Using Active Learning to Teach Concepts and Methods in Quantitative Biology.
This article provides a summary of the ideas discussed at the 2015 Annual Meeting of the Society for Integrative and Comparative Biology society-wide symposium on Leading Students and Faculty to Quantitative Biology through Active Learning. It also includes a brief review of the recent advancements in incorporating active learning approaches into quantitative biology classrooms. We begin with an overview of recent literature that shows that active learning can improve students' outcomes in Science, Technology, Engineering and Math Education disciplines. We then discuss how this approach can be particularly useful when teaching topics in quantitative biology. Next, we describe some of the recent initiatives to develop hands-on activities in quantitative biology at both the graduate and the undergraduate levels. Throughout the article we provide resources for educators who wish to integrate active learning and technology into their classrooms.


Participation of DNA polymerase zeta in replication of undamaged DNA in Saccharomyces cerevisiae.
Translesion synthesis DNA polymerases contribute to DNA damage tolerance by mediating replication of damaged templates. Due to the low fidelity of these enzymes, lesion bypass is often mutagenic. We have previously shown that, in Saccharomyces cerevisiae, the contribution of the error-prone DNA polymerase zeta (Polzeta) to replication and mutagenesis is greatly enhanced if the normal replisome is defective due to mutations in replication genes. Here we present evidence that this defective-replisome-induced mutagenesis (DRIM) results from the participation of Polzeta in the copying of undamaged DNA rather than from mutagenic lesion bypass. First, DRIM is not elevated in strains that have a high level of endogenous DNA lesions due to defects in nucleotide excision repair or base excision repair pathways. Second, DRIM remains unchanged when the level of endogenous oxidative DNA damage is decreased by using anaerobic growth conditions. Third, analysis of the spectrum of mutations occurring during DRIM reveals the characteristic error signature seen during replication of undamaged DNA by Polzeta in vitro. These results extend earlier findings in Escherichia coli indicating that Y-family DNA polymerases can contribute to the copying of undamaged DNA. We also show that exposure of wild-type yeast cells to the replication inhibitor hydroxyurea causes a Polzeta-dependent increase in mutagenesis. This suggests that DRIM represents a response to replication impediment per se rather than to specific defects in the replisome components.


NBD-based colorimetric and fluorescent turn-on probes for hydrogen sulfide.
Hydrogen sulfide (H2S) is an important endogenous signalling molecule and also an important environmental target for detection. New reaction-based colorimetric and fluorescent turn-on probes based on selective thiolyling of NBD (7-nitro-1,2,3-benzoxadiazole) ether were explored for sensing of H2S in aqueous buffer. The syntheses of both probes are simple and quite straightforward. The probes are highly sensitive and selective toward H2S over other biologically relevant species. Probe 1 can be used to directly visualize H2S by the naked eye and shows more than 1000-fold fluorescence increase upon reaction with H2S. Probe 2 is a near-infrared fluorescent sensor for H2S at physiological pH.


Academic Performance in Primary School Children With Common Emotional and Behavioral Problems.
Many emotional and behavioral problems first emerge in primary school and are the forerunners of mental health problems occurring in adolescence. However, the extent that these problems may be associated with academic failure has been explored less. We aimed to quantify the association between emotional and behavioral problems with academic performance. A stratified random sample of 8- to 9-year-olds (N = 1239) were recruited from schools in Australia. Data linkage was performed with a national assessment of academic performance to assess reading and numeracy. Parent report assessed emotional and behavioral problems with students dichotomized into "borderline/abnormal" and "normal" categories. One in 5 grade 3 students fell in the "borderline/abnormal" category. Boys with total difficulties (β = -47.8, 95% CI: -62.8 to -32.8), conduct problems, and peer problems scored lower on reading. Numeracy scores were lower in boys with total difficulties (β = -37.7, 95% CI: -53.9 to -21.5) and emotional symptoms. Children with hyperactivity/inattention scored lower in numeracy. Girls with peer problems scored lower in numeracy. Boys with emotional and behavioral problems in mid-primary school were 12 months behind their peers. Children with emotional and behavioral problems are at high risk for academic failure, and this risk is evident in mid-primary school.


Identification and characterization of two parathyroid hormone-like molecules in zebrafish.
Zebrafish (Danio rerio) have receptors homologous to the human PTH (hPTH)/PTHrP receptor (PTH1R) and PTH-2 receptor (PTH2R) and an additional receptor (PTH3R) with high homology to the PTH1R. To find natural ligands for zPTH1R and zPTH3R, we searched the zebrafish genomic database and discovered two distinct regions that, when translated (zPTH1 and zPTH2), showed high homology to hPTH. Isolation of cDNAs and determination of the intron/exon boundaries revealed genomic structures which were similar to known PTHs. Peptides consisting of the first 34 amino acids after the pre- and prosequences of the zebrafish PTHs (zPTHs) were synthesized and were shown to be fully active at the hPTH1R. zPTH2(1-34) was, however, approximately 30-fold less potent at the zPTH1R than hPTH(1-34), hPTHrP(1-36), and zPTH1(1-34). When tested with zPTH3R, zPTH1(1-34) and hPTHrP(1-36) showed similar potencies, whereas the potency of zPTH2(1-34) was moderately (3-fold) reduced. To determine whether other fishes have multiple PTHs, we searched the genomic database of the Japanese pufferfish (Takifugu rubripes) and identified zPTH1 and zPTH2 homologs. Phylogenetic analysis showed that PTHs from zebrafish and pufferfish are more closely related to each other than to known mammalian PTH homologs or to PTHrP and tuberoinfundibular peptide of 39 residues. This is consistent with evolution of two teleost PTH-like peptides occurring after the evolutionary divergence between fishes and mammals. Overall, the PTH system appears more complex in fishes than in mammals, providing evidence of continued evolution in nontetrapod species. The availability of multiple forms of fish PTH and their receptors provide additional tools for PTH ligand/receptor structure-function studies.


Emergency department attendance by children at risk of abuse.
Frequency of emergency department (ED) attendance has long been thought to be a risk factor for child abuse. The aim of this study was to test this assumption by comparing the ED attendances of at-risk children (before being placed on a child protection register) with the attendances of an age-matched control group (before an index attendance) Method: A group of 220 children (aged 0-12 years inclusive) were identified from the two child protection registers in the Shropshire area in 2006. The ED attendances of these children in the 2 years before registration were identified using the computer records of the two local EDs. A control group of 150 children for each year of age (0-12 years inclusive) was then identified from ED attendances between October and December 2006. The attendances of these children in the 2 years before this index attendance were obtained. The data for these two groups of children were then compared. The at-risk children did not attend the EDs more frequently than did the children in the control group. The identification of children who attend EDs frequently may be useful for other reasons but is unlikely to be an effective way to detect child abuse.


Apoptotic cells activate AMP-activated protein kinase (AMPK) and inhibit epithelial cell growth without change in intracellular energy stores.
Apoptosis plays an indispensable role in the maintenance and development of tissues. We have shown that receptor-mediated recognition of apoptotic target cells by viable kidney proximal tubular epithelial cells (PTECs) inhibits the proliferation and survival of PTECs. Here, we examined the effect of apoptotic targets on PTEC cell growth (cell size during G1 phase of the cell cycle). Using a cell culture model, we show that apoptotic cells potently activate AMP-activated protein kinase (AMPK), a highly sensitive sensor of intracellular energy stores. AMPK activation leads to decreased activity of its downstream target, ribosomal protein p70 S6 kinase (p70S6K), and concomitant inhibition of cell growth. Importantly, these events occur without detectable change in intracellular levels of AMP, ADP, or ATP. Inhibition of AMPK, either pharmacologically by compound C or molecularly by shRNA, diminishes the effects of apoptotic targets and largely restores p70S6K activity and cell size to normal levels. Apoptotic targets also inhibit Akt, a second signaling pathway regulating cell growth. Expression of a constitutively active Akt construct partially relieved cell growth inhibition but was less effective than inhibition of AMPK. Inhibition of cell growth by apoptotic targets is dependent on physical interaction between apoptotic targets and PTECs but independent of phagocytosis. We conclude that receptor-mediated recognition of apoptotic targets mimics the effects of intracellular energy depletion, activating AMPK and inhibiting cell growth. By acting as sentinels of environmental change, apoptotic death may enable nearby viable cells, especially nonmigratory epithelial cells, to monitor and adapt to local stresses.


Dual coordination modes of ethylene-linked NP2 ligands in cobalt(II) and nickel(II) iodides.
Here we report the syntheses and crystal structures of a series of cobalt(II) and nickel(II) complexes derived from (R)NP2 ligands (where R = OMe(Bz), H(Bz), Br(Bz), Ph) bearing ethylene linkers between a single N and two P donors. The Co(II) complexes generally adopt a tetrahedral configuration of general formula [(NP2)Co(I)(2)], wherein the two phosphorus donors are bound to the metal center but the central N-donor remains unbound. We have found one case of structural isomerism within a single crystal structure. The Co(II) complex derived from (Bz)NP2 displays dual coordination modes: one in the tetrahedral complex [((Bz)NP2)Co(I)(2)]; and the other in a square pyramidal variant, [((Bz)NP2)Co(I)(2)]. In contrast, the Ni(II) complexes adopt a square planar geometry in which the P(Et)N(Et)P donors in the ligand backbone are coordinated to the metal center, resulting in cationic species of formula [((R)NP2)Ni(I)](+) with iodide as counterion. All Ni(II) complexes exhibit sharp (1)H and (31)P spectra in the diamagnetic region. The Co(II) complexes are high-spin (S = 3/2) in the solid state as determined by SQUID measurements from 4 to 300 K. Solution electron paramagnetic resonance (EPR) experiments reveal a high-spin/low-spin Co(II) equilibrium that is dependent on solvent and ligand substituent.


Optical correlator production system neural net.
A new neural net is described that can easily and cost-effectively accommodate multiple objects in the field of view in parallel. The use of a correlator achieves shift invariance and accommodates multiple objects in parallel. Distortion-invariant filters provide aspect-invariant distortion. Symbolic encoding, the use of generic object parts, and a production system neural net allow large class problems to be addressed. Optical laboratory data on the production system inputs are provided and emphasized. Test data assume binary inputs, although analog (probability) input neurons are possible.


