Simultaneous transport of two bacterial strains in intact cores from Oyster, Virginia: biological effects and numerical modeling

The transport characteristics of two adhesion-deficient, indigenous groundwater strains, Comamonas sp. strain DA001 and Erwinia herbicola OYS2-A, were studied by using intact sediment cores (7 by 50 cm) from Oyster, Va. Both strains are gram-negative rods (1.10 by 0.56 and 1.56 by 0.46 microm, respectively) with strongly hydrophilic membranes and a slightly negative surface charge. The two strains exhibited markedly different behaviors when they were transported through granular porous sediment. To eliminate any effects of physical and chemical heterogeneity on bacterial transport and thus isolate the biological effect, the two strains were simultaneously injected into the same core. DA001 cells were metabolically labeled with (35)S and tagged with a vital fluorescent stain, while OYS2-A cells were metabolically labeled with (14)C. The fast decay of (35)S allowed deconvolution of the two isotopes (and therefore the two strains). Dramatic differences in the transport behaviors were observed. The breakthrough of DA001 and the breakthrough of OYS2-A both occurred before the breakthrough of a conservative tracer (termed differential advection), with effluent recoveries of 55 and 30%, respectively. The retained bacterial concentration of OYS2-A in the sediment was twofold higher than that of DA001. Among the cell properties analyzed, the statistically significant differences between the two strains were cell length and diameter. The shorter, larger-diameter DA001 cells displayed a higher effluent recovery than the longer, smaller-diameter OYS2-A cells. CXTFIT modeling results indicated that compared to the DA001 cells, the OYS2-A cells experienced lower pore velocity, higher porosity, a higher attachment rate, and a lower detachment rate. All these factors may contribute to the observed differences in transport.     

罗氏沼虾(Macrobrachium rosenbergii)养殖池塘沉积物中细菌、硫酸盐还原菌和硫氧化菌的垂直分布特征

沉积物中微生物介导的硫循环在有机物分解和养分循环中发挥重要作用,但目前我们对水产养殖生态系统中的参与硫酸盐还原和硫氧化过程的微生物多样性及其调控机制仍知之甚少。【目的】探究硫酸盐还原菌(sulfate-reducing bacteria,SRB)和硫氧化菌(sulfur-oxidizing bacteria,SOB)的垂直分布特征及其主要的环境驱动因素。【方法】本研究利用高通量测序和荧光定量PCR (qPCR)分析了罗氏沼虾(Macrobrachium rosenbergii)养殖池塘沉积物表层(0–1 cm)、中层(10–11 cm)和底层(20–21 cm)中的细菌、SRB和SOB的丰度、多样性和群落组成。【结果】细菌(16S rRNA)、硫酸盐还原菌(dsrB)和硫氧化菌(soxB)的基因拷贝数呈现着从表层到中层急剧骤降的趋势(ANOVA,P<0.05),但中层和底层样品之间的差异却并不显著(P>0.05),以及α多样性分析显示3个群体的物种丰富度和均匀度都随深度而逐步降低,这都说明硫循环过程主要发生于沉积物的表层。γ-、δ-和β-变形菌分别是细菌、SRB和SOB的优势类群;其中SRB以Desulfobacca属和脱硫八叠球菌属(Desulfosarcina)为主,前者在表层有着最低的比重,而后者却与之相反;硫杆菌(Thiobacillus)作为细菌和SOB的优势属,更广泛地分布于中层沉积物中。RDA分析和Mantel检验揭示了影响细菌群落的主要环境因子是NO₃^–、SO₄^2–、TOC和TON,而SRB的群落变异主要是由As、TON、NO₃^–和Pb所驱动,以及SOB的群落变化则主要响应了TC、NO₂^–、NH₄⁺和TON浓度。【结论】养殖池塘底栖的细菌、SRB和SOB的丰度、多样性和群落结构的垂直分布特征可能受到多种环境因素共同的影响。     

Application of a vital fluorescent staining method for simultaneous, near-real-time concentration monitoring of two bacterial strains in an Atlantic coastal plain aquifer in Oyster, Virginia

