Evolution and resurrection ecology of a foundational coastal marsh plant

Stratified storage of dormant seeds in soil can result in natural archives useful for studying evolutionary responses to environmental change. However, few studies leverage soil-stored seed banks as natural archives, in part because of concerns over attrition, bias, and sediment mixing. Here, I examine the persistent seed bank of Schoenoplectus americanus, a foundational brackish marsh sedge, to a) determine whether it can serve as a resource for reconstructing demographic and population genetic/genomic variation, b) whether and how evolution may be occurring across a century. After extracting seeds from radionuclide-dated soil cores taken across the Chesapeake Bay, I “resurrected” age cohorts spanning the 20th century. In Chapter 1, I use microsatellites to assess genetic diversity/differentiation among age cohorts, drawing comparisons to extant plants at the study site and to extant plants in nearby and more distant marshes. I found genotypic differences among cohorts and between cohorts and extant plants. Genetic diversity did not decline with depth, suggesting differentiation is likely not due to attrition. In Chapter 2, I use SNPs to examine population diversity/differentiation for resurrected plants taken from multiple marshes to understand how regional-scale geography interacts with temporal change. I found that location explained genetic clustering better than temporal differences, suggesting that habitat differences between marshes are consequential for S. americanus evolution. In Chapter 3, I deployed 2 resurrected age cohorts from one marsh in a greenhouse experiment to assess phenotypic differences. I cloned plants across triply-crossed conditions: simulated sea level rise (salinity, inundation) and competition. Biomass by treatment did not significantly differ when averaging by cohort. However, variance was smaller for young versus old cohorts, suggesting reduction in phenotypic plasticity across time. I also compared gene expression differences in response to salinity for stem/root tissue and between cohorts. I found that salinity treatment resulted in significantly different expression levels and some evidence of differentiation by age cohort, but only for root tissue. Overall, this work describes complex, geographically variable, and small evolutionary shifts across time in S americanus. This suggests that local population and/or habitat differences mitigate change through time, which may be in response to climate change corollaries

pH-Dependent Metal Ion Toxicity Influences the Antibacterial Activity of Two Natural Mineral Mixtures

Recent studies have demonstrated that several mineral products sold for medicinal purposes demonstrate antimicrobial activity, but little is known about the physicochemical properties involved in antibacterial activity. or other sequestered metal cations, in mineral toxicity. This conclusion was supported by NMR relaxation data, which indicated that BY07 and CB07 leachates contained higher concentrations of chemically accessible metal ions than leachates from non-bactericidal mineral samples.We conclude that the acidic environment of the hydrated minerals significantly contributes to antibacterial activity by increasing the availability and toxicity of metal ions. These findings provide impetus for further investigation of the physiological effects of mineral products and their applications in complementary antibacterial therapies

Relationship Between Foveal Cone Specialization and Pit Morphology in Albinism

Purpose.Albinism is associated with disrupted foveal development, though intersubject variability is becoming appreciated. We sought to quantify this variability, and examine the relationship between foveal cone specialization and pit morphology in patients with a clinical diagnosis of albinism. Methods. We recruited 32 subjects with a clinical diagnosis of albinism. DNA was obtained from 25 subjects, and known albinism genes were analyzed for mutations. Relative inner and outer segment (IS and OS) lengthening (fovea-to-perifovea ratio) was determined from manually segmented spectral domain-optical coherence tomography (SD-OCT) B-scans. Foveal pit morphology was quantified for eight subjects from macular SD-OCT volumes. Ten subjects underwent imaging with adaptive optics scanning light ophthalmoscopy (AOSLO), and cone density was measured. Results. We found mutations in 22 of 25 subjects, including five novel mutations. All subjects lacked complete excavation of inner retinal layers at the fovea, though four subjects had foveal pits with normal diameter and/or volume. Peak cone density and OS lengthening were variable and overlapped with that observed in normal controls. A fifth hyper-reflective band was observed in the outer retina on SD-OCT in the majority of the subjects with albinism. Conclusions. Foveal cone specialization and pit morphology vary greatly in albinism. Normal cone packing was observed in the absence of a foveal pit, suggesting a pit is not required for packing to occur. The degree to which retinal anatomy correlates with genotype or visual function remains unclear, and future examination of larger patient groups will provide important insight on this issue

