Examination of genes encoding for the virulence factors, hemolysin/cytolysin (vvhA) and capsular polysaccharide (CPS allele 1), during biofilm formation revealed that their expression was influenced by the maturity of the biofilm as well as by temperature. At 24 °C, expression of vvhA during biofilm formation was low between 4 and 12 h but increased 10-fold by 24 h to (5.1 × 104 ± 6.3 × 103mRNA copies/ml) as the biofilm matured. Compared to planktonic cells, expression of vvhA during biofilm formation at 24 °C was initially up-regulated at 4 h (1.07 ± 0.00-fold) but then was down-regulated almost four-fold during the intermediate and mature stages of biofilm formation. In contrast, vvhA expression at 37 °C was up-regulated almost four-fold in the early stages (4 and 6 h) of biofilm formation and remained two-fold up-regulated by 24 h even as the biofilm was deteriorating. CPS allele 1 expression at 24 °C during biofilm formation was up-regulated (1.50 ± 0.18-fold) during the initial attachment phase of the cells but was strongly down-regulated during the intermediate phases at 8 and 10 h (74.42 ± 42.16-fold and 453.76 ± 193.32-fold, respectively), indicating that capsular polysaccharide (CPS) is not important to intermediate biofilm architecture. Interestingly, as the biofilm matured by 24 h, expression of CPS allele 1 was again up-regulated (1.88 ± 1.07), showing that CPS plays a role in mature biofilm. At 37 °C, CPS allele 1 expression was significantly up-regulated (up to 105) during biofilm formation, indicating that the biofilm form of V. vulnificus may be preferred over the planktonic form in the human host.

The objective of this study was to find out if different enrichment media would influence the selection of clinical and environmental strains of Vibrio vulnificus. Therefore, traditional enrichment and maintenance medium for V. vulnificus were tested for selective preference for clinical or environmental strains. In the laboratory, Alkaline Peptone Water (APW) and Brain Heart Infusion Broth (BHIB), with adjusted salinity to that of APW, were inoculated with equal concentrations of clinical and environmental strains and incubated for 20 hr at 35 °C. BHIB selected for clinical strains (85%) as indicated by percentage of vcgC to vvhA genes, enumerated with quantitative PCR. In APW, the ratio of clinical (47%) to environmental strains (53%) was roughly equal. Enrichments of seawater with BHIB, APW, and Luria Bertani broth (LB) resulted in higher percentage of clinical strains in BHIB but not in APW or LB, in most samples. However seasonal differences in predominance of clinical or environmental strains in the seawater samples influenced the enrichment process. Our study suggests that BHIB10 might be advantageous for studying virulence factors and APW10 could be suitable for isolation and MPN enumeration in V. vulnificus. The selective preference of the BHIB10 medium for clinical strains indicates that iron plays an important role in the infection of the human host.

Due to the nature of Vibrio vulnificus infections (i.e., gastroenteritis and septicemia), only very few studies of a biofilm-associated form in this pathogen’s life cycle have been conducted. We proposed that biofilm production by clinical strains of V. vulnificus would be higher than by environmental strains. Biofilm formation by clinical and environmental reference strains was tested under different temperatures (24, 30, and 37 °C), pH (5.5, 7.5, and 8.5) and iron concentrations (18, 30, 50, 100, and 200 μM). Biofilm production by clinical strains was consistently higher (p < 0.001) at 24 °C than by environmental strains. Higher biofilm production was observed at pH 5.5 by all strains. Growth rates were lowest at pH 5.5 for environmental strains but for clinical strains there were no differences at pH 5.5, 7.5, and 8.5, demonstrating a tolerance to acidic and alkaline conditions. There was a strong, direct correlation between iron concentration in the growth medium and biofilm production by all strains tested. The current study indicates that biofilm formation might be important for the survival of V. vulnificus in vivo as well as in the marine environment. With regard to temperature and pH, higher biofilm production appears to be a trait of clinical strains and could be considered a virulence determinant in V. vulnificus.

Oysters from a reef in Galveston Bay, Texas, USA, were screened for more virulent clinical strains versus less virulent environmental strains of Vibrio vulnificus using a combination of quantitative PCR assays for the virulence correlating gene (clinical variant, vcgC) and 16S rRNA types A and B (type A = environmental, type B = clinical). The combination of vcgC and 16S rRNA type B loci to determine clinical type strains was suitable, as indicated by the strong correlation (R2 = 0.98; p < 0.001) between these gene counts over time and their relative proportion (up to 93.8% and 94.3%, respectively) to vvhA genes used to quantify all strains of V. vulnificus. A strong seasonal shift of V. vulnificus strain types was observed. Environmental strains (16S rRNA type A) predominated from April to mid-June as salinities increased from 22 to 27 PSU (practical salinity unit) and temperatures rose 20 to 28 °C, with peak gene quantities of 16 812 ± 56 CFU/g. As temperatures increased to ≥30 °C from mid-June to September and salinities rose above 27 PSU, clinical strains (16S rRNA type B; vcgC) predominated with peak quantities 31 868 ± 287 and 32 360 ± 178 CFU/g, respectively.

