Objective

To assess the risk of subsequent stroke among older patients discharged from an emergency department (ED) without a diagnosis of TIA or stroke.

OBJECTIVES:

To test the hypotheses that minority children with long-bone fractures are less likely to (1) receive analgesics, (2) receive opioid analgesics, and (3) achieve pain reduction.

BACKGROUND AND OBJECTIVES:

High-quality cardiopulmonary resuscitation (CPR) increases the likelihood of survival of pediatric out-of-hospital cardiac arrest (OHCA). Maintenance of high-quality CPR during transition of care between prehospital and pediatric emergency department (PED) providers is challenging. Our objective for this initiative was to minimize pauses in compressions, in alignment with American Heart Association recommendations, for patients with OHCA during the handoffs from prehospital to PED providers. We aimed to decrease interruptions in compressions during the first 2 minutes of PED care from 17 seconds (baseline data) to 10 seconds over 12 months. Our secondary aims were to decrease the length of the longest pause in compressions to <10 seconds and eliminate encounters in which time to defibrillator pad placement was >120 seconds.

Systemic sclerosis sine scleroderma (ssSSc) is a rare variant of systemic sclerosis, with only one pediatric case reported in the medical literature to date. Pulmonary arterial hypertension as the presenting feature of ssSSc is extremely rare, even in adults, and so far has never been reported in children. We report, for the first time, a case of pediatric ssSSc in a 3-year-old girl, who presented with interstitial lung disease and pulmonary hypertension. The patient was prescribed early aggressive pulmonary vasodilators combined with anti-inflammatory medications. The clinical response was good, and her current condition at 12 years of age is remarkable, considering the high mortality rates reported in adults. We underscore the importance of early aggressive treatment in future cases of similar presentation.

OBJECTIVES:

To determine if in utero selective serotonin reuptake inhibitor (SSRI) or selective serotonin norepinephrine inhibitor (SNRI) exposure is associated with developmental vulnerability in kindergarten among children whose mothers were diagnosed with prenatal mood or anxiety disorder.

Key Points

A Yersinia pestis mutant synthesizing an adjuvant form of lipid A (monophosphoryl lipid A, MPLA) displayed increased biogenesis of bacterial outer membrane vesicles (OMVs). To enhance the immunogenicity of the OMVs, we constructed an Asd-based balanced-lethal host-vector system that oversynthesized the LcrV antigen of Y. pestis, raised the amounts of LcrV enclosed in OMVs by the type II secretion system, and eliminated harmful factors like plasminogen activator (Pla) and murine toxin from the OMVs. Vaccination with OMVs containing MPLA and increased amounts of LcrV with diminished toxicity afforded complete protection in mice against subcutaneous challenge with 8 x 10⁵ CFU (80,000 50% lethal dose [LD₅₀]) and intranasal challenge with 5 x 10³ CFU (50 LD₅₀) of virulent Y. pestis. This protection was significantly superior to that resulting from vaccination with LcrV/alhydrogel or rF1-V/alhydrogel. At week 4 postimmunization, the OMV-immunized mice showed more robust titers of antibodies against LcrV, Y. pestis whole-cell lysate (YPL), and F1 antigen and more balanced IgG1:IgG2a/IgG2b-derived Th1 and Th2 responses than LcrV-immunized mice. Moreover, potent adaptive and innate immune responses were stimulated in the OMV-immunized mice. Our findings demonstrate that self-adjuvanting Y. pestis OMVs provide a novel plague vaccine candidate and that the rational design of OMVs could serve as a robust approach for vaccine development.

As the novel coronavirus spreads, communities across the United States are struggling to offer public testing. The need is urgent. Testing got off to a delayed start in the United States as a result of technical missteps and a slow response from government officials. Now cities across the country are...

Aerosol–radiation interaction (ARI) plays a significant role in the accumulation of fine particulate matter (PM2.5) by stabilizing the planetary boundary layer and thus deteriorating air quality during haze events. However, modification of photolysis by aerosol scattering or absorbing solar radiation (aerosol–photolysis interaction or API) alters the atmospheric oxidizing capacity, decreases...

Sub-Neptunes are common among the discovered exoplanets. However, lack of knowledge on the state of matter in H2O-rich setting at high pressures and temperatures (P−T) places important limitations on our understanding of this planet type. We have conducted experiments for reactions between SiO2 and H2O as archetypal materials for rock...

G-quadruplex, assembled from a square array of guanine (G) molecules, is an important structure with crucial biological roles in vivo but also a versatile template for ordered functional materials. Although the understanding of G-quadruplex structures is the focus of numerous studies, little is known regarding the control of G-quartet stacking...

The factors and mechanisms involved in vacuolar transport in plants, and in particular those directing vesicles to their target endomembrane compartment, remain largely unknown. To identify components of the vacuolar trafficking machinery, we searched for Arabidopsis modified transport to the vacuole (mtv) mutants that abnormally secrete the synthetic vacuolar cargo...

Proteasomes are essential protease complexes that maintain cellular homeostasis, and aberrant proteasomal activity supports cancer development. The regulatory mechanisms and biological function of the ubiquitin-26S proteasome have been studied extensively, while those of the ubiquitin-independent 20S proteasome system remain obscure. Here, we show that the cap ’n’ collar (CNC) family transcription factor NRF3 specifically enhances 20S proteasome assembly in cancer cells and that 20S proteasomes contribute to colorectal cancer development through ubiquitin-independent proteolysis of the tumor suppressor p53 and retinoblastoma (Rb) proteins. The NRF3 gene is highly expressed in many cancer tissues and cell lines and is important for cancer cell growth. In cancer cells, NRF3 upregulates the assembly of the 20S proteasome by directly inducing the gene expression of the 20S proteasome maturation protein POMP. Interestingly, NRF3 knockdown not only increases p53 and Rb protein levels but also increases p53 activities for tumor suppression, including cell cycle arrest and induction of apoptosis. Furthermore, protein stability and cell viability assays using two distinct proteasome inhibitor anticancer drugs, the 20S proteasome inhibitor bortezomib and the ubiquitin-activating enzyme E1 inhibitor TAK-243, show that the upregulation of the NRF3-POMP axis leads to ubiquitin-independent proteolysis of p53 and Rb and to impaired sensitivity to bortezomib but not TAK-243. More importantly, the NRF3-POMP axis supports tumorigenesis and metastasis, with higher NRF3/POMP expression levels correlating with poor prognoses in patients with colorectal or rectal adenocarcinoma. These results suggest that the NRF3-POMP-20S proteasome assembly axis is significant for cancer development via ubiquitin-independent proteolysis of tumor suppressor proteins.

