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Infection and Immunity, November 2001, p. 7074-7082, Vol. 69, No. 11
0019-9567/01/$04.00+0   DOI: 10.1128/IAI.69.11.7074-7082.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

agr Expression Precedes Escape of Internalized Staphylococcus aureus from the Host Endosome

Saara N. A. Qazi,1,2 Emilie Counil,3 Julie Morrissey,2 Catherine E. D. Rees,1 Alan Cockayne,2 Klaus Winzer,2 Weng C. Chan,4 Paul Williams,2,4 and Philip J. Hill1,2,*

School of Biosciences, University of Nottingham, Loughborough, Leicestershire LE12 5RD,1 Institute of Infections and Immunity, Queens Medical Centre, University of Nottingham, Nottingham NG7 2UH,2 and School of Pharmaceutical Sciences, University of Nottingham, Nottingham NG7 2RD,4 United Kingdom, and Institut National Agronomique, Paris-Grignon, France3

Received 11 April 2001/Returned for modification 1 June 2001/Accepted 23 July 2001

Staphylococcus aureus is a versatile pathogen capable of causing life-threatening infections. Many of its cell wall and exoproduct virulence determinants are controlled via the accessory gene regulator (agr). Although considered primarily as an extracellular pathogen, it is now recognized that S. aureus can be internalized by epithelial and endothelial cells. Traditional experimental approaches to investigate bacterial internalization are extremely time-consuming and notoriously irreproducible. We present here a new reporter gene method to assess intracellular growth of S. aureus in MAC-T cells that utilizes a gfp-luxABCDE reporter operon under the control of the Bacillus megaterium xylA promoter, which in S. aureus is expressed in a growth-dependent manner. This facilitates assessment of the growth of internalized bacteria in a nondestructive assay. The dual gfp-lux reporter cassette was also evaluated as a reporter of agr expression and used to monitor the temporal induction of agr during the MAC-T internalization process. The data obtained suggest that agr induction occurs prior to endosomal lysis and that agr-regulated exoproteins appear to be required prior to the release and replication of S. aureus within the infected MAC-T cells.

* Corresponding author. Mailing address: University of Nottingham, School of Biosciences, Sutton Bonington Campus, Loughborough, Leicestershire LE12 5RD, United Kingdom. Phone: 44-115-951-6169. Fax: 44-115-951-6162. E-mail: phil.hill{at}nottingham.ac.uk.

Infection and Immunity, November 2001, p. 7074-7082, Vol. 69, No. 11
0019-9567/01/$04.00+0   DOI: 10.1128/IAI.69.11.7074-7082.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


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