Sea ice microbial dynamics over an annual ice cycle in Prydz Bay, Antarctica

Microbial community dynamics within the fast sea ice of Prydz Bay (68°S?78°E) were investigated over an annual cycle at two sites (1 and 3?km offshore) between April and November 2008. There are few long-term sea ice studies, and few that cover the phase of winter darkness when autotrophic processes are curtailed. Mean chlorophyll a concentrations in the ice column ranged between 0.76 and 44.8?μg?L^?1 at the 1-km site (Site 1) and 3.11–144.6?μg?L^?1 at the 3-km site (Site 2). Highest chlorophyll a usually occurred at the base of the ice. Bacterial concentrations ranged between 0.30 and 2.08?×?10⁸?cells?L^?1, heterotrophic nanoflagellates (HNAN) between 0.21?×?10⁵ and 2.98?×?10⁵?cells?L^?1 and phototrophic nanoflagellates (PNAN) 0–1.06?×?10⁵?cells?L^?1. While HNAN occurred throughout the year, PNAN were largely absent in winter. Dinoflagellates were a conspicuous and occasionally an abundant element of the community (maximum 17,460?cells?L^?1), while ciliates were sparse. The bacterial community showed considerable morphological diversity with a dominance of filamentous forms. Bacterial production continued throughout the year ranging between 0 and 22.92?μg?C?L^?1?day^?1 throughout the ice column. Lowest rates occurred between late June and early August. The sea ice sustained an active and diverse microbial community through its annual extent. The data suggest that during winter darkness the microbial community is dominated by heterotrophic processes, sustained by a pool of dissolved organic carbon.     

Egg and larval load assessment and its influence on oviposition behaviour of the leaf beetle Galerucella nymphaeae

The oviposition behaviour of the water-lily beetle Galerucella nymphaeae was examined. This species is a specialist herbivore on the floating leaves of nymphaeids Nymphaeaceae and especially on the yellow water-lily, Nuphar lutea. Females lay their eggs in clutches on the leaves, and after hatching, the larvae feed on the leaves. The quality of the leaves decreases quickly after the larvae hatch, and eventually the leaves will sink below the water surface, whereupon the eggs, 1st-instar larvae and pupae are killed by drowning. The influence of conspecific eggs, larvae and feeding tracks on the oviposition preferences of the beetles was tested. Females were allowed to choose between fresh leaves and leaves with conspecific eggs and larvae as well as between leaves with larvae and leaves with feeding tracks but no larvae. An attempt was also made to determine whether eggs and larvae affect the oviposition rate of females when they are not given the opportunity to oviposit on untouched leaves. The results indicate that females tended to avoid leaves with conspecific larvae or to exhibit a decreased oviposition rate on such leaves. Females also avoided conspecific eggs, although the oviposition rate was not influenced by the presence of conspecific eggs. When females were allowed to choose between leaves with larvae and leaves with feeding tracks, possible discrimination against leaves with larvae just fails to reach the 5% level.     

