Attachment Stimulates Exopolysaccharide Synthesis by a Bacterium

This study examined the hypothesis that solid surfaces may stimulate attached bacteria to produce exopolymers. Addition of sand to shake-flask cultures seemed to induce exopolymer synthesis by a number of subsurface isolates, as revealed by optical microscopy. Several additional lines of evidence indicated that exopolymer production by attached cells (in continuous-flow sand-packed columns) was greater than by their free-living counterparts. Total carbohydrates and extracellular polysaccharides, both normalized to cell protein, were greater (2.5- and 5-fold, respectively) for attached cells than for free-living cells. Also, adsorption of a polyanion-binding dye to the exopolymer fraction was sixfold greater for attached cells than for unattached cells. When surface-grown cells were resuspended in fresh medium, exopolymer production decreased to the level characteristic of unattached cells, which ruled out the possibility that attached cells comprised a subpopulation of sticky mucoid variants. The mechanism by which attachment stimulated exopolymer synthesis did not involve changes of the specific growth rate, growth stage, or limiting nutrient.     

Distribution of the Luminous Bacterium Beneckea harveyi in a Semitropical Estuarine Environment

Bioluminescent bacteria were found in the water column, sediment, shrimp, and gastrointestinal tract of marine fishes from the semitropical estuarine environment of the East Lagoon, Galveston Island, Tex. Populations in the water column decreased during cold weather while sedimentary populations persisted. The highest percentages of luminous organisms were isolated from the gastrointestinal tract of marine fishes, where they persisted during 5 days of starvation. The presence of chitin temporarily increased intestinal populations. All isolates were Beneckea harveyi, whose natural habitat appears to be the gut of fishes and whose free-living reservoir appears to be marine sediments.     

Norovirus Distribution within an Estuarine Environment

Human norovirus (NoV) has been studied extensively as an important cause of gastroenteritis outbreaks worldwide. While oysters are a primary vehicle for infection, few studies have examined the wider distribution of NoV in the estuarine environment. Active shellfish-harvesting areas in Georgia were examined for the prevalence, genotype diversity, and concentrations of NoV in a variety of estuarine sample types over the course of 1 year. Of the 225 samples (9 oyster, 72 water, 72 63- to 200-μm plankton, and 72 >200-μm plankton) collected from 12 stations across two estuaries, 21 samples (9.3%) tested positive for NoV. By sample type, 55.0% (5/9) of oysters, 8.3% (6/72) of water samples, 11.1% (8/72) of 63- to 200-μm plankton samples, and 2.8% (2/72) of >200-μm plankton samples were positive for human NoV. The two NoV-positive >200-μm plankton samples, which contained mainly zooplankton, had the greatest quantity of NoV genomes (3.5 × 10¹³ and 1.7 × 10¹⁵ genomes g⁻¹) of any sample tested. The majority, 90.5% (19/21), of the samples tested positive for genogroup I NoV, and only 9.5% (2/21) of the samples tested positive for genogroup II. The high concentrations of NoV in plankton samples compared to water and oyster samples were unexpected and provide new insights into the presence and distribution of human NoV in the water environment.Human norovirus (NoV) is the leading cause of nonbacterial gastroenteritis worldwide (3). The Centers for Disease Control and Prevention (CDC) estimate that 23 million cases of acute gastroenteritis due to NoV occur each year, with symptoms including acute-onset vomiting, watery nonbloody diarrhea with abdominal cramps, and nausea (35). NoV outbreaks are pervasive for many reasons, but particularly because the virus is highly contagious and environmentally hardy (7). Additionally, infected individuals can excrete millions of viral particles in feces, leading to large numbers in sewage (16). Without proper removal or inactivation during wastewater treatment, the viruses can be released into recreational and shellfish-harvesting water bodies. Complete inactivation of NoV during sewage treatment is rare, and even in areas with proper wastewater treatment, contamination of oyster beds has been reported (5, 16, 17, 32, 38). Because bivalve molluscan shellfish are believed to act as filters for viruses and other microbes and because NoV is extremely infectious (as little as one viral particle is required for disease), the disease risk for consumption of raw oysters is high (27, 33, 40).Human NoV genogroup I (GI) and GII have been detected in oyster samples harvested from bays and estuaries worldwide (5, 10, 20). Ueki et al. (42) detected NoV in both shellfish and the surrounding river water in Japan and concluded that NoV contamination was most likely due to sewage and treated wastewater input into the river; however, no study has yet been able to characterize how NoV may be naturally distributed in an estuarine system, including in water, adhered to particles (including plankton), and in shellfish. The limitations are due in part to a lack of adequate detection methods specifically adapted to different environmental-sample types (8). Using a newly developed detection and quantification protocol (21), this study aimed to examine the distribution of NoV genogroups across a range of sample types within an estuarine system with the goal of better characterizing possible circulation of viruses between water, plankton, and oysters.     

