Macroecology meets macroevolution: evolutionary niche dynamics in the seaweed Halimeda

Aim Because of their broad distribution in geographical and ecological dimensions, seaweeds (marine macroalgae) offer great potential as models for marine biogeographical inquiry and exploration of the interface between macroecology and macroevolution. This study aims to characterize evolutionary niche dynamics in the common green seaweed genus Halimeda, use the observed insights to gain understanding of the biogeographical history of the genus and predict habitats that can be targeted for the discovery of species of special biogeographical interest. Location Tropical and subtropical coastal waters. Methods The evolutionary history of the genus is characterized using molecular phylogenetics and relaxed molecular clock analysis. Niche modelling is carried out with maximum entropy techniques and uses macroecological data derived from global satellite imagery. Evolutionary niche dynamics are inferred through application of ancestral character state estimation. Results A nearly comprehensive molecular phylogeny of the genus was inferred from a six‐locus dataset. Macroecological niche models showed that species distribution ranges are considerably smaller than their potential ranges. We show strong phylogenetic signal in various macroecological niche features. Main conclusions The evolution of Halimeda is characterized by conservatism for tropical, nutrient‐depleted habitats, yet one section of the genus managed to invade colder habitats multiple times independently. Niche models indicate that the restricted geographical ranges of Halimeda species are not due to habitat unsuitability, strengthening the case for dispersal limitation. Niche models identified hotspots of habitat suitability of Caribbean species in the eastern Pacific Ocean. We propose that these hotspots be targeted for discovery of new species separated from their Caribbean siblings since the Pliocene rise of the Central American Isthmus.     

Development and laboratory-scale testing of a fully automated online flow cytometer for drinking water analysis

Accurate and sensitive online detection tools would benefit both fundamental research and practical applications in aquatic microbiology. Here, we describe the development and testing of an online flow cytometer (FCM), with a specific use foreseen in the field of drinking water microbiology. The system incorporated fully automated sampling and fluorescent labeling of bacterial nucleic acids with analysis at 5-min intervals for periods in excess of 24 h. The laboratory scale testing showed sensitive detection (< 5% error) of bacteria over a broad concentration range (1 × 10(3) -1 × 10(6) cells mL(-1) ) and particularly the ability to track both gradual changes and dramatic events in water samples. The system was tested with bacterial pure cultures as well as indigenous microbial communities from natural water samples. Moreover, we demonstrated the possibility of using either a single fluorescent dye (e.g., SYBR Green I) or a combination of two dyes (SYBR Green I and Propidium Iodide), thus broadening the application possibilities of the system. The online FCM approach described herein has considerable potential for routine and continuous monitoring of drinking water, optimization of specific drinking water processes such as biofiltration or disinfection, as well as aquatic microbiology research in general.     

Cytochrome P-450-dependent H2O2 production demonstrated in vivo. Influence of phenobarbital and allylisopropylacetamide

By administration of allylisopropylacetamide, an inhibitor of cytochrome P-450, we demonstrated that cytochrome P-450 is involved in the production of H2O2 during aminopyrine metabolism and phenobarbital induction in both the unanaesthetized guinea pig and rat. In the guinea pig we also found evidence for the existence of a basal cytochrome P-450-dependent H2O2 production, i.e. in the absence of exogenous substrate. Catalase participates in the decomposition of H2O2 produced in the endoplasmic reticulum where cytochrome P-450 is localized.     

Gain of function of mutant p53 by coaggregation with multiple tumor suppressors

Many p53 missense mutations possess dominant-negative activity and oncogenic gain of function. We report that for structurally destabilized p53 mutants, these effects result from mutant-induced coaggregation of wild-type p53 and its paralogs p63 and p73, thereby also inducing a heat-shock response. Aggregation of mutant p53 resulted from self-assembly of a conserved aggregation-nucleating sequence within the hydrophobic core of the DNA-binding domain, which becomes exposed after mutation. Suppressing the aggregation propensity of this sequence by mutagenesis abrogated gain of function and restored activity of wild-type p53 and its paralogs. In the p53 germline mutation database, tumors carrying aggregation-prone p53 mutations have a significantly lower frequency of wild-type allele loss as compared to tumors harboring nonaggregating mutations, suggesting a difference in clonal selection of aggregating mutants. Overall, our study reveals a novel disease mechanism for mutant p53 gain of function and suggests that, at least in some respects, cancer could be considered an aggregation-associated disease.     