Thromboxane synthesis inhibitor in a beagle pup model of perinatal asphyxia.
During perinatal asphyxia, cerebral blood flow is markedly reduced in the gray and white matter of the telencephalon. Since previous work has implicated prostaglandins in the control of blood flow, we tested the hypothesis that a thromboxane synthesis inhibitor would improve cerebral blood flow and blunt the metabolic alterations that accompany asphyxia. Forty-three newborn beagles 2-7 days old were anesthetized, ventilated, and randomized to insult (5 minutes of asphyxia) or no insult and received treatment with either the thromboxane synthesis inhibitor CGS 13080 (CIBA-GEIGY Corp.) (0.06 mg/kg/hr i.v. infusion) or saline. Cerebral blood flow was measured in 25 pups. Pups received treatment 30 minutes before insult or no insult. In pups randomized to insult and receiving saline, cerebral blood flow increased during insult in the medulla but decreased elsewhere. Pups randomized to insult and treated with thromboxane synthesis inhibitor had increased cerebral blood flow during insult in all cerebral regions studied. In addition, these pups experienced a significantly higher incidence of intraventricular hemorrhage than did pups randomized to insult and receiving saline. In other experiments with 18 pups, brain extracts were prepared for proton nuclear magnetic resonance spectral analysis of high-energy phosphorylated compounds and lactate levels. In pups exposed to insult and receiving saline, mean +/- SD phosphocreatine concentration fell from 1.9 +/- 0.1 to 0.4 +/- 0.1 mmol/kg, lactate concentration increased from 2.0 +/- 0.5 to 3.3 +/- 0.4 mmol/kg, and the calculated pH fell 0.8 units. There were no differences between groups.(ABSTRACT TRUNCATED AT 250 WORDS)


Failure to vaccinate or failure of vaccine? Effectiveness of the 23-valent pneumococcal polysaccharide vaccine program in Indigenous adults in the Northern Territory of Australia.
Over the last decade, there has been no discernible reduction in Invasive Pneumococcal Disease (IPD) amongst Indigenous adults in the Northern Territory (NT) of Australia, despite increasing vaccination coverage. We examined the utility of two common methods, the screening method and the indirect method, to determine the 23-valent pneumococcal polysaccharide vaccine effectiveness (VE) in prevention of IPD amongst Indigenous adults in this setting. VE was calculated for the period 2001-2005 across two distinct geographical areas where the disease burden was known to differ. VE against vaccine-type IPD was 3.4% (95% CI -43, 35) for the NT. However, population vaccination coverage varied widely according to geographical region and where this was within the range appropriate for the use of the screening method, VE was within the expected range (67.2%, 95% CI 47, 80). VE according to the indirect cohort appeared unreliable in this setting due to the analysis being based on a very limited number of non-vaccine-type IPD cases. Surveillance based estimates of VE such as these need to be considered with caution, but the results suggest failure to vaccinate is the most likely reason vaccine-type IPD has not reduced in this setting.


Regenerating cochlear hair cells: quo vadis stem cell.
Many elderly people worldwide lose the neurosensory part of their ear and turn deaf. Cochlear implants to restore some hearing after neurosensory hearing loss are, at present, the only therapy for these people. In contrast to this therapy, replacement of hair cells via stem cell therapies holds the promise for a cure. We review here current insights into embryonic, adult, and inducible stem cells that might provide cells for seeding the cochlea with the hope of new hair cell formation. We propose a two-step approach using a first set of transcription factors to enhance the generation of inducible pluripotent stem (iPS) cells and a second set of factors to initiate the differentiation of hair cells. Recent evidence regarding ear development and stem cell research strongly suggest that microRNAs will be an important new regulatory factor in both iPS cell formation and differentiation to reprogram cells into hair cells. In addition, we highlight currently insurmountable obstacles to the successful transformation of stem cells into hair cell precursors and their injection into the cochlear canal to replace lost hair cells.


A single heterochronic blood exchange reveals rapid inhibition of multiple tissues by old blood.
Heterochronic parabiosis rejuvenates the performance of old tissue stem cells at some expense to the young, but whether this is through shared circulation or shared organs is unclear. Here we show that heterochronic blood exchange between young and old mice without sharing other organs, affects tissues within a few days, and leads to different outcomes than heterochronic parabiosis. Investigating muscle, liver and brain hippocampus, in the presence or absence of muscle injury, we find that, in many cases, the inhibitory effects of old blood are more pronounced than the benefits of young, and that peripheral tissue injury compounds the negative effects. We also explore mechanistic explanations, including the role of B2M and TGF-beta. We conclude that, compared with heterochronic parabiosis, heterochronic blood exchange in small animals is less invasive and enables better-controlled studies with more immediate translation to therapies for humans.


Inappropriate prescribing for older people admitted to an intermediate-care nursing home unit and hospital wards.
To identify inappropriate prescribing among older patients on admission to and discharge from an intermediate-care nursing home unit and hospital wards, and to compare changes during stay within and between these groups. Observational study. Altogether 400 community-dwelling people aged ≥ 70 years, on consecutive emergency admittance to hospital wards of internal medicine and orthopaedic surgery, were randomized to an intermediate-care nursing home unit or hospital wards; 290 (157 at the intermediate-care nursing home unit and 133 in hospital wards) were eligible for this sub-study. Prevalence on admission and discharge of potentially inappropriate medications (Norwegian general practice [NORGEP] criteria) and drug-drug interactions; changes during stay. The mean (SD) age was 84.7 (6.2) years; 71% were women. From admission to discharge, the mean numbers of drugs prescribed per person increased from 6.0 (3.3) to 9.3 (3.8), p < 0.01. The prevalence of potentially inappropriate medications increased from 24% to 35%, p < 0.01; concomitant use of ≥ 3 psychotropic/opioid drugs and drug combinations including non-steroid anti-inflammatory drugs (NSAIDs) increased significantly. Serious drug-drug interactions were scarce both on admission and discharge (0.7%). Inappropriate prescribing was prevalent among older people acutely admitted to hospital, and the prevalence was not reduced during stay at an intermediate-care nursing home unit specially designed for these patients.


Altered telomeres in tumors with ATRX and DAXX mutations.
The proteins encoded by ATRX and DAXX participate in chromatin remodeling at telomeres and other genomic sites. Because inactivating mutations of these genes are common in human pancreatic neuroendocrine tumors (PanNETs), we examined the telomere status of these tumors. We found that 61% of PanNETs displayed abnormal telomeres that are characteristic of a telomerase-independent telomere maintenance mechanism termed ALT (alternative lengthening of telomeres). All of the PanNETs exhibiting these abnormal telomeres had ATRX or DAXX mutations or loss of nuclear ATRX or DAXX protein. ATRX mutations also correlate with abnormal telomeres in tumors of the central nervous system. These data suggest that an alternative telomere maintenance function may operate in human tumors with alterations in the ATRX or DAXX genes.


A Review of Temperature, pH, and Other Factors that Influence the Survival of Salmonella in Mayonnaise and Other Raw Egg Products.
Salmonellosis is one of the main causes of foodborne illnesses worldwide, with outbreaks predominately linked to contamination of eggs and raw egg products, such as mayonnaise. This review explores previous studies that have investigated Salmonella control mechanisms utilized in the production of raw egg mayonnaise and other food products. Apart from the use of pasteurized eggs, the main control mechanism identified is the pH of the raw egg products, which plays an important role in the consistency and stability while affecting the survival of Salmonella spp. However, currently there is no consensus regarding the critical pH limit for the control of Salmonella. The effectiveness of pH as a control mechanism is influenced by the type of acid used, with the effectiveness of lemon juice compared with vinegar highly debated. Additionally, Salmonella susceptibility to pH stresses may also be influenced by storage temperature (in some studies refrigeration temperatures protected Salmonella spp. from acidulants) and is further complicated by the development of Salmonella cross-tolerance-induced responses, pH homeostasis achieved by the cellular antiport and symport systems, and acid tolerance response (ATR). These mechanisms all provide Salmonella with an added advantage to ensure survival under various pH conditions. Other confounding factors include the fat content, and the addition of NaCl, garlic and plant essential oils (PEOs) from mint, cinnamon, cardamom and clove.


Hepatocyte entry leads to degradation of autoreactive CD8 T cells.
Although most self-reactive T cells are eliminated in the thymus, mechanisms to inactivate or control T cells specific for extrathymic antigens are required and exist in the periphery. By investigating the site in which autoreactive T cells are tolerized, we identify a unique mechanism of peripheral deletion in which naïve autoreactive CD8 T cells are rapidly eliminated in the liver after intrahepatic activation. T cells actively invade hepatocytes, enter endosomal/lysosomal compartments, and are degraded. Blockade of this process leads to accumulation of autoreactive CD8 T cells in the liver and breach of tolerance, with the development of autoimmune hepatitis. Cell into cell invasion, or emperipolesis, is a long-observed phenomenon for which a physiological role has not been previously demonstrated. We propose that this "suicidal emperipolesis" is a unique mechanism of autoreactive T-cell deletion, a process critical for the maintenance of tolerance.


Incidence of retinal detachment after macular surgery: a retrospective study of 634 cases.
To evaluate the rate of retinal detachment after macular surgery. A retrospective non-randomised study of 634 macular surgery procedures was undertaken in two academic centres. Idiopathic macular hole (IMH) surgery (n = 272) and epiretinal membrane (ERM) surgery (n = 362) were performed between 2000 and 2003. We noted the anatomical and functional results of these procedures and we studied serious complications excluding cataract. Minimum follow up was 1 year. No retinal detachment (RD) occurred in patients presenting with an intraoperative or preoperative successfully treated retinal break (RB). The rate of RD occurring after IMH surgery was higher than after the ERM surgical procedure (6.6% vs 2.5%, p = 0.02). The rate of RD was higher in patients presenting with stage 2 and 3 IMH than with stage 4 IMH. However, lens status as well as preoperatively treated RD did not influence the rate of RD after macular surgery. Retinal detachment remains the most common serious complication of macular surgery. Surgical detachment of the posterior vitreous face and associated peripheral retina anomaly seem to increase the rate of this complication. Careful examination of the peripheral retina is a key issue in preventing retinal detachment occurring after macular surgery.


