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David A Stahl

Publications and source records attributed to David A Stahl.

At least 19 recordsLinked to original sources
PubMed2024

Ammonia-oxidizing bacteria and archaea exhibit differential nitrogen source preferences.

Ammonia-oxidizing microorganisms (AOM) contribute to one of the largest nitrogen fluxes in the global nitrogen budget. Four distinct lineages of AOM: ammonia-oxidizing archaea (AOA), beta- and gamma-proteobacterial ammonia-oxidizing bacteria (β-AOB and γ-AOB) and complete ammonia oxidizers (comammox), are thought to compete for ammonia as their primary nitrogen substrate. In addition, many AOM species can utilize urea as an alternative energy and nitrogen source through hydrolysis to ammonia. How the coordination of ammonia and urea metabolism in AOM influences their ecology remains poorly understood. Here we use stable isotope tracing, kinetics and transcriptomics experiments to show that representatives of the AOM lineages employ distinct regulatory strategies for ammonia or urea utilization, thereby minimizing direct substrate competition. The tested AOA and comammox species preferentially used ammonia over urea, while β-AOB favoured urea utilization, repressed ammonia transport in the presence of urea and showed higher affinity for urea than for ammonia. Characterized γ-AOB co-utilized both substrates. These results reveal contrasting niche adaptation and coexistence patterns among the major AOM lineages.

Archaea↗
PubMed2024

Environmental stress mediates groundwater microbial community assembly.

Community assembly describes how different ecological processes shape microbial community composition and structure. How environmental factors impact community assembly remains elusive. Here we sampled microbial communities and >200 biogeochemical variables in groundwater at the Oak Ridge Field Research Center, a former nuclear waste disposal site, and developed a theoretical framework to conceptualize the relationships between community assembly processes and environmental stresses. We found that stochastic assembly processes were critical (>60% on average) in shaping community structure, but their relative importance decreased as stress increased. Dispersal limitation and 'drift' related to random birth and death had negative correlations with stresses, whereas the selection processes leading to dissimilar communities increased with stresses, primarily related to pH, cobalt and molybdenum. Assembly mechanisms also varied greatly among different phylogenetic groups. Our findings highlight the importance of microbial dispersal limitation and environmental heterogeneity in ecosystem restoration and management.

Phylogeny↗
PubMed2006

Discrimination of shifts in a soil microbial community associated with TNT-contamination using a functional ANOVA of 16S rRNA hybridized to oligonucleotide microarrays.

A functional ANOVA analysis of the thermal dissociation of RNA hybridized to DNA microarrays was used to improve discrimination between two soil microbial communities. Following hybridization of in vitro transcribed 16S rRNA derived from uncontaminated and 2,4,6-trinitrotoluene contaminated soils to an oligonucleotide microarray containing group- and species-specific perfect match (PM) probes and mismatch (MM) variants, thermal dissociation was used to analyze the nucleic acid bound to each PM-MM probe set. Functional ANOVA of the dissociation curves generally discriminated PM-MM probe sets when Td values (temperature at 50% probe-target dissociation) could not. Maximum discrimination for many PM and MM probes often occurred at temperatures greaterthan the Td. Comparison of signal intensities measured prior to dissociation analysis from hybridizations of the two soil samples revealed significant differences in domain-, group-, and species-specific probes. Functional ANOVA showed significantly different dissociation curves for 11 PM probes when hybridizations from the two soil samples were compared, even though initial signal intensities for 3 of the 11 did not vary.

Analysis of Variance↗
PubMed2006

Recirculating flow accelerates DNA microarray hybridization in a microfluidic device.

A recirculating microfluidic device fabricated by laminating Mylar and glass was developed for the analysis of hybridization of oligonucleotides to DNA microarrays. The device is part of a system that provides controlled hybridization to DNA probes immobilized in a microarray of polyacrylamide gel pads using recirculation and temperature control. The system was used to obtain real-time kinetics of DNA hybridization and more accurate melting profiles of target-probe duplexes than possible using a static hybridization format. Recirculation shortened the time of perfect match target-probe hybridization from 6 hours to 2 hours and shifted the Td by 1.54 degrees C, relative to static conditions. The experimental results were consistent with a three-dimensional simulation of hybridization using a recirculating buffer system.

