A method for studying protistan diversity using massively parallel sequencing of V9 hypervariable regions of small-subunit ribosomal RNA genes

© 2009 The Authors. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in PLoS ONE 4 (2009): e6372, doi:10.1371/journal.pone.0006372.Massively parallel pyrosequencing of amplicons from the V6 hypervariable regions of small-subunit (SSU) ribosomal RNA (rRNA) genes is commonly used to assess diversity and richness in bacterial and archaeal populations. Recent advances in pyrosequencing technology provide read lengths of up to 240 nucleotides. Amplicon pyrosequencing can now be applied to longer variable regions of the SSU rRNA gene including the V9 region in eukaryotes. We present a protocol for the amplicon pyrosequencing of V9 regions for eukaryotic environmental samples for biodiversity inventories and species richness estimation. The International Census of Marine Microbes (ICoMM) and the Microbial Inventory Research Across Diverse Aquatic Long Term Ecological Research Sites (MIRADA-LTERs) projects are already employing this protocol for tag sequencing of eukaryotic samples in a wide diversity of both marine and freshwater environments. Massively parallel pyrosequencing of eukaryotic V9 hypervariable regions of SSU rRNA genes provides a means of estimating species richness from deeply-sampled populations and for discovering novel species from the environment.This work was supported by grants from the W.M. Keck Foundation and the Woods Hole Center for Oceans and Human Health from the National Institutes of Health and National Science Foundation (NIH/NIEHS 1 P50 ES012742-01 and NSF/OCE 0430724-J) (LAZ and SH)

Identification and quantification of microplastics in wastewater using focal plane array-based reflectance micro-FT-IR imaging

Microplastics (<5 mm) have been documented in environmental samples on a global scale. While these pollutants may enter aquatic environments via wastewater treatment facilities, the abundance of microplastics in these matrices has not been investigated. Although efficient methods for the analysis of microplastics in sediment samples and marine organisms have been published, no methods have been developed for detecting these pollutants within organic-rich wastewater samples. In addition, there is no standardized method for analyzing microplastics isolated from environmental samples. In many cases, part of the identification protocol relies on visual selection before analysis, which is open to bias. In order to address this, a new method for the analysis of microplastics in wastewater was developed. A pretreatment step using 30% hydrogen peroxide (H2O2) was employed to remove biogenic material, and focal plane array (FPA)-based reflectance micro-Fourier-transform (FT-IR) imaging was shown to successfully image and identify different microplastic types (polyethylene, polypropylene, nylon-6, polyvinyl chloride, polystyrene). Microplastic-spiked wastewater samples were used to validate the methodology, resulting in a robust protocol which was nonselective and reproducible (the overall success identification rate was 98.33%). The use of FPA-based micro-FT-IR spectroscopy also provides a considerable reduction in analysis time compared with previous methods, since samples that could take several days to be mapped using a single-element detector can now be imaged in less than 9 h (circular filter with a diameter of 47 mm). This method for identifying and quantifying microplastics in wastewater is likely to provide an essential tool for further research into the pathways by which microplastics enter the environment.This work is funded by a NERC (Natural Environment Research Council) CASE studentship (NE/K007521/1) with contribution from industrial partner Fera Science Ltd., United Kingdom. The authors would like to thank Peter Vale, from Severn Trent Water Ltd, for providing access to and additionally Ashley Howkins (Brunel University London) for providing travel and assistance with the sampling of the Severn Trent wastewater treatment plant in Derbyshire, UK. We are grateful to Emma Bradley and Chris Sinclair for providing helpful suggestions for our research

Impacts of Hurricanes Katrina and Rita on the microbial landscape of the New Orleans area

Author Posting. © The Author(s), 2007. This is the author's version of the work. It is posted here by permission of National Academy of Sciences of the USA for personal use, not for redistribution. The definitive version was published in Proceedings of the National Academy of Sciences 104 (2007): 9029-9034, doi:10.1073/pnas.0610552104.Floodwaters in New Orleans from Hurricanes Katrina and Rita were observed to contain high levels of fecal indicator bacteria and microbial pathogens, generating concern about long-term impacts of these floodwaters on the sediment and water quality of the New Orleans area and Lake Pontchartrain. We show here that fecal indicator microbe concentrations in offshore waters from Lake Pontchartrain returned to prehurricane concentrations within 2 months of the flooding induced by these hurricanes. Vibrio and Legionella species within the lake were more abundant in samples collected shortly after the floodwaters had receded compared with samples taken within the subsequent 3 months; no evidence of a long-term hurricane-induced algal bloom was observed. Giardia and Cryptosporidium were detected in canal waters. Elevated levels of fecal indicator bacteria observed in sediment could not be solely attributed to impacts from floodwaters, as both flooded and nonflooded areas exhibited elevated levels of fecal indicator bacteria. Evidence from measurements of Bifidobacterium and bacterial diversity analysis suggest that the fecal indicator bacteria observed in the sediment were from human fecal sources. Epidemiologic studies are highly recommended to evaluate the human health effects of the sediments deposited by the floodwaters.This work was funded by NSF-NIEHS Oceans and Human Health Program (NSF OCE0432368, OCE0432479, OCE0430724 and NIEHS P50 ES12736, ES012740, ES012742), the NSF-SGER Program (OCE 0554402, OCE 0554674, OCE 0554850, OCE0600130), the NSF-REU Program, and by the Georgia Sea Grant College Program (NA04OAR170033)

Spatial processing of visual information in the movement-detecting pathway of the fly