A Binding Domain on Mesothelin for CA125/MUC16

Ovarian cancer and malignant mesothelioma frequently express both mesothelin and CA125 (also known as MUC16) at high levels on the cell surface. The interaction between mesothelin and CA125 may facilitate the implantation and peritoneal spread of tumors by cell adhesion, whereas the detailed nature of this interaction is still unknown. Here, we used truncated mutagenesis and alanine replacement techniques to identify a binding site on mesothelin for CA125. We examined the molecular interaction by Western blot overlay assays and further quantitatively analyzed by enzyme-linked immunosorbent assay. We also evaluated the binding on cancer cells by flow cytometry. We identified the region (296–359) consisting of 64 amino acids at the N-terminal of cell surface mesothelin as the minimum fragment for complete binding activity to CA125. We found that substitution of tyrosine 318 with an alanine abolished CA125 binding. Replacement of tryptophan 321 and glutamic acid 324 with alanine could partially decrease binding to CA125, whereas mutation of histidine 354 had no effect. These results indicate that a conformation-sensitive structure of the region (296–359) is required and sufficient for the binding of mesothelin to CA125. In addition, we have shown that a single chain monoclonal antibody (SS1) recognizes this CA125-binding domain and blocks the mesothelin-CA125 interaction on cancer cells. The identified CA125-binding domain significantly inhibits cancer cell adhesion and merits evaluation as a new therapeutic agent for preventing or treating peritoneal malignant tumors.Ovarian cancer largely is confined to the peritoneal cavity for much of its natural history (1). Peritoneal mesothelioma is a highly invasive tumor originating from the mesothelial linings of the peritoneum (2). The development of effective drug regimens against ovarian cancer and mesothelioma has proven extremely difficult.Mesothelin was first identified in 1992 by the monoclonal antibody (mAb)² K1 that was generated by the immunization of mice with human ovarian carcinoma (OVCAR-3) cells (3). The mesothelin gene encodes a 71-kDa precursor protein that is processed to a 40-kDa protein termed mesothelin, which is a glycosylphosphatidylinositol (GPI)-anchored glycoprotein present on the cell surface (4). Mesothelin is a differentiation antigen that is present on a restricted set of normal adult tissues such as the mesothelium. In contrast, it is overexpressed in a variety of cancers including mesothelioma, ovarian cancer, and pancreatic cancer (5). In addition, mesothelin is also expressed on the surface of non-small cell lung cancer cells (6, 7), especially most lung adenocarcinomas (8).We and others have shown that mesothelin is shed from tumor cells (9, 10), and antibodies specific for mesothelin are elevated in the sera of patients with mesothelioma and ovarian cancer (11). Shed serum mesothelin has been approved by the United States Food and Drug Administration (FDA) as a new diagnostic biomarker in mesothelioma. In a Phase I clinical study of an intrapleural interferon-β gene transfer using an adenoviral vector in patients with mesotheliomas, we found that antitumor immune responses targeting mesothelin were elicited in several patients (12). A recent study indicated that anti-mesothelin antibodies and circulating mesothelin relate to the clinical state in ovarian cancer patients (13). Pastan and colleagues (14) developed an immunotoxin (SS1P) with a Fv for mesothelin. Two Phase I clinical trials were completed at the National Cancer Institute (National Institutes of Health, Bethesda, MD) and there was sufficient antitumor activity of SS1P to justify a Phase II trial. A chimeric antibody containing the mouse SS1 Fv for mesothelin was also developed and is currently examined in a Phase I clinical trial for ovarian cancer, mesothelioma, pancreatic cancer, and non-small cell lung cancer (15).Mucins are heavily glycosylated proteins found in the mucus layer or at the cell surface of many epitheliums (16). There are two structurally distinct families of mucins, secreted and membrane-bound forms. CA125 (also known as MUC16) was first identified in 1981 by OC125, a mAb that had been developed from mice immunized with human ovarian cancer cells (17). The first cDNA clones were reported in 2001 (18, 19). CA125 is a very large membrane-bound cell surface mucin, with an average molecular mass between 2.5 and 5 million daltons. It is also heavily glycosylated with both O-linked and N-linked oligosaccharides (20). The peptide backbone of CA125 is composed of the N-terminal region, extensive Ser/Thr/Pro-rich tandem repeats (TR) with 156 amino acids each with both N- and O-glycosylations, a SEA domain with high levels of O-glycosylation and a C-terminal region with a short cytoplasmic tail (19). The SEA domain was first identified as a module commonly found in sea urchin sperm protein, enterokinase and agrin (21, 22). The significance of the SEA domain in CA125 is not clear.CA125 was originally used as a biomarker in ovarian cancer due to its high expression in ovarian carcinomas and that it is shed into the serum (23). A majority (88%) of mesotheliomas are also CA125 positive on the cell membrane (24). It was shown that 25% of peritoneal mesotheliomas have high CA125 expression (25). The intensity of CA125 membranous expression is indistinguishable between ovarian carcinomas and peritoneal mesotheliomas. Gene expression analysis using the SAGE tag data base has shown that mesothelioma has the second highest co-expression of CA125 and mesothelin after ovarian cancer (26). Rump and colleagues (26) have shown that mesothelin binds to CA125 and that this interaction may mediate cell adhesion. Scholler et al. (27) recently showed that CA125/mesothelin-dependent cell attachment could be blocked with anti-CA125 antibodies. Because mesothelin is present on peritoneal mesothelium, there may be an important role for the mesothelin-CA125 interaction in the tumorigenesis of ovarian cancer and mesothelioma in the peritoneal cavity. The mesothelin binding site on CA125 may lie within the 156-amino acid TR units, indicating multimeric binding of mesothelin to CA125. It has been found that the extraordinarily abundant N-glycans on CA125, presumably in the TR region, are required for binding to both glycosylated and non-glycosylated mesothelin (28).Here, we identified the binding site of CA125 on mesothelin by use of truncated mutagenesis and alanine replacement approaches. We measured binding qualitatively by Western blot overlay assays and quantitatively by enzyme-linked immunosorbent assay (ELISA). We also evaluated the interaction of CA125 and mesothelin on cancer cells by flow cytometry. Furthermore, we have shown that a single chain mAb (SS1) recognized the CA125-binding domain and blocked the mesothelin-CA125 interaction on cancer cells. The identified CA125-binding domain-Fc fusion protein also significantly inhibited cancer cell adhesion. Our results suggest that conformation-sensitive structures of the region (296–359) are required and sufficient for specific binding of mesothelin to CA125. The domain proteins or the antibodies that block the mesothelin-CA125 interaction merit evaluation as new therapeutic agents in treating peritoneal malignant tumors.     

