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Biomedical subjects

David M Ward

Publications and source records attributed to David M Ward.

At least 19 recordsLinked to original sources
PubMed2007

Impact of carbon metabolism on 13C signatures of cyanobacteria and green non-sulfur-like bacteria inhabiting a microbial mat from an alkaline siliceous hot spring in Yellowstone National Park (USA).

Alkaline siliceous hot spring microbial mats in Yellowstone National Park are composed of two dominant phototropic groups, cyanobacteria and green non-sulfur-like bacteria (GNSLB). While cyanobacteria are thought to cross-feed low-molecular-weight organic compounds to support photoheterotrophic metabolism in GNSLB, it is unclear how this could lead to the heavier stable carbon isotopic signatures in GNSLB lipids compared with cyanobacterial lipids found in previous studies. The two groups of phototrophs were separated using percoll density gradient centrifugation and subsequent lipid and stable carbon isotopic analysis revealed that we obtained fractions with a approximately 60-fold enrichment in cyanobacterial and an approximately twofold enrichment in GNSLB biomass, respectively, compared with the mat itself. This technique was used to study the diel cycling and 13C content of the glucose pools in and the uptake of 13C-bicarbonate by the cyanobacteria and GNSLB, as well as the transfer of incorporated 13C from cyanobacteria to GNSLB. The results show that cyanobacteria have the highest bicarbonate uptake rates and accumulate glucose during the afternoon in full light conditions. In contrast, GNSLB have relatively higher bicarbonate uptake rates compared with cyanobacteria in the morning at low light levels. During the night GNSLB take up carbon that is likely derived through fermentation of cyanobacterial glucose enriched in 13C. The assimilation of 13C-enriched cyanobacterial carbon may thus lead to enriched 13C-contents of GNSLB cell components.

Bicarbonates↗
PubMed2006

Cyanobacterial ecotypes in the microbial mat community of Mushroom Spring (Yellowstone National Park, Wyoming) as species-like units linking microbial community composition, structure and function.

We have investigated microbial mats of alkaline siliceous hot springs in Yellowstone National Park as natural model communities to learn how microbial populations group into species-like fundamental units. Here, we bring together empirical patterns of the distribution of molecular variation in predominant mat cyanobacterial populations, theory-based modelling of how to demarcate phylogenetic clusters that correspond to ecological species and the dynamic patterns of the physical and chemical microenvironments these populations inhabit and towards which they have evolved adaptations. We show that putative ecotypes predicted by the theory-based model correspond well with distribution patterns, suggesting populations with distinct ecologies, as expected of ecological species. Further, we show that increased molecular resolution enhances our ability to detect ecotypes in this way, though yet higher molecular resolution is probably needed to detect all ecotypes in this microbial community.

Adaptation, Biological↗
PubMed2006

Microbial diversity in natural environments: focusing on fundamental questions.

Interactions with Gijs Kuenen and other Dutch scientists have led my lab to fundamental insights into the composition, structure and function of a hot spring cyanobacterial mat community that should influence our thinking about all microbial communities. By focusing on the distribution of molecular sequence variants of predominant mat phototrophs, we have discovered that small-scale sequence variation can be ecologically meaningful. By applying novel cultivation approaches, we have been able to obtain genetically relevant community members and thus to test the hypothesis that closely related sequence variants arose via adaptive evolutionary radiation. By applying the analytical tools of organic geochemistry we have gained insight into the metabolisms of major phototrophic members of the mat community as well as interactions between phototrophic guilds. These observations challenge traditional paradigms about prokaryotic species and cause us to consider evolutionary ecology theory as we develop genome-based methods for high-resolution analysis of the species-like fundamental units comprising microbial communities, and for investigating how such units coordinate the physiological activities within guilds of the community.

Autotrophic Processes↗
PubMed2006

In situ analysis of nitrogen fixation and metabolic switching in unicellular thermophilic cyanobacteria inhabiting hot spring microbial mats.

