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Models of conformity and anticonformity have typically focused on cultural traits with unordered variants, such as baby names, strategies (cooperate/defect), or the presence/absence of an innovation. There have been fewer studies of conformity to cultural traits with ordered variants, such as level of cooperation (low, medium, high) or proportion of time spent on a task (0% to 100%). In these studies of ordered cultural traits, conformity is defined as a preference for the mean trait value in a population even if no members of the population have variants near this mean; e.g., 50% of the population has variant 0 and 50% has variant 1, producing a mean of 0.5. Here, we introduce models of conformity to ordered traits, which can be either discrete or continuous. In these models, conformists prefer to adopt more popular cultural variants even if these variants are far from the population mean. To measure a variant's "popularity" in cases where no two individuals share precisely the same variant on a continuum, we introduce a metric called k -dispersal; this takes into account a variant's distance to its k closest neighbors, with more "popular" variants having lower distances to their neighbors. We demonstrate through simulations that conformity to ordered traits need not produce a homogeneous population, as has previously been claimed. Under some combinations of parameter values, conformity sustains substantial trait variation over many generations. Furthermore, anticonformity may produce a high level of polarization., Competing Interests: Competing interests statement:The authors declare no competing interest.

Direct-infusion mass spectrometry (DI-MS) and mass spectrometry imaging (MSI) are powerful techniques for lipidomics research. However, annotating isomeric and isobaric lipids with these methods is challenging due to the absence of chromatographic separation. Recently, cyclic ion mobility mass spectrometry (cIM-MS) has been proposed to overcome this limitation. However, fluctuations in room conditions can affect ion mobility multipass arrival times, potentially reducing annotation confidence. In this study, we developed a multipass arrival time correction method that proved effective across various dates, room temperatures, ion mobility settings, and laboratories using mixtures of reference standards. We observed slight variations in the linear correction lines between lipid and nonlipid molecules, underscoring the importance of choosing appropriate reference molecules. Based on these results, we demonstrated that an accurate multipass arrival time database can be constructed from corrected t 0 and t p for interlaboratory use and can effectively identify isomeric lipids in MSI using only a single measurement. This approach significantly simplifies the identification process compared to determining multipass collision cross-section, which requires multiple measurements that are both sample- and time-intensive for MSI. Additionally, we validated our multipass drift time correction method in shotgun lipidomics analyses of human and mouse serum samples and observed no matrix effect for the analysis. Despite variations in dates, room temperatures, instruments, and ion mobility settings, our approach reduced the mean drift time differences from over 2% to below 0.2%., Competing Interests: The authors declare no competing financial interest., (© 2024 The Authors. Published by American Chemical Society.)

The combination of enhanced resolving power and improved duty cycle on a multireflecting time-of-flight mass spectrometer is described. Resolving power increases are achieved by extending the effective ion path length from 47 m to greater than 200 m. Path length increases are achieved through containment of ions within the analyzer for up to N = 5 passes using a pulsed deflection electrode. Resolving power was shown to increase from 220,000 to 402,000 (fwhm) at m / z 785 for N = 1 and N = 4 analyzer passes, respectively. Due to the timing of the pulsed deflection electrode, the approach is particularly suited to high resolution analysis over a targeted m / z range. Duty cycle enhancements are achieved for ions of the targeted m / z range via accumulation prior to orthogonal acceleration, providing signal improvements of 2 orders of magnitude. Achieving such high resolving powers at fast scan rates (30 Hz) can yield additional information such as fine isotope structure; when combined with ppb mass measurement accuracy, high confidence in analyte identification can be achieved. The technique is applied for N = 2 analyzer passes, demonstrating fine isotope structure for a typical UHPLC metabolite identification experiment at a 10 Hz acquisition rate. Additionally, mass spectrometry imaging data is acquired using DESI, demonstrating the improved image clarity achieved at >300,000 (fwhm).

Background: Lenvatinib, a multikinase inhibitor, is an FDA-approved treatment for advanced hepatocellular carcinoma (HCC) in the first-line setting. Recent trial data have established atezolizumab plus bevacizumab as well as tremelimumab plus durvalumab as preferred first-line treatment options for advanced HCC. The role of lenvatinib following progression on immunotherapy in patients with advanced HCC remains unclear., Methods: We conducted a multicentric, retrospective analysis of patients with advanced HCC diagnosed between 2010 and 2021 at the Mayo Clinic in Minnesota, Arizona, and Florida who received immunotherapy followed by lenvatinib. Median overall survival and progression-free survival analyses were performed using the Kaplan-Meier method, and responses were determined using RECIST 1.1. Adverse events were determined using CTCAE v 4.0., Results: We identified 53 patients with advanced HCC who received lenvatinib following progression on immunotherapy. Forty five (85%) patients had a Child Pugh class A at diagnosis, while 30 (58%) patients were still Child Pugh A at time of lenvatinib initiation. Lenvatinib was administered as a second-line treatment in 85% of the patients. The median PFS was 3.7 months (95% CI: 3.2-6.6), and the median OS from the time of lenvatinib initiation was 12.8 months (95% CI: 6.7-19.5). In patients with Child Pugh class A, the median OS and PFS was 14 and 5.2 months, respectively. Race, gender, and Child Pugh class was associated with OS on multivariate analysis., Discussion: Our study, using real-world data, suggests that patients benefit from treatment with lenvatinib following progression on immunotherapy in advanced HCC. The optimal sequencing of therapy for patients with advanced HCC following progression on immunotherapy remains unknown, and these results need to be validated in a clinical trial.