Development of a sensory neuronal cell model for the estimation of mild eye irritation

In an attempt to improve the in vitro test strategy for the estimation of eye irritation, a neuronal cell model has been developed, with cells expressing vanilloid receptor type 1 (VR1) nociceptors. The currently accepted method for measuring eye irritancy is the ethically and scientifically criticised Draize rabbit eye test, despite the fact that alternative in vitro methods are available which have proved to be reliable and reproducible for predicting severe ocular toxicity. However, no alternative tests for measuring neuronal stimulation have yet been developed, and the prediction of eye irritation in the mild range is therefore insufficient. VR1 is a nociceptor localised in C-fibre neurons innervating the cornea and the surrounding tissue, and it responds to potentially damaging stimuli by releasing Ca2+ into the cytoplasm. As a sensory endpoint, [Ca2+]i was measured in VR1 transfected cells, as well as in control cells. Short-term cell cytotoxicity studies (cell membrane rupture and morphological divergence) were used to determine the non-corrosive concentrations of the test chemicals. Preliminary results indicated that hygiene products used daily may induce eye irritation via VR1 nociceptors. The lowest toxic concentration (0.025%) of liquid hand soap, as determined by morphologic divergences of cells, generated an 80% increase in [Ca2+]i over the basal [Ca2+]i in VR1 transfected cells, whereas the non-specific [Ca2+]i increased by 33%. Furthermore, all the endpoints studied indicated that shampoo for children was less active than shampoo for adults. If this method is successfully validated with standardised reference chemicals, the model could complete the test battery of in vitro alternatives, resulting in the saving of thousands of laboratory animals.     

Consequences of Sphaeropsis tip blight disease for the phytohormone profile and antioxidative metabolism of its pine host

Phytopathogenic fungi infections induce plant defence responses that mediate changes in metabolic and signalling processes with severe consequences for plant growth and development. Sphaeropsis tip blight, induced by the endophytic fungus Sphaeropsis sapinea that spreads from stem tissues to the needles, is the most widespread disease of conifer forests causing dramatic economic losses. However, metabolic consequences of this disease on bark and wood tissues of its host are largely unexplored. Here, we show that diseased host pines experience tissue dehydration in both bark and wood. Increased cytokinin and declined indole‐3‐acetic acid levels were observed in both tissues and increased jasmonic acid and abscisic acid levels exclusively in the wood. Increased lignin contents at the expense of holo‐cellulose with declined structural biomass of the wood reflect cell wall fortification by S. sapinea infection. These changes are consistent with H₂O₂ accumulation in the wood, required for lignin polymerization. Accumulation of H₂O₂ was associated with more oxidized redox states of glutathione and ascorbate pools. These findings indicate that S. sapinea affects both phytohormone signalling and the antioxidative defence system in stem tissues of its pine host during the infection process.     

Herbivory limits the yellow water lily in an overgrown lake and in flowing water

Aquatic macrophytes with floating leaves are often key ecological species that affect entire aquatic ecosystems. Here we describe an investigation of the importance of insect herbivory for population growth and leaf senescence in the yellow water lily (Nuphar lutea). In order to gain a general picture of the importance of herbivory under different conditions, we experimentally manipulated herbivory in a large lily population in natural still water and observed the natural development of 32 smaller populations in flowing water. Herbivory drastically increased leaf senescence, reducing leaf density. In the still water, over one summer, leaf density increased by a factor of 1.23 in the presence of water lily leaf beetles and 1.61 when herbivory was eliminated. In flowing water, population growth was restricted mainly by leaf crowdedness, which limited large dense populations. Herbivory by water lily leaf beetles also had a limiting effect on yellow water lily, again mainly in large dense populations. Small populations supported a lower density of beetles. Previous studies have not addressed population-level responses of vascular plants with floating leaves. Our results suggest that herbivory can result in greater light penetration into the water and reduce “enemy-free space” for aquatic species that find such space in water lily stands. We suggest that the water lily leaf beetle should be considered an “ecological engineer.”     