1. Spatial processing of visual signals in the fly's movement-detecting pathway was studied by recording the responses of directionally-selective movement-detecting (DSMD) neurons in the lobula plate. The summarized results pertain to a type of neuron which preferentially responds to horizontal movement directed toward the animal's midline. Three kinds of visual stimuli were used: moving gratings, reversing-contrast gratings and reversing-contrast bars.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/47087/1/359_2004_Article_BF00613743.pd

Shedding Light on Vampires: The Phylogeny of Vampyrellid Amoebae Revisited

With the advent of molecular phylogenetic techniques the polyphyly of naked filose amoebae has been proven. They are interspersed in several supergroups of eukaryotes and most of them already found their place within the tree of life. Although the ‘vampire amoebae’ have attracted interest since the middle of the 19th century, the phylogenetic position and even the monophyly of this traditional group are still uncertain. In this study clonal co-cultures of eight algivorous vampyrellid amoebae and the respective food algae were established. Culture material was characterized morphologically and a molecular phylogeny was inferred using SSU rDNA sequence comparisons. We found that the limnetic, algivorous vampyrellid amoebae investigated in this study belong to a major clade within the Endomyxa Cavalier-Smith, 2002 (Cercozoa), grouping together with a few soil-dwelling taxa. They split into two robust clades, one containing species of the genus Vampyrella Cienkowski, 1865, the other containing the genus Leptophrys Hertwig & Lesser, 1874, together with terrestrial members. Supported by morphological data these clades are designated as the two families Vampyrellidae Zopf, 1885, and Leptophryidae fam. nov. Furthermore the order Vampyrellida West, 1901 was revised and now corresponds to the major vampyrellid clade within the Endomyxa, comprising the Vampyrellidae and Leptophryidae as well as several environmental sequences. In the light of the presented phylogenetic analyses morphological and ecological aspects, the feeding strategy and nutritional specialization within the vampyrellid amoebae are discussed

A decadal view of biodiversity informatics: challenges and priorities

Biodiversity informatics plays a central enabling role in the research community's efforts to address scientific conservation and sustainability issues. Great strides have been made in the past decade establishing a framework for sharing data, where taxonomy and systematics has been perceived as the most prominent discipline involved. To some extent this is inevitable, given the use of species names as the pivot around which information is organised. To address the urgent questions around conservation, land-use, environmental change, sustainability, food security and ecosystem services that are facing Governments worldwide, we need to understand how the ecosystem works. So, we need a systems approach to understanding biodiversity that moves significantly beyond taxonomy and species observations. Such an approach needs to look at the whole system to address species interactions, both with their environment and with other species. It is clear that some barriers to progress are sociological, basically persuading people to use the technological solutions that are already available. This is best addressed by developing more effective systems that deliver immediate benefit to the user, hiding the majority of the technology behind simple user interfaces. An infrastructure should be a space in which activities take place and, as such, should be effectively invisible. This community consultation paper positions the role of biodiversity informatics, for the next decade, presenting the actions needed to link the various biodiversity infrastructures invisibly and to facilitate understanding that can support both business and policy-makers. The community considers the goal in biodiversity informatics to be full integration of the biodiversity research community, including citizens’ science, through a commonly-shared, sustainable e-infrastructure across all sub-disciplines that reliably serves science and society alike

Depleted dissolved organic carbon and distinct bacterial communities in the water column of a rapid-flushing coral reef ecosystem

Author Posting. © The Author(s), 2011. This is the author's version of the work. It is posted here by permission of Nature Publishing Group for personal use, not for redistribution. The definitive version was published in The ISME Journal 5 (2011): 1374–1387, doi:10.1038/ismej.2011.12.Coral reefs are highly productive ecosystems bathed in unproductive, low-nutrient oceanic waters, where microbially-dominated food webs are supported largely by bacterioplankton recycling of dissolved compounds. Despite evidence that benthic reef organisms efficiently scavenge particulate organic matter and inorganic nutrients from advected oceanic waters, our understanding of the role of bacterioplankton and dissolved organic matter in the interaction between reefs and the surrounding ocean remains limited. Here we present the results of a four-year study conducted in a well-characterized coral reef ecosystem (Paopao Bay, Moorea, French Polynesia) where changes in bacterioplankton abundance and dissolved organic carbon (DOC) concentrations were quantified and bacterial community structure variation was examined along spatial gradients of the reef:ocean interface. Our results illustrate that the reef is consistently depleted in concentrations of both DOC and bacterioplankton relative to offshore waters (averaging 79 µmol L-1 DOC and 5.5 X 108 cells L-1 offshore and 68 µmol L-1 DOC and 3.1 X 108 cells L-1 over the reef, respectively) across a four year time period. In addition, using a suite of culture-independent measures of bacterial community structure, we found consistent differentiation of reef bacterioplankton communities from those offshore or in a nearby embayment across all taxonomic levels. Reef habitats were enriched in Gamma-, Delta-, and Beta-proteobacteria, Bacteriodetes, Actinobacteria and Firmicutes. Specific bacterial phylotypes, including members of the SAR11, SAR116, Flavobacteria, and Synechococcus clades, exhibited clear gradients in relative abundance among nearshore habitats. Our observations indicate that this reef system removes oceanic DOC and exerts selective pressures on bacterioplankton community structure on timescales approximating reef water residence times, observations which are notable both because fringing reefs do not exhibit long residence times (unlike those characteristic of atoll lagoons) and because oceanic DOC is generally recalcitrant to degradation by ambient microbial assemblages. Our findings thus have interesting implications for the role of oceanic DOM and bacterioplankton in the ecology and metabolism of reef ecosystems.This project was supported by the US National Science Foundation Moorea Coral Reef Long Term Ecological Research project (NSF OCE-0417412) through minigrants to CAC and NSF OCE-0927411 to CAC as well as the MIRADA-LTERs program (NSF DEB-0717390 to LAZ)