Biomedical subjects

Tullis C Onstott

Publications and source records attributed to Tullis C Onstott.

5 recordsLinked to original sources

PubMed2004

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.

Biodegradation, Environmental↗

PubMed2003

Related assemblages of sulphate-reducing bacteria associated with ultradeep gold mines of South Africa and deep basalt aquifers of Washington State.

We characterized the diversity of sulphate-reducing bacteria (SRB) associated with South African gold mine boreholes and deep aquifer systems in Washington State, USA. Sterile cartridges filled with crushed country rock were installed on two hydrologically isolated and chemically distinct sites at depths of 3.2 and 2.7 km below the land surface (kmbls) to allow development of biofilms. Enrichments of sulphate-reducing chemolithotrophic (H2) and organotrophic (lactate) bacteria were established from each site under both meso- and thermophilic conditions. Dissimilatory sulphite reductase (Dsr) and 16S ribosomal RNA (rRNA) genes amplified from DNA extracted from the cartridges were most closely related to the Gram-positive species Desulfotomaculum thermosapovorans and Desulfotomaculum geothermicum, or affiliated with a novel deeply branching clade. The dsr sequences recovered from the Washington State deep aquifer systems affiliated closely with the South African sequences, suggesting that Gram-positive sulphate-reducing bacteria are widely distributed in the deep subsurface.

Biofilms↗

PubMed2003

Isolation of Halobacterium salinarum retrieved directly from halite brine inclusions.

Halite crystals were selected from a 186 m subsurface core taken from the Badwater salt pan, Death Valley, California to ascertain if halophilic Archaea and their associated 16S rDNA can survive over several tens of thousands of years. Using a combined microscope microdrill/micropipette system, fluids from brine inclusions were aseptically extracted from primary, hopper texture, halite crystals from 8 and 85 metres below the surface (mbls). U-Th disequilibrium dating indicates that these halite layers were deposited at 9,600 and 97,000 years before present (ybp) respectively. Extracted inclusions were used for polymerase chain reaction (PCR) analysis with haloarchaea-specific 16S rDNA primers or placed into haloarchaea culture medium. Enrichment cultures were obtained from 97 kyr halite crystal inclusion fluid and haloarchaea-containing prepared crystals (positive controls), whereas inclusions from crystals of 9.6 kyr halite and the haloarchaea-free halite crystals (negative controls) resulted in no growth. Phylogenetic analysis (16S rDNA) of the 97 kyr isolate, designated BBH 001, revealed a homology of 100% with Halobacterium salinarum. DNA-DNA hybridization experiments confirmed that BBH 001 was closely related to H. salinarum (81-75% hybridization) and its ascription to this haloarchaea species. The described method of retrieving particle-containing brine from fluid inclusions offers a robust approach for assessing the antiquity of microorganisms associated with evaporites.

Base Composition↗

PubMed2002

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.

Biological Transport↗

PubMedDate not supplied

A modular injection system, multilevel sampler, and manifold for tracer tests.

Ground water injection and sampling systems were developed for bacterial transport experiments in both homogenous and heterogeneous unconsolidated, surficial aquifers. Two types of injection systems, a large single tank and a dynamic mixing tank, were designed to deliver more than 800 L of amended ground water to the aquifer over 12 hours, without altering the ground water temperature, pH, Eh, or dissolved gas composition. Two types of multilevel samplers (MLSs) were designed and installed. Permanent MLSs performed well for the homogenous surficial aquifer, but their installation procedure promoted vertical mixing, which could obfuscate experimental data obtained from vertically stratified, heterogeneous aquifers. A novel, removable MLS was designed to fit in 2- and 4-inch wells. Expandable O-rings between each sampling port hydraulically isolated each port for sample collection when a nut was tightened at the land surface. A low-cost vacuum manifold system designed to work with both MLS designs used 50 mL centrifuge tubes to efficiently sample 12 MLS ports with one peristaltic pump head. The integrated system was developed and used during four field campaigns over a period of three years. During each campaign, more than 3000 ground water samples were collected in less than one week. This system should prove particularly useful for ground water tracer, injection, and push-pull experiments that require high-frequency and/or high-density sampling.

Bacteria↗