Long Lag Times and High Velocities in the Motility of Natural Assemblages of Marine Bacteria

The motility characteristics of natural assemblages of coastal marine bacteria were examined. Initially, less than 10% of the bacteria were motile. A single addition of tryptic soy broth caused an increase in the motile fraction of cells but only after 7 to 12 h. Motility peaked at 15 to 30 h, when more than 80% of cells were motile. These results support the proposal that energy limits motility in the marine environment. Cell speeds changed more than an order of magnitude on timescales of milliseconds and hours. The maximum community speed was 144 (mu)m s(sup-1), and the maximum individual burst velocity was 407 (mu)m s(sup-1). In uniform medium, speed was an inverse function of tryptic soy broth concentration, declining linearly over 0.001 to 1.0%. In media where concentration gradients existed, the mean speed was a function of position in a spatial gradient, changing from 69 to 144 (mu)m s(sup-1) over as little as 15 to 30 (mu)m. The results suggest that marine bacteria are capable of previously undescribed quick shifts in speed that may permit the bacteria to rapidly detect and keep up with positional changes in small nutrient sources. These high speeds and quick shifts may reflect the requirements for useful motility in a turbulent ocean.     

Formation of 30- to 40-Micrometer-Thick Laminations by High-Speed Marine Bacteria in Microbial Mats

The precision with which motile heterotrophic bacteria could position themselves in microbial mats was determined. This required the development of a technique to view motile bacteria in situ. This was successfully achieved by replacing a 1-cm-diameter minicore from the mat sediment with 210- to 300-(mu)m-diameter glass beads or sieved agar. After allowing 3 days for regrowth of the mat into the transparent medium, a cross section showed that bacteria formed a layer as thin as 30 to 40 (mu)m at a depth of 500 (mu)m below the surface. Bacterial concentrations in this microlamination were 20 times above background. Mean speeds were 200 (mu)m s(sup-1) inside and 60 (mu)m s(sup-1) outside the microlamination. The percentages of bacteria turning per 30 s were 93% inside and 10% outside the microlamination. Artificial chemical gradients were unsuccessful in stimulating microlamination formation or in eliciting the same extent of speed and turning responses. The significance of the results is that it is now possible to microscopically examine sedimentary bacteria in situ. Our first examination indicates that some bacteria form chemotactic microlaminations by increasing their turning frequency. This behavior is opposite that described in the enteric-based model of chemotactic movement, in which positive chemotaxis is achieved by decreasing the turning frequency.     

The influence of cell size on marine bacterial motility and energetics

The influence of Brownian motion on marine bacteria was examined. Due to their small size, marine bacteria rotate up to 1,400 degrees in one second. This rapid rotation makes directional swimming difficult or impossible, as a bacterium may point in a particular direction for only a few tens of milliseconds on average. Some directional movement, however, was found to be possible if swimming speed is sufficiently great, over approximately 100 μm sec⁻¹. This led to the testable hypothesis that marine bacteria with radiii less than about 0.75 μm should exceed this speed. The result of the increased speed is that marine bacteria may spend in excess of 10% of their total energy budget on movement. This expenditure is 100 times greater than values for enteric bacteria, and indicates that marine bacteria are likely to be immotile below critical size-specific nutrient concentrations.     

Evidence for Intracellular and Extracellular Dimethylsulfoniopropionate (DMSP) Lyases and DMSP Uptake Sites in Two Species of Marine Bacteria

The kinetics of dimethylsulfoniopropionate (DMSP) uptake and dimethylsulfide (DMS) production from DMSP in two bacterial species, Alcaligenes sp. strain M3A, an isolate from estuarine surface sediments, and Pseudomonas doudoroffii, from seawater, were investigated. In Alcaligenes cells induced for DMSP lyase (DL) activity, DMS production occurred without DMSP uptake. In DL-induced suspensions of P. doudoroffii, uptake of DMSP preceded the production of DMS, indicating an intracellular location of DL; intracellular DMSP levels reached ca. 7 mM. DMSP uptake rates in noninduced cells showed saturation at three concentrations (K(inft) [transport] values, 3.4, 127, and 500 (mu)M). In DL-induced cells of P. doudoroffii, DMSP uptake rates increased ca. threefold (V(infmax), 0.022 versus 0.065 (mu)mol of DMSP taken up min(sup-1) mg of cell protein(sup-1)), suggesting that the uptake binding proteins were inducible. DMSP uptake and DL activity in P. doudoroffii were both inhibited by CN(sup-), 2,4-dinitrophenol, and membrane-impermeable thiol-binding reagents, further indicating active uptake of DMSP by cell surface components. The respiratory inhibitors had limited or no effect on DL activity by the Alcaligenes sp. Of the structural analogs of DMSP tested for their effect on DMSP metabolism, glycine betaine (GBT), but not methyl-3-mercaptopropionic acid (MMPA), inhibited DMSP uptake by P. doudoroffii, suggesting that GBT shares a binding protein with DMSP and that MMPA is taken up at a separate site. Two models of DMSP uptake, induction, and DL location found in marine bacteria are presented.     