Genome sequences of two Synechococcus ecotypes inhabiting the Octopus Spring microbial mat in Yellowstone National Park revealed the presence of all genes required for nitrogenase biosynthesis. We demonstrate that nif genes of the Synechococcus ecotypes are expressed in situ in a region of the mat that varies in temperature from 53.5 degrees C to 63.4 degrees C (average 60 degrees C); transcripts are only detected at the end of the day when the mat becomes anoxic. Nitrogenase activity in mat samples was also detected in the evening. Hitherto, N2 fixation in hot spring mats was attributed either to filamentous cyanobacteria (not present at >50 degrees C in these mats) or to heterotrophic bacteria. To explore how energy-generating processes of the Synechococcus ecotypes track natural light and O2 conditions, we evaluated accumulation of transcripts encoding proteins involved in photosynthesis, respiration, and fermentation. Transcripts from photosynthesis (cpcF, cpcE, psaB, and psbB) and respiration (coxA and cydA) genes declined in the evening. In contrast, transcripts encoding enzymes that may participate in fermentation fell into two categories; some (ldh, pdhB, ald, and ackA) decreased in the evening, whereas others (pflB, pflA, adhE, and acs) increased at the end of the day and remained high into the night. Energy required for N2 fixation during the night may be derived from fermentation pathways that become prominent as the mat becomes anoxic. In a broader context, our data suggest that there are critical regulatory switches in situ that are linked to the diel cycle and that these switches alter many metabolic processes within the microbial mat.

Energy Metabolism↗
PubMed2006

Microbial population dynamics associated with crude-oil biodegradation in diverse soils.

Soil bacterial population dynamics were examined in several crude-oil-contaminated soils to identify those organisms associated with alkane degradation and to assess patterns in microbial response across disparate soils. Seven soil types obtained from six geographically distinct areas of the United States (Arizona, Oregon, Indiana, Virginia, Oklahoma, and Montana) were used in controlled contamination experiments containing 2% (wt/wt) crude oil spiked with [1-(14)C]hexadecane. Microbial populations present during hydrocarbon degradation were analyzed using both 16S rRNA gene sequence analysis and by traditional methods for cultivating hydrocarbon-oxidizing bacteria. After a 50-day incubation, all seven soils showed comparable hydrocarbon depletion, where >80% of added crude oil was depleted and approximately 40 to 70% of added [(14)C]hexadecane was converted to (14)CO(2). However, the initial rates of hydrocarbon depletion differed up to 10-fold, and preferential utilization of shorter-chain-length n-alkanes relative to longer-chain-length n-alkanes was observed in some soils. Distinct microbial populations developed, concomitant with crude-oil depletion. Phylogenetically diverse bacterial populations were selected across different soils, many of which were identical to hydrocarbon-degrading isolates obtained from the same systems (e.g., Nocardioides albus, Collimonas sp., and Rhodococcus coprophilus). In several cases, soil type was shown to be an important determinant, defining specific microorganisms responding to hydrocarbon contamination. However, similar Rhodococcus erythropolis-like populations were observed in four of the seven soils and were the most common hydrocarbon-degrading organisms identified via cultivation.

Alkanes↗
PubMed2006

Effect of temperature and light on growth of and photosynthesis by Synechococcus isolates typical of those predominating in the octopus spring microbial mat community of Yellowstone National Park.

Previous molecular analysis of the Octopus Spring cyanobacterial mat revealed numerous genetically distinct 16S rRNA sequences from predominant Synechococcus populations distantly related to the readily cultivated unicellular cyanobacterium Synechococcus lividus. Patterns in genotype distribution relative to temperature and light conditions suggested that the organisms contributing these 16S rRNA sequences may fill distinct ecological niches. To test this hypothesis, Synechococcus isolates were cultivated using a dilution and filtration approach and then shown to be genetically relevant to natural mat populations by comparisons of similarities of 16S rRNA genes and 16S-23S internal transcribed spacer (ITS) regions. Most isolates were identical or nearly identical at both loci to predominant mat genotypes; others showed 1- to 2-nucleotide differences at the 16S rRNA locus and even greater difference in ITS sequences. Isolates with predominant mat genotypes had distinct temperature ranges and optima for growth that were consistent with their distributions in the mat. Isolates with genotypes not previously detected or known to be predominant in the mat exhibited temperature ranges and optima that were not representative of predominant mat populations and also grew more slowly. Temperature effects on photosynthesis did not reflect temperature relations for growth. However, the isolate with the highest temperature optimum and upper limit was capable of performing photosynthesis at a higher temperature than other isolates. Growth rate and photosynthetic responses provided evidence for light acclimation but evidence of, at best, only subtle light adaptation.

