Phoca vitulina) is a widespread marine predator in Northern Hemisphere waters. British populations have been subject to rapid declines in recent years. Food supply or inter-specific competition may be implicated but basic ecological data are lacking and there are few studies of harbour seal foraging distribution and habits. In this study, satellite tagging conducted at the major seal haul outs around the British Isles showed both that seal movements were highly variable among individuals and that foraging strategy appears to be specialized within particular regions. We investigated whether these apparent differences could be explained by individual level factors: by modelling measures of trip duration and distance travelled as a function of size, sex and body condition. However, these were not found to be good predictors of foraging trip duration or distance, which instead was best predicted by tagging region, time of year and inter-trip duration. Therefore, we propose that local habitat conditions and the constraints they impose are the major determinants of foraging movements. Specifically the distance to profitable feeding grounds from suitable haul-out locations may dictate foraging strategy and behaviour. Accounting for proximity to productive foraging resources is likely to be an important component of understanding population processes. Despite more extensive offshore movements than expected, there was also marked fidelity to the local haul-out region with limited connectivity between study regions. These empirical observations of regional exchange at short time scales demonstrates the value of large scale electronic tagging programs for robust characterization of at-sea foraging behaviour at a wide spatial scale.The harbour seal ( Phoca vitulina vitulina, which occurs in the eastern Atlantic, from Brittany (France) to the Barents Sea (Russia) and as far north as Svalbard (Norway). This paper presents an analysis of movement and foraging data from harbour seals tagged at seven regions around Britain; some of the most extensive data collected on harbour seal movements to date.Harbour seals have a widespread distribution across the Northern Hemisphere, from temperate to polar regions. There are five recognised sub-species; our study focuses on P. v. vitulina in northern Britain have recently declined by around 50% in less than 10 years Improving our understanding of the movements of harbour seals is of interest ecologically but is also important to conservation and management. Subpopulations of Although British harbour seal populations are monitored via aerial surveys of the numbers present at haul-outs Obtaining data on movement patterns from a greater spatial domain is important for several reasons. Determining how often individuals move to other subpopulations is critical for developing realistic models that help understand the spread of epizootics Knowledge of movements and at-sea usage data are useful in broadening our understanding of the distribution of ecosystem impacts of top predators. Feeding in both pelagic and benthic habitats, harbour seals are considered generalist predators and take a wide variety of fish, cephalopods and crustaceans e.g. . Their dSex and size of harbour seals have been found to be predictors of foraging trip distance and duration, in the Moray Firth (Scotland) in summer We expect seals to forage in a manner optimised for local conditions. In this study we concentrate on characterizing variability in foraging duration and trip length, which may be related to a combination of factors. These might include intrinsic factors such as a seal size, age, sex or body condition. Alternatively, local habitat characteristics such as substrate or water-depth might shape foraging behaviour. Interactions between habitat and intrinsic physiological factors could also influence the foraging strategy employed by a seal  and 60/4009  issued by the UK Home Office under the Animals (Scientific Procedures) Act 1986.Seal capture and handling was also authorised by the UK Scottish Office under annual licences given to the Sea Mammal Research Unit in accordance with the UK Conservation of Seals Act 1970.Harbour seals were studied in seven regions around Britain. These areas represent the major British harbour seal population These sites span a wide range of at-sea foraging and haul-out habitats. Like many pinnipeds, harbour seals periodically haul out on land to pup and moult, and between foraging bouts. Haul-out habitats in the islands to the west and north of Scotland typically consist of rocky intertidal areas, whereas intertidal sandbanks are used on much of the east coast of Scotland and England. Foraging habitat also varies spatially. Relatively deep water, characterized by a high proportion of rocky substrate, occurs in close proximity to haul-outs in the northern and western regions of the British harbour seal distribution. In contrast, the western North Sea consists of relatively shallow water with low-relief and sedimentary substrates.A range of techniques were used to catch seals depending on local habitat conditions. Where seals hauled out on intertidal sandbanks , most seals were caught using a long net (120 m), set from a boat around the haul-out site. The net formed a barrier within which seals were trapped as they fled to the water Seals hauled out in rocky habitats  were mostly caught using the ‘rush and grab’ technique in which a small inflatable vessel was driven quickly up to the haul-out area, deploying people carrying hoop nets to catch animals before they reached the water. If ‘rush and grab’ was not successful tangle nets were deployed in the area around the haul-out sites. This tended to be in areas where seals can reach deep water before capturers can get close enough in a boat, or else shallow rocks prohibited running a vessel at speed to the haul-out site. These loose-mesh nets were deployed at different angles around the haul-out sites to entangle seals when they swam into them. Buoys attached to either end of the net indicated when an animal was captured. Captured seals were untangled and transferred into the boat and restrained with a hoop net.All seals were weighed and then anaesthetized using 0.05 ml of Zoletil per 10 kg body mass delivered intravenously Transmitters were attached to the fur on the neck of the seal at the base of the skull using a fast setting two-part epoxy adhesive http://www.smru.st-andrews.ac.uk/protected/downloads/SRDL9000X.pdf). The SRDLs measured 100×70×45 mm (excluding the 150 mm antenna) and weighed 305g. The SRDLs collected, compressed and transmitted data via the Argos system ; a detailed description is given in Satellite-Relay Data Loggers (SRDLs) comprise a data logger interfaced to an Argos transmitter unit and a pressure (depth) and conductivity (submergence) sensor  for more than 10 minutes the seal was recorded as hauled-out; a haul-out period was determined to have ended when the tag became wet or submerged for a minimum of 16 s and at a depth greater than 2 m. Otherwise, a seal was classified as ‘diving’ at depths greater than 2 m and ‘at the surface’ at depths less than 2 m. The average and maximum dive depth within a 2 h summary period were also transmitted. Data were stored and transmitted at random to prevent bias in the data received.http://www.smru.st-andrews.ac.uk/protected/downloads/GPS_Phone_Tag22.pdf). These tags were programmed to send all information detailed above for SRDLs, but used Fastloc GPS technology which enables higher quality positions to be obtained using GPS satellites over very short surface intervals GPS phone tags were deployed on three seals in the Outer Hebrides and the Thames (see th percentile accuracies range from 150 to >1000 m (Service Argos 1996), however real data collected from seals fitted with both Fastloc GPS and ARGOS tags show that it is common for errors to be an order of magnitude greater than this −1. The remaining data are assumed to contain errors approximately distributed as a two dimensional normal distribution, allowing the Kalman filter to be applied , it was found that the combined speed/Kalman filter yielded root-mean square errors between 6–12 km calibrated against concurrently collected GPS data Argos locations are subject to varying degrees of spatial error. Service Argos reports that 68lied see . BecauseFor GPS phone tags, calibration studies have shown that 95% of locations are accurate to +/−55 m and, even when information from fewer than four GPS satellites is received, the mean error is less than 150 meters P. vitulina is a central place forager, the duration of trips is likely to be a key descriptor of differences in foraging behaviour among regions. Therefore tracks from individuals were classified into a series of individual ‘trips’. This task is not straightforward because of the inaccuracies in location data and also because of the summarized nature of the SRDL diving data . Without such additional data on feeding events we cannot quantify how much foraging occurred in shallow waters but we assume here that our choice of a 10 m depth threshold should not impact our conclusions.In five of the ten tags deployed in the Moray Firth population, haul-out and dive information could not be processed into trips using the approach outlined above because of tag malfunction. Therefore, for these data sets, we used a simple rule that assigned a trip start time when a seal’s estimated location was at least 10 km from a haul-out site and ended when within 10 km of a haul-out site. This was only possible in this area because of the nature of the prolonged distance and duration of these foraging trips (see below). To check this did not cause any significant upwards bias in the overall findings of the study the analysis was repeated on just the five animals tagged in the Moray Firth whose tags were operating correctly and the same patterns were apparent with little difference in the observed mean foraging duration or distance travelled. Note that this simple distance-based rule could not be used to categorise trips for all seals in all areas because for animals that travelled small distances from the haul-out site the errors remaining in filtered tracks were of the same order as the distance travelled.Other studies have used a distance from haul-out site threshold to trigger the initiation of trips Only foraging trips returning to the same region were included in the analysis, because it is not clear to which region a between-region trip should be assigned and because there were only a few between-region movements, as discussed in the The main focus of this study was to examine differences in the distribution of trip distance and duration when seals were foraging, while accounting for differences in body condition within regions and by time of year. However because movements of seals will be confounded with breeding status, data from breeding season months (June to August) were excluded from analysis allowing us to focus on foraging behaviour alone. During breeding periods, females restrict their foraging range during pupping and lactation We investigated whether two indices of foraging effort; the time spent at sea (trip duration) and the distance covered on a foraging trip (trip distance), could be predicted by a range of explanatory variables: the time of year ; inter-trip duration (the time between the start of current trip and the end of the previous trip); tagging region; sex; and seal length (a proxy for age). Note that inter-trip duration is not the same as haul-out duration because, by our trip-splitting rule, the seal could remain in shallow (<10 m) water for long periods.For pinnipeds foraging outside the breeding season, it may not be immediately clear whether the duration of a foraging trip or the distance covered per trip is a better indicator of foraging effort, and each has strengths and weaknesses. Trip distance has the advantage of being spatially explicit but, unlike trip duration, its accuracy depends on the accuracy of Argos correction routines. In contrast, the accuracy of trip-duration depends on the veracity of the criterion for when a trip starts and stops, although diving behaviour was more reliably measured than location – particularly at small scales. Aside from these observational aspects, distance covered may under-represent foraging effort if the foraging strategy entails short distance trips because animals could forage intensively for long periods close to the haul-out site yielding relatively small measures of trip distance. On this basis, trip duration may be a better gauge of changes in foraging effort. Therefore, we use trip duration as an index of foraging effort but also report the regional differences in trip distances.Tdur, the foraging trip duration calculated as the elapsed time between the start and end of a foraging trip. We also calculated Tdist, the foraging trip distance calculated as distance between position locations. For