Objectives: To date, there are no published pharmacokinetic (PK) data for baricitinib in critically ill patients requiring continuous renal replacement therapy. This paper describes in detail the plasma PK and dialytic clearance of baricitinib in a patient infected with coronavirus disease 2019 (COVID-19) requiring continuous renal replacement therapy in order to suggest dosing strategies in this population., Methods: Baricitinib 2 mg daily was used for the treatment of COVID-19 in a critically ill patient on continuous venovenous haemodialysis (CVVHD). Prefiltration plasma drug concentrations of baricitinib were measured at hours 1, 2, 12, and 24 after drug administration. Postfiltration and ultrafiltrate concentrations were collected at hour 24., Results: Plasma PK parameters of baricitinib in this patient were as follows: maximum plasma concentration (C max ), 20.98 ng/mL; minimum plasma concentration (C min ), 9.84 ng/mL; half-life (t 1/2 ), 23.85 h; apparent volume of distribution at the steady state (V ss ), 99.42 L; total clearance at the steady state (CL ss ), 2.89 L/h; and area under the concentration-time curve (AUC 0-∞ ), 692.14 ng · h/mL. The saturation coefficient for baricitinib at 24 h after administration was 0.607. The transmembrane clearance of baricitinib by CVVHD running at a flow rate of 2 L/h was 1.21 L/h, representing 41.9% of the total clearance of baricitinib., Conclusions: In a critically ill COVID-19 patient on CVVHD, a 2-mg dose of baricitinib achieves a C max comparable with healthy subjects, but total clearance was reduced to about 20%. Larger studies exploring multiple patients and dialysis modes are needed to determine the optimal dosing strategy for baricitinib in this patient population., (Copyright © 2023 Elsevier Ltd and International Society of Antimicrobial Chemotherapy. All rights reserved.)

Background: Hypoperfusion leads to allograft injury during kidney transplantation. Catecholamine vasopressors are used to maintain blood pressure in the perioperative period but have demonstrated negative outcomes in the deceased-donor kidney transplant population. Little is known regarding living donor kidney transplants (LDKTs) and vasopressor use. The aim of this study is to describe the incidence of vasopressor use in LDKT and characterize its effects on allograft function and patient outcomes., Methods: This retrospective, observational cohort study included adult patients who underwent an isolated LDKT between August 1, 2017, and September 1, 2018. Patients were divided into those who received perioperative vasopressors and those who did not. The primary objective was to compare allograft function between LDKT recipients that received vasopressors and those who did not. Secondary outcomes included safety endpoints and the identification of clinical variables associated with vasopressor use., Results: A total of 67 patients received an LDKT during the study period. Of those, 25 (37%) received perioperative vasopressors, and 42 (62%) did not. Poor graft function, as defined by the development of slow or delayed graft function, occurred more frequently in patients receiving perioperative vasopressors compared with those who did not (6 [24%] vs 1 [2.4%], P = .016). In multivariable regression modeling, only perioperative vasopressors were statistically significantly associated with poor graft function. In addition, patients exposed to vasopressors experienced more postoperative arrhythmias (8 [32%] vs 1 [4.8%], P = .0025)., Conclusion: Using perioperative vasopressors was independently associated with worsened early renal allograft function, including delayed graft function and adverse events in the LDKT population., Competing Interests: Declaration of Competing Interest None., (Copyright © 2023 Elsevier Inc. All rights reserved.)

A novel mass spectrometry system is described here comprising a quadrupole-multireflecting time-of-flight design. The new multireflecting time-of-flight analyzer has an effective path length of 48 m and employs planar, gridless ion mirrors providing fourth-order energy focusing resulting in resolving power over 200 000 fwhm and sub-ppm mass accuracy. We show how these attributes are maintained with relatively fast acquisition speeds, setting the system apart from other high resolution mass spectrometers. We have integrated this new system into both liquid chromatography-mass spectrometry and mass spectrometry imaging workflows to demonstrate how the instrument characteristics are of benefit to these applications.

(1) Background: Cabozantinib, a multikinase inhibitor, is approved by the Food and Drug Administration (FDA) for the treatment of advanced hepatocellular carcinoma (HCC) following progression on sorafenib. Recently, atezolizumab plus bevacizumab has been approved in the first line setting for advanced HCC and has become the new standard of care. Whether cabozantinib improves outcomes following progression on immunotherapy remains unknown. We describe the clinical outcomes following treatment with immunotherapy in patients with advanced HCC who received cabozantinib. (2) Methods: We conducted a multicentric, retrospective analysis of patients with advanced HCC diagnosed between 2010-2021 at Mayo Clinic in Minnesota, Arizona, and Florida who received cabozantinib. Median overall survival and progression free survival analyses were performed using the Kaplan-Meier method. Adverse events were determined using Common Terminology Criteria for Adverse Events (CTCAE). (3). Results: We identified 26 patients with advanced HCC who received cabozantinib following progression on immunotherapy. Median progression free survival on cabozantinib therapy was 2.1 months (95% CI: 1.3-3.9) and median overall survival from time of cabozantinib initiation was 7.7 months (95% CI: 5.3-14.9). (4) Conclusion: The optimal sequencing of therapy for patients with advanced HCC following progression on immunotherapy remains unknown. Our study demonstrates that patients may benefit from treatment with cabozantinib following progression on immunotherapy.

A deep understanding of an antimicrobial's critical pharmacokinetic and pharmacodynamic properties is crucial towards optimizing its use in patients and bolstering the drug development program. With the growing threat of antimicrobial resistance and decline in antimicrobial development, the advancement of complex and rigorous pharmacokinetic and pharmacodynamic studies over a short time span has renewed confidence in the value of pharmacokinetic and pharmacodynamic studies and allowed it to become fundamental component of a robust drug development program with high chances of successful approval. In addition, recent guidance by various regulatory bodies have reinforced that a strong and dedicated focus on pharmacokinetics and pharmacodynamics throughout research and development lead to the use of an optimized dosing regimen in Phase 3 trials, improving the probability of drug approval. The objective of this review is to demonstrate the importance of pharmacokinetic and pharmacodynamic studies in the drug development decision-making process by highlighting the developments in pharmacokinetic and pharmacodynamic methods and discuss the role of pharmacokinetic and pharmacodynamic studies in antimicrobial successes and failures., Competing Interests: EW has received honoraria from Melinta Therapeutics, Abbvie Inc., and Shionogi Inc. The remaining 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., (Copyright © 2022 Palmer, Andrews, Abbey, Dahlquist and Wenzler.)