Biodegradation of Dichloromethane in an Estuarine Environment

Dichloromethane (DCM) is a toxic pollutant showing prolonged persistence in water. So far, biodegradation of DCM has only been reported in soils and freshwater systems. Herein, we studied whether or not biodegradation of DCM could occur in estuarine waters. Results showed over 90% mineralization of DCM in natural estuarine waters supplemented with DCM. Biodegradation of DCM in estuarine waters occurred by association of different bacterial species. Generally, two bacterial species participated in DCM degradation. Two bacterial consortia were obtained. Consortia were able to degrade around 80% of DCM in about 6 days. The species involved in the process were identified by 16S rRNA gene sequencing; a consortium was constituted by Pseudomonas sp. and Brevundimonas sp. and a second consortium was formed by Pseudomonas sp. and an Acinetobacter sp. Our results showed that DCM can be readily biodegraded in estuarine waters.     

Trophic Level Stability-Inducing Effects of Predaceous Early Juvenile Fish in an Estuarine Mesocosm Study

Background

Classically, estuarine planktonic research has focussed largely on the physico-chemical drivers of community assemblages leaving a paucity of information on important biological interactions.

Methodology/Principal Findings

Within the context of trophic cascades, various treatments using in situ mesocosms were established in a closed estuary to highlight the importance of predation in stabilizing estuarine plankton abundances. Through either the removal (filtration) or addition of certain planktonic groups, five different trophic systems were established. These treatments contained varied numbers of trophic levels and thus different “predators” at the top of the food chain. The abundances of zooplankton (copepod and polychaete), ciliate, micro-flagellate, nano-flagellate and bacteria were investigated in each treatment, over time. The reference treatment containing apex zooplanktivores (early juvenile mullet) and plankton at natural densities mimicked a natural, stable state of an estuary. Proportional variability (PV) and coefficient of variation (CV) of temporal abundances were calculated for each taxon and showed that apex predators in this experimental ecosystem, when compared to the other systems, induced stability. The presence of these predators therefore had consequences for multiple trophic levels, consistent with trophic cascade theory.

Conclusions/Significance

PV and CV proved useful indices for comparing stability. Apex predators exerted a stabilizing pressure through feeding on copepods and polychaetes which cascaded through the ciliates, micro-flagellates, nano-flagellates and bacteria. When compared with treatments without apex predators, the role of predation in structuring planktonic communities in closed estuaries was highlighted.     