the reasons outlined above and the non-independence of durT and distT  we calculated dist see  only TduTdur mandates the use of statistical models capable of handling non-Gaussian response variables and also capable of modelling individual variability between seals. We also needed to allow for the possibility of auto-correlation in Tdur. Therefore we used Generalized Additive Mixed Models (GAMMs). Our most complex or ‘saturated’ model can be expressed asThe strictly positive distribution of L), sex (S), inter-trip duration (ITD), day-of-the-year (DOY) and region (R) as explanatory variables of trip duration. The term DOY was calculated as days after 1 October, which indicated the start of seasonal data collection after breeding and moult period data had been discarded. In model (1) the subscript i indexes individual foraging trips, j indexes seals and k indexes the seven different tagging regions. In model (1), terms with two covariates denote interactions. The notation f(.) denotes non-parametric smooth terms in the GAMMs which were modelled using thin-plate splines and included seal length  functions. Similarly the effect of inter-trip duration was allowed to vary between regions. Finally the possibility of autocorrelation in trip-duration was modelled by a first-order autoregressive error structure, hereafter referred to as AR(1).Day of the year was included as an integer which equalled zero on 1 October, was modelled as a non-parametric smooth term and was assumed to interact with region, Because GAMMs were fitted using Penalized Quasi-Likelihood estimation The body condition of an individual seal might determine its ability to remain foraging at sea. We investigated whether there were differences in condition factor (length (cm)/mass (kg)) among different regions. This was important as differences in condition within region may alias any other regional signal. Initial inspection showed approximate linearity in the relationship log(mass) at length. Therefore on the basis of parsimony, we a used linear modelkβ, and via the interaction of Region/Sex we tested whether the slope of this relationship differed among regions and between sexes.to determine if there were regional differences in the Length/Mass relationship via the coefficient In total, 118 harbour seals were caught and tagged between November 2001 and October 2006. Of these, 115 seals were tagged with Satellite Relay Data Loggers (SRDLs) and the remaining three were tagged with GPS fastloc tags. The number of tags from each of the seven tagging locations is shown in Harbour seal movements were not restricted to near-shore waters; seals often undertook lengthy trips to offshore locations . RegionaThe majority of animals were site-faithful in their repeated use of the same or nearby haul-out sites, however a small proportion of animals did travel between regions and even between countries . In the Also apparent are what appear to be long distance exploratory movements into the North Sea . MovemenPatterns of regional trip durations were replicated in the distributions of trip distance, with long duration trips being made on the east coast of the UK and short trip durations in the Northern Isles and Outer Hebrides . Moray FComparison of trips made across regions stratified by sex  shows thThe relationship between trip distance and duration was approximately linear . However2 = 0.17–0.23, Comparison of a range of GAMMs resulted in selection of a model containing only inter-trip duration, DOY and the categorical variable region . The fitIt is important to note that making valid statistical inference is difficult with non-Gaussian random effects models The selected GAMM predicted that irrespective of the time of year, longer trips were expected in The Wash, St Andrews Bay and the Moray Firth relative to other regions. Notable differences were also apparent in the seasonal changes in expected trip duration. The longest trips for Orkney, Shetland and the Outer-Hebrides were predicted for January and February; trip length for the Moray Firth peaked in December. The longest trips in the Thames were predicted for January, although the variation here was smallest of all the regions and trip length could essentially be regarded as constant . Trip duInter-trip duration was retained in the selected model of trip duration. Despite the selection of the model with AR(1) correlations between trips, there was little evidence of temporal correlation in either the raw trip duration data or the residuals from the fitted models (AR(1) correlation parameter  = 0.03) which indicates only a small statistical dependence from one trip to the next. However, because inter-trip duration was retained as a significant explanatory variable this may indicate temporal dependence in overall foraging strategies. Additionally, retention of the smooth functions for day of the year suggest intra-annual trends in trip duration. However, there was considerable variation among regions in the timing of peaks in trip duration. The sites in the North Sea  showed a much larger signal of seasonal variation compared to the Outer Hebrides, Orkney and Shetland. This may indicate that the food resources in these latter regions tend to be more stable.2 = 0.53), as expected, but also that there were no differences in weight/length relationships between sex  or within region . Although the sample size (N = 118) is too small to characterize properly the weight-length relationship, it does suggest that within this dataset, condition did not vary significantly among regions and we therefore conclude that the apparent regional differences in foraging strategy were not aliasing regional differences in the condition of the seals. However, this does not preclude other aspects of foraging (such as diving ability or prey selection) being related to individual condition. Nonetheless, these data support the idea that a large component of the variability in foraging behaviour is due to region-specific variables. Additionally, models which did not contain region as a categorical factor variable were ranked poorly relative to models which did. This pattern, in which time of year and inter-trip duration were the most effective explanatory variables was also found when regional subsets of the data were modelled separately. For brevity these results are not given.Estimation of model (2) indicated a strong relationship between weight and length  Studies from other parts of this species’ range found similar variation. For example harbour seals tagged in Prince William Sound, Alaska were found to be moving between 5–25 km from haul-outs to at-sea foraging locations although some sub-adults made wider ranging movements of 300 to 500 km outside the sound where they were tagged The intrinsic factors of sex and length explained little of the large variability within regions. This result that the largest differences were mostly among regions is consistent with harbour seals adopting foraging strategies tailored to their local habitat. Obvious differences in habitat exist for animals hauling out on rocky shores in close proximity to deep water (around the north and west of Britain) and those sites where animals mostly haul out on intertidal sandbanks bordering the shallow and gently sloping sedimentary western North Sea (on the east coast of Britain). Relative to shallow water sedimentary sites, animals at the sites bordering deeper water, rocky bottomed sites, tended to undertake foraging trips of shorter distance and duration.The characteristics of the local habitat as prey refugia may partially explain increased foraging distances in areas of sedimentary habitat. In soft bottom habitats prey are more accessible to predation and thus may be more depleted. Areas with rocky bottom substrates may provide prey refugia preventing as much prey depletion close to the haul-out site.If we assume that seals aim to minimize energy expended during foraging trips, our results imply that harbour seals hauling out adjacent to shallow soft-sediment habitats need to travel greater distances to find sufficiently productive foraging areas from which they can maintain comparable levels of body condition, and presumably fitness.Further fine-scale analysis of foraging, such as examination of the diving behaviour of seals, also collected by tags in this study, may elucidate further on the characteristics of preferred foraging areas and whether foraging is largely benthic or pelagic within the study regions.Studies of foraging in breeding seabirds, another central place forager, have shown an increase in prey size, as well as a reduction in benthic foraging and increase in pelagic foraging, with distance from the colony The exception to this pattern was the Thames, where, despite being a gently sloping sedimentary site, animals were found to be travelling only short distances to forage. However, only small numbers of harbour seals haul out at this site, likely because it is a relatively poor quality haul-out site; the sandbanks are adversely affected by the prevailing weather and are exposed for only a small proportion of the tidal cycle. The sandbanks in the Thames may represent marginal haul-out habitat for harbour seals and it is possible that limited intra-specific competition for available prey allows the persistence of relatively small populations.Our measure of body condition did not differ significantly by region suggests that the different foraging strategies employed in different regions achieve similar levels of individual body condition. An important caveat is that our study animals are not representative of the whole population because animals smaller than 50 kg were not tagged and thus juveniles were not represented. Given the observed population declines around Britain Highly sexually dimorphic species of pinnipeds such as grey and elephant seals have been shown to exhibit differential movement and space use according to sex Differences between the sexes in harbour seal foraging distance and duration have been observed previously in the Moray Firth; however, that study was focused on late spring and early summer months when differences are likely to be most pronounced prior to the breeding season Time of year was found to be an important variable in explaining variation in foraging trip duration and distance travelled, a result that corroborates previous studies which found that harbour seals around the coast of Britain spend more time away from haul-out sites during the winter months The North Sea ecosystem has shifted markedly in recent decades, greatly affecting relative abundances of fish stocks For central-place foragers, intra-specific competition should play a part in determining the distance animals are travelling to forage as resources closest to the central place are expected to be most depleted. Consequently, in areas where relatively large numbers of harbour seals haul out, animals may be expected to travel further in order to obtain sufficient food. Although there is some suggestion of this in our data, , there are a number of complicating factors that probably obscure any relationship of increasing distance travelled with increasing population size. Most importantly, we do not have data on prey availability in the different regions. Also each region differs in the area available in which to search for nearby prey; colonies on islands have a larger area of water surrounding the haul-out; colonies within an enclosed coastline have a reduced area of water close to the haul-out. Finally, there are large differences between colonies in the depth of water surrounding the colony, obviously leading to a completely different species makeup in the prey resources.Intra-specific competition can also result in sex and age specific foraging strategies Regional differences in foraging behaviour could be influenced by competitive interactions with other species and competition with grey seals has been suggested as contributing to recent harbour seal declines. Grey seals have been tagged throughout their range around Britain A study comparing foraging of harbour and grey seals in the inner Moray Firth did find dietary and spatial overlap in the foraging of the two species although grey seals utilized a wider area and generally travelled further Unlike in other parts of their range, natural predators of seals are rare in British waters; however observations of predation on harbour seals by killer whales have been observed in northern and western Scotland, primarily in the pupping season and most frequently in northeast Scotland and Shetland Our study has shown that harbour seals forage more extensively in offshore waters than previously believed and should not be described as a purely coastal marine mammal. Our study also revealed notable interchange of individual seals between regions and countries. Conservation and management of the species needs to take this movement into account when considering population units. These new observations of at-sea distribution and connectivity may have an important bearing on our understanding of relationships among individuals primarily associated with different haul-out sites and how this affects susceptibility to epizootics and competition for prey, both of which are important for the conservation of a species in decline. The extensive distances travelled to forage mean that management of offshore fish stocks may be as important for harbour seals as management of inshore stocks.That our results show that variation in foraging movements was better accounted for by region and season than by sex, size, and body condition, may also have an important bearing on harbour seal conservation. It suggests that management action to reverse population declines may be best focussed in an area-specific way. The EU Natura 2000 initiative to establish networks of Special Areas of Conservation (SACs) for habitats and species may be particularly relevant here e.g. . SACs haFinally, this study demonstrates the value of a large dataset collected over a wide spatial range, without which our results would be of little more than local interest. Instead, the range of habitats and population sizes studied makes our results relevant to the ecology and conservation of harbour seal populations worldwide. The expansion of our view of harbour seal distribution in British and surrounding waters would be further developed by spatial modelling of seals’ distribution. This could identify important features of high-use areas for harbour seals, which could also aid conservation populations of this species and marine conservation planning in general.Appendix S1Tagging details for individual seals.(DOCX)Click here for additional data file.Appendix S2Regional trip distance and duration by sex.(DOCX)Click here for additional data file.


