Role of fungi in freshwater ecosystems

There are more than 600 species of freshwater fungi with a greater number known from temperate, as compared to tropical, regions. Three main groups can be considered which include Ingoldian fungi, aquatic ascomycetes and non-Ingoldian hyphomycetes, chytrids and, oomycetes. The fungi occurring in lentic habitats mostly differ from those occurring in lotic habitats. Although there is no comprehensive work dealing with the biogeography of all groups of freshwater fungi, their distribution probably follows that of Ingoldian fungi, which are either cosmopolitan, restricted to pantemperate or pantropical regions, or in a few cases, have a restricted distribution. Freshwater fungi are thought to have evolved from terrestrial ancestors. Many species are clearly adapted to life in freshwater as their propagules have specialised aquatic dispersal abilities. Freshwater fungi are involved in the decay of wood and leafy material and also cause diseases of plants and animals. These areas are briefly reviewed. Gaps in our knowledge of freshwater fungi are discussed and areas in need of research are suggested.     

Enzymes involved in fatty acid and polyketide biosynthesis in Streptomyces glaucescens: role of FabH and FabD and their acyl carrier protein specificity

Malonyl acyl carrier protein (ACP) is used as an extender unit in each of the elongation steps catalyzed by the type II dissociated fatty acid synthase (FAS) and polyketide synthase (PKS) of Streptomyces glaucescens. Initiation of straight-chain fatty acid biosynthesis by the type II FAS involves a direct condensation of acetyl-CoA with this malonyl-ACP to generate a 3-ketobutyryl-ACP product and is catalyzed by FabH. In vitro experiments with a reconstituted type II PKS system in the absence of FabH have previously shown that the acetyl-ACP (generated by decarboxylation of malonyl-ACP), not acetyl-CoA, is used to initiate tetracenomycin C (TCM C) biosynthesis. We have shown that sgFabH activity is present in S. glaucescens fermentations during TCM C production, suggesting that it could contribute to initiation of TCM C biosynthesis in vivo. Isotope incorporation studies with [CD3]acetate and [13CD3]acetate demonstrated significant intact retention of three deuteriums into the starter unit of palmitate and complete washout of deuterium label into the starter unit of TCM C. These observations provide evidence that acetyl-CoA is not used directly as a starter unit for TCM C biosynthesis in vivo and argue against an involvement of FabH in this process. Consistent with this conclusion, assays of the purified recombinant sgFabH with acetyl-CoA demonstrated activity using malonyl-ACP generated from either FabC (the S. glaucescens FAS ACP) (k(cat) 42.2 min(-1), K(m) 4.5 +/- 0.3 microM) or AcpP (the E. coli FAS ACP) (k(cat) 7.5 min(-1), K(m) 6.3 +/- 0.3 microM) but not TcmM (the S. glaucescens PKS ACP). In contrast, the sgFabD which catalyzes conversion of malonyl-CoA to malonyl-ACP for fatty acid biosynthesis was shown to be active with TcmM (k(cat) 150 min(-1), K(m) 12.2 +/- 1.2 microM), AcpP (k(cat) 141 min(-1), K(m) 13.2 +/- 1.6 microM), and FabC (k(cat) 560 min(-1), K(m) 12.7 +/- 2.6 microM). This enzyme was shown to be present during TCM C production and could play a role in generating malonyl-ACP for both processes. Previous demonstrations that the purified PKS ACPs catalyze self-malonylation and that a FabD activity is not required for polyketide biosynthesis are shown to be an artifact of the expression and purification protocols. The relaxed ACP specificity of FabD and the lack of a clear alternative are consistent with a role of FabD in providing malonyl-ACP precursors for PKS as well as FAS processes. In contrast, the ACP specificity of FabH, isotope labeling studies, and a demonstrated alternative mechanism for initiation of the PKS process provide unequivocal evidence that FabH is involved only in the FAS process.     

Transepithelial transport and enzymatic detoxification of gluten in gluten-sensitive rhesus macaques

Background and Aims

In a previous report, we characterized a condition of gluten sensitivity in juvenile rhesus macaques that is similar in many respects to the human condition of gluten sensitivity, celiac disease. This animal model of gluten sensitivity may therefore be useful toward studying both the pathogenesis and the treatment of celiac disease. Here, we perform two pilot experiments to demonstrate the potential utility of this model for studying intestinal permeability toward an immunotoxic gluten peptide and pharmacological detoxification of gluten in vivo by an oral enzyme drug candidate.

Methods

Intestinal permeability was investigated in age-matched gluten-sensitive and control macaques by using mass spectrometry to detect and quantify an orally dosed, isotope labeled 33-mer gluten peptide delivered across the intestinal epithelium to the plasma. The protective effect of a therapeutically promising oral protease, EP-B2, was evaluated in a gluten-sensitive macaque by administering a daily gluten challenge with or without EP-B2 supplementation. ELISA-based antibody assays and blinded clinical evaluations of this macaque and of an age-matched control were conducted to assess responses to gluten.

Results

Labeled 33-mer peptide was detected in the plasma of a gluten-sensitive macaque, both in remission and during active disease, but not in the plasma of healthy controls. Administration of EP-B2, but not vehicle, prevented clinical relapse in response to a dietary gluten challenge. Unexpectedly, a marked increase in anti-gliadin (IgG and IgA) and anti-transglutaminase (IgG) antibodies was observed during the EP-B2 treatment phase.

