Temperate Coastal Microbial Communities Rapidly Respond to Low Concentrations of Partially Weathered Diesel.

Diesel is frequently encountered in coastal ecosystems due to land run-off from road surfaces. The current study investigates how partially weathered diesel at environmentally relevant concentrations, as may be seen during a run-off event, affect coastal microbial communities. A mesocosm experiment using seawater from the Bedford Basin, Nova Scotia, was followed for 72 h after the addition of partially weathered diesel. Sequencing data suggests partially weathered diesel acts quickly to alter the prokaryotic community, as both opportunistic (Vibrio and Lentibacter) and oil-degrading (Colwellia, Sulfitobacter, and Pseudoalteromonas) bacteria proliferated after 24 h in comparison to the control. In addition, total prokaryotes seemed to recover in abundance after 24 h, where eukaryotes only ceased to decrease slightly at 72 h, likely because of an inability to adapt to the oil-laden conditions, unlike the prokaryotes. Considering there were no highly volatile components (benzene, toluene, ethylbenzene, and xylene) present in the diesel when the communities were exposed, the results indicate that even a relatively small concentration of diesel run-off can cause a drastic change to the microbial community under low energy conditions. Higher energy conditions due to wave action may mitigate the response of the microbial communities by dilution and additional weathering of the diesel.

River Flow Impacts Bacterial and Archaeal Community Structure in Surface Sediments in the Northern Gulf of Mexico.

Meiobenthic community structure in the northern Gulf of Mexico has been shown to be driven by geographical differences due to inshore-offshore gradients and location relative to river discharge. Samples collected along three transects spanning Mobile Bay, Alabama, showed significant differences in meiobenthic communities east of the bay compared to those sampled from the west. In contrast, analysis of bacterial and archaeal communities from the same sediment samples shows that the inshore-offshore gradient has minimal impact on their community structure. Significant differences in community structure were observed for Bacteria and Archaea between the east and west samples, but there was no difference in richness or diversity. Grouped by sediment type, higher richness was observed in silty samples compared to sandy samples. Significant differences were also observed among sediment types for community structure with bacteria communities in silty samples having more anaerobic sulfate reducers compared to aerobic heterotrophs, which had higher abundances in sandy sediments. This is likely due to increased organic matter in the silty sediments from the overlying river leading to low oxygen habitats. Most archaeal sequences represented poorly characterized high-level taxa, limiting interpretation of their distributions. Overlap between groups based on transect and sediment characteristics made determining which factor is more important in structuring bacterial and archaeal communities difficult. However, both factors are driven by discharge from the Mobile River. Although inshore-offshore gradients do not affect Bacteria or Archaea to the same extent as the meiobenthic communities, all three groups are strongly affected by sediment characteristics.

Metabarcoding reveals environmental factors influencing spatio-temporal variation in pelagic micro-eukaryotes.

Marine environments harbour a vast diversity of micro-eukaryotic organisms (protists and other small eukaryotes) that play important roles in structuring marine ecosystems. However, micro-eukaryote diversity is not well understood. Likewise, knowledge is limited regarding micro-eukaryote spatial and seasonal distribution, especially over long temporal scales. Given the importance of this group for mobilizing energy from lower trophic levels near the base of the food chain to larger organisms, assessing community stability, diversity and resilience is important to understand ecosystem health. Herein, we use a metabarcoding approach to examine pelagic micro-eukaryote communities over a 2.5-year time series. Bimonthly surface sampling (July 2009 to December 2011) was conducted at four locations within Mobile Bay (Bay) and along the Alabama continental shelf (Shelf). Alpha-diversity only showed significant differences in Shelf sites, with the greatest differences observed between summer and winter. Beta-diversity showed significant differences in community composition in relation to season and the Bay was dominated by diatoms, while the Shelf was characterized by dinoflagellates and copepods. The northern Gulf of Mexico is heavily influenced by the Mobile River Basin, which brings low-salinity nutrient-rich water mostly during winter and spring. Community composition was correlated with salinity, temperature and dissolved silicate. However, species interactions (e.g. predation and parasitism) may also contribute to the observed variation, especially on the Shelf, which warrants further exploration. Metabarcoding revealed clear patterns in surface pelagic micro-eukaryote communities that were consistent over multiple years, demonstrating how these techniques could be greatly beneficial to ecological monitoring and management over temporal scales.

