william ghiorse | Cornell University (original) (raw)
Papers by william ghiorse
Applied and Environmental Microbiology, 1985
Mn 2+ exerted various effects on the growth of Leptothrix discophora strain SS-1 in batch culture... more Mn 2+ exerted various effects on the growth of Leptothrix discophora strain SS-1 in batch cultures depending on the concentration added to the medium. Concentrations of 0.55 to 5.5 μM Mn 2+ , comparable to those in the environment from which strain SS-1 was isolated, decreased cell yield and prolonged stationary-phase survival, but did not affect growth rate. Elevated concentrations of 55 to 910 μM Mn 2+ also decreased cell yield and prolonged survival, but growth rate was decreased as well. The addition of 1,820 μM Mn 2+ caused a decline in cell numbers followed by an exponential rise after 80 h of incubation, indicating the development of a population of cells resistant to Mn 2+ toxicity. When 360 μM Mn 2+ or less was added to growth flasks, Mn 2+ was oxidized to manganese oxide (MnO x , where x is ∼2), which appeared as brown particles in the medium. Quantification of Mn oxidation during growth of cultures to which 55 μM Mn 2+ was added showed that nearly all of the Mn 2+ was oxi...
Applied and Environmental Microbiology, 1992
Leptothrix discophora SP-6 was isolated from the outflow reservoir of an artificial iron seep. It... more Leptothrix discophora SP-6 was isolated from the outflow reservoir of an artificial iron seep. Its sheathforming phenotype was maintained by slow growth in a mineral salts-vitamin-pyruvate medium under minimal aeration at 20 to 25°C. A sheathless variant, SP-6(sl), was isolated from smooth colonies that appeared on spread plates after rapid growth of SP-6 in well-aerated cultures. SP-6 and SP-6(sl) are closely related but not identical to the previously studied sheathless strain SS-1 (ATCC 43182). Increasing Mn 2+ concentrations in the growth medium of SP-6 increased the phase density of the sheath, indicating increased Mn oxide deposition in the sheath. Electron microscopy of cultures grown without added Mn 2+ revealed that the sheath consisted of a well-defined inner layer, 30 to 100 nm thick, and a diffuse outer capsular layer of variable thickness. Mn oxides were identified in the sheath by their characteristic ultrastructure, electron density, and X-ray-dispersive energy spectr...
Applied and Environmental Microbiology, 1985
The bacterial microflora of two shallow aquifers (saturated subsurface zones) in Oklahoma was cha... more The bacterial microflora of two shallow aquifers (saturated subsurface zones) in Oklahoma was characterized by direct observation with light and electron microscopy, by plating, and by examination of colony morphology and distribution. Isolated bacterial strains were also examined. Total cell counts varied only slightly (2.9 × 10 6 to 9.8 × 10 6 g [dry wt] −1 ) from sample to sample, whereas colony counts varied widely (6.3 × 10 2 to 6.5 × 10 6 CFU g [dry wt] −1 ). Colony counts on nutritionally rich media were lower than on low-nutrient media, especially in samples from the saturated zone. The variety of colony types growing on nutritionally rich media decreased with increasing depth and saturation. Colony counts of anaerobic bacteria also decreased with depth but were at least 100-fold lower than aerobic counts on most media. Cell morphologies of bacteria grown aerobically on plates included short rods, cocci, and actinomycete-like forms. Direct light microscopic observation of se...
Applied and environmental microbiology, 1997
A computer-assisted laser scanning microscope equipped for confocal laser scanning and color vide... more A computer-assisted laser scanning microscope equipped for confocal laser scanning and color video microscopy was used to examine Cryptosporidium parvum oocysts in two agricultural soils, a barnyard sediment, and calf fecal samples. An agar smear technique was developed for enumerating oocysts in soil and barnyard sediment samples. Enhanced counting efficiency and sensitivity (detection limit, 5.2 x 10(sup2) oocysts(middot)g [dry weight](sup-1)) were achieved by using a semiautomatic counting procedure and confocal laser scanning microscopy to enumerate immunostained oocysts and fragments of oocysts in the barnyard sediment. An agarose-acridine orange mounting procedure was developed for high-resolution confocal optical sectioning of oocysts in soil. Stereo images of serial optical sections revealed the three-dimensional spatial relationships between immunostained oocysts and the acridine orange-stained soil matrix material. In these hydrated, pyrophosphate-dispersed soil preparatio...
Applied and Environmental Microbiology, 1997
Horizontal transfer of genes responsible for pollutant biodegradation may play a key role in the ... more Horizontal transfer of genes responsible for pollutant biodegradation may play a key role in the evolution of bacterial populations and the adaptation of microbial communities to environmental contaminants. However, field evidence for horizontal gene transfer between microorganisms has traditionally been very difficult to obtain. In this study, the sequences of the 16S rRNA and naphthalene dioxygenase iron-sulfur protein (nahAc) genes of nine naphthalene-degrading bacteria isolated from a coal tar waste-contaminated site, as well as a naphthalene-degrading bacterium from a contaminated site in Washington state and two archetypal naphthalene-degrading strains, were compared. Seven strains from the study site had a single nahAc allele, whereas the 16S rRNA gene sequences of the strains differed by as much as 7.9%. No nahAc alleles from the site were identical to those of the archetypal strains, although the predominant allele was closely related to that of Pseudomonas putida NCIB 9816...
Applied and Environmental Microbiology, 1991
A bioreactor apparatus is described for studying bacterial attachment. A cyclic, on-off, flow reg... more A bioreactor apparatus is described for studying bacterial attachment. A cyclic, on-off, flow regime was imposed within the apparatus. Model calculations illustrate the utility of this flow pattern in the selection and maintenance of slow-growing, adherent organisms. The apparatus is believed to have general utility in testing bacterial attachment influenced by many types of experimental or environmental constraints, including variations in fluid dynamics, presence of toxic substances (metals or organics), nature of the substratum surface, concentrations of limiting nutrients, and competition between bacterial strains. As an example application, the apparatus was employed to test 14 bacterial strains for surface attachment in a nutrient-limited growth medium. The medium was developed, using the chemical equilibrium program MINEQL, for planned studies of biofilms in a solution with a chemically defined composition that permits calculation of trace metal speciation. The apparatus was ...