Cep57 has been characterized as a component of a pericentriolar complex containing Cep63 and Cep152. Interestingly, Cep63 and Cep152 self-assemble into a pericentriolar cylindrical architecture, and this event is critical for the orderly recruitment of Plk4, a key regulator of centriole duplication. However, the way in which Cep57 interacts with the Cep63-Cep152 complex and contributes to the structure and function of Cep63-Cep152 self-assembly remains unknown. We demonstrate that Cep57 interacts with Cep63 through N-terminal motifs and associates with Cep152 via Cep63. Three-dimensional structured illumination microscopy (3D-SIM) analyses suggested that the Cep57-Cep63-Cep152 complex is concentrically arranged around a centriole in a Cep57-in and Cep152-out manner. Cep57 mutant cells defective in Cep63 binding exhibited improper Cep63 and Cep152 localization and impaired Sas6 recruitment for procentriole assembly, proving the significance of the Cep57-Cep63 interaction. Intriguingly, Cep63 fused to a microtubule (MT)-binding domain of Cep57 functioned in concert with Cep152 to assemble around stabilized MTs in vitro. Thus, Cep57 plays a key role in architecting the Cep63-Cep152 assembly around centriolar MTs and promoting centriole biogenesis. This study may offer a platform to investigate how the organization and function of the pericentriolar architecture are altered by disease-associated mutations found in the Cep57-Cep63-Cep152 complex.

Recently, abundant evidence has clarified that long noncoding RNAs (lncRNAs) play an oncogenic or anticancer role in the tumorigenesis and development of diverse human cancers. Described as a crucial regulator in some cancers, MIR22HG has not yet been studied in laryngocarcinoma and therefore the underlying regulatory role of MIR22HG in laryngocarcinoma is worth detecting. In this study, MIR22HG expression in laryngocarcinoma cells was confirmed to be downregulated, and upregulated MIR22HG expression led to suppressive effects on laryngocarcinoma cell proliferation and migration. Molecular mechanism assays revealed that MIR22HG sponges miR-5000-3p in laryngocarcinoma cells. Besides, decreased expression of miR-5000-3p suppressed laryngocarcinoma cell proliferation and migration. Moreover, the FBXW7 gene was reported to be a downstream target gene of miR-5000-3p in laryngocarcinoma cells. More importantly, rescue assays verified that FBXW7 depletion or miR-5000-3p upregulation countervailed the repressive effects of MIR22HG overexpression on laryngocarcinoma progression. In addition, E2F6 was proved to be capable of inhibiting MIR22HG transcription in laryngocarcinoma cells. To sum up, E2F6-induced downregulation of MIR22HG promotes laryngocarcinoma progression through the miR-5000-3p/FBXW7 axis.

Microtubule-associated serine/threonine kinase like (MASTL), also known as Greatwall (Gwl) kinase, has an important role in the regulation of mitosis. By inhibiting protein phosphatase 2A (PP2A), it plays a crucial role in activating one of the most important mitotic kinases, known as cyclin-dependent kinase 1 (CDK1). MASTL has been seen to be upregulated in various types of cancers and is also involved in tumor recurrence. It is activated by CDK1 through phosphorylations in the activation/T-loop, but the complete mechanism of its activation is still unclear. Here, we report that AKT phosphorylates MASTL at residue T299, which plays a critical role in its activation. Our results suggest that AKT increases CDK1-mediated phosphorylation and hence the activity of MASTL, which, in turn, promotes mitotic progression through PP2A inhibition. We also show that the oncogenic potential of AKT is augmented by MASTL activation, since AKT-mediated proliferation in colorectal cell lines can be attenuated by inhibiting and/or silencing MASTL. In brief, we report that AKT plays an important role in the progression of mitosis in mammalian cells and that it does so through the phosphorylation and activation of MASTL.

The metabolic state of the brain can greatly impact neurologic function. Evidence of this includes the therapeutic benefit of a ketogenic diet in neurologic diseases, including epilepsy. However, brain lipid bioenergetics remain largely uncharacterized. The existence, capacity, and relevance of mitochondrial fatty acid β-oxidation (FAO) in the brain are highly controversial, with few genetic tools available to evaluate the question. We have provided evidence for the capacity of brain FAO using a pan-brain-specific conditional knockout (KO) mouse incapable of FAO due to the loss of carnitine palmitoyltransferase 2, the product of an obligate gene for FAO (CPT2^B–/–). Loss of central nervous system (CNS) FAO did not result in gross neuroanatomical changes or systemic differences in metabolism. Loss of CPT2 in the brain did not result in robustly impaired behavior. We demonstrate by unbiased and targeted metabolomics that the mammalian brain oxidizes a substantial quantity of long-chain fatty acids in vitro and in vivo. Loss of CNS FAO results in robust accumulation of long-chain acylcarnitines in the brain, suggesting that the mammalian brain mobilizes fatty acids for their oxidation, irrespective of diet or metabolic state. Together, these data demonstrate that the mammalian brain oxidizes fatty acids under normal circumstances with little influence from or on peripheral tissues.

Introduction:

Understanding patients’ perspectives about their diabetes and what causes those perspectives to shift is critical to building a treatment strategy with the patient and facilitating patient self-management behavior. Key "turning points" can provide crucial opportunities to enact a change in perspective. The goal of this study is to identify "turning points" that have significance to diabetes-related health.

JABFM seeks to widely disseminate its peer-reviewed publications, increasing article visibility for the purpose of advancing scientific knowledge. We describe the journal’s approach to dissemination and recommend a number of strategies for authors to implement, including press releases and social media. Providing the article’s digital object identifier (DOI) is most useful, compared with links that can break, or attaching the article PDF, which will depress reader metrics. All JABFM articles are freely accessible online worldwide.