Leveraging marine biotechnology for an All-Atlantic sustainable blue economy.

Despite the lack of research, development, and innovation funds, especially in South Atlantic countries, the Atlantic is suited to supporting a sustainable marine bioeconomy. Novel low-carbon mariculture systems can provide food security, new drugs, and climate mitigation. We suggest how to develop this sustainable marine bioeconomy across the entire Atlantic.

Environmental Drivers of Vibrio cholerae Abundances in Mobile Bay, Alabama.

Vibrio cholerae is the etiological agent of the illness cholera. However, there are non-O1/non-O139 V. cholerae (NOVC) strains that generally lack the toxin gene (ctx) and colonization factors that cause cholera. These NOVC strains are autochthonous members of estuarine environments and a significant cause of seafood-borne gastroenteritis in the United States. The objective of this study was to identify environmental parameters that correlate with NOVC prevalence in oysters, water, and sediment at three ecologically diverse locations in Mobile Bay, AL, including Dog River (DR), Fowl River (FR), and Cedar Point (CP). Oyster, water, and sediment samples were collected twice a month when conditions were favorable for NOVC growth and once a month when they were not. A most probable number (MPN)/real-time PCR assay was used to determine NOVC abundances. Environmental parameters were measured during sampling to determine their relationship, if any, with NOVC at each site. NOVC abundances in oysters at DR, FR, and CP were 0.87, 0.87, and -0.13 log MPN/g, respectively. In water, the median NOVC levels at DR, FR, and CP were 1.18, -0.13, and -0.82 log MPN/mL, and in sediment, the levels were 1.48, 1.87, and -0.03 log MPN/g, respectively. Correlations of NOVC abundances in oyster, water, and sediment samples with environmental parameters were largely site specific. For example, the levels of NOVC in oysters at DR had a positive correlation with temperature but a negative correlation with dissolved oxygen (DO) and nutrient concentrations, NO2-, NO3-, dissolved inorganic nitrogen (DIN), total dissolved nitrogen (TDN), and dissolved inorganic phosphorus (DIP). At FR, however, the levels of NOVC in oysters displayed only a negative correlation with NO2-. When grouping NOVC abundances by temperature, the main driving factor for prevalence, additional correlations with salinity, total cell counts, dissolved organic nitrogen (DON), and dissolved organic carbon (DOC) became evident regardless of the site. IMPORTANCE NOVC can cause gastrointestinal illness in humans, which typically occurs after the consumption of raw or undercooked seafood. Incidence rates of NOVC gastroenteritis have increased during the past decade. In this study, NOVC was enumerated from oysters, sediment, and water collected at three sites in Mobile Bay, with environmental parameters measured concurrently over the course of a year, to identify potential environmental drivers of NOVC abundances. The data from this study, from an area lacking in V. cholerae research, provide a useful baseline for risk analysis of V. cholerae infections. Defining correlations between NOVC and environmental attributes at different sites and temperatures within a dynamic system such as Mobile Bay provides valuable data to better understand the occurrence and proliferation of V. cholerae in the environment.

Factors that affect water column hydrocarbon concentrations have minor impacts on microbial responses following simulated diesel fuel spills.

Effects of season and mixing on hydrocarbon concentrations and the microbial community response was explored in a series of mesocosm experiments simulating surface spills of diesel into coastal waters. Mixing of any amount contributed to hydrocarbons entering the water column, but diesel fuel composition had a significant effect on hydrocarbon concentrations. Higher initial concentrations of aromatic hydrocarbons resulted in higher water column concentrations, with minimal differences among seasons due to high variability. Regardless of the concentrations of hydrocarbons, prokaryotes increased and there were higher relative abundances of hydrocarbon affiliated bacteria with indications of biodegradation within 4 d of exposure. As concentrations decreased over time, the eukaryote community shifted from the initial community to one which appeared to be composed of organisms with some resilience to hydrocarbons. This series of experiments demonstrates the wide range of conditions under which natural attenuation of diesel fuel is an effective response.

Proteomic analysis of Sulfolobus solfataricus during Sulfolobus Turreted Icosahedral Virus infection.

