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1.  Variation in spatial and temporal incidence of the crustacean pathogen Hematodinium perezi in environmental samples from Atlantic Coastal Bays 
Aquatic Biosystems  2013;9:11.
Hematodinium perezi, a parasitic dinoflagellate, infects and kills blue crabs, Callinectes sapidus, along the Atlantic and Gulf coasts of the United States. The parasite proliferates within host hemolymph and tissues, and also produces free-swimming biflagellated dinospores that emerge from infected crabs. Infections in C. sapidus recur annually, and it is not known if biotic or environmental reservoirs contribute to reinfection and outbreaks. To address this data gap, a quantitative PCR assay based on the internal transcribed spacer 2 (ITS2) region of H. perezi rRNA genes was developed to asses the temporal and spatial incidence of the parasite in Delaware and Maryland coastal bays.
A previously-used PCR assay for H. perezi, based on the small subunit rRNA gene sequence, was found to lack adequate species specificity to discriminate non-Hematodinium sp. dinoflagellate species in environmental samples. A new ITS2-targeted assay was developed and validated to detect H. perezi DNA in sediment and water samples using E. coli carrying the H. perezi rDNA genes. Application of the method to environmental samples identified potential hotspots in sediment in Indian River Inlet, DE and Chincoteague Bay, MD and VA. H. perezi DNA was not detected in co-occurring shrimp or snails, even during an outbreak of the parasite in C. sapidus.
H. perezi is present in water and sediment samples in Maryland and Delaware coastal bays from April through November with a wide spatial and temporal variability in incidence. Sampling sites with high levels of H. perezi DNA in both bays share characteristics of silty, organic sediments and low tidal currents. The environmental detection of H. perezi in spring, ahead of peak prevalence in crabs, points to gaps in our understanding of the parasite’s life history prior to infection in crabs as well as the mode of environmental transmission. To better understand the H. perezi life cycle will require further monitoring of the parasite in habitats as well as hosts. Improved understanding of potential environmental transmission to crabs will facilitate the development of disease forecasting.
PMCID: PMC3651331  PMID: 23641869
Blue crab; Hematodinium; Parasite; Disease reservoir; Fishery
2.  New Host Range for Hematodinium in Southern Australia and Novel Tools for Sensitive Detection of Parasitic Dinoflagellates 
PLoS ONE  2013;8(12):e82774.
Hematodinium is a parasitic dinoflagellate and emerging pathogen of crustaceans. It preferably manifests in haemolymph of marine decapod crustaceans, killing a large variety of genera with significant impacts on fisheries worldwide. There is, however, evidence that some crustacean stocks harbor high prevalence, low intensity infections that may not result in widespread host mortality and are therefore hard to detect. The most widely used methods for detection of Hematodinium are conventional blood smears and polymerase chain reaction (PCR) against ribosomal RNAs. Blood smears demand a trained investigator, are labor intensive and not readily scalable for high-throughput sampling. PCRs only detect parasite DNA and can also suffer from false negatives and positives. In order to develop alternative detection tools for Hematodinium cells in decapod crustaceans we employed an immunological approach against a newly identified, abundant dinoflagellate-specific nuclear protein—Dinoflagellate/Viral NucleoProtein (DVNP). Both immunofluorescence assay (IFA) and Western blot methods against DVNP showed high sensitivity of detection. The Western blot detects Hematodinium parasites to levels of 25 parasites per milliliter of crustacean haemolymph, with the potential for sample pooling and screening of large samples. Using both PCR and these new tools, we have identified Hematodinium cells present in three new host crab taxa, at high prevalence but with no sign of pathogenesis. This extends the known range of Hematodinium to southern Australia.
PMCID: PMC3855790  PMID: 24324829
3.  Hematodinium sp. and its bacteria-like endosymbiont in European brown shrimp (Crangon crangon) 
Aquatic Biosystems  2012;8:24.
Parasitic dinoflagellates of the genus Hematodinium are significant pathogens affecting the global decapod crustacean fishery. Despite this, considerable knowledge gaps exist regarding the life history of the pathogen in vivo, and the role of free living life stages in transmission to naïve hosts.
In this study, we describe a novel disease in European brown shrimp (Crangon crangon) caused by infection with a parasitic dinoflagellate of the genus Hematodinium. This is the second example host within the Infraorder Caridea (shrimp) and significantly, the first description within the superfamily Crangonoidea. Based upon analysis of the rRNA gene (SSU) and spacers (ITS1), the parasite in C. crangon is the same as that previously described infecting Nephrops norvegicus and Cancer pagurus from European seas, and to the parasite infecting several other commercially important crab species in the Northern Hemisphere. The parasite is however distinct from the type species, H. perezi, found infecting type hosts (Carcinus maenas and Liocarcinus depurator) from nearby sites within Europe. Despite these similarities, the current study has also described for the first time, a bacteria-like endosymbiont within dinospore stages of the parasite infecting shrimp. The endosymbionts were either contained individually within electron lucent vacuoles within the parasite cell cytoplasm, or remained in direct contact with the parasite cytoplasm or in some cases, the nucleoplasm. In all of these cases, no apparent detrimental effects of colonization were observed within the parasite cell.