Applied and Environmental Microbiology, 1994
This study reports improvements in two of the key steps, lysis of indigenous cells and DNA purifi... more This study reports improvements in two of the key steps, lysis of indigenous cells and DNA purification, required for achieving a rapid nonselective protocol for extracting nucleic acids directly from sodium dodecyl sulfate (SDS)-treated sediment rich in organic matter. Incorporation of bead-mill homogenization into the DNA extraction procedure doubled the densitometrically determined DNA yield (11.8 micrograms of DNA.g [dry weight] of sediment-1) relative to incorporation of three cycles of freezing and thawing (5.2 micrograms of DNA.g [dry weight] of sediment-1). The improved DNA extraction efficiency was attributed to increased cell lysis, measured by viable counts of sediment microorganisms which showed that 2 and 8%, respectively, survived the bead-mill homogenization and freeze-thaw procedures. Corresponding measurements of suspensions of viable Bacillus endospores demonstrated that 2 and 94% of the initial number survived. Conventional, laser scanning epifluorescence phase-co...
Applied and Environmental Microbiology, 1993
We report the amplification of bacterial genes from uninoculated surface and subsurface sediments... more We report the amplification of bacterial genes from uninoculated surface and subsurface sediments by the polymerase chain reaction (PCR). PCR amplification of indigenous bacterial 16S ribosomal DNA genes was unsuccessful when subsurface sediment containing approximately 10(7) cells.g-1 was added directly to a PCR mixture. However, when 10 mg of sediment was inoculated with approximately 10(5) cells of Pseudomonas putida G7, the nahAc naphthalene dioxygenase gene characteristic of the P. putida G7 NAH7 plasmid was detected by PCR amplification. Southern blotting of the PCR amplification product improved sensitivity to 10(3) to 10(4) cells from samples inoculated with P. putida G7, but controls with no sediment added showed that the PCR was partially inhibited by the sediments. Lysozyme-sodium dodecyl sulfate-freeze-thaw DNA extraction was combined with gel electrophoretic partial purification in the presence of polyvinylpyrrolidone to render DNA from indigenous bacteria in surface or...
Applied and Environmental Microbiology, 1999
Biogenic Mn oxides were produced by the bacterium Leptothrix discophora SS-1 (= ATCC 3182) in a c... more Biogenic Mn oxides were produced by the bacterium Leptothrix discophora SS-1 (= ATCC 3182) in a chemically defined mineral salts medium, and the Pb binding and specific surface area of these oxides were characterized. Growth of SS-1 in the defined medium with pyruvate as a carbon and energy source required the addition of vitamin B 12 . Complete oxidation of Mn(II) within 60 h required the addition of ≥0.1 μM FeSO 4 . Pb adsorption isotherms were determined for the biogenic Mn oxides (and associated cells with their extracellular polymer) and compared to the Pb adsorption isotherms of cells and exopolymer alone, as well as to abiotic Mn oxides. The Pb adsorption to cells and exopolymer with biogenic Mn oxides (0.8 mmol of Mn per g) at pH 6.0 and 25°C was 2 orders of magnitude greater than the Pb adsorption to cells and exopolymer alone (on a dry weight basis). The Pb adsorption to the biogenic Mn oxide was two to five times greater than the Pb adsorption to a chemically precipitated...
Deep Sea Research Part B. Oceanographic Literature Review, 1982
Hyphal budding bacteria were observed by electron microscopy in thin sections of surface material... more Hyphal budding bacteria were observed by electron microscopy in thin sections of surface material from Baltic Sea ferromanganese concretions. Similar bacteria were also observed in and isolated from enrichment cultures prepared from the same concretion material. Three morphologically similar strains of Mn-Fedepositing budding bacteria were isolated from the enrichment cultures. Strain B-4 possessed extracellular anionic polymers that accumulated Mn oxides. Mn deposition by B-4 was inhibited by elevated concentrations of Mn, 0.05% glutaraldehyde, 0.1 mM HgCl2, and heating at 93°C for 15 min, suggesting the participation of an enzyme protein in the Mn-depositing activity.
Journal of Bacteriology, 1993
Light microscopy and transmission electron microscopy of thin sections and metal-shadowed specime... more Light microscopy and transmission electron microscopy of thin sections and metal-shadowed specimens showed that the sheath of Leptothrix discophora SP-6 (ATCC 51168) is a tube-like extracellular polymeric structure consisting of a condensed fabric of 6.5-nm-diameter fibrils underlying a more diffuse outer capsular layer. In thin sections, outer membrane bridges seen to contact the inner sheath layer suggested that the sheath fabric was attached to the outer layer of the gram-negative cell wall. The capsular polymers showed an affinity for cationic colloidal iron and polycationic ferritin, indicating that they carry a negative charge. Cell-free sheaths were isolated by treatment with a mixture of lysozyme, EDTA, and N-lauroylsarcosine (Sarkosyl) or sodium dodecyl sulfate (SDS). Both Sarkosyl- and SDS-isolated sheaths were indistinguishable in microscopic appearance. However, the Mn-oxidizing activity of Sarkosyl-isolated sheaths was more stable than that of SDS-isolated sheaths. The ...
International Journal of Systematic Bacteriology, 1996
Transport and deposition of colloidal Fe, Mn and A! oxides play key roles in the cycling of toxic... more Transport and deposition of colloidal Fe, Mn and A! oxides play key roles in the cycling of toxic transition metals in aquatic environments because these colloids strongly bind transition metals. Further, attachment of biological cells and biofilm growth on surfaces can indirectly affect toxic metal distribution by influencing the deposition of colloidal oxides to surfaces. To elucidate the mechanisms governing these processes, deposition of colloidal oxides onto surfaces must be evaluated in the presence of suspended and adherent bacterial cells. Both particle size and concentration are expected to influence deposition. An experimental protocol was developed to determine the size distribution of iron colloids in mixtures with suspended ceils. A Ti(III) reagent was used to reduce and dissolve colloidal Fe(III) from mixtures containing both suspended cells and Fe colloids. The size distribution of Fe(III) colloids in the original solutl[on was then determined from the difference between size distributions before and after dissolution of Fe with Ti(III). The Ti(III) reagent dissolved over 95% of the Fe colloids without altering the size distribution of suspended bacterial ceils, and the method accurately determined the size distribution of Fe colloids added to cell suspensions. The applicability of this protocol was tested by applying it to a study of the deposition of Fe(III) oxide particles onto glass surfaces with and without biofilms of the bacterium Burkholdaria cepacia 17616. Experimental results using a laboratory biofilm reactor indicated that the deposition rate of Fe(III) colloids was not significantly affected by the presence of B. cepacia biofilms or by the presence of previously deposited Fe. However, deposition of Fe to reactor surfaces other than the glass surfaces may have interfered with the analyses, and atomic absorption measurements showed a slight increase in Fe deposition onto glass surfaces with biofilms present. Fe deposition to the composite of all reactor surfaces increased with increasing colloidal particle size, indicating a dominance of interception and/or sedimentation in controlling Fe deposition on surfaces in the biofilm reactor.
