Cholera toxin enhances vaccine-induced protection against Mycobacterium tuberculosis challenge in mice.
Interleukin (IL)-17 is emerging as an important cytokine in vaccine-induced protection against tuberculosis disease in animal models. Here we show that compared to parenteral delivery, BCG delivered mucosally enhances cytokine production, including interferon gamma and IL-17, in the lungs. Furthermore, we find that cholera toxin, delivered mucosally along with BCG, further enhances IL-17 production by CD4(+) T cells over mucosal BCG alone both in the lungs and systemically. This boosting effect of CT is also observed using a vaccine regimen of BCG followed by the candidate vaccine MVA85A. Using a murine Mycobacterium tuberculosis (M.tb) aerosol challenge model, we demonstrate the ability of cholera toxin delivered at the time of a priming BCG vaccination to improve protection against tuberculosis disease in a manner at least partially dependent on the observed increase in IL-17. This observed increase in IL-17 in the lungs has no adverse effect on lung pathology following M.tb challenge, indicating that IL-17 can safely be boosted in murine lungs in a vaccine/M.tb challenge setting.


Conservation, Duplication, and Divergence of Five Opsin Genes in Insect Evolution.
Opsin proteins covalently bind to small molecular chromophores and each protein-chromophore complex is sensitive to particular wavelengths of light. Multiple opsins with different wavelength absorbance peaks are required for color vision. Comparing opsin responses is challenging at low light levels, explaining why color vision is often lost in nocturnal species. Here, we investigated opsin evolution in 27 phylogenetically diverse insect species including several transitions between photic niches (nocturnal, diurnal, and crepuscular). We find widespread conservation of five distinct opsin genes, more than commonly considered. These comprise one c-opsin plus four r-opsins (long wavelength sensitive or LWS, blue sensitive, ultra violet [UV] sensitive and the often overlooked Rh7 gene). Several recent opsin gene duplications are also detected. The diversity of opsin genes is consistent with color vision in diurnal, crepuscular, and nocturnal insects. Tests for positive selection in relation to photic niche reveal evidence for adaptive evolution in UV-sensitive opsins in day-flying insects in general, and in LWS opsins of day-flying Lepidoptera specifically.


Effect of increased warfarin use on warfarin-related cerebral hemorrhage: a longitudinal population-based study.
Warfarin use has rapidly increased with the aging of the population. We investigated the temporal trends in the incidence and outcome of warfarin-related intracerebral hemorrhages (ICHs) in a defined population. We identified all subjects with first-ever primary ICH during 1993 to 2008 among the population of Northern Ostrobothnia, Finland. The number of warfarin users was obtained from the national register of prescribed medicines kept by the Social Insurance Institution of Finland. We calculated the annual incidence of warfarin-related ICHs, 28-day case fatality, and deaths from the primary bleed. The proportion of warfarin users among the population increased 3.6-fold from 0.68% in 1993 to 2.28% in 2008. Of a total of 982 patients with ICH, 182 (18.5%) had warfarin-related ICH. One-year survival rate after onset of stroke was 35.2% among warfarin users and 67.9% among nonusers. The annual incidence (P=0.062) and 28-day case fatality of warfarin-related ICHs (P=0.002) decreased during the observation period. Warfarin users were older (mean difference 6.6; 95% CI, 5.0 to 8.1; P<0.001) than nonusers. Admission international normalized ratio values above the therapeutic range (2.0 to 3.0) decreased through the observation period, suggesting improved control of anticoagulant therapy over time. The annual incidence and case fatality of warfarin-related ICHs decreased, although the proportion of warfarin users almost quadrupled in our population.


A low-fat diet enriched in fish oil increased lipogenesis and fetal outcome of C57BL/6 mice.
There is clear evidence that nutritional strategy employed during pregnancy has profound influence on the offspring health outcomes. However, the effect of the quality and the quantity of maternal fat intake on maternal metabolic profile during different stages of pregnancy and its impact on pregnancy sustainability is not known. Female C57BL/6 mice (7 weeks old) were fed diets varying in the quantity of fat (5% vs 11%) for two weeks prior to mating and throughout pregnancy. The 5% fat diet was enriched with longer chain omega (n)-3 polyunsaturated fatty acids (PUFA) from fish oil. Maternal plasma and tissues were collected before mating and during pregnancy at days 6.5, 12.5 and 18.5. Plasma lipids, glucose, insulin, progesterone and estradiol levels were measured. Cholesterol efflux capacity of maternal plasma as well as the mRNA expression of placental steroidogenic acute regulatory protein and hepatic lipogenic genes (acetyl-CoA carboxylase-1, fatty acid synthase, diacylglycerol acyltransferase-2 and stearoyl-CoA desaturase-1) was determined. Feto-placental weight and fetuses sustained throughout gestation were recorded. A low-fat maternal diet enriched with n-3 PUFA increased maternal plasma triacylglycerol and the mRNA expression of rate-limiting lipogenic enzymes, along with increasing cholesterol efflux capacity (P < 0.05), likely to meet fetal lipid demand during pregnancy. Furthermore, diet enriched with longer chain n-3 PUFA increased the maternal plasma concentration of progesterone and estradiol during pregnancy (P < 0.05), which coincides with an increase in the number of fetuses sustained till day 18.5. These novel findings may be important when designing dietary strategies to optimize reproductive capability and pregnancy outcomes.


Beyond the Exome: The Non-coding Genome and Enhancers in Neurodevelopmental Disorders and Malformations of Cortical Development.
The development of the human cerebral cortex is a complex and dynamic process, in which neural stem cell proliferation, neuronal migration, and post-migratory neuronal organization need to occur in a well-organized fashion. Alterations at any of these crucial stages can result in malformations of cortical development (MCDs), a group of genetically heterogeneous neurodevelopmental disorders that present with developmental delay, intellectual disability and epilepsy. Recent progress in genetic technologies, such as next generation sequencing, most often focusing on all protein-coding exons (e.g., whole exome sequencing), allowed the discovery of more than a 100 genes associated with various types of MCDs. Although this has considerably increased the diagnostic yield, most MCD cases remain unexplained. As Whole Exome Sequencing investigates only a minor part of the human genome (1-2%), it is likely that patients, in which no disease-causing mutation has been identified, could harbor mutations in genomic regions beyond the exome. Even though functional annotation of non-coding regions is still lagging behind that of protein-coding genes, tremendous progress has been made in the field of gene regulation. One group of non-coding regulatory regions are enhancers, which can be distantly located upstream or downstream of genes and which can mediate temporal and tissue-specific transcriptional control via long-distance interactions with promoter regions. Although some examples exist in literature that link alterations of enhancers to genetic disorders, a widespread appreciation of the putative roles of these sequences in MCDs is still lacking. Here, we summarize the current state of knowledge on cis-regulatory regions and discuss novel technologies such as massively-parallel reporter assay systems, CRISPR-Cas9-based screens and computational approaches that help to further elucidate the emerging role of the non-coding genome in disease. Moreover, we discuss existing literature on mutations or copy number alterations of regulatory regions involved in brain development. We foresee that the future implementation of the knowledge obtained through ongoing gene regulation studies will benefit patients and will provide an explanation to part of the missing heritability of MCDs and other genetic disorders.


Border formation in a Bmp gradient reduced to single dissociated cells.
Conversions of signaling gradients into sharp "all-or-none" borders are fundamental to tissue and organismal development. However, whether such conversions can be meaningfully reduced to dissociated cells in culture has been uncertain. Here we describe ultrasensitivity, the phenomenon equivalent to an all-or-none response, in dissociated neural precursor cells (NPCs) exposed to bone morphogenetic protein 4 (Bmp4). NPC ultrasensitivity is evident at the population and single-cell levels based on Msx1 induction, a well known Bmp target response, and occurs in the context of gene expression changes consistent with Bmp4 activity as a morphogen. Dissociated NPCs also display immediate early kinetics and irreversibility for Msx1 induction after brief Bmp4 exposure, which are attractive features for initial border formation. Relevance to border formation in vivo is provided by Bmp4 gain-of-function studies in explants and evidence for single-cell ultrasensitivity in normal and mutant Bmp gradient contexts in the developing forebrain. Together, these studies demonstrate relatively simple, robust, and reducible cell-intrinsic properties that contribute to developmental border formation within a signaling gradient.


Clinical presentation, processes and outcomes of care for patients with pneumococcal pneumonia.
To describe the presentation, resolution of symptoms, processes of care, and outcomes of pneumococcal pneumonia, and to compare features of the bacteremic and nonbacteremic forms of this illness. A prospective cohort study. Five medical institutions in 3 geographic locations. Inpatients and outpatients with community-acquired pneumonia (CAP). Sociodemographic characteristics, respiratory and nonrespiratory symptoms, and physical examination findings were obtained from interviews or chart review. Severity of illness was assessed using a validated prediction rule for short-term mortality in CAP. Pneumococcal pneumonia was categorized as bacteremic; nonbacteremic, pure etiology; or nonbacteremic, mixed etiology. One hundred fifty-eight (6.9%) of 2,287 patients (944 outpatients, 1,343 inpatients) with CAP had pneumococcal pneumonia. Sixty-five (41%) of the 158 with pneumococcal pneumonia were bacteremic; 74 (47%) were nonbacteremic with S. pneumoniae as sole pathogen; and 19 (12%) were nonbacteremic with S. pneumoniae as one of multiple pathogens. The pneumococcal bacteremia rate for outpatients was 2.6% and for inpatients it was 6.6%. Cough, dyspnea, and pleuritic pain were common respiratory symptoms. Hemoptysis occurred in 16% to 22% of the patients. A large number of nonrespiratory symptoms were noted. Bacteremic patients were less likely than nonbacteremic patients to have sputum production and myalgias (60% vs 82% and 33% vs 57%, respectively; P <.01 for both), more likely to have elevated blood urea nitrogen and serum creatinine levels, and more likely to receive penicillin therapy. Half of bacteremic patients were in the low risk category for short-term mortality (groups I to III), similar to the nonbacteremic patients. None of the 32 bacteremic patients in risk groups I to III died, while 7 of 23 (30%) in risk group V died. Intensive care unit admissions and pneumonia-related mortality were similar between bacteremic and nonbacteremic groups, although 46% of the bacteremic group had respiratory failure compared with 32% and 37% for the other groups. The nonbacteremic pure etiology patients returned to household activities faster than bacteremic patients. Symptoms frequently persisted at 30 days: cough (50%); dyspnea (53%); sputum production (48%); pleuritic pain (13%); and fatigue (63%). There were few differences in the presentation of bacteremic and nonbacteremic pneumococcal pneumonia. About half of bacteremic pneumococcal pneumonia patients were at low risk for mortality. Symptom resolution frequently was slow.