Flora Borne, Alexander Kovalev, Stanislav Gorb, and Virginie Courtier-Orgogozo

Insects produce a variety of adhesives for diverse functions such as locomotion, mating, and egg or pupal anchorage to substrates. Although they are important for the biology of organisms and potentially represent a great resource for developing new materials, insect adhesives have been little studied so far. Here, we examined the adhesive properties of the larval glue of Drosophila melanogaster. This glue is made of glycosylated proteins and allows the animal to adhere to a substrate during metamorphosis. We designed an adhesion test to measure the pull-off force required to detach a pupa from a substrate and to evaluate the contact area covered by the glue. We found that the pupa adheres with similar forces to a variety of substrates (with distinct roughness, hydrophilic and charge properties). We obtained an average pull-off force of 217 mN, corresponding to 15,500 times the weight of a pupa and an adhesion strength of 137–244 kPa. Surprisingly, the pull-off forces did not depend on the contact area. Our study paves the way for a genetic dissection of the components of D. melanogaster glue that confer its particular adhesive properties.

Gabriela Montejo-Kovacevich, Simon H. Martin, Joana I. Meier, Caroline N. Bacquet, Monica Monllor, Chris D. Jiggins, and Nicola J. Nadeau

Microclimatic variability in tropical forests plays a key role in shaping species distributions and their ability to cope with environmental change, especially for ectotherms. Nonetheless, currently available climatic datasets lack data from the forest interior and, furthermore, our knowledge of thermal tolerance among tropical ectotherms is limited. We therefore studied natural variation in the microclimate experienced by tropical butterflies in the genus Heliconius across their Andean range in a single year. We found that the forest strongly buffers temperature and humidity in the understorey, especially in the lowlands, where temperatures are more extreme. There were systematic differences between our yearly records and macroclimate databases (WorldClim2), with lower interpolated minimum temperatures and maximum temperatures higher than expected. We then assessed thermal tolerance of 10 Heliconius butterfly species in the wild and found that populations at high elevations had significantly lower heat tolerance than those at lower elevations. However, when we reared populations of the widespread H. erato from high and low elevations in a common-garden environment, the difference in heat tolerance across elevations was reduced, indicating plasticity in this trait. Microclimate buffering is not currently captured in publicly available datasets, but could be crucial for enabling upland shifting of species sensitive to heat such as highland Heliconius. Plasticity in thermal tolerance may alleviate the effects of global warming on some widespread ectotherm species, but more research is needed to understand the long-term consequences of plasticity on populations and species.

Lucy A. Winder, Stewart A. White, Andreas Nord, Barbara Helm, and Dominic J. McCafferty

During winter at temperate and high latitudes, the low ambient temperatures, limited food supplies and short foraging periods mean small passerines show behavioural, morphological and physiological adaptations to reduce the risk of facing energy shortages. Peripheral tissues vasoconstrict in low ambient temperatures to reduce heat loss and cold injury. Peripheral vasoconstriction has been observed with food restriction in captivity but has yet to be explored in free-ranging animals. We experimentally food restricted both wild and captive great tits (Parus major) during winter months and measured surface temperatures of the bill and eye region using thermal imaging, to investigate whether birds show rapid local heterothermic responses, which may reduce their thermoregulatory costs when facing a perceived imminent food shortage. Our results of a continuously filmed wild population showed that bill temperature was immediately reduced in response to food restriction compared with when food was available ad libitum, an apparent autonomic response. Such immediacy implies a ‘pre-emptive’ response before the bird experiences any shortfalls in energy reserves. We also demonstrate temporal variation in vasoconstriction of the bill, with bill temperature gradually rising throughout the food restriction after the initial drop. Eye-region temperature in the wild birds remained at similar levels throughout food restriction compared with unrestricted birds, possibly reflecting the need to maintain steady circulation to the central nervous and visual systems. Our findings provide evidence that birds selectively allow the bill to cool when a predictable food supply is suddenly disrupted, probably as a means of minimising depletion of body reserves for a perceived future shortage in energy.

Fredrik Andreasson, Arne Hegemann, Andreas Nord, and Jan-Ake Nilsson

The capacity to get rid of excess heat produced during hard work is a possible constraint on parental effort during reproduction [heat dissipation limit (HDL) theory]. We released hard-working blue tits (Cyanistes caeruleus) from this constraint by experimentally removing ventral plumage. We then assessed whether this changed their reproductive effort (feeding rate and nestling size) and levels of self-maintenance (change in body mass and innate immune function). Feather-clipped females reduced the number of feeding visits and increased levels of constitutive innate immunity compared with unclipped females but did not fledge smaller nestlings. Thus, they increased self-maintenance without compromising current reproductive output. In contrast, feather clipping did not affect the number of feeding visits or innate immune function in males, despite increased heat loss rate. Our results show that analyses of physiological parameters, such as constitutive innate immune function, can be important when trying to understand sources of variation in investment in self-maintenance versus reproductive effort and that risk of overheating can influence innate immune function during reproduction.

Ricardo Benini, Leandro A. Oliveira, Lucas Gomes-de-Souza, Bruno Rodrigues, and Carlos C. Crestani

This study evaluated the effect of exposure to either a chronic variable stress (CVS) protocol or social isolation, as well as treadmill exercise training, in the habituation of the cardiovascular response upon repeated exposure to restraint stress in rats. The habituation of the corticosterone response to repeated restraint stress was also evaluated. For this, animals were subjected to either acute or 10 daily sessions of 60 min of restraint stress. CVS and social isolation protocols lasted for 10 consecutive days, whereas treadmill training was performed for 1 h per day, 5 days per week for 8 weeks. We observed that the increase in serum corticosterone was reduced during both the stress and the recovery period of the 10th session of restraint. Habituation of the cardiovascular response was identified in terms of a faster return of heart rate to baseline values during the recovery period of the 10th session of restraint. The increase in blood pressure and the decrease in tail skin temperature were similar at the 1st and 10th session of restraint. Exposure to CVS, social isolation or treadmill exercise training inhibited the habituation of the restraint-evoked tachycardia. Additionally, CVS increased the blood pressure response at the 10th session of restraint, whereas social isolation enhanced both the tachycardia during the first session and the drop in skin temperature at the 10th session of restraint. Taken together, these findings provide new evidence that pathologies evoked by stress might be related to impairment in the habituation process to homotypic stressors.

