ppe25-pe19 region. Whereas the Mtbrv1794ko displayed no obvious phenotype, the other four mutants showed defects in secretion of the ESX-5-encoded EsxN and PPE41, a representative member of the large PPE protein family. Strikingly, the MtbeccD(5) ko mutant also showed enhanced sensitivity to detergents and hydrophilic antibiotics. When the virulence of the five mutants was evaluated, the MtbeccD(5) ko and MtbΔppe25-pe19 mutants were found attenuated both in macrophages and in the severe combined immune-deficient mouse infection model. Altogether these findings indicate an essential role of ESX-5 for transport of PPE proteins, cell wall integrity and full virulence of M. tuberculosis, thereby opening interesting new perspectives for the study of this human pathogen.