Sediments from a waste pit in Houston Ship Channel (HSC) were characterized using a number of molecular markers of natural organic matter fractions (e.g., pyrogenic carbon residues, PAHs, lignins), in addition to dioxins, in order to test the hypothesis that the dispersal and mobility of dioxins from the waste pit in the San Jacinto River is minimal. Station SG-6, sampled at the site of the submerged waste pit, had the highest dioxin/furan concentrations reported for the Houston Ship Channel/Galveston Bay (HSC/GB) system (10,000-46,000 pg/g), which translated into some of the highest reported World Health Organization Toxic Equivalents (TEQs: 2000-11,000 pg/g) in HSC sediments. Using a multi-tracer approach, this study confirmed our hypothesis that sludges from chlorinated pulps are a very likely source of dioxins/furans to this pit. However, this material also contained large quantities of additional hydrophobic organic contaminants (PAHs) and pyrogenic markers (soot-BC, levoglucosan), pointing to the co-occurrence of petroleum hydrocarbons and combustion byproducts. Comparison of dioxin/furan signatures in the waste pit with those from sediments of the HSC and a control site suggests that the remobilization of contaminated particles did not occur beyond the close vicinity of the pit itself. The dioxins/furans in sediments outside the waste pit within the HSC are rather from other diffuse inputs, entering the sedimentary environment through the air and water, and which are comprised of a mixture of industrial and municipal sources. Fingerprinting of waste pit dioxins indicates that their composition is typical of pulp and paper sources. Measured pore water concentrations were 1 order of magnitude lower than estimated values, calculated from a multiphase sorption model, indicating low mobility of dioxins within the waste pit. This is likely accomplished by co-occurring and strong sorbing pyrogenic and petrogenic residues in the waste pit, which tend to keep dioxins strongly sorbed to particles.

Sediments in the Houston Ship Channel and upper Galveston Bay, Texas, USA, are polluted with polychlorinated dibenzo-p-dioxins/furans (PCDD/F; ≤46,000 ng/kg dry weight (wt.)) with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), the most toxic congener, contributing >50 % of the total toxic equivalents (TEQ) at most locations. We measured PCDD/F concentrations in sediments and evaluated the potential for enhanced in situ biodegradation by surveying for Dehalococcoides mccartyi, an obligate organohalide respiring bacterium. Dehalococcoides spp. (98 % similar to D. mccartyi) and 22 other members of the class Dehalococcoidia were predominant 16S ribosomal RNA (rRNA) phylotypes. Dehalococcoides spp. were also present in the active fraction of the bacterial community. Presence/absence PCR screening detected D. mccartyi in sediment cores and sediment grab samples having at least 1 ng/kg dry wt. TEQ at salinities ranging from 0.6 to 19.5 PSU, indicating that they are widespread in the estuarine environment. Organic carbon-only and organic carbon + sulfate-amended sediment microcosm experiments resulted in ∼60 % reduction of ambient 2,3,7,8-TCDD in just 24 months leading to reductions in total TEQs by 38.4 and 45.0 %, respectively, indicating that 2,3,7,8-TCDD degradation is occurring at appreciable rates.

The bacterial diversity in ballast water from five general cargo ships calling at the Port of Houston was determined with ion semiconductor DNA sequencing (Ion Torrent PGM) of PCR amplified 16S rRNA genes. Phylogenetic analysis revealed that the composition of bacteria in ballast water did not resemble that of typical marine habitats or even open ocean waters where BWEs occur. The predominant group of bacteria in ships conducting BWEs was the Roseobacter clade within the Alphaproteobacteria. In contrast, Gammaproteobacteria were predominant in the ship that did not conduct a BWE. All the ships contained human, fish, and terrestrial plant pathogens as well as bacteria indicative of fecal or activated sludge contamination. Most of the 60 pathogens had not been detected in ballast water previously. Among these were the human pathogens Corynebacterium diptheriae and several Legionella species and the fish pathogens Francisella piscicida and Piscirickettsia salmonis.

