Methods and compositions for detecting serotypes of
Disclosed are methods and compositions for conducting assays utilizing real-time polymerase chain reactions (“PCRs”) in detection of serotypes L I, L II, and L III, but not stereotype B, of Chlamydia trachomatis , capable of causing lymphogranuloma venereum (“LGV”). These assays take advantage of a deletion occurring in the cytotoxin gene locus specific to the L I, L II, and L III serotypes. Each assay employs a first primer having a nucleotide sequence flanking one side of the deletion point and a second primer having a nucleotide sequence flanking the other side of the deletion point, wherein the first primer and the second primer are capable of hybridizing respectively to the plus strand and the minus strand of the genome of Chlamydia trachomatis during PCR. Synthesis during PCR of a sequence-specific amplicon containing this deletion point indicates that the sample contains nucleic acid specific to an LGV-causing serotype of Chlamydia trachomatis.
1. A method for detecting lymphogranuloma venereum-causing serotypes LI, LII, LIII, but not serotype B of Chlamydia trachomatis in a sample from a subject suspected of Chlamydia trachomatis infection, comprising the steps of:
(a) obtaining a sample from a subject suspected of Chlamydia trachomatis infection;
(b) isolating nucleic acid from said sample;
(c) mixing said isolated nucleic acid with a first primer and a second primer to form a mixture, said first primer consists of the nucleotide sequence of SEQ ID NO: 25 and said second primer consists of the nucleotide sequence of SEQ ID NO: 26;
(d) performing a PCR on said isolated nucleic acid mixture in step (c) under conditions that allow production of an amplicon; and
(e) detecting the presence of said amplicon,
wherein said presence of said amplicon is indicative of the presence of lymphogranuloma venereum-causing serotypes LI, LII, LIII, but not serotype B of Chlamydia trachomatis in said subject.
2. The method of claim 1 , wherein said PCR in step (d) is a real-time PCR.
3. The method of claim 2 , wherein said step (d) further comprises the step of using an oligonucleotide probe to detect said amplicon.
4. The method of claim 3 , wherein said oligonucleotide probe consists of the nucleotide sequence of SEQ ID NO: 27.
5. The method of claim 3 , wherein said oligonucleotide probe comprises a fluorescent moiety.
6. The method of claim 4 , wherein said fluorescent moiety is a 6-carboxy-fluorescein moiety.
7. The method of claim 1 , wherein said nucleic acid is DNA.