The 7.5-kb plasmid of Chlamydia trachomatis (CT) is believed to encode essential genes and might have a role in CT pathogenicity. Accordingly, analysis of plasmid-linked mutations in isolates from biovars with different pathogenic properties should help in identifying which plasmid-encoded genes, if any, may be involved in modulating virulence. For this purpose, the plasmid present in a low-virulence isolate (trachoma biovar, serotype D) was cloned and sequenced. Nucleotide changes were experimentally checked against the sequence of the plasmid variant from the highly virulent strain L2/434/Bu (LGV biovar). By aligning our data with two published sequences of different trachoma and LGV variants a general consensus structure was determined, comprising eight major open reading frames (ORF) and a number of points where there is consensus only between isolates of the same biovar (biovar-specific mutations). The degree of variation between different isolates is less than 1%. In particular, comparison of serotype-D and -L2 plasmids shows mutations which are generally silent or lead to few (one to four), often conservative, amino acid changes in ORFs 1, 2, 4, 5, 6, and 7. The protein encoded by ORF8 is completely conserved. In contrast, the polypeptide variants encoded by ORF3 show nine amino acid changes, seven of which are due to biovar-specific mutations.