The presence of bacteria-like endosymbionts within dinospore life stages presumes that the relationship between the dinoflagellate and the bacteria is extended beyond the period of liberation of spores from the infected host shrimp. In this context, a potential role of endosymbiosis in the survival of free-living stages of the parasite is possible. The finding offers a further intriguing insight into the life history of this enigmatic pathogen of marine crustacean hosts and highlights a potential for mixotrophy in the parasitic dinoflagellates contained within the genus Hematodinium.
PMCID: PMC3487949  PMID: 22958655
ITS1; Phylogenetics; Dinoflagellate; Bacteria; Crustacean; Disease; Fishery
4.  Characterization and Molecular Epidemiology of a Fungal Infection of Edible Crabs (Cancer pagurus) and Interaction of the Fungus with the Dinoflagellate Parasite Hematodinium 
This study reports on an emerging fungal disease of the edible crab, Cancer pagurus. Juvenile (prerecruit) crabs were found to be subject to this disease condition during the months of May to September at two intertidal sites in South Wales, United Kingdom. Histopathology revealed that the fungi overwhelm the host response in the tissues, leading to progressive septicemia. The causative agent of this infection was isolated and grown in pure culture and was identified as a member of the Ophiocordyceps clade by sequencing of the small subunit of the fungal ribosomal DNA (rDNA). Of the crabs naturally infected with the fungus, 94% had a coinfection with the parasitic dinoflagellate Hematodinium species. To determine if there was any interaction between the two disease-causing agents, apparently fungus-free crabs, both with and without natural Hematodinium infections, were challenged with the fungal isolate. The presence of Hematodinium caused a significant reduction in fungal multiplication in the hemocoel of the crabs in comparison to that in Hematodinium-free individuals. Histopathology of coinfected crabs showed a systemic multiplication of Hematodinium within host tissues, leading to a rapid death, while Hematodinium-free crabs experimentally infected with the fungal isolate died due to fungal sepsis (septicemia) with the same characteristic pathology as seen in natural infections.
PMCID: PMC3568540  PMID: 23160130
5.  Mosquitoes Associated with Ditch-Plugged and Control Tidal Salt Marshes on the Delmarva Peninsula 
A study was conducted during the summer of 2009 (from July to September) to characterize mosquito communities among different habitats in five historically ditched tidal salt marshes and three adjacent wooded areas in the E.A. Vaughn Wetland Management Area on the Maryland Delmarva Peninsula, USA. Study marshes are characteristic of Atlantic coastal salt marshes that had undergone grid ditching from the 1930s to 1950s. In the autumn of 2008 (October and November) ditches were plugged near their outlets in two (‘experimental’) marshes with the aim to restore their natural tidal hydrology. The three other marshes were not plugged. Marshes were sampled from July to September in 2009 by using standard dip count method. A total of 2,457 mosquito larvae representing six species were collected on 15.4% (86/557) of all sample occasions and 399 adults representing four mosquito species were collected from landing counts. Aedes sollicitans, Anopheles bradleyi and Culex salinarius were the most common species collected in larval habitats, and Ae. sollicitans was the most common adult collected. Wooded habitats had more total mosquitoes, were also more frequently occupied by mosquitoes and had higher densities of mosquitoes than marsh habitats. Almost all larvae collected from marshes were from one experimental and one control site. The majority of larvae at the control site were Ae. sollicitans in marsh pannes while Cx. salinarius, An. bradleyi, Ae. cantator, and Ae. sollicitans were collected in high numbers from ditches at the experimental site. We found a difference in the proportion of marsh pannes occupied by Ae. sollicitans but not total mosquitoes sampled 4–5 days after spring tide events than on other occasions. Salinity measures of 42 larval habitats showed lower median salinity in mosquito-occupied habitats (11.5 ppt) than unoccupied habitats (20.1 ppt), and in habitats in wooded areas followed by ditches and pannes in marsh areas. The results of this study suggest that wooded areas adjacent to salt marshes may be a substantial source of biting adult mosquitoes usually associated with salt marsh habitats and that ditch plugging may alter the productivity of mosquitoes on some marshes. We recommend future studies consider mosquito productivity from habitats surrounding salt marshes, and if assessments of marsh alterations are a goal, compare multiple experimental and control areas before and after treatments to determine if alterations have a consistent impact on regional mosquito production.