Normalized left ventricular workload using phase-contrast magnetic resonance imaging in patients with aortic stenosis.
Aortic stenosis (AS) severity contributes to the left ventricle (LV) deterioration due to the aortic valve narrowing and the alteration of systemic hemodynamic load. This load increment may also increase the LV stroke work (SW) which represent the required energy to deliver the blood at ejection. In this study, SW was derived from in-vivo cardiovascular magnetic resonance (CMR) velocity measurements (n=57) using a lumped-parametric model. Furthermore, normalized SW (N-SW) was evaluated as AS severity parameter. SW differentiated from normal flow (>35 mL/m(2)) and low flow (<35 mL/m(2)) states (p<0.05). N-SW showed a good association with valve effective orifice area (EOA, r=-0.5, p<0.001) and valvulo-arterial impedance (ZVA, r=0.65, p<0.001). A severity threshold for N-SW (1.5 cJ/mL) was found using an EOA=1 cm(2) as AS severity marker. CMR-derived SW and N-SW may be useful to the assessment and grading of AS patients.


Clearing asymptomatic parasitaemia increases the specificity of the definition of mild febrile malaria.
In clinical trials, the specificity of the disease endpoint is critical to an accurate estimate of vaccine efficacy. We used a logistic regression model to analyse parasite densities among children before and after treatment with antimalarials, in order to estimate the impact that clearing asymptomatic parasitaemia had on the specificity of the endpoint of febrile malaria. The malaria attributable fever fraction was higher after antimalarial treatment (i.e. fever and parasitaemia were more likely to be causally related), implying that drug treatment prior to monitoring decreased the misclassification of febrile malaria. In intervention studies with febrile malaria as an endpoint, clearing asymptomatic parasitaemia increases the study's power more effectively than raising the threshold parasitaemia.


Genetic control and evolutionary potential of a constitutive resistance mechanism against the spruce budworm (Choristoneura fumiferana) in white spruce (Picea glauca).
Insect herbivory may drive evolution by selecting for trees with heritable resistance against defoliation. The spruce budworm (Choristoneura fumiferana, SBW) is a highly damaging forest insect pest that can affect population structure of white spruce (Picea glauca) in North America. Resistance against SBW was recently described in white spruce and was linked to three constitutive resistance biomarkers: the phenolic compounds piceol and pungenol, and expression of a beta-glucosidase encoding gene (Pgβglu-1). We investigated the phenotypic variability and heritability of these resistance biomarkers and of picein, the precursor of piceol, in the foliage of 874 trees belonging to 33 full-sib families and 71 clonal lines under evaluation in seven field locations in Eastern Canada. We aimed to (i) determine their genetic control, (ii) estimate the genetic and phenotypic correlations among defense biomarkers, and (iii) determine whether their constitutive levels are associated with detrimental trade-offs on growth. Quantitative genetics analyses indicated that all four traits are moderately to highly heritable. The full-sib and clonal analyses showed that additive and non-additive genetic effects play major and minor roles, respectively. Positive genetic and phenotypic correlations between resistance biomarkers and primary growth indicated that there is no trade-off between total height and height increment and resistance traits, contradicting the GDBH (Growth Differentiation Balance Hypothesis). Our findings about the predominant additive genetic basis of the resistance biomarkers show that adaptive evolution of white spruce natural populations to resist to SBW is possible and that potentially important gains could also be expected from artificial selection.


Fetal outcomes of critically ill pregnant women admitted to the intensive care unit for nonobstetric causes.
The outcome of the fetus in critically ill mothers has been briefly reported as a part of descriptive studies focusing on maternal risk factors for admission to the intensive care unit. We evaluated the risk factors for adverse fetal outcomes in critically ill pregnant women admitted to the intensive care unit for nonobstetrical reasons. Retrospective cohort study of all critically ill pregnant patients >18 yr; admitted to four (medical, surgical, trauma, and mixed medical-surgical) intensive care units at the Mayo Clinic in Rochester, MN; during the period of January 1995 to December 2005. Only pregnant women admitted to the intensive care unit in the antepartum period for nonobstetrical indications were included. Main predictors for fetal outcomes included: maternal comorbidities, obstetrical history, intensive care unit interventions, and intensive care unit complications. Fetal outcomes were defined as spontaneous abortions, neonatal mortality, fetal deaths, admission to the neonatal intensive care unit, neonatal intensive care unit length of stay, and neonatal intensive care unit complications. A total of 153 adult women (>18 yr) with a diagnosis of pregnancy were admitted to the intensive care unit, of whom 93 pregnant women met the inclusion criteria. Median maternal age was 26 yr (interquartile range 22-33) and median gestational age was 25 wk (interquartile range 8-33). The median maternal Acute Physiologic and Chronic Health Evaluation III score was 27 (interquartile range 17-38). There were 32 fetal losses; 18 were spontaneous abortions and 14 were fetal deaths. Ten neonates required neonatal intensive care unit admission, five for respiratory distress syndrome; and only one neonate died. The median neonatal intensive care unit length of stay was 34 days (interquartile range 15-87). After multivariable logistic regression analysis, the risk factors associated with fetal loss were: presence of maternal shock, odds ratio 6.85 (95% confidence interval 1.16-58, p = 0.04); maternal transfusion of blood products, odds ratio 7.24 (95% confidence interval 1.4-49, p = 0.02); and gestational age, odds ratio 1.2 for every gestational week below 37 wk (95% confidence interval 1.1-1.3, p < 0.001). Nonobstetrical critical illness in pregnant women significantly affects fetal and neonatal outcomes. Maternal shock, maternal requirement of allogenic blood product transfusion and lower gestational age were associated with an increased risk of fetal loss.


Effect of Cryopreservation on Human Adipose Tissue and Isolated Stromal Vascular Fraction Cells: In Vitro and In Vivo Analyses.
Adipose tissue is a source of adipose-derived stromal/stem cells for tissue engineering and reconstruction and a tissue source for fat grafts. Although liposuction is a simple procedure for the harvest of adipose tissue, the repetition of this surgical intervention can cause adverse effects to the patient and can be a limiting factor for immediate use. Cryopreservation can avoid the morbidity associated with repetitive liposuction, allowing the use of stored tissue after the initial harvest procedure. This article focuses on the characterization of fresh and cryopreserved human adipose tissue. Lipoaspirates from eight donors were processed as fresh adipose tissue or cryopreserved for 4 to 6 weeks. Fresh and cryopreserved tissues were collagenase digested and the stromal vascular fraction cells were characterized immediately or cryopreserved. Characterization was based on stromal vascular fraction cell proliferation and immunophenotype. In vivo fat grafting was performed in C57BL/6 green fluorescent protein mice to analyze morphology of the tissue and its adiposity using confocal microscopy, histochemical staining (i.e., hematoxylin and eosin and Masson trichrome), and immunohistochemistry (i.e., green fluorescent protein, perilipin, and CD31). Although tissue and stromal vascular fraction cell cryopreservation reduced the total cell yield, the remaining viable cells retained their adhesive and proliferative properties. The stromal vascular fraction cell immunophenotype showed a significant reduction in the hematopoietic surface markers and increased expression of stromal and adipogenic markers following cryopreservation. In vivo cryopreserved fat grafts showed morphology similar to that of freshly implanted fat grafts. In this study, the authors demonstrated that cryopreserved adipose tissue is a potential source of stromal vascular fraction cells and a suitable source for fat grafts.


Water deficit improved the capacity of arbuscular mycorrhizal fungi (AMF) for inducing the accumulation of antioxidant compounds in lettuce leaves.
Lettuce, a major food crop within the European Union and the most used for the so-called 'Fourth Range' of vegetables, can associate with arbuscular mycorrhizal fungi (AMF). Mycorrhizal symbiosis can stimulate the synthesis of secondary metabolites, which may increase plant tolerance to stresses and enhance the accumulation of antioxidant compounds potentially beneficial to human health. Our objectives were to assess (1) if the application of a commercial formulation of AMF benefited growth of lettuce under different types and degrees of water deficits; (2) if water restrictions affected the nutritional quality of lettuce; and (3) if AMF improved the quality of lettuce when plants grew under reduced irrigation. Two cultivars of lettuce consumed as salads, Batavia Rubia Munguía and Maravilla de Verano, were used in the study. Four different water regimes were applied to both non-mycorrhizal and mycorrhizal plants: optimal irrigation (field capacity [FC]), a water regime equivalent to 2/3 of FC, a water regime equivalent to 1/2 of FC and a cyclic drought (CD). Results showed that mycorrhizal symbiosis improved the accumulation of antioxidant compounds, mainly carotenoids and anthocyanins, and to a lesser extent chlorophylls and phenolics, in leaves of lettuce. These enhancements were higher under water deficit than under optimal irrigation. Moreover, shoot biomass in mycorrhizal lettuces subjected to 2/3 of FC were similar to those of non-mycorrhizal plants cultivated under well-watered conditions. In addition, lettuces subjected to 2/3 FC had similar leaf RWC than their respective well-watered controls, regardless of mycorrhizal inoculation. Therefore, results suggest that mycorrhizal symbiosis can improve quality of lettuce and may allow restrict irrigation without reducing production.


Alteration in diel activity patterns as a thermoregulatory strategy in black wildebeest (Connochaetes gnou).
The nychthemeral activity patterns of a population of female black wildebeest inhabiting a shadeless environment were surveyed periodically over 1 year. The wildebeest fed mostly at night, with the proportion of feeding at night increasing when ambient conditions were hotter. Inactive periods were spent mostly lying during cooler weather but standing as days became hotter. We suggest that the entire suite of behavioural adjustments is beneficial to heat exchange with the environment. Behaviour patterns were markedly different during one warm weather survey, from the other warm weather surveys, when an 8-month dry spell had just been broken. We suggest that this may reflect the availability of water for autonomic thermoregulation, a consequent decreased reliance on behavioural thermoregulation, and a release of the thermal constraints on foraging. Our results help to explain the ability of black wildebeest to maintain body core temperature within a very narrow range despite being exposed to an environment with large nychthemeral variations in thermal conditions and offering little in the way of microclimate selection.