Buffers↗
PubMed2006

Technology development to explore the relationship between oral health and the oral microbial community.

The human oral cavity contains a complex microbial community that, until recently, has not been well characterized. Studies using molecular tools have begun to enumerate and quantify the species residing in various niches of the oral cavity; yet, virtually every study has revealed additional new species, and little is known about the structural dynamics of the oral microbial community or how it changes with disease. Current estimates of bacterial diversity in the oral cavity range up to 700 species, although in any single individual this number is much lower. Oral microbes are responsible for common chronic diseases and are suggested to be sentinels of systemic human diseases. Microarrays are now being used to study oral microbiota in a systematic and robust manner. Although this technology is still relatively young, improvements have been made in all aspects of the technology, including advances that provide better discrimination between perfect-match hybridizations from non-specific (and closely-related) hybridizations. This review addresses a core technology using gel-based microarrays and the initial integration of this technology into a single device needed for system-wide studies of complex microbial community structure and for the development of oral diagnostic devices.

Journal Article↗
PubMed2006

Using the stress response to monitor process control: pathways to more effective bioremediation.

Environmental contamination with a variety of pollutants has prompted the development of effective bioremediation strategies. But how can these processes be best monitored and controlled? One avenue under investigation is the development of stress response systems as tools for effective and general process control. Although the microbial stress response has been the subject of intensive laboratory investigation, the environmental reflection of the laboratory response to specific stresses has been little explored. However, it is only within an environmental context, in which microorganisms are constantly exposed to multiple changing environmental stresses, that there will be full understanding of microbial adaptive resiliency. Knowledge of the stress response in the environment will facilitate the control of bioremediation and other processes mediated by complex microbial communities.

Adaptation, Physiological↗
PubMed2006

Impact of prehybridization PCR amplification on microarray detection of nitrifying bacteria in wastewater treatment plant samples.

A gel-based microarray that included a set of 26 oligonucleotide probes targeting all nitrifying bacteria at varying levels of specificity suggested the presence of targeted microorganisms when hybridized to RNA isolated from a wastewater treatment plant, but could not discriminate between perfectly matched and mismatched sequences due in part to low signal intensity. To enhance sensitivity and improve discrimination, polymerase chain reaction was used to selectively amplify the 16S rRNA genes of specific nitrifier groups. RNA transcribed from these DNA templates was hybridized to the microarray and thermal dissociation analysis was used to characterize the specificity of hybridization. Amplification with Nitrospira-specific primers resulted in the selective amplification of this target group, confirmed by both a significant increase in signal intensity and a melting profile identical to the reference RNA. In contrast, Nitrobacter was not detected in the environmental samples with probe Nbac1000 despite pre-amplification with Nitrobacter-specific primers, indicating the absence of strains containing this Nitrobacter-specific sequence. Pre-amplification using primers specific for beta-Proteobacterial ammonia-oxidizing bacteria resulted in a significant increase in signal intensity for probe Nso190, but melting profiles for probe Nso190 showed a slight deviation between amplified RNA and the reference microorganism, suggesting that the amplification products contained some sequences that varied by a single nucleotide difference in the probe target region.

Bacteria↗
PubMed2006

Recovery of temperate Desulfovibrio vulgaris bacteriophage using a novel host strain.

A novel sulfate-reducing bacterium (strain DePue) closely related to Desulfovibrio vulgaris ssp. vulgaris strain Hildenborough was isolated from the sediment of a heavy-metal impacted lake using established techniques. Although few physiological differences between strains DePue and Hildenborough were observed, pulse-field gel electrophoresis (PFGE) revealed a significant genome reduction in strain DePue. Comparative whole-genome microarray and polymerase chain reaction analyses demonstrated that the absence of genes annotated in the Hildenborough genome as phage or phage-related contributed to the significant genome reduction in strain DePue. Two morphotypically distinct temperate bacteriophage from strain Hildenborough were recovered using strain DePue as a host for plaque isolation.