PMCID: PMC3166729  PMID: 21909293
ditch plugging; Delmarva Peninsula; tidal salt marsh; restoration; salinity
6.  Influence of Environmental Gradients on the Abundance and Distribution of Mycobacterium spp. in a Coastal Lagoon Estuary▿  
Applied and Environmental Microbiology  2009;75(23):7378-7384.
Environmental mycobacteria are of increasing concern in terms of the diseases they cause in both humans and animals. Although they are considered to be ubiquitous in aquatic environments, few studies have examined their ecology, and no ecological studies of coastal marine systems have been conducted. This study uses indirect gradient analysis to illustrate the strong relationships that exists between coastal water quality and the abundance of Mycobacterium spp. within a U.S. mid-Atlantic embayment. Mycobacterium species abundance and water quality conditions (based on 16 physical and chemical variables) were examined simultaneously in monthly samples obtained at 18 Maryland and Virginia coastal bay stations from August 2005 to November 2006 (n = 212). A quantitative molecular assay for Mycobacterium spp. was evaluated and applied, allowing for rapid, direct enumeration. By using indirect gradient analysis (environmental principal-components analysis), a strong linkage between eutrophic conditions, characterized by low dissolved-oxygen levels and elevated nutrient concentrations, and mycobacteria was determined. More specifically, a strong nutrient response was noted, with all nitrogen components and turbidity measurements correlating positively with abundance (r values of >0.30; P values of <0.001), while dissolved oxygen showed a strong negative relationship (r = −0.38; P = 0.01). Logistic regression models developed using salinity, dissolved oxygen, and total nitrogen showed a high degree of concordance (83%). These results suggest that coastal restoration and management strategies designed to reduce eutrophication may also reduce total mycobacteria in coastal waters.
PMCID: PMC2786408  PMID: 19801457
7.  Development of a Denaturing High-Performance Liquid Chromatography Method for Detection of Protist Parasites of Metazoans▿ †  
Applied and Environmental Microbiology  2008;74(14):4336-4345.
Increasingly, diseases of marine organisms are recognized as significant biotic factors affecting ecosystem health. However, the responsible disease agents are often unknown and the discovery and description of novel parasites most often rely on morphological descriptions made by highly trained specialists. Here, we describe a new approach for parasite discovery, utilizing denaturing high-performance liquid chromatography (DHPLC) reverse-phase ion-paring technology. Systematic investigations of major DHPLC variables, including temperature, gradient conditions, and target amplicon characteristics were conducted to develop a mechanistic understanding of DNA fragment separation by DHPLC. As a model system, 18S rRNA genes from the blue crab (Callinectes sapidus) and a parasitic dinoflagellate Hematodinium sp. were used. Binding of 18S rRNA gene PCR amplicons to the DNA separation column in the presence of triethylammonium acetate (TEAA) was inversely correlated with temperature and could be predicted based on the estimated DNA helicity of the PCR amplicon. Amplicons of up to 498 bp were resolved as single chromatographic peaks if they had high (>95%) DNA helicity. Amplicons that differed by as few as 2 bp could be resolved. Separation of 18S rRNA gene PCR amplicons was optimized by simultaneous manipulation of both temperature and solvent gradients. The optimal conditions included targeting regions of high DNA helicity (>95%), temperatures in the range of 57 to 63°C, and a linear acetonitrile gradient from 13.75 to 17.5% acetonitrile in 0.1 M TEAA (55 to 70% buffer B) over a 9-min period. Under these conditions, amplicons from a variety of parasites and their hosts can be separated and detected by DHPLC.
PMCID: PMC2493188  PMID: 18502933
8.  Environmental Gradient Favours Functionally Diverse Macrobenthic Community in a Placer Rich Tropical Bay 
The Scientific World Journal  2013;2013:750580.
The present paper examines the functional diversity-environment relation in a placer rich tropical bay. Understanding the environmental variables that determine the biodiversity pattern will help in the effective conservation plans of coastal habitat. However, few studies have been carried out on the biodiversity-environment relation from the diverse tropical coastal ecosystem. The geographic location of Kalbadevi Bay along the west coast of India provides an opportunity to study the functional diversity pattern of macrofauna along an environmental gradient. Additionally, the area is also a potential placer mining site. Seasonal sampling was carried out for macrofauna and environmental variables. Macrofaunal functional diversity showed significant temporal variation related to the environmental parameters. The most important environmental variables were organic matter and sediment texture. Filter feeders dominated during postmonsoon which is a period when the water column is enriched with sinking detritus. The deposit feeders which rapidly ingest the settled detritus and also transport it to deeper sediment for the subsurface deposit feeders dominated during premonsoon. Abundance of carnivores was high during premonsoon, a response to increase in food in terms of deposit feeders. The result thus indicates that the temporal environmental variation influenced the macrofaunal functional diversity pattern in the Kalbadevi Bay.
