is the primary etiological agent of aggressive periodontal disease. Identification of novel virulence factors at the genome-wide level is hindered by lack of efficient genetic tools to perform mutagenesis in this organism. The mariner transposon is known to yield a random distribution of insertions in an organism's genome with requirement for only a TA dinucleotide target and is independent of host-specific factors. However, the utility of this system in is unknown. In this study, we found that transposon mutagenesis occurs at a high frequency (×10 ), and can be universally applied to wild-type strains of serotypes a, b, and c. The transposon inserts were stably inherited in transconjugants in the absence of antibiotics. A library of 16,000 mutant colonies of was screened for reduced biofilm formation. Mutants with transposon inserts in genes encoding pilus, putative ion transporters, multidrug resistant proteins, transcription regulators and enzymes involved in the synthesis of extracellular polymeric substance, bacterial metabolism and stress response were discovered in this screen. Our results demonstrated the utility of the mutagenesis system as a novel genetic tool for functional genomic analysis in .