title: Characterization, distribution, and expression of novel genes among eight clinical isolates of Streptococcus pneumoniae.
abstract: Eight low-passage-number Streptococcus pneumoniae clinical isolates, each of a different serotype and a different multilocus sequence type, were obtained from pediatric participants in a pneumococcal vaccine trial. Comparative genomic analyses were performed with these strains and two S. pneumoniae reference strains. Individual genomic libraries were constructed for each of the eight clinical isolates, with an average insert size of approximately 1 kb. A total of 73,728 clones were picked for arraying, providing more than four times genomic coverage per strain. A subset of 4,793 clones were sequenced, for which homology searches revealed that 750 (15.6%) of the sequences were unique with respect to the TIGR4 reference genome and 263 (5.5%) clones were unrelated to any available streptococcal sequence. Hypothetical translations of the open reading frames identified within these novel sequences showed homologies to a variety of proteins, including bacterial virulence factors not previously identified in S. pneumoniae. The distribution and expression patterns of 58 of these novel sequences among the eight clinical isolates were analyzed by PCR- and reverse transcriptase PCR-based analyses, respectively. These unique sequences were nonuniformly distributed among the eight isolates, and transcription of these genes in planktonic cultures was detected in 81% (172/212) of their genic occurrences. All 58 novel sequences were transcribed in one or more of the clinical strains, suggesting that they all correspond to functional genes. Sixty-five percent (38/58) of these sequences were found in 50% or less of the clinical strains, indicating a significant degree of genomic plasticity among natural isolates.
authors.affiliation: Center for Genomic Sciences, Allegheny-Singer Research Institute, Allegheny General Hospital, 320 East North Avenue, 11th Floor South Tower, Pittsburgh, PA 15212, USA.
authors.forename: Kai
authors.initials: K
authors.lastname: Shen
authors.forename: John
authors.initials: J
authors.lastname: Gladitz
authors.forename: Patricia
authors.initials: P
authors.lastname: Antalis
authors.forename: Bethany
authors.initials: B
authors.lastname: Dice
authors.forename: Benjamin
authors.initials: B
authors.lastname: Janto
authors.forename: Randy
authors.initials: R
authors.lastname: Keefe
authors.forename: Jay
authors.initials: J
authors.lastname: Hayes
authors.forename: Azad
authors.initials: A
authors.lastname: Ahmed
authors.forename: Richard
authors.initials: R
authors.lastname: Dopico
authors.forename: Nathan
authors.initials: N
authors.lastname: Ehrlich
authors.forename: Jennifer
authors.initials: J
authors.lastname: Jocz
authors.forename: Laura
authors.initials: L
authors.lastname: Kropp
authors.forename: Shujun
authors.initials: S
authors.lastname: Yu
authors.forename: Laura
authors.initials: L
authors.lastname: Nistico
authors.forename: David P
authors.initials: DP
authors.lastname: Greenberg
authors.forename: Karen
authors.initials: K
authors.lastname: Barbadora
authors.forename: Robert A
authors.initials: RA
authors.lastname: Preston
authors.forename: J Christopher
authors.initials: JC
authors.lastname: Post
authors.forename: Garth D
authors.initials: GD
authors.lastname: Ehrlich
authors.forename: Fen Z
authors.initials: FZ
authors.lastname: Hu
mesh_terms: D001430:Bacteriocins / Q000235:genetics; D001792:Blood Platelets / Q000378:metabolism / Q000382:microbiology; D002352:Carrier Proteins / Q000235:genetics; D002648:Child; D003001:Cloning, Molecular; D020869:Gene Expression Profiling / Q000379:methods*; D006801:Humans; D008969:Molecular Sequence Data; D010455:Peptides / Q000235:genetics; D012697:Serine Endopeptidases / Q000235:genetics; D013296:Streptococcus pneumoniae / Q000201:enzymology / Q000235:genetics* / Q000302:isolation & purification* / Q000472:pathogenicity; D014774:Virulence; D000519:alpha-Galactosidase / Q000235:genetics
chemical_list: D001430:Bacteriocins; D002352:Carrier Proteins; D010455:Peptides; C069959:mersacidin; D000519:alpha-Galactosidase; D012697:Serine Endopeptidases; C021433:IgA-specific serine endopeptidase