Effects of administration of a growth promoting implant during the suckling phase or at weaning on growth, reproduction, and ovarian development in replacement heifers grazing native range

Management strategies utilized during pre-breeding development of replacement heifers can impact fertility and the ovarian reserve. Angus- Hereford crossbred heifers (n = 233) were utilized over a 3-yr period to determine the effects of administration of a growth promoting implant at either branding or weaning on growth, reproduction, and ovarian development. Heifer calves were randomly assigned to one of three treatments: 1) nonimplanted controls (CON; n = 79), 2) implanted at approximately 2 mo of age (average calf age = 58 d) with Synovex-C (BIMP, n = 82), or 3) implanted at approximately 7 mo of age (average calf age = 210 d) with Synovex-C (WIMP; n = 72). In years 2 and 3, a subset of heifers (year 2 n = 16; year 3 n = 14) were unilaterally ovariectomized. Heifers implanted at 2 mo of age were heavier at weaning, yearling (mid-February; average calf age = 332 d), and at the beginning of the breeding season (P \u3c 0.01) compared to CON and WIMP heifers. Average daily gain (ADG) was similar among treatments from weaning to yearling and weaning to the start of the breeding season (P ≥ 0.61); however, WIMP heifers had increased (P = 0.05) ADG from yearling to the start of the breeding season compared to BIMP heifers. Antral follicle count and reproductive tract scores were not influenced by implant treatment (P ≥ 0.18). Response to synchronization of estrus was increased (P = 0.02) in WIMP compared to CON heifers, with BIMP heifers similar to all other treatments. First service conception rates tended to be increased (P = 0.09) in CON heifers compared to WIMP heifers, with BIMP heifers similar to CON and WIMP. Final pregnancy rates were similar (P = 0.54) among treatments. A treatment × yr interaction was detected (P = 0.01) for the number of primordial follicles/section with increased primordial follicles in WIMP heifers in year 3 compared to BIMP and WIMP heifers in year 2 and CON heifers in year 3, as well as in BIMP compared to WIMP heifers in year 2. Utilization of growth promoting implants did not negatively impact postweaning reproductive development or compromise pregnancy rates in beef heifers. Based on these results, administration of a growth promoting Synovex-C implant at 2 mo of age may allow for increased body weight at weaning, without hindering reproductive performance

Expression of lymphotoxin-αβ on antigen-specific T cells is required for DC function

During an immune response, activated antigen (Ag)-specific T cells condition dendritic cells (DCs) to enhance DC function and survival within the inflamed draining lymph node (LN). It has been difficult to ascertain the role of the tumor necrosis factor (TNF) superfamily member lymphotoxin-αβ (LTαβ) in this process because signaling through the LTβ-receptor (LTβR) controls multiple aspects of lymphoid tissue organization. To resolve this, we have used an in vivo system where the expression of TNF family ligands is manipulated only on the Ag-specific T cells that interact with and condition Ag-bearing DCs. We report that LTαβ is a critical participant required for optimal DC function, independent of its described role in maintaining lymphoid tissue organization. In the absence of LTαβ or CD40L on Ag-specific T cells, DC dysfunction could be rescued in vivo via CD40 or LTβR stimulation, respectively, suggesting that these two pathways cooperate for optimal DC conditioning

Prescribing practices of primary-care veterinary practitioners in dogs diagnosed with bacterial pyoderma