DNA, Bacterial↗
PubMed2005

Diversity and functional analysis of bacterial communities associated with natural hydrocarbon seeps in acidic soils at Rainbow Springs, Yellowstone National Park.

In this paper we describe the bacterial communities associated with natural hydrocarbon seeps in nonthermal soils at Rainbow Springs, Yellowstone National Park. Soil chemical analysis revealed high sulfate concentrations and low pH values (pH 2.8 to 3.8), which are characteristic of acid-sulfate geothermal activity. The hydrocarbon composition of the seep soils consisted almost entirely of saturated, acyclic alkanes (e.g., n-alkanes with chain lengths of C15 to C30, as well as branched alkanes, predominately pristane and phytane). Bacterial populations present in the seep soils were phylogenetically characterized by 16S rRNA gene clone library analysis. The majority of the sequences recovered (>75%) were related to sequences of heterotrophic acidophilic bacteria, including Acidisphaera spp. and Acidiphilium spp. of the alpha-Proteobacteria. Clones related to the iron- and sulfur-oxidizing chemolithotroph Acidithiobacillus spp. were also recovered from one of the seep soils. Hydrocarbon-amended soil-sand mixtures were established to examine [14C]hexadecane mineralization and corresponding changes in the bacterial populations using denaturing gradient gel electrophoresis (DGGE) of 16S rRNA gene fragments. Approximately 50% of the [14C]hexadecane added was recovered as 14CO2 during an 80-day incubation, and this was accompanied by detection of heterotrophic acidophile-related sequences as dominant DGGE bands. An alkane-degrading isolate was cultivated, whose 16S rRNA gene sequence was identical to the sequence of a dominant DGGE band in the soil-sand mixture, as well as the clone sequence recovered most frequently from the original soil. This and the presence of an alkB gene homolog in this isolate confirmed the alkane degradation capability of one population indigenous to acidic hydrocarbon seep soils.

Bacteria↗
PubMed2005

Diel variations in carbon metabolism by green nonsulfur-like bacteria in alkaline siliceous hot spring microbial mats from Yellowstone National Park.

Green nonsulfur-like bacteria (GNSLB) in hot spring microbial mats are thought to be mainly photoheterotrophic, using cyanobacterial metabolites as carbon sources. However, the stable carbon isotopic composition of typical Chloroflexus and Roseiflexus lipids suggests photoautotrophic metabolism of GNSLB. One possible explanation for this apparent discrepancy might be that GNSLB fix inorganic carbon only during certain times of the day. In order to study temporal variability in carbon metabolism by GNSLB, labeling experiments with [13C]bicarbonate, [14C]bicarbonate, and [13C]acetate were performed during different times of the day. [14C]bicarbonate labeling indicated that during the morning, incorporation of label was light dependent and that both cyanobacteria and GNSLB were involved in bicarbonate uptake. 13C-labeling experiments indicated that during the morning, GNSLB incorporated labeled bicarbonate at least to the same degree as cyanobacteria. The incorporation of [13C]bicarbonate into specific lipids could be stimulated by the addition of sulfide or hydrogen, which both were present in the morning photic zone. The results suggest that GNSLB have the potential for photoautotrophic metabolism during low-light periods. In high-light periods, inorganic carbon was incorporated primarily into Cyanobacteria-specific lipids. The results of a pulse-labeling experiment were consistent with overnight transfer of label to GNSLB, which could be interrupted by the addition of unlabeled acetate and glycolate. In addition, we observed direct incorporation of [13C]acetate into GNSLB lipids in the morning. This suggests that GNSLB also have a potential for photoheterotrophy in situ.