Two differentially labeled bacterial strains were monitored in near-real time during two field-scale bacterial transport experiments in a shallow aquifer in July 2000 and July 2001. Comamonas sp. strain DA001 and Acidovorax sp. strain OY-107 were grown and labeled with the vital fluorescent stain TAMRA/SE (5 [and -6]-carboxytetramethylrhodamine, succinimidyl ester) or CFDA/SE (5 [and -6]-carboxyfluorescein diacetate, succinimidyl ester). Fluorescently labeled cells and a conservative bromide tracer were introduced into a suboxic superficial aquifer, followed by groundwater collection from down-gradient multilevel samplers. Cells were enumerated in the field by microplate spectrofluorometry, with confirmatory analyses for selected samples done in the laboratory by epifluorescence microscopy, flow cytometry, and ferrographic capture. There was general agreement in the results from all of the vital-stain-based enumeration methods, with differences ranging from <10% up to 40% for the analysis of identical samples between different tracking methods. Field analysis by microplate spectrofluorometry was robust and efficient, allowing thousands of samples to be analyzed in quadruplicate for both of the injected strains. The near-real-time data acquisition allowed adjustments to the predetermined sampling schedule to be made. The microplate spectrofluorometry data sets for the July 2000 and July 2001 experiments allowed the transport of the injected cells to be related to the site hydrogeology and injection conditions and enabled the assessment of differences in the transport of the two strains. This near-real-time method should prove effective for a number of microbial ecology applications.     

The Benthic Macrofauna Associated with the Oyster Reefs of the James River Estuary,Virginia, U.S.A.

Productive, natural oyster reefs within the James River estuary, were quantitatively sampled for associated macrofauna for a year. The 192 samples yielded 142 taxa. Mean faunal density on these reefs ranged from 5,757 to 57,857 m⁻². Over 770/0 of the individuals collected belonged to 11 taxa. The number of species and informational diversity (H') decreased up the estuarine gradient. The quality of available shell surface was deemed important in determining density. The spatial heterogeneity provided by the shell surface allowed development of a densely populated, diverse faunal assemblage which differs from soft-bottom assemblages principally in quantitative aspects. The assemblage is not spatially or temporally homogeneous and undergoes significant changes in community structure both seasonally and along the estuarine gradient.     

Simultaneous Transport of Two Bacterial Strains in Intact Cores from Oyster, Virginia: Biological Effects and Numerical Modeling

The transport characteristics of two adhesion-deficient, indigenous groundwater strains, Comamonas sp. strain DA001 and Erwinia herbicola OYS2-A, were studied by using intact sediment cores (7 by 50 cm) from Oyster, Va. Both strains are gram-negative rods (1.10 by 0.56 and 1.56 by 0.46 μm, respectively) with strongly hydrophilic membranes and a slightly negative surface charge. The two strains exhibited markedly different behaviors when they were transported through granular porous sediment. To eliminate any effects of physical and chemical heterogeneity on bacterial transport and thus isolate the biological effect, the two strains were simultaneously injected into the same core. DA001 cells were metabolically labeled with ³⁵S and tagged with a vital fluorescent stain, while OYS2-A cells were metabolically labeled with ¹⁴C. The fast decay of ³⁵S allowed deconvolution of the two isotopes (and therefore the two strains). Dramatic differences in the transport behaviors were observed. The breakthrough of DA001 and the breakthrough of OYS2-A both occurred before the breakthrough of a conservative tracer (termed differential advection), with effluent recoveries of 55 and 30%, respectively. The retained bacterial concentration of OYS2-A in the sediment was twofold higher than that of DA001. Among the cell properties analyzed, the statistically significant differences between the two strains were cell length and diameter. The shorter, larger-diameter DA001 cells displayed a higher effluent recovery than the longer, smaller-diameter OYS2-A cells. CXTFIT modeling results indicated that compared to the DA001 cells, the OYS2-A cells experienced lower pore velocity, higher porosity, a higher attachment rate, and a lower detachment rate. All these factors may contribute to the observed differences in transport.     