Complex phylogenetic distribution of a non-canonical genetic code in green algae

Background  

A non-canonical nuclear genetic code, in which TAG and TAA have been reassigned from stop codons to glutamine, has evolved independently in several eukaryotic lineages, including the ulvophycean green algal orders Dasycladales and Cladophorales. To study the phylogenetic distribution of the standard and non-canonical genetic codes, we generated sequence data of a representative set of ulvophycean green algae and used a robust green algal phylogeny to evaluate different evolutionary scenarios that may account for the origin of the non-canonical code.     

Towards a coupled paradigm of NH3‐CO2 biosphere–atmosphere exchange modelling

Stomatal conductance, one of the major plant physiological controls within NH₃ biosphere–atmosphere exchange models, is commonly estimated from semi‐empirical multiplicative schemes or simple light‐ and temperature‐response functions. However, due to their inherent parameterization on meteorological proxy variables, instead of a direct measure of stomatal opening, they are unfit for the use in climate change scenarios and of limited value for interpreting field‐scale measurements. Alternatives based on H₂O flux measurements suffer from uncertainties in the partitioning of evapotranspiration at humid sites, as well as a potential decoupling of transpiration from stomatal opening in the presence of hygroscopic particles on leaf surfaces. We argue that these problems may be avoided by directly deriving stomatal conductance from CO₂ fluxes instead. We reanalysed a data set of NH₃ flux measurements based on CO₂‐derived stomatal conductance, confirming the hypothesis that the increasing relevance of stomatal exchange with the onset of vegetation activity caused a rapid decrease of observed NH₃ deposition velocities. Finally, we argue that developing more mechanistic representations of NH₃ biosphere–atmosphere exchange can be of great benefit in many applications. These range from model‐based flux partitioning, over deposition monitoring using low‐cost samplers and inferential modelling, to a direct response of NH₃ exchange to climate change.     

Effect of alemtuzumab-based T-cell depletion on graft compositional change in vitro and immune reconstitution early after allogeneic stem cell transplantation

Background aimsTo reduce the risk of graft-versus-host disease (GVHD) after allogeneic stem cell transplantation (alloSCT), T-cell depletion (TCD) of grafts can be performed by the addition of alemtuzumab (ALT) “to the bag” (in vitro) before transplantation. In this prospective study, the authors analyzed the effect of in vitro incubation with 20 mg ALT on the composition of grafts prior to graft infusion. Furthermore, the authors assessed whether graft composition at the moment of infusion was predictive for T-cell reconstitution and development of GVHD early after TCD alloSCT.MethodsSixty granulocyte colony-stimulating factor-mobilized stem cell grafts were obtained from ≥9/10 HLA-matched related and unrelated donors. The composition of the grafts was analyzed by flow cytometry before and after in vitro incubation with ALT. T-cell reconstitution and incidence of severe GVHD were monitored until 12 weeks after transplantation.ResultsIn vitro incubation of grafts with 20 mg ALT resulted in an initial median depletion efficiency of T-cell receptor (TCR) α/β T cells of 96.7% (range, 63.5–99.8%), followed by subsequent depletion in vivo. Graft volumes and absolute leukocyte counts of grafts before the addition of ALT were not predictive for the efficiency of TCR α/β T-cell depletion. CD4^pos T cells were depleted more efficiently than CD8^pos T cells, and naive and regulatory T cells were depleted more efficiently than memory and effector T cells. This differential depletion of T-cell subsets was in line with their reported differential CD52 expression. In vitro depletion efficiencies and absolute numbers of (naive) TCR α/β T cells in the grafts after ALT incubation were not predictive for T-cell reconstitution or development of GVHD post- alloSCT.ConclusionsThe addition of ALT to the bag is an easy, fast and generally applicable strategy to prevent GVHD in patients receiving alloSCT after myeloablative or non-myeloablative conditioning because of the efficient differential depletion of donor-derived lymphocytes and T cells.     

Bulky DNA adduct levels in normal-appearing colon mucosa,and the prevalence of colorectal adenomas

Abstract

Purpose: Examine the association between bulky DNA adduct levels in colon mucosa and colorectal adenoma prevalence, and explore the correlation between adduct levels in leukocytes and colon tissue.