PMCID: PMC3703323  PMID: 23853540
9.  Edible Crabs “Go West”: Migrations and Incubation Cycle of Cancer pagurus Revealed by Electronic Tags 
PLoS ONE  2013;8(5):e63991.
Crustaceans are key components of marine ecosystems which, like other exploited marine taxa, show seasonable patterns of distribution and activity, with consequences for their availability to capture by targeted fisheries. Despite concerns over the sustainability of crab fisheries worldwide, difficulties in observing crabs’ behaviour over their annual cycles, and the timings and durations of reproduction, remain poorly understood. From the release of 128 mature female edible crabs tagged with electronic data storage tags (DSTs), we demonstrate predominantly westward migration in the English Channel. Eastern Channel crabs migrated further than western Channel crabs, while crabs released outside the Channel showed little or no migration. Individual migrations were punctuated by a 7-month hiatus, when crabs remained stationary, coincident with the main period of crab spawning and egg incubation. Incubation commenced earlier in the west, from late October onwards, and brooding locations, determined using tidal geolocation, occurred throughout the species range. With an overall return rate of 34%, our results demonstrate that previous reluctance to tag crabs with relatively high-cost DSTs for fear of loss following moulting is unfounded, and that DSTs can generate precise information with regards life-history metrics that would be unachievable using other conventional means.
PMCID: PMC3667170  PMID: 23734180
10.  Seagrass Restoration Enhances “Blue Carbon” Sequestration in Coastal Waters 
PLoS ONE  2013;8(8):e72469.
Seagrass meadows are highly productive habitats that provide important ecosystem services in the coastal zone, including carbon and nutrient sequestration. Organic carbon in seagrass sediment, known as “blue carbon,” accumulates from both in situ production and sedimentation of particulate carbon from the water column. Using a large-scale restoration (>1700 ha) in the Virginia coastal bays as a model system, we evaluated the role of seagrass, Zosteramarina, restoration in carbon storage in sediments of shallow coastal ecosystems. Sediments of replicate seagrass meadows representing different age treatments (as time since seeding: 0, 4, and 10 years), were analyzed for % carbon, % nitrogen, bulk density, organic matter content, and 210Pb for dating at 1-cm increments to a depth of 10 cm. Sediment nutrient and organic content, and carbon accumulation rates were higher in 10-year seagrass meadows relative to 4-year and bare sediment. These differences were consistent with higher shoot density in the older meadow. Carbon accumulation rates determined for the 10-year restored seagrass meadows were 36.68 g C m-2 yr-1. Within 12 years of seeding, the restored seagrass meadows are expected to accumulate carbon at a rate that is comparable to measured ranges in natural seagrass meadows. This the first study to provide evidence of the potential of seagrass habitat restoration to enhance carbon sequestration in the coastal zone.
PMCID: PMC3743776  PMID: 23967303
11.  Quantitative PCR Assay for Mycobacterium pseudoshottsii and Mycobacterium shottsii and Application to Environmental Samples and Fishes from the Chesapeake Bay▿  
Applied and Environmental Microbiology  2010;76(18):6171-6179.
Striped bass (Morone saxatilis) in the Chesapeake Bay are currently experiencing a very high prevalence of mycobacteriosis associated with newly described Mycobacterium species, Mycobacterium pseudoshottsii and M. shottsii. The ecology of these mycobacteria outside the striped bass host is currently unknown. In this work, we developed quantitative real-time PCR assays for M. pseudoshottsii and M. shottsii and applied these assays to DNA extracts from Chesapeake Bay water and sediment samples, as well as to tissues from two dominant prey of striped bass, Atlantic menhaden (Brevoortia tyrannus) and bay anchovy (Anchoa mitchilli). Mycobacterium pseudoshottsii was found to be ubiquitous in water samples from the main stem of the Chesapeake Bay and was also present in water and sediments from the Rappahannock River, Virginia. M. pseudoshottsii was also detected in menhaden and anchovy tissues. In contrast, M. shottsii was not detected in water, sediment, or prey fish tissues. In conjunction with its nonpigmented phenotype, which is frequently found in obligately pathogenic mycobacteria of humans, this pattern of occurrence suggests that M. shottsii may be an obligate pathogen of striped bass.
PMCID: PMC2937484  PMID: 20656856
12.  Description of Two Species of Early Branching Dinoflagellates, Psammosa pacifica n. g., n. sp. and P. atlantica n. sp 
PLoS ONE  2012;7(6):e34900.