Concern has been raised regarding the potential contributions of veterinary antimicrobial use to increasing levels of resistance in bacteria critically important to human health. Canine pyoderma is a frequent, often recurrent diagnosis in pet dogs, usually attributable to secondary bacterial infection of the skin. Lesions can range in severity based on the location, total area and depth of tissue affected and antimicrobial therapy is recommended for resolution. This study aimed to describe patient signalment, disease characteristics and treatment prescribed in a large number of UK, primary-care canine pyoderma cases and to estimate pyoderma prevalence in the UK vet-visiting canine population

Accounting for variability when resurrecting dormant propagules substantiates their use in eco-evolutionary studies

There has been a steady rise in the use of dormant propagules to study biotic responses to environmental change over time. This is particularly important for organisms that strongly mediate ecosystem processes, as changes in their traits over time can provide a unique snapshot into the structure and function of ecosystems from decades to millennia in the past. Understanding sources of bias and variation is a challenge in the field of resurrection ecology, including those that arise because often-used measurements like seed germination success are imperfect indicators of propagule viability. Using a Bayesian statistical framework, we evaluated sources of variability and tested for zero-inflation and overdispersion in data from 13 germination trials of soil-stored seeds of Schoenoplectus americanus, an ecosystem engineer in coastal salt marshes in the Chesapeake Bay. We hypothesized that these two model structures align with an ecological understanding of dormancy and revival: zero-inflation could arise due to failed germinations resulting from inviability or failed attempts to break dormancy, and overdispersion could arise by failing to measure important seed traits. A model that accounted for overdispersion, but not zero-inflation, was the best fit to our data. Tetrazolium viability tests corroborated this result: most seeds that failed to germinate did so because they were inviable, not because experimental methods failed to break their dormancy. Seed viability declined exponentially with seed age and was mediated by seed provenance and experimental conditions. Our results provide a framework for accounting for and explaining variability when estimating propagule viability from soil-stored natural archives which is a key aspect of using dormant propagules in eco-evolutionary studies

Caffeine Junkie: an Unprecedented Glutathione S-Transferase- Dependent Oxygenase Required for Caffeine Degradation by Pseudomonas putida CBB5 Downloaded from