The “positivity effect” refers to an age-related trend that favors positive over negative stimuli in cognitive processing. Relative to their younger counterparts, older people attend to and remember more positive than negative information. Since the effect was initially identified and the conceptual basis articulated (Mather and Carstensen,  The positivity effect refers to a relative preference in older adults (compared to younger adults) for positive over negative material in cognitive processing. Since the first explicit reference to the positivity effect in 2004  rather than bottom-up and fixed .Early in the last decade, our research team began to test hypotheses about the ways in which motivational changes postulated by SST may influence cognitive processing. These efforts expanded upon a large and rich literature in psychology documenting the powerful influence that goals exert on cognitive processing. From classic studies by Neisser and colleagues on inattentional blindness  control, when individuals are allowed to pursue chronically activated goals without external interference, and when regulating emotions contributes to well-being. Conversely, the effect should not appear when cognitive resources are limited, when information processing is automatic, when contexts impose situation-specific goals that conflict with chronically activated goals, and when prioritizing emotion regulation has significant risks. To assess the empirical support for these predictions we review the extensive research literature on the positivity effect.In reviewing the empirical literature we focus on three key conceptual issues: First, we present evidence suggesting that the positivity effect represents controlled processing not cognitive decline. Second, we critically examine experimental procedures and conditions under which the positivity effect is and is not observed, illustrating that the effect is malleable rather than unreliable. Finally, we consider whether effect is adaptive or maladaptive for older adults’ everyday functioning.According to the motivational perspective offered by SST, the positivity effect stems from age-related shifts in goal priorities that increase the salience of emotionally gratifying information in attention and memory. Because cognitive resources are required to direct information processing toward goal-relevant stimuli and away from less relevant stimuli Mather, , the posThe critical role of cognitive resources in the positivity effect is further highlighted by research comparing the emotional memory of healthy younger and older adults with older adults suffering from Alzheimer’s disease  implicated in the regulation of emotion to a greater extent when processing negative versus positive images  follows the opposite age-by-valence interaction  brain waves  and whether to invest retirement savings in a new company? Preliminary evidence suggests that the answer may be no. In a recent study by our research group  faces faster than other emotions , Hess et al.  decisions across age groups and as a function of information valenceDecision quality remains an important and understudied issue, and there are many reasons to expect that decision quality may suffer with age , when experimental tasks or stimuli elicit automatic processing or when situational demands supplant chronically activated goals. It appears increasingly that positivity may be reduced when the stakes are high. Taken together, the phenomenon appears to reflect a default cognitive processing approach in later life that favors information relevant to emotion-regulatory goals. Older people place high value on goals related to well-being and, all things being equal, cognitive processing operates under the influence of such goals.In less than a decade, the positivity effect has become a well-replicated empirical observation. As more evidence accumulates and the guiding theory becomes more nuanced and detailed, it will be possible to test its various aspects with greater precision.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


As the prevalence of diabetes mellitus continues to rise, innovative medical and surgical treatment options have increased dramatically to address diabetic-related foot and ankle complications. Among the most challenging clinical case scenarios is Charcot neuroarthropathy associated with soft tissue loss and/or osteomyelitis. In this review article, the authors present a review of the most common utilizations of negative-pressure wound therapy as an adjunctive therapy or combined with plastic surgery as it relates to the surgical management of diabetic Charcot foot and ankle wounds. Negative-pressure wound therapy (NPWT), also known as topical negative-pressure therapy or vacuum-assisted closure, has revolutionized the world of wound care. Morykwas et al. first published on this modality using an open-cell foam dressing with application of a controlled subatmospheric pressure for treatment of acute and chronic wounds in 1997 . Since tMuch of the existing evidence supports that the success of NPWT in wound healing is likely multifactorial. A study by Zöch on 10 patients found an increase in blood flow to diabetic foot wounds treated with NPWT measured by IC-view perfusography . Kamolz As the conventional option for care of diabetic foot wounds has been moist wound dressings, randomized controlled trials comparing NPWT to moist wound healing have populated the literature. Eginton et al. compared the rates of wound healing between NPWT and moist dressings, clearly showing improved healing rates in the diabetic foot wounds treated with NPWT . In regaNeuropathic wounds in the diabetic CN foot and ankle can manifest through a variety of different pathways and mechanisms. Most common clinical case scenarios that can lead into a CN deformity from a pre-existing neuropathic ulceration(s), osteomyelitis and/or surgical intervention may include but are not limited to the following two clinical presentations:  diabeticFor any other patient with diabetic CN, identification of the stage of CN is important in formulating a treatment strategy. In the acute stage of CN, inflammation may produce severe edema that when combined with ambulation in the insensate patient can cause significant skin breakdown and/or compromise a pre-existing wound. NPWT can be carefully applied during this stage if a wound is present but must be combined with a period of immobilization in order to halt the inflammatory process, limit peri-wound edema, and maintain the foot and/or ankle architecture to prevent further deformity . ImmobilAs CN can compromise the osseous architecture and stability of the foot and ankle, the chronic form of the condition typically presents with progressive deformity resulting from a cumulative effect of repeated injury that goes undetected or neglected during the acute process. Failure to properly accommodate and off-load braceable deformities or to recognize unstable and/or unbraceable deformities which require surgical intervention may potentially lead to an ulceration and/or infection. In this clinical scenario, patients often present to a medical facility because they have noticed a wound and possibly a misshapen foot, yet cannot recall any of the signs or symptoms of acute CN. For these patients in the absence of infection, the foot and ankle specialist should identify whether the deformity is stable or unstable in order to determine whether surgical intervention may be indicated for correction of an unstable deformity. In a wound with stable chronic CN, sharp debridement of the wound and use of NPWT can facilitate secondary healing, while re-ulceration can be avoided through accommodative shoe gear and/or bracing combined with frequent clinical examinations.Delayed or ineffective treatment of CN foot and ankle wounds in the diabetic population can potentially lead to soft tissue infection and osteomyelitis. Appropriate surgical debridement of infected soft tissue and bone often leaves large defects that may be amenable to treatment with non-biodegradable cemented antibiotic beads/spacers. In these surgical case scenarios, NPWT may only be considered for surgical wounds with eradicated bone or soft tissue infection, possess adequate perfusion, and are in need for frequent dressing changes. For exposed tendon structures, a non-adherent dressing can be applied first directly over the tendon prior to placement of the NPWT foam. Furthermore, commercially available open-cell foams of different compositions (white foam or silver-impregnated foam) are designed for placement over bone or in tunneling wounds; however, the efficacy of these materials is yet to be determined. Although NPWT is beneficial as an adjunct of treating these complex case scenarios, a thorough surgical debridement and appropriate antibiotic therapy are necessary to a successful wound healing in the diabetic CN foot or ankle .NPWT is also indicated as an adjunct to surgical wound closure utilizing plastic surgical techniques and osseous reconstruction for the unstable and/or unbraceable CN deformity. Due to the effects of NPWT mentioned earlier, this modality can be useful in wound bed preparation prior to definitive wound closure techniques such as skin grafts or flaps. NPWT can also be considered to promote wound healing at flap donor sites. When used cautiously, observed frequently and typically at lower pressure levels, NPWT can also be beneficial directly over skin grafts or flaps during early incorporation. In a clinical trial comparing NPWT with standard non-adherent gauze dressings for skin-grafted diabetic foot wounds, Paola et al. found that the percentage of subjects with complete skin graft healing was increased in the NPWT group . ReductiSimilarly, NPWT has also found a role in stabilizing closed incisions which is useful in extensive CN foot and ankle reconstructions where there is a high risk for wound dehiscence. Arthrodesis procedures often used for reconstruction of unstable CN foot or ankle fracture dislocations can produce tenuous incisions and may lead to significant post-operative edema. A study by Kilpadi et al. demonstrated that the use of NPWT over closed incisions can reduce and normalize tissue stresses and bolster appositional forces at the incision site . In CN rPrior to application of NPWT in any patient, one should consider the existing contraindications to this modality . AbsolutSince its initial development, NPWT has gained widespread acceptance for a broad range of wound indications, including those found among diabetic CN. NPWT can be used to treat CN wounds produced as a result of neuropathy and deformity, following debridement of infection or amputation, and in reconstructive soft tissue and osseous procedures. Several studies emphasize that treatment outcomes are often based on the specific techniques and materials used for NPWT application, therefore additional suitably powered, high-quality clinical trials are needed to fully determine efficacy. Careful patient and procedure selection along with appropriate technique is imperative for successful use of NPWT in the diabetic CN foot and ankle.