Bacteriophages↗
PubMed2006

Effects of endogenous substrates on adaptation of anaerobic microbial communities to 3-chlorobenzoate.

Lengthy adaptation periods in laboratory studies evaluating the potential for contaminant biodegradation in natural or engineered environments may indicate that the native microbial communities are not metabolizing the contaminants in situ. In this study, we characterized the adaptation period preceding the biodegradation of 3-chlorobenzoate in anaerobic communities derived from lake sediment and wastewater sludge digesters. The importance of alternative mechanisms of adaptation of the anaerobic communities to 3-chlorobenzoate was evaluated by monitoring the concentrations of metabolic substrates and products as well as the levels of total small subunit (SSU) rRNA and SSU rRNA from populations thought to be important in 3-chlorobenzoate mineralization. The anaerobic environments from which the 3-chlorobenzoate-degrading communities were derived contained different levels of endogenous substrates. Increasing methane levels in the digester and sediment communities and decreasing chemical oxygen demand concentrations in the sediment community during the adaptation periods revealed that endogenous substrates were preferentially utilized relative to 3-chlorobenzoate. Methane and chemical oxygen demand concentrations leveled off concomitantly with the onset of 3-chlorobenzoate biodegradation, suggesting that depletion of the preferentially degraded endogenous substrates stimulated 3-chlorobenzoate metabolism. Consistent with these observations, adaptation to 3-chlorobenzoate occurred more rapidly in digester samples that were depleted of endogenous substrates compared to samples that contained high levels of these biodegradable compounds. Other potential adaptation mechanisms, e.g., genetic change or selective population enrichment, appeared to be less important based on the reproducibility and relative lengths of the adaptation events, trends in the SSU rRNA levels, and/or amplification of SSU rRNA genes from key populations.

Adaptation, Physiological↗
PubMed2006

Recurring seasonal dynamics of microbial communities in stream habitats.

Recurring seasonal patterns of microbial distribution and abundance in three third-order temperate streams within the southeast Pennsylvania Piedmont were observed over 4 years. Populations associated with streambed sediments and rocks (epilithon) were identified using terminal restriction length polymorphism (tRFLP) and sequencing of 16S rRNA genes selectively amplified with primers for the bacterial domain. Analyses of the relative magnitudes of tRFLP peak areas by using nonmetric multidimensional scaling resolved clear seasonal trends in epilithic and sediment populations. Oscillations between two dominant groups of epilithic genotypes, explaining 86% of the seasonal variation in the data set, were correlated with temperature and dissolved organic carbon. Sequences affiliated with epilithic phototrophs (cyanobacteria and diatom chloroplasts), a Rhodoferax sp., and a Bacillus species clustered in the summer, whereas sequences most closely related to "Betaproteobacteria" (putative Burkholderia sp.), and a putative cyanobacterium clustered in the fall/spring. The sediment genotypes also clustered into two groups, and these explained 85% of seasonal variation but correlated only with temperature. A summer tRFLP pattern was characterized by prevalence of "Betaproteobacteria," "Gammaproteobacteria," and a Bacillus sp., whereas the winter/spring pattern was characterized by phylotypes most closely related to "Firmicutes," "Gammaproteobacteria," and "Nitrospirae." A close association between these headwater streams and their watersheds was suggested by the recovery of sequences related to microbial populations provisionally attributed to not only freshwaters but also terrestrial habitats.

Bacillus↗
PubMed2006

Transmission of nephridial bacteria of the earthworm Eisenia fetida.

