Nested polymerase chain reaction for diagnosis and monitoring treatment response in AIDS patients with tuberculous meningitis.
OBJECTIVE
To investigate the usefulness of polymerase chain reaction (PCR) from cerebrospinal fluid (CSF) for rapid diagnosis and assessing treatment response of tuberculous meningitis (TBM) in AIDS patients.


METHODS
Forty-four CSF samples from 10 patients with TBM confirmed by autopsy or by a culture of CSF (41 samples) and from two patients with highly probable TBM (three samples) were analysed. CSF specimens were collected before and during standard antituberculous treatment. CSF samples from 24 AIDS patients with autopsy evidence of other neurologic diseases were studied as controls.


METHODS
A nested PCR amplifying a 123 base-pair fragment of the IS6110 sequence was developed. Heating to 95 degrees C for 15 min was used for pre-PCR treatment of samples.


RESULTS
Detection limit was 10(2) colony-forming units per ml or 10 fg purified Mycobacterium tuberculosis DNA. M.tuberculosis DNA was detected in CSF from all the 12 confirmed or highly probable TBM cases. CSF was positive by nested PCR in 17 of 17 (100%) and 18 of 27 (67%) samples collected before and during therapy, respectively. Clinical and microbiological follow-up > or = 2 weeks was available for seven patients. PCR-positive CSF converted to M. tuberculosis DNA negative in four patients that showed improvement during treatment, but it remained positive in three patients who died of disseminated tuberculosis. All the CSF samples from the non-TBM controls were negative by nested PCR.


CONCLUSIONS
Nested PCR for detection of M. tuberculosis DNA is specific for diagnosis of TBM and more sensitive than conventional bacteriology. Moreover, nested PCR could be a useful method for assessing treatment response in AIDS patients with TBM.
Scarpellini
P
P
Infectious Diseases Department, San Raffaele Scientific Institute, Milan, Italy.
Racca
S
S
Cinque
P
P
Delfanti
F
F
Gianotti
N
N
Terreni
M R
MR
Vago
L
L
Lazzarin
A
A
D017088:AIDS-Related Opportunistic Infections / Q000175:diagnosis* / Q000382:microbiology; D001483:Base Sequence; D017931:DNA Primers / Q000235:genetics; D004269:DNA, Bacterial / Q000134:cerebrospinal fluid / Q000235:genetics; D005069:Evaluation Studies as Topic; D006801:Humans; D008969:Molecular Sequence Data; D009169:Mycobacterium tuberculosis / Q000235:genetics / Q000302:isolation & purification; D016133:Polymerase Chain Reaction / Q000379:methods* / Q000706:statistics & numerical data; D012680:Sensitivity and Specificity; D014390:Tuberculosis, Meningeal / Q000150:complications* / Q000175:diagnosis* / Q000382:microbiology
D017931:DNA Primers; D004269:DNA, Bacterial