A Nitrite Microsensor for Profiling Environmental Biofilms

A highly selective liquid membrane nitrite microsensor based on the hydrophobic ion-carrier aquocyanocobalt(III)-hepta(2-phenylethyl)-cobrynate is described. The sensor has a tip diameter of 10 to 15 (mu)m. The response is log-linear in freshwater down to 1 (mu)M NO(inf2)(sup-) and in seawater to 10 (mu)M NO(inf2)(sup-). A method is described for preparation of relatively large polyvinyl chloride (PVC)-gelled liquid membrane microsensors with a tip diameter of 5 to 15 (mu)m, having a hydrophilic coating on the tip. The coating and increased tip diameter resulted in more sturdy sensors, with a lower detection limit and a more stable signal than uncoated nitrite sensors with a tip diameter of 1 to 3 (mu)m. The coating protects the sensor membrane from detrimental direct contact with biomass and can be used for all PVC-gelled liquid membrane sensors meant for profiling microbial mats, biofilms, and sediments. Thanks to these improvements, liquid membrane sensors can now be used in complex environmental samples and in situ, e.g., in operating bioreactors. Examples of measurements in denitrifying, nitrifying, and nitrifying/denitrifying biofilms from wastewater treatment plants are shown. In all of these biofilms high nitrite concentrations were found in narrow zones of less than 1 mm.     

A New In Vivo Fluorimetric Technique To Measure Growth of Adhering Phototrophic Microorganisms

We developed a noninvasive rapid fluorimetric method for the investigation of growth of adhering (benthic) phototrophic microorganisms. The technique is based on the sensitive detection of the in vivo fluorescence of chlorophylls chlorophyll a and bacteriochlorophyll a and monitors increases in signal over time as an indicator for growth. The growth fluorimeter uses modulated excitation light of blue-light-emitting diodes and a photodiode as the detector. The light-emitting diodes are mounted geometrically in an aluminum housing for efficient and uniform illumination of the bottoms of the growth containers. The fluorimeter was characterized with respect to detection limit and dynamic range. This system is capable of resolving in vivo chlorophyll a concentrations of 0.5 (mu)g liter(sup-1) in cyanobacteria and 0.03 (mu)g liter(sup-1) in diatoms as well as in vivo bacteriochlorophyll a concentrations in phototrophic bacteria of 0.3 (mu)g liter(sup-1), which points to an extremely high sensitivity compared with that of similar available techniques. Thus, the new fluorimeter allows the determination of growth at extremely low cell densities. The instrument was used successfully to measure the growth of several adhering isolates of the filamentous cyanobacterium Microcoleus chthonoplastes from benthic microbial mats in seawater of different salinities. The data obtained demonstrate broad growth responses for all strains, which thus can be characterized as euryhaline organisms.     

Virus Decay and Its Causes in Coastal Waters

Recent evidence suggests that viruses play an influential role within the marine microbial food web. To understand this role, it is important to determine rates and mechanisms of virus removal and degradation. We used plaque assays to examine the decay of infectivity in lab-grown viruses seeded into natural seawater. The rates of loss of infectivity of native viruses from Santa Monica Bay and of nonnative viruses from the North Sea in the coastal seawater of Santa Monica Bay were determined. Viruses were seeded into fresh seawater that had been pretreated in various ways: filtration with a 0.2-(mu)m-pore-size filter to remove organisms, heat to denature enzymes, and dissolved organic matter enrichment to reconstitute enzyme activity. Seawater samples were then incubated in full sunlight, in the dark, or under glass to allow partitioning of causative agents of virus decay. Solar radiation always resulted in increased rates of loss of virus infectivity. Virus isolates which are native to Santa Monica Bay consistently degraded more slowly in full sunlight in untreated seawater (decay ranged from 4.1 to 7.2% h(sup-1)) than nonnative marine bacteriophages which were isolated from the North Sea (decay ranged from 6.6 to 11.1% h(sup-1)). All phages demonstrated susceptibility to degradation by heat-labile substances, as heat treatment reduced the decay rates to about 0.5 to 2.0% h(sup-1) in the dark. Filtration reduced decay rates by various amounts, averaging 20%. Heat-labile, high-molecular-weight dissolved material (>30 kDa, probably enzymes) appeared responsible for about 1/5 of the maximal decay. Solar radiation was responsible for about 1/3 to 2/3 of the maximal decay of nonnative viruses and about 1/4 to 1/3 of that of the native viruses, suggesting evolutionary adaptation to local light levels. Our results suggest that sunlight is an important contributing factor to virus decay but also point to the significance of particles and dissolved substances in seawater.     

