Inhibition of Phagocytosis by<i>Haemophilus ducreyi</i>Requires Expression of the LspA1 and LspA2 Proteins

Väkeväinen, Merja; Greenberg, Steven M.; Hansen, Eric J.

doi:10.1128/iai.71.10.5994-6003.2003

Cited by 48 publications

(54 citation statements)

References 58 publications

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“…This could represent a recombination event, either from externally acquired DNA or an internal rearrangement of the chromosome. LspA1, LspA2, and LspB are used by H. ducreyi to avoid phagocytic uptake and are critical virulence factors (42). The LspA1 and LspA2 predicted protein sequences are 86% identical to each other (43).…”

Section: Resultsmentioning

confidence: 99%

Rapid Divergence of Two Classes of Haemophilus ducreyi

et al. 2011

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Haemophilus ducreyi, the etiologic agent of chancroid, expresses variants of several key virulence factors. While previous reports suggested that H. ducreyi strains formed two clonal populations, the differences between, and diversity within, these populations were unclear. To assess their variability, we examined sequence diversity at 11 H. ducreyi loci, including virulence and housekeeping genes, augmenting published data sets with PCR-amplified genes to acquire data for at least 10 strains at each locus. While sequences from all 11 loci place strains into two distinct groups, there was very little variation within each group. The difference between alleles of the two groups was variable and large at 3 loci encoding surface-exposed proteins (0.4 < K S < 1.3, where K S is divergence at synonymous sites) but consistently small at genes encoding cytoplasmic or periplasmic proteins (K S < 0.09). The data suggest that the two classes have recently diverged, that recombination has introduced variant alleles into at least 3 distinct loci, and that these alleles have been confined to one of the two classes. In addition, recombination is evident among alleles within, but not between, classes. Rather than clones of the same species, these properties indicate that the two classes may form distinct species.Haemophilus ducreyi is a Gram-negative coccobacillus in the family Pasteurellaceae (2) and the causative agent of chancroid, a sexually transmitted, genital ulcer disease which facilitates transmission of 14,23,44). While H. ducreyi was placed in the polyphyletic genus Haemophilus due to its requirement for hemin, Haemophilus species do not form a cohesive group within the Pasteurellaceae (6, 20). The species that are most closely related to H. ducreyi include Actinobacter pleuropneumoniae and Nicoletella semolina (see Fig. S1 in the supplemental material) (16).Previous attempts to examine the diversity within the species H. ducreyi using immunotyping (36) and outer membrane profiling (32) showed very few differences among strains. Molecular analyses showed some differences, e.g., ribotyping of strains from two outbreaks yielded multiple restriction fragment length polymorphism (RFLP) patterns (19), but designations based on these differences have not been widely adopted or used for epidemiologic studies, in part due to the difficulty in culturing this fastidious organism. The lack of a standard typing system makes epidemiologic studies more difficult, and epidemiologic data for chancroid are therefore limited.One way to study diversity in H. ducreyi is to assess its population structure. Whereas the lack of recombination in some bacterial species generates stable clones (5), others recombine sufficiently rapidly to obscure clonal relationships (10, 11). While many Haemophilus species are naturally competent, H. ducreyi is considered nontransformable due to defects in the DNA uptake apparatus (16,34). This would likely decrease rates of recombination and generate stable clones. Initial studies of the clonal structure of...

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Section: Resultsmentioning

confidence: 99%

Rapid Divergence of Two Classes of Haemophilus ducreyi

et al. 2011

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“…In pustules and ulcers, H. ducreyi colocalizes with macrophages and neutrophils but resists phagocytosis and phagocytic killing and replicates extracellularly (6,7). H. ducreyi secretes two antiphagocytic proteins, LspA1 and LspA2, which inhibit Fc receptor-mediated uptake by macrophage-and neutrophil-like cell lines in vitro (50); a mutant lacking LspA1 and LspA2 expression is attenuated in human volunteers (20). The mechanism of H. ducreyi clearance is unknown, but clearance is likely mediated by bacterial uptake and killing by phagocytes.…”

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confidence: 99%

Haemophilus ducreyi-Induced Interleukin-10 Promotes a Mixed M1 and M2 Activation Program in Human Macrophages

Katz

Spinola

2012

Infect Immun

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“…The abilities of wild-type and mutant strains of H. ducreyi to inhibit phagocytic activity were measured by using the mouse monocytemacrophage cell line J774A.1 (TIB-67; American Type Culture Collection, Manassas, Va.) and opsonized fluorescent microspheres as described previously (38). Briefly, bacteria were incubated with the J774A.1 cell monolayers for 1 h at 33°C, and then the microspheres were added.…”

Section: Methodsmentioning

confidence: 99%

“…All H. ducreyi strains were grown on chocolate agar plates supplemented with 1% IsoVitaleX and incubated at 35°C with 5% CO 2 , or they were grown in broth consisting of proteose peptone, 50 g of hemin per ml, 1% IsoVitaleX, and 5% heat-inactivated fetal calf serum, or in supplemented Columbia broth as described elsewhere (38). Where appropriate, the medium was supplemented with kanamycin (30 g/ml) and chloramphenicol (1.5 g/ml).…”

Section: Methodsmentioning

confidence: 99%

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Expression of the LspA1 and LspA2 Proteins by Haemophilus ducreyi Is Required for Virulence in Human Volunteers

Janowicz¹,

Fortney²,

Katz³

et al. 2004

Infect Immun

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Haemophilus ducreyi colocalizes with polymorphonuclear leukocytes and macrophages and evades phagocytosis during experimental infection of human volunteers. H. ducreyi contains two genes, lspA1 and lspA2, which encode predicted proteins of 456 and 543 kDa, respectively. Compared to its wild-type parent, an lspA1 lspA2 double mutant does not inhibit phagocytosis by macrophage and myelocytic cell lines in vitro and is attenuated in an experimental rabbit model of chancroid. To test whether expression of LspA1 and LspA2 was necessary for virulence in humans, six volunteers were experimentally infected. Each volunteer was inoculated with three doses (ranging from 85 to 112 CFU) of the parent (35000HP) in one arm and three doses (ranging from 60 to 822 CFU) of the mutant (35000HP⍀12) in the other arm. The papule formation rates were 88% (95% confidence interval [95% CI], 76.8 to 99.9%) at 18 parent sites and 72% (95% CI, 44.4 to 99.9%) at 18 mutant sites (P ‫؍‬ 0.19). However, papules were significantly smaller at mutant sites (mean size, 24.8 mm 2 ) than at parent sites (mean size, 39.1 mm 2 ) 24 h after inoculation (P ‫؍‬ 0.0002). The pustule formation rates were 44% (95% CI, 5.8 to 77.6%) at parent sites and 0% (95% CI, 0 to 39.4%) at mutant sites (P ‫؍‬ 0.009). With the caveat that biosafety regulations preclude testing of a complemented mutant in human subjects, these results indicate that expression of LspA1 and LspA2 facilitates the ability of H. ducreyi to initiate disease and to progress to pustule formation in humans.

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Inhibition of Phagocytosis byHaemophilus ducreyiRequires Expression of the LspA1 and LspA2 Proteins

Cited by 48 publications

References 58 publications

Rapid Divergence of Two Classes of Haemophilus ducreyi

Rapid Divergence of Two Classes of Haemophilus ducreyi

Haemophilus ducreyi-Induced Interleukin-10 Promotes a Mixed M1 and M2 Activation Program in Human Macrophages

Expression of the LspA1 and LspA2 Proteins by Haemophilus ducreyi Is Required for Virulence in Human Volunteers

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