Evaluation of an isogenic major outer membrane protein-deficient mutant in the human model of Haemophilus ducreyi infection

Robert E. Throm, Jaffar A. Al-Tawfiq, Kate R. Fortney, Barry Katz, Antoinette F. Hood, Clive A. Slaughter, Eric J. Hansen, Stanley Spinola

Research output: Contribution to journalArticle

26 Citations (Scopus)

Abstract

Haemophilus ducreyi expresses 2 OmpA homologs, designated MOMP and OmpA2, whose genes are arranged in tandem on the chromosome. Northern blot analysis indicated that momp and ompA2 are transcribed independently. Sequences of the momp open reading frame (ORF) lacking the transcriptional start site were amplified by PCR, and an Ω-Km2 cassette was ligated into the ORF. A plasmid containing this construction was electroporated into H. ducreyi 35000HP, and an isogenic MOMP-deficient mutant (35000HP-SMS2) was generated by allele exchange. In Southern blotting, 35000HP-SMS2 contained one copy of the Ω-Km2 cassette in momp. 35000HP and 35000HP-SMS2 had similar outer membrane protein (OMP) and lipooligosaccharide profiles and growth rates except for up-regulation of a putative porin protein in the mutant. Five subjects were inoculated with three doses of live 35000HP-SMS2 on one arm and two doses of live 35000HP and one dose of a heat-killed ContrOl on the other arm in a double-blind escalating dose-response trial. Pustules developed at 7 of 10 sites inoculated with 35000HP and at 6 of 15 sites inoculated with 35000HP-SMS2 (P = 0.14). 35000HP and 35000HP-SMS2 were recovered at similar rates from daily surface cultures and semiquantitative cultures. The data suggest that expression of MOMP is not required for pustule formation by H. ducreyi in the human model of infection.

Original languageEnglish
Pages (from-to)2602-2607
Number of pages6
JournalInfection and Immunity
Volume68
Issue number5
DOIs
StatePublished - May 2000

Fingerprint

Haemophilus ducreyi
Haemophilus Infections
Membrane Proteins
Open Reading Frames
Porins
Mutant Proteins
Southern Blotting
Northern Blotting
Plasmids
Up-Regulation
Hot Temperature
Chromosomes
Alleles
Polymerase Chain Reaction
Growth
Infection
Genes

ASJC Scopus subject areas

  • Immunology

Cite this

Evaluation of an isogenic major outer membrane protein-deficient mutant in the human model of Haemophilus ducreyi infection. / Throm, Robert E.; Al-Tawfiq, Jaffar A.; Fortney, Kate R.; Katz, Barry; Hood, Antoinette F.; Slaughter, Clive A.; Hansen, Eric J.; Spinola, Stanley.

In: Infection and Immunity, Vol. 68, No. 5, 05.2000, p. 2602-2607.

Research output: Contribution to journalArticle

Throm, Robert E. ; Al-Tawfiq, Jaffar A. ; Fortney, Kate R. ; Katz, Barry ; Hood, Antoinette F. ; Slaughter, Clive A. ; Hansen, Eric J. ; Spinola, Stanley. / Evaluation of an isogenic major outer membrane protein-deficient mutant in the human model of Haemophilus ducreyi infection. In: Infection and Immunity. 2000 ; Vol. 68, No. 5. pp. 2602-2607.
@article{bff028ab337f49efb230d2bc56671142,
title = "Evaluation of an isogenic major outer membrane protein-deficient mutant in the human model of Haemophilus ducreyi infection",
abstract = "Haemophilus ducreyi expresses 2 OmpA homologs, designated MOMP and OmpA2, whose genes are arranged in tandem on the chromosome. Northern blot analysis indicated that momp and ompA2 are transcribed independently. Sequences of the momp open reading frame (ORF) lacking the transcriptional start site were amplified by PCR, and an Ω-Km2 cassette was ligated into the ORF. A plasmid containing this construction was electroporated into H. ducreyi 35000HP, and an isogenic MOMP-deficient mutant (35000HP-SMS2) was generated by allele exchange. In Southern blotting, 35000HP-SMS2 contained one copy of the Ω-Km2 cassette in momp. 35000HP and 35000HP-SMS2 had similar outer membrane protein (OMP) and lipooligosaccharide profiles and growth rates except for up-regulation of a putative porin protein in the mutant. Five subjects were inoculated with three doses of live 35000HP-SMS2 on one arm and two doses of live 35000HP and one dose of a heat-killed ContrOl on the other arm in a double-blind escalating dose-response trial. Pustules developed at 7 of 10 sites inoculated with 35000HP and at 6 of 15 sites inoculated with 35000HP-SMS2 (P = 0.14). 35000HP and 35000HP-SMS2 were recovered at similar rates from daily surface cultures and semiquantitative cultures. The data suggest that expression of MOMP is not required for pustule formation by H. ducreyi in the human model of infection.",
author = "Throm, {Robert E.} and Al-Tawfiq, {Jaffar A.} and Fortney, {Kate R.} and Barry Katz and Hood, {Antoinette F.} and Slaughter, {Clive A.} and Hansen, {Eric J.} and Stanley Spinola",
year = "2000",
month = "5",
doi = "10.1128/IAI.68.5.2602-2607.2000",
language = "English",
volume = "68",
pages = "2602--2607",
journal = "Infection and Immunity",
issn = "0019-9567",
publisher = "American Society for Microbiology",
number = "5",