Gene promoter methylation is associated with lung function in the elderly: The normative aging study

Lung function is a strong predictor of mortality. While inflammatory markers have been associated with lung function decrease, pathways are still poorly understood and epigenetic changes may participate in lung function decline mechanisms. We studied the cross-sectional association between DNA methylation in nine inflammatory genes and lung function in a cohort of 756 elderly men living in the metropolitan area of Boston. Participants donated a blood sample for DNA methylation analysis and underwent spirometry at each visit every 3 to 5 y from 1999–2006. We used separate multivariate mixed effects regression models to study the association between each lung function measurement and DNA methylation within each gene. Decreased CRAT, F3 and TLR2 methylation was significantly associated with lower lung function. One interquartile range (IQR) decrease in DNA methylation was associated with lower forced vital capacity (FVC) and forced expiratory volume in one second (FEV₁), respectively by 2.94% (p < 10⁻⁴) and 2.47% (p < 10⁻³) for F3 and by 2.10% (p < 10⁻²) and 2.42% (p < 10⁻³) for TLR2. Decreased IFNγ and IL6 methylation was significantly associated with better lung function. One IQR decrease in DNA methylation was associated with higher FEV₁ by 1.75% (p = 0.02) and 1.67% (p = 0.05) for IFNγ and IL6, respectively. These data demonstrate that DNA methylation may be part of the biological processes underlying the lung function decline and that IFNγ and IL6 may have ambivalent roles through activation of negative feedback.Key words: DNA methylation, genes, spirometry, FEV₁, lungs, TLR2, F3, INOS, GCR, OGG1     

The distribution of microplankton in the McMurdo Dry Valley Lakes,Antarctica: response to ecosystem legacy or present-day climatic controls?

Plankton abundance and biomass were investigated in five lakes of the McMurdo Dry Valleys, Antarctica: Lakes Bonney, Fryxell, Joyce, Hoare and Miers. Despite plankton communities being dominated by organisms <100 m in length, there were striking differences between the lakes, including large variations in plankton vertical distribution and differences in total plankton biomass. Bacterial biomass was highest in the anoxic monimolimnia of the meromictic lakes, reaching 191 g C l^–1 in Lake Fryxell. Photosynthetic nanoflagellates dominated phytoplankton in the five lakes studied. Highest chlorophyll a concentrations were recorded at the chemocline of Lake Fryxell (21 g chl a l^–1). Heterotrophic nanoflagellate concentrations were low, ranging from 2 cells ml^–1 in Hoare to 237 cells ml^–1 in Bonney. By Antarctic standards, ciliates were relatively successful in terms of biomass and diversity in Lakes Fryxell and Hoare. In contrast, Lake Miers possessed extremely low ciliate abundance (<0.04 cells ml^–1). On both sampling occasions, copepod nauplii were observed in Lake Joyce. This is the first recording of crustacean zooplankton within the McMurdo Dry Valley Lakes. Because the foodwebs of these lakes are structured by bottom-up forces, differences in plankton distributions could be related to the physicochemical characteristics of each lake. The effect of lake evolution (legacy) and present-day climate change on planktonic dynamics is discussed.     

Dispersal biology of Liatris scariosa var. novae-angliae (Asteraceae), a rare New England grassland perennial

Propagule dispersal biology is a crucial avenue of research for rare plant species, especially those adapted to disturbance, such as northern blazing star (Liatris scariosa var. novae-angliae), a rare, early-successional New England grassland perennial. We examined the dispersal ability of northern blazing star propagules collected from 14 populations covering the entire latitudinal range of the taxon. Multiple regression demonstrated that dispersal ability, as measured by drop time in still air and flight distance in a low-speed wind tunnel, decreased significantly with propagule size and achene length, and increased with achene width and (for flight distance) pappus length. We used this multiple regression model to test for differences in predicted dispersal capability among maternal families, populations, and inland, coastal, and island habitats. Dispersal capability differed significantly among families and populations but not regions, and allometric relationships between morphological measurements were consistent across populations. Overall, dispersal capability was negatively correlated with germination success in a common greenhouse environment. However, germination success for a given dispersal ability, as well as achene shape, differed among populations. These results suggest specific populations to be targeted for management efforts promoting dispersal and establishment.