With the advent of high-throughput technologies, the field of systems biology has amassed an abundance of “omics” data, quantifying thousands of cellular components across a variety of scales, ranging from mRNA transcript levels to metabolite quantities. Methods are needed to not only integrate this omics data but to also use this data to heighten the predictive capabilities of computational models. Several recent studies have successfully demonstrated how flux balance analysis (FBA), a constraint-based modeling approach, can be used to integrate transcriptomic data into genome-scale metabolic network reconstructions to generate predictive computational models. In this review, we summarize such FBA-based methods for integrating expression data into genome-scale metabolic network reconstructions, highlighting their advantages as well as their limitations. A central challenge in the development of systems biology is the integration of high-throughput data to generate predictive computational models. With the advent of high-throughput technologies, “omics” data types have provided quantitative data for thousands of cellular components across a variety of scales. Genomics provides data on a cell's DNA sequence, transcriptomics on the mRNA expression of cells, proteomics on a cell's protein composition, and metabolomics on a cell's metabolite abundance. Computational methods are needed to reduce this dimensionality across the wide spectrum of omics data to improve understanding of the underlying biological processes , a constraint-based modeling approach, can be used to probe these network reconstructions by predicting physiologically relevant growth rates as a function of the underlying biochemical networks  algorithm  and sets of lowly expressed reactions (RL). The algorithm then solves a mixed integer linear programming (MILP) problem according to stoichiometric and thermodynamic constraints while also maximizing RH and minimizing RL. Mirroring GIMME, the presence of reactions is allowed to deviate from the expression data in order for the algorithm to result in a functioning model. Thus, like GIMME, iMAT returns a functioning metabolic model that integrates expression data with a genome-scale metabolic network reconstruction by maximizing the number of highly expressed reactions and minimizing the number of lowly expressed reactions; however, unlike GIMME, this method does not demand that the user supply functionalities that the model is assumed to meet. Instead, iMAT requires that reactions catalyzed by the products of highly expressed genes are able to carry a minimum flux. By removing this need for user-specified objective functions, iMAT bypasses assumptions about metabolic functionalities of a particular network, which proves advantageous for models where there is no clear objective function, as in models of mammalian cells.Similar to GIMME, the Integrative Metabolic Analysis Tool (iMAT) results in a functioning model in which the fluxes of reactions correlated with high mRNA levels are maximized and the fluxes of reactions associated with low mRNA levels are minimized  uses statistically significant changes in gene expression measurements to determine sequences of highly and lowly expressed reactions  aims to fuse together metabolic networks and transcription regulatory networks with expression data (Chandrasekaran and Price, Each of the methods discussed hinges on the assumption that mRNA transcript levels are a strong indicator for the level of protein activity. For instance, GIMME and iMAT assume that mRNA levels below a certain threshold suggest that the corresponding reactions are inactive. MADE follows a similar logic, turning reactions on and off depending on the changes in mRNA transcript levels. E-Flux and PROM assume that transcript levels indicate the degree to which reactions are active, evident in the constraining of the upper bounds in the FBA optimization problems associated with these methods.However, recent studies have called into question the validity of this assumption which correlates mRNA transcript levels to protein levels. In one of the first large-scale screens to assess the correlation between mRNA and protein expression, the Pearson product moment correlation coefficient, a measure of the linear dependence between two variables ranging from −1 to 1, was calculated to be 0.356 for yeast data (Gygi et al., Acknowledging this questionable association between transcriptomic and proteomic data, the discussed methods claim to treat the assumption that mRNA levels serve as a strong indicator for protein activity as a “soft constraint.” Rather than requiring that the reconstruction mirror the expression data exactly, the methods allow for deviations in the FBA flux solution space in order to generate a functioning model that adheres to the specified constraints. In the case of GIMME, highly expressed reactions are prioritized relative to lowly expressed reactions; however, in the event that an optimal, functioning solution cannot be found, the assumption can be violated and lowly expressed reactions can be added back into the reconstruction. Thus, this assumption that mRNA transcript levels correlate to protein levels serves as a cue rather than a mandate.Mycobacterium tuberculosis (Colijn et al., The above methods have been used to not only integrate expression data from a variety of sources but to also make progress toward overcoming key challenges in the field of systems biology. For instance, iMAT, highlighting its applicability in multi-cellular organisms, was used to curate the human metabolic network reconstruction and predict tissue-specific gene activity levels in ten human tissues (Duarte et al., Given the recent success with using genome-scale metabolic network reconstructions as a platform for integrating expression data, efforts should focus on multi-omics data integration. A handful of methods have already been introduced that integrate two or more types of omics data into genome-scale metabolic network reconstructions. For example, despite the current dearth of quantitative metabolomics data, a method has been developed that demonstrates how semi-quantitative metabolomics data can be used with transcriptomic data to curate genome-scale metabolic network reconstructions and identify key reactions involved in the production of certain metabolites (Cakir et al., The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


The dorsal anterior cingulate cortex (dACC) has previously been implicated in processes that influence action initiation. In humans however, there has been little direct evidence connecting dACC to the temporal onset of actions. We studied reactive behavior in patients undergoing therapeutic bilateral cingulotomy to determine the immediate effects of dACC ablation on action initiation. In a simple reaction task, three patients were instructed to respond to a specific visual cue with the movement of a joystick. Within minutes of dACC ablation, the frequency of false starts increased, where movements occurred prior to presentation of the visual cue. In a decision making task with three separate patients, the ablation effect on action initiation persisted even when action selection was intact. These findings suggest that human dACC influences action initiation, apart from its role in action selection. The human dACC has been implicated in various aspects of action selection, including reward-dependent decision making Because action initiation and action selection are co-dependent in many tasks, separating these two roles in human dACC has been difficult. This is because initiation is modulated by components of action selection including reward expectancy Early anatomical work on dACC in monkeys established direct connections from neurons in the cingulate sulcus to primary motor cortex and spinal cord, and preliminary efforts were made through fMRI to identify homologous regions in human. In the seminal monkey work, a group of cingulate motor areas (CMA) were defined through cytoarchitectural examination and electrical stimulation studies on the basis of connections to primary motor cortex and the spinal cord The homology between monkey and human cingulate motor areas is widely employed, but this relationship is largely based on resemblance between PET and fMRI activation patterns in human and divisions of the medial frontal cortex in monkey In non-human primate, dACC has been linked to reward-dependent decision making Some work in rat dorsomedial prefrontal cortex (dmPFC) has attempted to provide causal statements about movement initiation, although the homology between dmPFC and human dACC is further removed and controversial Recently, several groups have used human cingulotomy, undertaken for purely therapeutic indications, as an opportunity to study the function of the human dACC Patients were enrolled in this study through informed written consent under a protocol approved by the Institutional Review Board and the cingulotomy assessment committee at Massachusetts General Hospital. The decision to offer surgery bore no relation to the current study, and adhered to the same evaluative and ethical guidelines used for all prior cingulotomy patients. At all time points, patients had the understanding that their participation bore no relation to the surgical outcome, and that they could withdraw from the study at any time.Six patients were studied before and after bilateral radiofrequency ablation of dACC  as a last-resort surgical treatment, illustrated in after two small bilateral burr-holes were made to introduce the ablation electrode. Post-ablation data was collected under identical intra-operative conditions, within 5 minutes after completing bilateral dACC ablation and approximately 30 minutes after the last pre-ablation trial. The task was run in blocks of 30 or 40 