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Infection and Immunity, April 2009, p. 1293-1303, Vol. 77, No. 4
0019-9567/09/$08.00+0     doi:10.1128/IAI.01181-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Pseudomonas aeruginosa Infection of Zebrafish Involves both Host and Pathogen Determinants

Anne E. Clatworthy,^1,2,3 Jenny See-Wai Lee,^1,2,3 Mark Leibman,^1,2,3 Zachary Kostun,^4,5 Alan J. Davidson,^4,5 and Deborah T. Hung^1,2,3,5*

Department of Molecular Biology and Center for Computational and Integrative Biology, Massachusetts General Hospital, 185 Cambridge St., Boston, Massachusetts 02114,¹ Department of Microbiology and Molecular Genetics, Harvard Medical School, Boston, Massachusetts 02115,² Broad Institute of MIT and Harvard, 7 Cambridge Center, Cambridge, Massachusetts 02142,³ Center for Regenerative Medicine, Massachusetts General Hospital, 185 Cambridge St., Boston, Massachusetts 02114,⁴ Department of Medicine, Harvard Medical School, Boston, Massachusetts 02114⁵

Received 23 September 2008/ Returned for modification 9 November 2008/ Accepted 12 January 2009

Zebrafish (Danio rerio) have a number of strengths as a host model for infection, including genetic tractability, a vertebrate immune system similar to that of mammals, ease and scale of laboratory handling, which allows analysis with reasonable throughput, and transparency, which facilitates visualization of the infection. With these advantages in mind, we examined whether zebrafish could be used to study Pseudomonas aeruginosa pathogenesis and found that infection of zebrafish embryos with live P. aeruginosa (PA14 or PAO1) by microinjection results in embryonic death, unlike infection with Escherichia coli or heat-killed P. aeruginosa, which has no effect. Similar to studies with mice, P. aeruginosa mutants deficient in type three secretion (pscD) or quorum sensing (lasR and mvfR) are attenuated in zebrafish embryos infected at 50 h postfertilization (hpf), a developmental stage when both macrophages and neutrophils are present. In contrast, embryos infected at 28 hpf, when only macrophages are initially present, succumb to lethal challenge with far fewer P. aeruginosa cells than those required for embryos infected at 50 hpf, are susceptible to infection with lasR and pscD deletion mutants, and are moderately resistant to infection with an mvfR mutant. Finally, we show that we can control the outcome of infection through the use of morpholinos, which allow us to shift immune cell numbers, or small molecules (antibiotics), which rescue embryos from lethal challenge. Thus, zebrafish are a novel host model that is well suited for studying the interactions among individual pathogenic functions of P. aeruginosa, the role of individual components of host immune defense, and small-molecule modulators of infection.

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* Corresponding author. Mailing address: Department of Molecular Biology and Center for Computational and Integrative Biology, Massachusetts General Hospital, 185 Cambridge St., Boston, MA 02114. Phone: (617) 643-3117. Fax: (617) 726-6893. E-mail: hung{at}molbio.mgh.harvard.edu

Published ahead of print on 21 January 2009.

Editor: S. R. Blanke

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Infection and Immunity, April 2009, p. 1293-1303, Vol. 77, No. 4
0019-9567/09/$08.00+0     doi:10.1128/IAI.01181-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.