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C. trachomatis
was detected by PCR and LCR in rectal samples processed using a
simple protocol.
Pilot study of
COBAS PCR and ligase chain reaction for detection of rectal
infections due to Chlamydia trachomatis.
Golden MR, Astete SG, Galvan R,
Lucchetti A, Sanchez J, Celum CL, Whittington WLH, Stamm WE, Holmes
KK, Totten PA.
Journal of Clinical
Microbiology 2003;41:2174-2175.
Summary:
Question
What is the agreement between PCR and LCR for the detection of C.
trachomatis in rectal samples from men who have sex with men? How do
different specimen processing methods affect the results?
Design
This study describes a comparison of the results of PCR, performed on
samples processed using 3 procedures, and LCR for the detection of C.
trachomatis in rectal specimens collected from men who have sex with
men (MSM).
Participants
One hundred eighty-six men participating in a study of MSM in Lima, Peru,
were tested.
Description of Tests and Diagnostic
Standard
Anorectal specimens were collected, placed into 2SP medium, and frozen
until testing. Specimen processing for PCR (Cobas PCR, Roche Diagnostics
Systems, Branchburg, NJ), performed according to the manufacturer's
instructions, included: 1) diluting 50 μl in lysis buffer and diluent
as described in the Roche CT/NG sample treatment kit for swab specimens,
2) centrifuging 125 μl and treating the specimen as described in the
Roche CT/NG sample treatment kit for urine specimens, and 3) treating 150
μl with the MasterPure DNA Purification kit (Epicenter, Madison, WI)
following the manufacturer's instructions, except that the pellet was
suspended in 30 μl of TE buffer. The amounts of original sample added
to each PCR were 12.5, 50, and 25 μl for methods 1, 2, and 3,
respectively. Specimen were processed for LCR (Abbott Laboratories, Abbott
Park, IL), performed according to the manufacturer's instructions, by
centrifuging 120 μl and suspending the pellet in the same volume of
urine suspension buffer (Abbott Laboratories). One hundred μl were
added to each LCR assay. Specimens were diluted 1:6 to confirm initial
results.
All specimens producing concordant
positive, all discordant, and 17 concordant negative samples were purified
using the MasterPure method and retested by PCR and LCR.
Main Outcome Measures
The results of the PCR, by specimen processing method, and LCR are
compared.
Main Results
The results of the initial PCR assays performed on samples processed by 3
methods and the initial LCR assay are shown in the table. Two of 2 samples
positive by all 3 PCR assays and negative by LCR, and 2 of 5 samples
positive by one PCR and negative by LCR were positive on retesting by LCR
after DNA purification using the MasterPure method. Agreement between PCR
and LCR was 178 of 184 (98%).
Authors' Conclusions
Given the low frequency of inhibition, the
high level of agreement between PCR and LCR on specimens processed using
the swab procedure, and the relative ease and low cost of this procedure,
the Roche swab processing procedure is recommended when testing rectal
specimens by PCR.
Source of funding: Roche Diagnostics
and the University of Washington NIH STD Cooperative Research Center.
For correspondence: Matthew R. Golden,
Harborview Medical Center Box 359777, 325 Ninth Ave., Seattle, WA 98104.
E-mail address: golden@u.washington.edu.
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