Evolutionary and Functional Relationships among the Nontypeable
            <i>Haemophilus influenzae</i>
            HMW Family of Adhesins

Buscher, Amy Z.; Burmeister, Katie; Barenkamp, Stephen J.; Geme, Joseph W. St.

doi:10.1128/jb.186.13.4209-4217.2004

Cited by 43 publications

(82 citation statements)

References 41 publications

(36 reference statements)

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“…HMWA is the structural component of the adhesin, HMWB has a role in transmembrane translocation, and HMWC is required for glycosylation of HMWA (7,9,37,110). Similarly, strain 86-028NP's two HMW gene clusters contain homologues of the hmwA, hmwB, and hmwC genes in the same gene context as in strain 12 (16). The HMW1A and HMW2A proteins from Hap is an autotransported protein with a domain homologous to the catalytic domain of immunoglobulin A1 proteases.…”

Section: Resultsmentioning

confidence: 99%

Genomic Sequence of an Otitis Media Isolate of Nontypeable Haemophilus influenzae : Comparative Study with H. influenzae Serotype d, Strain KW20

Harrison¹,

et al. 2005

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In 1995, the Institute for Genomic Research completed the genome sequence of a rough derivative of Haemophilus influenzae serotype d, strain KW20. Although extremely useful in understanding the basic biology of H. influenzae, these data have not provided significant insight into disease caused by nontypeable H. influenzae, as serotype d strains are not pathogens. In contrast, strains of nontypeable H. influenzae are the primary pathogens of chronic and recurrent otitis media in children. In addition, these organisms have an important role in acute otitis media in children as well as other respiratory diseases. Such strains must therefore contain a gene repertoire that differs from that of strain Rd. Elucidation of the differences between these genomes will thus provide insight into the pathogenic mechanisms of nontypeable H. influenzae. The genome of a representative nontypeable H. influenzae strain, 86-028NP, isolated from a patient with chronic otitis media was therefore sequenced and annotated. Despite large regions of synteny with the strain Rd genome, there are large rearrangements in strain 86-028NP's genome architecture relative to the strain Rd genome. A genomic island similar to an island originally identified in H. influenzae type b is present in the strain 86-028NP genome, while the mu-like phage present in the strain Rd genome is absent from the strain 86-028NP genome. Two hundred eighty open reading frames were identified in the strain 86-028NP genome that were absent from the strain Rd genome. These data provide new insight that complements and extends the ongoing analysis of nontypeable H. influenzae virulence determinants.In 1995 Haemophilus influenzae strain Rd, a rough derivative of H. influenzae serotype d strain KW20 (strain Rd hereafter), became the first free-living organism to have its genome sequenced to completion (34). Importantly, this also helped establish the large-scale shotgun approach, mated with the utilization of a scaffolding library and computer-assisted assembly, as a rational and expeditious approach for the sequencing of small bacterial genomes. Strain Rd was chosen as the prototypic bacterium for complete genome sequencing as it has a genome size representative of other bacteria and a GϩC content close to that of the human genome. Additionally, at the time of sequencing, a physical map of the strain Rd genome did not exist, so this genome was a good test for the approach of shotgun sequencing, scaffolding, and assembly (34).Although strain Rd is the exemplar organism for the current small-genome sequencing rationale and an important model organism for studying H. influenzae biology, strain Rd is a poor model for the study of pathogenicity caused by members of the genus Haemophilus. Serotype b strains of H. influenzae cause invasive diseases, for example, meningitis, and nontypeable H. influenzae (NTHi) strains principally have a role in localized respiratory disease, particularly in otitis media, acute sinusitis, and community-acquired pneumonia and have important conseque...