caused more rapid degradation ofthesheath thancyanide or sulfite. Treatment ofthesheath with1N Na... more caused more rapid degradation ofthesheath thancyanide or sulfite. Treatment ofthesheath with1N NaOH resulted inrapid breakdown, while treatment with1N HCIresulted inslowbut significant hydrolysis. Transmission electron microscopy showedthatthe6.5-nm fibrils previously showntobe an integral structural element ofthesheath fabric (D.EmersonandW.C.Ghiorse, J.Bacteriol. 175:7808-7818, 1993) were progressively dissociated intorandommasses duringDTT-induced degradation. Quantitation of disulfide bondswithDTT showedthatthesheaths contained approximately 2.2,umol ofdisulfides per mg of sheathprotein. Reaction with5,5'-dithio-bis-(2-nitrobenzoic acid)showedthatsheaths alsocontained approximately 0.8,umoloffreesulfhydryls per mg ofprotein. A sulfhydryl-specific fluorescent probe (fluorescein 5-maleimide) showedthatthefreesulfhydryls insheathed cell filaments were evenly distributed throughout thesheath. Sodiumdodecyl sulfate-polyacrylamide gelelectrophoresis autoradiography of ('4C)i...
Water Research, 2000
ÐSurface coatings (bio®lms and associated minerals) were collected on glass slides in the oxic su... more ÐSurface coatings (bio®lms and associated minerals) were collected on glass slides in the oxic surface waters of Cayuga Lake (New York State, U.S.A.) and were used to evaluate the relative contributions of Fe, Mn and Al oxides and organic material to total observed Pb and Cd adsorption by the surface coating materials. Several alternative selective extraction techniques were evaluated with respect to both selectivity and alteration of the residual unextracted material. Pb and Cd adsorption was measured under controlled laboratory conditions (mineral salts solution with de®ned metal speciation, ionic strength 0.05 M, 258C and pH 6.0) before and after extractions to determine by dierence the adsorptive properties of the extracted component(s). Hydroxylamine hydrochloride (0.01 M NH 2 OHÁHCl+0.01 M HNO 3) was used to selectively remove Mn oxides, sodium dithionite (0.3 M Na 2 S 2 O 4) was used to remove Mn and Fe oxides, and 10% oxalic acid was used to remove metal oxides and organic materials. Several other extractants were evaluated, but preliminary experiments indicated that they were not suitable for these experiments because of undesirable alterations of the residual, unextracted material. The selected extraction methods removed target components with eciencies between 71 and 83%, but signi®cant amounts of metal oxides and organic materials other than the target components were also removed by the extractants (up to 39%). Nonlinear regression analysis of the observed Pb and Cd adsorption based on the assumption of additive Langmuir adsorption isotherms was used to estimate the relative contributions of each surface coating constituent to total Pb and Cd binding of the bio®lms. Adsorption of Cd to the lake bio®lms was dominated by Fe oxides, with lesser roles attributed to adsorption by Mn and Al oxides and organic material. Adsorption of Pb was dominated by Mn oxides, with lesser roles indicated for adsorption to Fe oxides and organic material, and the estimated contribution of Al oxides to Pb adsorption was insigni®cant. The ®tted Pb adsorption isotherm for Fe oxides was in excellent agreement with those obtained through direct experiments and reported in independent investigations. The estimated Pb distribution between surface coating components also agreed well with that previously predicted by an additive adsorption model based on Pb adsorption isotherms for laboratory surrogates for Mn, Fe and Al oxides and de®ned biological components.
Water Research, 1995
A~tract-Biofilms influence the transport and fate of heavy metals in aquatic environments both di... more A~tract-Biofilms influence the transport and fate of heavy metals in aquatic environments both directly by adsorption and complexation reactions and indirectly via interactions with oxides of iron and manganese. These reactions were investigated by introducing lead into a continuous-flow biofilm reactor that was designed to simulate conditions in a flowing freshwater aquatic environment. The reactor provided controlled conditions, and use of a chemically-defined growth medium allowed calculation of lead speciation with a chemical equilibrium program (MINEQL). Pseudomonas cepacia was employed as a test cell strain because of its ability to grow and form biofilms in the defined medium. This bacterium affected lead distribution in the reactor by adsorbing lead both to adherent and suspended cells. When the aqueous bulk lead concentration was 1.4 0.1/~M and biofilm coverage (measured as chemical oxygen demand, COD) was 50 mequiv COD/m 2, lead adsorption was increased by about a factor of five relative to bare glass. Of the total lead in solution, only 1% was adsorbed to suspended cells (5 x 10 7 ceUs/ml). Lead adsorption to biofilms followed a Langmuir isotherm with a maximum adsorption (Fmax) of 56/~mol Pb/equiv COD and an adsorption equilibrium constant (K) of 0.64 liter/#mol Pb. Lead complexed with dissolved bacterial exopolymer was below detection limits. Pretreatment of glass slides with colloidal iron also significantly increased lead adsorption relative to bare glass. Lead adsorption to adsorbed iron fit a Langmuir isotherm with Fma x 50 #mol Pb/mol Fe, and K 1.3 liter/#mol Pb. Lead binding to glass coated with both cells and iron was additive, and could be predicted by summing adsorption predicted using isotherms for each constituent. The presence of iron surface coatings increased initial biofilm formation rates, but after reaching steady state conditions, biofilm coverage was similar for slides treated with iron and untreated slides. A concentration of 1 # M lead produced a transient reduction in suspended cell counts. Cell counts recovered to the original cell density over the course of five to ten reactor retention times. With iron present, the magnitude of the reduction in cell concentration in response to the addition of lead was greatly reduced, suggesting that toxic effects of lead may be reduced by iron.
Water Environment Research, 2003
When oocysts of the protozoan Cryptosporidium parvum contaminate drinking water supplies, they ca... more When oocysts of the protozoan Cryptosporidium parvum contaminate drinking water supplies, they can cause outbreaks of Cryptosporidiosis, a common waterborne disease. Of the different pathways by which oocysts can wind up in drinking water, one has received little attention to date; that is, because soils are often considered to be perfect filters, the transport of oocysts through the subsoil to groundwater is generally ignored. To evaluate the significance of this pathway, a series of laboratory experiments investigated subsurface transport of oocysts. Experiment 1 was carried out in a vertical 18-cm-long column filled either with glass beads or silica sand, under conditions known to foster fingered flow. Experiment 2 involved undisturbed, macroporous soil columns subjected to macropore flow. Experiment 3 aimed to study the lateral flow on an undisturbed soil block. The columns and soil samples were subjected to artificial rainfall and were allowed to reach steady state. At that point, feces of contaminated calves were applied at the surface along with a known amount of potassium chloride to serve as a tracer, and rainfall was continued at the same rate. The breakthrough of oocysts and chloride, monitored in the effluent, demonstrate the importance of preferential flow on the transport of oocysts. Compared with chloride, peak oocyst concentrations were not appreciably delayed and, in some cases, occurred even before the chloride peak. Recovery rates for oocysts were low, ranging from 0.1 to 10.4% of the oocysts originally applied on the columns. However, the numbers of oocysts present in the effluents were still orders of magnitude higher than 10 oocysts, the infectious dose considered by the U.S. Food and Drug Administration, Center for Food Safety and Applied Nutrition, to be sufficient to cause Cryptosporidiosis in healthy adults. These results suggest that the transport of oocysts in the subsurface via preferential flow may create a significant risk of groundwater contamination in some situations. Water Environ. Res., 75, 113 (2003).
