Biomedical subjects

Cheryl R Kuske

Publications and source records attributed to Cheryl R Kuske.

14 recordsLinked to original sources

PubMed2006

Current and emerging technologies for the study of bacteria in the outdoor air.

Pathogenic bacteria dispersed into the outdoor air from natural events or human activity are an important concern affecting public health, agriculture, ecological health and international security. Recent studies in atmospheric microbiology have contributed to our ability to detect bacterial pathogens in air samples, increased our knowledge of the spatial and temporal dispersal of aerosolized bacteria and furthered our understanding of the natural bacterial diversity and composition of air. Significant questions remain about bacterial dispersal and diversity in outdoor air that will benefit from continued technological advances. These include determining spatial and temporal patterns of aerosol dispersal, identifying the links between detection, concentration and viability of bacterial pathogens and disease incidence, and documenting the composition and regional movement of bacterial species in air.

Air Microbiology↗

PubMed2006

Microbial biogeography: putting microorganisms on the map.

We review the biogeography of microorganisms in light of the biogeography of macroorganisms. A large body of research supports the idea that free-living microbial taxa exhibit biogeographic patterns. Current evidence confirms that, as proposed by the Baas-Becking hypothesis, 'the environment selects' and is, in part, responsible for spatial variation in microbial diversity. However, recent studies also dispute the idea that 'everything is everywhere'. We also consider how the processes that generate and maintain biogeographic patterns in macroorganisms could operate in the microbial world.

Archaea↗

PubMed2006

Environmental survey for four pathogenic bacteria and closely related species using phylogenetic and functional genes.

Bacterial species with high DNA sequence similarity to pathogens could affect the specificity of assays designed to detect biological threat agents in environmental samples. The natural presence of four pathogenic bacteria, Bacillus anthracis, Clostridium perfringens, Francisella tularensis, and Yersinia pestis and their closely related species, was determined for a large collection of soil and aerosol samples. Polymerase chain reaction (PCR) and gene sequencing were used using group-specific 16S rRNA primers to identify pathogens and related species, and pathogen-specific virulence genes. Close relatives of B. anthracis (B. cereus group species) were detected in 37% of the soils and 25% of the aerosol samples. The B. anthracis protective antigen (pag) gene or a close homolog was detected in 16 of these samples. For the other three pathogen groups, the frequency of detection was much lower, and none of the samples were positive with both the phylogenetic and virulence gene primer sets.

Aerosols↗

PubMed2006

Composition of bacillus species in aerosols from 11 U.S. cities.

A PCR-based heteroduplex assay was used to determine the presence and composition of Bacillus species in 11,059 Environmental Protection Agency PM2.5 aerosol samples from 11 U.S. cities. The assay differentiated three groups: Type A containing Bacillus anthracis and very closely related, often pathogenic, Bacillus cereus and Bacillus thuringiensis strains; Type B containing other B. cereus and B. thuringiensis strains; and a third group of more-distantly related Bacillus species. Eight of the 11 cities were positive for Bacillus species in 50% or more of the samples, and the percent of aerosol samples that contained the HD Type A group ranged from 3% to 32%. Cities from the eastern half of the United States generally contained a higher frequency and broader diversity of Bacillus species than the western half of the United States. Positive samples were detected throughout the year. These results have implications for pathogen detection in environmental samples, understanding the natural evolution of new pathogenic strains, and incidence of infection caused by strains of the B. cereus subgroup.

Aerosols↗

PubMed2005

Changes in nitrogen-fixing and ammonia-oxidizing bacterial communities in soil of a mixed conifer forest after wildfire.

This study was undertaken to examine the effects of forest fire on two important groups of N-cycling bacteria in soil, the nitrogen-fixing and ammonia-oxidizing bacteria. Sequence and terminal restriction fragment length polymorphism (T-RFLP) analysis of nifH and amoA PCR amplicons was performed on DNA samples from unburned, moderately burned, and severely burned soils of a mixed conifer forest. PCR results indicated that the soil biomass and proportion of nitrogen-fixing and ammonia-oxidizing species was less in soil from the fire-impacted sites than from the unburned sites. The number of dominant nifH sequence types was greater in fire-impacted soils, and nifH sequences that were most closely related to those from the spore-forming taxa Clostridium and Paenibacillus were more abundant in the burned soils. In T-RFLP patterns of the ammonia-oxidizing community, terminal restriction fragments (TRFs) representing amoA cluster 1, 2, or 4 Nitrosospira spp. were dominant (80 to 90%) in unburned soils, while TRFs representing amoA cluster 3A Nitrosospira spp. dominated (65 to 95%) in fire-impacted soils. The dominance of amoA cluster 3A Nitrosospira spp. sequence types was positively correlated with soil pH (5.6 to 7.5) and NH(3)-N levels (0.002 to 0.976 ppm), both of which were higher in burned soils. The decreased microbial biomass and shift in nitrogen-fixing and ammonia-oxidizing communities were still evident in fire-impacted soils collected 14 months after the fire.

