An evaluation of iron bioavailability and speciation in western lake superior with the use of combined physical, chemical, and biological assessment

An iron-dependent cyanobacterial bioreporter (Synechococcus strain KAS101) was used in unison with sizefractionated iron content (.0.45, ,0.45, ,0.02 mm), and chemical characterization of iron complexation (C18 resin column) to elucidate the bioavailable forms of iron present in Lake Superior during periods of inverse thermal stratification (May) and strong thermal stratification (September) of the water column. The results provide evidence of organic complexation of iron in Lake Superior waters. Iron in most sampled water was complexed by organic compounds that behaved like fulvic acids, whereas some samples showed evidence for the presence of siderophore-like compounds. The presence of dissolved organic matter suppressed the cellular luminescence of the bioreporter, indicating an increased iron bioavailability. This effect could result either from the presence of siderophores forming iron complexes that are bioavailable to the bioreporter, or from more indirect effects because of the presence of other organic compounds, such as fulvic acids or polysaccharides. Model ligand additions, iron bioaccumulation, and photo-oxidation of dissolved organic matter were used to assess the bioavailability of organically complexed iron to the bioreporter. A significant fraction of the iron (40- 100%) was bioavailable to the bioreporter. Iron bioavailability was high enough for the bioreporter not to be iron limited in the water collected from Lake Superior. This measure of bioavailability to picocyanobacteria is relevant because picoplankton accounted for the majority of chlorophyll a in Lake Superior during this study. 2009, by the American Society of Limnology and Oceanography, Inc

Phylogeny of Parasitic Parabasalia and Free-Living Relatives Inferred from Conventional Markers vs. Rpb1, a Single-Copy Gene

Parabasalia are single-celled eukaryotes (protists) that are mainly comprised of endosymbionts of termites and wood roaches, intestinal commensals, human or veterinary parasites, and free-living species. Phylogenetic comparisons of parabasalids are typically based upon morphological characters and 18S ribosomal RNA gene sequence data (rDNA), while biochemical or molecular studies of parabasalids are limited to a few axenically cultivable parasites. These previous analyses and other studies based on PCR amplification of duplicated protein-coding genes are unable to fully resolve the evolutionary relationships of parabasalids. As a result, genetic studies of Parabasalia lag behind other organisms.Comparing parabasalid EF1α, α-tubulin, enolase and MDH protein-coding genes with information from the Trichomonas vaginalis genome reveals difficulty in resolving the history of species or isolates apart from duplicated genes. A conserved single-copy gene encodes the largest subunit of RNA polymerase II (Rpb1) in T. vaginalis and other eukaryotes. Here we directly sequenced Rpb1 degenerate PCR products from 10 parabasalid genera, including several T. vaginalis isolates and avian isolates, and compared these data by phylogenetic analyses. Rpb1 genes from parabasalids, diplomonads, Parabodo, Diplonema and Percolomonas were all intronless, unlike intron-rich homologs in Naegleria, Jakoba and Malawimonas.The phylogeny of Rpb1 from parasitic and free-living parabasalids, and conserved Rpb1 insertions, support Trichomonadea, Tritrichomonadea, and Hypotrichomonadea as monophyletic groups. These results are consistent with prior analyses of rDNA and GAPDH sequences and ultrastructural data. The Rpb1 phylogenetic tree also resolves species- and isolate-level relationships. These findings, together with the relative ease of Rpb1 isolation, make it an attractive tool for evaluating more extensive relationships within Parabasalia

Hypothesis for the evolution of three-helix Chl a/b and Chl a/c light-harvesting antenna proteins from two-helix and four-helix ancestors

The nuclear-encoded Chl a/b and Chl a/c antenna proteins of photosynthetic eukaryotes are part of an extended family of proteins that also includes the early light-induced proteins (ELIPs) and the 22 kDa intrinsic protein of PS II (encoded by psb S gene). All members of this family have three transmembrane helices except for the psb S protein, which has four. The amino acid sequences of these proteins are compared and related to the three-dimensional structure of pea LHC II Type I (Kühlbrandt and Wang, Nature 350: 130–134, 1991). The similarity of psb S to the three-helix members of the family suggests that the latter arose from a four-helix ancestor that lost its C-terminal helix by deletion. Strong internal similarity between the two halves of the psb S protein suggests that it in turn arose as the result of the duplication of a gene encoding a two-helix protein. Since psb S is reported to be present in at least one cyanobacterium, the ancestral four-helix protein may have been present prior to the endosymbiotic event or events that gave rise to the photosynthetic eukaryotes. The Chl a/b and Chl a/c antenna proteins, and the immunologically-related proteins in the rhodophytes may have had a common ancestor which was present in the early photosynthetic eukaryotes, and predated their division into rhodophyte, chromophyte and chlorophyte lineages. The LHC I-LHC II divergence probably occurred before the separation of higher plants from chlorophyte algae and euglenophytes, and the different Types of LHC I and LHC II proteins arose prior to the separation of angiosperms and gymnosperms.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/43538/1/11120_2004_Article_BF00029382.pd

Optimization of iron-dependent cyanobacterial (Synechococcus, Cyanophyceae) bioreporters to measure iron bioavailability

Complex chemistry and biological uptake pathways render iron bioavailability particularly difficult to assess in natural waters. Bioreporters are genetically modified organisms that are useful tools to directly sense the bioavailable fractions of solutes. In this study, three cyanobacterial bioreporters derived from Synechococcus PCC 7942 were examined for the purpose of optimizing the response to bioavailable Fe. Each bioreporter uses a Fe-regulated promoter (isiAB, irpA and mapA), modulated by distinct mechanisms under Fe deficiency, fused to a bacterial luciferase (luxAB). In order to provide a better understanding of the way natural conditions may affect the ability of the bioreporter to sense iron bioavailability, the effect of relevant environmental parameters on the response to iron was assessed. Optimal conditions (and limits of applicability) for the use of these bioreporters on the field were determined to be: a 12 h (12-24 h) exposure time, temperature of 15°C (15°C-22°C), photon flux density of 100 μmol photons·m-2·s-1 (37-200 μmol photons·m-2·s-1), initial biomass of 0.6-0.8 μg chlorophyll a (chl a)·L-1 (0.3-1.5 μg chl a·L-1) or approximately 105 bioreporter cells·mL-1, high phosphate (10 μM), and low micronutrients (absent). The measured luminescence was optimal with an exogenous addition of 60 μM aqueous decanal substrate allowing a 5 min reaction time in the dark before analysis. This study provides important considerations relating to the optimization in the use of bioreporters under field conditions that can be used for method development of other algal and cyanobacterial bioreporters in aquatic systems. © 2006 Phycological Society of America