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Section: Resultsmentioning

confidence: 99%

Genomic Sequence of an Otitis Media Isolate of Nontypeable Haemophilus influenzae : Comparative Study with H. influenzae Serotype d, Strain KW20

Harrison¹,

et al. 2005

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“…(4). Moreover, on the basis of the lengths of the amplified fragments, complete hmw1A-like and hmw2A-like genes were very likely present.…”

Section: Vol 74 2006 Variation In H Influenzae Hmw Binding Domainsmentioning

confidence: 99%

“…With a few exceptions, our results clearly showed clustering into HMW1-like and HMW2-like groups, in agreement with the hmw1A-like and hmw2A-like encoding genes. Recently, Buscher et al demonstrated that the chromosomal location of the hmw1A and hmw2A genes does not necessarily correlate with the binding phenotype of the associated adhesins, since, in some isolates, the hmw1A gene encodes a protein with HMW2 adherence properties and vice versa (4). In the present study, no investigation was conducted on the actual adherence properties of the hmw1A and hmw2A gene products to human ephithelial cells, our main object being to characterize hmwA alleles from several clinical NTHi isolates by assessing conservation and diversity at sequence level.…”

Section: Vol 74 2006 Variation In H Influenzae Hmw Binding Domainsmentioning

confidence: 99%

“…Recently, the presence of two physically distinct hmw loci has been demonstrated among genetically diverse NTHi isolates possessing hmwA sequences by Southern hybridization (4). Analysis of Escherichia coli transformants expressing these loci from two isolates indicated that both the isolates possess a protein with HMW1-like adherence properties and another with HMW2-like properties (4).…”

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confidence: 99%

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Conservation and Diversity of HMW1 and HMW2 Adhesin Binding Domains among Invasive Nontypeable Haemophilus influenzae Isolates

Giufrè¹,

Muscillo

Spigaglia³

et al. 2006

Infect Immun

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The pathogenesis of nontypeable Haemophilus influenzae (NTHi) begins with adhesion to the rhinopharyngeal mucosa. In almost 80% of NTHi clinical isolates, the HMW proteins are the major adhesins. The prototype HMW1 and HMW2 proteins, identified in NTHi strain 12, exhibit different binding specificities. The two binding domains have been localized in regions of maximal sequence dissimilarity (40% identity, 58% similarity). Two areas within these binding domains have been found essential for full level adhesive activity (designated the core-binding domains). To investigate the conservation and diversity of the HMW1 and HMW2 core-binding domains among isolates, PCR and DNA sequencing were used. First, we separately amplified the hmw1A-like and hmw2A-like structural genes in nine invasive NTHi isolates, discovering two new hmwA alleles, whose sequences are herein reported. Then, the hmw1A-like and hmw2A-like PCR products were used as the template in nested PCR to produce amplicons encompassing the encoding sequences of the two core-binding domains. In-depth sequence analysis was then performed among sequences of each group, with the support of specific computer programs. Overall, extensive sequence diversity among isolates was highlighted. However, similarity plots showed patterns consisting of peaks of relatively high similarity alternating with strongly divergent regions. The phylogenetic tree clearly indicated the HMW1-like and HMW2-like core-binding domain sequences as two clusters. Distinct sets of conserved amino acid motifs were identified within each group of sequences using the MEME/MOTIFSEARCH tool. Since HMW adhesins could represent candidates for future vaccines, identification of specific patterns of conserved motifs in otherwise highly variable regions is of great interest.Nontypeable Haemophilus influenzae (NTHi) is regarded as an important human pathogen, able to cause human respiratory tract disease and, especially in adults, invasive diseases such as meningitis and septicemia (11,12). The initial step in the pathogenesis of NTHi disease involves establishment of bacteria on the rhinopharyngeal respiratory mucosa followed by contiguous spread within the respiratory tract and, occasionally, to sterile sites. A number of adherence factors, including both pilus and nonpilus adhesins, have been identified (24). In almost 80% of NTHi clinical isolates, the HMW proteins are the major adhesins responsible for attachment to human epithelial cells (29). They were originally identified as a group of high-molecular-weight, surface-exposed proteins, predominant targets of human serum antibody response during acute otitis media (1).Given their functional role and their immunogenic character, HMW proteins have gained special consideration as NTHi vaccine candidates. The prototype proteins were detected in NTHi strain 12 and were designated HMW1 and HMW2 (2). They are encoded by separate chromosomal loci, hmw1 and hmw2, respectively, the hmwA gene encoding the structural protein, and two accessory genes, called hm...