A Novel Porous Nylon Biocarrier for Immobilized Bacteria

A highly porous nylon biocarrier was developed to support immobilized bacteria in bioreactors used to treat liquid wastes. Porosity analyses and scanning electron microscopy showed microbial colonization of accessible pores typically in the range of 100 to 1,200 (mu)m, with some as large as 3.9 mm. A bench-scale packed-bed reactor achieved a p-nitrophenol (PNP) removal rate of 5.95 kg of PNP m(sup-3) day(sup-1) for wastes containing 1,200 mg of PNP liter(sup-1). Complete mixing of the biocarrier bed to remove excess surface biomass was routinely achieved with simple air injection. These porous polymer biocarriers are promising as microbial supports in liquid-waste treatment and bioremediation applications.     

A Microsensor for Nitrate Based on Immobilized Denitrifying Bacteria

A biosensor for NO(inf3)(sup-) was constructed by attaching a 30- to 70-(mu)m-wide capillary with immobilized denitrifying bacteria in front of an N(inf2)O microsensor. These bacteria reduced O(inf2) so that only bacteria in the very tip of the sensor were exposed to O(inf2) whereas bacteria at a greater depth could carry out the anaerobic process of denitrification. In the presence of acetylene, which inhibits nitrous oxide reductase, bacteria reduced NO(inf3)(sup-) (or NO(inf2)(sup-)) from the surrounding medium to N(inf2)O and the concentration sensed by the N(inf2)O microsensor was directly proportional to the concentration of NO(inf3)(sup-) in the medium. By applying a 250-(mu)m-long capillary in front of the N(inf2)O microsensor, the 90% response time of the biosensor was 50 s. Biosensors may also be made with nitrous oxide-deficient strains so that acetylene inhibition can be omitted.     

Potential Significance of Lysogeny to Bacteriophage Production and Bacterial Mortality in Coastal Waters of the Gulf of Mexico

The potential effect that induction of lysogenic bacteria has on bacteriophage production and bacterial mortality in coastal waters was investigated, and we present estimates for the percentage of lysogenic cells in a natural aquatic bacterial community. Various concentrations of mitomycin C and exposure times to UV C radiation (UV-C) (wavelength of 254 nm) were used to induce the lytic cycle in lysogenic cells of natural communities of marine bacteria. UV-C treatment occasionally resulted in phage production, but phage production induced by UV-C was always less than that caused by the addition of mitomycin C. There was no evidence that high growth rates of bacteria resulted in lysogenic phage production. The burst size of cells induced by mitomycin C was determined by transmission electron microscopy and ranged from 11 to 45. Dividing the induced phage production by the burst size provided an estimate of the number of lysogenic bacterial cells, which ranged from 0.07 to 4.4% (average, 1.5%) of the total bacterial population. The percentages of lysogenic bacteria that were induced by mitomycin C were similar for samples collected nearshore from the pier of the Marine Science Institute (chlorophyll a, 1.6 to 2.9 (mu)g liter(sup-1)) and in relatively oligotrophic water (chlorophyll a, 0.2 to 0.9 (mu)g liter(sup-1)) collected 25 to 100 km offshore. By using a steady-state model, if all lysogenic bacteria were induced simultaneously, 0.14 to 8.8% (average, 3.0%) of the total bacterial mortality would result from induction of lysogenic cells. If mitomycin C induces all or the majority of lysogenized cells, our results imply that lysogenic phage production is generally not an important source of phage production or bacterial mortality in the coastal waters of the western Gulf of Mexico.     