Andreas Nord, Arne Hegemann, and Lars P. Folkow

Animals in seasonal environments must prudently manage energy expenditure to survive the winter. This may be achieved through reductions in the allocation of energy for various purposes (e.g. thermoregulation, locomotion, etc.). We studied whether such trade-offs also include suppression of the innate immune response, by subjecting captive male Svalbard ptarmigan (Lagopus muta hyperborea) to bacterial lipopolysaccharide (LPS) during exposure to either mild temperature (0°C) or cold snaps (acute exposure to –20°C), in constant winter darkness when birds were in energy-conserving mode, and in constant daylight in spring. The innate immune response was mostly unaffected by temperature. However, energy expenditure was below baseline when birds were immune challenged in winter, but significantly above baseline in spring. This suggests that the energetic component of the innate immune response was reduced in winter, possibly contributing to energy conservation. Immunological parameters decreased (agglutination, lysis, bacteriostatic capacity) or did not change (haptoglobin/PIT54) after the challenge, and behavioural modifications (anorexia, mass loss) were lengthy (9 days). While we did not study the mechanisms explaining these weak, or slow, responses, it is tempting to speculate they may reflect the consequences of having evolved in an environment where pathogen transmission rate is presumably low for most of the year. This is an important consideration if climate change and increased exploitation of the Arctic would alter pathogen communities at a pace outwith counter-adaption in wildlife.

Ana Gabriela Jimenez, Emily Cornelius Ruhs, Kailey J. Tobin, Katie N. Anderson, Audrey Le Pogam, Lyette Regimbald, and Francois Vezina

Seasonal changes in maximal thermogenic capacity (M_sum) in wild black-capped chickadees suggests that adjustments in metabolic performance are slow and begin to take place before winter peaks. However, when mean minimal ambient temperature (T_a) reaches –10°C, the chickadee phenotype appears to provide enough spare capacity to endure days with colder T_a, down to –20°C or below. This suggests that birds could also maintain a higher antioxidant capacity as part of their cold-acclimated phenotype to deal with sudden decreases in temperature. Here, we tested how environmental mismatch affected oxidative stress by comparing cold-acclimated (–5°C) and transition (20°C) phenotypes in chickadees exposed to an acute 15°C drop in temperature with that of control individuals. We measured superoxide dismutase, catalase and glutathione peroxidase activities, as well as lipid peroxidation damage and antioxidant scavenging capacity in pectoralis muscle, brain, intestine and liver. We generally found differences between seasonal phenotypes and across tissues, but no differences with respect to an acute cold drop treatment. Our data suggest oxidative stress is closely matched to whole-animal physiology in cold-acclimated birds compared with transition birds, implying that changes to the oxidative stress system happen slowly.

Irene Verhagen, Barbara M. Tomotani, Phillip Gienapp, and Marcel E. Visser

Phenotypic plasticity is an important mechanism by which an individual can adapt its seasonal timing to predictable, short-term environmental changes by using predictive cues. Identification of these cues is crucial to forecast the response of species to long-term environmental change and to study their potential to adapt. Individual great tits (Parus major) start reproduction early under warmer conditions in the wild, but whether this effect is causal is not well known. We housed 36 pairs of great tits in climate-controlled aviaries and 40 pairs in outdoor aviaries, where they bred under artificial contrasting temperature treatments or in semi-natural conditions, respectively, for two consecutive years, using birds from lines selected for early and late egg laying. We thus obtained laying dates in two different thermal environments for each female. Females bred earlier under warmer conditions in climate-controlled aviaries, but not in outdoor aviaries. The latter was inconsistent with laying dates from our wild population. Further, early selection line females initiated egg laying consistently ~9 days earlier than late selection line females in outdoor aviaries, but we found no difference in the degree of plasticity (i.e. the sensitivity to temperature) in laying date between selection lines. Because we found that temperature causally affects laying date, climate change will lead to earlier laying. This advancement is, however, unlikely to be sufficient, thereby leading to selection for earlier laying. Our results suggest that natural selection may lead to a change in mean phenotype, but not to a change in the sensitivity of laying dates to temperature.

Erin Gorrell, Ashley Shemery, Jesse Kowalski, Miranda Bodziony, Nhlalala Mavundza, Amber R. Titus, Mark Yoder, Sarah Mull, Lydia A. Heemstra, Jacob G. Wagner, Megan Gibson, Olivia Carey, Diamond Daniel, Nicholas Harvey, Meredith Zendlo, Megan Rich, Scott Everett, Chaitanya K. Gavini, Tariq I. Almundarij, Diane Lorton, and Colleen M. Novak

Non-shivering thermogenesis can promote negative energy balance and weight loss. In this study, we identified a contextual stimulus that induces rapid and robust thermogenesis in skeletal muscle. Rats exposed to the odor of a natural predator (ferret) showed elevated skeletal muscle temperatures detectable as quickly as 2 min after exposure, reaching maximum thermogenesis of >1.5°C at 10–15 min. Mice exhibited a similar thermogenic response to the same odor. Ferret odor induced a significantly larger and qualitatively different response from that of novel or aversive odors, fox odor or moderate restraint stress. Exposure to predator odor increased energy expenditure, and both the thermogenic and energetic effects persisted when physical activity levels were controlled. Predator odor-induced muscle thermogenesis is subject to associative learning as exposure to a conditioned stimulus provoked a rise in muscle temperature in the absence of the odor. The ability of predator odor to induce thermogenesis is predominantly controlled by sympathetic nervous system activation of β-adrenergic receptors, as unilateral sympathetic lumbar denervation and a peripherally acting β-adrenergic antagonist significantly inhibited predator odor-induced muscle thermogenesis. The potential survival value of predator odor-induced changes in muscle physiology is reflected in an enhanced resistance to running fatigue. Lastly, predator odor-induced muscle thermogenesis imparts a meaningful impact on energy expenditure as daily predator odor exposure significantly enhanced weight loss with mild calorie restriction. This evidence signifies contextually provoked, centrally mediated muscle thermogenesis that meaningfully impacts energy balance.

