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Characterisation of an acapsular mutant of Burkholderia pseudomallei identified by signature tagged mutagenesis

TIMOTHY ATKINS, RICHARD PRIOR, KERRI MACK, PAUL RUSSELL, MICHELLE NELSON, JOANN PRIOR, JILL ELLIS, PETRA C. F. OYSTON, GORDON DOUGAN^* and RICHARD W. TITBALL

Defence Science and Technology Laboratory, CBS Porton Down, Salisbury, Wiltshire, SP4 OJQ, *Department of Biological Sciences, Centre for Molecular Microbiology and Infection, Flowers Building, Imperial College of Science Technology and Medicine, Exhibition Road, London, SW7 2AY and Department of Infectious and Tropical Diseases, London School of Hygiene and Tropical Medicine, Keppel Street, London WC1E 7HT

Corresponding author: Dr T. Atkins (e-mail: TPATKINS{at}dstl.gov.uk).

Received 5 Nov. 2001; revised version accepted 21 Jan. 2002.

Abstract

A Burkholderia pseudomallei mutant which was attenuated in a mouse model of melioidosis was identified by a signature tagged mutagenesis approach. The transposon was shown to be inserted into a gene within the capsular biosynthetic operon. Compared with the wild-type bacteria this mutant demonstrated a 10⁵-fold increase in the median lethal dose in a mouse model and it did not react with a monoclonal antibody against high mol. wt polysaccharide of B. pseudomallei. To determine the kinetics of infection, mice were dosed intraperitoneally (i.p.) and intravenously (i.v.) with mutant and wild-type bacteria. After i.p challenge, the number of mutant bacteria in the peritoneal cavity declined, whereas wild-type bacteria proliferated. When administered by the i.v. route, the mutant was able to cause disease but the time to death was increased compared with the wild type. Mice were dosed with the mutant and subsequently challenged with wild-type B. pseudomallei, but the mutant failed to induce a protective immune response.

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