The characterization of enantiomers is an important analytical challenge in the chemical and life sciences. Thorough evaluation of the purity of chiral molecules is particularly required in the pharmaceutical industry where safety concerns are paramount. Assessment of the enantiomeric composition is still challenging and time-consuming, meaning that alternative approaches are required. In this study, we exploit the formation of dimers as diastereomeric pairs of enantiomers to affect separation by high resolution cyclic ion mobility-mass spectrometry. Using the example of ( R / S )-thalidomide, we show that even though this is not an enantiomer separation, we can determine which enantiomer is in excess and obtain quantitative information on the enantiomer composition without the need for a chiral modifier. Further examples of the approach are presented, including d/l-tryptophan and ( R / S )-propanolol, and demonstrate the need for mobility resolving power in excess of 400 (CCS/ΔCCS).

1-β- O -Acyl-glucuronides (AGs) are common metabolites of carboxylic acid-containing xenobiotics, including, e.g ., many nonsteroidal anti-inflammatory drugs (NSAIDs). They are of concern to regulatory authorities because of the association of these metabolites with the hepatotoxicity that has resulted in drug withdrawal. One factor in assessing the potential risk posed by AGs is the rate of transacylation of the biosynthetic 1-β- O -acyl form to the 2-, 3-, and 4- O -acyl isomers. While transacylation can be measured using 1 H NMR spectroscopy or liquid chromatography-mass spectrometry (LC-MS), the process can be time consuming and involve significant method development. The separation of these positional isomers by ion mobility spectrometry (IMS) has the potential to allow their rapid analysis, but conventional instruments lacked the resolving power to do this. Prediction of the collision cross section (CCS) using a machine learning model suggested that greater IMS resolution might be of use in this area. Cyclic IMS was evaluated for separating mixtures of isomeric AGs of diclofenac and was compared with a conventional ultraperformance liquid chromatography (UPLC)-MS method as a means for studying transacylation kinetics. The resolution of isomeric AGs was not seen using a conventional traveling wave IMS device; however, separation was seen after several passes around a cyclic IMS. The cyclic IMS enabled the degradation of the 1-β- O -acyl-isomer to be analyzed much more rapidly than by LC-MS. The ability of cyclic IMS to monitor the rate of AG transacylation at different pH values, without the need for a prior chromatographic separation, should allow high-throughput, real-time, monitoring of these types of reactions.

Rationale: Mass spectrometry (MS) is often employed in the characterisation of synthetic polymers. As polymer architecture becomes more complex, ion mobility (IM) is increasingly being coupled with MS to provide an additional dimension of separation, along with structural information. In this study, we explore the use of a novel cyclic ion mobility (cIM) mass spectrometer for the analysis of a co-polymer sample., Methods: A solution of poly(ethylene glycol)-poly(propylene glycol) random co-polymer (PEG-ran-PPG) was used as a representative polymer sample. The solution was infused into a cIM-enabled quadrupole time-of-flight mass spectrometer. An m/z region of interest, selected using the quadrupole, was passed around the cIM device multiple times. Subsequently, regions of an arrival time distribution were 'sliced' and subjected to tandem mass spectrometric (MS/MS) analysis., Results: Typical, multiply charged series were observed for the polymer under electrospray ionisation. Multiple passes of the cIM device resulted in the separation of otherwise-overlapping charge states within a narrow m/z window (~3 m/z units), allowing individual selection of ions. These isolated ions were then subjected to post-mobility fragmentation resulting in clean, high-resolution product ion spectra, with a significant reduction in interference., Conclusions: Scalable IM separation (IMS), brought about by passing ions multiple times around the cIM device, was demonstrated to provide increased IM resolution for ions in the selected m/z window. After multiple passes, deconvoluted high-resolution MS/MS product ion spectra were successfully acquired for ions that previously had interfering overlapping species present., (© 2019 John Wiley & Sons, Ltd.)

The advent of endothermy, which is achieved through the continuous homeostatic regulation of body temperature and metabolism 1,2 , is a defining feature of mammalian and avian evolution. However, when challenged by food deprivation or harsh environmental conditions, many mammalian species initiate adaptive energy-conserving survival strategies-including torpor and hibernation-during which their body temperature decreases far below its homeostatic set-point 3-5 . How homeothermic mammals initiate and regulate these hypothermic states remains largely unknown. Here we show that entry into mouse torpor, a fasting-induced state with a greatly decreased metabolic rate and a body temperature as low as 20 °C 6 , is regulated by neurons in the medial and lateral preoptic area of the hypothalamus. We show that restimulation of neurons that were activated during a previous bout of torpor is sufficient to initiate the key features of torpor, even in mice that are not calorically restricted. Among these neurons we identify a population of glutamatergic Adcyap1-positive cells, the activity of which accurately determines when mice naturally initiate and exit torpor, and the inhibition of which disrupts the natural process of torpor entry, maintenance and arousal. Taken together, our results reveal a specific neuronal population in the mouse hypothalamus that serves as a core regulator of torpor. This work forms a basis for the future exploration of mechanisms and circuitry that regulate extreme hypothermic and hypometabolic states, and enables genetic access to monitor, initiate, manipulate and study these ancient adaptations of homeotherm biology.

Mid-winter limnological surveys of Lake Erie captured extremes in ice extent ranging from expansive ice cover in 2010 and 2011 to nearly ice-free waters in 2012. Consistent with a warming climate, ice cover on the Great Lakes is in decline, thus the ice-free condition encountered may foreshadow the lakes future winter state. Here, we show that pronounced changes in annual ice cover are accompanied by equally important shifts in phytoplankton and bacterial community structure. Expansive ice cover supported phytoplankton blooms of filamentous diatoms. By comparison, ice free conditions promoted the growth of smaller sized cells that attained lower total biomass. We propose that isothermal mixing and elevated turbidity in the absence of ice cover resulted in light limitation of the phytoplankton during winter. Additional insights into microbial community dynamics were gleaned from short 16S rRNA tag (Itag) Illumina sequencing. UniFrac analysis of Itag sequences showed clear separation of microbial communities related to presence or absence of ice cover. Whereas the ecological implications of the changing bacterial community are unclear at this time, it is likely that the observed shift from a phytoplankton community dominated by filamentous diatoms to smaller cells will have far reaching ecosystem effects including food web disruptions., (© 2015 Society for Applied Microbiology and John Wiley & Sons Ltd.)