The lumbricid earthworms (annelid family Lumbricidae) harbor gram-negative bacteria in their excretory organs, the nephridia. Comparative 16S rRNA gene sequencing of bacteria associated with the nephridia of several earthworm species has shown that each species of worm harbors a distinct bacterial species and that the bacteria from different species form a monophyletic cluster within the genus Acidovorax, suggesting that there is a specific association resulting from radiation from a common bacterial ancestor. Previous microscopy and culture studies revealed the presence of bacteria within the egg capsules and on the surface of embryos but did not demonstrate that the bacteria within the egg capsule were the same bacteria that colonized the nephridia. We present evidence, based on curing experiments, in situ hybridizations with Acidovorax-specific probes, and 16S rRNA gene sequence analysis, that the egg capsules contain high numbers of the bacterial symbiont and that juveniles are colonized during development within the egg capsule. Studies exposing aposymbiotic hatchlings to colonized adults and their bedding material suggested that juvenile earthworms do not readily acquire bacteria from the soil after hatching but must be colonized during development by bacteria deposited in the egg capsule. Whether this is due to the developmental stage of the host or the physiological state of the symbiont remains to be investigated.

Animals↗
PubMed2005

Isolation of an autotrophic ammonia-oxidizing marine archaeon.

For years, microbiologists characterized the Archaea as obligate extremophiles that thrive in environments too harsh for other organisms. The limited physiological diversity among cultivated Archaea suggested that these organisms were metabolically constrained to a few environmental niches. For instance, all Crenarchaeota that are currently cultivated are sulphur-metabolizing thermophiles. However, landmark studies using cultivation-independent methods uncovered vast numbers of Crenarchaeota in cold oxic ocean waters. Subsequent molecular surveys demonstrated the ubiquity of these low-temperature Crenarchaeota in aquatic and terrestrial environments. The numerical dominance of marine Crenarchaeota--estimated at 10(28) cells in the world's oceans--suggests that they have a major role in global biogeochemical cycles. Indeed, isotopic analyses of marine crenarchaeal lipids suggest that these planktonic Archaea fix inorganic carbon. Here we report the isolation of a marine crenarchaeote that grows chemolithoautotrophically by aerobically oxidizing ammonia to nitrite--the first observation of nitrification in the Archaea. The autotrophic metabolism of this isolate, and its close phylogenetic relationship to environmental marine crenarchaeal sequences, suggests that nitrifying marine Crenarchaeota may be important to global carbon and nitrogen cycles.

Aerobiosis↗
PubMed2005

DNA microarray detection of nitrifying bacterial 16S rRNA in wastewater treatment plant samples.

A small scale DNA microarray containing a set of oligonucleotide probes targeting the 16S rRNAs of several groups of nitrifying bacteria was developed for the monitoring of wastewater treatment plant samples. The microarray was tested using reference rRNAs from pure cultures of nitrifying bacteria. Characterization of samples collected from an industrial wastewater treatment facility demonstrated that nitrifying bacteria could be detected directly by microarray hybridization without the need for PCR amplification. Specifically, the microarray detected Nitrosomonas spp. but did not detect Nitrobacter. The specificity and sensitivity of direct detection was evaluated using on-chip dissociation analysis, and by two independent analyses--an established membrane hybridization format and terminal restriction fragment length polymorphism fingerprinting (T-RFLP). The latter two analyses also revealed Nitrospira and Nitrobacter to be contributing populations in the treatment plant samples. The application of DNA microarrays to wastewater treatment systems, which has been demonstrated in the current work, should offer improved monitoring capabilities and process control for treatment systems, which are susceptible to periodic failures.

DNA Fingerprinting↗
PubMed2005

Functional marker genes for identification of sulfate-reducing prokaryotes.

Sulfate-reducing prokaryotes (SRPs) exploit sulfate as an electron acceptor for anaerobic respiration and exclusively catalyze this essential step of the world's sulfur cycle. Because SRPs are found in many prokaryotic phyla and are often closely related to non-SRPs, 16S rRNA gene-based analyses are inadequate to identify novel lineages of this guild in a cultivation-independent manner. This problem can be solved by comparative sequence analysis of environmentally retrieved gene fragments of the dissimilatory (bi)sulfite (dsrAB) and adenosine-5'-phosphosulfate reductases (apsA), which encode key enzymes of the SRP energy metabolism. This chapter provides detailed protocols for the application of these functional marker molecules for SRP diversity surveys in the environment. Data from the analysis of dsrAB sequence diversity in water samples from the Mariager Fjord in northeast Denmark are presented to illustrate the different steps of the protocols. Furthermore, this chapter describes a novel gel retardation-based technique, suitable for fingerprinting of the approximately 1.9-kb-large dsrAB polymerase chain reaction amplification products, which efficiently increases the chance of retrieving rare and novel dsrAB sequence types from environmental samples.

