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Journal of Bacteriology, September 2006, p. 6335-6345, Vol. 188, No. 17
0021-9193/06/$08.00+0     doi:10.1128/JB.00698-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Transposon Mutagenesis Identifies Genes Associated with Mycoplasma pneumoniae Gliding Motility

Benjamin M. Hasselbring, Clinton A. Page, Edward S. Sheppard, and Duncan C. Krause*

Department of Microbiology, University of Georgia, Athens, Georgia

Received 16 May 2006/ Accepted 13 June 2006

The wall-less prokaryote Mycoplasma pneumoniae, a common cause of chronic respiratory tract infections in humans, is considered to be among the smallest and simplest known cells capable of self-replication, yet it has a complex architecture with a novel cytoskeleton and a differentiated terminal organelle that function in adherence, cell division, and gliding motility. Recent findings have begun to elucidate the hierarchy of protein interactions required for terminal organelle assembly, but the engineering of its gliding machinery is largely unknown. In the current study, we assessed gliding in cytadherence mutants lacking terminal organelle proteins B, C, P1, and HMW1. Furthermore, we screened over 3,500 M. pneumoniae transposon mutants individually to identify genes associated with gliding but dispensable for cytadherence. Forty-seven transformants having motility defects were characterized further, with transposon insertions mapping to 32 different open reading frames widely distributed throughout the M. pneumoniae genome; 30 of these were dispensable for cytadherence. We confirmed the clonality of selected transformants by Southern blot hybridization and PCR analysis and characterized satellite growth and gliding by microcinematography. For some mutants, satellite growth was absent or developed more slowly than that of the wild type. Others produced lawn-like growth largely devoid of typical microcolonies, while still others had a dull, asymmetrical leading edge or a filamentous appearance of colony spreading. All mutants exhibited substantially reduced gliding velocities and/or frequencies. These findings significantly expand our understanding of the complexity of M. pneumoniae gliding and the identity of possible elements of the gliding machinery, providing a foundation for a detailed analysis of the engineering and regulation of motility in this unusual prokaryote.

* Corresponding author. Mailing address: Department of Microbiology, 523 Biological Sciences Building, University of Georgia, Athens, GA 30602. Phone: (706) 542-2671. Fax: (706) 542-2674. E-mail: dkrause{at}uga.edu.

Journal of Bacteriology, September 2006, p. 6335-6345, Vol. 188, No. 17
0021-9193/06/$08.00+0     doi:10.1128/JB.00698-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.



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