Location Is Everything: Evaluating the Effects of Terrestrial and Marine Resource Subsidies on an Estuarine Bivalve

Estuaries are amongst the world’s most productive ecosystems, lying at the intersection between terrestrial and marine environments. They receive substantial inputs from adjacent landscapes but the importance of resource subsidies is not well understood. Here, we test hypotheses for the effects of both terrestrial- and salmon-derived resource subsidies on the diet (inferred from stable isotopes of muscle tissue), size and percent nitrogen of the soft-shell clam (Mya arenaria), a sedentary estuarine consumer. We examine how these relationships shift across natural gradients among 14 estuaries that vary in upstream watershed size and salmon density on the central coast of British Columbia, Canada. We also test how assimilation and response to subsidies vary at smaller spatial scales within estuaries. The depletion and enrichment of stable isotope ratios in soft-shell clam muscle tissue correlated with increasing upstream watershed size and salmon density, respectively. The effects of terrestrial- and salmon-derived subsidies were also strongest at locations near stream outlets. When we controlled for age of individual clams, there were larger individuals with higher percent nitrogen content in estuaries below larger watersheds, though this effect was limited to the depositional zones below river mouths. Pink salmon exhibited a stronger effect on isotope ratios of clams than chum salmon, which could reflect increased habitat overlap as spawning pink salmon concentrate in lower stream reaches, closer to intertidal clam beds. However, there were smaller clams in estuaries that had higher upstream pink salmon densities, possibly due to differences in habitat requirements. Our study highlights the importance of upstream resource subsidies to this bivalve species, but that individual responses to subsidies can vary at smaller scales within estuaries.     

Some Effects of a Power Plant Effluent on Estuarine Epibenthic Organisms

A study using the submerged test panel method was conducted at the Oyster Creek Nuclear Generating Station near Barnegat Bay, New Jersey to investigate effects of the heated effluent on epibenthic communities. Test panels were placed in the intake and discharge canals and collected using SCUBA at 28, 84, 168 and 335-day intervals between October 1970 and October 1971. Exposed panels were replaced by clean panels upon collection. Faunal resemblance, both between stations during a sampling date and within stations from one sampling date to the next, was low. Numbers of species were higher at the discharge station than at the intake station from late fall to early summer and lower from midsummer to early fall. Overall differences in diversity at the two stations could not be detected. The optimum temperature for growth of Balanus sp. was approached more closely during winter in the discharge canal than in the intake canal where growth was retarded by cold water. Balanus eburneus appeared to have an extended breeding season. Because of dilution of the discharged water with cooler intake water through a plant bypass system during the summer, the effluent was not seriously harmful to amphipod reproduction. Season abundances of numerous invertebrates at both stations are reported and possible power plant effects are discussed.     

Persistence and Biodegradation of Spilled Residual Fuel on an Estuarine Beach

The enrichment of hydrocarbon-degrading bacteria and the persistence of petroleum hydrocarbons on an estuarine beach after a spill of residual fuel oil on 11 April 1973 in Upper Narragansett Bay, R.I. was investigated. A rapid enrichment occurred during days 4 to 16 after the oil spill and a significant population of hydrocarbon-degrading bacteria was maintained in the beach sand for at least a year. The concentration of petroleum hydrocarbons in the mid-tide area declined rapidly during the bacterial enrichment period, remained fairly constant throughout the summer, and then declined to a low concentration after 1 year. An increased concentration of branched and cyclic aliphatic hydrocarbons in the low-tide sediment 128 days after the spill suggested a migration of hydrocarbons during the summer. Hydrocarbon biodegradation was apparent during the winter months at a rate of less than 1 mug of hydrocarbon per g of dry sediment per day.     

核基质结合区的位置对转基因表达的影响

为研究核基质结合区 (MAR）序列不同插入位置对转基因表达作用的影响,PCR扩增人β 珠蛋白MAR分别插入到含氯霉素乙酰转移酶（chloramphenicol acetyltransferase,CAT）报告基因真核表达载体pCATG表达盒两侧、5′端及3′端.酶切鉴定后,用阳离子聚合物转染CHO细胞,G418筛选出阳性细胞克隆,ELISA分析CAT基因的表达水平,半定量PCR分析CAT基因相对拷贝数.结果表明,表达盒两侧含MAR序列的载体能提高介导的转基因表达水平平均提高10.4倍,5′端含MAR序列的载体表达水平平均提高3.9倍,3′端含MAR序列的载体反而降低转基因表达水平.5′端含MAR序列的表达载体其转基因相对拷贝数高于其它两组载体的基因拷贝数,转基因表达量与基因拷贝数不成正比.     

