Genes encoding high-molecular-weight adhesion proteins of nontypeable Haemophilus influenzae are part of gene clusters.

ABSTRACT The HMW1- and HMW2-like adhesion proteins of nontypeable Haemophilus influenzae are expressed by 75% of these strains, and antibodies directed against these proteins are protective in animal models of infection. The purpose of the present study was to define the functional activity of human antibodies specific for these proteins in an in vitro complement-dependent opsonophagocytic assay. Human promyelocytic cell line HL-60 served as the source of phagocytic cells, and a commercial preparation of intravenous immunoglobulin (IVIG) served as the source of human antibodies. High-molecular-weight (HMW) proteins were purified from four prototype nontypeable H. influenzae strains and used to prepare solid-phase affinity columns. IVIG was adsorbed on each column to remove strain-specific anti-HMW antibodies and to allow recovery of affinity-purified anti-HMW antibody fractions. Unadsorbed IVIG killed each of the prototype strains at titers of 1:80 to 1:320. HMW-adsorbed sera demonstrated fourfold decreases in opsonophagocytic titer against the homologous strains compared to unadsorbed IVIG. Affinity-purified anti-HMW antibody preparations demonstrated opsonophagocytic titers of 1:20 to 1:80 against the respective homologous strains and opsonophagocytic titers as high as 1:80 against heterologous strains. None of the affinity-purified anti-HMW antibody preparations was opsonophagocytic for a representative nontypeable H. influenzae strain that did not express HMW1- or HMW2-like proteins. These data demonstrate that human antibodies specific for the HMW1/HMW2-like adhesion proteins of nontypeable H. influenzae are opsonophagocytic and that such antibodies recognize epitopes shared by the HMW proteins of unrelated nontypeable H. influenzae strains. These results argue for continued investigation of the HMW1/HMW2-like proteins as potential vaccine candidates for prevention of disease due to nontypeable H. influenzae.

Download Full-text

Absence of high molecular weight proteins 1 and/or 2 is associated with decreased adherence among non-typeable Haemophilus influenzae clinical isolates

Journal of Medical Microbiology ◽

10.1099/jmm.0.058222-0 ◽

2013 ◽

Vol 62 (11) ◽

pp. 1649-1656 ◽

Cited By ~ 5

Author(s):

Jeni Vuong ◽

Xin Wang ◽

Jordan M. Theodore ◽

Jennifer Whitmon ◽

Patricia Gomez de Leon ◽

...

Keyword(s):

Molecular Weight ◽

Haemophilus Influenzae ◽

High Molecular Weight ◽

Surface Proteins ◽

Clinical Isolates ◽

Type Iv ◽

High Adherence ◽

Genes Encoding ◽

Type Iv Pilin ◽

Pilin Protein

High molecular weight (Hmw) proteins 1 and 2, type IV pilin protein (PilA), outer-membrane protein P5 (OmpP5), Haemophilus protein D (Hpd) and Haemophilus adhesive protein (Hap) are surface proteins involved in the adherence of non-typeable Haemophilus influenzae. One hundred clinical isolates were evaluated for the presence of the genes encoding these proteins by PCR and for their adherence capacity (AC) to Detroit 562 nasopharyngeal cells (D562). The majority of isolates were from blood (77/100); other sites were also represented. Confluent D562 monolayers (1.2×105 cells per well) were inoculated with standardized minimal infective doses (m.o.i.) of 102, 103 or 104 c.f.u. per well. The AC was categorized as low (<10 %) or high (≥10 %) depending on the percentage of c.f.u. adhering per well. All the isolates evaluated showed adherence: 69/100 (69 %) demonstrated high adherence, while 31/100 (31 %) showed low adherence. Of all the genes evaluated, hmw1A and/or hmw2A were detected in 69/100 (69 %) of isolates. The presence of hmw1A and/or hmw2A was associated with increased adherence to D562 cells (P≤0.001). Dot immunoblots were performed to detect protein expression using mAbs 3D6, AD6 and 10C5. Among the high-adherence isolates (n = 69), 72 % reacted with 3D6 and 21 % with 10C5. Our data indicate that the absence of Hmw1 and/or Hmw2 was associated with decreased adherence to D562 cells.

Download Full-text