An Integrated Statistical Approach to Compare Transcriptomics Data Across Experiments: A Case Study on the Identification of Candidate Target Genes of the Transcription Factor PPARα.
An effective strategy to elucidate the signal transduction cascades activated by a transcription factor is to compare the transcriptional profiles of wild type and transcription factor knockout models. Many statistical tests have been proposed for analyzing gene expression data, but most tests are based on pair-wise comparisons. Since the analysis of microarrays involves the testing of multiple hypotheses within one study, it is generally accepted that one should control for false positives by the false discovery rate (FDR). However, it has been reported that this may be an inappropriate metric for comparing data across different experiments. Here we propose an approach that addresses the above mentioned problem by the simultaneous testing and integration of the three hypotheses (contrasts) using the cell means ANOVA model. These three contrasts test for the effect of a treatment in wild type, gene knockout, and globally over all experimental groups. We illustrate our approach on microarray experiments that focused on the identification of candidate target genes and biological processes governed by the fatty acid sensing transcription factor PPARα in liver. Compared to the often applied FDR based across experiment comparison, our approach identified a conservative but less noisy set of candidate genes with same sensitivity and specificity. However, our method had the advantage of properly adjusting for multiple testing while integrating data from two experiments, and was driven by biological inference. Taken together, in this study we present a simple, yet efficient strategy to compare differential expression of genes across experiments while controlling for multiple hypothesis testing.


Do women's provider-role attitudes moderate the links between work and family?
The authors examined the links between mothers' work qualities and their individual well-being and marital quality, as well as adolescent daughters' and sons' gender-role attitudes, as a function of mothers' provider-role attitudes, in 134 dual-earner families. In home interviews, mothers described their work, provider-role attitudes, family relationships, and mental health; their offspring reported gender-role attitudes. Women's attitudes about breadwinning were coded into main-secondary, coprovider, and ambivalent coprovider groups. Mothers' provider-role attitudes moderated the links between status indicators and mothers' depression, marital conflict, and daughters' gender-role attitudes. For example, depression and marital conflict were negatively related to coprovider mothers' earnings and occupational prestige. The same was not true for main-secondary and ambivalent coprovider mothers. These findings underscore the importance of considering employed women's interpretation of their work roles when exploring work-family links.


Malignant lymphomas (ML) and HIV infection in Tanzania.
HIV infection is reported to be associated with some malignant lymphomas (ML) so called AIDS-related lymphomas (ARL), with an aggressive behavior and poor prognosis. The ML frequency, pathogenicity, clinical patterns and possible association with AIDS in Tanzania, are not well documented impeding the development of preventive and therapeutic strategies. Sections of 176 archival formalin-fixed paraffin-embedded biopsies of ML patients at Muhimbili National Hospital (MNH)/Muhimbili University of Health and Allied Sciences (MUHAS), Tanzania from 1996-2001 were stained for hematoxylin and eosin and selected (70) cases for expression of pan-leucocytic (CD45), B-cell (CD20), T-cell (CD3), Hodgkin/RS cell (CD30), histiocyte (CD68) and proliferation (Ki-67) antigen markers. Corresponding clinical records were also evaluated. Available sera from 38 ML patients were screened (ELISA) for HIV antibodies. The proportion of ML out of all diagnosed tumors at MNH during the 6 year period was 4.2% (176/4200) comprising 77.84% non-Hodgkin (NHL) including 19.32% Burkitt's (BL) and 22.16% Hodgkin's disease (HD). The ML tumors frequency increased from 0.42% (1997) to 0.70% (2001) and 23.7% of tested sera from these patients were HIV positive. The mean age for all ML was 30, age-range 3-91 and peak age was 1-20 years. The male:female ratio was 1.8:1. Supra-diaphragmatic presentation was commonest and histological sub-types were mostly aggressive B-cell lymphomas however, no clear cases of primary effusion lymphoma (PEL) and primary central nervous system lymphoma (PCNSL) were diagnosed. Malignant lymphomas apparently, increased significantly among diagnosed tumors at MNH between 1996 and 2001, predominantly among the young, HIV infected and AIDS patients. The frequent aggressive clinical and histological presentation as well as the dominant B-immunophenotype and the HIV serology indicate a pathogenic association with AIDS. Therefore, routine HIV screening of all malignant lymphoma patients at MNH is necessary to enable comprehensive ARL diagnosis and formulation of preventive and therapeutic protocols.


Secretome of in vitro cultured human embryos contains extracellular vesicles that are uptaken by the maternal side.
Communication between embryo and maternal endometrium occurs during a specific time frame in which implantation is possible. Here we demonstrate for the first time that conditioned media from non-manipulated human embryos cultured in vitro for 3 days or up to the blastocyst stage contain extracellular vesicles (EVs) with a diameter of 50 to 200 nm and bearing the traditional microvesicle and exosome marker proteins CD63, CD9 and ALIX. The embryonic origin of these EVs has been confirmed by the presence of stemness gene transcripts and their enrichment in the non-classical HLA-G protein. NANOG and POU5F1 transcripts were shown to be contained in vesicles deriving from embryos at different stages of development. In line with a higher detection rate of the HLA-G protein in blastocysts compared to cleavage stage embryos, a significantly higher amount of HLA-G was found in vesicles accumulated in spent media from day 3 to day 5 of development compared to those isolated from the earlier stage. Uptake of dye-labeled embryo-derived EVs by human primary endometrial epithelial and stromal cells was also demonstrated with a fluorescence intensity signal significantly higher for cells treated with vesicles derived from blastocysts. Based on these findings, EV exchange may be suggested as an emerging way of communication at the maternal-fetal interface.


International Nosocomial Infection Control Consortium report, data summary for 2002-2007, issued January 2008.
We report the results of an International Nosocomial Infection Control Consortium (INICC) surveillance study from 2002 through 2007 in 98 intensive care units (ICUs) in Latin America, Asia, Africa, and Europe. During the 6-year study, using Centers for Disease Control and Prevention (CDC) National Nosocomial Infections Surveillance System (NNIS) definitions for device-associated health care-associated infection, we collected prospective data from 43,114 patients hospitalized in the Consortium's hospital ICUs for an aggregate of 272,279 days. Although device utilization in the INICC ICUs was remarkably similar to that reported from US ICUs in the CDC's National Healthcare Safety Network, rates of device-associated nosocomial infection were markedly higher in the ICUs of the INICC hospitals: the pooled rate of central line-associated bloodstream infections (CLABs) in the INICC ICUs, 9.2 per 1000 CL-days, is nearly 3-fold higher than the 2.4-5.3 per 1000 CL-days reported from comparable US ICUs, and the overall rate of ventilator-associated pneumonia was also far higher, 19.5 vs 1.1-3.6 per 1000 ventilator-days, as was the rate of catheter-associated urinary tract infection, 6.5 versus 3.4-5.2 per 1000 catheter-days. Most strikingly, the frequencies of resistance of Staphylococcus aureus isolates to methicillin (MRSA) (80.8% vs 48.1%), Enterobacter species to ceftriaxone (50.8% vs 17.8%), and Pseudomonas aeruginosa to fluoroquinolones (52.4% vs 29.1%) were also far higher in the Consortium's ICUs, and the crude unadjusted excess mortalities of device-related infections ranged from 14.3% (CLABs) to 27.5% (ventilator-associated pneumonia).


Leveraging Structural Diversity and Allosteric Regulatory Mechanisms of Protein Kinases in the Discovery of Small Molecule Inhibitors.
Protein kinases are versatile molecule switches that govern functional processes in signal transduction networks and regulate fundamental biological processes of cell cycle and organism development. The continuous growth of biological information and a remarkable breath of structural, genetic, and pharmacological studies on protein kinase genes have significantly advanced our knowledge of the kinase activation, drug binding and allosteric mechanisms underlying kinase regulation and interactions in signaling cascades.. Structural and biochemical studies of the genetic and molecular determinants of protein kinases binding with inhibitors have been the cornerstone of drug discovery efforts in clinical oncology leading to proliferation of effective anticancer therapies. Recent advances in understanding allosteric regulation of protein kinases have fueled unprecedented efforts aiming in the discovery of targeted and allosteric kinase inhibitors that can combat cancer mutants and are at the forefront of the precision medicine initiative in oncology. Despite diversity of regulatory scenarios underlying kinase functions, dimerization-driven activation is a common mechanism of allosteric regulation that is shared by many protein kinase families, most notably ErbB and BRAF kinases that play a central role in growth factor signaling and human disease. In this review, we focused on structural, biochemical and computational studies of the ErbB and BRAF kinases and discuss how diversity of the structural landscape for these kinase genes and dimerization- dependent mechanisms of their regulation can be leveraged in the design and discovery of kinase inhibitors and allosteric modulators of kinase activation. The lessons from this analysis could inform discovery of specific targeted therapies and robust drug combinations for cancer treatment.


The role of extracellular matrix quality in pulmonary fibrosis.
This review discusses the role of extracellular matrix (ECM) quality in the pathogenesis of pulmonary fibrosis (PF). In PF, the highly ordered structure of collagens and elastin within the ECM of the lung is severely disrupted and lacks its original tissue quality. Discussions about the ECM have focused on the role of protein quantity in relation to the progression of PF, while the importance of lung ECM quality, defined by the levels of ECM protein modifications and by the protein distribution in lung tissue, has not been properly addressed. The quality and function of proteins may be altered by different post-translational modifications (PTMs), such as cross-linking, proteolytic cleavage, citrullination, misfolding and glycosylation. This paper is the first to review key data from the literature related to the lung ECM at the molecular level, relate these to changes observed at a macroscopic level and evaluate which PTMs most likely contribute to PF. This paper also reviews the role of novel neo-epitope-specific biomarkers in the early diagnosis and prognosis of fibrotic disorders. We discuss and argue that the altered quality of the individual ECM proteins contributes to the progression of PF and may also lead to the increased quantity of lung proteins. Thus, both quantity and quality appear to be of utmost importance.