In alveolate evolution, dinoflagellates have developed many unique features, including the cell that has epicone and hypocone, the undulating transverse flagellum. However, it remains unclear how these features evolved. The early branching dinoflagellates so far investigated such as Hematodinium, Amoebophrya and Oxyrrhis marina differ in many ways from of core dinoflagellates, or dinokaryotes. Except those handful of well studied taxa, the vast majority of early branching dinoflagellates are known only by environmental sequences, and remain enigmatic. In this study we describe two new species of the early branching dinoflagellates, Psammosa pacifica n. g., n. sp. and P. atlantica n. sp. from marine intertidal sandy beach. Molecular phylogeny of the small subunit (SSU) ribosomal RNA and Hsp90 gene places Psammosa spp. as an early branch among the dinoflagellates. Morphologically (1) they lack the typical dinoflagellate epicone–hypocone structure, and (2) undulation in either flagella. Instead they display a mosaïc of dinokaryotes traits, i.e. (3) presence of bi-partite trychocysts; Oxyrrhis marina–like traits, i.e. (4) presence of flagellar hairs, (5) presence of two-dimensional cobweb scales ornamenting both flagella (6) transversal cell division; a trait shared with some syndineansand Parvilucifera spp. i.e. (7) a nucleus with a conspicuous nucleolus and condensed chromatin distributed beneath the nuclear envelope; as well as Perkinsus marinus -like features i.e. (8) separate ventral grooves where flagella emerge and (9) lacking dinoflagellate-type undulating flagellum. Notably Psammosa retains an apical complex structure, which is shared between perkinsids, colpodellids, chromerids and apicomplexans, but is not found in dinokaryotic dinoflagellates.
PMCID: PMC3377698  PMID: 22719825
13.  Analysis of bacterial and archaeal diversity in coastal microbial mats using massive parallel 16S rRNA gene tag sequencing 
The ISME Journal  2011;5(11):1701-1712.
Coastal microbial mats are small-scale and largely closed ecosystems in which a plethora of different functional groups of microorganisms are responsible for the biogeochemical cycling of the elements. Coastal microbial mats play an important role in coastal protection and morphodynamics through stabilization of the sediments and by initiating the development of salt-marshes. Little is known about the bacterial and especially archaeal diversity and how it contributes to the ecological functioning of coastal microbial mats. Here, we analyzed three different types of coastal microbial mats that are located along a tidal gradient and can be characterized as marine (ST2), brackish (ST3) and freshwater (ST3) systems. The mats were sampled during three different seasons and subjected to massive parallel tag sequencing of the V6 region of the 16S rRNA genes of Bacteria and Archaea. Sequence analysis revealed that the mats are among the most diverse marine ecosystems studied so far and consist of several novel taxonomic levels ranging from classes to species. The diversity between the different mat types was far more pronounced than the changes between the different seasons at one location. The archaeal community for these mats have not been studied before and revealed a strong reaction on a short period of draught during summer resulting in a massive increase in halobacterial sequences, whereas the bacterial community was barely affected. We concluded that the community composition and the microbial diversity were intrinsic of the mat type and depend on the location along the tidal gradient indicating a relation with salinity.
PMCID: PMC3197164  PMID: 21544102
microbial diversity; microbial mats; 16S tag sequencing
14.  Dynamics of the Methanogenic Archaea in Tropical Estuarine Sediments 
Archaea  2013;2013:582646.
Methanogenesis may represent a key process in the terminal phases of anaerobic organic matter mineralization in sediments of coastal lagoons. The aim of the present work was to study the temporal and spatial dynamics of methanogenic archaea in sediments of tropical coastal lagoons and their relationship with environmental changes in order to determine how these influence methanogenic community. Sediment samples were collected during the dry (February, May, and early June) and rainy seasons (July, October, and November). Microbiological analysis included the quantification of viable methanogenic archaea (MA) with three substrates and the evaluation of kinetic activity from acetate in the presence and absence of sulfate. The environmental variables assessed were temperature, pH, Eh, salinity, sulfate, solids content, organic carbon, and carbohydrates. MA abundance was significantly higher in the rainy season (106–107 cells/g) compared with the dry season (104–106 cells/g), with methanol as an important substrate. At spatial level, MA were detected in the two layers analyzed, and no important variations were observed either in MA abundance or activity. Salinity, sulfate, solids, organic carbon, and Eh were the environmental variables related to methanogenic community. A conceptual model is proposed to explain the dynamics of the MA.
PMCID: PMC3562609  PMID: 23401664
15.  Uncertainty in Model Predictions of Vibrio vulnificus Response to Climate Variability and Change: A Chesapeake Bay Case Study 
PLoS ONE  2014;9(5):e98256.
