Anna Shevtsova

Growth response of subarctic dwarf shrubs, Empetrum nigrum and Vaccinium vitis-idaea, to manipulated environmental conditions and species removal

We investigated the growth responses of two evergreen dwarf shrubs Empetrum nigrum and Vaccinium vitis-idaea to environmental manipulations (elevated temperature, increased precipitation and simulated acid rain) and to experimentally altered species composition in a three-year field experiment in the Finnish Subarctic. The responses of these species to experimental manipulations were highly complex, due to species-specific patterns of growth, great inter-annual variation and a high number of interactions between temperature, water and community composition. Both species showed increased shoot growth in the second and third season in response to elevated temperature. Elevated temperature also accelerated the vegetative bud break and shifted the peak shoot growth to an earlier time. The shoot growth of V. vitis-idaea was generally enhanced by additional irrigation, while the growth of E. nigrum tended to decrease in watered plots. Removal of heterospecific neighbours significantly modified growth responses to environmental manipulations. The positive effect of elevated temperature on E. nigrum was usually more profound on mixed species than on removal plots. The shoot growth of V. vitis-idaea was increased most when warming and watering were applied to plots of mixed species composition. The effect of the removal of companion species in turn depended greatly on environmental manipulations. Following the removal of V. vitis-idaea, E. nigrum generally increased branching, especially on open plots, and it increased the size of current shoots on open irrigated plots. However, when temperature was elevated, removal of V. vitis-idaea had negative effect on shoot growth and mass in E. nigrum. In V. vitis-idaea, current shoot size decreased after removal of E. nigrum on plots where either temperature or precipitation was increased. Only the most favourable conditions, on plots with both warming and watering, eliminated the negative effect of companion species removal on V. vitis-idaea. Acid rain treatment had no consistent effect on shoot growth of E. nigrum, while it greatly increased the production of adventitious shoots in V. vitis-idaea. The three years of experimental perturbations modified the shoot architecture markedly due to the effects of treatments on vegetative production per current shoot and the rate of lateral meristems activation. Changes in shoot architecture and phenology may be caused even by a short-term modification of climate and community composition, and may govern the success of these dwarf shrubs in a changing climate.

Plant community type and small‐scale disturbances, but not altitude, influence the invasibility in subarctic ecosystems

Little of our knowledge about invasibility comes from arctic and alpine ecosystems, despite increasing plant migration and invasion in those regions. Here, we examine how community type, altitude, and small-scale disturbances affect invasibility in a subarctic ecosystem. Over a period of 4 yr, we studied seedling emergence and establishment in 17 species sown in gaps or undisturbed vegetation in four subarctic community types (Salix scrub, meadow, rich heath, poor heath) along an elevation gradient. Invasibility was lowest in rich heath and highest in Salix scrub. Small disturbances significantly increased the invasibility in most communities, thereby showing the importance of biotic resistance to invasion in subarctic regions. Unexpectedly, invasibility did not decrease with increasing elevation, and it was also not related to summer temperature. Our data suggest that biotic resistance might be more important than abiotic stress for invasibility in subarctic tundra and that low temperatures do not necessarily limit seedling establishment at high altitudes. High elevations are therefore potentially more vulnerable to invasion than was originally thought. Changes in community composition as a result of species migration or invasion are most likely to occur in Salix scrub and meadow, whereas Empetrum-dominated rich heath will largely remain unchanged.

Structural Insights into Polymorphic ABO Glycan Binding by Helicobacter pylori.

The Helicobacter pylori adhesin BabA binds mucosal ABO/Le(b) blood group (bg) carbohydrates. BabA facilitates bacterial attachment to gastric surfaces, increasing strain virulence and forming a recognized risk factor for peptic ulcers and gastric cancer. High sequence variation causes BabA functional diversity, but the underlying structural-molecular determinants are unknown. We generated X-ray structures of representative BabA isoforms that reveal a polymorphic, three-pronged Le(b) binding site. Two diversity loops, DL1 and DL2, provide adaptive control to binding affinity, notably ABO versus O bg preference. H. pylori strains can switch bg preference with single DL1 amino acid substitutions, and can coexpress functionally divergent BabA isoforms. The anchor point for receptor binding is the embrace of an ABO fucose residue by a disulfide-clasped loop, which is inactivated by reduction. Treatment with the redox-active pharmaceutic N-acetylcysteine lowers gastric mucosal neutrophil infiltration in H. pylori-infected Le(b)-expressing mice, providing perspectives on possible H. pylori eradication therapies.