}

TY - JOUR

T1 - Evaluation of an isogenic major outer membrane protein-deficient mutant in the human model of Haemophilus ducreyi infection

AU - Throm, Robert E.

AU - Al-Tawfiq, Jaffar A.

AU - Fortney, Kate R.

AU - Katz, Barry

AU - Hood, Antoinette F.

AU - Slaughter, Clive A.

AU - Hansen, Eric J.

AU - Spinola, Stanley

PY - 2000/5

Y1 - 2000/5

N2 - Haemophilus ducreyi expresses 2 OmpA homologs, designated MOMP and OmpA2, whose genes are arranged in tandem on the chromosome. Northern blot analysis indicated that momp and ompA2 are transcribed independently. Sequences of the momp open reading frame (ORF) lacking the transcriptional start site were amplified by PCR, and an Ω-Km2 cassette was ligated into the ORF. A plasmid containing this construction was electroporated into H. ducreyi 35000HP, and an isogenic MOMP-deficient mutant (35000HP-SMS2) was generated by allele exchange. In Southern blotting, 35000HP-SMS2 contained one copy of the Ω-Km2 cassette in momp. 35000HP and 35000HP-SMS2 had similar outer membrane protein (OMP) and lipooligosaccharide profiles and growth rates except for up-regulation of a putative porin protein in the mutant. Five subjects were inoculated with three doses of live 35000HP-SMS2 on one arm and two doses of live 35000HP and one dose of a heat-killed ContrOl on the other arm in a double-blind escalating dose-response trial. Pustules developed at 7 of 10 sites inoculated with 35000HP and at 6 of 15 sites inoculated with 35000HP-SMS2 (P = 0.14). 35000HP and 35000HP-SMS2 were recovered at similar rates from daily surface cultures and semiquantitative cultures. The data suggest that expression of MOMP is not required for pustule formation by H. ducreyi in the human model of infection.

AB - Haemophilus ducreyi expresses 2 OmpA homologs, designated MOMP and OmpA2, whose genes are arranged in tandem on the chromosome. Northern blot analysis indicated that momp and ompA2 are transcribed independently. Sequences of the momp open reading frame (ORF) lacking the transcriptional start site were amplified by PCR, and an Ω-Km2 cassette was ligated into the ORF. A plasmid containing this construction was electroporated into H. ducreyi 35000HP, and an isogenic MOMP-deficient mutant (35000HP-SMS2) was generated by allele exchange. In Southern blotting, 35000HP-SMS2 contained one copy of the Ω-Km2 cassette in momp. 35000HP and 35000HP-SMS2 had similar outer membrane protein (OMP) and lipooligosaccharide profiles and growth rates except for up-regulation of a putative porin protein in the mutant. Five subjects were inoculated with three doses of live 35000HP-SMS2 on one arm and two doses of live 35000HP and one dose of a heat-killed ContrOl on the other arm in a double-blind escalating dose-response trial. Pustules developed at 7 of 10 sites inoculated with 35000HP and at 6 of 15 sites inoculated with 35000HP-SMS2 (P = 0.14). 35000HP and 35000HP-SMS2 were recovered at similar rates from daily surface cultures and semiquantitative cultures. The data suggest that expression of MOMP is not required for pustule formation by H. ducreyi in the human model of infection.

UR - http://www.scopus.com/inward/record.url?scp=0034058060&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0034058060&partnerID=8YFLogxK

U2 - 10.1128/IAI.68.5.2602-2607.2000

DO - 10.1128/IAI.68.5.2602-2607.2000

M3 - Article

C2 - 10768950

AN - SCOPUS:0034058060

VL - 68

SP - 2602

EP - 2607

JO - Infection and Immunity

JF - Infection and Immunity

SN - 0019-9567

IS - 5

ER -