ERIC-PCR fingerprinting-based community DNA hybridization to pinpoint genome-specific fragments as molecular markers to identify and track populations common to healthy human guts.
Bacterial populations common to healthy human guts may play important roles in human health. A new strategy for discovering genomic sequences as markers for these bacteria was developed using Enterobacterial Repetitive Intergenic Consensus (ERIC)-PCR fingerprinting. Structural features within microbial communities are compared with ERIC-PCR followed by DNA hybridization to identify genomic fragments shared by samples from healthy human individuals. ERIC-PCR profiles of fecal samples from 12 diseased or healthy human and piglet subjects demonstrated stable, unique banding patterns for each individual tested. Sequence homology of DNA fragments in bands of identical size was examined between samples by hybridization under high stringency conditions with DIG-labeled ERIC-PCR products derived from the fecal sample of one healthy child. Comparative analysis of the hybridization profiles with the original agarose fingerprints identified three predominant bands as signatures for populations associated with healthy human guts with sizes of 500, 800 and 1000 bp. Clone library profiling of the three bands produced 17 genome fragments, three of which showed high similarity only with regions of the Bacteroides thetaiotaomicron genome, while the remainder were orphan sequences. Association of these sequences with healthy guts was validated by sequence-selective PCR experiments, which showed that a single fragment was present in all 32 healthy humans and 13 healthy piglets tested. Two fragments were present in the healthy human group and in 18 children with non-infectious diarrhea but not in eight children with infectious diarrhea. Genome fragments identified with this novel strategy may be used as genome-specific markers for dynamic monitoring and sequence-guided isolation of functionally important bacterial populations in complex communities such as human gut microflora.
Laboratory of Molecular Microbial Ecology and Ecogenomics, College of Life Science and Biotechnology, Shanghai Jiao Tong University, Shanghai 200240, China.
Guifang
G
Wei
Li
L
Pan
Huimin
H
Du
Junyi
J
Chen
Liping
L
Zhao
D000328:Adult; D000818:Animals; D001483:Base Sequence; D002648:Child; D016000:Cluster Analysis; D016172:DNA Fingerprinting / Q000379:methods; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D003967:Diarrhea / Q000382:microbiology; D004064:Digestive System / Q000382:microbiology*; D004755:Enterobacteriaceae / Q000145:classification / Q000235:genetics* / Q000254:growth & development; D005183:Failure to Thrive / Q000382:microbiology; D005243:Feces / Q000382:microbiology; D005260:Female; D006801:Humans; D007223:Infant; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D016133:Polymerase Chain Reaction / Q000379:methods*; D013552:Swine
D004269:DNA, Bacterial