The effect that climate change and variability will have on waterborne bacteria is a topic of increasing concern for coastal ecosystems, including the Chesapeake Bay. Surface water temperature trends in the Bay indicate a warming pattern of roughly 0.3–0.4°C per decade over the past 30 years. It is unclear what impact future warming will have on pathogens currently found in the Bay, including Vibrio spp. Using historical environmental data, combined with three different statistical models of Vibrio vulnificus probability, we explore the relationship between environmental change and predicted Vibrio vulnificus presence in the upper Chesapeake Bay. We find that the predicted response of V. vulnificus probability to high temperatures in the Bay differs systematically between models of differing structure. As existing publicly available datasets are inadequate to determine which model structure is most appropriate, the impact of climatic change on the probability of V. vulnificus presence in the Chesapeake Bay remains uncertain. This result points to the challenge of characterizing climate sensitivity of ecological systems in which data are sparse and only statistical models of ecological sensitivity exist.
PMCID: PMC4038616  PMID: 24874082
16.  Global Patterns of Bacterial Beta-Diversity in Seafloor and Seawater Ecosystems 
PLoS ONE  2011;6(9):e24570.
Marine microbial communities have been essential contributors to global biomass, nutrient cycling, and biodiversity since the early history of Earth, but so far their community distribution patterns remain unknown in most marine ecosystems.
Methodology/Principal Findings
The synthesis of 9.6 million bacterial V6-rRNA amplicons for 509 samples that span the global ocean's surface to the deep-sea floor shows that pelagic and benthic communities greatly differ, at all taxonomic levels, and share <10% bacterial types defined at 3% sequence similarity level. Surface and deep water, coastal and open ocean, and anoxic and oxic ecosystems host distinct communities that reflect productivity, land influences and other environmental constraints such as oxygen availability. The high variability of bacterial community composition specific to vent and coastal ecosystems reflects the heterogeneity and dynamic nature of these habitats. Both pelagic and benthic bacterial community distributions correlate with surface water productivity, reflecting the coupling between both realms by particle export. Also, differences in physical mixing may play a fundamental role in the distribution patterns of marine bacteria, as benthic communities showed a higher dissimilarity with increasing distance than pelagic communities.
This first synthesis of global bacterial distribution across different ecosystems of the World's oceans shows remarkable horizontal and vertical large-scale patterns in bacterial communities. This opens interesting perspectives for the definition of biogeographical biomes for bacteria of ocean waters and the seabed.
PMCID: PMC3169623  PMID: 21931760
17.  Development of a 3D Coupled Physical-Biogeochemical Model for the Marseille Coastal Area (NW Mediterranean Sea): What Complexity Is Required in the Coastal Zone? 
PLoS ONE  2013;8(12):e80012.
Terrestrial inputs (natural and anthropogenic) from rivers, the atmosphere and physical processes strongly impact the functioning of coastal pelagic ecosystems. The objective of this study was to develop a tool for the examination of these impacts on the Marseille coastal area, which experiences inputs from the Rhone River and high rates of atmospheric deposition. Therefore, a new 3D coupled physical/biogeochemical model was developed. Two versions of the biogeochemical model were tested, one model considering only the carbon (C) and nitrogen (N) cycles and a second model that also considers the phosphorus (P) cycle. Realistic simulations were performed for a period of 5 years (2007–2011). The model accuracy assessment showed that both versions of the model were able of capturing the seasonal changes and spatial characteristics of the ecosystem. The model also reproduced upwelling events and the intrusion of Rhone River water into the Bay of Marseille well. Those processes appeared to greatly impact this coastal oligotrophic area because they induced strong increases in chlorophyll-a concentrations in the surface layer. The model with the C, N and P cycles better reproduced the chlorophyll-a concentrations at the surface than did the model without the P cycle, especially for the Rhone River water. Nevertheless, the chlorophyll-a concentrations at depth were better represented by the model without the P cycle. Therefore, the complexity of the biogeochemical model introduced errors into the model results, but it also improved model results during specific events. Finally, this study suggested that in coastal oligotrophic areas, improvements in the description and quantification of the hydrodynamics and the terrestrial inputs should be preferred over increasing the complexity of the biogeochemical model.
PMCID: PMC3851166  PMID: 24324589
18.  No evidence of infection with avian influenza viruses among US poultry workers in the Delmarva Peninsula Maryland Virginia, USA 
Journal of agromedicine  2011;16(1):52-57.
Industrial poultry workers may be at elevated risk of avian influenza infection due to intense occupational contact with live poultry. Serum samples from poultry workers and community members in the Delmarva Peninsula, one of the densest regions of poultry production in the United States, were analyzed for antibodies to strains of five avian influenza subtypes using microneutralization assays. No evidence of infection was found, suggesting inefficient transmission to humans or the absence of virus in these premises. Continued serological surveillance of workers in industrial food animal facilities is necessary to prevent the transmission of influenza A viruses.