Where there is life, there are viruses. The impact of viruses on evolution, global nutrient cycling, and disease has driven research on their cellular and molecular biology. Knowledge exists for a wide range of viruses; however, a major exception are viruses with archaeal hosts. Archaeal virus-host systems are of great interest because they have similarities to both eukaryotic and bacterial systems and often live in extreme environments. Here we report the first proteomics-based experiments on archaeal host response to viral infection. Sulfolobus Turreted Icosahedral Virus (STIV) infection of Sulfolobus solfataricus P2 was studied using 1D and 2D differential gel electrophoresis (DIGE) to measure abundance and redox changes. Cysteine reactivity was measured using novel fluorescent zwitterionic chemical probes that, together with abundance changes, suggest that virus and host are both vying for control of redox status in the cells. Proteins from nearly 50% of the predicted viral open reading frames were found along with a new STIV protein with a homologue in STIV2. This study provides insight to features of viral replication novel to the archaea, makes strong connections to well-described mechanisms used by eukaryotic viruses such as ESCRT-III mediated transport, and emphasizes the complementary nature of different omics approaches.

Microbial community response to simulated diluted bitumen spills in coastal seawater and implications for oil spill response.

Oil spills in coastal waters can have devastating impacts on local ecosystems, from the microscopic base through to mammals and seabirds. Increasing transport of diluted bitumen has led to concerns about how this novel product might impact coastal ecosystems. A mesocosm study determined that the type of diluent and the season can affect the concentrations of hydrocarbons entering the water column from a surface spill. Those same mesocosms were sampled to determine whether diluent type and season also affected the microbial response to a surface spill. Overall, there were no differences in impacts among the three types of diluted bitumen, but there were consistent responses to all products within each season. Although microbial abundances with diluted bitumen rarely differed from unoiled controls, community structure in these organisms shifted in response to hydrocarbons, with hydrocarbon-degrading bacteria becoming more abundant. The relative abundance of heterotrophic eukaryotes also increased with diluted bitumen, with few photosynthetic organisms responding positively to oil. Overall shifts in the microbial communities were minimal relative to spills of conventional oil products, with low concentrations of hydrocarbons in the water column. Oil spill response should focus on addressing the surface slick to prevent sinking or stranding to minimize ecosystem impacts.

The architecture and chemical stability of the archaeal Sulfolobus turreted icosahedral virus.

Viruses utilize a diverse array of mechanisms to deliver their genomes into hosts. While great strides have been made in understanding the genome delivery of eukaryotic and prokaryotic viruses, little is known about archaeal virus genome delivery and the associated particle changes. The Sulfolobus turreted icosahedral virus (STIV) is a double-stranded DNA (dsDNA) archaeal virus that contains a host-derived membrane sandwiched between the genome and the proteinaceous capsid shell. Using cryo-electron microscopy (cryo-EM) and different biochemical treatments, we identified three viral morphologies that may correspond to biochemical disassembly states of STIV. One of these morphologies was subtly different from the previously published 27-A-resolution electron density that was interpreted with the crystal structure of the major capsid protein (MCP). However, these particles could be analyzed at 12.5-A resolution by cryo-EM. Comparing these two structures, we identified the location of multiple proteins forming the large turret-like appendages at the icosahedral vertices, observed heterogeneous glycosylation of the capsid shell, and identified mobile MCP C-terminal arms responsible for tethering and releasing the underlying viral membrane to and from the capsid shell. Collectively, our studies allow us to propose a fusogenic mechanism of genome delivery by STIV, in which the dismantled capsid shell allows for the fusion of the viral and host membranes and the internalization of the viral genome.

Particle assembly and ultrastructural features associated with replication of the lytic archaeal virus sulfolobus turreted icosahedral virus.

Little is known about the replication cycle of archaeal viruses. We have investigated the ultrastructural changes of Sulfolobus solfataricus P2 associated with infection by Sulfolobus turreted icosahedral virus (STIV). A time course of a near synchronous STIV infection was analyzed using both scanning and transmission electron microscopy. Assembly of STIV particles, including particles lacking DNA, was observed within cells, and fully assembled STIV particles were visible by 30 h postinfection (hpi). STIV was determined to be a lytic virus, causing cell disruption beginning at 30 hpi. Prior to cell lysis, virus infection resulted in the formation of pyramid-like projections from the cell surface. These projections, which have not been documented in any other host-virus system, appeared to be caused by the protrusion of the cell membrane beyond the bordering S-layer. These structures are thought to be sites at which progeny virus particles are released from infected cells. Based on these observations of lysis, a plaque assay was developed for STIV. From these studies we propose an overall assembly model for STIV.

