Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

H. ducreyi 35000HP grown in the presence or absence of FCS

ABSTRACT: Haemophilus ducreyi 35000HP was subjected to different in vitro growth conditions that resemble the in vivo host environment (presence of serum), in order to ascertain the overall changes in gene expression. This information might identify genes important for colonization and/or survival inside the host. H. ducreyi 35000HP was grown for 8 hours in Columbia broth in the presence or absence of FCS. After 8 hours total RNA was isolated and processed for DNA microarray analysis. This study includes four biological replicates (paired samples of bacteria grown in the presence/absence of FCS), one of which (Rep1) was selected for dye swap.

ORGANISM(S): Haemophilus ducreyi 35000HP

SUBMITTER: Maria Labandeira-Rey

PROVIDER: E-GEOD-13851 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Regulation of expression of the Haemophilus ducreyi LspB and LspA2 proteins by CpxR.

Labandeira-Rey Maria M Mock Jason R JR Hansen Eric J EJ

Infection and immunity 20090518 8

The LspA1, LspA2, and LspB proteins of Haemophilus ducreyi comprise a two-partner secretion system that has been shown to be necessary for H. ducreyi to inhibit phagocytosis by immune cells in vitro. Inactivation of lspA1 resulted in increased levels of LspA2, suggesting that these two proteins are differentially controlled (C. J. Ward et al., Infect. Immun. 71:2478-2486, 2003). Expression of LspA2 but not LspA1 was shown to be both growth phase dependent and affected by the presence of fetal ca ...[more]

PMID: 19451237

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Project description:Comparison of gene expression from subjects who resolved or formed pustules to H.ducreyi. In human inoculation experiments, the cutaneous immune response to Haemophilus ducreyi consists of serum, PMN, macrophages, T cells and myeloid DC. In reinfection experiments, some subjects form pustules twice (PP group) or resolve infection twice (RR group). Although pustule formation is associated with serum resistance and phagocytic failure, there are no differences in the ability of isolated phagocytes or serum obtained from PP and RR subjects to ingest or kill H. ducreyi. To identify the basis for differential host susceptibility to H. ducreyi, we used microarrays to profile gene expression in infected and uninfected tissue and monocyte-derived DC obtained from PP and RR subjects. In infected tissue, both groups had a core response to H. ducreyi. Many additional transcripts that signify active immune function were upregulated exclusively in RR tissue, while PP tissue exclusively contained differentially regulated transcripts consistent with immune dysregulation. The core response of infected DC from both groups was typical of a DC1 response. RR DC exclusively expressed many additional transcripts indicative of DC1 function, while PP DC uniquely expressed differentially regulated transcripts characteristic of both DC1 and DCreg. The data suggest that DC from PP and RR subjects are prewired to respond differentially to H. ducreyi. Experiment Overall Design: Six healthy adult volunteers (5 females, 1 male, 36 Â± 13 yrs, mean age Â± SD) who had been infected with H. ducreyi twice were inoculated a third time. Each volunteer was inoculated at 3 sites on the upper arm with live H. ducreyi 35000HP (a human passaged isolate of strain 35000) and at 1 site with sterile PBS. Forty-eight hours after inoculation, lesion size was measured and RNA was isolated from the infected site that had the largest diameter. RNA was also obtained from the PBS control site (uninfected site). We obtained peripheral blood 6 to 12 months after inoculation of the 6th subject and derived myeloid DC in vitro. In brief, CD14+ peripheral blood monocytes were isolated from PBMC by positive selection magnetic beads (Miltenyi Biotec, Auburn, CA) and grown in the presence of recombinant human (rh) IL-4 (1 ng/ml) and rhGM-CSF (0.2 ng/ml) (R&D Systems, Minneapolis, MN). The cells were HLA-DR+, CD86+ CD40+, CD3-, CD14-, and CD19- by flow cytometry. The experiment was done in pairs so that DC from 1 RR subject were exposed to the same inoculum as DC from 1 PP subject. DC were incubated with nonopsonized H. ducreyi for 90 minutes at an MOI of 30:1. The DC were washed to remove non-associated bacteria, and incubated an additional 22.5 hours. Cells were collected and used for microarray analysis. Supernatants were collected and analyzed for cytokines using the Human Th1/Th2 II Cytometric Bead Array Kit per manufacturerâs instructions (BD Biosciences).

Dataset Information

H. ducreyi 35000HP grown in the presence or absence of FCS

Publications

Regulation of expression of the Haemophilus ducreyi LspB and LspA2 proteins by CpxR.

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