Osmotic Effects on Membrane Permeability in a Marine Bacterium

When cells of Alteromonas haloplanktis 214 (ATCC 19855) were preloaded with α-[¹⁴C]aminoisobutyric acid or the K⁺ in the cells was labeled with ⁴²K by incubation in a buffered salt solution containing 0.05 M MgSO₄, 0.01 M KCl, and 0.3 M NaCl, the cells retained their radioactivity when resuspended in the same salt solution. When NaCl was omitted from the solution, 80 to 90% of the radioactivity was lost from the cells. Cells suspended at intermediate concentrations of NaCl also lost radioactivity. New steady-state levels of the intracellular solutes were established within 15 s of suspending the cells; the percentage of radioactivity retained at each level decreased proportionately as the osmolality of the NaCl in the suspending solution decreased. With minor variations in effectiveness, MgCl₂, LiCl, and sucrose could substitute for NaCl on an equiosmolal basis for the retention of radioactivity by the cells. KCl, RbCl, and CsCl were appreciably less effective as replacements for NaCl, particularly when their osmolalities in the suspending solutions were low. The amount of α-[¹⁴C]aminoisobutyric acid taken up by the cells at the steady-state level increased to a maximum as the NaCl concentration in the suspending medium increased to 0.3 M. At suboptimal levels of NaCl, either LiCl or sucrose could substitute for NaCl in increasing the steady-state levels. The results obtained indicate that the porosity of the cytoplasmic membrane of this organism is determined by the difference between the osmotic pressure of the cytoplasm and the suspending medium. The lesser effectiveness of K⁺, Rb⁺, and Cs⁺ than Na⁺, Li, or Mg²⁺ in permitting the retention of solutes by the cells is attributed to the greater penetrability of the hydrated ions of the former group through the dilated pores of a stretched cytoplasmic membrane.     

Role of Chitin-Binding Proteins in the Specific Attachment of the Marine Bacterium Vibrio harveyi to Chitin

We examined the mechanism of attachment of the marine bacterium Vibrio harveyi to chitin. Wheat germ agglutinin and chitinase bind to chitin and competitively inhibited the attachment of V. harveyi to chitin, but not to cellulose. Bovine serum albumin and cellulase do not bind to chitin and had no effect on bacterial attachment to chitin. These data suggest that this bacterium recognizes specific attachment sites on the chitin particle. The level of attachment of a chitinase-overproducing mutant of V. harveyi to chitin was about twice as much as that of the uninduced wild type. Detergent-extracted cell membranes inhibited attachment and contained a 53-kDa peptide that was overproduced by the chitinase-overproducing mutant. Three peptides (40, 53, and 150 kDa) were recovered from chitin which had been exposed to membrane extracts. Polyclonal antibodies raised against extracellular chitinase cross-reacted with the 53- and 150-kDa chitin-binding peptides and inhibited attachment, probably by sterically hindering interactions between the chitin-binding peptides and chitin. The 53- and 150-kDa chitin-binding peptides did not have chitinase activity. These results suggest that chitin-binding peptides, especially the 53-kDa chitin-binding peptide and chitinase and perhaps the 150-kDa peptide, mediate the specific attachment of V. harveyi to chitin.     

Induction of Chitin-Binding Proteins during the Specific Attachment of the Marine Bacterium Vibrio harveyi to Chitin

Previous work has shown that attachment of Vibrio harveyi to chitin is specific and involves at least two chitin-binding peptides. However, the roles and regulation of these chitin-binding peptides in attachment are still unclear. Here we show that preincubation with the oligomeric sugars composing chitin stimulated chitinase activity, cellular attachment to chitin, and production of chitin-binding peptides. One of these peptides, a 53-kDa peptide, is produced constitutively and appears to mediate initial attachment to chitin. Synthesis of another peptide, a 150-kDa chitin-binding peptide, is induced by chitin and thus may be involved in time-dependent attachment. Coordinated regulation of attachment and degradation of chitin may give bacteria like V. harveyi a selective advantage over other bacteria in nutrient-poor aquatic environments.     