Descriptors for Electron and Hole Charge Carriers in Metal Oxides.
Metal oxides can act as insulators, semiconductors, or metals depending on their chemical composition and crystal structure. Metal oxide semiconductors, which support equilibrium populations of electron and hole charge carriers, have widespread applications including batteries, solar cells, and display technologies. It is often difficult to predict in advance whether these materials will exhibit localized or delocalized charge carriers upon oxidation or reduction. We combine data from first-principles calculations of the electronic structure and dielectric response of 214 metal oxides to predict the energetic driving force for carrier localization and transport. We assess descriptors based on the carrier effective mass, static polaron binding energy, and Fröhlich electron-phonon coupling. Numerical analysis allows us to assign p- and n-type transport of a metal oxide to three classes: (i) band transport with high mobility; (ii) small polaron transport with low mobility; and (iii) intermediate behavior. The results of this classification agree with observations regarding carrier dynamics and lifetimes and are used to predict 10 candidate p-type oxides.


Ultrasound guided corticosteroid injection for plantar fasciitis: randomised controlled trial.
To investigate the effectiveness of ultrasound guided corticosteroid injection in the treatment of plantar fasciitis. Randomised, investigator and participant blinded, placebo controlled trial. University clinic in Melbourne, Australia. 82 people with a clinical and ultrasound diagnosis of plantar fasciitis unrelated to systemic inflammatory disease. Participants were randomly allocated to ultrasound guided injection of the plantar fascia with either 1 mL of 4 mg/mL dexamethasone sodium phosphate (experimental group) or 1 mL normal saline (placebo). Before injection the participants were given an ultrasound guided posterior tibial nerve block with 2% lidocaine (lignocaine). Primary outcomes were pain, as measured by the foot health status questionnaire (0-100 point scale), and plantar fascia thickness, measured by ultrasound at 4, 8, and 12 weeks. Reduction in pain at four weeks favoured the dexamethasone group by 10.9 points (95% confidence interval 1.4 to 20.4, P=0.03). Between group differences for pain scores at eight and 12 weeks were not statistically significant. Plantar fascia thickness measured at four weeks favoured the dexamethasone group by -0.35 mm (95% confidence interval -0.67 to -0.03, P=0.03). At eight and 12 weeks, between group differences for plantar fascia thickness also favoured dexamethasone, at -0.39 mm (-0.73 to -0.05, P=0.02) and -0.43 mm (-0.85 to -0.01, P=0.04), respectively. The number needed to treat with dexamethasone for one successful outcome for pain at four weeks was 2.93 (95% confidence interval 2.76 to 3.12). There were no reported adverse events associated with the intervention. A single ultrasound guided dexamethasone injection is a safe and effective short term treatment for plantar fasciitis. It provides greater pain relief than placebo at four weeks and reduces abnormal swelling of the plantar fascia for up to three months. However, clinicians offering this treatment should also note that significant pain relief did not continue beyond four weeks. Australian New Zealand Clinical Trials Registry ACTRN12610000239066.


Investigation of hybrid microlens integration with vertical-cavity surface-emitting lasers for free-space optical links.
In a novel one-step process, a vertical-cavity surface-emitting laser (VCSEL, operation wavelength of 980 nm) is integrated with a hybrid microdiffractive lens by focused ion beam milling (FIBM) for use in freespace optical links. A hybrid microlens with a diameter of 100 m, numerical aperture of 0.56, and sag height of 4.196 m, combined with a diffractive lens with continuous relief and 6 annuli, was designed and fabricated in one step by FIBM on the back of a VCSEL with a GaAs substrate for beam collimation. A previous VCSEL integrated with a pure diffractive lens had a half-divergence angle of 0.6 degrees ; the half-divergence angle of the VCSEL with the hybrid microlens was improved to 0.3 degrees . Test results show that athermalization with respect to the variation in operating temperature can be realized with the hybrid microlens.


5-Benzylidene-4-oxazolidinones potently inhibit biofilm formation in Methicillin-resistant Staphylococcus aureus.
Investigation into the biological function of 5-benzylidene-4-oxazolidinones revealed dose-dependent inhibition of biofilm formation in Methicillin-resistant S. aureus (MRSA). This structurally unusual class of small molecules inhibit up to 89% of biofilm formation with IC50 values as low as 0.78 μM, and disperse pre-formed biofilms with IC50 values as low as 4.7 μM. Together, these results suggest that 4-oxazolidinones represent new chemotypes to enable the study of bacterial biofilms with small molecule chemical probes.


Oncolytic viruses: finally delivering.
Oncolytic viruses can be found at the confluence of virology, genetic engineering and pharmacology where versatile platforms for molecularly targeted anticancer agents can be designed and optimised. Oncolytic viruses offer several important advantages over traditional approaches, including the following. (1) Amplification of the active agent (infectious virus particles) within the tumour. This avoids unnecessary exposure to normal tissues experienced during delivery of traditional stoichiometric chemotherapy and maximises the therapeutic index. (2) The active cell-killing mechanisms, often independent of programmed death mechanisms, should decrease the emergence of acquired drug resistance. (3) Lytic death of cancer cells provides a pro-inflammatory microenvironment and the potential for induction of an anticancer vaccine response. (4) Tumour-selective expression and secretion of encoded anticancer biologics, providing a new realm of potent and cost-effective-targeted therapeutics.


The dynamic in vivo distribution of bone marrow-derived mesenchymal stem cells after infusion.
Bone marrow-derived mesenchymal stem cells (MSCs) have the potential to differentiate along different mesenchymal lineages including those forming bone, cartilage, tendon, fat, muscle and marrow stroma that supports hematopoiesis. This differentiation potential makes MSCs candidates for cell-based therapeutic strategies for mesenchymal tissue injuries and for hematopoietic disorders by both local and systemic application. In the present study, rat marrow-derived MSCs were ex vivo culture-expanded, labeled with (111)In-oxine, and infused into syngeneic rats via intra-artery (i.a.), intravenous (i.v.) and intraperitoneal cavity (i.p.) infusions. In addition, for i.a. and i.v. infusions, a vasodilator, sodium nitroprusside, was administered prior to the cell infusion and examined for its effect on MSC circulation. The dynamic distribution of infused MSCs was monitored by real-time imaging using a gamma camera immediately after infusion and at 48 h postinfusion. After 48 h, radioactivity in excised organs, including liver, lungs, kidneys, spleen and long bones, was measured in a gamma well counter and expressed as a percentage of injected doses. After both i.a. and i.v. infusion, radioactivity associated with MSCs was detected primarily in the lungs and then secondarily in the liver and other organs. When sodium nitroprusside was used, more labeled MSCs cleared the lungs resulting in a larger proportion detected in the liver. Most importantly, the homing of labeled MSCs to the marrow of long bones was significantly increased by the pretreatment with vasodilator. These results indicate multiple homing sites for injected MSCs and that the distribution of MSCs can be influenced by administration of vasodilator.


The birth of the American Physician Scientists Association--the next generation of Young Turks.
The American Society for Clinical Investigation (ASCI) was started a century ago to foster and to address the needs of the younger physician-scientists. A hundred years later, ASCI remains one of the premier organizations for physician-scientists and one of most well-respected organizations in the medical community. I have had the opportunity and pleasure to interact with the ASCI not only as an organization through my tenure as president of the American Physician Scientists Association, but also with its members over the last four years. In my view, the same characteristics that permeate ASCI the organization also define ASCI the membership--mentorship, exemplary role models, advocacy, and leadership.


Cannabidiol-based natural health products for companion animals: Recent advances in the management of anxiety, pain, and inflammation.
Recent advances in cannabidiol (CBD) use in canines and felines for anxiety management, pain management, and anti-inflammatory effects were reviewed using a literature search conducted with the following keywords: CBD, anxiety, inflammation, pain, dogs, cats, and companion animals. For decades, research on CBD has been hindered due to the status of cannabis (C. sativa L.) as an illicit drug. Limited safety data show that CBD is well-tolerated in dogs, with insufficient information on the safety profile of CBD in cats. Upon oral supplementation of CBD, elevation in liver enzymes was observed for both dogs and cats, and pharmacokinetics of CBD are different in the two species. There is a significant gap in the literature on the therapeutic use of CBD in cats, with no feline data on anxiety, pain, and inflammation management. There is evidence that chronic osteoarthritic pain in dogs can be reduced by supplementation with CBD. Furthermore, experiments are required to better understand whether CBD has an influence on noise-induced fear and anxiolytic response. Preliminary evidence exists to support the analgesic properties of CBD in treating chronic canine osteoarthritis; however, there are inter- and intra-species differences in pharmacokinetics, tolerance, dosage, and safety of CBD. Therefore, to validate the anxiety management, pain management, and anti-inflammatory efficacy of CBD, it is essential to conduct systematic, randomized, and controlled trials. Further, the safety and efficacious dose of CBD in companion animals warrants investigation.


Reduced Self-Referential Source Memory Performance is Associated with Interpersonal Dysfunction in Borderline Personality Disorder.
Source memory is impaired in schizophrenia, and this deficit is related to symptoms of interpersonal antagonism such as suspiciousness and hostility. The present study evaluated source memory in borderline personality disorder (BPD) and its relation to interpersonal antagonism. Forty-one noninpatient adults with BPD according to the DSM-IV and 26 healthy control subjects performed a verbal source memory test requiring completion of sentences with and without emotional content ("Hot" vs. "Cold" sentences). Subjects also completed self-report measures of suspiciousness and interpersonal antagonism (Buss-Durkee Hostility Inventory) and depression (Beck Depression Inventory). The BPD group showed no significant difference from the control group in self-referential source memory, recognition memory, response bias, and performance enhancement for items with emotion content. However, in the BPD group, poorer self-referential source memory was significantly related to Hostility measures including suspiciousness, but not with Depression scores. In contrast, generic item recognition memory was unrelated to Hostility. Heterogeneity in source memory function may be specifically related to some of the hallmark interpersonal disturbances of BPD, independent of the effects of general negative affect or general memory impairment.