Vadose Zone Journal, 2004
in the form of 4-to 6-m-long ovoid-shaped oocysts, with a double wall that is resistant to most o... more in the form of 4-to 6-m-long ovoid-shaped oocysts, with a double wall that is resistant to most oxidation As a result of Cryptosporidium parvum in drinking water, several processes such as ozonation and chlorination (Current, outbreaks of cryptosporidiosis have occurred in the last 10 yr. Al-1986; Atwill et al., 1997). though it is generally believed that movement of pathogens through the soil is minimal, recent research has shown that appreciable num-During the past two decades, the presence of C. parbers of C. parvum oocysts may be transported via preferential or vum in surface-and groundwaters in the United States fingered flow to groundwater. The objective of the present research and Great Britain (Galbraith et al., 1987; Rose et al., was to further investigate and model the transport of oocysts through 1991; Craun et al., 1998) has been associated with several preferential flow paths in the vadose zone under a "worst-case" scemajor outbreaks of cryptosporidiosis (Hayes et al., 1989; nario. This was studied by adding calves feces containing C. parvum MacKenzie et al., 1994). Among the different pathways oocysts with a Cl Ϫ tracer to undisturbed silt loam columns and disfor the transport of oocysts to drinking water sources, turbed sand columns during a simulated steady-state rain. The sand columns exhibited preferential flow in the form of fingers whereas downward percolation is usually considered to be insigmacropore flow occurred in the undisturbed cores. In the columns nificant, because soils are generally assumed to be an with fingered flow, oocysts and Cl were transported rapidly with the effective filter for a wide range of pathogens. Studies same velocity through the columns. Although only 14 to 86% of the of packed columns with saturated flow by Brush et al. amount applied, the number of oocysts transported across the columns (1999) and Harter et al. (2000) and undisturbed columns was several orders of magnitude above an infective dose. The macwith unsaturated flow (Mawdsley et al., 1996), however, ropore columns had only a very limited breakthrough of oocysts, showed that C. parvum oocysts could be transported which appeared several pore volumes after the Cl broke through initially. A simulation model for the transport of oocysts via preferen-rapidly downward through the soil. Although transport tial flow was developed on the basis of an existing preferential flow of C. parvum oocysts in saturated flow has been studied model for nonadsorbing solutes, with addition of a first-order sink experimentally and described mathematically (Brush et term for adsorbance of the C. parvum to the air-water-solid (AWS) al., 1999; Harter et al., 2000), detailed observations of interfaces, and with velocity and dispersivity parameters derived from the transport and persistence of C. parvum oocysts in Cl Ϫ transport. The breakthrough of C. parvum oocysts could be deunsaturated soils with preferential flow are still lacking, scribed realistically for the sand columns. However, the model could particularly in the presence of preferential flow pronot describe oocyst transport in the columns with macropores. Three sets of experiments were performed in which C. parvum oocysts contained in the feces of infected calves were
Limnology and Oceanography, 1999
The role of the composition of surface-coating materials in controlling trace metal adsorption in... more The role of the composition of surface-coating materials in controlling trace metal adsorption in aquatic environments was investigated using natural biofilms that developed on glass slides in three New York State lakes and a water-supply well. Adsorption isotherms were obtained for Pb binding to each of the biofilms in solutions with defined Pb speciation at 25ЊC and pH 6.0, with Pb concentrations ranging from 0.2 to 2.0 M. Adsorption isotherms for Pb binding to laboratory-derived metal oxides and surrogate organic materials were determined under the same conditions. These isotherms, combined with characterization of natural biofilm composition, were used to estimate the relative contributions of the organic and metal oxide surface-coating constituents by assuming additivity of adsorption to discrete adsorbing phases. Cells of a diatom (Navicula peliculosa), a green alga (Chlorella vulgaris), the bacterium Leptothrix discophora, and extracellular polymer of the bacterium Burkholdaria cepacia were tested as laboratory analogs for the organic phase of the biofilms. Amorphous Fe oxyhydroxide, ␥Al 2 O 3 , and a laboratoryderived biogenic Mn oxyhydroxide were used as laboratory surrogates for biofilm minerals. The sum total of predicted Pb binding to the defined surrogates accounted for at least 90% of the total observed Pb binding in the three lake biofilms and 60% of that observed for the well biofilms. For the lake biofilms, Pb adsorption to Fe and Mn oxides was significantly greater than that estimated for organic materials. The use of biogenic Mn oxide as a model component resulted in an estimated Pb adsorption to Mn oxyhydroxides in the lake biofilms up to four times greater than that estimated for Fe oxyhydroxide. Estimated Pb binding by Al oxide was negligible for all four biofilms. These results suggest that Fe and biogenic Mn oxides exert the greatest influence on Pb adsorption in oxic freshwater environments at pH 6.0. This research was supported by NSF grants BES-97067715 and CHE-9708093 and through a STAR fellowship from the U.S. EPA (Y.N.).
Journal of Parasitology, 2001
Cryptosporidium parvum oocysts were examined to ascertain excystation requirements and the effect... more Cryptosporidium parvum oocysts were examined to ascertain excystation requirements and the effects of gamma irradiation. Oocysts and excysted sporozoites were examined for dye permeability and infectivity. Maximum excystation occurred when oocysts were pretreated with acid and incubated with bile salts, and potassium or sodium bicarbonate. Pretreatment with Hanks' balanced salt solution or NaCl lowered excystation; however, this effect was overcome with acid. Sodium ions were replaceable with potassium ions, and sodium bicarbonate was replaceable with sodium phosphate. Oocysts that received 200 krad irradiation excysted at the same rates as nonirradiated oocysts (95%), the excystation rates were lowered (50%) by 2,000 krad, and no excystation was observed by 5,000 krad. No differences were observed between the propidium iodide (PI) permeability of untreated oocysts and oocysts treated with 200 krad, while 92% of oocysts were PI positive after 2,000 krad. Most of the sporozoites exposed to 2,000 krad were not viable as indicated by the dye permeability assay. The oocysts irradiated with 200 and 2,000 krad infected cells, but no replication was observed. The results suggest that gamma-irradiated oocysts may still be capable of excystation and apparent infection; however, because the sporozoites could not reproduce they must not have been viable.