Fluorescently Labeled Virus Probes Show that Natural Virus Populations Can Control the Structure of Marine Microbial Communities

Fluorescently stained viruses were used as probes to label, identify, and enumerate specific strains of bacteria and cyanobacteria in mixed microbial assemblages. Several marine virus isolates were fluorescently stained with YOYO-1 or POPO-1 (Molecular Probes, Inc.) and added to seawater samples that contained natural microbial communities. Cells to which the stained viruses adsorbed were easily distinguished from nonhost cells; typically, there was undetectable binding of stained viruses to natural microbial assemblages containing >10(sup6) bacteria ml(sup-1) but to which host cells were not added. Host cells that were added to natural seawater were quantified with 99% (plusmn) 2% (mean (plusmn) range) efficiency with fluorescently labeled virus probes (FLVPs). A marine bacterial isolate (strain PWH3a), tentatively identified as Vibrio natriegens, was introduced into natural microbial communities that were either supplemented with nutrients or untreated, and changes in the abundance of the isolate were monitored with FLVPs. Simultaneously, the concentrations of viruses that infected strain PWH3a were monitored by plaque assay. Following the addition of PWH3a, the concentration of viruses infecting this strain increased from undetectable levels (<1 ml(sup-1)) to 2.9 x 10(sup7) and 8.3 x 10(sup8) ml(sup-1) for the untreated and nutrient-enriched samples, respectively. The increase in viruses was associated with a collapse in populations of strain PWH3a from ca. 30 to 2% and 43 to 0.01% of the microbial communities in untreated and nutrient-enriched samples, respectively. These results clearly demonstrate that FLVPs can be used to identify and quantify specific groups of bacteria in mixed microbial communities. The data show as well that viruses which are present at low abundances in natural aquatic viral communities can control microbial community structure.     

Highly Active Microbial Communities in the Ice and Snow Cover of High Mountain Lakes

An exploratory study carried out in Pyrenean and Alpine lakes shows that a rich, active microbial community lives in the slush layers of the winter cover of such lakes in spite of the low temperature and the seasonal occurrence of the habitat. Bacteria were very diverse in morphology, with filaments reaching up to 100 (mu)m long; flagellates, both autotrophic (chrysophytes, cryptophytes, dinoflagellates, and volvocales) and heterotrophic, and ciliates were abundant, reaching biovolume values up to 2.7 x 10(sup6) (mu)m(sup3) ml(sup-1). Species composition was very variable, with dominance depending on date and depth. Although many species were typical of lake plankton communities, some were restricted to the slush, for instance the predatory ciliates Dileptus sp. and Lacrymaria sp., and others were restricted to the surface pools, such as the snow algae Chlamydomonas nivalis. Microbial biomasses and usually bacterial and algal activities were greater in the slush layers than in the lake water. Photosynthesis rate in the upper cover layers reached values up to 0.5 (mu)g of C liter(sup-1) h(sup-1), and high bacterial activities up to 226 pmol of leucine incorporated liter(sup-1) h(sup-1) and 25 pmol of thymidine incorporated liter(sup-1) h(sup-1) were measured. For most species, lake water flooding the ice and snow cover could provide an inoculum. Differential growth depending on the environmental conditions (nutrients, organic matter, light) of a particular slush layer could provide dominance of different groups or species. However, there was no obvious colonizing mechanism for those species not appearing either in plankton or in communities on top of the snowpack.     

Effects of Nitrate Availability and the Presence of Glyceria maxima on the Composition and Activity of the Dissimilatory Nitrate-Reducing Bacterial Community

