Prevalence of the sodC Gene in Nontypeable Haemophilus influenzae and Haemophilus haemolyticus by Microarray-Based Hybridization

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Abstract
The sodC gene has been reported to be a useful marker for differentiating nontypeable (NT) Haemophilus influenzae from Haemophilus haemolyticus in respiratory-tract samples, but discrepancies exist as to the prevalence of sodC in NT H. influenzae . Therefore, we used a microarray-based, “library-on-a-slide” method to differentiate the species and found that 21 of 169 (12.4%) NT H. influenzae strains and all 110 (100%) H. haemolyticus strains possessed the sodC gene. Multilocus sequence analysis confirmed that the 21 NT H. influenzae strains were H. influenzae and not H. haemolyticus . An inactive sodC gene has been reported in encapsulated H. influenzae strains belonging to phylogenetic division II. Capsule-specific Southern hybridization and PCR and a lack of copper/zinc-cofactored superoxide dismutase (CuZnSOD) expression indicated that 6 of the 21 sodC -containing NT H. influenzae strains in our study were likely capsule-deficient mutants belonging to phylogenetic division II. DNA sequence comparisons of the 21 H. influenzae sodC genes with sodC from H. haemolyticus or encapsulated H. influenzae demonstrated that the sodC genes of the six H. influenzae capsule-deficient mutants were, on average, 99% identical to sodC from encapsulated H. influenzae but only 85% identical to sodC from H. haemolyticus . The sodC genes from 2/15 NT H. influenzae strains were similarly more closely related to sodC from encapsulated strains, while sodC genes from 13 NT H. influenzae strains were almost 95% identical to sodC genes from H. haemolyticus , suggesting the possibility of interspecies recombination in these strains. In summary, this study demonstrates that sodC is not completely absent (9.2%) in true NT H. influenzae strains.

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