Ballast water (BW) discharge by shipping vessels is a known transport vector of harmful species of dinoflagellates and diatoms. With a steady growth in global commerce, ship traffic to ports worldwide has intensified, increasing the risk of invasion by nonindigenous species. From 2008-12, >140 million metric tons of BW was discharged into Galveston Bay, Texas, much more than reported in other highly invaded Bays: San Francisco (96x10(6) mt) and Chesapeake (25x10(6) mt) during the same period. Studies conducted specifically on the dinoflagellate and diatom communities within Galveston Bay have been lacking until the present effort, which used both microscopic and genetic methods. Within one year of sampling, 35 genera of dinoflagellates and diatoms were identified from the two deepwater ports of Houston and Galveston. Seven of the genera are known toxin producers, three of which have formed harmful algal blooms within the Bay: Alexandrium, Gymnodinium, and Prorocentrum. Two genera identified from the ports (Takayama and Woloszynskia) have not been previously reported. This study provides a baseline of the phytoplankton community within these major ports in Galveston Bay before foreign shipping traffic increases due to the expansion of the Panama Canal.

Global movement of nonindigenous species, within ballast water tanks across natural barriers, threatens coastal and estuarine ecosystem biodiversity. In 2012, the Port of Houston ranked 10th largest in the world and 2nd in the US (waterborne tonnage). Ballast water was collected from 13 vessels to genetically examine the eukaryotic microorganism diversity being discharged into the Port of Houston, Texas (USA). Vessels took ballast water onboard in North Atlantic Ocean between the Port of Malabo, Africa and Port of New Orleans, Louisiana, (USA). Twenty genera of Protists, Fungi and Animalia were identified from at least 10 phyla. Dinoflagellates were the most diverse and dominant identified (Alexandrium, Exuviaella, Gyrodinium, Heterocapsa, Karlodinium, Pfiesteria and Scrippsiella). We are reporting the first detection of Picobiliphytes, Apusozoa (Amastigomonas) and Sarcinomyces within ballast water. This study supports that global commerce by shipping contributes to long-distance transportation of eukaryotic microorganisms, increasing propagule pressure and invasion supply on ecosystems.

Urbanization and stream-channel modifications affect organic matter concentrations and quality in streams, by altering allochthonous organic matter input and in-stream transformation. This study uses multiple tracers (δ(13)C, δ(15)N, C/N ratio, and chlorophyll-a) to track sources of organic matter in two highly urbanized bayous in Houston (Texas, USA). Wastewater treatment plants (WWTPs) are located in headwaters of both bayous and contribute more than 75% to water flow. Low isotopic relatedness to natural end-members and enriched δ(15)N values suggest the influence of WWTPs on the composition of all organic matter fractions. The two bayous differ in degree of channel improvement resulting in different responses to hydrological conditions. During high flow conditions, the influence of terrestrial organic matter and sediment resuspension was much more pronounced in the Buffalo Bayou than in the concrete-lined White Oak Bayou. Particulate organic matter (POM) in White Oak Bayou had similar values of enriched δ(15)N in all subsegments, whereas in Buffalo Bayou, the degree of δ(15)N enrichment was less in the subsegments of the lower watershed. The difference in riparian zone contributions and interactions with sediments/soils was likely responsible for the compositional differences between the two bayous. Phytoplankton inputs were significantly higher in the bayous, especially in slow-flowing sections, relative to the reference sites, and elevated phytoplankton inputs accounted for the observed stable C isotope differences between FPOM and high molecular weight dissolved organic matter (HMW DOM). Relative to POM, HMW DOM in the bayous was similar to WWTP effluents and showed minor longitudinal variability in both streams suggesting that WWTPs contribute much of the DOM in the systems. Urbanization has a major influence on organic matter sources and quality in these urban water bodies and these changes seem further enhanced by stream channel modifications.

The purpose of this research was to determine if Escherichia coli and enterococci in streambed and bank sediments from two urban bayous, Buffalo Bayou and White Oak Bayou, in Houston, TX, USA are a significant source of the chronically high levels of these bacteria in the overlying water. The watersheds of the bayous lie within highly urbanized areas of Greater Houston and there is primary recreational contact with the public. Extensive sampling of the watersheds was conducted from 2008 to 2010. Both fecal indicator bacteria were found at ≥104MPNgdry wt.−1 concentrations in the upper 1cm of sediment cores with declines by orders of magnitude at 15 and 30cm sediment horizons and in some cases 60cm, but, nonetheless, indicating that they can remain viable even at depth. No interannual variation was observed. And, there was no correlation with percent organic matter, however there was moderate correlation (R2=0.12; p=0.001) of E. coli with sediment moisture. In sediments, most E. coli and enterococci in Buffalo Bayou (76%) and White Oak Bayou (87.5%) were associated with fine sand grains (60 to 250μm). In the water column, E. coli was associated, in roughly equal percentages, with particle sizes <10, 10–25, 25–63, and ≥63μm (21.9, 25.6, 30.4, and 32.9%, respectively). Enterococci were mostly attached to particle sizes in the ranges of 10–25μm (36.0%) and 25–63μm (31.1%) as well as ≥63μm (37.7%) (p=0.0001). Fingerprinting of E. coli isolates from both bayous with Rep-PCR and the BOX A1R primer was used to demonstrate translocation of sediments from the upper to lower watersheds.