Comparative analysis of allyl isothiocyanate (AITC)-induced carbohydrate oxidation changes via TRPV1 between mice and chickens.
Postprandial hyperglycemia is a risk factor for cardiovascular diseases. It has been reported that intragastric administration of allyl isothiocyanate (AITC), which is one of the pungent ingredients of wasabi and horseradish but it is not included in hot chili pepper, increased carbohydrate oxidation and reduced postprandial increase of blood glucose via transient receptor potential vanilloid 1 (TRPV1)in mice. However, the action site of AITC on TRPV1 for increasing carbohydrate oxidation is unclear. Both mammalian and chicken TRPV1 (cTRPV1) are activated by heat and acid, but unlike its mammalian counterpart, cTRPV1 is only faintly activated by capsaicin. This difference is due to the 8 chicken-specific amino acid residues around transmembrane 3, which is the main site of capsaicin-binding in rat TRPV1. Moreover, AITC-induced activation of mouse TRPV1 (mTRPV1) is largely dependent on S513, a residue that is involved in capsaicin-binding. Thus, we hypothesized that the increase of carbohydrate oxidation by AITC in mammals is induced by the binding of AITC to the capsaicin-binding site of TRPV1. In this study, we performed a comparative study using chickens and mice, since chickens are thought to partly lack the capsaicin-binding site of TRPV1. We examined the effects of AITC on the respiratory quotient (RQ), the index of carbohydrate oxidation and fat oxidation, in chickens and mice. Respiratory gas analysis revealed that AITC does not increase the RQ in chickens, and Ca2+ imaging methods and a whole cell-patch clamp analysis showed that AITC does not activate cTRPV1. These results implied that the capsaicin-binding site is an important region for increasing carbohydrate oxidation by AITC administration in animals.


Characterization of the elongating alpha-D-mannosyl phosphate transferase from three species of Leishmania using synthetic acceptor substrate analogues.
Leishmania express lipophosphoglycans and proteophosphoglycans that contain Galbeta1-4Manalpha1-P phosphosaccharide repeat structures assembled by the sequential addition of Manalpha1-P and betaGal. The synthetic acceptor substrate Galbeta1-4Manalpha1-P-decenyl and a series of analogues were used to probe Leishmania alpha-D-mannosyl phosphate transferase activity. We show that the activity detected with Galbeta1-4Manalpha1-P-decenyl is the elongating alpha-D-mannosyl phosphate transferase associated with lipophosphoglycan biosynthesis (eMPT(LPG)). Differences in the apparent K(m) values for the donor and acceptor substrates were found using L. major, L. mexicana, and L. donovani promastigote membranes, but total activity correlated with the number of lipophosphoglycan repeats. Further comparisons showed that lesion-derived L. mexicana amastigotes, that do not express lipophosphoglycan, lack eMPT(LPG) and that nondividing L. major metacyclic promastigotes contain 5-fold less eMPT(LPG) activity than dividing procyclic promastigotes. The fine specificity of promastigote eMPT(LPG) activity was determined using 24 synthetic analogues of Galbeta1-4Manalpha1-P-decenyl. The three species gave similar results: the negative charge of the phosphodiester and the C-6 hydroxyl of the alphaMan residue are essential for substrate recognition, the latter most likely acting as a hydrogen bond acceptor. The C-6' hydroxyl of the betaGal residue is required for substrate recognition as well as for catalysis. The rate of Manalpha1-P transfer declines with increasing acceptor substrate chain length. The presence of a monosaccharide substituent at the C-3 position of the terminal betaGal residue abrogates Man-P transfer, showing that chain elongation must precede side chain modification during lipophosphoglycan biosynthesis. In contrast, substitution of the penultimate phosphosaccharide repeat does not abrogate transfer but is slightly stimulatory in L. mexicana and inhibitory in L. major.


A manganese transporter, BB0219 (BmtA), is required for virulence by the Lyme disease spirochete, Borrelia burgdorferi.
Borrelia burgdorferi (Bb), the causative agent of Lyme disease, is transmitted to mammalian hosts through an arthropod (tick) vector. To establish infection, Bb must acquire essential nutrients, including transition metals, from its mammalian and tick hosts. Thus far, no metal transporter has been identified in Bb. Here, we report the identification of the first metal transporter, BmtA (BB0219), in Bb. BmtA-deficient mutants of virulent Bb were readily generated, and the mutants grew slightly slower than wild-type Bb in vitro. However, BmtA mutants were sensitive to the chelating actions of EDTA, suggesting a role for BmtA in metal utilization. Intracellular accumulation of manganese (Mn) was substantially diminished in the bmtA mutant, indicating that BmtA was operative in Mn uptake. Given that BmtA lacks homology to any known Mn transporter, we postulate that BmtA is part of a novel mechanism for Mn acquisition by a bacterial pathogen. BmtA also was essential to the infectious life cycle of Bb in ticks and mammals, thereby qualifying BmtA as a new borrelial virulence factor. In addition, the bmtA mutant was sensitive to treatment with t-butyl hydroperoxide, implying that BmtA, and thus Mn, is important to Bb for detoxifying reactive oxygen species, including those potentially liberated by immune effector cells during the innate immune response. Our discovery of the first molecule involved in metal transport in Bb provides a foundation for further elucidating metal homeostasis in this important human pathogen, which may lead to new strategies for thwarting Lyme disease.


Biological Processes Highlighted in Saccharomyces cerevisiae during the Sparkling Wines Elaboration.
Sparkling wines elaboration has been studied by several research groups, but this is the first report on analysis of biological processes according to the Gene Ontology terms (GO terms) and related to proteins expressed by yeast cells during the second fermentation of sparkling wines. This work provides a comprehensive study of the most relevant biological processes in Saccharomyces cerevisiae P29, a sparkling wine strain, during the second fermentation under two conditions (without and with endogenous CO2 overpressure) in the middle and the end of second fermentation. Consequently, a proteomic analysis with the OFFGEL fractionator and protein identification with LTQ Orbitrap XL coupled to HPLC were performed. The classification of biological processes was carried out using the tools provided by the Saccharomyces Genome Database. Results indicate that a greater number of biological processes were identified under condition without CO2 overpressure and in the middle of the fermentation versus the end of the second fermentation. The biological processes highlighted under condition without CO2 overpressure in the middle of the fermentation were involved in the carbohydrate and lipid metabolic processes and catabolic and biosynthetic processes. However, under CO2 overpressure, specific protein expression in response to stress, transport, translation, and chromosome organization and specific processes were not found. At the end of fermentation, there were higher specific processes under condition without CO2 overpressure; most were related to cell division, growth, biosynthetic process, and gene transcription resulting in increased cell viability in this condition. Under CO2 overpressure condition, the most representative processes were related to translation as tRNA metabolic process, chromosome organization, mRNA processing, ribosome biogenesis, and ribonucleoprotein complex assembly, probably in response to the stress caused by the hard fermentation conditions. Therefore, a broader knowledge of the adaptation of the yeast, and its behavior under typical conditions to produce sparkling wine, might improve and favor the wine industry and the selection of yeast for obtaining a high-quality wine.


Different brain circuits underlie motor and perceptual representations of temporal intervals.
In everyday life, temporal information is used for both perception and action, but whether these two functions reflect the operation of similar or different neural circuits is unclear. We used functional magnetic resonance imaging to investigate the neural correlates of processing temporal information when either a motor or a perceptual representation is used. Participants viewed two identical sequences of visual stimuli and used the information differently to perform either a temporal reproduction or a temporal estimation task. By comparing brain activity evoked by these tasks and control conditions, we explored commonalities and differences in brain areas involved in reproduction and estimation of temporal intervals. The basal ganglia and the cerebellum were commonly active in both temporal tasks, consistent with suggestions that perception and production of time are subserved by the same mechanisms. However, only in the reproduction task was activity observed in a wider cortical network including the right pre-SMA, left middle frontal gyrus, left premotor cortex, with a more reliable activity in the right inferior parietal cortex, left fusiform gyrus, and the right extrastriate visual area V5/MT. Our findings point to a role for the parietal cortex as an interface between sensory and motor processes and suggest that it may be a key node in translation of temporal information into action. Furthermore, we discuss the potential importance of the extrastriate cortex in processing visual time in the context of recent findings.


Correlation between gestational age and level of sensory block in spinal anesthesia.
To identify a specific correlation between progression of gestational age and level of sensory block in spinal anesthesia in patients undergoing cervical cerclage. The medical records of patients who underwent cervical cerclage under spinal anesthesia using hyperbaric bupivacaine between March 2016 and May 2018 were retrospectively reviewed. The primary outcome measure was the correlation between gestational age and sensory block level of spinal anesthesia. Secondary outcomes included the correlations between gestational age and reduction in blood pressure and length of the postanesthesia care unit (PACU) stay. The Pearson correlation test, partial correlation test, and linear regression were used to identify relationships and to adjust for confounding variables. Of the 261 cases reviewed, there was a linear and significant correlation between gestational age and sensory block level, reduction in systolic blood pressure during surgery, reduction in systolic blood pressure during PACU, and length of PACU stay, after adjusting for confounding factors (partial correlation coefficient=0.71, p<0.001; 0.27, p<0.001; 0.25, p<0.001; 0.21, p=0.001, respectively). A 1-day increase in gestational age corresponded to an increase of 0.04 dermatomes in sensory block level (95% CI 0.035 to 0.045, p<0.001). Results demonstrated that as the gestational age progressed, sensory block level increased, blood pressure was decreased, and length of PACU stay increased after spinal anesthesia. Sensory block increased by 0.28 dermatomes when the pregnancy progressed 1 week. Further controlled prospective studies are needed to support this relationship. KCT0000538.