Applied and Environmental Microbiology, 1985
Mn 2+ exerted various effects on the growth of Leptothrix discophora strain SS-1 in batch culture... more Mn 2+ exerted various effects on the growth of Leptothrix discophora strain SS-1 in batch cultures depending on the concentration added to the medium. Concentrations of 0.55 to 5.5 μM Mn 2+ , comparable to those in the environment from which strain SS-1 was isolated, decreased cell yield and prolonged stationary-phase survival, but did not affect growth rate. Elevated concentrations of 55 to 910 μM Mn 2+ also decreased cell yield and prolonged survival, but growth rate was decreased as well. The addition of 1,820 μM Mn 2+ caused a decline in cell numbers followed by an exponential rise after 80 h of incubation, indicating the development of a population of cells resistant to Mn 2+ toxicity. When 360 μM Mn 2+ or less was added to growth flasks, Mn 2+ was oxidized to manganese oxide (MnO x , where x is ∼2), which appeared as brown particles in the medium. Quantification of Mn oxidation during growth of cultures to which 55 μM Mn 2+ was added showed that nearly all of the Mn 2+ was oxi...
Applied and Environmental Microbiology, 1992
Leptothrix discophora SP-6 was isolated from the outflow reservoir of an artificial iron seep. It... more Leptothrix discophora SP-6 was isolated from the outflow reservoir of an artificial iron seep. Its sheathforming phenotype was maintained by slow growth in a mineral salts-vitamin-pyruvate medium under minimal aeration at 20 to 25°C. A sheathless variant, SP-6(sl), was isolated from smooth colonies that appeared on spread plates after rapid growth of SP-6 in well-aerated cultures. SP-6 and SP-6(sl) are closely related but not identical to the previously studied sheathless strain SS-1 (ATCC 43182). Increasing Mn 2+ concentrations in the growth medium of SP-6 increased the phase density of the sheath, indicating increased Mn oxide deposition in the sheath. Electron microscopy of cultures grown without added Mn 2+ revealed that the sheath consisted of a well-defined inner layer, 30 to 100 nm thick, and a diffuse outer capsular layer of variable thickness. Mn oxides were identified in the sheath by their characteristic ultrastructure, electron density, and X-ray-dispersive energy spectr...
Applied and Environmental Microbiology, 1985
The bacterial microflora of two shallow aquifers (saturated subsurface zones) in Oklahoma was cha... more The bacterial microflora of two shallow aquifers (saturated subsurface zones) in Oklahoma was characterized by direct observation with light and electron microscopy, by plating, and by examination of colony morphology and distribution. Isolated bacterial strains were also examined. Total cell counts varied only slightly (2.9 × 10 6 to 9.8 × 10 6 g [dry wt] −1 ) from sample to sample, whereas colony counts varied widely (6.3 × 10 2 to 6.5 × 10 6 CFU g [dry wt] −1 ). Colony counts on nutritionally rich media were lower than on low-nutrient media, especially in samples from the saturated zone. The variety of colony types growing on nutritionally rich media decreased with increasing depth and saturation. Colony counts of anaerobic bacteria also decreased with depth but were at least 100-fold lower than aerobic counts on most media. Cell morphologies of bacteria grown aerobically on plates included short rods, cocci, and actinomycete-like forms. Direct light microscopic observation of se...
Applied and environmental microbiology, 1997
A computer-assisted laser scanning microscope equipped for confocal laser scanning and color vide... more A computer-assisted laser scanning microscope equipped for confocal laser scanning and color video microscopy was used to examine Cryptosporidium parvum oocysts in two agricultural soils, a barnyard sediment, and calf fecal samples. An agar smear technique was developed for enumerating oocysts in soil and barnyard sediment samples. Enhanced counting efficiency and sensitivity (detection limit, 5.2 x 10(sup2) oocysts(middot)g [dry weight](sup-1)) were achieved by using a semiautomatic counting procedure and confocal laser scanning microscopy to enumerate immunostained oocysts and fragments of oocysts in the barnyard sediment. An agarose-acridine orange mounting procedure was developed for high-resolution confocal optical sectioning of oocysts in soil. Stereo images of serial optical sections revealed the three-dimensional spatial relationships between immunostained oocysts and the acridine orange-stained soil matrix material. In these hydrated, pyrophosphate-dispersed soil preparatio...
Applied and Environmental Microbiology, 1997
Horizontal transfer of genes responsible for pollutant biodegradation may play a key role in the ... more Horizontal transfer of genes responsible for pollutant biodegradation may play a key role in the evolution of bacterial populations and the adaptation of microbial communities to environmental contaminants. However, field evidence for horizontal gene transfer between microorganisms has traditionally been very difficult to obtain. In this study, the sequences of the 16S rRNA and naphthalene dioxygenase iron-sulfur protein (nahAc) genes of nine naphthalene-degrading bacteria isolated from a coal tar waste-contaminated site, as well as a naphthalene-degrading bacterium from a contaminated site in Washington state and two archetypal naphthalene-degrading strains, were compared. Seven strains from the study site had a single nahAc allele, whereas the 16S rRNA gene sequences of the strains differed by as much as 7.9%. No nahAc alleles from the site were identical to those of the archetypal strains, although the predominant allele was closely related to that of Pseudomonas putida NCIB 9816...
Applied and Environmental Microbiology, 1991
A bioreactor apparatus is described for studying bacterial attachment. A cyclic, on-off, flow reg... more A bioreactor apparatus is described for studying bacterial attachment. A cyclic, on-off, flow regime was imposed within the apparatus. Model calculations illustrate the utility of this flow pattern in the selection and maintenance of slow-growing, adherent organisms. The apparatus is believed to have general utility in testing bacterial attachment influenced by many types of experimental or environmental constraints, including variations in fluid dynamics, presence of toxic substances (metals or organics), nature of the substratum surface, concentrations of limiting nutrients, and competition between bacterial strains. As an example application, the apparatus was employed to test 14 bacterial strains for surface attachment in a nutrient-limited growth medium. The medium was developed, using the chemical equilibrium program MINEQL, for planned studies of biofilms in a solution with a chemically defined composition that permits calculation of trace metal speciation. The apparatus was ...
Applied and Environmental Microbiology, 1994
This study reports improvements in two of the key steps, lysis of indigenous cells and DNA purifi... more This study reports improvements in two of the key steps, lysis of indigenous cells and DNA purification, required for achieving a rapid nonselective protocol for extracting nucleic acids directly from sodium dodecyl sulfate (SDS)-treated sediment rich in organic matter. Incorporation of bead-mill homogenization into the DNA extraction procedure doubled the densitometrically determined DNA yield (11.8 micrograms of DNA.g [dry weight] of sediment-1) relative to incorporation of three cycles of freezing and thawing (5.2 micrograms of DNA.g [dry weight] of sediment-1). The improved DNA extraction efficiency was attributed to increased cell lysis, measured by viable counts of sediment microorganisms which showed that 2 and 8%, respectively, survived the bead-mill homogenization and freeze-thaw procedures. Corresponding measurements of suspensions of viable Bacillus endospores demonstrated that 2 and 94% of the initial number survived. Conventional, laser scanning epifluorescence phase-co...
