What traits are carried on mobile genetic elements, and why?

Although similar to any other organism, prokaryotes can transfer genes vertically from mother cell to daughter cell, they can also exchange certain genes horizontally. Genes can move within and between genomes at fast rates because of mobile genetic elements (MGEs). Although mobile elements are fundamentally self-interested entities, and thus replicate for their own gain, they frequently carry genes beneficial for their hosts and/or the neighbours of their hosts. Many genes that are carried by mobile elements code for traits that are expressed outside of the cell. Such traits are involved in bacterial sociality, such as the production of public goods, which benefit a cell's neighbours, or the production of bacteriocins, which harm a cell's neighbours. In this study we review the patterns that are emerging in the types of genes carried by mobile elements, and discuss the evolutionary and ecological conditions under which mobile elements evolve to carry their peculiar mix of parasitic, beneficial and cooperative genes

Process evaluation for complex interventions in primary care: understanding trials using the normalization process model

Background: the Normalization Process Model is a conceptual tool intended to assist in understanding the factors that affect implementation processes in clinical trials and other evaluations of complex interventions. It focuses on the ways that the implementation of complex interventions is shaped by problems of workability and integration.Method: in this paper the model is applied to two different complex trials: (i) the delivery of problem solving therapies for psychosocial distress, and (ii) the delivery of nurse-led clinics for heart failure treatment in primary care.Results: application of the model shows how process evaluations need to focus on more than the immediate contexts in which trial outcomes are generated. Problems relating to intervention workability and integration also need to be understood. The model may be used effectively to explain the implementation process in trials of complex interventions.Conclusion: the model invites evaluators to attend equally to considering how a complex intervention interacts with existing patterns of service organization, professional practice, and professional-patient interaction. The justification for this may be found in the abundance of reports of clinical effectiveness for interventions that have little hope of being implemented in real healthcare setting

An accurate and efficient identification of children with psychosocial problems by means of computerized adaptive testing

Background: Questionnaires used by health services to identify children with psychosocial problems are often rather short. The psychometric properties of such short questionnaires are mostly less than needed for an accurate distinction between children with and without problems. We aimed to assess whether a short Computerized Adaptive Test (CAT) can overcome the weaknesses of short written questionnaires when identifying children with psychosocial problems. Method: We used a Dutch national data set obtained from parents of children invited for a routine health examination by Preventive Child Healthcare with 205 items on behavioral and emotional problems (n = 2,041, response 84%). In a random subsample we determined which items met the requirements of an Item Response Theory (IRT) model to a sufficient degree. Using those items, item parameters necessary for a CAT were calculated and a cut-off point was defined. In the remaining subsample we determined the validity and efficiency of a Computerized Adaptive Test using simulation techniques, with current treatment status and a clinical score on the Total Problem Scale (TPS) of the Child Behavior Checklist as criteria. Results: Out of 205 items available 190 sufficiently met the criteria of the underlying IRT model. For 90% of the children a score above or below cut-off point could be determined with 95% accuracy. The mean number of items needed to achieve this was 12. Sensitivity and specificity with the TPS as a criterion were 0.89 and 0.91, respectively. Conclusion: An IRT-based CAT is a very promising option for the identification of psychosocial problems in children, as it can lead to an efficient, yet high-quality identification. The results of our simulation study need to be replicated in a real-life administration of this CAT

Total Shoulder Arthroplasty

Shoulder arthroplasty has been the subject of marked advances over the last few years. Modern implants provide a wide range of options, including resurfacing of the humeral head, anatomic hemiarthroplasty, total shoulder arthroplasty, reverse shoulder arthroplasty and trauma-specific implants for fractures and nonunions. Most humeral components achieve successful long-term fixation without bone cement. Cemented all-polyethylene glenoid components remain the standard for anatomic total shoulder arthroplasty. The results of shoulder arthroplasty vary depending on the underlying diagnosis, the condition of the soft-tissues, and the type of reconstruction. Total shoulder arthroplasty seems to provide the best outcome for patients with osteoarthritis and inflammatory arthropathy. The outcome of hemiarthroplasty for proximal humerus fractures is somewhat unpredictable, though it seems to have improved with the use of fracture-specific designs, more attention to tuberosity repair, and the selective use of reverse arthroplasty, as well as a shift in indications towards internal fixation. Reverse shoulder arthroplasty has become extremely popular for patients with cuff-tear arthropathy, and its indications have been expanded to the field of revision surgery. Overall, shoulder arthroplasty is a very successful procedure with predictable pain relief and substantial improvements in motion and function

Expression and functional activity of nucleoside transporters in human choroid plexus