The Bay of Quinte, a Z-shaped embayment at the northeastern end of Lake Ontario, has a long history of eutrophication problems primarily manifested as spatially extensive algal blooms and predominance of toxic cyanobacteria. The purpose of this study was to identify the structural changes of the phytoplankton community induced by two environmental alterations: point-source phosphorus (P) loading reduction in the late 1970s and establishment of dreissenid mussels in the mid-1990s. A combination of statistical techniques was used to draw inference about compositional shifts of the phytoplankton assemblage, the consistency of the seasonal succession patterns along with the mechanisms underlying the algal biovolume variability in the Bay of Quinte over the past three decades. Based on a number of diversity and similarity indices, the algal assemblages in the upper and middle segments of the Bay are distinctly different from those typically residing in the outer segments. Our analysis also identified significant differences among the phytoplankton communities, representing the pre- and post-P control as well as the pre- and post-dreissenid invasion periods. Recent shifts in phytoplankton community composition were mainly associated with increased frequency of occurrence of toxin-producing Microcystis outbreaks and reduced biovolume of N 2 fixers, such as Aphanizomenon and Anabaena. Bayesian hierarchical models were developed to elucidate the importance of different abiotic factors (light attenuation, water temperature, phosphorus, and ammonium) on total cyanobacteria, Microcystis, Aphanizomenon, and Anabaena relative biovolume. Our modelling exercise suggests that there is significant spatial heterogeneity with respect to the role of the factors examined, and thus total phosphorus alone cannot always explain the year-to-year variability of cyanobacteria succession patterns in the system. The lessons learned from the present analysis will be helpful to the water quality criteria setting process and could influence the management decisions in order to delist the system as an Area of Concern., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Although single genes underlying several evolutionary adaptations have been identified, the genetic basis of complex, polygenic adaptations has been far more challenging to pinpoint. Here we report that the budding yeast Saccharomyces paradoxus has recently evolved resistance to citrinin, a naturally occurring mycotoxin. Applying a genome-wide test for selection on cis-regulation, we identified five genes involved in the citrinin response that are constitutively up-regulated in S. paradoxus. Four of these genes are necessary for resistance, and are also sufficient to increase the resistance of a sensitive strain when over-expressed. Moreover, cis-regulatory divergence in the promoters of these genes contributes to resistance, while exacting a cost in the absence of citrinin. Our results demonstrate how the subtle effects of individual regulatory elements can be combined, via natural selection, into a complex adaptation. Our approach can be applied to dissect the genetic basis of polygenic adaptations in a wide range of species.

Calcium (Ca) concentrations are decreasing in softwater lakes across eastern North America and western Europe. Using long-term contemporary and palaeo-environmental field data, we show that this is precipitating a dramatic change in Canadian lakes: the replacement of previously dominant pelagic herbivores (Ca-rich Daphnia species) by Holopedium glacialis, a jelly-clad, Ca-poor competitor. In some lakes, this transformation is being facilitated by increases in macro-invertebrate predation, both from native (Chaoborus spp.) and introduced (Bythotrephes longimanus) zooplanktivores, to which Holopedium, with its jelly coat, is relatively invulnerable. Greater representation by Holopedium within cladoceran zooplankton communities will reduce nutrient transfer through food webs, given their lower phosphorus content relative to daphniids, and greater absolute abundances may pose long-term problems to water users. The dominance of jelly-clad zooplankton will likely persist while lakewater Ca levels remain low., (© 2014 The Author(s) Published by the Royal Society. All rights reserved.)

It has long been debated whether natural selection acts primarily upon individual organisms, or whether it also commonly acts upon higher-level entities such as lineages. Two arguments against the effectiveness of long-term selection on lineages have been (i) that long-term evolutionary outcomes will not be sufficiently predictable to support a meaningful long-term fitness and (ii) that short-term selection on organisms will almost always overpower long-term selection. Here, we use a computational model of protein folding and binding called 'lattice proteins'. We quantify the long-term evolutionary success of lineages with two metrics called the k-fitness and k-survivability. We show that long-term outcomes are surprisingly predictable in this model: only a small fraction of the possible outcomes are ever realized in multiple replicates. Furthermore, the long-term fitness of a lineage depends only partly on its short-term fitness; other factors are also important, including the 'evolvability' of a lineage-its capacity to produce adaptive variation. In a system with a distinct short-term and long-term fitness, evolution need not be 'short-sighted': lineages may be selected for their long-term properties, sometimes in opposition to short-term selection. Similar evolutionary basins of attraction have been observed in vivo, suggesting that natural biological lineages will also have a predictive long-term fitness.

Unlabelled: Simple sequence repeat (SSR) tracts produce stochastic on-off switching, or phase variation, in the expression of a panoply of surface molecules in many bacterial commensals and pathogens. A change to the number of repeats in a tract may alter the phase of the translational reading frame, which toggles the on-off state of the switch. Here, we construct an in silico SSR locus with mutational dynamics calibrated to those of the Haemophilus influenzae mod locus. We simulate its evolution in a regimen of two alternating environments, simultaneously varying the selection coefficient, s, and the epoch length, T. Some recent work in a simpler (two-locus) model suggested that stochastic switching in a regimen of two alternating environments may be evolutionarily favored only if the selection coefficients in the two environments are nearly equal ("symmetric") or selection is very strong. This finding was puzzling, as it greatly restricted the conditions under which stochastic switching might evolve. Instead, we find agreement with other recent theoretical work, observing selective utility for stochastic switching if the product sT is large enough for the favored state to nearly fix in both environments. Symmetry is required neither in s nor in sT. Because we simulate finite populations and use a detailed model of the SSR locus, we are also able to examine the impact of population size and of several SSR locus parameters. Our results indicate that conditions favoring evolution and maintenance of SSR loci in bacteria are quite broad., Importance: Bacteria experience frequent changes of environment during the infection cycle. One means to rapidly adapt is stochastic switching: a bacterial lineage will stochastically produce a variety of genotypes, so that some descendants will survive if the environment changes. Stochastic switching mediated by simple sequence repeat (SSR) loci is widespread among bacterial commensals and pathogens and influences critical interactions with host surfaces or immune effectors, thereby affecting host persistence, transmission, and virulence. Here, we use the most detailed in silico model of an SSR locus to date, with its phase variation calibrated to match the mod locus of Haemophilus influenzae. The type III restriction-modification system encoded by mod participates in the regulation of multiple other genes; thus, SSR-mediated phase variation of mod has far-reaching cis-regulatory effects. This coupling of phase-variable switching to complex phenotypic effects has been described as the "phasevarion" and is central to understanding the infection cycle of bacterial commensals and pathogens.