Distribution of nutrients,trace elements,PAHs and radionuclides in sediment cores from Lake Varese (N. Italy)

Sediment cores were collected at 5 stations in Lake Varese. They were analyzed for organic matter, N, P, organic C, Cd, Cr, Cu, Hg, Pb, Zn and PAHs. A sedimentological approach has been applied to estimate the ecological risk from identified pollutants.As in other eutrophic lakes in Northern Italy, this lake is also at considerable risk from heavy metal pollution. Cr, Cu and Cd showed the highest enrichment factors for the last 5 years, 23.4, 8.0 and 7.6 respectively. Other metals had enrichment factors ranging from 1 to 3. Fluoranthene was chosen as a representative PAH, derived from combustion products; its average value in surficial sediments ranged from 100 to 220 ng l^-1 dw. This compound can be a hazard to human health and aquatic life. An evaluation of radionuclide distributions after the recent Chernobyl accident in the USSR (¹³⁴Cs, ¹³⁷C s, ¹³¹I, ¹⁰⁶Ru) provided useful tracers to follow the cycling of pollutants bound to particulate matter in the aquatic ecosystem.     

Ultrastructural, physiological, and cytochemical characterization of cores in group D streptococci.

Cores are large, rod-shaped structures that have been found almost exclusively in group D streptococci, measure 0.1 to 0.16 mum in diameter, and extend the width or length of cells. This study has shown that cores are produced in the cells at a reproducible point in early stationary growth after extensive mesosomal formation and after the pH has dropped below 6.5. When cells containing cores were introduced into a fresh medium with a pH above 6.5, the structures disappeared within 5 min. The structures were not found in young, logarithmically growing cells but formed in these cells upon autolysis or treatment with penicillin. Cores that were forming or disintegrating appeared to have a lamellar substructure. When chloramphenicol was added to the medium before the culture reached stationary phase, no cores were found in the cells. Cytochemical studies indicated that cores contain protein and are not composed of cell wall material or other polysaccharides that contain 1,2-glycol groups.     

Electron microscopic detection and in situ characterization of bacterial nanoforms in extreme biotopes

The morphology, ultrastructure, and quantity of bacterial nanoforms were studied in extreme biotopes: East Siberia permafrost soil (1-3 Ma old), petroleum-containing slimes (35 years old), and biofilms from subsurface oil pipelines. The morphology and ultrastructure of microbial cells in natural biotopes in situ were investigated by high-resolution transmission electron microscopy and various methods of sample preparation: ultrathin sectioning, cell replicas, and cryofractography. It was shown that the biotopes under study contained high numbers of bacterial nanoforms (29-43% of the total number of microorganisms) that could be assigned to ultramicrobacteria due to their size (diameter of < or =0.3 microm and volume of < or =0.014 microm3) and structural characteristics (the presence of the outer and cytoplasmic membranes, nucleoid, and cell wall, as well as their division patterns). Seven different morphostructural types of nanoforms of vegetative cells, as well as nanospores and cyst-like cells were described, potentially representing new species of ultramicrobacteria. In petroleum-containing slimes, a peculiar type of nanocells was discovered, gram-negative cells mostly 0.18-0.20 x 0.20-0.30 microm in size, forming spherical aggregates (microcolonies) of dividing cells in situ. The data obtained promoted the isolation of pure cultures of ultramicrobacteria from petroleum-containing slimes; they resembled the ultramicrobacterium observed in situ in their morphology and ultrastructure.     

Electron microscopic detection and in situ characterization of bacterial nanoforms in extreme biotopes

The morphology, ultrastructure, and quantity of bacterial nanoforms were studied in extreme biotopes: East Siberia permafrost soil (1–3 Ma old), petroleum-containing slimes (35 years old), and biofilms from subsurface oil pipelines. The morphology and ultrastructure of microbial cells in natural biotopes in situ were investigated by high-resolution transmission electron microscopy and various methods of sample preparation: ultrathin sectioning, cell replicas, and cryofractography. It was shown that the biotopes under study contained high numbers of bacterial nanoforms (29–43% of the total number of microorganisms) that could be assigned to ultramicrobacteria due to their size (diameter of ≤ 0.3 μm and volume of ≤ 0.014 μm³) and structural characteristics (the presence of the outer and cytoplasmic membranes, nucleoid, and cell wall, as well as their division patterns). Seven different morphostructural types of nanoforms of vegetative cells, as well as nanospores and cyst-like cells were described, potentially representing new species of ultramicrobacteria. In petroleum-containing slimes, a peculiar type of nanocells was discovered, gram-negative cells mostly 0.18–0.20 × 0.20–0.30 μm in size, forming in situ spherical aggregates (microcolonies) of dividing cells. The data obtained promoted the isolation of pure cultures of ultramicrobacteria from petroleum-containing slimes; they resembled the ultramicrobacterium observed in situ in their morphology and ultrastructure.     