Peter J. Edmunds and Scott C. Burgess

Experiments with coral fragments (i.e. nubbins) have shown that net calcification is depressed by elevated P_CO2. Evaluating the implications of this finding requires scaling of results from nubbins to colonies, yet the experiments to codify this process have not been carried out. Building from our previous research demonstrating that net calcification of Pocillopora verrucosa (2–13 cm diameter) was unaffected by P_CO2 (400 and 1000 µatm) and temperature (26.5 and 29.7°C), we sought generality to this outcome by testing how colony size modulates P_CO2 and temperature sensitivity in a branching acroporid. Together, these taxa represent two of the dominant lineages of branching corals on Indo-Pacific coral reefs. Two trials conducted over 2 years tested the hypothesis that the seasonal range in seawater temperature (26.5 and 29.2°C) and a future P_CO2 (1062 µatm versus an ambient level of 461 µatm) affect net calcification of an ecologically relevant size range (5–20 cm diameter) of colonies of Acropora hyacinthus. As for P. verrucosa, the effects of temperature and P_CO2 on net calcification (mg day^–1) of A. verrucosa were not statistically detectable. These results support the generality of a null outcome on net calcification of exposing intact colonies of branching corals to environmental conditions contrasting seasonal variation in temperature and predicted future variation in P_CO2. While there is a need to expand beyond an experimental culture relying on coral nubbins as tractable replicates, rigorously responding to this need poses substantial ethical and logistical challenges.

Surendra Singh Patel, Sanyami Zunjarrao, and Beena Pillai

Eisenia fetida, the common vermicomposting earthworm, shows robust regeneration of posterior segments removed by amputation. During the period of regeneration, the newly formed tissue initially contains only undifferentiated cells but subsequently differentiates into a variety of cell types including muscle, nerve and vasculature. Transcriptomics analysis, reported previously, provided a number of candidate non-coding RNAs that were induced during regeneration. We found that one such long non-coding RNA (lncRNA) is expressed in the skin, only at the base of newly formed chaetae. The spatial organization and precise arrangement of the regenerating chaetae and the cells expressing the lncRNA on the ventral side clearly support a model wherein the regenerating tissue contains a zone of growth and cell division at the tip and a zone of differentiation at the site of amputation. The temporal expression pattern of the lncRNA, named Neev, closely resembled the pattern of chitin synthase genes, implicated in chaetae formation. We found that the lncRNA has 49 sites for binding a set of four microRNAs (miRNAs) while the chitin synthase 8 mRNA has 478 sites. The over-representation of shared miRNA sites suggests that lncRNA Neev may act as a miRNA sponge to transiently de-repress chitin synthase 8 during formation of new chaetae in the regenerating segments of Eisenia fetida.

Vanessa Corby-Harris, Megan E. Deeter, Lucy Snyder, Charlotte Meador, Ashley C. Welchert, Amelia Hoffman, and Bethany T. Obernesser

Recent widespread honey bee (Apis mellifera) colony loss is attributed to a variety of stressors, including parasites, pathogens, pesticides and poor nutrition. In principle, we can reduce stress-induced declines in colony health by either removing the stressor or increasing the bees' tolerance to the stressor. This latter option requires a better understanding than we currently have of how honey bees respond to stress. Here, we investigated how octopamine, a stress-induced hormone that mediates invertebrate physiology and behavior, influences the health of young nurse-aged bees. Specifically, we asked whether octopamine induces abdominal lipid and hypopharyngeal gland (HG) degradation, two physiological traits of stressed nurse bees. Nurse-aged workers were treated topically with octopamine and their abdominal lipid content, HG size and HG autophagic gene expression were measured. Hemolymph lipid titer was measured to determine whether tissue degradation was associated with the release of nutrients from these tissues into the hemolymph. The HGs of octopamine-treated bees were smaller than control bees and had higher levels of HG autophagy gene expression. Octopamine-treated bees also had higher levels of hemolymph lipid compared with control bees. Abdominal lipids did not change in response to octopamine. Our findings support the hypothesis that the HGs are a rich source of stored energy that can be mobilized during periods of stress.

Ewa Komar, Dina K. N. Dechmann, Nicolas J. Fasel, Marcin Zegarek, and Ireneusz Ruczynski

Balancing energy budgets can be challenging, especially in periods of food shortage, adverse weather conditions and increased energy demand due to reproduction. Bats have particularly high energy demands compared to other mammals and regularly use torpor to save energy. However, while torpor limits energy expenditure, it can also downregulate important processes, such as sperm production. This constraint could result in a trade-off between energy saving and future reproductive capacity. We mimicked harsh conditions by restricting food and tested the effect on changes in body mass, torpor use and seasonal sexual maturation in male parti-coloured bats (Vespertilio murinus). Food-restricted individuals managed to maintain their initial body mass, while in well-fed males, mass increased. Interestingly, despite large differences in food availability, there were only small differences in torpor patterns. However, well-fed males reached sexual maturity up to half a month earlier. Our results thus reveal a complex trade-off in resource allocation; independent of resource availability, males maintain a similar thermoregulation strategy and favour fast sexual maturation, but limited resources and low body mass moderate this latter process.

Fredrik Christiansen, Kate R. Sprogis, Jasmin Gross, Juliana Castrillon, Hunter A. Warick, Eva Leunissen, and Susan Bengtson Nash

An animal's body condition provides valuable information for ecophysiological studies, and is an important measure of fitness in population monitoring and conservation. While both the external body shape of an animal and its internal tissues (i.e. fat content) can be used as a measure of body condition, the relationship between the two is not always linear. We compared the morphological body condition (external metric obtained through aerial photogrammetry) of migrating humpback whales (Megaptera novaeangliae) with their outer blubber lipid concentration (internal metric obtained through blubber biopsy sampling) off the coast of south-west Australia early and late in the breeding season (spanning ~4.5 months). The external body condition index of juvenile and adult humpback whales decreased by 26.9 (from 18.8% to –8.1%) and 12.0 percentage points (from 8.6% to –3.4%), respectively, between the early and late phase. In contrast, we found no intra-seasonal change in blubber lipid concentration, and no difference between reproductive classes (juveniles, adults and lactating females); however, the small sample size prevented us from effectively testing these effects. Importantly, however, in the 33 animals for which paired metrics were obtained, we found no correlation between the morphometric body condition index and the blubber lipid concentration of individual whales. The lack of a linear relationship suggests that changes in outer blubber lipid concentration do not reflect external changes in body shape, thus limiting the utility of outer blubber lipid reserves for individual body condition evaluation. The wider spectrum of change in body morphometry captured with aerial photogrammetry supports the use of body morphometry as a reliable and well-tested method.