Effects of Grazing by Estuarine Gammaridean Amphipods on the Microbiota of Allochthonous Detritus

Estuarine gammaridean amphipods grazing at natural population density on detrital microbiota affected the microbial community composition, biomass, and metabolic activity without affecting the physical structure of the leaves. Total microbial biomass estimated by adenosine triphosphate and lipid phosphate or observed by scanning electron microscopy was greater on grazed than on ungrazed detritus. The rates of oxygen consumption, poly-β-hydroxybutyrate synthesis, total lipid biosynthesis, and release of ¹⁴CO₂ from radioactively prelabeled microbiota were higher on grazed than on ungrazed leaves, indicating stimulation of the metabolic activity of grazed detrital microbes. This was true with rates based either on the dry leaf weight or microbial biomass. Alkaline phosphatase activity was lower in the grazed system, consistent with enhanced inorganic phosphate cycling. The loss of ¹⁴C from both total lipid and poly-β-hydroxybutyrate of microorganisms prelabeled with ¹⁴C was greater from grazed than ungrazed microbes. There was a faster decrease in the ¹⁴C-glycolipid than in the ¹⁴C-neutral lipid or ¹⁴C-phospholipid fractions. Analysis of specific phospholipids showed losses of the metabolically stable [¹⁴C]glycerolphosphorylcholine derived from phosphatidylcholine and much more rapid metabolism of the bacterial lipid phosphatidylglycerol measured as [¹⁴C]glycerolphosphorylglycerol with amphipod grazing. The biochemical data supported scanning electron microscopy observations of a shift as the grazing proceeded from a bacterial/fungal community to one dominated by bacteria.     

Influence of an Oyster Reef on Development of the Microbial Heterotrophic Community of an Estuarine Biofilm

We characterized microbial biofilm communities developed over two very closely located but distinct benthic habitats in the Pensacola Bay estuary using two complementary cultivation-independent molecular techniques. Biofilms were grown for 7 days on glass slides held in racks 10 to 15 cm over an oyster reef and an adjacent muddy sand bottom. Total biomass and optical densities of dried biofilms showed dramatic differences for oyster reef versus non-oyster reef biofilms. This study assessed whether the observed spatial variation was reflected in the heterotrophic prokaryotic species composition. Genomic biofilm DNA from both locations was isolated and served as a template to amplify 16S rRNA genes with universal eubacterial primers. Fluorescently labeled PCR products were analyzed by terminal restriction fragment length polymorphism, creating a genetic fingerprint of the composition of the microbial communities. Unlabeled PCR products were cloned in order to construct a clone library of 16S rRNA genes. Amplified ribosomal DNA restriction analysis was used to screen and define ribotypes. Partial sequences from unique ribotypes were compared with existing database entries to identify species and to construct phylogenetic trees representative of community structures. A pronounced difference in species richness and evenness was observed at the two sites. The biofilm community structure from the oyster reef setting had greater evenness and species richness than the one from the muddy sand bottom. The vast majority of the bacteria in the oyster reef biofilm were related to members of the γ- and δ-subdivisions of Proteobacteria, the Cytophaga-Flavobacterium -Bacteroides cluster, and the phyla Planctomyces and Holophaga-Acidobacterium. The same groups were also present in the biofilm harvested at the muddy sand bottom, with the difference that nearly half of the community consisted of representatives of the Planctomyces phylum. Total species richness was estimated to be 417 for the oyster reef and 60 for the muddy sand bottom, with 10.5% of the total unique species identified being shared between habitats. The results suggest dramatic differences in habitat-specific microbial diversity that have implications for overall microbial diversity within estuaries.     