Ammonia↗

PubMed2005

Detection of diverse new Francisella-like bacteria in environmental samples.

Following detection of putative Francisella species in aerosol samples from Houston, Texas, we surveyed soil and water samples from the area for the agent of tularemia, Francisella tularensis, and related species. The initial survey used 16S rRNA gene primers to detect Francisella species and related organisms by PCR amplification of DNA extracts from environmental samples. This analysis indicated that sequences related to Francisella were present in one water and seven soil samples. This is the first report of the detection of Francisella-related species in soil samples by DNA-based methods. Cloning and sequencing of PCR products indicated the presence of a wide variety of Francisella-related species. Sequences from two soil samples were 99.9% similar to previously reported sequences from F. tularensis isolates and may represent new subspecies. Additional analyses with primer sets developed for detection and differentiation of F. tularensis subspecies support the finding of very close relatives to known F. tularensis strains in some samples. While the pathogenicity of these organisms is unknown, they have the potential to be detected in F. tularensis-specific assays. Similarly, a potential new subspecies of Francisella philomiragia was identified. The majority of sequences obtained, while more similar to those of Francisella than to any other genus, were phylogenetically distinct from known species and formed several new clades potentially representing new species or genera. The results of this study revise our understanding of the diversity and distribution of Francisella and have implications for tularemia epidemiology and our ability to detect bioterrorist activities.

Bacterial Proteins↗

PubMed2004

Virulence signatures: microarray-based approaches to discovery and analysis.

Rapid, accurate, and sensitive detection of biothreat agents requires a broad-spectrum assay capable of discriminating between closely related microbial or viral pathogens. Moreover, in cases where a biological agent release has been identified, forensic analysis demands detailed genetic signature data for accurate strain identification and attribution. To date, nucleic acid sequences have provided the most robust and phylogentically illuminating signature information. Nucleic acid signature sequences are not often linked to genomic or extrachromosomal determinants of virulence, a link that would further facilitate discrimination between pathogens and closely related species. Inextricably coupling genetic determinants of virulence with highly informative nucleic acid signatures would provide a robust means of identifying human, livestock, and agricultural pathogens. By means of example, we present here an overview of two general applications of microarray-based methods for: (1) the identification of candidate virulence factors; and (2) the analysis of genetic polymorphisms that are coupled to Bacillus anthracis virulence factors using an accurate, low cost solid-phase mini-sequencing assay. We show that microarray-based analysis of gene expression can identify potential virulence associated genes for use as candidate signature targets, and, further, that microarray-based single nucleotide polymorphism assays provide a robust platform for the detection and identification of signature sequences in a manner independent of the genetic background in which the signature is embedded. We discuss the strategy as a general approach or pipeline for the discovery of virulence-linked nucleic acid signatures for biothreat agents.

Bacillus anthracis↗

PubMed2004

Diazotrophic community structure and function in two successional stages of biological soil crusts from the Colorado Plateau and Chihuahuan Desert.

The objective of this study was to characterize the community structure and activity of N2-fixing microorganisms in mature and poorly developed biological soil crusts from both the Colorado Plateau and Chihuahuan Desert. Nitrogenase activity was approximately 10 and 2.5 times higher in mature crusts than in poorly developed crusts at the Colorado Plateau site and Chihuahuan Desert site, respectively. Analysis of nifH sequences by clone sequencing and the terminal restriction fragment length polymorphism technique indicated that the crust diazotrophic community was 80 to 90% heterocystous cyanobacteria most closely related to Nostoc spp. and that the composition of N2-fixing species did not vary significantly between the poorly developed and mature crusts at either site. In contrast, the abundance of nifH sequences was approximately 7.5 times greater (per microgram of total DNA) in mature crusts than in poorly developed crusts at a given site as measured by quantitative PCR. 16S rRNA gene clone sequencing and microscopic analysis of the cyanobacterial community within both crust types demonstrated a transition from a Microcoleus vaginatus-dominated, poorly developed crust to mature crusts harboring a greater percentage of Nostoc and Scytonema spp. We hypothesize that ecological factors, such as soil instability and water stress, may constrain the growth of N2-fixing microorganisms at our study sites and that the transition to a mature, nitrogen-producing crust initially requires bioengineering of the surface microenvironment by Microcoleus vaginatus.