The effects of nitrate availability and the presence of Glyceria maxima on the composition and activity of the dissimilatory nitrate-reducing bacterial community were studied in the laboratory. Four different concentrations of NO(inf3)(sup-), 0, 533, 1434, and 2,905 (mu)g of NO(inf3)(sup-)-N g of dry sediment(sup-1), were added to pots containing freshwater sediment, and the pots were then incubated for a period of 69 days. Upon harvest, NH(inf4)(sup+) was not detectable in sediment that received 0 or 533 (mu)g of NO(inf3)(sup-)-N g of dry sediment(sup-1). Nitrate concentrations in these pots ranged from 0 to 8 (mu)g of NO(inf3)(sup-)-N g of dry sediment(sup-1) at harvest. In pots that received 1,434 or 2,905 (mu)g of NO(inf3)(sup-)-N g of dry sediment(sup-1), final concentrations varied between 10 and 48 (mu)g of NH(inf4)(sup+)-N g of dry sediment(sup-1) and between 200 and 1,600 (mu)g of NO(inf3)(sup-)-N g of dry sediment(sup-1), respectively. Higher input levels of NO(inf3)(sup-) resulted in increased numbers of potential nitrate-reducing bacteria and higher potential nitrate-reducing activity in the rhizosphere. In sediment samples from the rhizosphere, the contribution of denitrification to the potential nitrate-reducing capacity varied from 8% under NO(inf3)(sup-)-limiting conditions to 58% when NO(inf3)(sup-) was in ample supply. In bulk sediment with excess NO(inf3)(sup-), this percentage was 44%. The nitrate-reducing community consisted almost entirely of NO(inf2)(sup-)-accumulating or NH(inf4)(sup+)-producing gram-positive species when NO(inf3)(sup-) was not added to the sediment. The addition of NO(inf3)(sup-) resulted in an increase of denitrifying Pseudomonas and Moraxella strains. The factor controlling the composition of the nitrate-reducing community when NO(inf3)(sup-) is limited is the presence of G. maxima. In sediment with excess NO(inf3)(sup-), nitrate availability determines the composition of the nitrate-reducing community.     

Investigation of 0.2 µm filterable bacteria from the Western Mediterranean Sea using a molecular approach: dominance of potential starvation forms

Although the existence of 0.2 μm filterable bacteria has been known since the early 80's, they are not taken into consideration when modeling microbial food webs, due to an overall lack of information concerning this specific size class. According to physiological studies on starvation forms and investigations on small bacterial cells in marine ecosystems, a 0.2 μm filtrate may consist of different phenotypes: starvation forms of typical marine bacteria, ultramicrobacteria or bacterial cells, even larger than 0.2 μm, but flexible enough to pass the nominal filter pore-size. In this pilot study we examined three filtered seawater fractions from the Western Mediterranean Sea (Bay of Calvi, Corsica/France) - the total bacterial population, the bacterial fraction above 0.2 μm and the 0.2 μm filtrate - to investigate the bacterial community structure of each of those fractions by the molecular approach of denaturing gradient gel electrophoresis (DGGE) of 16S rDNA fragments. The analysis of the resulting DGGE profiles revealed different patterns of dominant bands for the 0.2 μm filterable and the total bacterial populations within the samples. Additionally the 0.2 μm filterable bacterial compartment exhibited obvious differences in band patterns for winter and summer samples, which were not observed for the total bacterial fraction. According to the current knowledge concerning the status of 0.2 μm filterable bacteria, DGGE patterns indicate that most of the fragments representing 0.2 μm filterable bacteria were rather starvation forms of marine bacteria than ultramicrobacteria. The sequencing of excised and cloned DNA bands of the DGGE profiles characterized the phylogenetic affiliation of the corresponding 0.2 μm filterable bacteria, clustering mainly with known, typical marine isolates of both alpha-subclass and gamma-subclass of the Proteobacteria and the Cytophaga-Flavobacterium-Bacteroides branch.     

Activity and Distribution of Methane-Oxidizing Bacteria in Flooded Rice Soil Microcosms and in Rice Plants (Oryza sativa)

The activity and distribution of CH(inf4)-oxidizing bacteria (MOB) in flooded rice (Oryza sativa) soil microcosms was investigated. CH(inf4) oxidation was shown to occur in undisturbed microcosms by using (sup14)CH(inf4), and model calculations indicated that almost 90% of the oxidation measured had taken place at a depth where only roots could provide the O(inf2) necessary. Slurry from soil planted with rice had an apparent K(infm) for CH(inf4) of 4 (mu)M and a V(infmax) of 0.1 (mu)mol g (dry weight)(sup-1) h(sup-1). At a depth of 1 to 2 cm, there was no significant difference (P > 0.05) in numbers of MOB between soil from planted and nonplanted microcosms (mean, 7.7 x 10(sup5) g [fresh weight](sup-1)). Thus, the densely rooted soil at 1 to 2 cm deep did not represent rhizospheric soil with respect to the number of MOB. A significantly increased number of MOB was found only in soil immediately around the roots (1.2 x 10(sup6) g [fresh weight](sup-1)), corresponding to a layer of 0.1 to 0.2 mm. Plant-associated CH(inf4) oxidation was shown in a double chamber with carefully washed intact rice plants. Up to 90% of the CH(inf4) supplied to the root compartment was oxidized in the plants. CH(inf4) oxidation on isolated roots was higher and had a larger variability than that in soil slurries. Roots had an apparent K(infm) for CH(inf4) of 6 (mu)M and a V(infmax) of 5 (mu)mol g (dry weight)(sup-1) h(sup-1). The average number of MOB in homogenized roots was larger than on the rhizoplane and increased with plant age. MOB also were found in surface-sterilized roots and basal culms, indicating the ability of these bacteria to colonize the interior of roots and culms.     