亚硝化细菌的分离和特性的研究

从厌氧/好氧(A/O)工艺的废水处理系统活性污泥中分离出亚硝化细菌A5,鉴定为亚硝化单胞菌(Nitrosomonas sp．)．并对其特性进行了研究。得出氨氧化作用最适的温度为32℃,最适的pH值为8．0,需要充足的溶解氧。另外．亚硝化细菌A5具有较高的耐盐能力,NaCl或Na2SO4浓度为50g/L范围内,其氨氧化作用没有受到明显的抑制,适合于投加到处理含有较高的溶解性盐和较高离子强度的废水生化处理系统中去除氨氮。     

一株嗜酸硫氧化菌的分离

从江西德兴分离得到一株嗜酸硫氧化杆菌DX-2,采用双层固体培养基进行分离纯化,对分离菌株进行了形态、生理生化特性研究及16SrRNA序列分析.该菌株为革兰氏阴性细菌,短杆状,菌体大小（0.4～0.5）μm×（1～2）μm,化能自养,可利用硫磺和硫代硫酸盐为能源生长,不能利用亚铁进行生长.以16SrRNA序列同源性为基础构建了相关种属在内的系统发育树,结果表明,DX-2与喜温硫杆菌（Acidithiobacillus caldus）处于同一进化树分支中,相似性达99%以上.考察了不同重金属离子对DX-2的生长的影响.     

Native and Introduced Ecosystem Engineers Produce Contrasting Effects on Estuarine Infaunal Communities

Cordgrasses in the genus Spartina are good examples of ecosystem engineers that modify habitat structure in estuaries throughout the world. In San Francisco Bay, California, USA, marshes containing native California cordgrass (Spartina foliosa) are being invaded by a hybrid (S. alterniflora × S. foliosa) formed after introduction of S. alterniflora. This study compared vegetation, sediment structure, and infaunal invertebrates in native and invaded marshes. We hypothesized that differences in the physical structure between S. foliosa and hybrid Spartina would be reflected in differences in density, biomass, diversity, and taxonomic composition of infauna. Hybrid Spartina modifies habitat structure more than S. foliosa by producing taller stems, and greater plant biomass both above- and belowground while occupying a much wider tidal range, thereby transforming open mudflats to a vegetated habitat. In general, S. foliosa areas contained significantly higher densities of benthic infauna than adjacent mudflats, while hybrid Spartina areas never contained greater infaunal densities than mudflats. This is because S. foliosa produces a moderate level of structure that can facilitate benthic invertebrates, whereas hybrid Spartina produces so much structure, particularly belowground, that it actually excludes invertebrates. Therefore, we suggest that these two closely related species both act as ecosystem engineers, but with opposing effects on invertebrate communities.     

Effects of Inorganic Particles on Metabolism by a Periphytic Marine Bacterium

Measurements were made of adsorption of a periphytic marine bacterium, glucose, and glutamic acid to inorganic particles in seawater and defined bacterial growth medium. Measurements of the metabolism of bacteria were made in the presence and absence of particles by microcalorimetry and radiorespirometry. It was found that hydroxyapatite adsorbs glutamic acid, but not glucose, from the experimental medium. It was also found that hydroxyapatite adsorbs essentially all of the bacteria from the medium when the bacterial concentration is approximately 6 × 10⁵ bacteria per ml. If the bacterial concentration is approximately 6 × 10⁷, then only a small fraction of cells become attached. It was therefore possible to select bacterial concentrations and organic nutrients so that bacterial attachment, organic nutrient adsorption, or both would occur in different experiments. In this experimental system the metabolism by attached and nonattached bacteria of adsorbing and nonadsorbing organic nutrients was measured. The results show that bacterial activity in this model system was not enhanced by the particles, regardless of whether the bacteria, the organic nutrient, or both were associated with the surface. In fact, the respiratory activity of the attached bacteria was diminished in comparison with that of free bacteria.