Applied and Environmental Microbiology, 1993
We report the amplification of bacterial genes from uninoculated surface and subsurface sediments... more We report the amplification of bacterial genes from uninoculated surface and subsurface sediments by the polymerase chain reaction (PCR). PCR amplification of indigenous bacterial 16S ribosomal DNA genes was unsuccessful when subsurface sediment containing approximately 10(7) cells.g-1 was added directly to a PCR mixture. However, when 10 mg of sediment was inoculated with approximately 10(5) cells of Pseudomonas putida G7, the nahAc naphthalene dioxygenase gene characteristic of the P. putida G7 NAH7 plasmid was detected by PCR amplification. Southern blotting of the PCR amplification product improved sensitivity to 10(3) to 10(4) cells from samples inoculated with P. putida G7, but controls with no sediment added showed that the PCR was partially inhibited by the sediments. Lysozyme-sodium dodecyl sulfate-freeze-thaw DNA extraction was combined with gel electrophoretic partial purification in the presence of polyvinylpyrrolidone to render DNA from indigenous bacteria in surface or...
Applied and Environmental Microbiology, 1999
Biogenic Mn oxides were produced by the bacterium Leptothrix discophora SS-1 (= ATCC 3182) in a c... more Biogenic Mn oxides were produced by the bacterium Leptothrix discophora SS-1 (= ATCC 3182) in a chemically defined mineral salts medium, and the Pb binding and specific surface area of these oxides were characterized. Growth of SS-1 in the defined medium with pyruvate as a carbon and energy source required the addition of vitamin B 12 . Complete oxidation of Mn(II) within 60 h required the addition of ≥0.1 μM FeSO 4 . Pb adsorption isotherms were determined for the biogenic Mn oxides (and associated cells with their extracellular polymer) and compared to the Pb adsorption isotherms of cells and exopolymer alone, as well as to abiotic Mn oxides. The Pb adsorption to cells and exopolymer with biogenic Mn oxides (0.8 mmol of Mn per g) at pH 6.0 and 25°C was 2 orders of magnitude greater than the Pb adsorption to cells and exopolymer alone (on a dry weight basis). The Pb adsorption to the biogenic Mn oxide was two to five times greater than the Pb adsorption to a chemically precipitated...
Deep Sea Research Part B. Oceanographic Literature Review, 1982
Hyphal budding bacteria were observed by electron microscopy in thin sections of surface material... more Hyphal budding bacteria were observed by electron microscopy in thin sections of surface material from Baltic Sea ferromanganese concretions. Similar bacteria were also observed in and isolated from enrichment cultures prepared from the same concretion material. Three morphologically similar strains of Mn-Fedepositing budding bacteria were isolated from the enrichment cultures. Strain B-4 possessed extracellular anionic polymers that accumulated Mn oxides. Mn deposition by B-4 was inhibited by elevated concentrations of Mn, 0.05% glutaraldehyde, 0.1 mM HgCl2, and heating at 93°C for 15 min, suggesting the participation of an enzyme protein in the Mn-depositing activity.
Journal of Bacteriology, 1993
Light microscopy and transmission electron microscopy of thin sections and metal-shadowed specime... more Light microscopy and transmission electron microscopy of thin sections and metal-shadowed specimens showed that the sheath of Leptothrix discophora SP-6 (ATCC 51168) is a tube-like extracellular polymeric structure consisting of a condensed fabric of 6.5-nm-diameter fibrils underlying a more diffuse outer capsular layer. In thin sections, outer membrane bridges seen to contact the inner sheath layer suggested that the sheath fabric was attached to the outer layer of the gram-negative cell wall. The capsular polymers showed an affinity for cationic colloidal iron and polycationic ferritin, indicating that they carry a negative charge. Cell-free sheaths were isolated by treatment with a mixture of lysozyme, EDTA, and N-lauroylsarcosine (Sarkosyl) or sodium dodecyl sulfate (SDS). Both Sarkosyl- and SDS-isolated sheaths were indistinguishable in microscopic appearance. However, the Mn-oxidizing activity of Sarkosyl-isolated sheaths was more stable than that of SDS-isolated sheaths. The ...
International Journal of Systematic Bacteriology, 1996
Transport and deposition of colloidal Fe, Mn and A! oxides play key roles in the cycling of toxic... more Transport and deposition of colloidal Fe, Mn and A! oxides play key roles in the cycling of toxic transition metals in aquatic environments because these colloids strongly bind transition metals. Further, attachment of biological cells and biofilm growth on surfaces can indirectly affect toxic metal distribution by influencing the deposition of colloidal oxides to surfaces. To elucidate the mechanisms governing these processes, deposition of colloidal oxides onto surfaces must be evaluated in the presence of suspended and adherent bacterial cells. Both particle size and concentration are expected to influence deposition. An experimental protocol was developed to determine the size distribution of iron colloids in mixtures with suspended ceils. A Ti(III) reagent was used to reduce and dissolve colloidal Fe(III) from mixtures containing both suspended cells and Fe colloids. The size distribution of Fe(III) colloids in the original solutl[on was then determined from the difference between size distributions before and after dissolution of Fe with Ti(III). The Ti(III) reagent dissolved over 95% of the Fe colloids without altering the size distribution of suspended bacterial ceils, and the method accurately determined the size distribution of Fe colloids added to cell suspensions. The applicability of this protocol was tested by applying it to a study of the deposition of Fe(III) oxide particles onto glass surfaces with and without biofilms of the bacterium Burkholdaria cepacia 17616. Experimental results using a laboratory biofilm reactor indicated that the deposition rate of Fe(III) colloids was not significantly affected by the presence of B. cepacia biofilms or by the presence of previously deposited Fe. However, deposition of Fe to reactor surfaces other than the glass surfaces may have interfered with the analyses, and atomic absorption measurements showed a slight increase in Fe deposition onto glass surfaces with biofilms present. Fe deposition to the composite of all reactor surfaces increased with increasing colloidal particle size, indicating a dominance of interception and/or sedimentation in controlling Fe deposition on surfaces in the biofilm reactor.
caused more rapid degradation ofthesheath thancyanide or sulfite. Treatment ofthesheath with1N Na... more caused more rapid degradation ofthesheath thancyanide or sulfite. Treatment ofthesheath with1N NaOH resulted inrapid breakdown, while treatment with1N HCIresulted inslowbut significant hydrolysis. Transmission electron microscopy showedthatthe6.5-nm fibrils previously showntobe an integral structural element ofthesheath fabric (D.EmersonandW.C.Ghiorse, J.Bacteriol. 175:7808-7818, 1993) were progressively dissociated intorandommasses duringDTT-induced degradation. Quantitation of disulfide bondswithDTT showedthatthesheaths contained approximately 2.2,umol ofdisulfides per mg of sheathprotein. Reaction with5,5'-dithio-bis-(2-nitrobenzoic acid)showedthatsheaths alsocontained approximately 0.8,umoloffreesulfhydryls per mg ofprotein. A sulfhydryl-specific fluorescent probe (fluorescein 5-maleimide) showedthatthefreesulfhydryls insheathed cell filaments were evenly distributed throughout thesheath. Sodiumdodecyl sulfate-polyacrylamide gelelectrophoresis autoradiography of ('4C)i...