trials. Extra-operative and training trials were excluded from analysis. Total trial numbers are listed in Patients were trained on the tasks prior to surgery. Pre-ablation data was collected while the patient was situated on the operating table delay), and the feedback screen was presented after a movement was detected, or 3 seconds from the go cue, whichever came first. Patients were instructed to pull the joystick, initiating movement as rapidly as possible following appearance of the red circle, but not prematurely. This instruction to the patients was identical for all trial types, including exit and non-exit trials. Patients were not explicitly cued whether trials were exit or non-exit type. The feedback screen provided a pictorial representation of premature response or non-premature reaction time. The onset of the white circle was accompanied by a brief acoustic tone. To choose trial type and Tdelay, a number Texp was drawn from an exponential distribution. For Texp>1, an exit trial was presented. For Texp<1, a non-exit trial was presented with Tdelay = 1+Texp. The exponential distribution parameter was chosen such that 40 percent of trials were exit trials. Reaction tasks commonly elicit anticipatory rather than reactive behavior because go cues are predictable The simple reaction time task  consisteThe reward-dependent decision making task  consistePatients were instructed and trained in their respective tasks for a cumulative time of less than 30 minutes in a day preceding surgery, or in the minutes preceding entry into the operating room. There was no evidence for learning, including the more complex reward-dependent decision making task as reported previously http://svi.cps.utexas.edu/software.shtml). Visual stimulus times were detected with a photoresistor, and movement onset time was recorded for feedback as the first detectable displacement from origin based on a simple threshold. Joystick displacements and photoresistor outputs were separately recorded with a data acquisition system  for off-line detection of stimulus and movement onset times based on simple thresholding in MATLAB. Incidence of false start fractions was estimated with the Clopper-Pearson interval. Significance testing of differences in false start fractions between pre- and post-lesion sessions was performed using a two-tailed Pearson Chi-square test at p<0.05.Behavioral control software was written in Matlab with the Data Acquisition Toolbox and Space Variant Imaging Toolbox (As shown in a representative high resolution (MPRAGE) T1-weighted MRI scan of a patient different from S1–S6 , each heWe examined the ablation extent with relation to the previously defined discrete motor areas of the human medial frontal cortex We first examined the fraction of movements generated prior to the go cue. We also analyzed false start fractions in comparison to action selection errors in the reward-dependent decision making task (S4–S6). We previously reported for these patients that bilateral dACC ablation profoundly increased action selection errors in the reduced reward and double arrow conditions, but not in the standard reward condition In In order to determine the extent of statistical dependence between choice errors and false starts, we asked what percent of false start movements also registered correct choice answers, relative to the overall percentage of false start movements. In technical terms, recall that statistical independence between two random variables x and y is equivalent to p(x) = p(x|y). In reduced reward and double arrow trials following dACC ablation, choice errors were under-represented in false start trials, inconsistent with statistical independence in reduced reward and double arrow conditions. Specifically, only 16+/−14 percent of false start trials resulted in choice error, despite an overall choice error of 46+/−2 percent in these trials (p<0.05). For standard reward trials, the rate of choice error in false start trials was indistinguishable (p<0.05) from the overall rate of choice error (nearly 0 percent), consistent with statistical independence in standard reward conditions. There, only 1 trial out of 42 trials resulted in choice error, which was also a false start trial.Finally, we examined the effect of ablation on late responses in the reaction time task. A late response was defined as a response where no movement was registered when a go cue was issued. Intuitively, the percentage of late response is a measure of task engagement. In S1 and S2, no late responses were registered in any testing. In subject S3, late response percentages were also not significantly changed following ablation. In this subject, percentage late response measured 7% pre-ablation  and 15% post-ablation  post-ablation.”In navigating daily life, we are frequently called upon to make choices (action selection), and to act upon those choices (action initiation). Previous research has implicated the dACC in action selection, with other correlative studies suggesting it may also be involved in action initiation. Our results provide evidence that dACC is causally involved in action initiation, separate from its involvement in action selection. We demonstrated that anatomically and temporally focused dACC ablation immediately disrupted action initiation even when action selection is intact or when actions are pre-selected. Below, we discuss issues of correlation versus causality, and address possible confounds in interpretation. We conclude with a discussion of our results in the context of the various prevalent models of dACC function.Establishing causality between human dACC and elements of behavior has been challenging for a variety of reasons. Imaging and electrophysiology studies are fundamentally correlative. Transcranial magnetic stimulation (TMS) has been useful in establishing causal roles for more superficial cortical structures In comparison, cingulotomy provides several comparative advantages to pathological lesion studies. The ablation is focal, surgically controlled in position, extent, temporal onset, and mechanism (thermocoagulation), and readily visualized by MRI. The effects of dACC ablation can be studied intra-operatively, immediately preceding and following ablation. This minimizes potential variability due to cortical re-organization, mass effect from post-ablation edema, or behavioral adaptation.Could increased premature responses following ablation be entirely explained by dACC's causal role in reward-dependent decision making The experimental design helps to address several other potential confounds. It is unlikely that the action initiation deficit is a simple placebo effect of the ablation, as patients were not told to expect any action initiation deficit and the action initiation deficit did not relate to the expected therapeutic benefits for OCD, depression, or bipolar affective disorder. The effect is unlikely to be specific to a particular experimental paradigm because the action initiation effect persisted across two entirely separate tasks. Finally, the ablation effect on action initiation is unlikely to be attributable to operating room conditions or neurosurgery. This is because our experiment allowed each patient to serve as their own internal control where pre- and post-ablation data was collected under the same intra-operative conditions. Additionally, the pre-ablation data were collected following placement of a stereotactic frame and the start of surgery, so that effects of these neurosurgical interventions would have been present in the pre-ablation data. The post-ablation trials occurred within 5 minutes of the ablation procedure, temporally associating the observed deficits with the ablation event itself.Because this experiment represents a surgically invasive human study, performing the lesion experiment on healthy volunteers is not an option. As a result, we cannot be certain that the effects of dACC ablation on action initiation necessarily generalize to the healthy brain. However, we can be reassured that the ablation effects were not specific to any particular disorder. Specifically, two of these patients received dACC ablation for otherwise refractory major depression or bipolar affective disorder while the remainder had OCD. Moreover, as discussed above, the surgical ablation procedure mitigated concerns raised by retrospective stroke or tumor studies, such as effects of cortical plasticity or paraneoplastic phenomena. Finally, prior work that implicated dACC in action initiation used healthy volunteers Our analysis confirms dACC ablation results in action timing deficits even when decision making is intact (standard reward condition of the reward-dependent decision making task) or when decision making is entirely removed from a task (simple reaction time task). Consequently, deficits in action timing are not simply an epiphenomenon of dACC's role in decision making. Paradoxically, when choice errors and false starts coexist (reduced reward and double arrow conditions), choice error is decreased in false start trials relative to the overall choice performance. This is contrary to what might be expected from noisy integration in an integrate-to-threshold model, where false starts would be enriched with choice errors. Instead, under these conditions, dACC ablation may actually allow choice certainty to potentiate false starts.With regards to reaction times, our prior work on reward-dependent decision making showed that S4–S6 together demonstrated no significant difference in reaction times between correct and incorrect decisions One prevalent theory suggests that areas of medial frontal cortex are variously involved in response inhibition, arising from early studies of anatomically non-specific medial frontal lesions in humans during go/no-go and related tasks Is it possible that damage to SMA could explain the ablation