Dorsal and pectoral fin spines from two species of sea catfishes (Bagre marinus and Ariopsis felis) landed at 54 sites in Galveston Bay, Texas, and its sub-bays from June to October 2005 were screened with traditional cultivation-based assays and quantitative PCR assays for Vibrio vulnificus and Vibrio parahaemolyticus. V. vulnificus was present on 51.2% of fish (n = 247), with an average of 403 ± 337 SD cells g(-1). V. parahaemolyticus was present on 94.2% (n = 247); 12.8% tested positive for the virulence-conferring tdh gene, having an average 2,039 ± 2,171 SD cells g(-1). The increasing trend in seafood consumption of "trash fishes" from lower trophic levels, such as sea catfishes, warrants evaluation of their life histories for association with pathogens of concern for human consumption.

Ballast water exchange processes facilitate the dispersal and unnatural geographic expansion of phytoplankton, including harmful algal bloom species. From 2005 to 2010, over 45,000 vessels (≈ 8000 annually) travelled across Galveston Bay (Texas, USA) to the deep-water ports of Houston (10th largest in the world), Texas City and Galveston. These vessels (primarily tankers and bulkers) discharged ≈ 1.2 × 10(8) metrictons of ballast water; equivalent to ≈ 3.4% of the total volume of the Bay. Over half of the ballast water discharged had a coastwise origin, 96% being from US waters. Galveston Bay has fewer non-indigenous species but receives a higher volume of ballast water discharge, relative to the highly invaded Chesapeake and San Francisco Bays. Given the magnitude of shipping traffic, the role of Galveston Bay, both as a recipient and donor region of non-indigenous phytoplankton species is discussed here in terms of the invasibility risk to this system by way of ballast water.

129I is a major by-product of nuclear fission and had become one of the major radiation risk drivers at Department of Energy (DOE) sites. 129I is present at elevated levels in the surface soils of the Savannah River Site (SRS) F-Area and was found to be bound predominantly to soil organic matter (SOM). Naturally bound 127I and 129I to sequentially extracted humic acids (HAs), fulvic acids (FAs) and a water extractable colloid (WEC) were measured in a 129I-contaminated wetland surface soil located on the SRS. WEC is a predominantly colloidal organic fraction obtained from soil re-suspension experiments to mimic the fraction that may be released during groundwater exfiltration, storm water or surface runoff events. For the first time, NMR techniques were applied to infer the molecular environment of naturally occurring stable iodine and radioiodine binding to SOM. Iodine uptake partitioning coefficients (Kd) by these SOM samples at ambient iodine concentrations were also measured and related to quantitative structural analyses by 13C DPMAS NMR and solution state 1H NMR on the eight humic acid fractions. By assessing the molecular environment of iodine, it was found that it was closely associated with the aromatic regions containing esterified products of phenolic and formic acids or other aliphatic carboxylic acids, amide functionalities, quinone-like structures activated by electron-donating groups (e.g., NH2), or a hemicellulose–lignin-like complex with phenyl-glycosidic linkages. However, FAs and WEC contained much greater concentrations of 127I or 129I than HAs. The contrasting radioiodine contents among the three different types of SOM (HAs, FAs and WEC) suggest that the iodine binding environment cannot be explained solely by the difference in the amount of their reactive binding sites. Instead, indirect evidence indicates that the macro-molecular conformation, such as the hydrophobic aliphatic periphery hindering the active aromatic cores and the hydrophilic polysaccharides favoring the access by hydrophilic iodine species, also influences iodine–SOM interactions.