Abstract Background Human equilibrative nucleoside transporters (hENTs) 1-3 and human concentrative nucleoside transporters (hCNTs) 1-3 in the human choroid plexus (hCP) play a role in the homeostasis of adenosine and other naturally occurring nucleosides in the brain; in addition, hENT1, hENT2 and hCNT3 mediate membrane transport of nucleoside reverse transcriptase inhibitors that could be used to treat HIV infection, 3'-azido-3'-deoxythymidine, 2'3'-dideoxycytidine and 2'3'-dideoxyinosine. This study aimed to explore the expression levels and functional activities of hENTs 1-3 and hCNTs 1-3 in human choroid plexus. Methods Freshly-isolated pieces of lateral ventricle hCP, removed for various clinical reasons during neurosurgery, were obtained under Local Ethics Committee approval. Quantification of mRNAs that encoded hENTs and hCNTs was performed by the hydrolysis probes-based reverse transcription real time-polymerase chain reaction (RT-qPCR); for each gene of interest and for 18 S ribosomal RNA, which was an endogenous control, the efficiency of PCR reaction (E) and the quantification cycle (Cq) were calculated. The uptake of [3H]inosine by the choroid plexus pieces was investigated to explore the functional activity of hENTs and hCNTs in the hCP. Results RT-qPCR revealed that the mRNA encoding the intracellularly located transporter hENT3 was the most abundant, with E-Cq value being only about 40 fold less that the E-Cq value for 18 S ribosomal RNA; mRNAs encoding hENT1, hENT2 and hCNT3 were much less abundant than mRNA for the hENT3, while mRNAs encoding hCNT1 and hCNT2 were of very low abundance and not detectable. Uptake of [3H]inosine by the CP samples was linear and consisted of an Na+-dependent component, which was probably mediated by hCNT3, and Na+-independent component, mediated by hENTs. The latter component was not sensitive to inhibition by S-(4-nitrobenzyl)-6-thioinosine (NBMPR), when used at a concentration of 0.5 μM, a finding that excluded the involvement of hENT1, but it was very substantially inhibited by 10 μM NBMPR, a finding that suggested the involvement of hENT2 in uptake. Conclusion Transcripts for hENT1-3 and hCNT3 were detected in human CP; mRNA for hENT3, an intracellularly located nucleoside transporter, was the most abundant. Human CP took up radiolabelled inosine by both concentrative and equilibrative processes. Concentrative uptake was probably mediated by hCNT3; the equilibrative uptake was mediated only by hENT2. The hENT1 transport activity was absent, which could suggest either that this protein was absent in the CP cells or that it was confined to the basolateral side of the CP epithelium.</p

Single Molecule In Vivo Analysis of Toll-Like Receptor 9 and CpG DNA Interaction

Toll-like receptor 9 (TLR9) activates the innate immune system in response to oligonucleotides rich in CpG whereas DNA lacking CpG could inhibit its activation. However, the mechanism of how TLR9 interacts with nucleic acid and becomes activated in live cells is not well understood. Here, we report on the successful implementation of single molecule tools, constituting fluorescence correlation/cross-correlation spectroscopy (FCS and FCCS) and photon count histogram (PCH) with fluorescence lifetime imaging (FLIM) to study the interaction of TLR9-GFP with Cy5 labeled oligonucleotide containing CpG or lacking CpG in live HEK 293 cells. Our findings show that i) TLR9 predominantly forms homodimers (80%) before binding to a ligand and further addition of CpG or non CpG DNA does not necessarily increase the proportion of TLR9 dimers, ii) CpG DNA has a lower dissociation constant (62 nM±9 nM) compared to non CpG DNA (153 nM±26 nM) upon binding to TLR9, suggesting that a motif specific binding affinity of TLR9 could be an important factor in instituting a conformational change-dependant activation, and iii) both CpG and non CpG DNA binds to TLR9 with a 1∶2 stoichiometry in vivo. Collectively, through our findings we establish an in vivo model of TLR9 binding and activation by CpG DNA using single molecule fluorescence techniques for single cell studies

Cyclic AMP signalling pathways in the regulation of uterine relaxation

Studying the mechanism(s) of uterine relaxation is important and will be helpful in the prevention of obstetric difficulties such as preterm labour, which remains a major cause of perinatal mortality and morbidity. Multiple signalling pathways regulate the balance between maintaining relative uterine quiescence during gestation, and the transition to the contractile state at the onset of parturition. Elevation of intracellular cyclic AMP promotes myometrial relaxation, and thus quiescence, via effects on multiple intracellular targets including calcium channels, potassium channels and myosin light chain kinase. A complete understanding of cAMP regulatory pathways (synthesis and hydrolysis) would assist in the development of better tocolytics to delay or inhibit preterm labour. Here we review the enzymes involved in cAMP homoeostasis (adenylyl cyclases and phosphodiesterases) and possible myometrial substrates for the cAMP dependent protein kinase. We must emphasise the need to identify novel pharmacological targets in human pregnant myometrium to achieve safe and selective uterine relaxation when this is indicated in preterm labour or other obstetric complications

