Caught in the Act: Incipient Speciation at the Southern limit of Viburnum in the Central Andes.

A fundamental objective of evolutionary biology is to understand the origin of independently evolving species. Phylogenetic studies of species radiations rarely are able to document ongoing speciation; instead, modes of speciation, entailing geographic separation and/or ecological differentiation, are posited retrospectively. The Oreinotinus clade of Viburnum has radiated recently from north to south through the cloud forests of Mexico and Central America to the Central Andes. Our analyses support a hypothesis of incipient speciation in Oreinotinus at the southern edge of its geographic range, from central Peru to northern Argentina. Although several species and infraspecific taxa of have been recognized in this area, multiple lines of evidence and analytical approaches (including analyses of phylogenetic relationships, genetic structure, leaf morphology, and climatic envelopes) favor the recognition of just a single species, V. seemenii. We show that what has previously been recognized as V. seemenii f. minor has recently occupied the drier Tucuman-Bolivian forest region from Samaipata in Bolivia to Salta in northern Argentina. Plants in these populations form a well-supported clade with a distinctive genetic signature and they have evolved smaller, narrower leaves. We interpret this as the beginning of a within-species divergence process that has elsewhere in the neotropics resulted repeatedly in Viburnum species with a particular set of leaf ecomorphs. Specifically, the southern populations are in the process of evolving the small, glabrous, and entire leaf ecomorph that has evolved in four other montane areas of endemism. As predicted based on our studies of leaf ecomorphs in Chiapas, Mexico, these southern populations experience generally drier conditions, with large diurnal temperature fluctuations. In a central portion of the range of V. seemenii, characterized by wetter climatic conditions, we also document what may be the initial differentiation of the leaf ecomorph with larger, pubescent, and toothy leaves. The emergence of these ecomorphs thus appears to be driven by adaptation to subtly different climatic conditions in separate geographic regions, as opposed to parapatric differentiation along elevational gradients as suggested by Viburnum species distributions in other parts of the neotropics.

Paraventricular Thalamic MC3R Circuits Link Energy Homeostasis with Anxiety-Related Behavior.

The hypothalamic melanocortin system is critically involved in sensing stored energy and communicating this information throughout the brain, including to brain regions controlling motivation and emotion. This system consists of first-order agouti-related peptide (AgRP) and pro-opiomelanocortin (POMC) neurons located in the hypothalamic arcuate nucleus and downstream neurons containing the melanocortin-3 (MC3R) and melanocortin-4 receptor (MC4R). Although extensive work has characterized the function of downstream MC4R neurons, the identity and function of MC3R-containing neurons are poorly understood. Here, we used neuroanatomical and circuit manipulation approaches in mice to identify a novel pathway linking hypothalamic melanocortin neurons to melanocortin-3 receptor neurons located in the paraventricular thalamus (PVT) in male and female mice. MC3R neurons in PVT are innervated by hypothalamic AgRP and POMC neurons and are activated by anorexigenic and aversive stimuli. Consistently, chemogenetic activation of PVT MC3R neurons increases anxiety-related behavior and reduces feeding in hungry mice, whereas inhibition of PVT MC3R neurons reduces anxiety-related behavior. These studies position PVT MC3R neurons as important cellular substrates linking energy status with neural circuitry regulating anxiety-related behavior and represent a promising potential target for diseases at the intersection of metabolism and anxiety-related behavior such as anorexia nervosa.SIGNIFICANCE STATEMENT Animals must constantly adapt their behavior to changing internal and external challenges, and impairments in appropriately responding to these challenges are a hallmark of many neuropsychiatric disorders. Here, we demonstrate that paraventricular thalamic neurons containing the melanocortin-3 receptor respond to energy-state-related information and external challenges to regulate anxiety-related behavior in mice. Thus, these neurons represent a potential target for understanding the neurobiology of disorders at the intersection of metabolism and psychiatry such as anorexia nervosa.

Network dynamics of hypothalamic feeding neurons.

