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Journal of Bacteriology, February 2009, p. 822-831, Vol. 191, No. 3
0021-9193/09/$08.00+0     doi:10.1128/JB.00782-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Delineation of the Species Haemophilus influenzae by Phenotype, Multilocus Sequence Phylogeny, and Detection of Marker Genes{triangledown} ,{dagger}

Niels Nørskov-Lauritsen,1* Merete D. Overballe,1 and Mogens Kilian2

Department of Clinical Microbiology, Aarhus University Hospital Skejby, DK-8200 Aarhus N,1 Institute of Medical Microbiology and Immunology, Aarhus University, DK-8000 Aarhus C, Denmark2

Received 4 June 2008/ Accepted 26 November 2008

To obtain more information on the much-debated definition of prokaryotic species, we investigated the borders of Haemophilus influenzae by comparative analysis of H. influenzae reference strains with closely related bacteria including strains assigned to Haemophilus haemolyticus, cryptic genospecies biotype IV, and the never formally validated species "Haemophilus intermedius". Multilocus sequence phylogeny based on six housekeeping genes separated a cluster encompassing the type and the reference strains of H. influenzae from 31 more distantly related strains. Comparison of 16S rRNA gene sequences supported this delineation but was obscured by a conspicuously high number of polymorphic sites in many of the strains that did not belong to the core group of H. influenzae strains. The division was corroborated by the differential presence of genes encoding H. influenzae adhesion and penetration protein, fuculokinase, and Cu,Zn-superoxide dismutase, whereas immunoglobulin A1 protease activity or the presence of the iga gene was of limited discriminatory value. The existence of porphyrin-synthesizing strains ("H. intermedius") closely related to H. influenzae was confirmed. Several chromosomally encoded hemin biosynthesis genes were identified, and sequence analysis showed these genes to represent an ancestral genotype rather than recent transfers from, e.g., Haemophilus parainfluenzae. Strains previously assigned to H. haemolyticus formed several separate lineages within a distinct but deeply branching cluster, intermingled with strains of "H. intermedius" and cryptic genospecies biotype IV. Although H. influenzae is phenotypically more homogenous than some other Haemophilus species, the genetic diversity and multicluster structure of strains traditionally associated with H. influenzae make it difficult to define the natural borders of that species.

* Corresponding author. Mailing address: Department of Clinical Microbiology, Aarhus University Hospital Skejby, Brendstrupgaardsvej, DK-8200 Aarhus N, Denmark. Phone: 45 8949 5603. Fax: 45 8949 5611. E-mail: nielnoer{at}rm.dk

{triangledown} Published ahead of print on 5 December 2008.

{dagger} Supplemental material for this article may be found at http://jb.asm.org/.

Journal of Bacteriology, February 2009, p. 822-831, Vol. 191, No. 3
0021-9193/09/$08.00+0     doi:10.1128/JB.00782-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Norskov-Lauritsen, N. (2009). Detection of Cryptic Genospecies Misidentified as Haemophilus influenzae in Routine Clinical Samples by Assessment of Marker Genes fucK, hap, and sodC. J. Clin. Microbiol. 47: 2590-2592 [Abstract] [Full Text]  


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