To develop an understanding of the role that microorganisms play in the transport of (129)I in soil-water systems, bacteria isolated from subsurface sediments were assessed for iodide oxidizing activity. Spent liquid medium from 27/84 bacterial cultures enhanced iodide oxidation 2-10 fold in the presence of H(2)O(2). Organic acids secreted by the bacteria were found to enhance iodide oxidation by (1) lowering the pH of the spent medium, and (2) reacting with H(2)O(2) to form peroxy carboxylic acids, which are extremely strong oxidizing agents. H(2)O(2)-dependent iodide oxidation increased exponentially from 8.4 to 825.9 μM with decreasing pH from 9 to 4. Organic acids with ≥2 carboxy groups enhanced H(2)O(2)-dependent iodide oxidation (1.5-15-fold) as a function of increasing pH above pH 6.0, but had no effect at pH ≤ 5.0. The results indicate that as pH decreases (≤5.0), increasing H(2)O(2) hydrolysis is the driving force behind iodide oxidation. However, at pH ≥ 6.0, spontaneous decomposition of peroxy carboxylic acids, generated from H(2)O(2) and organic acids, contributes significantly to iodide oxidation. The results reveal an indirect microbial mechanism, organic acid secretion coupled to H(2)O(2) production, that could enhance iodide oxidation and organo-iodine formation in soils and sediments.

Field and laboratory studies were carried out to understand the cause for steady increases in (129)I concentrations emanating from radiological basins located on the Savannah River Site, South Carolina. The basins were closed in 1988 by adding limestone and slag and then capping with a low permeability engineered cover. Groundwater (129)I concentrations in a well near the basins in 1993 were 200 pCi L(-1) and are presently between 400 and 1000 pCi L(-1). Iodine speciation in the plume contained wide ranges of iodide, iodate, and organo-iodine concentrations. First-order calculations based on a basin sediment desorption study indicate that the modest increase of 0.7 pH units detected in the study site groundwater over the last 17 years since closure of the basins may be sufficient to produce the observed increased groundwater (129)I concentrations near the basins. Groundwater monitoring of the plume at the basins has shown that the migration of many of the high risk radionuclides originally present at this complex site has been attenuated. However, (129)I continues to leave the source at a rate that may have been exacerbated by the initial remediation efforts. This study underscores the importance of identifying the appropriate in situ stabilization technologies for all source contaminants, especially if their geochemical behaviors differ.

ABSTRACT 129 I is of major concern because of its mobility in the environment, excessive inventory, toxicity (it accumulates in the thyroid), and long half-life (∼16 million years). The aim of this study was to determine if bacteria from a 129 I-contaminated oxic aquifer at the F area of the U.S. Department of Energy's Savannah River Site, SC, could accumulate iodide at environmentally relevant concentrations (0.1 μM I − ). Iodide accumulation capability was found in 3 out of 136 aerobic bacterial strains isolated from the F area that were closely related to Streptomyces / Kitasatospora spp., Bacillus mycoides , and Ralstonia/Cupriavidus spp. Two previously described iodide-accumulating marine strains, a Flexibacter aggregans strain and an Arenibacter troitsensis strain, accumulated 2 to 50% total iodide (0.1 μM), whereas the F-area strains accumulated just 0.2 to 2.0%. Iodide accumulation by FA-30 was stimulated by the addition of H 2 O 2 , was not inhibited by chloride ions (27 mM), did not exhibit substrate saturation kinetics with regard to I − concentration (up to 10 μM I − ), and increased at pH values of <6. Overall, the data indicate that I − accumulation likely results from electrophilic substitution of cellular organic molecules. This study demonstrates that readily culturable, aerobic bacteria of the F-area aquifer do not accumulate significant amounts of iodide; however, this mechanism may contribute to the long-term fate and transport of 129 I and to the biogeochemical cycling of iodine over geologic time.

In order to investigate the distributions and speciation of (129)I (and (127)I) in a contaminated F-Area groundwater plume of the Savannah River Site that cannot be explained by simple transport models, soil resuspension experiments simulating surface runoff or stormflow and erosion events were conducted. Results showed that 72-77% of the newly introduced I(-) or IO(3)(-) were irreversibly sequestered into the organic-rich riparian soil, while the rest was transformed by the soil into colloidal and truly dissolved organo-iodine, resulting in (129)I remobilization from the soil greatly exceeding the 1 pCi/L drinking water permit. This contradicts the conventional view that only considers I(-) or IO(3)(-) as the mobile forms. Laboratory iodination experiments indicate that iodine likely covalently binds to aromatic structures of the soil organic matter (SOM). Under very acidic conditions, abiotic iodination of SOM was predominant, whereas under less acidic conditions (pH ≥5), microbial enzymatically assisted iodination of SOM was predominant. The organic-rich soil in the vadose zone of F-Area thus acts primarily as a "sink," but may also behave as a potentially important vector for mobile radioiodine in an on-off carrying mechanism. Generally the riparian zone provides as a natural attenuation zone that greatly reduces radioiodine release.