Mutations in the melanocortin 4 receptor (MC4R) result in hyperphagia and obesity and are the most common cause of monogenic obesity in humans. Preclinical rodent studies have determined that the critical role of the MC4R in controlling feeding can be mapped in part to its expression in the paraventricular nucleus of the hypothalamus (paraventricular nucleus [PVN]), where it regulates the activity of anorexic neural circuits. Despite the critical role of PVN MC4R neurons in regulating feeding, the in vivo neuronal activity of these cells remains largely unstudied, and the network activity of PVN MC4R neurons has not been determined. Here, we utilize in vivo single-cell endomicroscopic and mathematical approaches to determine the activity and network dynamics of PVN MC4R neurons in response to changes in energy state and pharmacological manipulation of central melanocortin receptors. We determine that PVN MC4R neurons exhibit both quantitative and qualitative changes in response to fasting and refeeding. Pharmacological stimulation of MC4R with the therapeutic MC4R agonist setmelanotide rapidly increases basal PVN MC4R activity, while stimulation of melanocortin 3 receptor (MC3R) inhibits PVN MC4R activity. Finally, we find that distinct PVN MC4R neuronal ensembles encode energy deficit and energy surfeit and that energy surfeit is associated with enhanced network connections within PVN MC4R neurons. These findings provide valuable insight into the neural dynamics underlying hunger and energy surfeit.

Comparison of circulating tumor DNA between African American and Caucasian patients with metastatic castrate-resistant prostate cancer post-abiraterone and/or enzalutamide.

BACKGROUND: African American men are much more likely than Caucasian men to be diagnosed with and to die of prostate cancer. Genetic differences likely play a role. The cBioPortal database reveals that African American men with prostate cancer have higher rates of CDK12 somatic mutations compared to Caucasian men. However, this does not account for prior prostate cancer treatments, which are particularly important in the castrate-resistant setting. We aimed to compare somatic mutations based on circulating tumor DNA (ctDNA) in metastatic castration-resistant prostate cancer (mCRPC) between African American and Caucasian men after exposure to abiraterone and/or enzalutamide. METHODS: This single-institution retrospective study characterizes the somatic mutations detected on ctDNA for African American and Caucasian men with mCRPC who had progressed after abiraterone and/or enzalutamide from 2015 through 2022. We evaluated the gene mutations and types of mutations in this mCRPC cohort. RESULTS: There were 50 African American and 200 Caucasian men with CRPC with available ctDNA data. African American men were younger at the time of diagnosis (p = 0.008) and development of castration resistance (p = 0.006). African American men were more likely than Caucasian men to have pathogenic/likely pathogenic (P/LP) mutations in CDK12 (12% vs. 1.5%; p = 0.003) and copy number amplifications and P/LP mutations in KIT (8.0% vs. 1.5%; p = 0.031). African American men were also significantly more likely to have frameshift mutations (28% vs. 14%; p = 0.035). CONCLUSIONS: Compared to Caucasian men, African American men with mCRPC after exposure to abiraterone and/or enzalutamide had a higher incidence of somatic CDK12 P/LP mutations and KIT amplifications and P/LP mutations based on ctDNA. African American men also had more frameshift mutations. We hypothesize that these findings have potential implications for tumor immunogenicity.

Total shoulder arthroplasty in patients with a B2 glenoid addressed with corrective reaming: mean 8-year follow-up.

BACKGROUND: The management of Walch B2 glenoid deformities in primary glenohumeral osteoarthritis is frequently debated. Previous literature has shown that the treatment of B2 glenoids with high-side reaming and anatomic total shoulder arthroplasty (aTSA) perform well in the short-term but is associated with an increased glenoid component failure rate in severe deformities. Therefore, many have explored alternative options, including augmented anatomic glenoid components and reverse shoulder arthroplasty. Our goal in this study is to provide mid-term radiographic and clinical outcomes after high-side reaming and aTSA for B2 glenoids. METHODS: Patients were followed both clinically and radiographically. Preoperative computed tomography scans were used for retrospective analysis of deformity. Both preoperative and postoperative visual analog scale pain, American Shoulder and Elbow Surgeons score, and Simple Shoulder Test scores were collected. Radiographs were analyzed preoperatively and postoperatively for humeral head decentering relative to the glenoid vault, immediate glenoid seating, and final glenoid peg radiolucency. RESULTS: Of the original cohort of 59 patients (6 now deceased) reported at a mean radiographic follow-up of 3.4 years, 34 shoulders in 33 patients with B2 glenoids (mean retroversion 18.9°, range 4°-32°) were available for follow-up at a mean of 8.6 years (range 5.5-11.2) after high-side glenoid reaming with aTSA. Three (5.1%) of the original 59 shoulder were revised. At final follow-up, 3 of 30 (10.0%) shoulders had radiographic glenoid component failure, but were unrevised. Glenoid component failure was associated with worse initial glenoid component seating (mean Lazarus score 1.2 vs. 2.0, P = .002). Glenoid failure was also associated with increased posterior humeral head subluxation at 2-4 year follow-up (mean 5.6% vs. 12.6%, P = .045) and at final follow-up (mean 7.0% vs. 21%, P = .002). There was no association between glenoid component failure and preoperative retroversion, inclination, or humeral head subluxation (all P > .05). Glenoid component failure was associated with worse American Shoulder and Elbow Surgeons (88 vs. 73) and visual analog scale pain (0.8 vs. 2.1) scores (both P = .03). CONCLUSIONS: At a mean of 8.6 years, 88% of shoulders available for follow-up had well-fixed glenoid components. Glenoid component failure was associated with poor initial glenoid component seating, with failed components having an average of 25% of the glenoid component not seated. Preoperative deformity such as glenoid retroversion, inclination, or humeral head subluxation did not predict glenoid component failure. This study supports that initial glenoid component seating and recurrence of posterior humeral head subluxation may be the most important factors for mid-term glenoid component survival in aTSA in patients with B2 glenoids.