Synergistic activation of the fibroblast growth factor 4 enhancer by Sox2 and Oct-3 depends on protein-protein interactions facilitated by a specific spatial arrangement of factor binding sites.
Octamer binding and Sox factors are thought to play important roles in development by potentiating the transcriptional activation of specific gene subsets. The proteins within these factor families are related by the presence of highly conserved DNA binding domains, the octamer binding protein POU domain or the Sox factors HMG domain. We have previously shown that fibroblast growth factor 4 (FGF-4) gene expression in embryonal carcinoma cells requires a synergistic interaction between Oct-3 and Sox2 on the FGF-4 enhancer. Sox2 and Oct-3 bind to adjacent sites within this enhancer to form a ternary protein-DNA complex (Oct-3*) whose assembly correlates with enhancer activity. We now demonstrate that increasing the distance between the octamer and Sox binding sites by base pair insertion results in a loss of enhancer function. Significantly, those enhancer "spacing mutants" which failed to activate transcription were also compromised in their ability to form the Oct* complexes even though they could still bind both Sox2 and the octamer binding proteins, suggesting that a direct interaction between Sox2 and Oct-3 is necessary for enhancer function. Consistent with this hypothesis, Oct-3 and Sox2 can participate in a direct protein-protein interaction in vitro in the absence of DNA, and both this interaction and assembly of the ternary Oct* complexes require only the octamer protein POU and Sox2 HMG domains. Assembly of the ternary complex by these two protein domains occurs in a cooperative manner on FGF-4 enhancer DNA, and the loss of this cooperative interaction contributes to the defect in Oct-3* formation observed for the enhancer spacing mutants. These observations indicate that Oct-3* assembly results from protein-protein interactions between the domains of Sox2 and Oct-3 that mediate their binding to DNA, but it also requires a specific arrangement of the binding sites within the FGF-4 enhancer DNA. Thus, these results define one parameter that is fundamental to synergistic activation by Sox2 and Oct-3 and further emphasize the critical role of enhancer DNA sequences in the proper assembly of functional activation complexes.


Quantitative electroencephalography: preliminary report--tinnitus.
This preliminary report is an account of 21 consecutive patients who had tinnitus of the severe disabling type and were examined with quantitative electroencephalography (QEEG). A multimetric analysis of the raw data was highlighted by an abnormal incidence of significant central nervous system electrical dysfunction identified in each patient (21 of 21). Relative power was increased or decreased in the temporal region in 10 of 21 patients. Relative power was reported to be increased or decreased in temporal frontal regions in 20 of 21 patients. Coherence irregularity was identified in all 21 patients. QEEG preliminary data support the hypothesis of a final common pathway for tinnitus and the significant role of the temporal and temporofrontal regions of interest in patients with tinnitus of the severe disabling type. QEEG with multimetric analysis is considered a significant addition to the medical audiological tinnitus patient protocol as one of a battery of electrophysiological tests for the clinical identification of a predominantly central type of tinnitus.


Interactive perceptual and attentional limits in visual extinction.
We report a case-study of YE, a 54-year-old person who suffered multiple shell injuries that caused a right-parietal lesion and left hemianopia, almost 30 years ago. We conducted 4 experiments using a basic extinction paradigm in which YE had to report single stimuli presented on the left or right or two stimuli presented simultaneously in both visual fields. We show that extinction was selectively affected both by increasing the relative perceptual salience of the contralesional stimulus and by cueing attention to the contralesional side. The effects of perceptual salience and attentional cueing interacted, with cueing being more effective when the stimuli had relatively high perceptual salience. The data are consistent with attentional and perceptual factors interacting to determine the competition between left and right side stimuli that underlies extinction.


Resonant Zener tunnelling via zero-dimensional states in a narrow gap diode.
Interband tunnelling of carriers through a forbidden energy gap, known as Zener tunnelling, is a phenomenon of fundamental and technological interest. Its experimental observation in the Esaki p-n semiconductor diode has led to the first demonstration and exploitation of quantum tunnelling in a condensed matter system. Here we demonstrate a new type of Zener tunnelling that involves the resonant transmission of electrons through zero-dimensional (0D) states. In our devices, a narrow quantum well of the mid-infrared (MIR) alloy In(AsN) is placed in the intrinsic (i) layer of a p-i-n diode. The incorporation of nitrogen in the quantum well creates 0D states that are localized on nanometer lengthscales. These levels provide intermediate states that act as "stepping stones" for electrons tunnelling across the diode and give rise to a negative differential resistance (NDR) that is weakly dependent on temperature. These electron transport properties have potential for the development of nanometre-scale non-linear components for electronics and MIR photonics.


Does exercise impact on sleep for people who have rheumatoid arthritis? A systematic review.
To systematically search for the availability of evidence for exercise impacting on sleep for people who have rheumatoid arthritis. Two reviewers independently searched seven electronic databases, identified and extracted relevant studies by applying eligibility criteria. Sources of bias were assessed independently by two reviewers using the Cochrane bias assessment tool for randomized controlled trials (RCTs) and Newcastle-Ottawa Quality Assessment Scale for non-RCTs. Data were synthesized using a level of evidence approach. Meta-analyses were deemed to be inappropriate due to the heterogeneity of study designs, measurement tools and interventions. Five studies were included: one RCT; two pilot RCTs and two samples of convenience. A total of 262 people with RA were included. Interventions used were difficult to assess due to the heterogeneity of study designs and the inclusion of two using different types of yoga as an intervention. Different sleep outcome measures were used thus, it was not feasible to pool results. Studies had a high risk of bias. This review could find no consistent or conclusive evidence on whether exercise impacts on sleep in people who have rheumatoid arthritis, therefore no firm conclusions can be made. However, there is some indication that exercise may have positive benefits on sleep in people who have rheumatoid arthritis. Further studies with improved study designs, using subjective and objective measures, are needed.


Berberine alleviates symptoms of anxiety by enhancing dopamine expression in rats with post-traumatic stress disorder.
Post-traumatic stress disorder (PTSD) is a trauma-induced psychiatric disorder characterized by impaired fear extermination, hyperarousal, anxiety, depression, and amnesic symptoms that may involve the release of monoamines in the fear circuit. The present study measured several anxiety-related behavioral responses to examine the effects of berberine (BER) on symptoms of anxiety in rats after single prolonged stress (SPS) exposure, and to determine if BER reversed the dopamine (DA) dysfunction. Rats received BER (10, 20, or 30 mg/kg, intraperitoneally, once daily) for 14 days after SPS exposure. BER administration significantly increased the time spent in the open arms and reduced grooming behavior during the elevated plus maze test, and increased the time spent in the central zone and the number of central zone crossings in the open field test. BER restored neurochemical abnormalities and the SPS-induced decrease in DA tissue levels in the hippocampus and striatum. The increased DA concentration during BER treatment may partly be attributed to mRNA expression of tyrosine hydroxylase and the DA transporter in the hippocampus, while BER exerted no significant effects on vesicular monoamine transporter mRNA expression in the hippocampus of rats with PTSD. These results suggest that BER had anxiolytic-like effects on behavioral and biochemical measures associated with anxiety. These findings support a role for reduced anxiety altered DAergic transmission and reduced anxiety in rats with PTSD. Thus, BER may be a useful agent to treat or alleviate psychiatric disorders like those observed in patients with PTSD.


Strategies and challenges associated with recruiting retirement village communities and residents into a group exercise intervention.
Randomized controlled trials (RCTs) provide the highest level of scientific evidence, but successful participant recruitment is critical to ensure the external and internal validity of results. This study describes the strategies associated with recruiting older adults at increased falls risk residing in retirement villages into an 18-month cluster RCT designed to evaluate the effects of a dual-task exercise program on falls and physical and cognitive function. Recruitment of adults aged ≥65 at increased falls risk residing within retirement villages (size 60-350 residents) was initially designed to occur over 12 months using two distinct cohorts (C). Recruitment occurred via a three-stage approach that included liaising with: 1) village operators, 2) independent village managers, and 3) residents. To recruit residents, a variety of different approaches were used, including distribution of information pack, on-site presentations, free muscle and functional testing, and posters displayed in common areas. Due to challenges with recruitment, three cohorts were established between February 2014 and April 2015 (14 months). Sixty retirement villages were initially invited, of which 32 declined or did not respond, leaving 28 villages that expressed interest. A total of 3947 individual letters of invitation were subsequently distributed to residents of these villages, from which 517 (13.1%) expressions of interest (EOI) were received. Across three cohorts with different recruitment strategies adopted there were only modest differences in the number of EOI received (10.5 to 15.3%), which suggests that no particular recruitment approach was most effective. Following the initial screening of these residents, 398 (77.0%) participants were deemed eligible to participate, but a final sample of 300 (58.0% of the 517 EOI) consented and was randomized; 7.6% of the 3947 residents invited. Principal reasons for not participating, despite being eligible, were poor health, lack of time and no GP approval. This study highlights that there are significant challenges associated with recruiting sufficient numbers of older adults from independent living retirement villages into an exercise intervention designed to improve health and well-being. Australian New Zealand Clinical Trials Registry: ACTRN12613001 161718 . Date registered 23rd October 2013.


Healthy aging: is smaller better? - a mini-review.
A recent report of virtually complete protection from diabetes and cancer in a population of people with hereditary dwarfism revived interest in elucidating the relationships between growth, adult body size, age-related disease and longevity. In many species, smaller individuals outlive those that are larger and a similar relationship was shown in studies of various human populations. Adult body size is strongly dependent on the actions of growth hormone (GH) and the absence of GH or GH receptor in mice leads to a remarkable extension of longevity. Many mechanisms that may account for, or contribute to, this association have been identified. It is suggested that modest modifications of the diet at different ages may extend human healthspan and lifespan by reducing levels of hormones that stimulate growth.


Calibration of diffuse correlation spectroscopy with a time-resolved near-infrared technique to yield absolute cerebral blood flow measurements.
A primary focus of neurointensive care is the prevention of secondary brain injury, mainly caused by ischemia. A noninvasive bedside technique for continuous monitoring of cerebral blood flow (CBF) could improve patient management by detecting ischemia before brain injury occurs. A promising technique for this purpose is diffuse correlation spectroscopy (DCS) since it can continuously monitor relative perfusion changes in deep tissue. In this study, DCS was combined with a time-resolved near-infrared technique (TR-NIR) that can directly measure CBF using indocyanine green as a flow tracer. With this combination, the TR-NIR technique can be used to convert DCS data into absolute CBF measurements. The agreement between the two techniques was assessed by concurrent measurements of CBF changes in piglets. A strong correlation between CBF changes measured by TR-NIR and changes in the scaled diffusion coefficient measured by DCS was observed (R(2) = 0.93) with a slope of 1.05 ± 0.06 and an intercept of 6.4 ± 4.3% (mean ± standard error).