Ayelen M. Blanco, Juan I. Bertucci, Jose L. Soengas, and Suraj Unniappan

This research assessed the direct effects of insulin on nutrient-sensing mechanisms in the brain of rainbow trout (Oncorhynchus mykiss) using an in vitro approach. Cultured hypothalamus and hindbrain were exposed to 1 µmol l^–1 insulin for 3 h, and signals involved in appetite regulation and nutrient-sensing mechanisms were measured. Additionally, the involvement of the phosphatidylinositide 3-kinase (PI3K)/protein kinase B (Akt) signaling pathway in the actions of insulin was studied by using the inhibitor wortmannin. Treatment with insulin alone did not elicit many changes in the appetite regulators and nutrient-sensing-related genes and enzymes tested in the hypothalamus and hindbrain. However, we found that, when insulin and nutrients were added together, insulin reversed most of the effects exerted by nutrients alone, suggesting that insulin changes responsiveness to nutrients at the central level. Effects reversed by insulin included expression levels of genes related to the sensing of both glucose (slc2a2, slc5a1, gck, pck1, pklr, g6pcb, gys1, tas1r3 and nr1h3 in the hindbrain, and slc2a2, pklr and pck1 in the hypothalamus) and fatty acid (cd36 in the hindbrain, and cd36 and acly in the hypothalamus). Nutrient-induced changes in the activity of Acly and Cpt-1 in the hindbrain and of Pepck, Acly, Fas and Hoad in the hypothalamus were also reversed by insulin. Most of the insulin effects disappeared in the presence of wortmannin, suggesting the PI3K/Akt pathway is a mediator of the effects of insulin reported here. This study adds new information to our knowledge of the mechanisms regulating nutrient sensing in fish.

Yves Bötsch, Zulima Tablado, Bettina Almasi, and Lukas Jenni

Outdoor recreational activities are booming and most animals perceive humans as predators, which triggers behavioural and/or physiological reactions [e.g. heart rate increase, activation of the hypothalamic–pituitary–adrenal (HPA) axis]. Physiological stress reactions have been shown to affect the immune system of an animal and therefore may also affect the amount of maternal antibodies a female transmits to her offspring. A few studies have revealed that the presence of predators affects the amount of maternal antibodies deposited into eggs of birds. In this study, using Eurasian blue and great tit offspring (Cyanistes caeruleus and Parus major) as model species, we experimentally tested whether human recreation induces changes in the amount of circulating antibodies in young nestlings and whether this effect is modulated by habitat and competition. Moreover, we investigated whether these variations in antibody titre in turn have an impact on hatching success and offspring growth. Nestlings of great tit females that had been disturbed by experimental human recreation during egg laying had lower antibody titres compared with control nestlings. Antibody titre of nestling blue tits showed a negative correlation with the presence of great tits, rather than with human disturbance. The hatching success was positively correlated with the average amount of antibodies in great tit nestlings, independent of the treatment. Antibody titre in the first days of life in both species was positively correlated with body mass, but this relationship disappeared at fledging and was independent of treatment. We suggest that human recreation may have caused a stress-driven activation of the HPA axis in breeding females, chronically increasing their circulating corticosterone, which is known to have an immunosuppressive function. Either, lower amounts of antibodies are transmitted to nestlings or impaired transfer mechanisms lead to lower amounts of immunoglobulins in the eggs. Human disturbance could, therefore, have negative effects on nestling survival at early life-stages, when nestlings are heavily reliant on maternal antibodies, and in turn lead to lower breeding success and parental fitness. This is a so far overlooked effect of disturbance on early life in birds.

Daniel P. Small, Piero Calosi, Samuel P. S. Rastrick, Lucy M. Turner, Stephen Widdicombe, and John I. Spicer

Regulation of extracellular acid–base balance, while maintaining energy metabolism, is recognised as an important aspect when defining an organism's sensitivity to environmental changes. This study investigated the haemolymph buffering capacity and energy metabolism (oxygen consumption, haemolymph [l-lactate] and [protein]) in early benthic juveniles (carapace length <40 mm) of the European lobster, Homarus gammarus, exposed to elevated temperature and P_CO2. At 13°C, H. gammarus juveniles were able to fully compensate for acid–base disturbances caused by the exposure to elevated seawater P_CO2 at levels associated with ocean acidification and carbon dioxide capture and storage (CCS) leakage scenarios, via haemolymph [HCO₃^–] regulation. However, metabolic rate remained constant and food consumption decreased under elevated P_CO2, indicating reduced energy availability. Juveniles at 17°C showed no ability to actively compensate haemolymph pH, resulting in decreased haemolymph pH particularly under CCS conditions. Early benthic juvenile lobsters at 17°C were not able to increase energy intake to offset increased energy demand and therefore appear to be unable to respond to acid–base disturbances due to increased P_CO2 at elevated temperature. Analysis of haemolymph metabolites suggests that, even under control conditions, juveniles were energetically limited. They exhibited high haemolymph [l-lactate], indicating recourse to anaerobic metabolism. Low haemolymph [protein] was linked to minimal non-bicarbonate buffering and reduced oxygen transport capacity. We discuss these results in the context of potential impacts of ongoing ocean change and CCS leakage scenarios on the development of juvenile H. gammarus and future lobster populations and stocks.