Incorporation of glucose under anoxic conditions by bacterioplankton from coastal North Sea surface waters

It has been hypothesized that the potential for anaerobic metabolism might be a common feature of bacteria in coastal marine waters (L. Riemann and F. Azam, Appl. Environ. Microbiol. 68: 5554-5562, 2002). Therefore, we investigated whether different phylogenetic groups of heterotrophic picoplankton from the coastal North Sea were able to take up a simple carbon source under anoxic conditions. Oxic and anoxic incubations (4 h) or enrichments (24 h) of seawater with radiolabeled glucose were performed in July and August 2003. Bacteria with incorporated substrate were identified by using a novel protocol in which we combined fluorescence in situ hybridization and microautoradiography of cells on membrane filters. Incorporation of glucose under oxic and anoxic conditions was found in alpha-Proteobacteria, gamma-Proteobacteria, and the Cytophaga-Flavobacterium cluster of the Bacteroidetes at both times, but not in marine Euryarchaeota. In July, the majority of cells belonging to the alpha-proteobacterial Roseobacter clade showed tracer incorporation both in oxic incubations and in oxic and anoxic enrichments. In August, only a minority of the Roseobacter cells, but most bacteria affiliated with Vibrio spp., were able to incorporate the tracer under either condition. A preference for glucose uptake under anoxic conditions was observed for bacteria related to Alteromonas and the Pseudoalteromonas-Colwellia group. These genera are commonly considered to be strictly aerobic, but facultatively fermentative strains have been described. Our findings suggest that the ability to incorporate substrates anaerobically is widespread in pelagic marine bacteria belonging to different phylogenetic groups. Such bacteria may be abundant in fully aerated coastal marine surface waters.     

Responses to Stress and Nutrient Availability by the Marine Ultramicrobacterium Sphingomonas sp. Strain RB2256

Sphingomonas sp. strain RB2256 was isolated from Resurrection Bay in Alaska and possibly represents the dominant bacterial species in some oligotrophic marine environments. Strain RB2256 has a high-affinity nutrient uptake system when growing under nutrient-limiting conditions, and growing cells are very small (<0.08 (mu)m(sup3)). These characteristics indicate that RB2256 is highly evolved for withstanding nutrient limitations and grazing pressure by heterotrophic nanoflagellates. In this study, strain RB2256 was subjected to nutrient starvation and other stresses (high temperature, ethanol, and hydrogen peroxide). It was found that growing cells were remarkably resistant, being able to survive at a temperature of 56(deg)C, in 25 mM hydrogen peroxide, or in 20% ethanol. In addition, growing cells were generally as resistant as starved cells. The fact that vegetative cells of this strain are inherently resistant to such high levels of stress-inducing agents indicates that they possess stress resistance mechanisms which are different from those of other nondifferentiating bacteria. Only minor changes in cell volume (0.03 to 0.07 (mu)m(sup3)) and maximum specific growth rate (0.13 to 0.16 h(sup-1)) were obtained for cells growing in media with different organic carbon concentrations (0.8 to 800 mg of C per liter). Furthermore, when glucose-limited, chemostat-grown cultures or multiple-nutrient-starved batch cultures were suddenly subjected to excess glucose, maximum growth rates were reached immediately. This immediate response to nutrient upshift suggests that the protein-synthesizing machinery is constitutively regulated. In total, these results are strong evidence that strain RB2256 possesses novel physiological and molecular strategies that allow it to predominant in natural seawater.     