Predominance of HLA-restricted cytotoxic T-lymphocyte responses to serotype-cross-reactive epitopes on nonstructural proteins following natural secondary dengue virus infection.
We examined the memory cytotoxic T-lymphocytic (CTL) responses of peripheral blood mononuclear cells (PBMC) obtained from patients in Thailand 12 months after natural symptomatic secondary dengue virus infection. In all four patients analyzed, CTLs were detected in bulk culture PBMC against nonstructural dengue virus proteins. Numerous CD4+ and CD8+ CTL lines were generated from the bulk cultures of two patients, KPP94-037 and KPP94-024, which were specific for NS1.2a (NS1 and NS2a collectively) and NS3 proteins, respectively. All CTL lines derived from both patients were cross-reactive with other serotypes of dengue virus. The CD8+ NS1.2a-specific lines from patient KPP94-037 were HLA B57 restricted, and the CD8+ NS3-specific lines from patient KPP94-024 were HLA B7 restricted. The CD4+ CTL lines from patient KPP94-037 were HLA DR7 restricted. A majority of the CD8+ CTLs isolated from patient KPP94-024 were found to recognize amino acids 221 to 232 on NS3. These results demonstrate that in Thai patients after symptomatic secondary natural dengue infections, CTLs are mainly directed against nonstructural proteins and are broadly cross-reactive.


2-Methylene analogs of 1alpha-hydroxy-19-norvitamin D3: synthesis, biological activities and docking to the ligand binding domain of the rat vitamin D receptor.
In continuing efforts towards the synthesis of biologically active vitamin D compounds of potential therapeutic value, new 2-methylene-1alpha-hydroxy-19-norvitamin D(3) analogs 3 and 4 with modified alkyl side chains have been synthesized. The key synthetic step involved Lythgoe-type Wittig-Horner coupling of Windaus-Grundmann type ketones 9, possessing different 17beta-alkyl substituents, with the phosphine oxide 10 prepared from (-)-quinic acid. The prepared vitamins 3 and 4 were ca. eight times less potent than 1alpha,25-dihydroxyvitamin D(3) (1alpha,25-(OH)(2)D(3)) (1) in binding to the rat intestinal vitamin D receptor (VDR). In comparison with the hormone 1 they exhibited slightly lower cellular HL-60 differentiation activity. When tested in vivo; the analog 3 was characterized by very high bone calcium mobilizing potency and intestinal calcium transport activity. Unexpectedly, the 25-methyl compound 4 showed marked calcemic activity in both assays. Computational docking of the vitamin 3 into the binding pocket of the rat vitamin D receptor is also reported.


DIVA vaccine properties of the live chimeric pestivirus strain CP7_E2gif.
Live modified vaccines to protect against classical swine fever virus (CSFV), based on chimeric pestiviruses, have been developed to enable serological Differentiation of Infected from Vaccinated Animals (DIVA). In this context, the chimeric virus CP7_E2gif vaccine candidate is unique as it does not include any CSFV components. In the present study, the DIVA vaccine properties of CP7_E2gif were evaluated in comparison to the conventional live attenuated Riemser C-strain vaccine. Sera and tonsil samples obtained from pigs immunised with these two vaccines were analysed. No viral RNA was found in serum after vaccination with CP7_E2gif, whereas some serum samples from C-strain vaccinated animals were positive. In both vaccinated groups, individual viral RNA-positive tonsil samples were detected in animals euthanised between 7 and 21 days post vaccination. Furthermore, serum samples from these animals, together with archival samples from pigs vaccinated with CP7_E2gif and subsequently CSFV challenged, were analysed for specific antibodies using ELISAs and for homologous neutralising antibodies. In animals vaccinated with CP7_E2gif, neutralising antibodies were detected from day 10. However, the sera remained negative for anti-CSFV E2-specific antibodies whereas pigs vaccinated with C-strain seroconverted against CSFV by 14 days after vaccination, as determined by a CSFV-E2 specific blocking ELISA. One week after subsequent CSFV challenge, a strong anti-CSFV E2 reaction was detected in CP7_E2gif vaccinated pigs and anti-E(rns) antibodies were detected from 10 days after infection. In conclusion, CP7_E2gif has the potential to be used as a DIVA vaccine in combination with detection of anti-CSFV E2-specific antibodies.


HIV-related symptoms and patient clusters among Chileans living with HIV.
Identifying both Human immunodeficiency virus (HIV)-related and co-morbid symptoms experienced by people living with HIV (PLWH) who are receiving antiretroviral therapy (ART) treatment is a major challenge for healthcare providers globally. Yet, little research to date has examined the symptoms of illness experienced by PLWH including patients living in Central and South American. To address this gap, this study was designed to identify symptoms of HIV by socio-demographic and/or clinical characteristics among Chilean patients living with the virus. A convenience sample of 209 Chilean PLWH was recruited from an outpatient clinic in Santiago, Chile. A structured interview was used to elicit socio-demographic information and HIV symptoms status. Additional clinical information was obtained through a review of the participants' medical records. Results show that patients' most commonly reported HIV-related symptoms were fear/worries (66%), anxiety (52%), gas/bloating (50%), and thirst (50%). Multivariate analysis revealed a positive association between the number of reported HIV-related symptoms and number of years living with HIV. Having completed college was negatively associated with number of symptoms. Latent class analysis indicated that PLWH in the sample who had completed college were two times more likely to experience a mild intensity of HIV-related symptoms than their lesser educated counterparts. Similarly, logistic regression revealed that college-educated PLWH were twice as likely to be classified in the subgroup reporting mild intensity of symptoms than those who lacked a college degree. Overall, the study's results reveal that many Chilean PLWH, even those with high CD4 counts and low or undetectable viral loads, are not symptom free. The findings point to the need for clinicians to tailor a plan of care for individuals living with HIV that is based on their symptomatology.


Improvement of Physical Decline Through Combined Effects of Muscle Enhancement and Mitochondrial Activation by a Gastric Hormone Ghrelin in Male 5/6Nx CKD Model Mice.
Because a physical decline correlates with an increased risk of a wide range of disease and morbidity, an improvement of physical performance is expected to bring significant clinical benefits. The primary cause of physical decline in 5/6 nephrectomized (5/6Nx) chronic kidney disease model mice has been regarded as a decrease in muscle mass; however, our recent study showed that a decrease in muscle mitochondria plays a critical role. In the present study, we examined the effects of a gastric hormone ghrelin, which has been reported to promote muscle mitochondrial oxidation, on the physical decline in the chronic kidney disease model mice, focusing on the epigenetic modulations of a mitochondrial activator gene, peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α). Ghrelin treatment improved a decline in exercise endurance of 5/6Nx mice, associated with an increase in both of the muscle mass and mitochondrial amount. The expression level of PGC-1α was decreased in the skeletal muscle of 5/6Nx mice, which was associated with an increase in the methylation ratio of the cytosine residue at 260 base pairs upstream of the initiation point. Conversely, ghrelin treatment de-methylated the cytosine residue and increased the expression of PGC-1α. A representative muscle anabolic factor, IGF-1, did not affect the expression of PGC-1α and muscle mitochondrial amount, although it increased muscle mass. As a result, IGF-1 treatment in 5/6Nx mice did not increase the decreased exercise endurance as effectively as ghrelin treatment did. These findings indicate an advantage of ghrelin treatment for a recovery of physical decline.


Population-Based Incidence Rates of Diarrheal Disease Associated with Norovirus, Sapovirus, and Astrovirus in Kenya.
Diarrheal diseases remain a major cause of mortality in Africa and worldwide. While the burden of rotavirus is well described, population-based rates of disease caused by norovirus, sapovirus, and astrovirus are lacking, particularly in developing countries. Data on diarrhea cases were collected through a population-based surveillance platform including healthcare encounters and household visits in Kenya. We analyzed data from June 2007 to October 2008 in Lwak, a rural site in western Kenya, and from October 2006 to February 2009 in Kibera, an urban slum. Stool specimens from diarrhea cases of all ages who visited study clinics were tested for norovirus, sapovirus, and astrovirus by RT-PCR. Of 334 stool specimens from Lwak and 524 from Kibera, 85 (25%) and 159 (30%) were positive for norovirus, 13 (4%) and 31 (6%) for sapovirus, and 28 (8%) and 18 (3%) for astrovirus, respectively. Among norovirus-positive specimens, genogroup II predominated in both sites, detected in 74 (87%) in Lwak and 140 (88%) in Kibera. The adjusted community incidence per 100,000 person-years was the highest for norovirus (Lwak: 9,635; Kibera: 4,116), followed by astrovirus (Lwak: 3,051; Kibera: 440) and sapovirus (Lwak: 1,445; Kibera: 879). For all viruses, the adjusted incidence was higher among children aged <5 years (norovirus: 22,225 in Lwak and 17,511 in Kibera; sapovirus: 5,556 in Lwak and 4,378 in Kibera; astrovirus: 11,113 in Lwak and 2,814 in Kibera) compared to cases aged ≥5 years. Although limited by a lack of controls, this is the first study to estimate the outpatient and community incidence rates of norovirus, sapovirus, and astrovirus across the age spectrum in Kenya, suggesting a substantial disease burden imposed by these viruses. By applying adjusted rates, we estimate approximately 2.8-3.3 million, 0.45-0.54 million, and 0.77-0.95 million people become ill with norovirus, sapovirus, and astrovirus, respectively, every year in Kenya.


Troponin T- and troponin I-like proteins in bovine vascular smooth muscle.
We have tested the hypothesis whether proteins with biochemical and immunochemical properties similar to those of troponin T (TnT) and troponin I (TnI) are expressed in bovine vascular smooth muscle (SM). Three monoclonal anti-TnT antibodies (TT-1, TT-2, and RV-C2) specific for the two isoforms of TnT present in the bovine cardiac muscle and two monoclonal antibodies (TI-1 and TI-5) reacting with cardiac TnI were used in this study. Anti-TnT antibodies were found to be unreactive with 1) skeletal and nonmuscle isoforms of glyceraldehyde-3-phosphate dehydrogenase, a glycolytic enzyme that shares some structural homologies with skeletal TnT, and 2) calponin, a TnT-like calmodulin/tropomyosin binding protein with some antigenic properties in common with TnT. When tested on SM extracts from aorta or coronary arteries by Western blotting, the anti-TnT antibodies were able to react exclusively with one or two polypeptides whose electrophoretic mobility corresponds to the cardiac TnT subunits. Similarly, anti-TnI antibodies specifically recognized a component in the aortic or coronary SM extracts with electrophoretic properties identical to the cardiac TnI. Immunofluorescence analysis performed on the vascular SM cells of bovine aorta, coronary arteries, and intramural branches of coronary vessels confirmed the existence of cardiac troponin immunoreactivity in these tissues. In addition, differences in the distribution of cardiac TnT- and TnI-like proteins were evidenced in nonvascular and vascular SM cells. This study shows for the first time that polypeptides with some structural properties in common with cardiac TnT and TnI can be found in the vascular SM system.