PMCID: PMC3041272  PMID: 21213164
Agricultural workers; influenza A viruses; avian; poultry; occupational exposure; zoonoses
19.  Recombination Shapes the Structure of an Environmental Vibrio cholerae Population ▿ †  
Vibrio cholerae consists of pathogenic strains that cause sporadic gastrointestinal illness or epidemic cholera disease and nonpathogenic strains that grow and persist in coastal aquatic ecosystems. Previous studies of disease-causing strains have shown V. cholerae to be a primarily clonal bacterial species, but isolates analyzed have been strongly biased toward pathogenic genotypes, while representing only a small sample of the vast diversity in environmental strains. In this study, we characterized homologous recombination and structure among 152 environmental V. cholerae isolates and 13 other putative Vibrio isolates from coastal waters and sediments in central California, as well as four clinical V. cholerae isolates, using multilocus sequence analysis of seven housekeeping genes. Recombinant regions were identified by at least three detection methods in 72% of our V. cholerae isolates. Despite frequent recombination, significant linkage disequilibrium was still detected among the V. cholerae sequence types. Incongruent but nonrandom associations were observed for maximum likelihood topologies from the individual loci. Overall, our estimated recombination rate in V. cholerae of 6.5 times the mutation rate is similar to those of other sexual bacteria and appears frequently enough to restrict selection from purging much of the neutral intraspecies diversity. These data suggest that frequent recombination among V. cholerae may hinder the identification of ecotypes in this bacterioplankton population.
PMCID: PMC3020551  PMID: 21075874
20.  Impact of parasites on salmon recruitment in the Northeast Atlantic Ocean 
Parasites may have large effects on host population dynamics, marine fisheries and conservation, but a clear elucidation of their impact is limited by a lack of ecosystem-scale experimental data. We conducted a meta-analysis of replicated manipulative field experiments concerning the influence of parasitism by crustaceans on the marine survival of Atlantic salmon (Salmo salar L.). The data include 24 trials in which tagged smolts (totalling 283 347 fish; 1996–2008) were released as paired control and parasiticide-treated groups into 10 areas of Ireland and Norway. All experimental fish were infection-free when released into freshwater, and a proportion of each group was recovered as adult recruits returning to coastal waters 1 or more years later. Treatment had a significant positive effect on survival to recruitment, with an overall effect size (odds ratio) of 1.29 that corresponds to an estimated loss of 39 per cent (95% CI: 18–55%) of adult salmon recruitment. The parasitic crustaceans were probably acquired during early marine migration in areas that host large aquaculture populations of domesticated salmon, which elevate local abundances of ectoparasitic copepods—particularly Lepeophtheirus salmonis. These results provide experimental evidence from a large marine ecosystem that parasites can have large impacts on fish recruitment, fisheries and conservation.
PMCID: PMC3574446  PMID: 23135680
recruitment; salmon; parasites; conservation; fisheries
21.  Shifts in the Microbial Community Composition of Gulf Coast Beaches Following Beach Oiling 
PLoS ONE  2013;8(9):e74265.
Microorganisms associated with coastal sands serve as a natural biofilter, providing essential nutrient recycling in nearshore environments and acting to maintain coastal ecosystem health. Anthropogenic stressors often impact these ecosystems, but little is known about whether these disturbances can be identified through microbial community change. The blowout of the Macondo Prospect reservoir on April 20, 2010, which released oil hydrocarbons into the Gulf of Mexico, presented an opportunity to examine whether microbial community composition might provide a sensitive measure of ecosystem disturbance. Samples were collected on four occasions, beginning in mid-June, during initial beach oiling, until mid-November from surface sand and surf zone waters at seven beaches stretching from Bay St. Louis, MS to St. George Island, FL USA. Oil hydrocarbon measurements and NOAA shoreline assessments indicated little to no impact on the two most eastern beaches (controls). Sequence comparisons of bacterial ribosomal RNA gene hypervariable regions isolated from beach sands located to the east and west of Mobile Bay in Alabama demonstrated that regional drivers account for markedly different bacterial communities. Individual beaches had unique community signatures that persisted over time and exhibited spatial relationships, where community similarity decreased as horizontal distance between samples increased from one to hundreds of meters. In contrast, sequence analyses detected larger temporal and less spatial variation among the water samples. Superimposed upon these beach community distance and time relationships, was increased variability in bacterial community composition from oil hydrocarbon contaminated sands. The increased variability was observed among the core, resident, and transient community members, indicating the occurrence of community-wide impacts rather than solely an overprinting of oil hydrocarbon-degrading bacteria onto otherwise relatively stable sand population structures. Among sequences classified to genus, Alcanivorax, Alteromonas, Marinobacter, Winogradskyella, and Zeaxanthinibacter exhibited the largest relative abundance increases in oiled sands.