The recent history of the chironomid fauna of Lough Neagh, from the analysis of remains in sediment cores

SUMMARY. Analysis of sub-fossil chironomid remains in the top of Lough Neagh sediments shows a change from a mainly Tanytarsus fauna to a mainly Chironomus one. This has been accompanied by an exponential increase in numbers of head capsules deposited per m² per year. Tentative dating places this change within the last 130 years, with the most rapid increase in numbers post-1945     

Construction and characterization of bacterial artificial chromosome libraries from the silkworm, Bombyx mori.

Two bacterial artificial chromosome (BAC) libraries were constructed using nuclear DNA from posterior silkglands of the silkworm (Bombyx mori) strains p50 and C108. The libraries contain a total of 36,864 clones, or approximately 9 genome equivalents. The average insert sizes in the libraries were 134.5 kb and 120.8 kb, respectively. PCR-based screening was performed on the p50 library using probes for 34 sequence-tagged sites (STSs). Between 3 and 11 (6.1 hits on average) clones were isolated with each STS, in good agreement with the library size, 5.8 genome equivalents. The previously reported close linkage between the Bombyx homologs of the invected (Bm in) and engrailed (Bm en) genes was confirmed by construction of a BAC contig that contained both. Moreover, screening revealed novel information about the chromosomal organization of the sericin-1 and DH-PBAN genes, which were localized within a 22-kb interval and are divergently oriented. These results show that it is possible to construct contigs and analyze chromosome organization using these libraries. Received: 12 October 1998 / Accepted: 10 February 1999     

Improved technique for the isolation of magnetotactic spirilla from a freshwater sediment and their phylogenetic characterization.

An improved technique for the isolation of magnetotactic bacteria was used for the axenic cultivation of microaerophilic magnetotactic spirilla. Magnetotactic bacteria were first separated from non-magnetic contaminants by exploiting their active migration along magnetic field lines by a capillary "racetrack" method. The purified magnetic cells were then inoculated into a two-layer isolation medium with opposing oxygen and sulfide gradients. Several strains of magnetotactic spirilla were isolated from a freshwater sediment sample using this method. Based on their morphology, physiology and comparative analysis of almost complete 16S rRNA gene sequences, all newly isolated strains were identified as members of the genus Magnetospirillum. While five of the isolates were closely related to previously described species (> 99% sequence similarity), two isolates appear to represent a third phylogenetic cluster within the genus Magnetospirillum.     

Transport of bacterial inoculants through intact cores of two different soils as affected by water percolation and the presence of wheat plants

Abstract Water flow-innduced transport of Burkholderia cepacia strain P2 and Pseudomonas fluorescens strain R2f cells through intact cores of loamy sand and silt loam field soils was measured for two percolation regimes, 0.9 and 4.4 mm h⁻¹, applied daily during 1 hour. For each strain, transport was generally similar between the two water regimes. Translocation of B. cepacia , with 4.4 mm h⁻¹, did occur initially in both soils. In the loamy sand soil, no change in the bacterial distribution occurred during the experiment (51 days). In the silt loam, B. cepacia cell numbers in the lower soil layers were significantly reduced, to levels at or below the limit of detection. Transport of P. fluorescens in both soils also occurred initially and was comparable to that of B. cepacia . Later in the experiment, P. fluorescens was not detectable in the lower soil layers of the loamy sand cores, due to a large decrease in surviving cell numbers. In the silt loam, the inoculant cell distribution did not change with time. Pre-incubation of the inoculated cores before starting percolation reduced B. cepacia inoculant transport in the loamy sand soil measured after 5 days, but not that determined after 54 days. Delayed percolation in the silt loam soil affected bacterial transport only after 54 days. The presence of growing wheat plants overall enhanced bacterial translocation as compared to that in unplanted soil cores, but only with percolating water. Percolation water from silt loam cores appeared the day after the onset of percolation and often contained inoculant bacteria. With loamy sand, percolation water appeared only 5 days after the start of percolation, and no inoculant bacteria were found. The results presented aid in predicting the fate of genetically manipulated bacteria in a field experiment.     