Colorado↗

PubMed2003

Statistical analysis of microarray data: a Bayesian approach.

The potential of microarray data is enormous. It allows us to monitor the expression of thousands of genes simultaneously. A common task with microarray is to determine which genes are differentially expressed between two samples obtained under two different conditions. Recently, several statistical methods have been proposed to perform such a task when there are replicate samples under each condition. Two major problems arise with microarray data. The first one is that the number of replicates is very small (usually 2-10), leading to noisy point estimates. As a consequence, traditional statistics that are based on the means and standard deviations, e.g. t-statistic, are not suitable. The second problem is that the number of genes is usually very large (approximately 10,000), and one is faced with an extreme multiple testing problem. Most multiple testing adjustments are relatively conservative, especially when the number of replicates is small. In this paper we present an empirical Bayes analysis that handles both problems very well. Using different parametrizations, we develop four statistics that can be used to test hypotheses about the means and/or variances of the gene expression levels in both one- and two-sample problems. The methods are illustrated using experimental data with prior knowledge. In addition, we present the result of a simulation comparing our methods to well-known statistics and multiple testing adjustments.

Algorithms↗

PubMed2003

Fluorescent heteroduplex assay for monitoring Bacillus anthracis and close relatives in environmental samples.

A fluorescent heteroduplex method was developed to assess the presence of 16S rRNA gene (rDNA) sequences from Bacillus anthracis and close relatives in PCR-amplified 16S rDNA sequence mixtures from environmental samples. The method uses a single-stranded, fluorescent DNA probe, 464 nucleotides in length, derived from a B. anthracis 16S rRNA gene. The probe contains a unique, engineered deletion such that all probe-target duplexes are heteroduplexes with an unpaired G at position 343 (deltaG343). Heteroduplex profiles of sequences >/=85% similar to the probe were produced using an ABI 377 sequencer in less than 3 h. The method divides strains of the Bacillus cereus-Bacillus thuringiensis-B. anthracis group into two subgroups. Each subgroup is defined by a specific 16S rRNA gene sequence type. Sequence type A, containing one mismatch with the probe, occurs in B. anthracis and a small number of closely related clonal lineages represented mostly by food-borne pathogenic isolates of B. cereus and B. thuringiensis. Sequence type B, containing two mismatches with the probe, is found in the majority of B. cereus and B. thuringiensis strains examined to date. Sequence types A and B, when hybridized to the probe, generate two easily differentiated heteroduplexes. Thus, from heteroduplex profiles, the presence of B. cereus-B. thuringiensis-B. anthracis subgroups in environmental samples can be inferred unambiguously. The results show that fluorescent heteroduplex analysis is an effective profiling technique for detection and differentiation of sequences representing small phylogenetic or functional groups in environmental samples.

Bacillus↗

PubMed2002

DNA sequence conservation between the Bacillus anthracis pXO2 plasmid and genomic sequence from closely related bacteria.

BACKGROUND: Complete sequencing and annotation of the 96.2 kb Bacillus anthracis plasmid, pXO2, predicted 85 open reading frames (ORFs). Bacillus cereus and Bacillus thuringiensis isolates that ranged in genomic similarity to B. anthracis, as determined by amplified fragment length polymorphism (AFLP) analysis, were examined by PCR for the presence of sequences similar to 47 pXO2 ORFs. RESULTS: The two most distantly related isolates examined, B. thuringiensis 33679 and B. thuringiensis AWO6, produced the greatest number of ORF sequences similar to pXO2; 10 detected in 33679 and 16 in AWO6. No more than two of the pXO2 ORFs were detected in any one of the remaining isolates. Dot-blot DNA hybridizations between pXO2 ORF fragments and total genomic DNA from AWO6 were consistent with the PCR assay results for this isolate and also revealed nine additional ORFs shared between these two bacteria. Sequences similar to the B. anthracis cap genes or their regulator, acpA, were not detected among any of the examined isolates. CONCLUSIONS: The presence of pXO2 sequences in the other Bacillus isolates did not correlate with genomic relatedness established by AFLP analysis. The presence of pXO2 ORF sequences in other Bacillus species suggests the possibility that certain pXO2 plasmid gene functions may also be present in other closely related bacteria.