Temperature and ultraviolet B radiation (UVB 290-320 nm) are inextricably linked to global climate change. These two variables may act separately, additively, or synergistically on specific aspects of fish biochemistry. We raised Atlantic Salmon (Salmo salar) parr for 54 days in outdoor tanks held at 12 and 19 °C and, at each temperature, we exposed them to three spectral treatments differing in UV radiation intensity. We quantified individual fatty acid (FA) mass fractions in four tissues (dorsal muscle, dorsal and ventral skin, and ocular tissue) at each temperature × UV combination. FA composition of dorsal muscle and dorsal and ventral skin was not affected by UV exposure. Mass fractions of 16:0, 18:0, and saturated fatty acids (SFA) were greater in dorsal muscle of warm-reared fish whereas 18:3n-3, 20:2, 20:4n-6, 22:5n-3, 22:6n-3, n-3, n-6, polyunsaturated fatty acids (PUFA), and total FA were significantly higher in cold-reared fish. Mass fractions of most of the FA were greater in the dorsal and ventral skin of warm-reared fish. Cold-reared salmon exposed to enhanced UVB had higher ocular tissue mass fractions of 20:2, 20:4n-6, 22:6n-3, n-3, n-6, and PUFA compared to fish in which UV had been removed. These observations forecast a host of ensuing physiological and ecological responses of juvenile Atlantic Salmon to increasing temperatures and UVB levels in native streams and rivers where they mature before smolting and returning to the sea.

Mutability as mechanism for rapid adaptation to environmental challenge is an alluringly simple concept whose apotheosis is realized in simple sequence repeats (SSR). Bacterial genomes of several species contain SSRs with a proven role in adaptation to environmental fluctuations. SSRs are hypermutable and generate reversible mutations in localized regions of bacterial genomes, leading to phase variable ON/OFF switches in gene expression. The application of genetic, bioinformatic, and mathematical/computational modeling approaches are revolutionizing our current understanding of how genomic molecular forces and environmental factors influence SSR-mediated adaptation and led to evolution of this mechanism of localized hypermutation in bacterial genomes., (© 2012 New York Academy of Sciences.)

An ecosystem perspective to restoring beneficial uses in Areas of Concern can be interpreted as a shift from the traditional elucidation of simple cause-effect relationships to a multicausal way of thinking that more effectively accommodates ecosystem complexity. This holistic management paradigm has also pervaded the contemporary ecological modeling practice, making compelling the adoption of more sophisticated ecosystem modeling tools. In this study, our primary objective is to develop a Bayesian hierarchical network of simple ecological models for Lake Simcoe, Ontario, Canada, aiming to establish a realistic representation of the causal connections among exogenous nutrient loading, ambient nutrient conditions, and epilimnetic plankton dynamics. In particular, we used a spatially explicit simple mass-balance model forced with idealized sinusoidal loading to predict total phosphorus concentrations. A structural equation model was then used to delineate the interplay among nutrients, ambient light conditions, phytoplankton, and herbivorous biomass. Our analysis highlights the strength of the causal linkages between total phosphorus and water clarity with phytoplankton as well as the capacity of zooplankton grazing to modulate the algal standing crop. Our Bayesian network is also used to examine the exceedance frequency of threshold values for total phosphorus (15 μg/L) and chlorophyll a (4 μg/L) concentrations under scenarios of phosphorus loading reduction. Our study suggests that a 15% phosphorus loading decrease will still result in >25% violations of the 4 μg chla/L value in the two embayments of Lake Simcoe (Cook's Bay and Kempenfelt Bay). The TP levels will decrease in response to the exogenous loading reductions and this improvement will be primarily manifested in the northcentral segments of the system.

For a lineage to survive over long time periods, it must sometimes change. This has given rise to the term evolvability, meaning the tendency to produce adaptive variation. One lineage may be superior to another in terms of its current standing variation, or it may tend to produce more adaptive variation. However, evolutionary outcomes depend on more than standing variation and produced adaptive variation: deleterious variation also matters. Evolvability, as most commonly interpreted, is not predictive of evolutionary outcomes. Here, we define a predictive measure of the evolutionary success of a lineage that we call the k-survivability, defined as the probability that the lineage avoids extinction for k generations. We estimate the k-survivability using multiple experimental replicates. Because we measure evolutionary outcomes, the initial standing variation, the full spectrum of generated variation, and the heritability of that variation are all incorporated. Survivability also accounts for the decreased joint likelihood of extinction of sub-lineages when they 1) disperse in space, or 2) diversify in lifestyle. We illustrate measurement of survivability with in silico models, and suggest that it may also be measured in vivo using multiple longitudinal replicates. The k-survivability is a metric that enables the quantitative study of, for example, the evolution of 1) mutation rates, 2) dispersal mechanisms, 3) the genotype-phenotype map, and 4) sexual reproduction, in temporally and spatially fluctuating environments. Although these disparate phenomena evolve by well-understood microevolutionary rules, they are also subject to the macroevolutionary constraint of long-term survivability.