Joint Phylogenetic Estimation of Geographic Movements and Biome Shifts during the Global Diversification of Viburnum.

Phylogeny, molecular sequences, fossils, biogeography, and biome occupancy are all lines of evidence that reflect the singular evolutionary history of a clade, but they are most often studied separately, by first inferring a fossil-dated molecular phylogeny, then mapping on ancestral ranges and biomes inferred from extant species. Here we jointly model the evolution of biogeographic ranges, biome affinities, and molecular sequences, while incorporating fossils to estimate a dated phylogeny for all of the 163 extant species of the woody plant clade Viburnum (Adoxaceae) that we currently recognize in our ongoing worldwide monographic treatment of the group. Our analyses indicate that while the major Viburnum lineages evolved in the Eocene, the majority of extant species originated since the Miocene. Viburnum radiated first in Asia, in warm, broad-leaved evergreen (lucidophyllous) forests. Within Asia, we infer several early shifts into more tropical forests, and multiple shifts into forests that experience prolonged freezing. From Asia, we infer two early movements into the New World. These two lineages probably first occupied warm temperate forests and adapted later to spreading cold climates. One of these lineages (Porphyrotinus) occupied cloud forests and moved south through the mountains of the Neotropics. Several other movements into North America took place more recently, facilitated by prior adaptations to freezing in the Old World. We also infer four disjunctions between Asia and Europe: the Tinus lineage is the oldest and probably occupied warm forests when it spread, whereas the other three were more recent and in cold-adapted lineages. These results variously contradict published accounts, especially the view that Viburnum radiated initially in cold forests and, accordingly, maintained vessel elements with scalariform perforations. We explored how the location and biome assignments of fossils affected our inference of ancestral areas and biome states. Our results are sensitive to, but not entirely dependent upon, the inclusion of fossil biome data. It will be critical to take advantage of all available lines of evidence to decipher events in the distant past. The joint estimation approach developed here provides cautious hope even when fossil evidence is limited. [Biogeography; biome; combined evidence; fossil pollen; phylogeny; Viburnum.].

Imbalance in Axial-plane Rotator Cuff Fatty Infiltration in Posteriorly Worn Glenoids in Primary Glenohumeral Osteoarthritis: An MRI-based Study.