Antiadhesive Properties of Abelmoschus esculentus (Okra) Immature Fruit Extract against Helicobacter pylori Adhesion

Background Traditional Asian and African medicine use immature okra fruits (Abelmoschus esculentus) as mucilaginous food to combat gastritis. Its effectiveness is due to polysaccharides that inhibit the adhesion of Helicobacter pylori to stomach tissue. The present study investigates the antiadhesive effect in mechanistic detail. Methodology A standardized aqueous fresh extract (Okra FE) from immature okra fruits was used for a quantitative in vitro adhesion assay with FITC-labled H. pylori J99, 2 clinical isolates, AGS cells, and fluorescence-activated cell sorting. Bacterial adhesins affected by FE were pinpointed using a dot-blot overlay assay with immobilized Lewisb, sialyl-Lewisa, H-1, laminin, and fibronectin. 125I-radiolabeled Okra FE polymer served for binding studies to different H. pylori strains and interaction experiments with BabA and SabA. Iron nanoparticles with different coatings were used to investigate the influence of the charge-dependence of an interaction on the H. pylori surface. Principal findings Okra FE dose-dependently (0.2 to 2 mg/mL) inhibited H. pylori binding to AGS cells. FE inhibited the adhesive binding of membrane proteins BabA, SabA, and HpA to its specific ligands. Radiolabeled compounds from FE bound non-specifically to different strains of H. pylori, as well as to BabA/SabA deficient mutants, indicating an interaction with a still-unknown membrane structure in the vicinity of the adhesins. The binding depended on the charge of the inhibitors. Okra FE did not lead to subsequent feedback regulation or increased expression of adhesins or virulence factors. Conclusion Non-specific interactions between high molecular compounds from okra fruits and the H. pylori surface lead to strong antiadhesive effects.

The response of forest plant regeneration to temperature variation along a latitudinal gradient

BACKGROUND AND AIMS The response of forest herb regeneration from seed to temperature variations across latitudes was experimentally assessed in order to forecast the likely response of understorey community dynamics to climate warming. METHODS Seeds of two characteristic forest plants (Anemone nemorosa and Milium effusum) were collected in natural populations along a latitudinal gradient from northern France to northern Sweden and exposed to three temperature regimes in growth chambers (first experiment). To test the importance of local adaptation, reciprocal transplants were also made of adult individuals that originated from the same populations in three common gardens located in southern, central and northern sites along the same gradient, and the resulting seeds were germinated (second experiment). Seedling establishment was quantified by measuring the timing and percentage of seedling emergence, and seedling biomass in both experiments. KEY RESULTS Spring warming increased emergence rates and seedling growth in the early-flowering forb A. nemorosa. Seedlings of the summer-flowering grass M. effusum originating from northern populations responded more strongly in terms of biomass growth to temperature than southern populations. The above-ground biomass of the seedlings of both species decreased with increasing latitude of origin, irrespective of whether seeds were collected from natural populations or from the common gardens. The emergence percentage decreased with increasing home-away distance in seeds from the transplant experiment, suggesting that the maternal plants were locally adapted. CONCLUSIONS Decreasing seedling emergence and growth were found from the centre to the northern edge of the distribution range for both species. Stronger responses to temperature variation in seedling growth of the grass M. effusum in the north may offer a way to cope with environmental change. The results further suggest that climate warming might differentially affect seedling establishment of understorey plants across their distribution range and thus alter future understorey plant dynamics.

A latitudinal gradient in seed nutrients of the forest herb Anemone nemorosa

The nutrient concentration in seeds determines many aspects of potential success of the sexual reproductive phase of plants, including the seed predation probability, efficiency of seed dispersal and seedling performance. Despite considerable research interest in latitudinal gradients of foliar nutrients, a similar gradient for seeds remains unexplored. We investigated a potential latitudinal gradient in seed nutrient concentrations within the widespread European understorey forest herb Anemone nemorosa L. We sampled seeds of A. nemorosa in 15 populations along a 1900-km long latitudinal gradient at three to seven seed collection dates post-anthesis and investigated the relative effects of growing degree-hours >5 °C, soil characteristics and latitude on seed nutrient concentrations. Seed nitrogen, nitrogen:phosphorus ratio and calcium concentration decreased towards northern latitudes, while carbon:nitrogen ratios increased. When taking differences in growing degree-hours and measured soil characteristics into account and only considering the most mature seeds, the latitudinal decline remained particularly significant for seed nitrogen concentration. We argue that the decline in seed nitrogen concentration can be attributed to northward decreasing seed provisioning due to lower soil nitrogen availability or greater investment in clonal reproduction. This pattern may have large implications for the reproductive performance of this forest herb as the degree of seed provisioning ultimately co-determines seedling survival and reproductive success.