Bacteria, Anaerobic↗
PubMed2005

Hydrothermal vent gastropods from the same family (Provannidae) harbour epsilon- and gamma-proteobacterial endosymbionts.

The discovery of new hydrothermal vent systems in the back-arc basins of the Western Pacific revealed chemosynthesis-based faunal communities distinct from those of other vents. These vents are dominated by two related gastropods (Alviniconcha spp. and Ifremeria nautilei) that harbour symbiotic bacteria in their gills. We used comparative 16S ribosomal RNA (rRNA) gene sequencing and in situ hybridization with rRNA-targeted probes to characterize the bacterial symbionts of Alviniconcha sp. and I. nautilei from the Manus Basin in the Western Pacific. The analyses revealed that these two gastropod species, although affiliated with the same family, harbour phylogenetically distant chemosymbionts, suggesting independent origins of these endosymbioses. The I. nautilei endosymbiont clusters with sulfur-oxidizing bacteria within the gamma-Proteobacteria, as is the case for all previously characterized endosymbionts from a wide diversity of host taxa harbouring thioautotrophic prokaryotes. In contrast, the Alviniconcha endosymbiont is affiliated with sulfur-oxidizing bacteria within the epsilon-Proteobacteria. These results show that bacteria from the epsilon-Proteobacteria are also capable of forming endosymbiotic associations with marine invertebrates from chemosynthetic environments. More generally, the endosymbiotic lifestyle is now shown to be distributed throughout all recognized classes of the Proteobacteria.

Animals↗
PubMed2005

Loss of diversity of ammonia-oxidizing bacteria correlates with increasing salinity in an estuary system.

Ammonia-oxidizing bacteria (AOB) play an important role in nitrogen cycling in estuaries, but little is known about AOB diversity, distribution and activity in relation to the chemical and physical changes encountered in estuary systems. Although estuarine salinity gradients are well recognized to influence microbial community structure, few studies have examined the influence of varying salinity on the diversity and stability of AOB populations. To investigate these relationships, we collected sediment samples from low-, mid- and high-salinity sites in Plum Island Sound estuary, MA, during spring and late summer over 3 years. Ammonia-oxidizing bacteria distribution and diversity were assessed by terminal restriction fragment length polymorphism (TRFLP) analysis of the ammonia monooxygenase (amoA) gene, and fragments were identified by screening amoA clone libraries constructed from each site. Most striking was the stability and low diversity of the AOB community at the high-salinity site, showing little variability over 3 years. Ammonia-oxidizing bacteria at the high-salinity site were not closely related to any cultured AOB, but were most similar to Nitrosospira spp. Ammonia-oxidizing bacteria at the mid- and low-salinity sites were distributed among Nitrosospira-like sequences and sequences related to Nitrosomonas ureae/oligotropha and Nitrosomonas sp. Nm143. Our study suggests that salinity is a strong environmental control on AOB diversity and distribution in this estuary.

Ammonia↗
PubMed2005

Lateral gene transfer of dissimilatory (bi)sulfite reductase revisited.

In contrast to previous findings, we demonstrate that the dissimilatory (bi)sulfite reductase genes (dsrAB) of Desulfobacula toluolica were vertically inherited. Furthermore, Desulfobacterium anilini and strain mXyS1 were identified, by dsrAB sequencing of 17 reference strains, as members of the donor lineage for those gram-positive Desulfotomaculum species which laterally acquired dsrAB.

Bacteria, Anaerobic↗