BACKGROUND: Fatty infiltration of the rotator cuff evaluated with CT has been associated with asymmetric glenoid wear and humeral head subluxation in patients with glenohumeral arthritis. The relationship between rotator cuff pathologic findings and abnormal glenoid wear plays an important role in determining the optimal surgical management of advanced glenohumeral osteoarthritis. Compared with CT, MRI has increased sensitivity for identifying rotator cuff conditions; therefore, prior studies using CT may have underestimated the association between fatty infiltration of the rotator cuff and abnormal glenoid wear. QUESTIONS/PURPOSES: (1) Compared with Type A glenoids, which muscles in which Walch subtypes have a greater degree of fatty infiltration using Goutallier scores? (2) What glenoid type is associated with greater imbalance in fatty infiltration, as measured by comparing Goutallier scores between the posterior and anterior rotator cuff muscles? (3) What is the correlation between glenoid version and fatty infiltration of the rotator cuff muscles? (4) Comparing Type B2 and B3 glenoids with Type A glenoids, after accounting for age and sex, is there an increase in fatty infiltration of the infraspinatus muscle? METHODS: A total of 129 shoulders from 129 patients undergoing anatomic total shoulder arthroplasty to treat primary glenohumeral osteoarthritis were retrospectively reviewed. Patients had an average age of 66.4 ± 9.3 years and an average BMI of 30.6 ± 6.7 kg/m2, and 53% (69 of 129) were men. All patients underwent MRI within 12 months before total shoulder arthroplasty to assess glenoid morphology and rotator cuff pathologic findings. Three reviewers assessed the images, and glenoid morphology was assigned using the modified Walch classification system (Types A1, A2, B1, B2, B3, C, and D). Fatty infiltration of the rotator cuff was classified using Goutallier scores. The examiners demonstrated moderate-to-good reliability using these classification systems; the Walch classification system had interrater reliability kappa coefficients (κ) from 0.54 to 0.69 and intrarater reliability κ from 0.60 to 0.64. Goutallier scores using the simplified classification system had interrater reliability κ from 0.64 to 0.68 and intrarater reliability κ from 0.64 to 0.79. Thirty-six percent (46 of 129) of the shoulders had posterior wear patterns (18% [23] were Type B2 glenoids; 18% [23] were Type B3 glenoids). The average Goutallier scores for each rotator cuff muscle were determined, and the amount of fatty infiltration was compared between the various Walch subtypes using independent t-tests. Axial-plane imbalance in fatty infiltration of the rotator cuff was assessed by determining the difference in the average fatty infiltration of the posterior rotator cuff muscles (infraspinatus and teres minor) and anterior rotator cuff muscles (subscapularis) and comparing the differences among the Walch subtypes using independent t-tests. The association between glenoid version and fatty infiltration was assessed using Pearson correlations. Finally, a multivariate logistic regression model was used to assess fatty infiltration of the rotator cuff among the various Walch subtypes while accounting for patient age and sex. RESULTS: Compared with Type A1 glenoids, Type B2 and B3 glenoids had an increased amount of fatty infiltration of the infraspinatus (1.6 ± 0.7 versus 0.7 ± 0.4; mean difference 0.9 [95% CI 0.7-1.2]; p < 0.001 and 1.8 ± 0.4 versus 0.7 ± 0.4; mean difference 1.1 [95% CI 0.9-1.4]; p < 0.001, respectively) and teres minor (1.3 ± 0.7 versus 0.6 ± 0.5; mean difference 0.7 [95% CI 0.4-1.0]; p < 0.001 and 1.6 ± 0.6 versus 0.6 ± 0.5; mean difference 1.0 [95% CI 0.7-1.2]; p < 0.001, respectively). There was greater imbalance in fatty infiltration between the posterior and anterior rotator cuff muscles for Type B2 (0.5 ± 0.3) and B3 (0.6 ± 0.5) glenoids than for Type A1 (0.1 ± 0.3) and A2 (0.1 ± 0.6) glenoids (p < 0.001). Only the infraspinatus's fatty infiltration was strongly correlated with glenoid version (r = 0.64; p < 0.001), while fatty infiltration of the other muscles only correlated weakly or moderately. After accounting for age and sex, fatty infiltration in the infraspinatus was associated with Type B2 (OR 66.1 [95% CI 7.6-577.9]; p < 0.001) and Type B3 glenoids (OR 59.5 [95% CI 5.4-661.3]; p < 0.001) compared with Type A glenoids. CONCLUSION: Compared with concentric wear, posteriorly worn glenoids had an imbalance in axial-plane rotator cuff fatty infiltration and an increased amount of fatty infiltration of the infraspinatus and teres minor compared with the subscapularis. These imbalances may contribute to the higher rates of failure after anatomic total shoulder arthroplasty in patients with posterior wear compared with those with concentric wear. Future research should be directed toward investigating the temporal relationship of these findings, as well as understanding the clinical outcomes for patients undergoing anatomic total shoulder arthroplasty who have posteriorly worn glenoids with a high degree of fatty infiltration of the posterior rotator cuff musculature. CLINICAL RELEVANCE: Providers should consider the increased likelihood of higher-grade fatty infiltration of the posterior rotator cuff in the setting of posteriorly worn glenoids, particularly when treating patients without using MRI. These patients have higher rates of failure postoperatively and may benefit from closer monitoring and altered postoperative rehabilitation protocols that target the posterior rotator cuff.