Unlabelled: The stress-responsive alternative sigma factor σ(B) is conserved across diverse Gram-positive bacterial genera. In Listeria monocytogenes, σ(B) regulates transcription of >150 genes, including genes contributing to virulence and to bacterial survival under host-associated stress conditions, such as those encountered in the human gastrointestinal lumen. An inhibitor of L. monocytogenes σ(B) activity was identified by screening ~57,000 natural and synthesized small molecules using a high-throughput cell-based assay. The compound fluoro-phenyl-styrene-sulfonamide (FPSS) (IC(50) = 3.5 µM) downregulated the majority of genes previously identified as members of the σ(B) regulon in L. monocytogenes 10403S, thus generating a transcriptional profile comparable to that of a 10403S ΔsigB strain. Specifically, of the 208 genes downregulated by FPSS, 75% had been identified previously as positively regulated by σ(B). Downregulated genes included key virulence and stress response genes, such as inlA, inlB, bsh, hfq, opuC, and bilE. From a functional perspective, FPSS also inhibited L. monocytogenes invasion of human intestinal epithelial cells and bile salt hydrolase activity. The ability of FPSS to inhibit σ(B) activity in both L. monocytogenes and Bacillus subtilis indicates its utility as a specific inhibitor of σ(B) across multiple Gram-positive genera., Importance: The σ(B) transcription factor regulates expression of genes responsible for bacterial survival under changing environmental conditions and for virulence; therefore, this alternative sigma factor is important for transmission of L. monocytogenes and other Gram-positive bacteria. Regulation of σ(B) activity is complex and tightly controlled, reflecting the key role of this factor in bacterial metabolism. We present multiple lines of evidence indicating that fluoro-phenyl-styrene-sulfonamide (FPSS) specifically inhibits activity of σ(B) across Gram-positive bacterial genera, i.e., in both Listeria monocytogenes and Bacillus subtilis. Therefore, FPSS is an important new tool that will enable novel approaches for exploring complex regulatory networks in L. monocytogenes and other Gram-positive pathogens and for investigating small-molecule applications for controlling pathogen transmission.

Previous studies have shown that temporally fluctuating environments can create indirect selection for modifiers of evolvability. Here, we use a simple computational model to investigate whether spatially varying environments (multiple demes with limited migration among them, and a different, static selective optimum in each) can also create indirect selection for increased evolvability. The answer is surprisingly complicated. Spatial variation in the environment can sharply reduce the survival rate of migrants, because migrants may be maladapted to their new deme, relative to incumbents. The incumbent advantage can be removed by occasional extinctions in single demes. After all incumbents in a particular deme die, incoming migrants from other demes will, on average, be similarly maladapted to the new environment. This sets off a race to adapt rapidly. Over many extinction events, and the subsequent invasions by maladapted immigrants into a new environment, indirect selection for the ability to adapt rapidly, also known as high evolvability, may result., (© 2011 The Author(s). Evolution© 2011 The Society for the Study of Evolution.)

A set of seven Listeria monocytogenes 10403S mutant strains, each bearing an in-frame null mutation in a gene encoding a key regulatory protein, was used to characterize transcriptional networks in L. monocytogenes; the seven regulatory proteins addressed include all four L. monocytogenes alternative sigma factors (σ(B), σ(C), σ(H), and σ(L)), the virulence gene regulator PrfA, and the heat shock-related negative regulators CtsR and HrcA. Whole-genome microarray analyses, used to identify regulons for each of these 7 transcriptional regulators, showed considerable overlap among regulons. Among 188 genes controlled by more than one regulator, 176 were coregulated by σ(B), including 92 genes regulated by both σ(B) and σ(H) (with 18 of these genes coregulated by σ(B), σ(H), and at least one additional regulator) and 31 genes regulated by both σ(B) and σ(L) (with 10 of these genes coregulated by σ(B), σ(L), and at least one additional regulator). Comparative phenotypic characterization measuring acid resistance, heat resistance, intracellular growth in J774 cells, invasion into Caco-2 epithelial cells, and virulence in the guinea pig model indicated contributions of (i) σ(B) to acid resistance, (ii) CtsR to heat resistance, and (iii) PrfA, σ(B), and CtsR to virulence-associated characteristics. Loss of the remaining transcriptional regulators (i.e., sigH, sigL, or sigC) resulted in limited phenotypic consequences associated with stress survival and virulence. Identification of overlaps among the regulons provides strong evidence supporting the existence of complex regulatory networks that appear to provide the cell with regulatory redundancies, along with the ability to fine-tune gene expression in response to rapidly changing environmental conditions.

The ability of the foodborne pathogen Listeria monocytogenes to survive antimicrobial treatments is a public health concern; therefore, this study was designed to investigate genetic mechanisms contributing to antimicrobial response in L. monocytogenes. In previous studies, the putative bacteriocin immunity gene lmo2570 was predicted to be regulated by the stress responsive alternative sigma factor, sigma(B). As the alternative sigma factor sigma(L) controls expression of genes important for resistance to some antimicrobial peptides, we hypothesized roles for lmo2570, sigma(B), and sigma(L) in L. monocytogenes antimicrobial response. Results from phenotypic characterization of a L. monocytogenes lmo2570 null mutant suggested that this gene does not contribute to resistance to nisin or to SdpC, an antimicrobial peptide produced by some strains of Bacillus subtilis. While lmo2570 transcript levels were confirmed to be sigma(B) dependent, they were sigma(L) independent and were not affected by the presence of nisin under the conditions used in this study. In spot-on-lawn assays with the SdpC-producing B. subtilis EG351, the L. monocytogenes DeltasigB, DeltasigL, and DeltasigB/DeltasigL strains all showed increased sensitivity to SdpC, indicating that both sigma(B) and sigma(L) regulate genes contributing to SdpC resistance. Nisin survival assays showed that sigma(B) and sigma(L) both affect L. monocytogenes sensitivity to nisin in broth survival assays; that is, a sigB null mutant is more resistant than the parent strain to nisin, while a sigB null mutation in DeltasigL background leads to reduced nisin resistance. In summary, while the sigma(B)-dependent lmo2570 does not contribute to resistance of L. monocytogenes to nisin or SdpC, both sigma(B) and sigma(L) contribute to the L. monocytogenes antimicrobial response.