Conclusions

Gluten-sensitive rhesus macaques may be an attractive resource for investigating important aspects of celiac disease, including enhanced intestinal permeability and pharmacology of oral enzyme drug candidates. Orally dosed EP-B2 exerts a protective effect against ingested gluten. Limited data suggest that enhanced permeability of short gluten peptides generated by gastrically active glutenases may trigger an elevated antibody response, but that these antibodies are not necessarily causative of clinical illness.     

The non-native chironomid Eretmoptera murphyi in Antarctica: erosion of the barriers to invasion

Antarctica is the continent least affected by invasive species, but climate change and increasing human activity are increasing this threat. Antarctic terrestrial ecosystems generally have low biodiversity with simple community structures and little competition for resources. Consequently, species with pre-adaptations or capabilities that allow them to tolerate polar conditions may have disproportionately large ecosystem impacts when introduced to Antarctica compared with other regions of the Earth. Here we investigate the invasion risk associated with the flightless chironomid midge, Eretmoptera murphyi, which was accidentally introduced from South Georgia (54°S) to Signy Island, South Orkney Islands (61°S), probably during plant transplantation experiments in the 1960s. Larval size class distribution analysis indicated that E. murphyi has a 2 year life cycle on Signy Island, supporting previous suggestions. Estimates of litter turnover show that recent large increases in E. murphyi population density and extent are likely to increase nutrient cycling rates on Signy Island substantially. Existing physiological adaptations may allow E. murphyi to colonise higher latitude locations. Growth rate and microhabitat climatic modelling show that temperature constraints on larval development on Anchorage Island (68°S) are theoretically similar to those on Signy Island even though it is ~750 km further south. Establishment of this non-native midge at climatically similar intervening locations along the western Antarctic Peninsula is therefore plausible. Currently, lack of effective natural dispersal mechanisms is probably limiting the spread of the midge. However, dispersal to other areas of the Antarctic Peninsula may occur via human-assisted transportation, highlighting the importance of appropriate biosecurity measures.     

Beyond ecosystem modeling: A roadmap to community cyberinfrastructure for ecological data‐model integration

In an era of rapid global change, our ability to understand and predict Earth's natural systems is lagging behind our ability to monitor and measure changes in the biosphere. Bottlenecks to informing models with observations have reduced our capacity to fully exploit the growing volume and variety of available data. Here, we take a critical look at the information infrastructure that connects ecosystem modeling and measurement efforts, and propose a roadmap to community cyberinfrastructure development that can reduce the divisions between empirical research and modeling and accelerate the pace of discovery. A new era of data‐model integration requires investment in accessible, scalable, and transparent tools that integrate the expertise of the whole community, including both modelers and empiricists. This roadmap focuses on five key opportunities for community tools: the underlying foundations of community cyberinfrastructure; data ingest; calibration of models to data; model‐data benchmarking; and data assimilation and ecological forecasting. This community‐driven approach is a key to meeting the pressing needs of science and society in the 21st century.     

Identification of a complex genetic network underlying Saccharomyces cerevisiae colony morphology

When grown on solid substrates, different microorganisms often form colonies with very specific morphologies. Whereas the pioneers of microbiology often used colony morphology to discriminate between species and strains, the phenomenon has not received much attention recently. In this study, we use a genome‐wide assay in the model yeast Saccharomyces cerevisiae to identify all genes that affect colony morphology. We show that several major signalling cascades, including the MAPK, TORC, SNF1 and RIM101 pathways play a role, indicating that morphological changes are a reaction to changing environments. Other genes that affect colony morphology are involved in protein sorting and epigenetic regulation. Interestingly, the screen reveals only few genes that are likely to play a direct role in establishing colony morphology, with one notable example being FLO11, a gene encoding a cell‐surface adhesin that has already been implicated in colony morphology, biofilm formation, and invasive and pseudohyphal growth. Using a series of modified promoters for fine‐tuning FLO11 expression, we confirm the central role of Flo11 and show that differences in FLO11 expression result in distinct colony morphologies. Together, our results provide a first comprehensive look at the complex genetic network that underlies the diversity in the morphologies of yeast colonies.     

C-terminal domain of the Uup ATP-binding cassette ATPase is an essential folding domain that binds to DNA

The Uup protein belongs to a subfamily of soluble ATP-binding cassette (ABC) ATPases that have been implicated in several processes different from transmembrane transport of molecules, such as transposon precise excision. We have demonstrated previously that Escherichia coli Uup is able to bind DNA. DNA binding capacity is lowered in a truncated Uup protein lacking its C-terminal domain (CTD), suggesting a contribution of CTD to DNA binding. In the present study, we characterize the role of CTD in the function of Uup, on its overall stability and in DNA binding. To this end, we expressed and purified isolated CTD and we investigated the structural and functional role of this domain. The results underline that CTD is essential for the function of Uup, is stable and able to fold up autonomously. We compared the DNA binding activities of three versions of the protein (Uup, UupΔCTD and CTD) by an electrophoretic mobility shift assay. CTD is able to bind DNA although less efficiently than intact Uup and UupΔCTD. These observations suggest that CTD is an essential domain that contributes directly to the DNA binding ability of Uup.