Machine Learning Using Digitized Herbarium Specimens to Advance Phenological Research.

Machine learning (ML) has great potential to drive scientific discovery by harvesting data from images of herbarium specimens-preserved plant material curated in natural history collections-but ML techniques have only recently been applied to this rich resource. ML has particularly strong prospects for the study of plant phenological events such as growth and reproduction. As a major indicator of climate change, driver of ecological processes, and critical determinant of plant fitness, plant phenology is an important frontier for the application of ML techniques for science and society. In the present article, we describe a generalized, modular ML workflow for extracting phenological data from images of herbarium specimens, and we discuss the advantages, limitations, and potential future improvements of this workflow. Strategic research and investment in specimen-based ML methods, along with the aggregation of herbarium specimen data, may give rise to a better understanding of life on Earth.

Restriction-Site-Associated DNA Sequencing Reveals a Cryptic Viburnum Species on the North American Coastal Plain.

Species are the starting point for most studies of ecology and evolution, but the proper circumscription of species can be extremely difficult in morphologically variable lineages, and there are still few convincing examples of molecularly informed species delimitation in plants. Here, we focus on the Viburnum nudum complex, a highly variable clade that is widely distributed in eastern North America. Taxonomic treatments have mostly divided this complex into northern (V. nudum var. cassinoides) and southern (V. nudum var. nudum) entities, but additional names have been proposed. We used multiple lines of evidence, including RADseq, morphological, and geographic data, to test how many independently evolving lineages exist within the V. nudum complex. Genetic clustering and phylogenetic methods revealed three distinct groups-one lineage that is highly divergent, and two others that are recently diverged and morphologically similar. A combination of evidence that includes reciprocal monophyly, lack of introgression, and discrete rather than continuous patterns of variation supports the recognition of all three lineages as separate species. These results identify a surprising case of cryptic diversity in which two broadly sympatric species have consistently been lumped in taxonomic treatments. The clarity of our findings is directly related to the dense sampling and high-quality genetic data in this study. We argue that there is a critical need for carefully sampled and integrative species delimitation studies to clarify species boundaries even in well-known plant lineages. Studies following the model that we have developed here are likely to identify many more cryptic lineages and will fundamentally improve our understanding of plant speciation and patterns of species richness.

Appetite suppressive role of medial septal glutamatergic neurons.

Feeding behavior is controlled by diverse neurons and neural circuits primarily concentrated in the hypothalamus and hindbrain in mammals. In this study, by using chemo/optogenetic techniques along with feeding assays, we investigate how neurons within the medial septal complex (MSc), a brain area implicated in emotion and cognition, contribute to food intake. We find that chemo/optogenetic activation of MSc glutamatergic neurons profoundly reduces food intake during both light and dark periods of the rodent light cycle. Furthermore, we find that selective activation of MSc glutamatergic projections in paraventricular hypothalamus (PVH) reduces food intake, suggesting that MSc glutamatergic neurons suppress feeding by activating downstream neurons in the PVH. Open-field behavioral assays reveal that these neurons do not overtly affect anxiety levels and locomotion. Collectively, our findings demonstrate that septal glutamatergic neurons exert anorexigenic effects by projecting to the PVH without affecting anxiety and physical activities.

Assessment of Flexion Strength Following Single- Versus Double-Hamstring Tendon Harvest for Anterior Cruciate Ligament Reconstruction.