Zhi-Jun Zhao, Davina Derous, Abby Gerrard, Jing Wen, Xue Liu, Song Tan, Catherine Hambly, and John R. Speakman

Lactating mice increase food intake 4- to 5-fold, reaching an asymptote in late lactation. A key question is whether this asymptote reflects a physiological constraint, or a maternal investment strategy (a ‘restraint’). We exposed lactating mice to periods of food restriction, hypothesizing that if the limit reflected restraint, they would compensate by breaching the asymptote when refeeding. In contrast, if it was a constraint, they would by definition be unable to increase their intake on refeeding days. Using isotope methods, we found that during food restriction, the females shut down milk production, impacting offspring growth. During refeeding, food intake and milk production rose again, but not significantly above unrestricted controls. These data provide strong evidence that asymptotic intake in lactation reflects a physiological/physical constraint, rather than restraint. Because hypothalamic neuropeptide Y (Npy) was upregulated under both states of restriction, this suggests the constraint is not imposed by limits in the capacity to upregulate hunger signalling (the saturated neural capacity hypothesis). Understanding the genetic basis of the constraint will be a key future goal and will provide us additional information on the nature of the constraining factors on reproductive output, and their potential links to life history strategies.

Samantha M. Logan, Kama E. Szereszewski, Nigel C. Bennett, Daniel W. Hart, Barry van Jaarsveld, Matthew E. Pamenter, and Kenneth B. Storey

Mole-rats are champions of self-preservation, with increased longevity compared to other rodents their size, strong antioxidant capabilities, and specialized defenses against endogenous oxidative stress. However, how the brains of these subterranean mammals handle acute in vivo hypoxia is poorly understood. This study is the first to examine the molecular response to low oxygen in six different species of hypoxia-tolerant mole-rats from sub-Saharan Africa. Protein carbonylation, a known marker of DNA damage (hydroxy-2’-deoxyguanosine), and antioxidant capacity did not change following hypoxia but HIF-1 protein levels increased significantly in the brains of two species. Nearly 30 miRNAs known to play roles in hypoxia-tolerance were differentially regulated in a species-specific manner. The miRNAs exhibiting the strongest response to low oxygen stress inhibit apoptosis and regulate neuroinflammation, likely providing neuroprotection. A principal component analysis using a subset of the molecular targets assessed herein revealed differences between control and hypoxic groups for two solitary species (Georychus capensis and Bathyergus suillus), which are ecologically adapted to a normoxic environment, suggesting a heightened sensitivity to hypoxia relative to species that may experience hypoxia more regularly in nature. By contrast, all molecular data were included in the PCA to detect a difference between control and hypoxic populations of eusocial Heterocephalus glaber, indicating they may require many lower-fold changes in signaling pathways to adapt to low oxygen settings. Finally, none of the Cryptomys hottentotus subspecies showed a statistical difference between control and hypoxic groups, presumably due to hypoxia-tolerance derived from environmental pressures associated with a subterranean and social lifestyle.

Juha Nuutila, Anna E. Honkanen, Kyösti Heimonen, and Matti Weckström

Using tethered American cockroaches walking on a trackball in a spherical virtual reality environment, we tested optomotor responses to horizontally moving black-and-white gratings of different vertical extent under six different light intensities. We found that shortening the vertical extent of the wide-field stimulus grating within a light level weakened response strength, reduced average velocity, and decreased angular walking distance. Optomotor responses with the vertically shortened stimuli persisted down to light intensity levels of 0.05 lx. Response latency seems to be independent of both the height of the stimulus and light intensity. The optomotor response started saturating at the light intensity of 5 lx, where the shortest behaviourally significant stimulus was 1°. This indicates that the number of vertical ommatidial rows needed to elicit an optomotor response at 5 lx and above is in the single digits, maybe even just one. Our behavioural results encourage further inquiry into the interplay of light intensity and stimulus size in insect dim-light vision.

Kei Jokura, Junko M. Nishino, Michio Ogasawara, and Atsuo Nishino

Ciliary movement is a fundamental process to support animal life, and the movement pattern may be altered in response to external stimuli under the control of nervous systems. Juvenile and adult ascidians have ciliary arrays around their pharyngeal gill slits (stigmata), and continuous beating is interrupted for seconds by mechanical stimuli on other parts of the body. Although it has been suggested that neural transmission to evoke ciliary arrest is cholinergic, its molecular basis has not yet been elucidated in detail. We herein attempted to clarify the molecular mechanisms underlying this neurociliary transmission in the model ascidian Ciona. Acetylcholinesterase histochemical staining showed strong signals on the laterodistal ciliated cells of stigmata, hereafter referred to as trapezial cells. The direct administration of acetylcholine (ACh) and other agonists of nicotinic ACh receptors (nAChRs) onto ciliated cells reliably evoked ciliary arrest that persisted for seconds in a dose-dependent manner. Only one isoform among all nAChR subunits encoded in the Ciona genome, called nAChR-A7/8-1, a relative of vertebrate α7 nAChRs, was expressed by trapezial cells. Exogenously expressed nAChR-A7/8-1 on Xenopus oocytes responded to ACh and other agonists with consistent pharmacological traits to those observed in vivo. Further efforts to examine signaling downstream of this receptor revealed that an inhibitor of phospholipase C (PLC) hampered ACh-induced ciliary arrest. We herein propose that homomeric α7-related nAChR-A7/8-1 mediates neurociliary transmission in Ciona stigmata to elicit persistent ciliary arrest by recruiting intracellular Ca²⁺ signaling.

Krishnamoorthy Krishnan, Asif Shahriar Nafi, Roi Gurka, and Roi Holzman

Fish larvae are the smallest self-sustaining vertebrates. As such, they face multiple challenges that stem from their minute size, and from the hydrodynamic regime in which they dwell. This regime, of intermediate Reynolds numbers, was shown to affect the swimming of larval fish and impede their ability to capture prey. Prey capture is impeded because smaller larvae produce weaker suction flows, exerting weaker forces on the prey. Previous observations on feeding larvae also showed prey exiting the mouth after initially entering it (hereafter "in-and-out"), although the mechanism causing such failures had been unclear. In this study, we used numerical simulations to investigate the hydrodynamic mechanisms responsible for the failure to feed caused by this in-and-out prey movement. Detailed kinematics of the expanding mouth during prey capture by larval Sparus aurata were used to parameterize age-specific numerical models of the flows inside the mouth. These models revealed that for small larvae which expand their mouth slowly, fluid entering the mouth cavity is expelled through the mouth before it is closed, resulting in flow reversal at the orifice. This relative efflux of water through the mouth was >8% of the influx through the mouth for younger ages. However similar effluxes were found when we simulated slow strikes by larger fish. The simulations can explain the observations of larval fish failing to fish due to the in-and-out movement of the prey. These results further highlight the importance of transporting the prey from the gape deeper into the mouth cavity in determining suction-feeding success.