Journal Article↗

PubMed2002

Comparison of soil bacterial communities in rhizospheres of three plant species and the interspaces in an arid grassland.

Soil bacteria are important contributors to primary productivity and nutrient cycling in arid land ecosystems, and their populations may be greatly affected by changes in environmental conditions. In parallel studies, the composition of the total bacterial community and of members of the Acidobacterium division were assessed in arid grassland soils using terminal restriction fragment length polymorphism (TRF, also known as T-RFLP) analysis of 16S rRNA genes amplified from soil DNA. Bacterial communities associated with the rhizospheres of the native bunchgrasses Stipa hymenoides and Hilaria jamesii, the invading annual grass Bromus tectorum, and the interspaces colonized by cyanobacterial soil crusts were compared at three depths. When used in a replicated field-scale study, TRF analysis was useful for identifying broad-scale, consistent differences in the bacterial communities in different soil locations, over the natural microscale heterogeneity of the soil. The compositions of the total bacterial community and Acidobacterium division in the soil crust interspaces were significantly different from those of the plant rhizospheres. Major differences were also observed in the rhizospheres of the three plant species and were most apparent with analysis of the Acidobacterium division. The total bacterial community and the Acidobacterium division bacteria were affected by soil depth in both the interspaces and plant rhizospheres. This study provides a baseline for monitoring bacterial community structure and dynamics with changes in plant cover and environmental conditions in the arid grasslands.

Bacteria↗

PubMed2002

Empirical and theoretical bacterial diversity in four Arizona soils.

Understanding patterns of biodiversity in microbial communities is severely constrained by the difficulty of adequately sampling these complex systems. We illustrate the problem with empirical data from small surveys (200-member 16S rRNA gene clone libraries) of four bacterial soil communities from two locations in Arizona. Among the four surveys, nearly 500 species-level groups ( Dunbar et al., Appl. Environ. Microbiol. 65:662-1669, 1999) and 21 bacterial divisions were documented, including four new candidate divisions provisionally designated SC1, SC2, SC3, and SC4. We devised a simple approach to constructing theoretical null models of bacterial species abundance. These null models provide, for the first time, detailed descriptions of soil bacterial community structure that can be used to guide experimental design. Models based on a lognormal distribution were consistent with the observed sizes of the four communities and the richness of the clone surveys. Predictions from the models showed that the species richness of small surveys from complex communities is reproducible, whereas the species composition is not. By using the models, we can now estimate the required survey scale to document specified fractions of community diversity. For example, documentation of half the species in each model community would require surveys of 16,284 to 44,000 individuals. However, quantitative comparisons of half the species in two communities would require surveys at least 10-fold larger for each community.

Arizona↗

PubMed2002

Bacillus anthracis pXO1 plasmid sequence conservation among closely related bacterial species.

The complete sequencing and annotation of the 181.7-kb Bacillus anthracis virulence plasmid pXO1 predicted 143 genes but could only assign putative functions to 45. Hybridization assays, PCR amplification, and DNA sequencing were used to determine whether pXO1 open reading frame (ORF) sequences were present in other bacilli and more distantly related bacterial genera. Eighteen Bacillus species isolates and four other bacterial species were tested for the presence of 106 pXO1 ORFs. Three ORFs were conserved in most of the bacteria tested. Many of the pXO1 ORFs were detected in closely related Bacillus species, and some were detected only in B. anthracis isolates. Three isolates, Bacillus cereus D-17, B. cereus 43881, and Bacillus thuringiensis 33679, contained sequences that were similar to more than one-half of the pXO1 ORF sequences examined. The majority of the DNA fragments that were amplified by PCR from these organisms had DNA sequences between 80 and 98% similar to that of pXO1. Pulsed-field gel electrophoresis revealed large potential plasmids present in both B. cereus 43881 (341 kb) and B. thuringiensis ATCC 33679 (327 kb) that hybridized with a DNA probe composed of six pXO1 ORFs.

Bacillus↗