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Infection and Immunity, September 2008, p. 4046-4054, Vol. 76, No. 9
0019-9567/08/$08.00+0     doi:10.1128/IAI.00283-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

A Listeria monocytogenes Mutant Defective in Bacteriophage Attachment Is Attenuated in Orally Inoculated Mice and Impaired in Enterocyte Intracellular Growth{triangledown}

Patricia A. Spears,1 M. Mitsu Suyemoto,1 Angela M. Palermo,1 John R. Horton,1 Terri S. Hamrick,2 Edward A. Havell,1 and Paul E. Orndorff1*

Department of Population Health and Pathobiology, College of Veterinary Medicine, North Carolina State University, Raleigh, North Carolina 27606,1 Department of Pharmaceutical Sciences, School of Pharmacy, Campbell University, Buies Creek, North Carolina 275602

Received 29 February 2008/ Returned for modification 9 April 2008/ Accepted 9 June 2008

A Listeria monocytogenes bacteriophage was used to identify a phage-resistant Tn917 insertion mutant of the mouse-virulent listerial strain F6214-1. The mutant was attenuated when it was inoculated orally into female A/J mice and failed to replicate efficiently in cultured mouse enterocytes. Phage binding studies indicated that the mutant had a cell surface alteration that precluded phage attachment. All phenotypes associated with the mutation could be complemented in trans by a single open reading frame (ORF) that corresponded to the ORF interrupted by the Tn917 insertion. The complementation effected was, in all cases, at a level indistinguishable from that of the parent. The Tn917 insertion interrupted a gene that is predicted to encode a group 2 glycosyl transferase (provisionally designated glcV). A similar glcV gene is present in Listeria welshimeri and Listeria innocua and in some serotypes of L. monocytogenes. We speculate that the loss of the glcV product results in a defective phage receptor and that this alteration coincidentally influences a feature of the normal host-pathogen interaction required for virulence. Interestingly, the glcV lesion, while preventing phage attachment, did not alter the mutant's ability to bind to cultured mouse enterocyte monolayers. Rather, the mutation appeared to alter a subsequent step in intracellular replication measured by a reduction in plaque-forming efficiency and plaque size. In vivo, the mutant was undetectable in the liver and spleen 48 h after oral inoculation. The mutation is significant in part because it is one of the few that produce attenuation when the mutant is delivered orally.

* Corresponding author. Mailing address: College of Veterinary Medicine, North Carolina State University, 4700 Hillsborough Street, Raleigh, NC 27606. Phone: (919) 513-7019. Fax: (919) 515-3044. E-mail: Paul_Orndorff{at}ncsu.edu

{triangledown} Published ahead of print on 16 June 2008.

Editor: V. J. DiRita

Infection and Immunity, September 2008, p. 4046-4054, Vol. 76, No. 9
0019-9567/08/$08.00+0     doi:10.1128/IAI.00283-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.





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