The outcome of extreme prematurity.
Significant advances in perinatology and neonatology in the last decade have resulted in increased survival of extremely premature infants. Survival rates for infants born in tertiary perinatal and neonatal care centers in the United States in the 1990s increase with each week of gestational age from 22 through 26 weeks. Reported survival rates at 22 weeks range from 0% to 21% in the few reporting studies. Reported survival rates at 23 and 24 weeks range from 5% to 46% and from 40% to 59%, respectively. These may not be the maximum survival rates possible because at these gestational ages information is either insufficient to determine that obstetric and neonatal intensive care strategies to maximize neonatal survival were used or it is specified that such strategies were not used. Reported survival rates at 25 and 26 weeks range from 60% to 82% and from from 75% to 93%, respectively. The literature regarding the prevalence of major neurodevelopmental disabilities among extremely premature survivors in the last 25 years is heteogeneous, and the reported prevalances of major disability vary much more than do survival rates. However, the majority of extremely premature infants who survive will be free of major disability. Overall, approximately one fifth to one quarter of survivors have at least one major disability-impaired mental development, cerebral palsy, blindness, or deafness. Impaired mental development is the most prevalent disability (17%-21% [95% CI] of survivors affected), followed by cerebral palsy (12%-15% of survivors affected). Blindness and deafness are less common (5% to 8% and 3% to 5% of survivors affected, respectively). Approximately one half of disabled survivors have more than one major disability. Based on studies of infants less than 750 to 1,000 grams birth weight, it can be anticipated that approximately another half of all extremely premature survivors will have one or more subtle neurodevelopmental disabilities in the school and teenage years. There is little evidence to suggest that long-term neurodevelopmental outcome has changed from the late 1970s to the early 1990s or with increasing survival. Survival of individual extremely premature infants cannot be accurately predicted in the immediate perinatal period. Major disability cannot be accurately predicted for individual survivors during the course in the newborn intensive care unit.


An online mindfulness intervention targeting socioemotional regulation in fibromyalgia: results of a randomized controlled trial.
Patients with fibromyalgia (FM) experience pain as well as deficits in positive affect and social relations that are not explicitly addressed in most behavioral treatments. The purpose of this study is to compare the effects of a 12-module online intervention targeting socioemotional regulation via mindful awareness/acceptance (MSER) with those of an attention-control treatment, healthy lifestyle tips (HT). Seventy-nine FM patients were randomly assigned to MSER or HT, with outcomes assessed via online diary reports of pain, coping efficacy, affect, and social relations. Multilevel analyses revealed greater improvements in social functioning, positive affect, and coping efficacy for pain and stress (all ps < .05) in MSER versus HT across the 6-week trial. FM patients experience increases in self-efficacy for coping with pain and positive engagement in relationships, marginal increases in positive affect, and decreases in relationship stress from an automated online intervention that targets socioemotional regulation skills. Findings highlight the potential utility of widely accessible, low-cost intervention methods for fibromyalgia (Clinicaltrials.gov number NCT01748786).


Sympathetic activation of leptin via the ventromedial hypothalamus: leptin-induced increase in catecholamine secretion.
Leptin is an adipocyte-derived blood-borne satiety factor that acts directly on the hypothalamus, thereby regulating food intake and energy expenditure. We have demonstrated that the hypothalamic arcuate nucleus (Arc) is a primary site of the satiety effect of leptin (Neurosci Lett 224:149-152, 1997). To explore the hypothalamic pathway of sympathetic activation of leptin, we examined the effects of a single intravenous or intracerebroventricular injection of recombinant human leptin on catecholamine secretion in rats. We also examined the effects of direct microinjection of leptin into the ventromedial hypothalamus (VMH), Arc, paraventricular nucleus (PVN), and dorsomedial hypothalamus (DMH) in rats. To further assess whether sympathetic activation of leptin is mediated via the VMH, we also examined the effects of a single intravenous injection of leptin in VMH-lesioned rats. A single injection of leptin (0.25-1.0 mg i.v./rat or 0.5-2.0 pg i.c.v./rat) increased plasma norepinephrine (NE) and epinephrine (EPI) concentrations in a dose-dependent manner. Plasma NE and EPI concentrations were increased significantly when leptin was injected directly into the VMH but were unchanged when injected into the Arc, PVN, and DMH. Plasma NE and EPI concentrations were unchanged in VMH-lesioned rats that received a single intravenous injection of leptin. The present study provides evidence that a leptin-induced increase in catecholamine secretion is mediated primarily via the VMH and suggests the presence of distinct hypothalamic pathways mediating the satiety effect and sympathetic activation of leptin.


The social welfare function and individual responsibility: some theoretical issues and empirical evidence.
The literature on income distribution has attempted to evaluate different degrees of inequality using a social welfare function (SWF) approach. However, it has largely ignored the source of such inequalities, and has thus failed to consider different degrees of inequity. The literature on egalitarianism has addressed issues of equity, largely in relation to individual responsibility. This paper builds upon these two literatures, and introduces individual responsibility into the SWF. Results from a small-scale study of people's preferences in relation to the distribution of health benefits are presented to illustrate how the parameter values of a SWF might be determined.


Anti-cancer drugs elicit re-expression of UDP-glucuronosyltransferases in melanoma cells.
The UDP-glucuronosyltransferase (UGT) family of enzymes plays a vital role in the detoxification of carcinogens as well as clearance of anti-cancer drugs. In humans, 19 UGT family members have been identified and are expressed in a tissue specific manner throughout the body. However, the UGTs have not been previously characterized in melanocytes or melanoma. In the present study, UGT2B7, UGT2B10, and UGT2B15 were identified as being normally expressed in human melanocytes. The same three UGT family members were also expressed in the primary melanoma cell line WM115. No UGT expression was detected in another primary melanoma cell line, WM3211, or in any metastatic melanoma cell line examined. These results suggest that UGT expression is lost during melanoma progression. Treatment of WM3211 or metastatic melanoma cell lines with anti-cancer agents (including vemurafenib) induced expression of UGT2B7, UGT2B10 and UGT2B15 demonstrating that melanoma cells retain the ability to re-express these same three UGTs. The corresponding increase in glucuronidation activity in melanoma cells following anti-cancer treatment was also observed. Furthermore, knockdown of UGT2B7 in WM115 cells sensitized these cells to treatment by adriamycin and epirubicin indicating that UGT2B7 is involved in resistance to these drugs. However, knockdown of UGT2B7 had no effect on temozolomide toxicity. Taken together, these results clearly demonstrate a role for UGTs in melanoma etiology. Since the UGTs are drug metabolism enzymes, we propose that re-expression of the UGTs constitutes a previously unsuspected mechanism for intratumoral drug resistance in melanoma.


Individual variation in the spatial reference and working memory assessed under allothetic and idiothetic orientation cues in rat.
The present study was designed to examine which kind of memory: reference or working, better correlates with individual variation in rats' spatial learning abilities. To answer this question two groups of rats were trained to an arbitrary criterion in a partially baited 12-arm radial maze under two different experimental conditions: with or without allothetic cues. After 10 days break, rats were examined under the same conditions for memory retention. Within- and between-group variation in the length of training to criterion, and in the frequency of reference and working memory errors were analysed. The present experiment confirmed the facilitating effect of the presence of distal visual cues on place learning in rats. Task-dependent (between-group) differences in the rate of learning were attributed to differences in the frequency of reference memory errors. Conversely, within-group variation in the rate of task acquisition reflected individual variation in the frequency of working memory errors. These results were looked upon from an evolutionary perspective. Low correlation between reference and working memory errors confirms that these two types of memory have different mechanisms. The fact that differences in the rate of learning were not paralleled by the differences in the memory retention supports the notion that memory acquisition and memory retention are two independent processes.


Inhibition of human immunodeficiency virus type 1 multiplication by transforming growth factor beta 1 and AZT in HIV-1-infected myeloid cells.
Myeloid cells are important reservoirs of HIV-1 infection. In response to pathogenic agents, macrophages secrete inflammatory cytokines that can modulate viral replication and contribute to AIDS pathogenesis. Because HIV replication is dependent on cellular activation, immunosuppressive cytokines that deactivate macrophages and T cells may be important modulators of an antiviral effect. We tested the anti-HIV potential of the immunosuppressive cytokine-transforming growth factor beta (TGF-beta 1) alone and in combination with AZT in a new monomyeloblastic model of HIV-1 infection. The PLB-985 cell model was infected with HIV IIIB strain, and the course of HIV-1 infection and replication was monitored by reverse transcriptase assay, p24 immunofluorescence, and northern blot analysis of HIV-1-specific mRNA. TGF-beta 1 as a single agent had no effect on the multiplication of HIV-IIIB in de novo-infected PLB 985 monomyeloblastic cells. However, cotreatment with TGF-beta 1 and AZT synergistically slowed virus multiplication within the first week following infection, as determined by reverse transcriptase measurement, p24 antigen detection, and northern blot analysis of viral RNA. The synergistic actions of TGF-beta 1 and AZT were also observed in PLB 985 cells infected with an AZT-resistant strain of HIV-1 (HIV 1393). Synergism between nucleoside analogs and cytokines may be an important therapeutic approach to HIV-1 infection. Elucidation of the role of cytokines in controlling the degree of HIV multiplication may have an impact on both clinical treatments and understanding the progression to AIDS.


Allostery in Its Many Disguises: From Theory to Applications.
Allosteric regulation plays an important role in many biological processes, such as signal transduction, transcriptional regulation, and metabolism. Allostery is rooted in the fundamental physical properties of macromolecular systems, but its underlying mechanisms are still poorly understood. A collection of contributions to a recent interdisciplinary CECAM (Center Européen de Calcul Atomique et Moléculaire) workshop is used here to provide an overview of the progress and remaining limitations in the understanding of the mechanistic foundations of allostery gained from computational and experimental analyses of real protein systems and model systems. The main conceptual frameworks instrumental in driving the field are discussed. We illustrate the role of these frameworks in illuminating molecular mechanisms and explaining cellular processes, and describe some of their promising practical applications in engineering molecular sensors and informing drug design efforts.