Carbon↗
PubMed2004

Evaluation and validation of the ABI 3700, ABI 3100, and the MegaBACE 1000 capillary array electrophoresis instruments for use with short tandem repeat microsatellite typing in a forensic environment.

The demand for high-throughput DNA profiling has increased with the introduction of national DNA databases and has led to the development of automated methods of short tandem repeat (STR) profile production; however, a potential bottleneck still exists at the gel electrophoresis stage. Capillary electrophoresis sequencers capable of processing 96 samples with considerably reduced manual intervention are now available. In this paper, we compare the ABI Prism 377 slab-gel sequencer currently used by the Forensic Science Service with three leading capillary array electrophoresis instruments: the ABI Prism 3700, the Amersham MegaBACE 1000 and the 16-capillary ABI Prism 3100. We describe the experiments used to evaluate and validate these platforms for forensic use with the STR multiplex Ampf/STR SGMplus [1, 2], along with comparative data from the ABI Prism 377 sequencer.

Databases, Nucleic Acid↗
PubMed2004

Treatment costs for youths receiving multisystemic therapy or hospitalization after a psychiatric crisis.

OBJECTIVE: The authors conducted a cost analysis for multisystemic therapy, an evidence-based treatment that is used as an intensive community-based alternative to the hospitalization of youths presenting with psychiatric emergencies. METHODS: Data from a randomized clinical trial that compared multisystemic therapy with usual inpatient services followed by community aftercare were used to compare Medicaid costs and clinical outcomes during a four-month period postreferral and a 12-month follow-up period. Data were from 115 families receiving Medicaid (out of 156 families in the clinical trial). RESULTS: During the four months postreferral, multisystemic therapy was associated with an average net savings per youth treated of $1,617 compared with usual services. Costs during the 12-month follow-up period were similar between treatments. Multisystemic therapy demonstrated better short-term cost-effectiveness for each of the clinical outcomes (externalizing behavior, internalizing behavior, and global severity of symptoms) than did usual inpatient care and community aftercare. The two treatments demonstrated equivalent long-term cost-effectiveness. CONCLUSIONS: Among youths presenting with psychiatric emergencies, multisystemic therapy was associated with better outcomes at a lower cost during the initial postreferral period and with equivalent costs and outcomes during the 12-month follow-up period.

Adolescent↗
PubMed2003

Geographical isolation in hot spring cyanobacteria.

It has been proposed that free-living microorganisms exhibit ubiquitous dispersal, do not form geographically isolated populations and rarely (if ever) speciate via allopatry. We studied island-like hot spring cyanobacterial communities in which geographical isolation should be prominent and detectable if it influences the evolution of bacteria. The genetic diversity of cyanobacteria indigenous to North American, Japanese, New Zealand and Italian springs was surveyed by (i) amplification and cloning of 16S rRNA and 16S-23S internal transcribed spacer regions; (ii) lineage-specific oligonucleotide probing (used to verify the predominance of cloned sequences); and (iii) lineage-specific polymerase chain reaction (PCR) (used to search for possible rare genotypes). Phylogenetic and distribution patterns were found to be consistent with the occurrence of geographical isolation at both global and local spatial scales, although different cyanobacterial lineages were found to vary in their distribution. A lack of correspondence between biological patterning and the chemical character of springs sampled suggested that the geographical distribution of thermophilic cyanobacteria cannot be explained by the 20 potential niche-determining chemical parameters that we assayed. Thus, geographical isolation (i.e. genetic drift) must in part be responsible for driving the observed evolutionary divergences. Geographical isolation may be an important underestimated aspect of microbial evolution.

Biological Evolution↗
PubMed2003

One-year follow-up of multisystemic therapy as an alternative to the hospitalization of youths in psychiatric crisis.