PURPOSE: To compare isometric hamstring strength deficits, knee laxity, functional outcomes, and patient-reported outcomes between patients who underwent anterior cruciate ligament (ACL) reconstruction with doubled semitendinosus and gracilis tendon autograft (ST/G) versus quadrupled semitendinosus autograft (ST), at a minimum follow-up of 1-year postoperatively. METHODS: Patients who underwent ACL reconstruction with ST/G or ST hamstring autografts were retrospectively identified. Isometric hamstring strength was tested with a hand-held dynamometer at 30, 60, and 90° of knee flexion. Anterior knee laxity was assessed using a KT-1000 arthrometer. Functional outcomes were collected using the single-leg hop test and single-leg squat test. Side-to-side differences were determined and compared between the ST/G and ST groups. Patient-reported outcomes were collected on all patients. RESULTS: Eighty-four patients who underwent ST/G (n = 34) or ST (n = 50) autograft ACL reconstruction were recruited to participate in this study. There was no difference in knee laxity between the groups. Side-to-side hamstring strength deficits increased with increased flexion angles. At 90° of flexion, the ST/G group had a significantly greater flexion strength deficit compared with the ST group (37.8 ± 15.1% vs 24.7 ± 12.5%, P < .001). Aside from a significant difference in the KOOS pain Score (P .045), no other significant differences in functional or patient reported outcomes between the groups were identified. CONCLUSIONS: Patients who underwent ACL reconstruction with ST/G compared with ST autograft have a significantly greater isometric flexion strength deficit at 90° of flexion. Future investigations are required to determine the clinical relevance of this difference and whether specialized therapy protocols can mitigate this deficit. LEVEL OF EVIDENCE: Level III, retrospective comparative study.

Sterile marginal flowers increase visitation and fruit set in the hobblebush (Viburnum lantanoides, Adoxaceae) at multiple spatial scales.

Background and Aims: Enlarged sterile flowers on the periphery of inflorescences increase the attractiveness of floral displays, and previous studies have generally demonstrated that these have positive effects on insect visitation and/or reproductive success. However, experiments have not specifically been designed to examine the benefits of sterile flowers under conditions that reflect the early stages in their evolution, i.e. when plants that produce sterile flowers are at low frequency. Methods: Over three years, three experiments were performed in natural populations of Viburnum lantanoides, which produces sterile marginal flowers (SMFs). The first experiment established that fruit production in V. lantanoides increases with the receipt of outcross pollen. The second tested the role of SMFs under extant conditions, comparing fruit production in two populations composed entirely of intact plants or entirely of plants with the SMFs removed. The third was designed to mimic the presumed context in which SMFs first evolved; here, SMFs were removed from all but a few plants in a population, and rates of insect visitation and fruit set were compared between plants with intact and denuded SMFs. Key Results: In comparing whole populations, the presence of SMFs nearly doubled fruit set. Under simulated 'ancestral' conditions within a population, plants with intact SMFs received double the insect visits and produced significantly more fruits than denuded plants. There was no significant effect of the number of inflorescences or fertile flowers on insect visitation or fruit set, indicating that the presence of SMFs accounted for these differences. Conclusions: The presence of SMFs significantly increased pollinator attraction and female reproductive success both in contemporary and simulated ancestral contexts, indicating that stabilizing selection is responsible for their maintenance, and directional selection likely drove their evolution when they first appeared. This study demonstrates a novel approach to incorporating historically relevant scenarios into experimental studies of floral evolution.

Differences in flowering time maintain species boundaries in a continental radiation of Viburnum.

PREMISE: We take an integrative approach in assessing how introgression and Pleistocene climate fluctuations have shaped the diversification of the core Lentago clade of Viburnum, a group of five interfertile species with broad areas of sympatry. We specifically tested whether flowering time plays a role in maintaining species isolation. METHODS: RAD-seq data for 103 individuals were used to infer the species relationships and the genetic structure within each species. Flowering times were compared among species on the basis of historical flowering dates documented by herbarium specimens. RESULTS: Within each species, we found a strong relationship between flowering date and latitude, such that southern populations flower earlier than northern ones. In areas of sympatry, the species flower in sequence rather than simultaneously, with flowering dates offset by ≥9 d for all species pairs. In two cases it appears that the offset in flowering times is an incidental consequence of adaptation to differing climates, but in the recently diverged sister species V. prunifolium and V. rufidulum, we find evidence that reinforcement led to reproductive character displacement. Long-term trends suggest that the two northern-most species are flowering earlier in response to recent climate change. CONCLUSIONS: We argue that speciation in the Lentago clade has primarily occurred through ecological divergence of allopatric populations, but differences in flowering time were essential to maintain separation of incipient species when they came into secondary contact. This combination of factors may underlie diversification in many other plant clades.

Widespread sampling biases in herbaria revealed from large-scale digitization.