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“…The two products were not clearly resolved by gel electrophoresis, so the lack of one hmw locus would not have been detected. Both hmw and hia are known to differ in sequence from strain to strain (4,33). We did not attempt to assess the sequence heterogeneity of these loci; rather the internal primers that we used were selected to hybridize with conserved regions of the hia and hmwA sequences.…”

Section: Phylogenetic Analysis (I) Mlstmentioning

confidence: 99%

Analysis of Genetic Relatedness of Haemophilus influenzae Isolates by Multilocus Sequence Typing

et al. 2008

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The gram-negative bacterium Haemophilus influenzae is a human-restricted commensal of the nasopharynx that can also be associated with disease. The majority of H. influenzae respiratory isolates lack the genes for capsule production and are nontypeable (NTHI). Whereas encapsulated strains are known to belong to serotype-specific phylogenetic groups, the structure of the NTHI population has not been previously described. A total of 656 H. influenzae strains, including 322 NTHI strains, have been typed by multilocus sequence typing and found to have 359 sequence types (ST). We performed maximum-parsimony analysis of the 359 sequences and calculated the majority-rule consensus of 4,545 resulting equally most parsimonious trees. Eleven clades were identified, consisting of six or more ST on a branch that was present in 100% of trees. Two additional clades were defined by branches present in 91% and 82% of trees, respectively. Of these 13 clades, 8 consisted predominantly of NTHI strains, three were serotype specific, and 2 contained distinct NTHI-specific and serotype-specific clusters of strains. Sixty percent of NTHI strains have ST within one of the 13 clades, and eBURST analysis identified an additional phylogenetic group that contained 20% of NTHI strains. There was concordant clustering of certain metabolic reactions and putative virulence loci but not of disease source or geographic origin. We conclude that well-defined phylogenetic groups of NTHI strains exist and that these groups differ in genetic content. These observations will provide a framework for further study of the effect of genetic diversity on the interaction of NTHI with the host.Haemophilus influenzae is a small (1 to 2 m in length) gram-negative bacterium that is found only in humans. The polysaccharide-protein conjugate vaccines against serotype b H. influenzae have almost eliminated H. influenzae as a cause of pediatric meningitis in the western world. However, unencapsulated (nontypeable) H. influenzae (NTHI) remains an important pathogen, particularly in children and the elderly (5,8,23). NTHI infections are usually limited to respiratory mucosal sites such as the middle ear or bronchi but are occasionally systemic. It is not known whether NTHI isolates associated with localized or systemic disease are genetically distinct from each other or distinct from isolates associated with asymptomatic colonization of the nasopharynx.Efforts to understand and control NTHI disease have been hampered by the diversity of these bacteria. Many of the surface antigens that have been studied display interstrain and intrastrain heterogeneity as a result of both sequence divergence and phase variation. It is increasingly recognized that NTHI isolates also vary in genetic content. We use the term island to refer to a genetic locus (one or more genes) that occurs in some but not all strains. As used here, the term does not imply that the locus is known to be readily transferred between strains or is thought to have been recently acquired. Islands whose functi...

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Evolutionary and Functional Relationships among the Nontypeable Haemophilus influenzae HMW Family of Adhesins

Cited by 43 publications

References 41 publications

Genomic Sequence of an Otitis Media Isolate of Nontypeable Haemophilus influenzae : Comparative Study with H. influenzae Serotype d, Strain KW20

Genomic Sequence of an Otitis Media Isolate of Nontypeable Haemophilus influenzae : Comparative Study with H. influenzae Serotype d, Strain KW20

Conservation and Diversity of HMW1 and HMW2 Adhesin Binding Domains among Invasive Nontypeable Haemophilus influenzae Isolates

Analysis of Genetic Relatedness of Haemophilus influenzae Isolates by Multilocus Sequence Typing

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