effects? Prior work in monkey had investigated spiking neural activity in SMA versus CMAd and CMAv in a classical delayed instructed center-out reaching task A complementary theory based on primate and human studies states that medial frontal cortex participates selectively in voluntary rather than reflexive actions Accumulator models describe reaction time as a consequence of the integration of a noisy signal, with actions initiated when this integration crosses a threshold. These models are mathematically compact. However, accumulator models are black box, i.e. they are not biophysically detailed. These models are not specifically related to anatomy of dACC or any other brain region. This is problematic because it limits the model's practical utility in predicting the effects of lesions immediately beyond the vicinity of the area studied. At their core, these models are parametric descriptions of the data for a specific experimental setting, with limited predictive power beyond the conditions of the experiment.A counter argument in support of accumulator models is that by mapping changes in reaction time distributions onto changes in parameters of an accumulator model, results can be interpreted in terms of this model's language. For example, we might say that the effect of dACC lesions was to lower the decision threshold, or to increase the noisiness of integration. Even if the equivalent effect on accumulator model parameters was relevant in some context, pre-ablation false start rates in this experiment were nearly zero in 4 out of 5 patients that demonstrated the effect see . As a reThe theory by Frank, et. al in Interestingly, the authors posit that STN derives the level of conflict from cingulate and neighboring areas, supported by anatomical connectivity and incorporation of the separate finding that cingulate fMRI activation correlates with conflict level Both previous studies and our current results suggest that dACC is not the unique, exclusive determinant of action initiation. Ablation studies of comparable spatial and temporal precision are not readily available for other regions of human frontal cortex. However, retrospective studies of heterogeneous frontal lesions have demonstrated response inhibition as being affected by various areas of medial and lateral frontal cortices


Idiopathic autism, caused by genetic susceptibility interacting with unknown environmental triggers, has increased dramatically in the past 25 years. Identifying environmental triggers has been difficult due to poorly understood pathophysiology and subjective definitions of autism. The use of antidepressants by pregnant women has been associated with autism. These and other unmetabolized psychoactive pharmaceuticals (UPPs) have also been found in drinking water from surface sources, providing another possible exposure route and raising questions about human health consequences. Here, we examined gene expression patterns of fathead minnows treated with a mixture of three psychoactive pharmaceuticals  in dosages intended to be similar to the highest observed conservative estimates of environmental concentrations. We conducted microarray experiments examining brain tissue of fish exposed to individual pharmaceuticals and a mixture of all three. We used gene-class analysis to test for enrichment of gene sets involved with ten human neurological disorders. Only sets associated with idiopathic autism were unambiguously enriched. We found that UPPs induce autism-like gene expression patterns in fish. Our findings suggest a new potential trigger for idiopathic autism in genetically susceptible individuals involving an overlooked source of environmental contamination. Autism spectrum disorders (ASD) are characterized by stereotyped behaviors and impaired social skills, typically diagnosed by three years of age In order to serve as such a trigger, a candidate teratogen must have a biologically plausible etiological mechanism, exist in sufficient environmental concentrations, be capable of passing from mother to fetus and across the fetal blood-brain barrier , and have experienced historical increases in environmental concentration parallel with observed increases in ASD prevalence.Coincident with the observed increase in ASD prevalence is the introduction of modern rationally designed psychoactive pharmaceuticals, beginning with selective serotonin re-uptake inhibitors (SSRIs) in 1987 , and serotonin–norepinephrine reuptake inhibitors (SNRIs) in 1994 . These pharmaceuticals result in an increase in the neurotransmitter serotonin, which is responsible for the regulation of neural activity and other physiological functions The use of SSRIs and SNRIs by pregnant women to treat depression and other psychological disorders has been associated with low APGAR  scores -1), human health consequences of UPPs remain controversial. While the highest observed concentrations of UPPs have biological effects in fish There is an alternative source of exposure to antidepressants: Unmetabolized psychoactive pharmaceuticals (UPPs) found in raw sewage, effluent from sewer treatment facilities, rivers downstream of such facilities, and, ultimately, drinking water Here, we describe results of an experiment that explores a potential association between UPPs and idiopathic ASD. We tested whether chronic exposure to a mixture of UPPs induced autism-like gene expression profiles in a model organism using gene-class analysis. We used treatments involving a mixture of UPPs similar to that observed in aquatic systems and examined expression of genes expressed by individuals with various forms of idiopathic ASD. For comparison, we examined sets of genes expressed by individuals with other neurological disorders.Pimephales promelas, to FLX, VNX, and CBZ in a 3-component mixture. FLX, VNX, and CBZ were chosen because they represent modern pharmaceutical classes that are highly prescribed and are among the UPPs with the highest observed environmental concentrations  known to be caused by specific mutations. This is significant, because it indicates that enrichment in our treatments involve only idiopathic forms of ASD.There was no enrichment of other neurological disorders except MS (in one of two sets) and Parkinson’s. This is significant because it indicates that enrichment of the idiopathic ASD set was not simply associated with general neurological processes, pathways or systems generally common to neurological disorders. The MS set has a low NES (<1.40), and a second MS set see  is not eA number of the genes contributing to enrichment of ASD gene sets have been implicated in other recent studies not included in our analysis. For example, Suda et al. The results presented here are consistent with several recent lines of inquiry: First, the hypothesis that hyperserotonemia plays a role in autism, affecting the developing fetus and potentially involving SSRIs Second, recent evidence supports an association of antidepressants, including SSRIs, with autism Third, there is evidence for an unambiguous environmental component involved in the etiology of autism Fourth, there is evidence of demographic changes that may have increased the proportion of genetically susceptible individuals in contemporary populations Given the conserved nature  of the genes involved in the observed expression profiles, and given that the genes on the Fathead array are homologous to highly conserved human genes, it is reasonable to expect induction of humans gene expression profiles similar to the Fathead profiles. This sort of approach has been effectively used for other models of human disorders The concentrations used in this study were higher than observed environmental concentrations in order to account for conservative concentration estimates and the presence of related formulations and active metabolites These results provide a new perspective on the etiology of idiopathic ASD and suggest new directions for research into autism’s environmental “exposome” All fish handling and treatments were performed at the Great Lakes WATER Institute  using appropriate UWM Institutional Animal Care and Use Committee (IACUC) approved protocols .Full details of the fish treatments are described in a previous report Fish mRNA was pooled within a tank for microarray work (for 3 replicates per treatment) but not for qPCR validation (for 15 replicates per treatment). Details of microarray experiments, including validation by qPCR analysis of 9 genes with high rank correlation and all data files, are described in a previous report The previous study Here, two groups of gene sets gene-class analyses were conducted: ND  and ASD (“autism spectrum disorder”). The gene sets in the ND collection, known to be associated with a variety of human neurological disorders, are described in Gene-class analyses used GSEA release 2.06 and MSigDB release 2.5. Weighted enrichment scores were calculated using gene expression lists ranked by signal-to-noise ratio. The genes on the array were ranked by correlation between the MIX and CTL treatments .  containing a list of HUGO symbols with accepted aliases. Gene set components listed as aliases in this file were replaced with the appropriate HUGO symbol. For additional details of the annotation and GSEA implementation using the EcoArray 15k Fathead Minnow arrays, see Thomas et al. Table S1ND gene list. A list of the neurological disorders gene sets, in the GSEA gene matrix (.GMX) file format.(GMX)Click here for additional data file.Table S2ASD gene list. A list of the autism spectrum disorders gene sets, in the GSEA gene matrix (.GMX) file format.(GMX)Click here for additional data file.Table S3Ranked gene list. A Microsoft Excel spreadsheet containing all annotated array gene elements sorted by signal-to-noise ratio for the mixture v. control comparison.(XLS)Click here for additional data file.