Phylogenetic identification and metabolism of marine dimethylsulfide-consuming bacteria

Microbial consumption is one of the main processes, along with photolysis and ventilation, that remove the biogenic trace gas dimethylsulfide (DMS) from the surface ocean. Although a few isolates of marine bacteria have been studied for their ability to utilize DMS, little is known about the characteristics or phylogenetic affiliation of DMS consumers in seawater. We enriched coastal and open-ocean waters with different carbon sources to stimulate different bacterial communities (glucose-consuming bacteria, methyl group-consuming bacteria and DMS consumers) in order to test how this affected DMS consumption and to examine which organisms might be involved. Dimethylsulfide consumption was greatly stimulated in the DMS addition treatments whereas there was no stimulation in the other treatments. Analysis of microbial DNA by two different techniques (sequenced bands from DGGE gels and clone libraries) showed that bacteria grown specifically with the presence of DMS were closely related to the genus Methylophaga. We also followed the fate of consumed DMS in some of the enrichments. Dimethylsulfide was converted mostly to DMSO in glucose or methanol enrichments, whereas it was converted mostly to sulfate in DMS enrichments, the latter suggesting use of DMS as a carbon and energy source. Our results indicate that unlike the biochemical precursor of DMS, dimethylsulfoniopropionate (DMSP), which is consumed by a broad spectrum of marine microorganisms, DMS seems to be utilized as a carbon and electron source by specialists. This is consistent with the usual observation that DMSP turns over at much higher rates than DMS.     

Dissolution of Barium from Barite in Sewage Sludges and Cultures of Desulfovibrio desulfuricans

High concentrations of total barium, ranging from 0.42 to 1.58 mg(middot)g(sup-1) (dry weight) were found in sludges of two sewage treatment plants near Florence, Italy. Barium concentrations in the suspended matter decreased as redox potential values changed from negative to positive. An anoxic sewage sludge sample was aerated, and 30% of the total barium was removed in 24 h. To demonstrate that barium was solubilized from barite by sulfate-reducing bacteria, a strain of Desulfovibrio desulfuricans was used to study the solubilization of barium from barite under laboratory conditions. During cell growth with different concentrations of barite from 0.01 to 0.3 g(middot)liter(sup-1) (the latter is the MIC) as the only source of sulfates in the cultures, the D. desulfuricans strain accumulated barium up to 0.58 (mu)g(middot)mg(sup-1) (dry weight). Three times the quantity of barium was dissolved by bacteria than in the uninoculated medium (control). The unexpectedly low concentration of soluble barium (1.2 mg of Ba(middot)liter(sup-1)) with respect to the quantity expected (109 mg of Ba(middot)liter(sup-1)), calculated on the basis of the free H(inf2)S evolved from the dissimilatory reduction of sulfate from barite, was probably due to the formation of other barium compounds, such as witherite (BaCO(inf3)) and the transient species barium sulfide (BaS). The D. desulfuricans strain, growing on barite, formed visible aggregates. Confocal microscopy analysis showed that aggregates consisted of bacteria and barite. After 3 days of incubation, several autofluorescent crystals surrounded by a dissolution halo were observed. The crystals were identified as BaS by comparison with the commercial compound.     

Role of Pore Size Location in Determining Bacterial Activity during Predation by Protozoa in Soil

The predation of a luminescence-marked strain of Pseudomonas fluorescens by the soil ciliate Colpoda steinii was studied in soil microcosms. Bacterial cells were introduced in either small (neck diameter, <6 (mu)m) or intermediate-sized (neck diameter, 6 to 30 (mu)m) pores in the soil by inoculation at appropriate matric potentials, and ciliates were introduced into large pores (neck diameter, 30 to 60 (mu)m). Viable cell concentrations of bacteria introduced into intermediate-sized pores decreased at a greater rate than those in small pores, with reductions in bacterial populations being accompanied by an increase in viable cell numbers of the ciliate. The data indicate that the location of bacteria in small pores provides significant protection from predation. In the absence of C. steinii, the level of metabolic activity of the bacterial population, measured by luminometry, decreased at a greater rate than cell number, and the level of luminescence cell(sup-1) consequently decreased. The decrease in levels of luminescence indicates a loss of activity due to starvation. During predation by C. steinii, the level of the activity of cells introduced into small pores fell in a similar manner. The level of cell activity was, however, significantly greater for cells introduced into intermediate-sized pores, despite their greater susceptibility to predation. The data suggest that increased activity arises from a release of nutrients by the predator and the greater accessibility of bacteria to nutrients in larger pores. Nutrient amendment of microcosms resulted in increases in bacterial populations to sustained, higher levels, while levels of luminescence increased transiently. The predation of cells introduced into intermediate-sized pores was greater, and there was also evidence that the level of activity of surviving bacteria was greater for bacteria in intermediate-sized but not small pores.