Rohini Singh and Timothy A. Linksvayer

Wolbachia is a widespread group of maternally-transmitted endosymbiotic bacteria that often manipulates the reproductive strategy and life history of its hosts to favor its own transmission. Wolbachia mediated phenotypic effects are well characterized in solitary hosts, but effects in social hosts are unclear. The invasive pharaoh ant, Monomorium pharaonis, shows natural variation in Wolbachia infection between colonies and can be readily bred under laboratory conditions. We previously showed that Wolbachia-infected pharaoh ant colonies had more queen-biased sex ratios than uninfected colonies, which is expected to favor the spread of maternally-transmitted Wolbachia. Here, we further characterize the effects of Wolbachia on the short- and longer-term reproductive and life history traits of pharaoh ant colonies. First, we characterized the reproductive differences between naturally infected and uninfected colonies at three discrete time points and found that infected colonies had higher reproductive investment (i.e. infected colonies produced more new queens), particularly when existing colony queens were three months old. Next, we compared the long-term growth and reproduction dynamics of infected and uninfected colonies across their whole life cycle. Infected colonies had increased colony-level growth and early colony reproduction, resulting in a shorter colony life cycle, when compared to uninfected colonies.

Meike Stumpp, Inga Petersen, Femke Thoben, Jia-Jiun Yan, Matthias Leippe, and Marian Y. Hu

Larval stages of the abulacraria superphylum including echinoderms and hemichordates have highly alkaline midguts. To date the reason for the evolution of such extreme pH conditions in the gut of these organisms remains unknown. Here, we test the hypothesis that analogous to the acidic stomachs of vertebrates, these alkaline conditions may represent a first defensive barrier to protect from environmental pathogens.

pH-optimum curves for five different species of marine bacteria demonstrated a rapid decrease in proliferation rates by 50-60% between pH 8.5 and 9.5. Using the marine bacterium Vibrio diazotrophicus which elicits a coordinated immune response in the sea urchin larva of Strongylocentrotus purpuratus, we studied the physiological responses of the midgut pH regulatory machinery to this pathogen. Gastroscopic microelectrode measurements demonstrate a stimulation of midgut alkalization upon infection with V. diazotrophicus accompanied by an upregulation of acid-base transporter transcripts of the midgut. Pharmacological inhibition of midgut alkalization resulted in an increased mortality rate of larvae during Vibrio infection. Reductions in seawater pH resembling ocean acidification (OA) conditions lead to moderate reductions in midgut alkalization. However, these reductions in midgut pH do not affect the immune response and resilience of sea urchin larvae to a Vibrio infection under OA conditions.

Our study addressed the evolutionary benefits of the alkaline midgut of ambulacraria larval stages. The data indicate that alkaline conditions in the gut may serve as a first defensive barrier against environmental pathogens and that this mechanism can compensate for changes in seawater pH.

Brian K. McNab and Kerry A. Weston

The thermal physiology of the endangered New Zealand rockwren (Xenicus gilviventris) is examined. It is a member of the Acanthisittidae, a family unique to New Zealand. This family, derived from Gondwana, is thought to be the sister taxon to all other passerines. Rockwrens permanently reside above the climatic timberline at altitudes from 1,000 to 2,900 meters in the mountains of South Island. They feed on invertebrates and in winter face ambient temperatures far below freezing and deep deposits of snow. Their body temperature and rate of metabolism are highly variable. Rockwrens regulate body temperature at ca. 36.4°C, which in one individual decreased to 33.1°C at an ambient temperature of 9.4°C. Its rate of metabolism decreased by 30%; body temperature spontaneously returned to 36°C. The rate of metabolism in a second individual twice decreased by 35%, nearly to the basal rate expected from mass without a decrease in body temperature. The New Zealand rockwren's food habits, entrance into torpor, and continuous residence in a thermally demanding environment suggest that it may hibernate. For that conclusion to be accepted, evidence of its use of torpor for extended periods is required. Those data are not presently available. Acanthisittids are distinguished from other passerines by the combination of their permanent temperate distribution, thermal flexibility, and a propensity to evolve a flightless condition. These characteristics may principally reflect their geographical isolation in a temperate environment isolated from Gondwana for 82 million years in the absence of mammalian predators.

Yi Huang, Jazmin Osorio Mendoza, Catherine Hambly, Baoguo Li, Zengguang Jin, Li Li, Moshen Madizi, Sumei Hu, and John R. Speakman

The heat dissipation limit theory predicts lactating female mice consuming diets with lower specific dynamic action (SDA) should have enhanced lactation performance. Dietary fat has lower SDA than other macronutrients. Here we tested the effects of graded dietary fat levels on lactating Swiss mice. We fed females five diets varying in fat content from 8.3 to 66.6%. Offspring of mothers fed diets of 41.7% fat and above were heavier and fatter at weaning compared to those of 8.3% and 25% fat diets. Mice on dietary fat contents of 41.7% and above had greater metabolizable energy intake at peak lactation (8.3%: 229.4±39.6, 25%: 278.8±25.8, 41.7%: 359.6±51.5, 58.3%: 353.7±43.6, 66.6%: 346±44.7 kJ day^–1), lower daily energy expenditure (8.3%: 128.5±16, 25%: 131.6±8.4, 41.7%: 124.4±10.8, 58.3%: 115.1±10.5, 66.6%: 111.2±11.5 kJ day^–1) and thus delivered more milk energy to their offspring (8.3%: 100.8±27.3, 25%: 147.2±25.1, 41.7%: 225.1±49.6, 58.3%: 238.6±40.1, 66.6%: 234.8±41.1 kJ day^–1). Milk fat content (%) was unrelated to dietary fat content, indicating females on higher fat diets (> 41.7%) produced more rather than richer milk. Mothers consuming diets with 41.7% fat or above enhanced their lactation performance compared to those on 25% or less, probably by diverting dietary fat directly into the milk, thereby avoiding the costs of lipogenesis. At dietary fat contents above 41.7% they were either unable to transfer more dietary fat to the milk, or they chose not to do so, potentially because of a lack of benefit to the offspring that were increasingly fatter as maternal dietary fat increased.