Identification and characterization of an in vivo regulated D15/Oma87 homologue in Shigella flexneri using differential display polymerase chain reaction

Shigella genes expressed during infection likely contribute to adaptation a nd virulence in the host. Using differential display PCR (DDPCR), a cDNA fr agment from Shigella flexneri serotype 5 that showed enhanced expression in a murine model was identified, cloned and sequenced. Enhanced expression w as verified by RNA dot blot. The full-length gene was cloned using PCR and sequenced. The complete gene sequence was BLAST searched against GenBank, a nd exhibited strong homology to genes encoding Haemophilus influenzae D15 a nd Pasteurella multocida Oma87 protective outer membrane antigens. The S. f lexneri gene putatively encodes a similar to 90-kDa protein and was termed oma90. The deduced amino acid sequence from oma90 was analyzed and compared to the D15/Oma87 antigens. Additionally, oma90 mapped to a cluster of orth ologous groups, and probably contains tin ancient conserved domain. The chr omosomal organization of oma90 was similar to that for H. influenzae and P. multocida as well as for other known homologues. Northern blot revealed th at the oma90 transcript encoded only oma90. This report represents the firs t description of a S. flexneri gene identified based on enhanced expression in the host. Furthermore, we report the first evidence demonstrating in vi vo regulation of a member of the d15/oma87 gene family. (C) 2001 Elsevier S cience B.V. All rights reserved.