Water Research, 2000
ÐSurface coatings (bio®lms and associated minerals) were collected on glass slides in the oxic su... more ÐSurface coatings (bio®lms and associated minerals) were collected on glass slides in the oxic surface waters of Cayuga Lake (New York State, U.S.A.) and were used to evaluate the relative contributions of Fe, Mn and Al oxides and organic material to total observed Pb and Cd adsorption by the surface coating materials. Several alternative selective extraction techniques were evaluated with respect to both selectivity and alteration of the residual unextracted material. Pb and Cd adsorption was measured under controlled laboratory conditions (mineral salts solution with de®ned metal speciation, ionic strength 0.05 M, 258C and pH 6.0) before and after extractions to determine by dierence the adsorptive properties of the extracted component(s). Hydroxylamine hydrochloride (0.01 M NH 2 OHÁHCl+0.01 M HNO 3) was used to selectively remove Mn oxides, sodium dithionite (0.3 M Na 2 S 2 O 4) was used to remove Mn and Fe oxides, and 10% oxalic acid was used to remove metal oxides and organic materials. Several other extractants were evaluated, but preliminary experiments indicated that they were not suitable for these experiments because of undesirable alterations of the residual, unextracted material. The selected extraction methods removed target components with eciencies between 71 and 83%, but signi®cant amounts of metal oxides and organic materials other than the target components were also removed by the extractants (up to 39%). Nonlinear regression analysis of the observed Pb and Cd adsorption based on the assumption of additive Langmuir adsorption isotherms was used to estimate the relative contributions of each surface coating constituent to total Pb and Cd binding of the bio®lms. Adsorption of Cd to the lake bio®lms was dominated by Fe oxides, with lesser roles attributed to adsorption by Mn and Al oxides and organic material. Adsorption of Pb was dominated by Mn oxides, with lesser roles indicated for adsorption to Fe oxides and organic material, and the estimated contribution of Al oxides to Pb adsorption was insigni®cant. The ®tted Pb adsorption isotherm for Fe oxides was in excellent agreement with those obtained through direct experiments and reported in independent investigations. The estimated Pb distribution between surface coating components also agreed well with that previously predicted by an additive adsorption model based on Pb adsorption isotherms for laboratory surrogates for Mn, Fe and Al oxides and de®ned biological components.
Water Research, 1995
A~tract-Biofilms influence the transport and fate of heavy metals in aquatic environments both di... more A~tract-Biofilms influence the transport and fate of heavy metals in aquatic environments both directly by adsorption and complexation reactions and indirectly via interactions with oxides of iron and manganese. These reactions were investigated by introducing lead into a continuous-flow biofilm reactor that was designed to simulate conditions in a flowing freshwater aquatic environment. The reactor provided controlled conditions, and use of a chemically-defined growth medium allowed calculation of lead speciation with a chemical equilibrium program (MINEQL). Pseudomonas cepacia was employed as a test cell strain because of its ability to grow and form biofilms in the defined medium. This bacterium affected lead distribution in the reactor by adsorbing lead both to adherent and suspended cells. When the aqueous bulk lead concentration was 1.4 0.1/~M and biofilm coverage (measured as chemical oxygen demand, COD) was 50 mequiv COD/m 2, lead adsorption was increased by about a factor of five relative to bare glass. Of the total lead in solution, only 1% was adsorbed to suspended cells (5 x 10 7 ceUs/ml). Lead adsorption to biofilms followed a Langmuir isotherm with a maximum adsorption (Fmax) of 56/~mol Pb/equiv COD and an adsorption equilibrium constant (K) of 0.64 liter/#mol Pb. Lead complexed with dissolved bacterial exopolymer was below detection limits. Pretreatment of glass slides with colloidal iron also significantly increased lead adsorption relative to bare glass. Lead adsorption to adsorbed iron fit a Langmuir isotherm with Fma x 50 #mol Pb/mol Fe, and K 1.3 liter/#mol Pb. Lead binding to glass coated with both cells and iron was additive, and could be predicted by summing adsorption predicted using isotherms for each constituent. The presence of iron surface coatings increased initial biofilm formation rates, but after reaching steady state conditions, biofilm coverage was similar for slides treated with iron and untreated slides. A concentration of 1 # M lead produced a transient reduction in suspended cell counts. Cell counts recovered to the original cell density over the course of five to ten reactor retention times. With iron present, the magnitude of the reduction in cell concentration in response to the addition of lead was greatly reduced, suggesting that toxic effects of lead may be reduced by iron.
Water Environment Research, 2003
When oocysts of the protozoan Cryptosporidium parvum contaminate drinking water supplies, they ca... more When oocysts of the protozoan Cryptosporidium parvum contaminate drinking water supplies, they can cause outbreaks of Cryptosporidiosis, a common waterborne disease. Of the different pathways by which oocysts can wind up in drinking water, one has received little attention to date; that is, because soils are often considered to be perfect filters, the transport of oocysts through the subsoil to groundwater is generally ignored. To evaluate the significance of this pathway, a series of laboratory experiments investigated subsurface transport of oocysts. Experiment 1 was carried out in a vertical 18-cm-long column filled either with glass beads or silica sand, under conditions known to foster fingered flow. Experiment 2 involved undisturbed, macroporous soil columns subjected to macropore flow. Experiment 3 aimed to study the lateral flow on an undisturbed soil block. The columns and soil samples were subjected to artificial rainfall and were allowed to reach steady state. At that point, feces of contaminated calves were applied at the surface along with a known amount of potassium chloride to serve as a tracer, and rainfall was continued at the same rate. The breakthrough of oocysts and chloride, monitored in the effluent, demonstrate the importance of preferential flow on the transport of oocysts. Compared with chloride, peak oocyst concentrations were not appreciably delayed and, in some cases, occurred even before the chloride peak. Recovery rates for oocysts were low, ranging from 0.1 to 10.4% of the oocysts originally applied on the columns. However, the numbers of oocysts present in the effluents were still orders of magnitude higher than 10 oocysts, the infectious dose considered by the U.S. Food and Drug Administration, Center for Food Safety and Applied Nutrition, to be sufficient to cause Cryptosporidiosis in healthy adults. These results suggest that the transport of oocysts in the subsurface via preferential flow may create a significant risk of groundwater contamination in some situations. Water Environ. Res., 75, 113 (2003).
