Limits...
A functional gene array for detection of bacterial virulence elements.

Jaing C, Gardner S, McLoughlin K, Mulakken N, Alegria-Hartman M, Banda P, Williams P, Gu P, Wagner M, Manohar C, Slezak T - PLoS ONE (2008)

Bottom Line: When tested with organisms at varying phylogenetic distances from the four target strains, the array detected orthologs for the majority of targeted gene families present in bacteria belonging to the same taxonomic family.In combination with whole-genome amplification, the array detects femtogram concentrations of purified DNA, either spiked in to an aerosol sample background, or in combinations from one or more of the four target organisms.By targeting virulence gene families as well as genes unique to specific biothreat agents, these arrays will provide important data about the pathogenic potential and drug resistance profiles of unknown organisms in environmental samples.

View Article: PubMed Central - PubMed

Affiliation: Chemistry, Materials, Earth and Life Sciences, Lawrence Livermore National Laboratory, Livermore, California, United States of America. jaing2/at/llnl.gov

ABSTRACT
Emerging known and unknown pathogens create profound threats to public health. Platforms for rapid detection and characterization of microbial agents are critically needed to prevent and respond to disease outbreaks. Available detection technologies cannot provide broad functional information about known or novel organisms. As a step toward developing such a system, we have produced and tested a series of high-density functional gene arrays to detect elements of virulence and antibiotic resistance mechanisms. Our first generation array targets genes from Escherichia coli strains K12 and CFT073, Enterococcus faecalis and Staphylococcus aureus. We determined optimal probe design parameters for gene family detection and discrimination. When tested with organisms at varying phylogenetic distances from the four target strains, the array detected orthologs for the majority of targeted gene families present in bacteria belonging to the same taxonomic family. In combination with whole-genome amplification, the array detects femtogram concentrations of purified DNA, either spiked in to an aerosol sample background, or in combinations from one or more of the four target organisms. This is the first report of a high density NimbleGen microarray system targeting microbial antibiotic resistance and virulence mechanisms. By targeting virulence gene families as well as genes unique to specific biothreat agents, these arrays will provide important data about the pathogenic potential and drug resistance profiles of unknown organisms in environmental samples.

Show MeSH

Related in: MedlinePlus

Effect of the position of mismatches on the hybridization of target and probe.The mean mismatch (MM) probe intensity vs perfect match (PM) probe intensity ratio, averaged over 60 PM probes and their corresponding MM probes, from 10 arrays is plotted vs the position of the MM.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2367441&req=5

pone-0002163-g005: Effect of the position of mismatches on the hybridization of target and probe.The mean mismatch (MM) probe intensity vs perfect match (PM) probe intensity ratio, averaged over 60 PM probes and their corresponding MM probes, from 10 arrays is plotted vs the position of the MM.

Mentions: To more comprehensively assess the factors influencing the balance between probe sensitivity and specificity, we analyzed data from two series of probes, containing single and multiple mismatches respectively. We first examined data from probes that perfectly matched the hybridized strain, except for a single mismatch (MM) base placed at a known position. We investigated the effect of the MM position on the probe intensity, relative to the intensity of the corresponding perfect match (PM) probe for the same hybridization. Figure 5 shows mean intensity ratios (MM/PM) with error bars corresponding to 95% confidence intervals, averaged over 60 PM probes and their cognate MM probes, from 10 arrays hybridized to a variety of samples. The MM positions are numbered in 5′ to 3′ order, and probe lengths range from 30 to 66.


A functional gene array for detection of bacterial virulence elements.

Jaing C, Gardner S, McLoughlin K, Mulakken N, Alegria-Hartman M, Banda P, Williams P, Gu P, Wagner M, Manohar C, Slezak T - PLoS ONE (2008)

Effect of the position of mismatches on the hybridization of target and probe.The mean mismatch (MM) probe intensity vs perfect match (PM) probe intensity ratio, averaged over 60 PM probes and their corresponding MM probes, from 10 arrays is plotted vs the position of the MM.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2367441&req=5

pone-0002163-g005: Effect of the position of mismatches on the hybridization of target and probe.The mean mismatch (MM) probe intensity vs perfect match (PM) probe intensity ratio, averaged over 60 PM probes and their corresponding MM probes, from 10 arrays is plotted vs the position of the MM.
Mentions: To more comprehensively assess the factors influencing the balance between probe sensitivity and specificity, we analyzed data from two series of probes, containing single and multiple mismatches respectively. We first examined data from probes that perfectly matched the hybridized strain, except for a single mismatch (MM) base placed at a known position. We investigated the effect of the MM position on the probe intensity, relative to the intensity of the corresponding perfect match (PM) probe for the same hybridization. Figure 5 shows mean intensity ratios (MM/PM) with error bars corresponding to 95% confidence intervals, averaged over 60 PM probes and their cognate MM probes, from 10 arrays hybridized to a variety of samples. The MM positions are numbered in 5′ to 3′ order, and probe lengths range from 30 to 66.

Bottom Line: When tested with organisms at varying phylogenetic distances from the four target strains, the array detected orthologs for the majority of targeted gene families present in bacteria belonging to the same taxonomic family.In combination with whole-genome amplification, the array detects femtogram concentrations of purified DNA, either spiked in to an aerosol sample background, or in combinations from one or more of the four target organisms.By targeting virulence gene families as well as genes unique to specific biothreat agents, these arrays will provide important data about the pathogenic potential and drug resistance profiles of unknown organisms in environmental samples.

View Article: PubMed Central - PubMed

Affiliation: Chemistry, Materials, Earth and Life Sciences, Lawrence Livermore National Laboratory, Livermore, California, United States of America. jaing2/at/llnl.gov

ABSTRACT
Emerging known and unknown pathogens create profound threats to public health. Platforms for rapid detection and characterization of microbial agents are critically needed to prevent and respond to disease outbreaks. Available detection technologies cannot provide broad functional information about known or novel organisms. As a step toward developing such a system, we have produced and tested a series of high-density functional gene arrays to detect elements of virulence and antibiotic resistance mechanisms. Our first generation array targets genes from Escherichia coli strains K12 and CFT073, Enterococcus faecalis and Staphylococcus aureus. We determined optimal probe design parameters for gene family detection and discrimination. When tested with organisms at varying phylogenetic distances from the four target strains, the array detected orthologs for the majority of targeted gene families present in bacteria belonging to the same taxonomic family. In combination with whole-genome amplification, the array detects femtogram concentrations of purified DNA, either spiked in to an aerosol sample background, or in combinations from one or more of the four target organisms. This is the first report of a high density NimbleGen microarray system targeting microbial antibiotic resistance and virulence mechanisms. By targeting virulence gene families as well as genes unique to specific biothreat agents, these arrays will provide important data about the pathogenic potential and drug resistance profiles of unknown organisms in environmental samples.

Show MeSH
Related in: MedlinePlus