Long-term toxicity of surface-charged polystyrene nanoplastics to marine planktonic species Dunaliella tertiolecta and Artemia franciscana.
Plastic pollution has been globally recognized as a critical issue for marine ecosystems and nanoplastics constitute one of the last unexplored areas to understand the magnitude of this threat. However, current difficulties in sampling and identifying nano-sized debris make hard to assess their occurrence in marine environment. Polystyrene nanoparticles (PS NPs) are largely used as nanoplastics in ecotoxicological studies and although acute exposures have been already investigated, long-term toxicity on marine organisms is unknown. Our study aims at evaluating the effects of 40nm PS anionic carboxylated (PS-COOH) and 50nm cationic amino-modified (PS-NH2) NPs in two planktonic species, the green microalga Dunaliella tertiolecta and the brine shrimp Artemia franciscana, respectively prey and predator. PS NP behaviour in exposure media was determined through DLS, while their toxicity to microalgae and brine shrimps evaluated through 72h growth inhibition test and 14 d long-term toxicity test respectively. Moreover, the expression of target genes (i.e. clap and cstb), having a role in brine shrimp larval growth and molting, was measured in 48h brine shrimp larvae. A different behaviour of the two PS NPs in exposure media as well as diverse toxicity to the two planktonic species was observed. PS-COOH formed micro-scale aggregates (Z-Average>1μm) and did not affect the growth of microalgae up to 50μg/ml or that of brine shrimps up to 10μg/ml. However, these negatively charged NPs were adsorbed on microalgae and accumulated (and excreted) in brine shrimps, suggesting a potential trophic transfer from prey to predator. On the opposite, PS-NH2-formed nano-scale aggregates (Z-Average<200nm), caused inhibition of algal growth (EC50=12.97μg/ml) and mortality in brine shrimps at 14 d (LC50=0.83μg/ml). Moreover, 1μg/ml PS-NH2 significantly induced clap and cstb genes, explaining the physiological alterations (e.g. increase in molting) previously observed in 48h larvae, but also suggesting an apoptotic pathway triggered by cathepsin L-like protease in brine shrimps upon PS-NH2 exposure. These findings provide a first insight into long-term toxicity of nanoplastics to marine plankton, underlining the role of the surface chemistry in determining the behaviour and effects of PS NPs, in terms of adsorption, growth inhibition, accumulation, gene modulation and mortality. The use of long-term end-point has been identified as valuable tool for assessing the impact of nanoplastics on marine planktonic species, being more predictable of real exposure scenarios for risk assessment purposes.


Mutations in NCAPG2 Cause a Severe Neurodevelopmental Syndrome that Expands the Phenotypic Spectrum of Condensinopathies.
The use of whole-exome and whole-genome sequencing has been a catalyst for a genotype-first approach to diagnostics. Under this paradigm, we have implemented systematic sequencing of neonates and young children with a suspected genetic disorder. Here, we report on two families with recessive mutations in NCAPG2 and overlapping clinical phenotypes that include severe neurodevelopmental defects, failure to thrive, ocular abnormalities, and defects in urogenital and limb morphogenesis. NCAPG2 encodes a member of the condensin II complex, necessary for the condensation of chromosomes prior to cell division. Consistent with a causal role for NCAPG2, we found abnormal chromosome condensation, augmented anaphase chromatin-bridge formation, and micronuclei in daughter cells of proband skin fibroblasts. To test the functional relevance of the discovered variants, we generated an ncapg2 zebrafish model. Morphants displayed clinically relevant phenotypes, such as renal anomalies, microcephaly, and concomitant increases in apoptosis and altered mitotic progression. These could be rescued by wild-type but not mutant human NCAPG2 mRNA and were recapitulated in CRISPR-Cas9 F0 mutants. Finally, we noted that the individual with a complex urogenital defect also harbored a heterozygous NPHP1 deletion, a common contributor to nephronophthisis. To test whether sensitization at the NPHP1 locus might contribute to a more severe renal phenotype, we co-suppressed nphp1 and ncapg2, which resulted in significantly more dysplastic renal tubules in zebrafish larvae. Together, our data suggest that impaired function of NCAPG2 results in a severe condensinopathy, and they highlight the potential utility of examining candidate pathogenic lesions beyond the primary disease locus.


Wanted: fully operational definitions of dissociations in single-case studies.
In contrast to the careful consideration given to the issue of what we can infer from dissociations in single-case studies, the more basic question of how we decide whether a dissociation is present has been relatively neglected. Proposals are made for fully operational definitions of a deficit, classical and strong dissociations, and double dissociations. In developing these definitions it was assumed that they should be based on the use of inferential rather than descriptive statistical methods. The scope of these definitions is limited to typical single-case studies in which patients are compared to control samples of a modest size. The operational definition of a classical dissociation incorporates a requirement that a patient's performance on Task X is significantly different from Task Y, in addition to the "standard" requirement that the patient has a deficit on Task X and is within normal limits on Task Y. We ran a simulation to estimate the Type I error rates when the criteria for dissociations are applied and found these to be low (Type I errors were defined as identifying an individual from the control population as having a dissociation). The inferential methods for testing whether the various criteria are met make use of t-distributions. These methods are contrasted with the widespread use of z to test for a deficit or a difference between tasks. In the latter approach the statistics of the control sample are treated as parameters; this is not appropriate when, as is normally the case, the control sample size is modest in size.


Reversal of PSA progression on abiraterone acetate through the administration with food in men with metastatic castration-resistant prostate cancer.
Owing to efficacy and tolerability, abiraterone acetate (AA) is a leading treatment for men with metastatic castration-resistant prostate cancer. Increased serum concentrations of AA, such as by taking AA with food, may lead to the inhibition of additional enzymes in the androgen synthesis pathway implicated in castration-resistant prostate cancer progression. Medical records of men with metastatic castration-resistant prostate cancer (mCRPC) who received AA between 1 April 2011 and 31 December 2013 were retrospectively reviewed. The primary outcome was the percent of men with a rising PSA on AA who experienced any PSA decline within 3 months after changing the administration of AA from without food to with food. Secondary outcomes were median time on AA therapy in men who received AA therapy without food versus those that switched administration from without food to with food at PSA progression, and the percent of men who experienced any decline in serum testosterone concentration, and the rate of adverse events observed while taking AA with food. Nineteen men who switched AA administration from without food to with food and 41 patients who administered AA without food only were included in the study. Of those patients who took AA with food at PSA progression, a PSA decline was observed in 3 of the 19 (16%) men, including 3 of the 14 men who had an initial response to AA (21%). Testosterone declined in five out of seven patients from pre-food levels. The median time on AA therapy was increased by nearly 100 days in patients who switched AA administration from without food to with food. No increases in toxicity were observed. Some men with mCRPC may benefit from taking AA with food. Further prospective comparative studies are needed to determine if changing AA administration is beneficial.


World Heart Federation Cholesterol Roadmap.
The World Heart Federation has undertaken an initiative to develop a series of Roadmaps. The aim of these is to promote development of national policies and health systems approaches and identify potential roadblocks on the road to effective prevention, detection and management of cardiovascular disease (CVD) in low-and middle-income countries (LMIC), and strategies for overcoming these. This Roadmap focuses on elevated blood cholesterol, a leading risk factor for myocardial infarction, stroke, and peripheral arterial disease. Through a review of published guidelines and research papers, and consultation with a committee composed of experts in clinical management of cholesterol and health systems research in LMIC, this Roadmap identifies (1) key interventions for primordial, primary and secondary prevention of CVD through detection, treatment, and management of elevated cholesterol and familial hypercholesterolemia (FH); (2) gaps in implementation of these interventions (knowledge-practice gaps); (3) health system roadblocks to treatment of elevated cholesterol in LMIC; and (4) potential strategies for overcoming these. Despite strong evidence of the importance of cholesterol levels in primary or secondary prevention of CVD, and the effectiveness of statin therapy for cholesterol lowering and reduction of CVD risk, gaps exist in the detection, treatment, and management of high cholesterol globally. Some potential roadblocks include poor access to laboratory facilities or trained professionals for cholesterol management, low awareness of FH among the general population and health professionals, unaffordability of statins for patient households, and low awareness of the importance of persistent adherence to lipid-lowering medication. Potential solutions include point-of-care testing, provision of free or subsidized lipid-lowering medication, and treatment adherence support using text message reminders. Known effective strategies for detection, treatment, and management of elevated cholesterol and FH exist, but there are barriers to their implementation in many low-resource settings. Priorities for health system intervention should be identified at the national level, and the feasibility and effectiveness of proposed solutions should be assessed in specific contexts. Many solutions proposed in this Roadmap may apply to other cardiovascular conditions and present opportunities for integration of CVD care in LMIC.


The Therapeutic Environment Screening Survey for Nursing Homes (TESS-NH): an observational instrument for assessing the physical environment of institutional settings for persons with dementia.
To develop an observational instrument that describes the ability of physical environments of institutional settings to address therapeutic goals for persons with dementia. A National Institute on Aging workgroup identified and subsequently revised items that evaluated exit control, maintenance, cleanliness, safety, orientation/cueing, privacy, unit autonomy, outdoor access, lighting, noise, visual/tactile stimulation, space/seating, and familiarity/homelikeness. The final instrument contains 84 discrete items and one global rating. A summary scale, the Special Care Unit Environmental Quality Scale (SCUEQS), consists of 18 items. Lighting items were validated using portable light meters. Concurrent criterion validation compared SCUEQS scores with the Professional Environmental Assessment Protocol (PEAP). Interrater kappa statistics for 74% of items were above.60. For another 10% of items, kappas could not be calculated due to empty cells, but interrater agreement was above 80%. The SCUEQS demonstrated an interrater reliability of.93, a test--retest reliability of.88, and an internal consistency of.81--.83. Light meter ratings correlated significantly with the Therapeutic Environment Screening Survey for Nursing Homes (TESS-NH) lighting items (r =.29--.38, p =.01--.04), and the SCUEQS correlated significantly with global PEAP ratings (r =.52, p <.01). The TESS-NH efficiently assesses discrete elements of the physical environment and has strong reliability and validity. The SCUEQS provides a quantitative measure of environmental quality in institutional settings.