OBJECTIVE: This study presents findings from a 1-year follow-up to a randomized clinical trial comparing multisystemic therapy (MST), modified for use with youths presenting psychiatric emergencies, with inpatient psychiatric hospitalization. METHOD: One hundred fifty-six children and adolescents approved for emergency psychiatric hospitalization were randomly assigned to home-based MST or inpatient hospitalization followed by usual services. Assessments examining mental health symptoms, out-of-home placement, school attendance, and family relations were conducted at five times: within 24 hours of recruitment, shortly after the hospitalized youth was released from the hospital (1-2 weeks after recruitment), at the completion of MST (average of 4 months postrecruitment), and 10 and 16 months postrecruitment. RESULTS: Based on placement and youth-report measures, MST was initially more effective than emergency hospitalization and usual services at decreasing youths' symptoms and out-of-home placements and increasing school attendance and family structure, but these differences generally dissipated by 12 to 16 months postrecruitment. Hospitalization produced a rapid, but short-lived, decrease in externalizing symptoms based on caregiver reports. CONCLUSION: Findings suggest that youths with serious emotional disturbance might benefit from continuous access to a continuum of evidence-based practices titrated to clinical need.

Adolescent↗
PubMed2003

Compound-specific isotopic fractionation patterns suggest different carbon metabolisms among Chloroflexus-like bacteria in hot-spring microbial mats.

Stable carbon isotope fractionations between dissolved inorganic carbon and lipid biomarkers suggest photoautotrophy by Chloroflexus-like organisms in sulfidic and nonsulfidic Yellowstone hot springs. Where co-occurring, cyanobacteria appear to cross-feed Chloroflexus-like organisms supporting photoheterotrophy as well, although the relatively small 13C fractionation associated with cyanobacterial sugar biosynthesis may sometimes obscure this process.

Biomass↗
PubMed2003

Cyanobacterial ecotypes in different optical microenvironments of a 68 degrees C hot spring mat community revealed by 16S-23S rRNA internal transcribed spacer region variation.

We examined the population of unicellular cyanobacteria (Synechococcus) in the upper 3-mm vertical interval of a 68 degrees C region of a microbial mat in a hot spring effluent channel (Yellowstone National Park, Wyoming). Fluorescence microscopy and microsensor measurements of O(2) and oxygenic photosynthesis demonstrated the existence of physiologically distinct Synechococcus populations at different depths along a light gradient quantified by scalar irradiance microprobes. Molecular methods were used to evaluate whether physiologically distinct populations could be correlated with genetically distinct populations over the vertical interval. We were unable to identify patterns in genetic variation in Synechococcus 16S rRNA sequences that correlate with different vertically distributed populations. However, patterns of variation at the internal transcribed spacer locus separating 16S and 23S rRNA genes suggested the existence of closely related but genetically distinct populations corresponding to different functional populations occurring at different depths.

Base Sequence↗
PubMed2003

Evidence-based financial management.

Like the practice of evidence-based medicine, evidence-based financial management can be used by providers to improve results. The concept provides a framework that managers and researchers can use to help direct efforts in gathering and using evidence to support management decisions in health care.

Accounting↗
PubMed2002

Alkane-1,2-diol-based glycosides and fatty glycosides and wax esters in Roseiflexus castenholzii and hot spring microbial mats.

The lipid composition of Roseiflexus castenholzii, a thermophilic filamentous phototrophic bacterium related to uncultivated filamentous phototrophic bacteria that predominate in hot spring microbial mats, is reported. R. castenholzii lipid extracts were dominated by components characterized by alkane-1-ol-2-alkanoate moieties glycosidically bonded to a C(6) sugar. Similar fatty glycosides, with an additional fatty acid esterified, were detected by HPLC-MS. R. castenholzii also produces a suite of wax esters ranging from 37 to 40 carbon atoms in length. In lipid extracts from two nonsulfidic hot spring microbial mats, similar alkane-1,2-diol-based lipids were detected in minor amounts. R. castenholzii lipids are compared to lipids of mats and other thermophilic mat isolates.

Alkanes↗