After an ancestral population splits into two allopatric populations, different mutations may fix in each. When pairs of mutations are brought together in a hybrid offspring, epistasis may cause reduced fitness. Such pairs are known as Bateson-Dobzhansky-Muller (BDM) incompatibilities. A well-known model of BDM incompatibility due to Orr suggests that the fitness load on hybrids should initially accelerate, and continue to increase as the number of potentially incompatible substitutions increases (the "snowball effect"). In the gene networks model, which violates a key assumption of Orr's model (independence of fixation probabilities), the snowball effect often does not occur. Instead, we describe three possible dynamics in a constant environment: (1) Stabilizing selection can constrain two allopatric populations to remain near-perfectly compatible. (2) Despite constancy of environment, punctuated evolution may obtain; populations may experience rare adaptations asynchronously, permitting incompatibility. (3) Despite stabilizing selection, developmental system drift may permit genetic change, allowing two populations to drift in and out of compatibility. We reinterpret Orr's model in terms of genetic distance. We extend Orr's model to the finite loci case, which can limit incompatibility. Finally, we suggest that neutral evolution of gene regulation in nature, to the point of speciation, is a distinct possibility.

Calcium concentrations are now commonly declining in softwater boreal lakes. Although the mechanisms leading to these declines are generally well known, the consequences for the aquatic biota have not yet been reported. By examining crustacean zooplankton remains preserved in lake sediment cores, we document near extirpations of calcium-rich Daphnia species, which are keystone herbivores in pelagic food webs, concurrent with declining lake-water calcium. A large proportion (62%, 47 to 81% by region) of the Canadian Shield lakes we examined has a calcium concentration approaching or below the threshold at which laboratory Daphnia populations suffer reduced survival and fecundity. The ecological impacts of environmental calcium loss are likely to be both widespread and pronounced.

Listeria monocytogenes HrcA and CtsR negatively regulate class I and III stress response genes, respectively, while sigma(B) positively regulates the transcription of class II stress response genes. To define the HrcA regulon and identify interactions between HrcA, CtsR, and sigma(B), we characterized newly generated L. monocytogenes DeltahrcA, DeltactsR DeltahrcA, and DeltahrcA DeltasigB strains, along with previously described DeltasigB, DeltactsR, and DeltactsR DeltasigB strains, using phenotypic assays (i.e., heat resistance, acid resistance, and invasion of human intestinal epithelial cells) and performed whole-genome transcriptome analysis of the DeltahrcA strain. The hrcA and sigB deletions had significant effects on heat resistance. While the hrcA deletion had no significant effect on acid resistance or invasion efficiency in Caco-2 cells, a linear regression model revealed a significant (P = 0.0493) effect of interactions between the hrcA deletion and the ctsR deletion on invasiveness. Microarray-based transcriptome analyses and promoter searches identified (i) 25 HrcA-repressed genes, including two operons (the groESL and dnaK operons, both confirmed as HrcA regulated by quantitative real-time PCR) and one gene directly repressed by HrcA, and (ii) 36 genes that showed lower transcript levels in the DeltahrcA strain and thus appear to be indirectly upregulated by HrcA. A number of genes were found to be coregulated by either HrcA and CtsR (2 genes), HrcA and sigma(B) (31 genes), or all three regulators (5 genes, e.g., gadCB). Combined with previous evidence that sigma(B) appears to directly regulate hrcA transcription, our data suggest that HrcA and sigma(B), as well as CtsR, form a regulatory network that contributes to the transcription of a number of L. monocytogenes genes.

The question of how natural selection affects asexual mutation rates has been considered since the 1930s, yet our understanding continues to deepen. The distribution of mutation rates observed in natural bacteria remains unexplained. It is well known that environmental constancy can favor minimal mutation rates. In contrast, environmental fluctuation (e.g., at period T) can create indirect selective pressure for stronger mutators: genes modifying mutation rate may "hitchhike" to greater frequency along with environmentally favored mutations they produce. This article extends a well-known model of Leigh to consider fitness genes with multiple mutable sites (call the number of such sites alpha). The phenotypic effect of such a gene is enabled if all sites are in a certain state and disabled otherwise. The effects of multiple deleterious loci are also included (call the number of such loci gamma). The analysis calculates the indirect selective effects experienced by a gene inducing various mutation rates for given values of alpha, gamma, and T. Finite-population simulations validate these results and let us examine the interaction of drift with hitchhiking selection. We close by commenting on the importance of other factors, such as spatiotemporal variation, and on the origin of variation in mutation rates.

Cuttlefish have a large repertoire of body patterns that are used for camouflage and interspecific signaling. Intraspecific signaling by male cuttlefish has been well documented but studies on signaling by females are lacking. We found that females displayed a newly described body pattern termed Splotch toward their mirror image and female conspecifics, but not to males, prey or inanimate objects. Female cuttlefish may use the Splotch body pattern as an intraspecific signal, possibly to reduce agonistic interactions. The ability of females to produce a consistent body pattern in response to conspecifics and mirrors suggests that they can recognize same-sex conspecifics using visual cues, despite the lack of sexual dimorphism visible to human observers.

We present a novel enhancement to matrix-assisted laser desorption ionization (MALDI) post-source decay (PSD) analysis whereby fragment ions from multiple precursor ions are acquired into the same spectrum without employing a timed ion gate to preselect each parent ion. Fragment ions are matched to their corresponding precursor ions by comparing spectra acquired at slightly different reflectron electric fields. By measuring the difference in time-of-flight (TOF) between the two spectra for each fragment, it is possible to calculate the mass of the fragment ion and its parent. This new "parallel PSD" technique reduces analysis time and consumes less sample than conventional PSD, which requires an ion gate for serial preselection of precursor ions.

The use of capillary zone electrophoresis (CZE) and capillary zone electrophoresis/mass spectrometry (CZE/MS) has been demonstrated, in principle, for the separation of nicotine and nicotine metabolites. The buffer system developed for separation and detection by CZE/UV was modified for use in CZE/MS analysis. Several of the metabolites are isobaric and tandem mass spectrometric (MS/MS) techniques have been used to differentiate such analytes., (Copyright 2001 John Wiley & Sons, Ltd.)