Periodic whole body acceleration: a novel therapy for cardiovascular disease.
Periodic whole body acceleration in the spinal axis (pGz) applied by a motion platform is a novel treatment modality that induced endothelial nitric oxide release into the circulation of animals, healthy subjects and patients with inflammatory diseases during single treatment sessions in previous studies. We hypothesized that patients with advanced arteriosclerotic diseases who are not candidates for a surgical intervention would clinically benefit from repeated pGz treatments over several weeks through improvement of endothelial function. 11 patients, 5 men (37 to 71 y) with stable ischemic heart disease, LVEF < 35%, NYHA stage > II, and 6 patients (51 to 83 y, 1 woman) with intermittent leg claudication, Fontaine stage II, were enrolled after optimization of pharmacological therapy. PGz was applied for 40 min, 5 days/week during 5 weeks. Quality of life (SF-36 questionnaire), exercise performance, and endothelial function were assessed at baseline, during the treatment period, and 4 weeks after discontinuation of pGz. PGz was well tolerated. In heart failure paitents, pGz therapy improved quality of life, increased 6 min walking distance by a mean +/- SE of 105 +/- 24 m, and improved postischemic skin hyperemia (p < .05 in all instances). In 4 of 6 patients with intermittent claudication, quality of life, treadmill walking distance and post-ischemic skin hyperemia improved with pGz therapy (p < .05). Four weeks after discontinuation of pGz, most therapeutic effects had vanished in both patient groups. In patients with severe heart failure and with leg claudication who remain symptomatic despite maximal medical therapy and who were not candidates for surgery, periodic acceleration applied over several weeks improved quality of life and exercise capacity. The clinical benefits appear to be mediated through improved endothelial function.


AI-Enabled, Ultrasound-Guided Handheld Robotic Device for Femoral Vascular Access.
Hemorrhage is a leading cause of trauma death, particularly in prehospital environments when evacuation is delayed. Obtaining central vascular access to a deep artery or vein is important for administration of emergency drugs and analgesics, and rapid replacement of blood volume, as well as invasive sensing and emerging life-saving interventions. However, central access is normally performed by highly experienced critical care physicians in a hospital setting. We developed a handheld AI-enabled interventional device, AI-GUIDE (Artificial Intelligence Guided Ultrasound Interventional Device), capable of directing users with no ultrasound or interventional expertise to catheterize a deep blood vessel, with an initial focus on the femoral vein. AI-GUIDE integrates with widely available commercial portable ultrasound systems and guides a user in ultrasound probe localization, venous puncture-point localization, and needle insertion. The system performs vascular puncture robotically and incorporates a preloaded guidewire to facilitate the Seldinger technique of catheter insertion. Results from tissue-mimicking phantom and porcine studies under normotensive and hypotensive conditions provide evidence of the technique's robustness, with key performance metrics in a live porcine model including: a mean time to acquire femoral vein insertion point of 53 ± 36 s (5 users with varying experience, in 20 trials), a total time to insert catheter of 80 ± 30 s (1 user, in 6 trials), and a mean number of 1.1 (normotensive, 39 trials) and 1.3 (hypotensive, 55 trials) needle insertion attempts (1 user). These performance metrics in a porcine model are consistent with those for experienced medical providers performing central vascular access on humans in a hospital.


Use of colour-coded labels for intravenous high-risk medications and lines to improve patient safety.
Labelling of high-risk drug infusions and lines is a well-recognised safety strategy to prevent medication errors. Although hospital wards characterised by multiple high-risk drug infusions use different types of labelling, little is known about the contribution of a colour-coded label (CCL) to patient safety. A quality improvement programme audit at a tertiary care facility, the Hadassah University Medical Center Ein Kerem, Jerusalem, Israel. A CCL for intravenous (IV) high-risk medications and lines was designed to promote safer medication administration at the intensive care unit bedside and in other acute wards. The purpose of the study was to compare a new CCL method (intervention) with the current labelling method (control). Laboratory simulation, imitating an intensive care unit, was designed. Safety of the medication treatment and overall duration of nurses' orientation with drugs and lines at the patient's bedside were measured. The use of the new CCL improved proper identification of IV bags (p<0.0001), reduced the time required for description of overall drugs and lines (p = 0.04), improved identification of errors at the treatment setting-drugs and lines (p = 0.03) and reduced the average performance time for overall tasks (p<0.0001). The use of CCLs for IV high-risk medications and lines can improve patient safety and improve medical staff efficiency.


Dysregulation in retinal para-inflammation and age-related retinal degeneration in CCL2 or CCR2 deficient mice.
We have shown previously that a para-inflammatory response exists at the retinal/choroidal interface in the aging eye; and this response plays an important role in maintaining retinal homeostasis under chronic stress conditions. We hypothesized that dysregulation of the para-inflammatory response may result in an overt pro-inflammatory response inducing retinal degeneration. In this study, we examined this hypothesis in mice deficient in chemokine CCL2 or its cognate receptor CCR2. CCL2- or CCR2-deficient mice developed retinal degenerative changes with age, characterized as retinal pigment epithelial (RPE) cell and photoreceptor cell death. Retinal cell death was associated with significantly more subretinal microglial accumulation and increased complement activation. In addition, monocytes from CCL2- or CCR2-deficient mice had reduced capacity for phagocytosis and chemotaxis, expressed less IL-10 but more iNOS, IL-12 and TNF-α when compared to monocytes from WT mice. Complement activation at the site of RPE cell death resulted in C3b/C3d but not C5b-9 deposition, indicating only partial activation of the complement pathway. Our results suggest that altered monocyte functions may convert the protective para-inflammatory response into an overtly harmful inflammation at the retina/choroidal interface in CCL2- or CCR2-deficient mice, leading to RPE and photoreceptor degeneration. These data support a concept whereby a protective para-inflammatory response relies upon a normally functioning innate immune system. If the innate immune system is deficient chronic stress may tip the balance towards an overt inflammatory response causing cell/tissue damage.


Eating Disorders In weight-related Therapy (EDIT) Collaboration: rationale and study design.
The cornerstone of obesity treatment is behavioural weight management, resulting in significant improvements in cardio-metabolic and psychosocial health. However, there is ongoing concern that dietary interventions used for weight management may precipitate the development of eating disorders. Systematic reviews demonstrate that, while for most participants medically supervised obesity treatment improves risk scores related to eating disorders, a subset of people who undergo obesity treatment may have poor outcomes for eating disorders. This review summarises the background and rationale for the formation of the Eating Disorders In weight-related Therapy (EDIT) Collaboration. The EDIT Collaboration will explore the complex risk factor interactions that precede changes to eating disorder risk following weight management. In this review, we also outline the programme of work and design of studies for the EDIT Collaboration, including expected knowledge gains. The EDIT studies explore risk factors and the interactions between them using individual-level data from international weight management trials. Combining all available data on eating disorder risk from weight management trials will allow sufficient sample size to interrogate our hypothesis: that individuals undertaking weight management interventions will vary in their eating disorder risk profile, on the basis of personal characteristics and intervention strategies available to them. The collaboration includes the integration of health consumers in project development and translation. An important knowledge gain from this project is a comprehensive understanding of the impact of weight management interventions on eating disorder risk.


Neuropathic Arthropathy: Review of Current Knowledge.
Neuropathic arthropathy is a chronic, progressive degenerative disorder affecting one or more peripheral or vertebral articulations, which develops as the result of a disturbance in the normal sensory (pain or proprioceptive) innervation of joints. Diabetes, syphilis, and syringomelia are the most commonly associated clinical entities. When neuropathic arthropathy is suspected, careful clinical evaluation should be performed to identify an underlying neurologic disorder. Patient education, joint protection, and early recognition of fractures are the most important general management principles. Surgery can be considered in cases of advanced joint destruction when there is significant disability.


Enhanced Performance of Plasmid DNA Polyplexes Stabilized by a Combination of Core Hydrophobicity and Surface PEGylation.
Nonviral gene therapy has high potential for safely promoting tissue restoration and for treating various genetic diseases. One current limitation is that conventional transfection reagents such as polyethylenimine (PEI) form electrostatically stabilized plasmid DNA (pDNA) polyplexes with poor colloidal stability. In this study, a library of poly(ethylene glycol-b-(dimethylaminoethyl methacrylate-co-butyl methacrylate)) [poly(EG-b-(DMAEMA-co-BMA))] polymers were synthesized and screened for improved colloidal stability and nucleic acid transfection following lyophilization. When added to pDNA in the appropriate pH buffer, the DMAEMA moieties initiate formation of electrostatic polyplexes that are internally stabilized by hydrophobic interactions of the core BMA blocks and sterically stabilized against aggregation by a PEG corona. The BMA content was varied from 0% to 60% in the second polymer block in order to optimally tune the balance of electrostatic and hydrophobic interactions in the polyplex core, and polymers with 40 and 50 mol% BMA achieved the highest transfection efficiency. Diblock copolymers were more stable than PEI in physiologic buffers. Consequently, diblock copolymer polyplexes aggregated more slowly and followed a reaction-limited colloidal aggregation model, while fast aggregation of PEI polyplexes was governed by a diffusion-limited model. Polymers with 40% BMA did not aggregate significantly after lyophilization and produced up to 20-fold higher transfection efficiency than PEI polyplexes both before and after lyophilization. Furthermore, poly(EG-b-(DMAEMA-co-BMA)) polyplexes exhibited pH-dependent membrane disruption in a red blood cell hemolysis assay and endosomal escape as observed by confocal microscopy.Lyophilized polyplexes made with the lead candidate diblock copolymer (40% BMA) also successfully transfected cells in vitro following incorporation into gas-foamed polymeric scaffolds. In summary, the enhanced colloidal stability, endosomal escape, and resultant high transfection efficiency of poly(EG-b-(DMAEMA-co-BMA))-pDNA polyplexes underscores their potential utility both for local delivery from scaffolds as well as systemic, intravenous delivery.