Different methods for extracting bacteria from freshwater sediment and a simple method to measure bacterial production in sediment samples

The efficiency of different treatments was tested to extract bacterial cells from freshwater sediment samples. The influence of sonication, density gradient centrifugation, fixation by formalin and centrifugation speed on bacterial recovery was investigated. The method developed by Smith and Azam [Mar. Microb. Food Webs 6 (1992) 107] to measure microbial activity on bacterioplankton (3H-leucine incorporation), was also evaluated in sediment samples. After 1 min of sonication bacterial abundance was reduced by about 47% in diluted sediments with tetrasodium pyrophosphate. With the addition of Percoll after sonication, bacterial counts were not significantly different (P<0.05). Fixation by formalin increased bacterial counts using sonication. However, higher bacterial abundance was estimated in non-sonicated samples. Bacterial abundance in samples centrifuged at 7000xg with and without Percoll was not significantly different (P<0.05). Highest bacterial abundance was obtained after centrifugation at low speed (750xg). Bacterial abundance decreased with higher centrifugation speed (750, 1500 and 3000xg), the difference, however, was not significant. Bacterial production ranged from 0.10 microg C cm(-3) d(-1) in autoclaved sediment to 0. 27 microg C cm(-3) d(-1) in untreated sediment. The radioactivity measured in controls of both untreated and autoclaved sediment was high (70 and 91%, respectively), indicating a high level of leucine adsorption in sediment particles. In contrast, radioactivity in control samples previously centrifuged was markedly lower (6%). Despite the high values of radioactivity in the controls, bacterial production in untreated sediment was significantly higher than in centrifuged sediment (P<0.05).     

Monosaccharide transport in the mammary gland of the intact lactating rat.

The Michaelis-Menten equation for the utilization of competing substrates was applied to the uptake of 2-deoxy[3H]glucose into the mammary gland of anaesthetized lactating rats. Intracellular water was calculated from total tissue water and sucrose space. Fed rats had a mean transport capacity of 2.2 mumol/min per g of tissue, giving an actual glucose transport in vivo of 1.1 mumol/min per g. Transport decreased by 90% on overnight starvation and returned to normal by 2 h of re-feeding. Similar changes were observed in the 1 min or 5 min transport of circulating 3-O-methylglucose. Transport of 3-O-methylglucose in starved rats was restored towards normal by insulin. In fed rats it increased between parturition and day 12 of lactation. The findings support the proposal that transport is a rate-limiting factor in the mammary utilization of carbohydrate.     

Isolation and characterization of the intact gastrin precursor from a gastrinoma

Antibodies to the extreme C-terminal region of human progastrin have been used to monitor the isolation of high-M_r immunoreactive material in a gastrinoma extract. Microsequence analysis of the product revealed amino acid residues in the first 18 positions corresponding to those predicted from the cDNA sequence for preprogastrin starting at position 22; the sequence and immunochemical data together allow the identification of this material as intact progastrin. Implications for gastrin biosynthesis are discussed.     

Fluxes of mercury to lake sediments in central and northern Canada inferred from dated sediment cores

Sediment cores were collected from lakes in central and northern Canada and from Hudson Bay to compare current and historic net depositions of mercury. Cores from most locations were enriched in mercury in the upper layers deposited recently relative to deeper, historic layers. The lakes with the greatest enrichments in mercury were located in central/southern Canada. This enrichment was interpreted as being of anthropogenic origin. Mercury inputs at the Experimental Lakes Area (ELA) in northwestern Ontario inferred from a core profile agreed well with inputs calculated independently from precipitation and runoff data. Anthropogenic inputs of mercury to northwestern Ontario were calculated to be about 9 µg m^-2 y^-1. Considering all the locations over the geographic range, the core profiles infer that fluxes of mercury have increased on average by about 2 fold over the past half century. This is consistent with results from other sites in North America and Europe.