There is an increasingly growing trend towards integrating scientific research training into undergraduate medical education. The importance and compulsoriness of this trend has been greatly highlighted at the Boyer Commission on Educating Undergraduates in the Research University. Despite the importance and benefits of undergraduate research, attempts of medical schools to encourage undergraduates to take part in formal research training during undergraduate medical education remain unsatisfactory. This article serves as a ‘reminder call’ highlighting the requisite to integrate scientific research training into undergraduate medical curricula. There is an increasingly growing trend towards integrating scientific research training into undergraduate medical education. The importance and compulsoriness of this trend has been greatly highlighted at the Boyer commission on educating undergraduates in the research university .The Boyer commission report has dramatically reformed how universities should approach undergraduate education. In particular, it was advocated that research-based learning should be the standard for any given undergraduate education. Since then, the integration of teaching and scientific research into undergraduate medical curricula has been greatly emphasized and employed in Western countries .Medical schools have different methods of engaging its students in scholarly undergraduate research activities. These methods include research-driven curricula , researcExposure to scientific research promotes scientific output  encouragLittle is known about how medical students perceive undergraduate research . StudentAt present, the practice of medicine is largely dependent on utilization of the contemporary evidence-based medicine. Evidence-based medicine is defined as ‘the conscientious, explicit and judicious use of current best evidence in making decisions about the care of individual patients’ . As evidAs ‘practice makes perfect’, medical students should be introduced to scientific research as early as possible in undergraduate medical education as well as engaged in as many various curricular/extracurricular undergraduate research activities. This is because in the settings of early introduction to undergraduate research, knowledge of scientific research and familiarity with basic research skills, medical students will be increasingly self-assured to continue further research activities and gradually solidify and build up their scientific research competencies over time.It must be noted that no prospective medical student must be obliged to do research, as this may promote a tendency for ‘bad research’ negatively influencing the academic and research institutes. However, basics of scientific research to largely comprehend research studies, critically appraise findings and wisely deduce conclusions should be offered in undergraduate medical curricula. Research-related ethical matters should be also considered in undergraduate medical curricula.It must be recognized that educators of undergraduate medical curricula hold the chief responsibility in nurturing scientific research competencies in medical students. Such nurturing of scientific research competencies should start early in undergraduate medical education and span throughout the pre-clinical and clinical years of undergraduate medical education. This is because medical students, with the passage of time, tend to forget the taught research competencies if they are not somehow revisited, revised or utilized appropriately. Failure to develop positive attitudes of scientific research will influence negatively on the curiosity of students in considering further research activities or on commencing research-focused careers. Moreover, as proper execution of evidence-based medicine is largely based on solid acquisition of scientific research competencies, failure to develop such competencies will most likely weaken the proper practice of medicine.Teaching scientific research in undergraduate medical curricula  is highly advised and has been coupled with positive attitudes towards research, advancement of research competencies, and smooth facilitation to engagement in future research undertakings . Basic kMany medical students fail to appreciate the important concept of ‘translational research’ and its key players ‘physician–scientists’. Translational research integrates advances in biomedical and clinical sciences, and translates these findings from the ‘bench-to-bedside’ – that is, from laboratory benches and clinical trials to pertinently relevant and beneficial patient care applications . PhysiciTaking into account the importance of translational research and the absolute need to produce physician–scientist generations, there has been a rapidly evolving emphasis towards incorporating a ‘translational research’ instructional component into medical curricula. The aim of such emphasis is to sufficiently introduce prospective medical doctors to the fundamental scientific and ethical standards of translational research, including the means to how such research is properly conducted, critically evaluated, appropriately explained to patients, and effectively/productively applied to patient care.In conclusion, despite the importance and benefits of undergraduate research, attempts of medical schools to encourage undergraduates to take part in formal research training during undergraduate medical education remain unsatisfactory. It is highly recommended to integrate formal research training into undergraduate medical curricula and to provide a diversity of high-quality well-mentored undergraduate research opportunities. Instruction of research knowledge and skills in undergraduate medical curricula is absolutely indispensable to the proper practice of evidence-based medicine and effective carrying out of clinical practice. Furthermore, it is very important to drive students to develop positive attitudes towards undergraduate research and to resolve any barrier that may hinder medical students from productive engagement in undergraduate research activities.


The effectiveness of DDT and other insecticides when properly used as indoor residual spray (IRS) to combat malaria is not in question . HoweverPolicy formulation, negotiating fora, and the development of research priorities via consensus (some possibly burdened with other agendas) seem not to be good platforms to deal with intractable paradoxes. IRS with chemicals seems out of vogue and often relegated in favour of the enticing promises of high-tech or new methods.inter alia: better application, more selective areas of indoor application, mosquito irritability and repellency, better formulations, and new chemicals [IRS as a method has remained almost unchanged since de Meillon pioneered it in South Africa in 1936 . CombiniMaintaining a proven top-down IRS strategy supported by an effective hospital and clinic system requires a minor inconvenience but no other behavioural changes by the inhabitants , ecologi


Without treatment with a bisphosphonate, 50% of patients with metastases to bone will develop a skeletal related event  and in terms of repeated clinic visits for intravenous treatment in patients entering the terminal phase of their illness, a time during which the derivation of any true benefit from continuing bisphosphonate therapy is unproven.As is the case for all clinical trials, placebo-controlled studies of bisphosphonates have specific inclusion and exclusion criteria that could potentially limit generalizability. The patient population selected for a study are typically those who are most likely to gain maximum benefit from treatment. Several important differences exist between patients treated on study and those treated in non-study clinical practice.First, the benefits of bisphosphonates are time-dependent, with maximum benefit gained after 6 months sres or bone progression sres while on a bisphosphonate produces continued patient benefit.Second, patients with bone-only disease are the ones who benefit most from treatment. Clinical trial populations therefore tend to contain mainly patients in whom bone-only disease predominates . In actusres. The definition of sre does not include variables that are important to patients, such as pain or immobility Third, the most frequently used primary endpoint in bisphosphonate trials is an analysis of These differences between the two populations may limit the drawing of meaningful conclusions regarding the margin of benefit for bisphosphonate treatment outside of a trial setting.sres as pathologic bone fracture , spinal cord compression, surgery or radiation to bone, or development of hypercalcemia. To estimate the amount of benefit gained from bisphosphonate use in clinical practice, we compared our earlier non-study patients in terms of demographics and outcomes to the clinical trial populations (treatment arms) of the placebo-controlled trials.We previously reviewed the use of bisphosphonates in three Canadian centres sre (sre sres per patient was 1.2 (range: 0–13 sres). Time to first sre was much shorter in the off-study population . Breast cancer patients had the highest risk  Markers of bone resorption have shown great promise in both of these areas bismark (Bisphosphonate Therapy Directed by Bone Resorption Markers) trial. Breast cancer patients with metastatic bone disease are randomized to receive either a fixed dose every 3–4 weeks or a modified schedule as determined by NTx level. This study will test the hypothesis that patients with normal levels of bone turnover  can receive less frequent treatment .Dosing schedules of zoledronate are currently being evaluated in the Given the inherent differences, as outlined here, between trial populations and patients treated in clinical practice, it is unlikely that we will ever know the true benefit of bisphosphonate treatment. What we can do is optimize our current use of these agents by means of innovative strategies. Bone markers offer the greatest hope of for optimization. Ongoing research should improve our understanding of these markers’ role in guiding treatment.


Where hemispheric lateralization was once considered an exclusively human trait, it is increasingly recognized that hemispheric asymmetries are evident throughout the animal kingdom. Emotion is a prime example of a lateralized function: given its vital role in promoting adaptive behavior and hence survival, a growing body of research in affective neuroscience is working to illuminate the cortical bases of emotion processing. Presuming that human and non-human primates evolved from a shared ancestor, one would anticipate evidence of organizational continuity in the neural substrate supporting emotion processing. This paper thus reviews research examining the patterns of lateralization for the expression and perception of facial emotion in non-human primates, aiming to determine whether the patterns of hemispheric asymmetry that characterize the human brain are similarly evident in other primate species. As such, this review seeks to enhance understanding of the evolution of hemispheric specialization for emotion, using emotion lateralization in non-human primates as a window through which to view emotion lateralization in humans. For much of the past 150 years, hemispheric lateralization has been considered an exclusive characteristic of the human brain. However, where once cerebral asymmetry was thought “the most imposing difference between man and animal,” . Subsequent observations of clinical patients offer further support for the right hemisphere hypothesis, demonstrating that damage to the right hemisphere compromises both the perception . A visual means of emotional expression is widely used by diurnal social mammals, including primates , the data from those investigations indicate a left visual field (right hemisphere) bias for the perception of emotion in both conspecific , however it is important to note an alternate possibility. Hopkins and Cantalupo  point ouBeyond the suggestion of continuity in emotion lateralization for human and non-human primates, this review makes it strikingly apparent that there are significant gaps in the non-human primate literature. Whilst emotional expression has been subject to comparatively greater investigation than emotion perception in non-human primates, even there the studies are restricted to only a few species . Whether the right hemisphere’s dominance for emotion extends to prosimians remains an open question. Though it appears probable that the left side expressivity bias seen in Old World monkeys, New World Monkeys, great apes, and humans will be similarly evident in their more phylogenetically distant primate relatives, only investigation of facial displays in more primitive primates, such as lemurs, galagos, and lorises, will resolve the question.As Ward  notes, eAotus  also offers intriguing possibilities. Being a nocturnal anthropoid, this group of non-human primates has evolved to possess comparatively less differentiated facial musculature than diurnal primates and is reported to have virtually no facial expressions  or cathmeral anthropoid species , one may speculate on the influence of experience on emotion lateralization. Observation of others’ emotion expressions may increase the magnitude of the hemispheric asymmetry for emotion processing in diurnal primates via experience-dependent processes; such influences may be less likely to induce changes in nocturnal and cathmeral primates like Aotus.Investigation of emotion lateralization in the owl monkey family s Huber, . ExaminaThe fact that hemispheric asymmetries are evident across primate species has implications beyond the lateralization of emotion. Where once lateralization was thought to be a defining human attribute (e.g., Pruner-Bey, The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