Disulfiram modulated ROS–MAPK and NFκB pathways and targeted breast cancer cells with cancer stem cell-like properties

BACKGROUND: Previous studies indicate that disulfiram (DS), an anti-alcoholism drug, is cytotoxic to cancer cell lines and reverses anticancer drug resistance. Cancer stem cells (CSCs) are the major cause of chemoresistance leading to the failure of cancer chemotherapy. This study intended to examine the effect of DS on breast cancer stem cells (BCSCs). METHODS: The effect of DS on BC cell lines and BCSCs was determined by MTT, western blot, CSCs culture and CSCs marker analysis. RESULTS: Disulfiram was highly toxic to BC cell lines in vitro in a copper (Cu)-dependent manner. In Cu-containing medium (1 mu M), the IC50 concentrations of DS in BC cell lines were 200-500 nM. Disulfiram/copper significantly enhanced (3.7-15.5-fold) cytotoxicity of paclitaxel (PAC). Combination index isobologram analysis demonstrated a synergistic effect between DS/Cu and PAC. The increased Bax and Bcl2 protein expression ratio indicated that intrinsic apoptotic pathway may be involved in DS/Cu-induced apoptosis. Clonogenic assay showed DS/Cu-inhibited clonogenicity of BC cells. Mammosphere formation and the ALDH1(+VE) and CD24(Low)/CD44(High) CSCs population in mammospheres were significantly inhibited by exposure to DS/Cu for 24 h. Disulfiram/copper induced reactive oxygen species (ROS) generation and activated its downstream apoptosis-related cJun N-terminal kinase and p38 MAPK pathways. Meanwhile, the constitutive NF kappa B activity in BC cell lines was inhibited by DS/Cu. CONCLUSION: Disulfiram/copper inhibited BCSCs and enhanced cytotoxicity of PAC in BC cell lines. This may be caused by simultaneous induction of ROS and inhibition of NF kappa B. British Journal of Cancer (2011) 104, 1564-1574. doi: 10.1038/bjc.2011.126 www.bjcancer.com Published online 12 April 2011 (C) 2011 Cancer Research U

Obscured phylogeny and possible recombinational dormancy in Escherichia coli

<p>Abstract</p> <p>Background</p> <p><it>Escherichia coli </it>is one of the best studied organisms in all of biology, but its phylogenetic structure has been difficult to resolve with current data and analytical techniques. We analyzed single nucleotide polymorphisms in chromosomes of representative strains to reconstruct the topology of its emergence.</p> <p>Results</p> <p>The phylogeny of <it>E. coli </it>varies according to the segment of chromosome analyzed. Recombination between extant <it>E. coli </it>groups is largely limited to only three intergroup pairings.</p> <p>Conclusions</p> <p>Segment-dependent phylogenies most likely are legacies of a complex recombination history. However, <it>E. coli </it>are now in an epoch in which they no longer broadly share DNA. Using the definition of species as organisms that freely exchange genetic material, this recombinational dormancy could reflect either the end of <it>E. coli </it>as a species, or herald the coalescence of <it>E. coli </it>groups into new species.</p

Immune Subversion and Quorum-Sensing Shape the Variation in Infectious Dose among Bacterial Pathogens

Many studies have been devoted to understand the mechanisms used by pathogenic bacteria to exploit human hosts. These mechanisms are very diverse in the detail, but share commonalities whose quantification should enlighten the evolution of virulence from both a molecular and an ecological perspective. We mined the literature for experimental data on infectious dose of bacterial pathogens in humans (ID50) and also for traits with which ID50 might be associated. These compilations were checked and complemented with genome analyses. We observed that ID50 varies in a continuous way by over 10 orders of magnitude. Low ID50 values are very strongly associated with the capacity of the bacteria to kill professional phagocytes or to survive in the intracellular milieu of these cells. Inversely, high ID50 values are associated with motile and fast-growing bacteria that use quorum-sensing based regulation of virulence factors expression. Infectious dose is not associated with genome size and shows insignificant phylogenetic inertia, in line with frequent virulence shifts associated with the horizontal gene transfer of a small number of virulence factors. Contrary to previous proposals, infectious dose shows little dependence on contact-dependent secretion systems and on the natural route of exposure. When all variables are combined, immune subversion and quorum-sensing are sufficient to explain two thirds of the variance in infectious dose. Our results show the key role of immune subversion in effective human infection by small bacterial populations. They also suggest that cooperative processes might be important for successful infection by bacteria with high ID50. Our results suggest that trade-offs between selection for population growth-related traits and selection for the ability to subvert the immune system shape bacterial infectiousness. Understanding these trade-offs provides guidelines to study the evolution of virulence and in particular the micro-evolutionary paths of emerging pathogens