Restraint of angiogenesis by zinc finger transcription factor CTCF-dependent chromatin insulation.
Angiogenesis is meticulously controlled by a fine balance between positive and negative regulatory activities. Vascular endothelial growth factor (VEGF) is a predominant angiogenic factor and its dosage is precisely regulated during normal vascular formation. In cancer, VEGF is commonly overproduced, resulting in abnormal neovascularization. VEGF is induced in response to various stimuli including hypoxia; however, very little is known about the mechanisms that confine its induction to ensure proper angiogenesis. Chromatin insulation is a key transcription mechanism that prevents promiscuous gene activation by interfering with the action of enhancers. Here we show that the chromatin insulator-binding factor CTCF binds to the proximal promoter of VEGF. Consistent with the enhancer-blocking mode of chromatin insulators, CTCF has little effect on basal expression of VEGF but specifically affects its activation by enhancers. CTCF knockdown cells are sensitized for induction of VEGF and exhibit elevated proangiogenic potential. Cancer-derived CTCF missense mutants are mostly defective in blocking enhancers at the VEGF locus. Moreover, during mouse retinal development, depletion of CTCF causes excess angiogenesis. Therefore, CTCF-mediated chromatin insulation acts as a crucial safeguard against hyperactivation of angiogenesis.


Changing Tides: Mothers' Supportive Emotion Socialization Relates Negatively to Third-Grade Children's Social Adjustment in School.
Parents' supportive reactions to children's negative emotions are thought to promote children's social adjustment. Research heretofore has implicitly assumed that such reactions are equally supportive of children's adjustment across ages. Recent findings challenge this assumption, suggesting that during middle childhood, socialization practices previously understood as supportive may in fact impede children's social adjustment. We explored this possibility in a sample of 203 third-grade children and their mothers. Using structural equation modeling, we tested associations between mothers' supportive (i.e., problem- and emotion-focused) reactions to children's negative emotions and children's social skills and problems as reported by mothers and teachers. Mothers' supportive reactions predicted greater social adjustment in children as reported by mothers. Inverse associations, however, were found with teachers' reports of children's social adjustment: mothers' supportive reactions predicted fewer socioemotional skills and more problem behaviors. These contrasting patterns suggest potential unperceived costs associated with mothers' supportiveness of children's negative emotions for third-grade children's social adjustment in school and highlight the importance of considering associations between socialization practices and children's various social contexts. The findings also highlight a need for greater consideration of what supportiveness means across different developmental periods.


Nitric Oxide and Related Aspects Underlying Angina.
Increased number of patients affected by metabolic syndrome (MS) has prompted the necessity of better understanding what is involved in such syndrome. Nevertheless, the establishment of promising therapies depends on the knowledge about the interaction of molecules within MS. In such context, Nitric Oxide (NO) emerges from a bulk of works relating its roles on aspects of MS, including cardiovascular diseases, their symptoms and comorbidities, which are thought to be triggered by similar sources. NO, nitric oxide synthase and enzymatic chains are keys for those disease and symptoms processes. NO has been separately described as part of hypertensive, ischemic and pain signaling. Although there are similar pathways likely shared for generating cardiovascular symptoms such angina, they are barely associated to NO in literature. The present review aims to clarify the patterns of NO alteration in metabolic syndrome directly concerned to cardiovascular symptoms, especially angina.


Anemone - an Additional Perennial Ornamental Host of Tobacco Rattle Virus in the U.S.
A previous report (1) drew attention to the occurrence in the U.S. of tobacco rattle tobravirus (TRV) in several perennial ornamentals that move freely in international trade. Here we report the occurrence of TRV in an additional host plant of this type. The virus was identified in anemone (Anemone × hybrida cv. Honorine Jobert) with leaf symptoms consisting of chlorotic blotches, chlorotic line patterns, and distortion. Characteristic tobravirus-like particles with modal lengths of 50, 80, 130, and 180 nm were observed by electron microscopy in partially purified extracts of symptomatic but not of asymptomatic plants. These particles reacted specifically with antibodies to TRV and pea early-browning (PEBV) tobraviruses in immunoelectron microscopic (IEM) assays, produced typical TRV-induced symptoms in Nicotiana benthamiana and N. clevelandii, in which similar particles were detected by IEM, but, like other TRV isolates, did not infect pea (2). No other viruslike particles were observed in partially purified extracts of symptomatic anemone plants. This is the first report of TRV infection in anemone. While TRV occurs widely in Europe, it had been identified previously in the U.S. only in Oregon, Washington, and California (1,2,3). This virus is an important pathogen of several crops, including potato. Several areas of the Midwest, including Minnesota and Wisconsin, have significant seed-potato industries, and further introduction and possible dissemination of TRV pose potential regulatory and quarantine issues. This example of the movement of an exotic pathogen of potential economic importance into new areas underlines the need for closer monitoring of plant material entering international trade in an era of increasing globalization. References: (1) B. E. Lockhart et al. Plant Dis. 79:1249, 1995. (2) B. E. L. Lockhart and H. U. Fischer. Phytopathology 66:1391, 1976. (3) J. M. Crosslin and P. E. Thomas. Am. Potato J. 72:605, 1995.


Mutational neighbourhood and mutation supply rate constrain adaptation in Pseudomonas aeruginosa.
Understanding adaptation by natural selection requires understanding the genetic factors that determine which beneficial mutations are available for selection. Here, using experimental evolution of rifampicin-resistant Pseudomonas aeruginosa, we show that different genotypes vary in their capacity for adaptation to the cost of antibiotic resistance. We then use sequence data to show that the beneficial mutations associated with fitness recovery were specific to particular genetic backgrounds, suggesting that genotypes had access to different sets of beneficial mutations. When we manipulated the supply rate of beneficial mutations, by altering effective population size during evolution, we found that it constrained adaptation in some selection lines by restricting access to rare beneficial mutations, but that the effect varied among the genotypes in our experiment. These results suggest that mutational neighbourhood varies even among genotypes that differ by a single amino acid change, and this determines their capacity for adaptation as well as the influence of population biology processes that alter mutation supply rate.


Job strain and smoking cessation among Japanese male employees: a two-year follow-up study.
The purpose of the present study was to assess the association between job strain and smoking cessation among Japanese male employees. In 1997, a baseline questionnaire was given to 2,625 (2,113 males and 512 females) employees of an electronics firm in Gifu Prefecture, Japan. The self-administered questionnaire was a set of questions on smoking habits and consisted of items on socio-demographic variables and smoking habits, including the Japanese version of the Job Content Questionnaire (JCQ). The JCQ consists of scales of job control, job demand, supervisory support, coworker support, job insecurity, physical demands, and isometric load. A total of 733 male smokers were then followed for 2 years, with 446 completing a follow-up questionnaire in 1999 (follow-up rate, 61%). Logistic regression analyses were performed to examine associations between job strain and smoking cessation. Among the 446 participants, 38 had quit smoking. After adjusting for age ((odds ratio: OR) = 0.38, 95% (contidence interval: CI) = 0.15-0.94), men with a high level of physical demands at baseline showed a lower smoking cessation rate at follow-up than did those with a low level. However, when adjustments were made for age and other socio-demographic variables, the odds ratio of smoking cessation showed marginal significance (OR = 0.45, 95% CI = 0.16-1.29). There was no significant association between other job strain variables and smoking cessation at the 2-year follow-up. No significant association was found between job strain and change in the number of smoked cigarettes per day. The present study did not support the hypothesis that higher levels of job stressors are associated with a lower rate of smoking cessation among men.


Decreased absolute amygdala volume in cocaine addicts.
The amygdala is instrumental to a set of brain processes that lead to cocaine consumption, including those that mediate reward and drug craving. This study examined the volumes of the amygdala and hippocampus in cocaine-addicted subjects and matched healthy controls and determined that the amygdala but not the hippocampus was significantly reduced in volume. The right-left amygdala asymmetry in control subjects was absent in the cocaine addicts. Topological analysis of amygdala isosurfaces (population averages) revealed that the isosurface of the cocaine-dependent group undercut the anterior and superior surfaces of the control group, implicating a difference in the corticomedial and basolateral nuclei. In cocaine addicts, amygdala volume did not correlate with any measure of cocaine use. The amygdala symmetry coefficient did correlate with baseline but not cocaine-primed craving. These findings argue for a condition that predisposes the individual to cocaine dependence by affecting the amygdala, or a primary event early in the course of cocaine use.


Breast-milk substitutes: a new old-threat for breastfeeding policy in developing countries. A case study in a traditionally high breastfeeding country.
Developing countries with traditionally breastfeeding are now experiencing the increasing pressure of formula milk marketing. This may endanger lives and undermine the efforts of national policies in achieving the objectives of the Millennium Development Goals. We examined the use of, and factors for use, of all available breast-milk substitutes (BMS) in a country with a traditionally high rate of breastfeeding. Randomised multi-stage sampling surveys in 90 villages in 12/17 provinces in Laos. 1057 mothers with infants under 24 months of age. 50-query questionnaire and a poster of 22 BMS (8 canned or powdered milk; 6 non-dairy; 6 formulas; 2 non-formulas). OUTCOME MEASURES INCLUDED: prevalence of use and age of starting BMS in relation to socio-demographic characteristics and information sources, by univariate and multivariate analyses. Of 1057 mothers: 72.5% currently breastfed; 25.4% gave BMS (10.6% infant formula); 19.6% gave BMS before 6 months of age (of them: 83% non-dairy or cereals; mean age: 2.9 months; 95% Confidence interval: 2.6-3.2). One formula and one non-formula product accounted for 85% of BMS. BMS were considered as milk by the majority of mothers. Thai TV was the main source of information on BMS for mothers. Lao Loum mothers (the main ethnic group) living in concrete houses with good sanitary conditions, were more likely than others to use BMS before 6 months (OR: 1.79, (1.15-2.78), p<0.009). Mothers who fed their infants colostrum at birth were less likely to use BMS before 6 months of age (OR: 0.63, (0.41-0.99), p = 0.04). Unemployed mothers living in rural areas were less likely to consider BMS better than breast milk. In Laos, mothers with the highest socio-economic status are showing a tendency to give up breastfeeding. Successful educational strategies and advocacy measures should be urgently developed to promote and sustain breastfeeding in developing countries.