Oligonucleotide microarray for identification ofEnterococcusspecies

Abstract

For detection of most members of the Enterococcaceae, the specificity of a novel oligonucleotide microarray (ECC-PhyloChip) consisting of 41 hierarchically nested 16S or 23S rRNA gene-targeted probes was evaluated with 23 pure cultures (including 19 Enterococcus species). Target nucleic acids were prepared by PCR amplification of a 4.5-kb DNA fragment containing large parts of the 16S and 23S rRNA genes and were subsequently labeled fluorescently by random priming. Each tested member of the Enterococcaceae was correctly identified on the basis of its unique microarray hybridization pattern. The evaluated ECC-PhyloChip was successfully applied for identification of Enterococcus faecium and Enterococcus faecalis in artificially contaminated milk samples demonstrating the utility of the ECC-PhyloChip for parallel identification and differentiation of Enterococcus species in food samples.

Development of a DNA Microarray for Enterococcal Species, Virulence, and Antibiotic Resistance Gene Determinations among Isolates from Poultry

“…Taxonomic microarrays generally rely on 16S or 23S rRNA genes or the intervening sequence to identify bacterial species (31,58). A previous enterococcus-specific array used 18-mer sequences to define species-specific probes.…”

“…Eighteen Enterococcus species most commonly used for the development of new identification methods (28,31) were targeted for the Enteroarray. Confirmation of selected enterococcal isolates was performed by sequencing a segment of the cpn60 gene amplified by the universal cpn60 primers H729 (5Ј-CGC CAG GGT TTT CCC AGT CAC GAC GAI III GCI GGI GAY GGI ACI ACI AC-3Ј) and H730 (5Ј-AGC GGA TAA CAA TTT CAC ACA GGA YKI YKI TCI CCR AAI CCI GGI GCY TT-3Ј), including the M13F and M13R sequences, respectively (underlined) (34).…”

Greengenes, a Chimera-Checked 16S rRNA Gene Database and Workbench Compatible with ARB

“…Comparative analysis of 16S small-subunit rRNA genes is commonly used to survey the constituents of microbial communities (4,13,23,24), to infer bacterial and archaeal evolution (14,19), and to design monitoring and analysis tools, such as microarrays (5,10,17,20,29,30). Because the rate of production of 16S small-subunit rRNA gene sequence records for uncultured organisms now exceeds the rate of production for their cultured counterparts, taxonomic placement of sequences lags behind.…”

Application of a High-Density Oligonucleotide Microarray Approach To Study Bacterial Population Dynamics during Uranium Reduction and Reoxidation

“…To validate this approach, we also analyzed identical samples using a more routine clone library approach in addition to confirmatory tests using quantitative PCR. Microarrays targeting functional genes have been employed for analysis of biodegradative capabilities in contaminated sites (43), while 16S rRNA gene microarrays have been used successfully to differentiate bacteria in specific groups, such as Enterococcus (26), Cyanobacteria, (4), nitrifying bacteria (24), fish pathogens (61), and human colon microflora (37). Here we report the first application of high-density array technology in profiling the complex microbial communities of soils or sediments.…”