The interfacing of capillary electrophoresis (CE) with mass spectrometry (MS) is well established and may be accomplished by use of either a coaxial arrangement or by employing a liquid T-junction. In both these interfaces a make-up flow is introduced. This is required because of the mismatch in flow rates for capillary electrophoresis approximately nL/min and 'true' electrospray approximately 2-10 microL/min. Electrical connectivity may also be established where the liquid flows meet (the introduction of nanospray renders the use of make-up flow unnecessary). Hydrogen/deuterium (H/D) exchange occurs in solution when there are labile hydrogen atoms present in a molecule. The establishment of the presence and the number of such exchangeable hydrogen atoms may be of importance in the identification and differentiation of compounds. It may also be an aid in the structural elucidation of unknown materials. We have investigated the feasibility of carrying out H/D exchange via a CE/MS interface. This involved the addition of D2O to the sheath flow and our preliminary results showing the separations of drug substances, subsequently undergoing exchange, are presented.

The influences of medium type, inoculum density, and a cold incubation on antimicrobial assay sensitivity were tested. The largest and most distinct zones were produced using nutrient agar and the 1/10 4 inoculum density for Staphylococcus aureus and Proteus mirabilis but a 1/10 12 inoculum density for Pseudomonas aeruginosa and Escherichia coli . The greatest number of zones were detected without cold incubation. Using this method, eight plants from Belize were screened for antibacterial activity. Six plants showed activity against the four organisms tested. Both inoculum density and medium type played important roles in assay sensitivity; however, inoculum density was of more practical significance.

An accurate and robust method involving liquid liquid extraction and capillary gas chromatographic (GC) assay with nitrogen phosphorus detection (NPD) was developed and validated for the quantitative determination of topiramate [2,3:4,5-bis-O-(-1-methylethylidene)-beta-D-fructopyranose sulfamate], Topamax, an anticonvulsant drug, in human plasma, urine, and whole blood. The galactopyranose analog of topiramate was used as the internal standard. A DB-5, fused silica capillary column (J&W Scientific, Folsom, CA) was used, yielding typical retention times of 4.95 min for topiramate and 5.32 min for the internal standard in human plasma. The assay involved organic extraction with methyl t-butyl ether (MTBE) from base, a back extraction into acid and a second extraction in MTBE. The organic solvent was evaporated, and the residue was redissolved and injected for analysis. The standard curve was validated from 0.5 to 50 microg/ml(-1) for human plasma and whole blood, and from 1.0 to 50 microg/ml(-1) for urine. Peak area ratios of drug to internal standard were determined and used to construct a standard curve. The resulting chromatograms showed no endogenous interfering peaks with the respective blank human fluids. Chromatograms corresponding to topiramate and the internal standard produced sharp peaks that were well resolved. This assay showed precision and accuracy of < or = 5%. Two minor human metabolites of topiramate did not interfere with the assay. This assay was successfully applied to determine the pharmacokinetics of topiramate during the development of this drug.

This study investigated expression of the key antioxidant enzyme copper/zinc superoxide dismutase in the villous trophoblast of the human placenta at different gestational ages from 8 weeks (last menstrual period) to term. Immunostaining for the enzyme was observed in the cytotrophoblast cells at all stages. Staining was generally absent from the syncytiotrophoblast at 8 weeks, except for small isolated areas close to the basal surface. The size and location of these areas suggested they were the result of recent cytotrophoblastic fusion. By 10 weeks, examples were more frequent and diffuse staining throughout most of the syncytiotrophoblast was observed at 12 weeks. The intensity of the immunostaining within the syncytiotrophoblast continued to increase until 14 weeks, by which time it matched generally that within the cytotrophoblast cells. A similar pattern of staining was observed within term material. These results are entirely consistent with the hypothesis that the oxygen tension within the intervillous space is low throughout the first trimester of pregnancy. They support the idea that an effective maternal circulation to the human placenta is only established at the start of the second trimester.

We have recently shown using dansyl-L-lysine exclusion studies that the release of human chorionic gonadotrophin (HCG) in conjunction with L-lactate dehydrogenase (LDH) from first trimester villi during organ culture is symptomatic of syncytiotrophoblast degeneration. The purpose of this study was to examine chorionic villi at the ultrastructural level in order to determine events occurring during organ culture. The tissue was sampled after 0, 24, 48 and 120 h in culture and processed for electron microscopy. In addition to confirming the previously recorded syncytial degeneration, the electron micrographs showed clearly the generation of a new syncytiotrophoblast layer. The new layer, derived from differentiating cytotrophoblast cells, was largely formed by 48 h and was maintained for at least 120 h in culture. This study demonstrates a model which provides an opportunity to study the differentiation of cytotrophoblast cells whilst they retain their anatomical relationships within the villous structure.

The placenta operates as a vital interface between the mother and fetus. In addition to facilitating fetal nourishment, it acts as a barrier both to potentially deleterious agents and to contact between their two immune systems. As a consequence, damage to the placenta, even on a relatively small scale, could be very dangerous to the fetus. Therefore, wound repair mechanisms are likely to be of great importance in ensuring that an intact placental barrier is re-established as soon as possible. By use of an in vitro method for injuring and subsequently culturing small pieces of first trimester villous tissue, we have observed a number of indications that a wound response is initiated. Pronounced expression of transforming growth factor-beta1, heavy infiltration of macrophages and late deposition of tenascin in the region of the wound all provide good evidence of some form of healing activity. Furthermore, we have noted that these indicators are suggestive of 'adult-type' rather than 'fetal-type' repair processes.

The use of human chorionic gonadotrophin (HCG) secretion as a measure of viability during the organ culture of human first trimester placental tissue has become a popular practice. It has been suggested that if cultured tissue is releasing large amounts of this protein hormone, there is a high level of viability. We have found, however, that the cytosolic enzyme L-lactate dehydrogenase is released into the culture supernatant in a similar daily pattern as HCG, suggesting that tissue disruption may be occurring, resulting in some of the observed hormone release. In addition, we have shown that the uptake of the fluorescent dye dansyl-L-lysine into the syncytium increases significantly from day 0 to day 4, suggesting a loss of syncytial membrane integrity. Electron micrographs show further evidence of the syncytial degeneration at the ultrastructural level, displaying extensive vacuolation and poor microvillous cover. In contrast to the degenerated state of the syncytiotrophoblast, a high level of bromodeoxyuridine incorporation is observed for cytotrophoblasts and, in particular, stromal cells up to 5 days in culture. Overall, the results suggest that the use of HCG release as a determinant of tissue viability in placental organ culture should be treated with a degree of caution.