Nonrandom collecting practices may bias conclusions drawn from analyses of herbarium records. Recent efforts to fully digitize and mobilize regional floras online offer a timely opportunity to assess commonalities and differences in herbarium sampling biases. We determined spatial, temporal, trait, phylogenetic, and collector biases in c. 5 million herbarium records, representing three of the most complete digitized floras of the world: Australia (AU), South Africa (SA), and New England, USA (NE). We identified numerous shared and unique biases among these regions. Shared biases included specimens collected close to roads and herbaria; specimens collected more frequently during biological spring and summer; specimens of threatened species collected less frequently; and specimens of close relatives collected in similar numbers. Regional differences included overrepresentation of graminoids in SA and AU and of annuals in AU; and peak collection during the 1910s in NE, 1980s in SA, and 1990s in AU. Finally, in all regions, a disproportionately large percentage of specimens were collected by very few individuals. We hypothesize that these mega-collectors, with their associated preferences and idiosyncrasies, shaped patterns of collection bias via 'founder effects'. Studies using herbarium collections should account for sampling biases, and future collecting efforts should avoid compounding these biases to the extent possible.

Worldwide Engagement for Digitizing Biocollections (WeDigBio): The Biocollections Community's Citizen-Science Space on the Calendar.

The digitization of biocollections is a critical task with direct implications for the global community who use the data for research and education. Recent innovations to involve citizen scientists in digitization increase awareness of the value of biodiversity specimens; advance science, technology, engineering, and math literacy; and build sustainability for digitization. In support of these activities, we launched the first global citizen-science event focused on the digitization of biodiversity specimens: Worldwide Engagement for Digitizing Biocollections (WeDigBio). During the inaugural 2015 event, 21 sites hosted events where citizen scientists transcribed specimen labels via online platforms (DigiVol, Les Herbonautes, Notes from Nature, the Smithsonian Institution's Transcription Center, and Symbiota). Many citizen scientists also contributed off-site. In total, thousands of citizen scientists around the world completed over 50,000 transcription tasks. Here, we present the process of organizing an international citizen-science event, an analysis of the event's effectiveness, and future directions-content now foundational to the growing WeDigBio event.

An Inhibitory Septum to Lateral Hypothalamus Circuit That Suppresses Feeding.

Feeding behavior is orchestrated by neural circuits primarily residing in the hypothalamus and hindbrain. However, the relative influence of cognitive and emotional brain circuits to the feeding circuitry in the hypothalamus and hindbrain remains unclear. Here, using the cell-type selectivity of genetic methods, circuit mapping, and behavior assays, we sought to decipher neural circuits emanating from the septal nucleus to the lateral hypothalamus (LH) that contribute to neural regulation of food intake in mice. We found that chemogenetic and optogenetic activation of septal vesicular GABA transporter (vGAT)-containing neurons or their projections in the LH reduced food intake in mice. Consistently, chemogenetic inhibition of septal vGAT neurons increased food intake. Furthermore, we investigated a previously unknown neural circuit originating from septal vGAT neurons to a subset of vGAT neurons in the LH, an area involved in homeostatic and hedonic control of energy states. Collectively, our data reveal an inhibitory septohypothalamic feeding circuit that might serve as a therapeutic target for the treatment of eating disorders such as anorexia nervosa. SIGNIFICANCE STATEMENT: Our results demonstrate that top-down projections from the septum to the hypothalamus control food intake negatively. Given the known role for both of these brain regions in the control of feeding and emotion-related behaviors, these findings reveal previously unknown neural circuitry that is likely implicated in emotional aspects of food intake and provide new insights into the development of therapeutic targets for the treatment of eating disorders.

Activation of hypothalamic astrocytes suppresses feeding without altering emotional states.

Emerging evidence shows that hypothalamic astrocytes react to and counteract energy surfeit produced by high-fat diet (HFD) feeding. However, the functional role of astrocytes in the control of energy states and the underlying molecular mechanism(s) during physiological conditions remain largely underexplored. In the present study, by taking advantage of spatiotemporally precise optogenetic approaches, real-time measurements of extracellular adenosine, and behavioral assays, we find that optogenetic stimulation of astrocytes localized in the medial basal hypothalamus (MBH) suppresses food intake in a frequency dependent manner with high frequency, but not low frequency, stimulation of astrocytes reducing food intake. Furthermore, stimulation of MBH astrocytes diminishes orexigenic ghrelin or fasting-induced hyperphagia without effecting anxiety-related behavior. Consistent with a frequency dependent role for MBH astrocytes in feeding behavior, optogenetic stimulation of MBH astrocytes increases extracellular levels of adenosine in a frequency dependent manner. Collectively, our results provide new insights into the role of astrocytes in physiological functions during naturally occurring behaviors, such as feeding. GLIA 2016;64:2263-2273.