Virus movement maintains local virus population diversity.

Viruses are the largest reservoir of genetic material on the planet, yet little is known about the population dynamics of any virus within its natural environment. Over a 2-year period, we monitored the diversity of two archaeal viruses found in hot springs within Yellowstone National Park (YNP). Both temporal phylogeny and neutral biodiversity models reveal that virus diversity in these local environments is not being maintained by mutation but rather by high rates of immigration from a globally distributed metacommunity. These results indicate that geographically isolated hot springs are readily able to exchange viruses. The importance of virus movement is supported by the detection of virus particles in air samples collected over YNP hot springs and by their detection in metacommunity sequencing projects conducted in the Sargasso Sea. Rapid rates of virus movement are not expected to be unique to these archaeal viruses but rather a common feature among virus metacommunities. The finding that virus immigration rather than mutation can dominate community structure has significant implications for understanding virus circulation and the role that viruses play in ecology and evolution by providing a reservoir of mobile genetic material.

Measuring the fate of different diluted bitumen products in coastal surface waters.

Diluted bitumens are produced by adding lower viscosity diluent to highly viscous bitumen to enable it to flow through pipelines and thus may behave differently than conventional oils when spilled into coastal seawater. Simulated surface spills using three different diluted bitumen products were carried out in May, July and November and water column hydrocarbons were monitored over a 14 day period. Volatile and total petroleum hydrocarbons varied in the water column depending on season and type of diluent. In summer, products diluted with synthetic crude or a mixture of condensate and crude released droplets into the water column. Diluted bitumen did not sink to the bottom of the enclosures with surface slicks showing a range of weathering after 14 d. With most of the diluted bitumen product remaining on the surface for 14 d, a rapid conventional clean up response may be effective in low energy, coastal waters.

Transcriptome analysis of infection of the archaeon Sulfolobus solfataricus with Sulfolobus turreted icosahedral virus.

Microarray analysis of infection by Sulfolobus turreted icosahedral virus (STIV) revealed insights into the timing and extent of virus transcription, as well as differential regulation of host genes. Using a microarray containing genes from both the host and the virus, the infection cycle of STIV was studied. Following infection of Sulfolobus solfataricus strain 2-2-12 with STIV, transcription of virus genes was first detected at 8 h postinfection (p.i.), with a peak at 24 h p.i. Lysis of cells was first detected at 32 h p.i. There was little temporal control of the transcription of virus genes, although the three open reading frames on the noncoding strand were transcribed later in the infection process. During the infection, 177 host genes were determined to be differentially expressed, with 124 genes up-regulated and 53 genes down-regulated. The up-regulated genes were dominated by genes associated with DNA replication and repair and those of unknown function, while the down-regulated genes, mostly detected at 32 h p.i., were associated with energy production and metabolism. Examination of infected cells by transmission electron microscopy revealed alterations in cell ultrastructure consistent with the microarray analysis. The observed patterns of transcription suggest that up-regulated genes are likely used by the virus to reprogram the cell for virus replication, while the down-regulated genes reflect the imminent lysis of the cells.

Hot crenarchaeal viruses reveal deep evolutionary connections.

The discovery of archaeal viruses provides insights into the fundamental biochemistry and evolution of the Archaea. Recent studies have identified a wide diversity of archaeal viruses within the hot springs of Yellowstone National Park and other high-temperature environments worldwide. These viruses are often morphologically unique and code for genes with little similarity to other known genes in the biosphere, a characteristic that has complicated efforts to trace their evolutionary history. Comparative genomics combined with structural analysis indicate that spindle-shaped virus lineages might be unique to the Archaea, whereas other icosahedral viruses might share a common lineage with viruses of Bacteria and Eukarya. These studies provide insights into the evolutionary history of viruses in all three domains of life.

Determination of virus abundance by epifluorescence microscopy.

Determination of virus abundance using epifluorescence microscopy is a rapid and accurate method. The protocol requires the concentration of virus particles by collection on a filter. The nucleic acid in the virus particles is then stained with a fluorescent stain and the sample viewed with an epifluorescence microscope. The method was originally developed to determine the abundance of virus particles in water samples, however the protocol has been adapted for cultures and sediment samples. Although the method provides total counts of all virus-sized particles, regardless of infectivity, the method can be used for rapidly screening samples for further study.

Inorganic nutrients have a significant, but minimal, impact on a coastal microbial community's response to fresh diluted bitumen.