During the past two decades growing evidence indicates that brain oscillations in the alpha band (~10 Hz) not only reflect an “idle” state of cortical activity, but also take a more active role in the generation of complex cognitive functions. A recent study shows that more than 60% of the observed inter-subject variability in perceptual learning can be ascribed to ongoing alpha activity. This evidence indicates a significant role of alpha oscillations for perceptual learning and hence motivates to explore the potential underlying mechanisms. Hence, it is the purpose of this review to highlight existent evidence that ascribes intrinsic alpha oscillations a role in shaping our ability to learn. In the review, we disentangle the alpha rhythm into different neural signatures that control information processing within individual functional building blocks of perceptual learning. We further highlight computational studies that shed light on potential mechanisms regarding how alpha oscillations may modulate information transfer and connectivity changes relevant for learning. To enable testing of those model based hypotheses, we emphasize the need for multidisciplinary approaches combining assessment of behavior and multi-scale neuronal activity, active modulation of ongoing brain states and computational modeling to reveal the mathematical principles of the complex neuronal interactions. In particular we highlight the relevance of multi-scale modeling frameworks such as the one currently being developed by “The Virtual Brain” project. Perceptual learning, a form of implicit learning and adult brain plasticity, allows us to tune our perception to efficiently select relevant sensory signals. It ultimately determines our success when adapting and interacting with the dynamic and complex environment. A glimpse into our daily life is enough to realize that learning efficacy varies greatly across human beings but also changes over time in a single person. A recent study on perceptual learning has shown that ongoing brain activity, more specifically, electrical oscillations in the alpha frequency band (~8–12 Hz), are able to predict up to 64% of the observed variability in the learning outcome in a perceptual task  context. In other words, alpha oscillations may play different roles during the sequence of processes. They indicate or encode different aspects of information processing in the brain. In the present article we address how alpha oscillations may modulate perceptual learning. Based on recent computational models implementing biophysical plausible mechanisms, we hypothesize that high alpha ongoing activity in learning-related areas could play an “active” role, promoting learning by improving the encoding capabilities and memory formation. In more general terms, we illustrate the iterative process of empirical and modeling work that is necessary to infer knowledge about the interplay of intrinsic brain activity and external stimuli. This interplay constitutes the foundation of perceptual learning and in general of human cognition.Generally, learning occurs in different forms. Some forms explicitly require memorizing information , while others occur implicitly by exposition or practice  . Generally, learning, attention, and memory appear to be strongly related  or electroencephalography (EEG) recordings. The majority of studies have focused on the visual modality  in 1929 Berger, . OpeningAlthough alpha oscillations are most prominent in visual areas, i.e., they exhibit highest amplitudes in electrodes placed over occipital brain areas, they are generally widespread in the cortex but regionally attenuated depending on different stimuli and tasks Buzsaki, , Ch. 9, Initial evidence suggested that alpha oscillations originate solely from thalamo-cortical interactions  over posterior electrodes contralateral to the attended side . To explain the role of alpha in non-directed attention Sadaghiani et al.  proposedAlpha activity correlates with important processes underlying information processing, including perceptual learning. Yet, whether these correlations indicate a causal relation between behavior and the alpha rhythm remains unknown. Intervening brain activity through neurofeeedback and/or non-invasive brain stimulation can shed light on the causal relation between alpha rhythm and perceptual learning. Transcranial magnetic stimulation (TMS) has proven to successfully modulate ongoing alpha oscillations, eventually modulating visual perception  through purely cortical connectivity , provides the necessary tools to perform full-brain simulations , changes in the structural neural connectivity induced by learning are likely to alter ongoing oscillatory activity. At the same time, plastic mechanisms underlying learning can be boosted during specific brain states defined by spatial patterns of ongoing oscillatory activity . In fact, several computational models have successfully linked STDP . Currently, such models are able to reproduce coarse/general aspects of spatiotemporal brain dynamics that are present in the majority of experimental data. In these models, brain activity can be constrained by individual large-scale connectivity parameters derived from dwMRI. This allows scientists to simulate the effects that large-scale plastic changes have on emerging brain dynamics. Our hypothesis of how plastic changes in sensory areas elicited by perceptual learning affect large-scale information processing is based on a recent study of our group, that shows results of full-brain simulations . In this study, Roy et al. (in preparation) propose ways to incorporate plasticity mechanisms in existing computational models that are capable of generating ongoing spontaneous activity as a function of transmission delays, noise and connectivity. In those simulations, plasticity in local populations can change the dynamical stability of global functional networks distributed across multiple brain areas. The main findings are: (1) Local network activity in the absence of plasticity is characterized by irregular oscillations between a low-amplitude asynchronous and a high amplitude synchronous state. (2) Alterations in local synapses (due to STDP), in the order of few milliseconds, induce changes in the local connectivity of the brain areas where plasticity is implemented. Such changes alters distinct features of the global functional connectivity (FC). (3) The interaction between those regions is organized systematically in correlated and anti-correlated networks depending on the choice of the model parameters. These parameters include plasticity parameters as well as the amplitude, frequency of the background oscillatory state. Anti-correlated networks after time dependent plasticity show significantly and highly correlated BOLD spatiotemporal activity. In particular, simulations show that the intrinsic alpha oscillations generated by local cortical neurons efficiently influence the learning outcome of brain areas connected structurally. While this model does not target exclusively brain areas involved in perceptual learning, it proposes a general mechanism able to explain the effect of perceptual learning on resting-state activity, as it is the case in some of the studies presented above. In the near future, based on individual subject/patient parameters, this kind of model simulations will help to predict the impact that interventional actions, which evoke plasticity will have on evolving brain dynamics. Ultimately, similar brain simulations will aid scientists in the planning of experimental learning protocols as well as clinicians developing therapeutic strategies, in order to reveal the complex relation between perceptual learning and large-scale ongoing brain activity.The current view on how alpha oscillations relate to cognitive abilities, such as perceptual learning, is becoming far more complex compared to the initial view which associates alpha activity to an “idle” brain state. Although the role of alpha oscillations in spatial attention, working memory, and perception is well documented, and despite initial evidence indicating that alpha oscillations influence perceptual learning, the detailed role of alpha rhythm in perceptual learning and its contribution to the observed variability in learning outcome needs further empirical and theoretical assessment. Changes in perception due to perceptual learning develop under a variety of conditions such as training, sensory or even central stimulation, with and without attention, showing complex degrees of spatial specificity and temporal persistence. This suggests that a variety of neural processes and brain areas are implicated in perceptual learning (Sasaki et al., The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


The formation of the Amazon Dark Earths was a model of sustainable soil management that involved intensive composting and charcoal (biochar) application. Biochar has been the focus of increasing research attention for carbon sequestration, although the role of compost or humic substances (HS) as they interact with biochar has not been much studied. We provide a perspective that biochar and HS may facilitate extracellular electron transfer (EET) reactions in soil, which occurs under similar conditions that generate the greenhouse gases methane and nitrous oxide. Facilitating EET may constitute a viable strategy to mitigate greenhouse gas emission. In general, we lack knowledge in the mechanisms that link the surface chemical characteristics of biochar to the physiology of microorganisms that are involved in various soil processes including those that influence soil organic matter dynamics and methane and nitrous oxide emissions. Most studies view biochar as a mostly inert microbial substrate that offers little other than a high sorptive surface area. Synergism between biochar and HS resulting in enhanced EET provides a mechanism to link electrochemical properties of these materials to microbial processes in sustainable soils. Sadly, the original populace that created terra preta also became victim to the onslaught of disease after first contact. However, their legacy in the form of the land they managed lives on.Microbes have played prominent roles in the history of ancient American civilizations. To the Spanish conquistadors, they became unwitting allies – smallpox, typhus, diphtheria, measles – these and other diseases contributed to the deaths of perhaps 95% of the indigenous people in the Americas within 130 years after first contact with Europeans . Within Terra preta was first described by modern soil science in the mid-1960s  are extremely complex and heterogeneous high molecular weight organic materials that often contain polyaromatic structures that include oxygen-containing functional groups, including quinone moieties. Humus is commonly viewed as an essential component of rich soil, although stable humus no longer contributes nutrients and is commonly viewed as important mainly to improve the physical structure of soil. However, recent findings suggest that dissolved HS are redox-active and contribute to anaerobic microbial metabolism by mediating extracellular electron transfer (EET), in effect shuttling electrons between metal-respiring bacteria and metal oxides ; through microbially synthesized electron shuttles . Charcoal itself is a source of humic acids that are formed during pyrolysis . In the presence of dissolved HS, these metal oxides in effect become more available for electron transfer reactions due to electron shuttling by HS.Enhanced EET in soil may facilitate electron flow to electron acceptors other than oxygen. For example, a phylogenetically diverse range of bacteria including members of the alpha-, beta-, gamma-, and delta-Proteobacteria utilize HS as electron donor and nitrate as electron acceptor in forest soil, freshwater and marine sediments . SimilarOryza sativa) soils  soils . Enhancia) soils .terra preta have been studied almost entirely in the tropical setting, there is a need to research long-term effects in other settings. Gradual application will most likely be based on cost-benefit analyses. Similar to the original terra preta, the most promising application of this ancient technology will be on marginal or degraded soils, such as those contaminated by organic or inorganic pollutants . These lands are often poorly developed with little or no vegetation. There are numerous studies documenting both beneficial and negative effects of charcoal on plant growth (Rhizobia (With the societal and environmental benefits associated with biochar, there is now increasing interest in widespread application. However, since the long-term effects of creating t growth  and utilt growth . The post growth . In connt growth , increast growth , and enhRhizobia . Aside fRhizobia . Much ofRhizobia  is a proThe author declares that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.