Early life stress is associated with default system integrity and emotionality during infancy.
Extensive animal research has demonstrated the vulnerability of the brain to early life stress (ELS) with consequences for emotional development and mental health. However, the influence of moderate and common forms of stress on early human brain development is less well-understood and precisely characterized. To date, most work has focused on severe forms of stress, and/or on brain functioning years after stress exposure. In this report we focused on conflict between parents (interparental conflict), a common and relatively moderate form of ELS that is highly relevant for children's mental health outcomes. We used resting state functional connectivity MRI to examine the coordinated functioning of the infant brain (N = 23; 6-12-months-of-age) in the context of interparental conflict. We focused on the default mode network (DMN) due to its well-characterized developmental trajectory and implications for mental health. We further examined DMN strength as a mediator between conflict and infants' negative emotionality. Higher interparental conflict since birth was associated with infants showing stronger connectivity between two core DMN regions, the posterior cingulate cortex (PCC) and the anterior medial prefrontal cortex (aMPFC). PCC to amygdala connectivity was also increased. Stronger PCC-aMPFC connectivity mediated between higher conflict and higher negative infant emotionality. The developing DMN may be an important marker for effects of ELS with relevance for emotional development and subsequent mental health. Increasing understanding of the associations between common forms of family stress and emerging functional brain networks has potential to inform intervention efforts to improve mental health outcomes.


Intranasal vaccination of rabbits with Pasteurella multocida A:3 outer membranes that express iron-regulated proteins.
To determine efficacy of intranasal vaccination of rabbits with Pasteurella multocida A:3 outer membrane proteins (OMP) expressing iron-regulated OMP (IROMP) in conferring protection against experimental challenge exposure. 52 male New Zealand White rabbits. Rabbits were vaccinated intranasally on days 0, 7, and 14; some vaccines included cholera toxin (CT) as an adjuvant. Concentrations of intranasal IgA and serum IgG antibodies against P multocida OMP were determined. In experiment A, rabbits were vaccinated with either phospate-buffered saline solution (PBSS), PBSS-CT, OMP-CT, or IROMP-CT, challenge-exposed intranasally on day 16, and euthanatized and necropsied on day 28. Rabbits were also vaccinated with OMP or IROMP without CT and were not challenge-exposed. In experiment B, rabbits were vaccinated with PBSS, PBSS-CT, IROMP, or IROMP-CT. On day 17, rabbits were challenge-exposed intranasally. Nasal bacteria and antibodies were determined on day 24. In experiment A, OMP-CT vaccination stimulated mucosal and systemic antibody responses to the bacterium and enhanced resistance against challenge exposure. Intranasal bacterial counts were not significantly reduced. Vaccination with IROMP-CT stimulated mucosal and systemic antibodies, enhanced resistance to challenge exposure, and significantly reduced nasal bacterial counts. In experiment B, natural infection was detected in several rabbits at challenge exposure; however, IROMP-CT-vaccinated rabbits had significantly higher serum and nasal antibody responses, compared with other rabbits IROMP-CT-vaccinated rabbits had significantly lower nasal bacterial counts compared to control rabbits. Intranasal vaccination of rabbits with P multocida outer membranes containing IROMP and CT stimulated immunity against experimental pneumonic pasteurellosis.


Revising the recent evolutionary history of equids using ancient DNA.
The rich fossil record of the family Equidae (Mammalia: Perissodactyla) over the past 55 MY has made it an icon for the patterns and processes of macroevolution. Despite this, many aspects of equid phylogenetic relationships and taxonomy remain unresolved. Recent genetic analyses of extinct equids have revealed unexpected evolutionary patterns and a need for major revisions at the generic, subgeneric, and species levels. To investigate this issue we examine 35 ancient equid specimens from four geographic regions (South America, Europe, Southwest Asia, and South Africa), of which 22 delivered 87-688 bp of reproducible aDNA mitochondrial sequence. Phylogenetic analyses support a major revision of the recent evolutionary history of equids and reveal two new species, a South American hippidion and a descendant of a basal lineage potentially related to Middle Pleistocene equids. Sequences from specimens assigned to the giant extinct Cape zebra, Equus capensis, formed a separate clade within the modern plain zebra species, a phenotypicically plastic group that also included the extinct quagga. In addition, we revise the currently recognized extinction times for two hemione-related equid groups. However, it is apparent that the current dataset cannot solve all of the taxonomic and phylogenetic questions relevant to the evolution of Equus. In light of these findings, we propose a rapid DNA barcoding approach to evaluate the taxonomic status of the many Late Pleistocene fossil Equidae species that have been described from purely morphological analyses.


Mitochondrial DNA mutations in preneoplastic lesions of the gastrointestinal tract: a biomarker for the early detection of cancer.
Somatic mutations of mitochondrial DNA (mtDNA) are common in many human cancers. We have described an oligonucleotide microarray ("MitoChip") for rapid sequencing of the entire mitochondrial genome (Zhou et al, J Mol Diagn 2006), facilitating the analysis of mtDNA mutations in preneoplastic lesions. We examined 14 precancerous lesions, including seven Barrett esophagus biopsies, with or without associated dysplasia; four colorectal adenomas; and three inflammatory colitis-associated dysplasia specimens. In all cases, matched normal tissues from the corresponding site were obtained as germline control. MitoChip analysis was performed on DNA obtained from cryostat-embedded specimens. A total of 513,639 bases of mtDNA were sequenced in the 14 samples, with 490,224 bases (95.4%) bases assigned by the automated genotyping software. All preneoplastic lesions examined demonstrated at least one somatic mtDNA sequence alteration. Of the 100 somatic mtDNA alterations observed in the 14 cases, 27 were non-synonymous coding region mutations (i.e., resulting in an amino acid change), 36 were synonymous, and 37 involved non-coding mtDNA. Overall, somatic alterations most commonly involved the COI, ND4 and ND5 genes. Notably, somatic mtDNA alterations were observed in preneoplastic lesions of the gastrointestinal tract even in the absence of histopathologic evidence of dysplasia, suggesting that the mitochondrial genome is susceptible at the earliest stages of multistep cancer progression. Our findings further substantiate the rationale for exploring the mitochondrial genome as a biomarker for the early diagnosis of cancer, and confirm the utility of a high-throughput array-based platform for this purpose from a clinical applicability standpoint.


Surgical Management of Aorto-Ventricular Tunnel. A Multicenter Study.
Aorto-ventricular tunnel (AoVT), a rare congenital anomaly, is a channel originating in the ascending aorta just above the sinotubular junction and leading to the cavity of the left ventricle (AoLVT), or, rarely, the right (AoRVT). This study reviews our collective 30-year experience with the surgical treatment of AoVT. Data were submitted by 15 participating centers on 42 patients who underwent correction of AoVT between 1987 and 2018. Of these, 36 had AoLVT, and 6 AoRVT. The tunnel originated above the right coronary sinus in 28 (77.8%) patients. For AoLVT, most operations were performed early (median age 25 days, range: 1 day-25 years). In contrast, AoRVT was diagnosed and repaired later (median age 6 years, range: 1 month-12 years). Surgically important coronary ostial displacement was common. Patch closure of the aortic orifice only was the commonest surgical repair for AoLVT (23 patients), while in AoRVT, both orifices or only the ventricular one was closed. Aortic valvar insufficiency, severe or moderate, coexisted in 11 (30.5%) patients with AoLVT, and aortic valvuloplasty was performed in 8, mainly due to aortic valve stenosis. Aortic valvar insufficiency at discharge ranged from trivial to mild in almost all patients. Early mortality was 7.14%, with 3 patients with AoLVT succumbing to cardiac failure. There were 2 early reoperations and 1 late death. AoVT is a rare malformation. AoLVT usually necessitates surgery in early life. AoRVT is rarer, diagnosed and repaired later in life. Surgical repair by patch closure, with concomitant aortic valve repair as needed, is associated with good results.


Recent advances on hepatitis C virus in dialysis population.
Hepatitis C virus (HCV) infection remains common among patients undergoing regular dialysis and good evidence supports the detrimental role of HCV on survival in patients undergoing maintenance dialysis. According to an updated meta-analysis of clinical studies (n=15; 195,370 unique patients on maintenance dialysis), the summary estimate for adjusted relative risk (all-cause mortality) with anti-HCV across the published studies was 1.32 with a 95% Confidence Intervals of 1.24; 1.42, homogeneity assumption was not rejected. Various mechanisms support the excess death risk of HCV-infected patients on regular dialysis, in addition to liver disease-related mortality. The adjusted relative risk for cardiovascular mortality among HCV-infected patients on regular dialysis was 1.26 (95% Confidence Intervals, 1.10; 1.45); the increased cardiovascular mortality in anti-HCV positive patients has been associated in part to malnutrition and chronic inflammation. The current standard of care for HCV in dialysis population is combined antiviral therapy (pegylated interferon plus ribavirin) with a rate of viral response of around 60%. Triple therapy with telaprevir proved to be effective and safe in dialysis patients with HCV but only anecdotal evidence exists. Antiviral treatment of HCV-infected patients on maintenance dialysis could lead to cure the liver damage and the extrahepatic complications. The future availability of all-oral interferon/ribavirin free regimens for antiviral treatment of HCV will help nephrologists to improve survival in this high-risk group.


