Thirty urogenital Chlamydia trachomatis isolates collected in Moscow in 2005 were typed using newly developed molecular typing approaches: (1) multilocus sequence typing (MLST(7)) based on sequences of seven housekeeping genes (http://pubmlst.org/chlamydiales/), (2) MLST(5) based on the investigation of five target regions of the chlamydial genome and (3) ompA gene sequencing supplemented with three variable number tandem repeat (VNTR) loci of the genome. ompA typing divided all isolates into 11 groups with E serotype dominating, while MLST7, MLST5 and VNTR analysis divided them into eight, 20 and 18 groups, respectively. The discriminatory power of each method calculated using the Hunter-Gaston discriminatory index was found to be 0.83 for the ompA typing scheme, 0.82 for MLST(7) and 0.95 for MLST(5). A novel sequence type combining 13% of all strains was discovered, as well as new alleles of genes. This is the first study characterizing the genetic diversity of the urogenital C. trachomatis population in Central Russia using MLST. We conclude that the MLST(7) scheme is the best possible choice for global epidemiological purposes, whereas MLST(5) is more appropriate for tracing local outbreaks.