PMCID: PMC3769389  PMID: 24040219
22.  13C-Carrier DNA Shortens the Incubation Time Needed To Detect Benzoate-Utilizing Denitrifying Bacteria by Stable-Isotope Probing 
The active bacterial community able to utilize benzoate under denitrifying conditions was elucidated in two coastal sediments using stable-isotope probing (SIP) and nosZ gene amplification. The SIP method employed samples from Norfolk Harbor, Virginia, and a Long-Term Ecosystem Observatory (no. 15) off the coast of Tuckerton, New Jersey. The SIP method was modified by use of archaeal carrier DNA in the density gradient separation. The carrier DNA significantly reduced the incubation time necessary to detect the 13C-labeled bacterial DNA from weeks to hours in the coastal enrichments. No denitrifier DNA was found to contaminate the archaeal 13C-carrier when [12C]benzoate was used as a substrate in the sediment enrichments. Shifts in the activity of the benzoate-utilizing denitrifying population could be detected throughout a 21-day incubation. These results suggest that temporal analysis using SIP can be used to illustrate the initial biodegrader(s) in a bacterial population and to document the cross-feeding microbial community.
PMCID: PMC1214685  PMID: 16151104
23.  Ambient-temperature primary nonselective enrichment for isolation of Salmonella spp. from an estuarine environment. 
A primary, nonselective, ambient-temperature enrichment procedure for isolation of Salmonella spp. is described. The procedure was superior to elevated-temperature selective enrichment for Salmonella when estuarine water samples were examined. Five Chesapeake Bay stations were monitored, over an 8-month period, for the presence of salmonellae. Of 72 water and sediment samples collected, 17 (23.6%) yielded Salmonella spp. Seven serotypes were identified among the isolates. A seasonal pattern was noted for the incidence of the salmonellae. A most probable number procedure, performed by membrane filtration and nonselective enrichment, yielded Salmonella most probable number indices as high as 110 per 100 g of sediment. The results suggest that new methods, such as the one described in this report, are required for isolation of human intestinal pathogens from estuaries and coastal waters.
PMCID: PMC170775  PMID: 326187
24.  Incidence of Vibrio species associated with blue crabs (Callinectes sapidus) collected from Galveston Bay, Texas. 
Bacteria were readily isolated from the hemolymph of a majority (88%) of the blue crabs collected from Galveston Bay, Texas. The hemolymph of most crabs contained moderate (greater than 10(3) bacteria/ml) to heavy (greater than (10(5) bacteria/ml) infections. Large variances were observed in the bacterial number associated with individual crabs, but no significant difference was observed between the mean bacterial levels in the hemolymph of crabs collected during different seasons of the sampling year. Vibrio spp. were the predominant bacterial types in the hemolymph of infected crabs and increased in number significantly during the summer season. Warmer water temperatures were thought to be responsible for this increase. Bacterial numbers and the percentage of Vibrio spp. were highest in the interior of the crab bodies, especially in the digestive tract. The exterior of the crabs did not appear to be the source of the hemolymph's bacterial flora. Bacteria taxonomically identical to Vibrio cholerae. V. vulnificus, and V. parahaemolyticus were routinely isolated from the crab hemolymph and external carapace. V. parahaemolyticus was the most prevalent of the pathogenic Vibrio spp. and was isolated from 23% of the hemolymph samples. V. vulnificus (7%) and V. cholerae (2%) occurred less commonly in the hemolymph. The incidences of V. parachaemolyticus and V. vulnificus were related and increased in the summer months. Both organisms were frequently isolated from the same crab.
PMCID: PMC244191  PMID: 7103475
25.  Investigation of Spatial and Temporal Trends in Water Quality in Daya Bay, South China Sea 
The objective is to identify the spatial and temporal variability of the hydrochemical quality of the water column in a subtropical coastal system, Daya Bay, China. Water samples were collected in four seasons at 12 monitoring sites. The Southeast Asian monsoons, northeasterly from October to the next April and southwesterly from May to September have also an important influence on water quality in Daya Bay. In the spatial pattern, two groups have been identified, with the help of multidimensional scaling analysis and cluster analysis. Cluster I consisted of the sites S3, S8, S10 and S11 in the west and north coastal parts of Daya Bay. Cluster I is mainly related to anthropogenic activities such as fish-farming. Cluster II consisted of the rest of the stations in the center, east and south parts of Daya Bay. Cluster II is mainly related to seawater exchange from South China Sea.
PMCID: PMC3138029  PMID: 21776234
Daya Bay; principal component analysis; multidimensional scaling analysis; statistical techniques; water quality

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