Pathological alterations in GABAergic interneurons and reduced tonic inhibition in the basolateral amygdala during epileptogenesis.
An acute brain insult such as traumatic head/brain injury, stroke, or an episode of status epilepticus can trigger epileptogenesis, which, after a latent, seizure-free period, leads to epilepsy. The discovery of effective pharmacological interventions that can prevent the development of epilepsy requires knowledge of the alterations that occur during epileptogenesis in brain regions that play a central role in the induction and expression of epilepsy. In the present study, we investigated pathological alterations in GABAergic interneurons in the rat basolateral amygdala (BLA), and the functional impact of these alterations on inhibitory synaptic transmission, on days 7 to 10 after status epilepticus induced by kainic acid. Using design-based stereology combined with glutamic acid decarboxylase (GAD) 67 immunohistochemistry, we found a more extensive loss of GABAergic interneurons compared to the loss of principal cells. Fluoro-Jade C staining showed that neuronal degeneration was still ongoing. These alterations were accompanied by an increase in the levels of GAD and the alpha1 subunit of the GABA(A) receptor, and a reduction in the GluK1 (previously known as GluR5) subunit, as determined by Western blots. Whole-cell recordings from BLA pyramidal neurons showed a significant reduction in the frequency and amplitude of action potential-dependent spontaneous inhibitory postsynaptic currents (IPSCs), a reduced frequency but not amplitude of miniature IPSCs, and impairment in the modulation of IPSCs via GluK1-containing kainate receptors (GluK1Rs). Thus, in the BLA, GABAergic interneurons are more vulnerable to seizure-induced damage than principal cells. Surviving interneurons increase their expression of GAD and the alpha1 GABA(A) receptor subunit, but this does not compensate for the interneuronal loss; the result is a dramatic reduction of tonic inhibition in the BLA circuitry. As activation of GluK1Rs by ambient levels of glutamate facilitates GABA release, the reduced level and function of these receptors may contribute to the reduction of tonic inhibitory activity. These alterations at a relatively early stage of epileptogenesis may facilitate the progress towards the development of epilepsy.


Preparation, fabrication and biocompatibility of novel injectable temperature-sensitive chitosan/glycerophosphate/collagen hydrogels.
This paper introduces a novel type of injectable temperature-sensitive chitosan/glycerophosphate/collagen (C/GP/Co) hydrogel that possesses great biocompatibility for the culture of adipose tissue-derived stem cells. The C/GP/Co hydrogel is prepared by mixing 2.2% (v/v) chitosan with 50% (w/w) β-glycerophosphate at different proportions and afterwards adding 2 mg/ml of collagen. The gelation time of the prepared solution at 37°C was found to be of around 12 min. The inner structure of the hydrogel presented a porous spongy structure, as observed by scanning electron microscopy. Moreover, the osmolality of the medium in contact with the hydrogel was in the range of 310-330 mmol kg(-1). These analyses have shown that the C/GP/Co hydrogels are structurally feasible for cell culture, while their biocompatibility was further examined. Human adipose tissue-derived stem cells (ADSCs) were seeded into the developed C/GP and C/GP/Co hydrogels (The ratios of C/GP and C/GP/Co were 5:1 and 5:1:6, respectively), and the cellular growth was periodically observed under an inverted microscope. The proliferation of ADSCs was detected using cck-8 kits, while cell apoptosis was determined by a Live/Dead Viability/Cytotoxicity kit. After 7 days of culture, cells within the C/GP/Co hydrogels displayed a typical adherent cell morphology and good proliferation with very high cellular viability. It was thus demonstrated that the novel C/GP/Co hydrogel herein described possess excellent cellular compatibility, representing a new alternative as a scaffold for tissue engineering, with the added advantage of being a gel at the body's temperature that turns liquid at room temperature.


Transforming clinical imaging and 3D data for virtual reality learning objects: HTML5 and mobile devices implementation.
Web deployable anatomical simulations or "virtual reality learning objects" can easily be produced with QuickTime VR software, but their use for online and mobile learning is being limited by the declining support for web browser plug-ins for personal computers and unavailability on popular mobile devices like Apple iPad and Android tablets. This article describes complementary methods for creating comparable, multiplatform VR learning objects in the new HTML5 standard format, circumventing platform-specific limitations imposed by the QuickTime VR multimedia file format. Multiple types or "dimensions" of anatomical information can be embedded in such learning objects, supporting different kinds of online learning applications, including interactive atlases, examination questions, and complex, multi-structure presentations. Such HTML5 VR learning objects are usable on new mobile devices that do not support QuickTime VR, as well as on personal computers. Furthermore, HTML5 VR learning objects can be embedded in "ebook" document files, supporting the development of new types of electronic textbooks on mobile devices that are increasingly popular and self-adopted for mobile learning.


The impact of metabolic control and tetrahydrobiopterin treatment on health related quality of life of patients with early-treated phenylketonuria: A PKU-COBESO study.
The aim of this study was to examine Health-Related Quality of Life (HRQoL) of patients with Phenylketonuria (PKU) in three different age groups and to investigate the impact of metabolic control and tetrahydrobiopterin (BH4) treatment on HRQoL of these patients. Participants were 90 early-treated patients aged 7 to 40 years (M = 21.0, SD = 10.1) and 109 controls aged 7 to 40.8 years (M = 19.4, SD = 8.6). HRQoL was assessed with the (generic) TNO-AZL questionnaires. Overall, good HRQoL was reported for children below 12 years of age, although they were judged to be less autonomic than their healthy counterparts. Adolescents aged 12-15 years showed poorer HRQoL in the domain "cognitive functioning" compared to controls. For adults ≥16 years, poorer age-controlled HRQoL was found for the domains cognition, depressive moods, and anger, with a further trend for the domain "pain". With respect to metabolic control, only for adult PKU-patients robust associations were observed, indicating poorer functioning, most notably in the domains cognition, sleep, pain, sexuality and anger, with higher historical and concurrent Phe-levels. With respect to BH4-use, effects on HRQoL were again only observed for adult PKU-patients. After controlling for age and historical Phe-levels, small but significant differences in favor of adult BH4-users compared to non-users were observed for HRQoL-categories happiness, anger, and social functioning. Together, these results show that, particularly for adult PKU-patients, HRQoL-problems are evident and that many of these problems are related to (history of) metabolic control. Beneficial effects of BH4-use appear to be limited to those associated with relief from the practical burdens related to the strict dietary treatment regimen, i.e. general mood and sociability, whereas metabolic control is more strongly related to basic physical and cognitive functioning.


Rates of Cognitive and Functional Impairments in Older Adults Residing in a Continuing Care Senior Housing Community.
The current cross-sectional study examined cognition and performance-based functional abilities in a continuing care senior housing community (CCSHC) that is comparable to other CCSHCs in the US with respect to residents' demographic characteristics. Participants were 110 older adult residents of the independent living unit. We assessed sociodemographics, mental health, neurocognitive functioning, and functional capacity. Compared to normative samples, participants performed at or above expectations in terms of premorbid functioning, attention span and working memory, processing speed, timed set-shifting, inhibitory control, and confrontation naming. They performed below expectation in verbal fluency and verbal and visual learning and memory, with impairment rates [31.4% (>1 SD below the mean) and 18.49% (>1.5 SD below the mean)] well above the general population (16% and 7%, respectively). Within the cognitive test battery, two tests of delayed memory were most predictive of a global deficit score. Most cognitive test scores correlated with performance-based functional capacity. Overall, results suggest that a subset of older adults in the independent living sector of CCSHCs are cognitively and functionally impaired and are at risk for future dementia. Results also argue for the inclusion of memory tests in abbreviated screening batteries in this population. We suggest that CCSHCs implement regular cognitive screening procedures to identify and triage those older adults who could benefit from interventions and, potentially, a transition to a higher level of care.


The predictive adaptive response: modeling the life-history evolution of the butterfly Bicyclus anynana in seasonal environments.
A predictive adaptive response (PAR) is a type of developmental plasticity where the response to an environmental cue is not immediately advantageous but instead is later in life. The PAR is a way for organisms to maximize fitness in varying environments. Insects living in seasonal environments are valuable model systems for testing the existence and form of PAR. Previous manipulations of the larval and the adult environments of the butterfly Bicyclus anynana have shown that individuals that were food restricted during the larval stage coped better with forced flight during the adult stage compared to those with optimal conditions in the larval stage. Here, we describe a state-dependent energy allocation model, which we use to test whether such a response to food restriction could be adaptive in nature where this butterfly exhibits seasonal cycles. The results from the model confirm the responses obtained in our previous experimental work and show how such an outcome was facilitated by resource allocation patterns to the thorax during the pupal stage. We conclude that for B. anynana, early-stage cues can direct development toward a better adapted phenotype later in life and, therefore, that a PAR has evolved in this species.


Road safety legislation in the Americas.
Legislating five of the main risk factors for road traffic injuries (RTIs), as much as enforcing the law, is essential in forging an integral culture of road safety. Analysis of the level of progression in law enforcement allows for an evaluation of the state of world regions. A secondary analysis of the 2009 Global status report on road safety: time for action survey was undertaken to evaluate legislation on five risk factors (speed management, drinking and driving, motorcycle helmet use, seatbelt use, and use of child restraints) in the Americas. Laws were classified depending on their level of progression: the existence of legislation, whether the legislation is adequate, a level of law enforcement > 6 (on a scale of 0-10), and whether the law is considered comprehensive. A descriptive analysis was performed. The totality of the countries has national or subnational legislation for at least one of the five risk factors. However, 63% have laws on the five risk factors studied, and none of them has comprehensive laws for all five. Seatbelt use appears to be the most extended enforced legislation, while speeding laws appear to be the least enforced. There are positive efforts that should be recognized in the region. However, the region stands in different stages of progression. Law enforcement remains the main issue to be tackled. Laws should be based on evidence about what is already known to be effective.


Fast label-free microscopy technique for 3D dynamic quantitative imaging of living cells.
The refractive index (RI) is an important optical characteristic that is often exploited in label-free microscopy for analysis of biological objects. A technique for 3D RI reconstruction of living cells has to be fast enough to capture the cell dynamics and preferably needs to be compatible with standard wide-field microscopes. To solve this challenging problem, we present a technique that provides fast measurement and processing of data required for real-time 3D visualization of the object RI. Specifically, the 3D RI is reconstructed from the measurement of bright-field intensity images, axially scanned by a high-speed focus tunable lens mounted in front of a sCMOS camera, by using a direct deconvolution approach designed for partially coherent light microscopy in the non-paraxial regime. Both the measurement system and the partially coherent illumination, that provides optical sectioning and speckle-noise suppression, enable compatibility with wide-field microscopes resulting in a competitive and affordable alternative to the current holographic laser microscopes. Our experimental demonstrations show video-rate 3D RI visualization of living bacteria both freely swimming and optically manipulated by using freestyle laser traps allowing for their trapping and transport along 3D trajectories. These results prove that is possible to conduct simultaneous 4D label-free quantitative imaging and optical manipulation of living cells, which is promising for the study of the cell biophysics and biology.