Methods: Bulky DNA adduct levels were measured using ³²P-postlabelling in biopsies of normal-appearing colon tissue and blood donated by 202 patients. Multivariable logistic regression was used to examine associations between DNA adducts, and interactions of DNA adduct-DNA repair polymorphisms, with the prevalence of colorectal adenomas. Correlation between blood and tissue levels of DNA adducts was evaluated using Spearman’s correlation coefficient.

Results: An interaction between bulky DNA adduct levels and XPA rs1800975 on prevalence of colorectal adenoma was observed. Among individuals with lower DNA repair activity, increased DNA adduct levels were associated with increased colorectal adenoma prevalence (OR?=?1.41 per SD increase, 95%CI: 0.92–2.18). Conversely, among individuals with normal DNA activity, an inverse association was observed (OR?=?0.60 per SD increase, 95%CI: 0.34–1.07). Blood and colon DNA adduct levels were inversely correlated (ρ?=??0.20).

Conclusions: Among genetically susceptible individuals, higher bulky DNA adducts in the colon was associated with the prevalence of colorectal adenomas. The inverse correlation between blood and colon tissue measures demonstrates the importance of quantifying biomarkers in target tissues.     

Development of polyploidy of scale-building cells in the wings of Manduca sexta

The developing wings of butterflies and moths are composed of two epithelial monolayers. Each epithelial sheet is made up of two kinds of cells, diploid cells that make up the epidermal surface and body of the wing, and large polyploid cells that become the scale-building cells whose cytoplasmic projections develop into the scales that will cover the adult wing and bear the pigment pattern. We studied the development of polyploidization of the scale-building cells during the pupal stage of the tobacco hornworm moth, Manduca sexta. The endomitotic divisions of the presumptive scale-building cells and the mitotic divisions of the diploid epithelial cells begin on day 3 of the pupal stage and continue until day 7. We show that scales of different colors and positions on the wing differ in size, and that the size of the scale is proportional to the ploidy of the scale-building cell. Scale-building cells are arranged in irregular rows and within each row there is an alternation of ploidy levels, with the lower ploidy cells giving rise to the underscales and the higher ploidy cells giving rise to the cover scales that carry the color pattern. Along the wing there is a proximo-distal decreasing gradient of average ploidy and scale size. Scale-building cells of high ploidy are surrounded by fewer epidermal cells than those of low ploidy. This inverse relationship is known as Henke's compensation principle, which posits that the number of endomitoses of a pre-polyploid cell and the number of mitotic divisions of its diploid daughter cell add up to a constant. We show that the inverse relationship fits the predictions of the compensation principle and does not fit constraints imposed by packing density, and we discuss mechanisms that could give rise to the inverse relationship.     

Pre-Analytical Conditions in Non-Invasive Prenatal Testing of Cell-Free Fetal RHD

Background

Non-invasive prenatal testing of cell-free fetal DNA (cffDNA) in maternal plasma can predict the fetal RhD type in D negative pregnant women. In Denmark, routine antenatal screening for the fetal RhD gene (RHD) directs the administration of antenatal anti-D prophylaxis only to women who carry an RhD positive fetus. Prophylaxis reduces the risk of immunization that may lead to hemolytic disease of the fetus and the newborn. The reliability of predicting the fetal RhD type depends on pre-analytical factors and assay sensitivity. We evaluated the testing setup in the Capital Region of Denmark, based on data from routine antenatal RHD screening.

Methods

Blood samples were drawn at gestational age 25 weeks. DNA extracted from 1 mL of plasma was analyzed for fetal RHD using a duplex method for exon 7/10. We investigated the effect of blood sample transportation time (n = 110) and ambient outdoor temperatures (n = 1539) on the levels of cffDNA and total DNA. We compared two different quantification methods, the delta Ct method and a universal standard curve. PCR pipetting was compared on two systems (n = 104).

Results

The cffDNA level was unaffected by blood sample transportation for up to 9 days and by ambient outdoor temperatures ranging from -10°C to 28°C during transport. The universal standard curve was applicable for cffDNA quantification. Identical levels of cffDNA were observed using the two automated PCR pipetting systems. We detected a mean of 100 fetal DNA copies/mL at a median gestational age of 25 weeks (range 10–39, n = 1317).

Conclusion

The setup for real-time PCR-based, non-invasive prenatal testing of cffDNA in the Capital Region of Denmark is very robust. Our findings regarding the transportation of blood samples demonstrate the high stability of cffDNA. The applicability of a universal standard curve facilitates easy cffDNA quantification.