Vadose Zone Journal, 2004
in the form of 4-to 6-m-long ovoid-shaped oocysts, with a double wall that is resistant to most o... more in the form of 4-to 6-m-long ovoid-shaped oocysts, with a double wall that is resistant to most oxidation As a result of Cryptosporidium parvum in drinking water, several processes such as ozonation and chlorination (Current, outbreaks of cryptosporidiosis have occurred in the last 10 yr. Al-1986; Atwill et al., 1997). though it is generally believed that movement of pathogens through the soil is minimal, recent research has shown that appreciable num-During the past two decades, the presence of C. parbers of C. parvum oocysts may be transported via preferential or vum in surface-and groundwaters in the United States fingered flow to groundwater. The objective of the present research and Great Britain (Galbraith et al., 1987; Rose et al., was to further investigate and model the transport of oocysts through 1991; Craun et al., 1998) has been associated with several preferential flow paths in the vadose zone under a "worst-case" scemajor outbreaks of cryptosporidiosis (Hayes et al., 1989; nario. This was studied by adding calves feces containing C. parvum MacKenzie et al., 1994). Among the different pathways oocysts with a Cl Ϫ tracer to undisturbed silt loam columns and disfor the transport of oocysts to drinking water sources, turbed sand columns during a simulated steady-state rain. The sand columns exhibited preferential flow in the form of fingers whereas downward percolation is usually considered to be insigmacropore flow occurred in the undisturbed cores. In the columns nificant, because soils are generally assumed to be an with fingered flow, oocysts and Cl were transported rapidly with the effective filter for a wide range of pathogens. Studies same velocity through the columns. Although only 14 to 86% of the of packed columns with saturated flow by Brush et al. amount applied, the number of oocysts transported across the columns (1999) and Harter et al. (2000) and undisturbed columns was several orders of magnitude above an infective dose. The macwith unsaturated flow (Mawdsley et al., 1996), however, ropore columns had only a very limited breakthrough of oocysts, showed that C. parvum oocysts could be transported which appeared several pore volumes after the Cl broke through initially. A simulation model for the transport of oocysts via preferen-rapidly downward through the soil. Although transport tial flow was developed on the basis of an existing preferential flow of C. parvum oocysts in saturated flow has been studied model for nonadsorbing solutes, with addition of a first-order sink experimentally and described mathematically (Brush et term for adsorbance of the C. parvum to the air-water-solid (AWS) al., 1999; Harter et al., 2000), detailed observations of interfaces, and with velocity and dispersivity parameters derived from the transport and persistence of C. parvum oocysts in Cl Ϫ transport. The breakthrough of C. parvum oocysts could be deunsaturated soils with preferential flow are still lacking, scribed realistically for the sand columns. However, the model could particularly in the presence of preferential flow pronot describe oocyst transport in the columns with macropores. Three sets of experiments were performed in which C. parvum oocysts contained in the feces of infected calves were
Limnology and Oceanography, 1999
The role of the composition of surface-coating materials in controlling trace metal adsorption in... more The role of the composition of surface-coating materials in controlling trace metal adsorption in aquatic environments was investigated using natural biofilms that developed on glass slides in three New York State lakes and a water-supply well. Adsorption isotherms were obtained for Pb binding to each of the biofilms in solutions with defined Pb speciation at 25ЊC and pH 6.0, with Pb concentrations ranging from 0.2 to 2.0 M. Adsorption isotherms for Pb binding to laboratory-derived metal oxides and surrogate organic materials were determined under the same conditions. These isotherms, combined with characterization of natural biofilm composition, were used to estimate the relative contributions of the organic and metal oxide surface-coating constituents by assuming additivity of adsorption to discrete adsorbing phases. Cells of a diatom (Navicula peliculosa), a green alga (Chlorella vulgaris), the bacterium Leptothrix discophora, and extracellular polymer of the bacterium Burkholdaria cepacia were tested as laboratory analogs for the organic phase of the biofilms. Amorphous Fe oxyhydroxide, ␥Al 2 O 3 , and a laboratoryderived biogenic Mn oxyhydroxide were used as laboratory surrogates for biofilm minerals. The sum total of predicted Pb binding to the defined surrogates accounted for at least 90% of the total observed Pb binding in the three lake biofilms and 60% of that observed for the well biofilms. For the lake biofilms, Pb adsorption to Fe and Mn oxides was significantly greater than that estimated for organic materials. The use of biogenic Mn oxide as a model component resulted in an estimated Pb adsorption to Mn oxyhydroxides in the lake biofilms up to four times greater than that estimated for Fe oxyhydroxide. Estimated Pb binding by Al oxide was negligible for all four biofilms. These results suggest that Fe and biogenic Mn oxides exert the greatest influence on Pb adsorption in oxic freshwater environments at pH 6.0. This research was supported by NSF grants BES-97067715 and CHE-9708093 and through a STAR fellowship from the U.S. EPA (Y.N.).
Journal of Parasitology, 2001
Cryptosporidium parvum oocysts were examined to ascertain excystation requirements and the effect... more Cryptosporidium parvum oocysts were examined to ascertain excystation requirements and the effects of gamma irradiation. Oocysts and excysted sporozoites were examined for dye permeability and infectivity. Maximum excystation occurred when oocysts were pretreated with acid and incubated with bile salts, and potassium or sodium bicarbonate. Pretreatment with Hanks' balanced salt solution or NaCl lowered excystation; however, this effect was overcome with acid. Sodium ions were replaceable with potassium ions, and sodium bicarbonate was replaceable with sodium phosphate. Oocysts that received 200 krad irradiation excysted at the same rates as nonirradiated oocysts (95%), the excystation rates were lowered (50%) by 2,000 krad, and no excystation was observed by 5,000 krad. No differences were observed between the propidium iodide (PI) permeability of untreated oocysts and oocysts treated with 200 krad, while 92% of oocysts were PI positive after 2,000 krad. Most of the sporozoites exposed to 2,000 krad were not viable as indicated by the dye permeability assay. The oocysts irradiated with 200 and 2,000 krad infected cells, but no replication was observed. The results suggest that gamma-irradiated oocysts may still be capable of excystation and apparent infection; however, because the sporozoites could not reproduce they must not have been viable.