c Caffeine and other N-methylated xanthines are natural products found in many foods, beverages, and pharmaceuticals. Therefore, it is not surprising that bacteria have evolved to live on caffeine as a sole carbon and nitrogen source. The caffeine degradation pathway of Pseudomonas putida CBB5 utilizes an unprecedented glutathione-S-transferase-dependent Rieske oxygenase for demethylation of 7-methylxanthine to xanthine, the final step in caffeine N-demethylation. The gene coding this function is unusual, in that the iron-sulfur and non-heme iron domains that compose the normally functional Rieske oxygenase (RO) are encoded by separate proteins. The non-heme iron domain is located in the monooxygenase, ndmC, while the Rieske [2Fe-2S] domain is fused to the RO reductase gene, ndmD. This fusion, however, does not interfere with the interaction of the reductase with N 1 -and N 3 -demethylase RO oxygenases, which are involved in the initial reactions of caffeine degradation. We demonstrate that the N 7 -demethylation reaction absolutely requires a unique, tightly bound protein complex composed of NdmC, NdmD, and NdmE, a novel glutathione-S-transferase (GST). NdmE is proposed to function as a noncatalytic subunit that serves a structural role in the complexation of the oxygenase (NdmC) and Rieske domains (NdmD). Genome analyses found this gene organization of a split RO and GST gene cluster to occur more broadly, implying a larger function for RO-GST protein partners. C affeine (1,3,7-trimethylxanthine) and other N-methylated xanthines are well known for applications in food and as pharmaceuticals that improve lung function for asthmatics and chronic obstructive pulmonary disease (COPD) sufferers. More recently, these compounds have been investigated for use as natural insecticides and in treatments for cancer, septic shock, and functional neutrophil disorders (1-3). Enzymatic methods for producing and degrading these N-methylated xanthines could have broader applications for health through both biosynthesis and environmental remediation of waste and by-products. Therefore, bacteria that have evolved to live on caffeine as the sole carbon and nitrogen source are of interest, as are their metabolic pathways toward N-methylated xanthines. Pseudomonas putida CBB5 degrades caffeine, theophylline (1,3-dimethylxanthine), and related methylxanthines via sequential N-demethylation to xanthine (4-6). The ordered N-demethylation of caffeine to xanthine occurs in three steps catalyzed by enzymes belonging to the Rieske oxygenase (RO) family (5, 6), which are encoded by the Alx operon. Initially, two Rieske, nonheme Fe(II) monooxygenases, NdmA and NdmB, remove the N 1 -and N 3 -methyl groups, respectively, from caffeine to form 7-methylxanthine. Both enzymes require an unusually large 65-kDa redox-dense RO reductase, NdmD, which transfers electrons from NADH to NdmA and NdmB for oxygen activation. The final step in the caffeine degradation pathway is N 7 -demethylation of 7-methylxanthine to xanthine. This N 7 -demethylation activity was inseparable from NdmD after four chromatographic steps (6). A highly enriched protein fraction containing this activity was comprised of NdmD and two additional major protein bands, as visualized by SDS-PAGE. These two additional peptides are encoded by two genes in the Alx operon, labeled orf7 and orf8, which flank ndmD on the CBB5 genome Here, we report that ndmE encodes a new type of GST that is absolutely required for N 7 -demethylation of 7-methylxanthine, the final step of caffeine degradation in P. putida CBB5. The N 7 -demethylase RO is unusual in itself because the iron-sulfur and non-heme iron domains that compose the normally functional oxygenase are encoded by two separate genes. The non-heme iron is contained in NdmC, while the iron-sulfur domain is fused to NdmD. NdmE is proposed to facilitate the formation of the NdmCDE complex, which catalyzes the N 7 -demethylation. This is the first report of a new class of GST-dependent ROs. Additional identification of similar uncharacterized gene clusters within genome databases suggests that there is a more generalized role for GSTs in oxygenation and/or biodegradation

A large annotated medical image dataset for the development and evaluation of segmentation algorithms

Semantic segmentation of medical images aims to associate a pixel with a label in a medical image without human initialization. The success of semantic segmentation algorithms is contingent on the availability of high-quality imaging data with corresponding labels provided by experts. We sought to create a large collection of annotated medical image datasets of various clinically relevant anatomies available under open source license to facilitate the development of semantic segmentation algorithms. Such a resource would allow: 1) objective assessment of general-purpose segmentation methods through comprehensive benchmarking and 2) open and free access to medical image data for any researcher interested in the problem domain. Through a multi-institutional effort, we generated a large, curated dataset representative of several highly variable segmentation tasks that was used in a crowd-sourced challenge - the Medical Segmentation Decathlon held during the 2018 Medical Image Computing and Computer Aided Interventions Conference in Granada, Spain. Here, we describe these ten labeled image datasets so that these data may be effectively reused by the research community

Healthcare recommendations for Joubert syndrome

Joubert syndrome (JS) is a recessive neurodevelopmental disorder defined by a characteristic cerebellar and brainstem malformation recognizable on axial brain magnetic resonance imaging as the "Molar Tooth Sign". Although defined by the neurological features, JS is associated with clinical features affecting many other organ systems, particularly progressive involvement of the retina, kidney, and liver. JS is a rare condition; therefore, many affected individuals may not have easy access to subspecialty providers familiar with JS (e.g., geneticists, neurologists, developmental pediatricians, ophthalmologists, nephrologists, hepatologists, psychiatrists, therapists, and educators). Expert recommendations can enable practitioners of all types to provide quality care to individuals with JS and know when to refer for subspecialty care. This need will only increase as precision treatments targeting specific genetic causes of JS emerge. The goal of these recommendations is to provide a resource for general practitioners, subspecialists, and families to maximize the health of individuals with JS throughout the lifespan