DNA REPEATS IDENTIFY NOVEL VIRULENCE GENES IN HAEMOPHILUS-INFLUENZAE

The whole genome sequence (1.83 Mbp) of Haemophilus influenzae strain Rd was searched to identify tandem oligonucleotide repeat sequences, L oss or gain of one or more nucleotide repeats through a recombination- independent slippage mechanism is known to mediate phase variation of surface molecules of pathogenic bacteria, including H. influenzae. Thi s facilitates evasion of host defenses and adaptation to the varying m icroenvironments of the host, We reasoned that iterative nucleotides c ould identify novel genes relevant to microbe-host interactions, Our s earch of the Rd genome sequence identified 9 novel loci with multiple (range 6-36, mean 22) tandem tetranucleotide repeats, All were found t o be located within putative open reading frames and included homologu es of hemoglobin-binding proteins of Neisseria, a glycosyltransferase (lgtC gene product) of Neisseria, and an adhesin of Yersinia. These te tranucleotide repeat sequences were also shown to be present in two ot her epidemiologically different H. influenzae type b strains, although the number and distribution of repeats was different, Further charact erization of the lgtC gene showed that it was involved in phenotypic s witching of a lipopolysaccharide epitope and that this variable expres sion was associated with changes in the number of tetranucleotide repe ats, Mutation of lgtC resulted in attenuated virulence of H. influenza e in an infant rat model of invasive infection, These data indicate th e rapidity, economy, and completeness with which whole genome sequence s can be used to investigate the biology of pathogenic bacteria.