Limonene: a versatile chemical of the bioeconomy.
(+)-Limonene is a renewable chemical with numerous and growing applications. Its traditional uses such as flavor, fragrance and green solvent are rapidly expanding to include its utilization as a platform chemical, extraction solvent for natural products and an active agent for functionalized products. We anticipate that the expansion in uses for limonene will translate into increasing production and use of this relevant natural product, especially for advanced applications.


The effect of human mobility and control measures on the COVID-19 epidemic in China.
The ongoing COVID-19 outbreak has expanded rapidly throughout China. Major behavioral, clinical, and state interventions are underway currently to mitigate the epidemic and prevent the persistence of the virus in human populations in China and worldwide. It remains unclear how these unprecedented interventions, including travel restrictions, have affected COVID-19 spread in China. We use real-time mobility data from Wuhan and detailed case data including travel history to elucidate the role of case importation on transmission in cities across China and ascertain the impact of control measures. Early on, the spatial distribution of COVID-19 cases in China was well explained by human mobility data. Following the implementation of control measures, this correlation dropped and growth rates became negative in most locations, although shifts in the demographics of reported cases are still indicative of local chains of transmission outside Wuhan. This study shows that the drastic control measures implemented in China have substantially mitigated the spread of COVID-19.


Failures in top-down control in schizophrenia revealed by patterns of saccadic eye movements.
Successful execution of many behavioral goals relies on well-organized patterns of saccadic eye movements, and in complex tasks, these patterns can reveal the component processes underlying task performance. The present study examined the pattern of eye movements in a visual search task to provide evidence of attentional control impairments in people with schizophrenia (PSZ). We tested PSZ(N = 38) and nonpsychiatric control subjects (NCS, N = 35) in a task that was designed to stress top-down control by pitting task goals against bottom-up salience. Participants searched for either a low-contrast (nonsalient) or a high-contrast (salient) target among low- and high-contrast distractors. By examining fixations of the low- and high-contrast items, we evaluated the ability of PSZ and NCS to focus on low-salience targets and filter out high-salience distractors (or vice versa). When participants searched for a salient target, both groups successfully focused on relevant, high-contrast stimuli and filtered out target-mismatched, low-contrast stimuli. However, when searching for a nonsalient target, PSZ were impaired at efficiently suppressing high-contrast (salient) distractors. Specifically, PSZ were more likely than NCS to fixate and revisit salient distractors, and they dwelled on these items longer than did NCS. The results provide direct evidence that PSZ are impaired in their ability to utilize top-down goals to overcome the prepotent tendency to focus attention on irrelevant but highly salient information. (PsycINFO Database Record (c) 2019 APA, all rights reserved).


Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.
miR-122, an abundant liver-specific microRNA (miRNA), regulates cholesterol metabolism and promotes hepatitis C virus (HCV) replication. Reduced miR-122 expression in hepatocellular carcinoma (HCC) correlates with metastasis and poor prognosis. Nevertheless, the consequences of sustained loss of function of miR-122 in vivo have not been determined. Here, we demonstrate that deletion of mouse Mir122 resulted in hepatosteatosis, hepatitis, and the development of tumors resembling HCC. These pathologic manifestations were associated with hyperactivity of oncogenic pathways and hepatic infiltration of inflammatory cells that produce pro-tumorigenic cytokines, including IL-6 and TNF. Moreover, delivery of miR-122 to a MYC-driven mouse model of HCC strongly inhibited tumorigenesis, further supporting the tumor suppressor activity of this miRNA. These findings reveal critical functions for miR-122 in the maintenance of liver homeostasis and have important therapeutic implications, including the potential utility of miR-122 delivery for selected patients with HCC and the need for careful monitoring of patients receiving miR-122 inhibition therapy for HCV.


Colonization with Staphylococcus aureus and Klebsiella pneumoniae causes infections in a Vietnamese intensive care unit.
Pre-existing colonization with Staphylococcus aureus or Klebsiella pneumoniae has been found to increase the risk of infection in intensive care patients. We previously conducted a longitudinal study to characterize colonization of these two organisms in patients admitted to intensive care in a hospital in southern Vietnam. Here, using genomic and phylogenetic analyses, we aimed to assess the contribution these colonizing organisms made to infections. We found that in the majority of patients infected with S. aureus or K. pneumoniae, the sequence type of the disease-causing (infecting) isolate was identical to that of corresponding colonizing organisms in the respective patient. Further in-depth analysis revealed that in patients infected by S. aureus ST188 and by K. pneumoniae ST17, ST23, ST25 and ST86, the infecting isolate was closely related to and exhibited limited genetic variation relative to pre-infection colonizing isolates. Multidrug-resistant S. aureus ST188 was identified as the predominant agent of colonization and infection. Colonization and infection by K. pneumoniae were characterized by organisms with limited antimicrobial resistance profiles but extensive repertoires of virulence genes. Our findings augment the understanding of the link between bacterial colonization and infection in a low-resource setting, and could facilitate the development of novel evidence-based approaches to prevent and treat infections in high-risk patients in intensive care.


A mechanistically relevant cytotoxicity assay based on the detection of cellular GFP.
Cell-based assays for measuring ribosome inhibition by proteins such as the plant toxin ricin are important for characterizing decontamination strategies and developing detection technologies for field use. We report here an assay for ricin that provides a response that is relevant to the mechanism of ricin activity and permits a much faster readout than the commonly used assays for cytotoxicity. The assay relies on the response of an engineered reporter cell line that was produced by stably transfecting Vero cells to express green fluorescent protein (GFP) under the control ofa cytomegalovirus (CMV) promoter. The results of the GFP-based assay were compared with the assay results from three commercially available cytotoxicity assays. The GFP assay reports a sensitive response to ricin after 6 h of treatment while the other assays require a 24-h incubation. Unlike the other assays, monitoring cellular GFP on a per-cell basis allows detection of reduced ribosome activity before significant cell death occurs, and the results are not convoluted by the numbers of cells being assayed.


Prescription naloxone: a novel approach to heroin overdose prevention.
The mortality and morbidity from heroin overdose have increased in the United States and internationally in the last decade. The lipid solubility allows the rapid deposition of heroin and its metabolites into the central nervous system and accounts for the "rush" experienced by users and for the toxicity. Risk factors for fatal and nonfatal heroin overdoses such as recent abstinence, decreased opiate tolerance, and polydrug use have been identified. Opiate substitution treatment such as methadone or buprenorphine is the only proven method of heroin overdose prevention. Death from a heroin overdose most commonly occurs 1 to 3 hours after injection at home in the company of other people. Numerous communities have taken advantage of this opportunity for treatment by implementing overdose prevention education to active heroin users, as well as prescribing naloxone for home use. Naloxone is a specific opiate antagonist without agonist properties or potential for abuse. It is inexpensive and nonscheduled and readily reverses the respiratory depression and sedation caused by heroin, as well as causing transient withdrawal symptoms. Program implementation considerations, legal ramifications, and research needs for prescription naloxone are discussed.


Amiodarone hepatotoxicity.
Potential hepatotoxicity related to amiodarone therapy is often a concern when deciding whether to initiate or continue treatment with this medication. While mostly associated with long-term oral administration of the drug, toxicity has also been reported early during intravenous administration and months after discontinuation of therapy. In the majority of patients, it is discovered incidentally during routine testing of liver biochemistry and rarely do the hepatic effects develop into symptomatic liver injury or failure. Despite the widespread use of amiodarone, prospective clinical studies have been sparse and there has been little consensus among experts in the field regarding optimum monitoring for adverse effects in patients receiving this drug. In order to examine the current state of knowledge surrounding the incidence, pathogenesis and mechanism of liver effects associated with amiodarone, the existing literature was reviewed, with particular emphasis on clinical recommendations for monitoring.


Metabolisable Energy, In situ Rumen Degradation and In vitro Fermentation Characteristics of Linted Cottonseed Hulls, Delinted Cottonseed Hulls and Cottonseed Linter Residue.
Dietary supplementation with conventional linted cottonseed hulls (LCSH) is a common practice in livestock production all over the world. However, supplementation with mechanically delinted cottonseed hulls (DCSH) and cottonseed linter residue (CLR) is uncommon. Cottonseed by-products, including LCSH, DCSH and CLR, were assessed by chemical analysis, an in situ nylon bag technique, an in vitro cumulative gas production technique and in vitro enzyme procedure. The crude protein (CP) content of CLR (302 g/kg dry matter (DM)) was approximately 3 times that of LCSH and 5 times that of DCSH. The crude fat content was approximately 3 times higher in CLR (269 g/kg DM) than in LCSH and 4 times higher than in DCSH. Neutral detergent fibre (311 g/kg DM) and acid detergent fibre (243 g/kg DM) contents of CLR were less than half those of DCSH or LCSH. Metabolisable energy, estimated by in vitro gas production and chemical analyses, ranked as follows: CLR (12.69 kJ/kg DM)>LCSH (7.32 kJ/kg DM)>DCSH (5.82 kJ/kg DM). The in situ degradation trial showed that the highest values of effective degradability of DM and CP were obtained for CLR (p<0.05). The in vitro disappearance of ruminal DM ranked as follows: CLR>LCSH>DCSH (p<0.05). The lowest digestibility was observed for DCSH with a two-step in vitro digestion procedure (p<0.05). The potential gas production in the batch cultures did not differ for any of the three cottonseed by-product feeds. The highest concentration of total volatile fatty acids was observed in CLR after a 72 h incubation (p<0.05). The molar portions of methane were similar between all three treatments, with an average gas production of 22% (molar). The CLR contained a higher level of CP than did LCSH and DCSH, and CLR fermentation produced more propionate. The DCSH and LCSH had more NDF and ADF, which fermented into greater amounts of acetate.


Diode-laser-pumped Nd:YAG laser injection seeding system.
We have designed and tested a compact injection seeding system consisting of a diode-laser-pumped Nd:YAG master oscillator and a permanent-magnet Faraday isolator. With active resonator frequency stabilization, this system permits highly reliable single-axial-mode operation of a Q-switched Nd:YAG laser over a period of hours. The system is capable of injection seeding both stable and unstable resonator designs and is suitable for injection seeding commercial lasers with only minor modifications.