Antiadhesive Properties of Arabinogalactan Protein from Ribes nigrum Seeds against Bacterial Adhesion of Helicobacter pylori

Fruit extracts from black currants (Ribes nigrum L.) are traditionally used for treatment of gastritis based on seed polysaccharides that inhibit the adhesion of Helicobacter pylori to stomach cells. For detailed investigations an arabinogalactan protein (F2) was isolated from seeds and characterized concerning molecular weight, carbohydrate, amino acid composition, linkage, configuration and reaction with β-glucosyl Yariv. Functional testing of F2 was performed by semiquantitative in situ adhesion assay on sections of human gastric mucosa and by quantitative in vitro adhesion assay with FITC-labled H. pylori strain J99 and human stomach AGS cells. Bacterial adhesins affected were identified by overlay assay with immobilized ligands. 125I-radiolabeled F2 served for binding studies to H. pylori and interaction experiments with BabA and SabA. F2 had no cytotoxic effects against H. pylori and AGS cells; but inhibited bacterial binding to human gastric cells. F2 inhibited the binding of BabA and fibronectin-binding adhesin to its specific ligands. Radiolabeled F2 bound non-specifically to different strains of H. pylori; and to BabA deficient mutant. F2 did not lead to subsequent feedback regulation or increased expression of adhesins or virulence factors. From these data the non-specific interactions between F2 and the H. pylori lead to moderate antiadhesive effects.

Muc5ac gastric mucin glycosylation is shaped by FUT2 activity and functionally impacts Helicobacter pylori binding

The gastrointestinal tract is lined by a thick and complex layer of mucus that protects the mucosal epithelium from biochemical and mechanical aggressions. This mucus barrier confers protection against pathogens but also serves as a binding site that supports a sheltered niche of microbial adherence. The carcinogenic bacteria Helicobacter pylori colonize the stomach through binding to host glycans present in the glycocalyx of epithelial cells and extracellular mucus. The secreted MUC5AC mucin is the main component of the gastric mucus layer, and BabA-mediated binding of H. pylori to MUC5AC confers increased risk for overt disease. In this study we unraveled the O-glycosylation profile of Muc5ac from glycoengineered mice models lacking the FUT2 enzyme and therefore mimicking a non-secretor human phenotype. Our results demonstrated that the FUT2 determines the O-glycosylation pattern of Muc5ac, with Fut2 knock-out leading to a marked decrease in α1,2-fucosylated structures and increased expression of the terminal type 1 glycan structure Lewis-a. Importantly, for the first time, we structurally validated the expression of Lewis-a in murine gastric mucosa. Finally, we demonstrated that loss of mucin FUT2-mediated fucosylation impairs gastric mucosal binding of H. pylori BabA adhesin, which is a recognized feature of pathogenicity.

Host Determinants of Expression of the Helicobacter pylori BabA Adhesin

Expression of the Helicobacter pylori blood group antigen binding adhesin A (BabA) is more common in strains isolated from patients with peptic ulcer disease or gastric cancer, rather than asymptomatic colonization. Here we used mouse models to examine host determinants that affect H. pylori BabA expression. BabA expression was lost by phase variation as frequently in WT mice as in RAG2−/− mice that do not have functional B or T cells, and in MyD88−/−, TLR2−/− and TLR4−/− mice that are defective in toll like receptor signaling. The presence of other bacteria had no effect on BabA expression as shown by infection of germ free mice. Moreover, loss of BabA expression was not dependent on Leb expression or the capacity of BabA to bind Leb. Surprisingly, gender was the host determinant most associated with loss of BabA expression, which was maintained to a greater extent in male mice and was associated with greater bacterial load. These results suggest the possibility that loss of BabA expression is not driven by adaptive immunity or toll-like receptor signaling, and that BabA may have other, unrecognized functions in addition to serving as an adhesin that binds Leb.