Microbes capable of degrading hydrocarbons in oil are present in low abundances in coastal waters, but quickly respond to oil following a spill. When estimating potential biodegradation rates in the laboratory, high concentrations of inorganic nutrients are often added to prevent nutrient limitation. In this study, we tested the short term response of coastal microbes to fresh diluted bitumen under varying nutrient conditions in a cold water regime. Total hydrocarbon concentrations changed minimally over five days; however, oil composition changed over time and the abundance of microbes increased in all treatments. Addition of phosphate, with or without nitrogen, resulted in rapid changes in community composition, but after three days treatments no longer differed. Nutrients were never depleted in any treatment suggesting that, even at low inorganic nutrient concentrations, microbial communities can quickly respond to hydrocarbons following a spill.

Spatial and temporal patterns in the Pelagibacteraceae across an estuarine gradient.

Marine bacterial communities show strong spatial and seasonal patterns, often characterized by changes at high taxonomic levels. The Pelagibacteraceae are common members of bacterial communities, with well-documented biogeography at the subclade level. To identify patterns within the subclades, the abundance and diversity of Pelagibacteraceae were analyzed over a two-year period at four stations across an estuarine gradient. Pelagibacteraceae was the most abundant bacterial family, averaging 27% of the community, but varying from 1% to 57% in any one sample. Highest abundances were detected in autumn and winter. Pelagibacteraceae richness was lowest at the most inshore site, and highest in autumn and winter at all sites. Shannon diversity decreased in winter, when a few OTUs dominated the community. Dissolved oxygen, dissolved silicate and prokaryote abundance explained most of the variability in the Pelagibacteraceae communities, with salinity differentiating low salinity communities. The 10 most abundant OTUs included OTUs that varied across sites, with little seasonality as well as those with small site effects, but strong seasonal patterns indicating differences in the niches of individual OTUs. While salinity was important in structuring low salinity communities, higher salinity communities appear to be responding to additional environmental parameters including oxygen, nutrients and other organisms.

Genetic assessment of meiobenthic community composition and spatial distribution in coastal sediments along northern Gulf of Mexico.

Meiobenthic (meiofauna and micro-eukaryotes) organisms are important contributors to ecosystem functioning in aquatic environments through their roles in nutrient transport, sediment stability, and food web interactions. Despite their ecological importance, information pertaining to variation of these communities at various spatial and temporal scales is not widely known. Many studies in the Gulf of Mexico (GOM) have focused either on deep sea or continental shelf areas, while little attention has been paid to bays and coastal regions. Herein, we take a holistic approach by using high-throughput sequencing approaches to examine spatial variation in meiobenthic communities within Alabama bays and the coastal northern GOM region. Sediment samples were collected along three transects (Mississippi Sound: MS, FOCAL: FT, and Orange Beach: OB) from September 2010 to April 2012 and community composition was determined by metabarcoding the V9 hypervariable region of the nuclear18S rRNA gene. Results showed that Stramenopiles (diatoms), annelids, arthropods (copepods), and nematodes were the dominate groups within samples, while there was presence of other phyla throughout the dataset. Location played a larger role than time sampled in community composition. However, samples were collected over a short temporal scale. Samples clustered in reference to transect, with the most eastern transect (OB) having a distinct community composition in comparison to the other two transects (MS and FT). Communities also differed in reference to region (Bay versus Shelf). Bulk density and percent inorganic carbon were the only measured environmental factors that were correlated with community composition.

Initial community and environment determine the response of bacterial communities to dispersant and oil contamination.

Bioremediation of seawater by natural bacterial communities is one potential response to coastal oil spills, but the success of the approach may vary, depending on geographical location, oil composition and the timing of spill. The short term response of coastal bacteria to dispersant, oil and dispersed oil was characterized using 16S rRNA gene tags in two mesocosm experiments conducted two months apart. Despite differences in the amount of oil-derived alkanes across the treatments and experiments, increases in the contributions of hydrocarbon degrading taxa and decreases in common estuarine bacteria were observed in response to dispersant and/or oil. Between the two experiments, the direction and rates of changes in particulate alkane concentrations differed, as did the magnitude of the bacterial response to oil and/or dispersant. Together, our data underscore large variability in bacterial responses to hydrocarbon pollutants, implying that bioremediation success varies with starting biological and environmental conditions.