Caffeine increases food intake while reducing anxiety-related behaviors.

The objective of this study was to determine the effects of different doses of caffeine on appetite and anxiety-related behavior. Additionally, we sought to determine if withdrawal from chronic caffeine administration promotes anxiety. In this study, we utilized rodent open field testing and feeding behavior assays to determine the effects of caffeine on feeding and anxiety-related behavior (n = 8 mice; 4-8 weeks old). We also measured 2 h and 24 h food intake and body-weight during daily administration of caffeine (n = 12 mice; 4-8 weeks old). To test for caffeine withdrawal induced anxiety, anxiety-related behavior in rodents was quantified following withdrawal from four consecutive days of caffeine administration (n = 12 mice; 4-8 weeks old). We find that acute caffeine administration increases food intake in a dose-dependent manner with lower doses of caffeine more significantly increasing food intake than higher doses. Acute caffeine administration also reduced anxiety-related behaviors in mice without significantly altering locomotor activity. However, we did not observe any differences in 24 h food intake or body weight following chronic caffeine administration and there were no observable differences in anxiety-related behaviors during caffeine withdrawal. In conclusion, we find that caffeine can both increase appetite and decrease anxiety-related behaviors in a dose dependent fashion. Given the complex relationship between appetite and anxiety, the present study provides additional insights into potential caffeine-based pharmacological mechanisms governing appetite and anxiety disorders, such as bulimia nervosa.

Temperate radiations and dying embers of a tropical past: the diversification of Viburnum.

We used a near-complete phylogeny for the angiosperm clade Viburnum to assess lineage diversification rates, and to examine possible morphological and ecological factors driving radiations. Maximum-likelihood and Bayesian approaches identified shifts in diversification rate and possible links to character evolution. We inferred the ancestral environment for Viburnum and changes in diversification dynamics associated with subsequent biome shifts. Viburnum probably diversified in tropical forests of Southeast Asia in the Eocene, with three subsequent radiations in temperate clades during the Miocene. Four traits (purple fruits, extrafloral nectaries, bud scales and toothed leaves) were statistically associated with higher rates of diversification. However, we argue that these traits are unlikely to be driving diversification directly. Instead, two radiations were associated with the occupation of mountainous regions and a third with repeated shifts between colder and warmer temperate forests. Early-branching depauperate lineages imply that the rare lowland tropical species are 'dying embers' of once more diverse lineages; net diversification rates in Viburnum likely decreased in these tropical environments after the Oligocene. We suggest that 'taxon pulse' dynamics might characterize other temperate plant lineages.

A chloroplast tree for Viburnum (Adoxaceae) and its implications for phylogenetic classification and character evolution.

• Premise of the study: Despite recent progress, significant uncertainties remain concerning relationships among early-branching lineages within Viburnum (Adoxaceae), prohibiting a new classification and hindering studies of character evolution and the increasing use of Viburnum in addressing a wide range of ecological and evolutionary questions. We hoped to resolve these issues by sequencing whole plastid genomes for representative species and combining these with molecular data previously obtained from an expanded taxon sample.• Methods: We performed paired-end Illumina sequencing of plastid genomes of 22 Viburnum species and combined these data with a 10-gene data set to infer phylogenetic relationships for 113 species. We used the results to devise a comprehensive phylogenetic classification and to analyze the evolution of eight morphological characters that vary among early-branching lineages.• Key results: With greatly increased levels of confidence in most of the early branches, we propose a phylogenetic classification of Viburnum, providing formal phylogenetic definitions for 30 clades, including 13 with names recognized under the International Code of Nomenclature for Algae, Fungi, and Plants, eight with previously proposed informal names, and nine newly proposed names for major branches. Our parsimony reconstructions of bud structure, leaf margins, inflorescence form, ruminate endosperm, extrafloral nectaries, glandular trichomes, palisade anatomy, and pollen exine showed varying levels of homoplasy, but collectively provided morphological support for some, though not all, of the major clades.• Conclusions: Our study demonstrates the value of next-generation plastid sequencing, the ease of creating a formal phylogenetic classification, and the utility of such a system in describing patterns of character evolution.