Performance of nested PCR in the specific detection of Mycobacterium tuberculosis complex in blood samples of pediatric patients.
OBJECTIVE
To evaluate the performance of nested PCR (nPCR) in detecting the Mycobacterium tuberculosis complex in blood samples of patients suspected of having TB, in order to determine its potential for use as an auxiliary tool in the laboratory diagnosis of TB in children.


METHODS
Detection of the M. tuberculosis complex in blood samples using as a target the insertion sequence IS6110 of the genomic DNA of the bacillus. Blood samples of 120 patients were evaluated. All of the patients were under 15 years of age at the time of their treatment at public hospitals in the city of Recife, Brazil (between January of 2003 and August of 2005). Attending physicians at the hospitals diagnosed TB based on the criteria recommended by the American Thoracic Society. The nPCR amplified a 123-bp fragment with outer oligonucleotides (IS1/IS2) and, in the subsequent reaction, using inner oligonucleotides (IS3/IS4), generating an 81-bp amplicon.


RESULTS
Active or latent TB was found in 65 patients, TB was ruled out in 28 suspected cases, and 27 patients were TB-free (controls). The sensitivity of nPCR was 26.15% and was significantly higher for the extrapulmonary form of the disease (55.56%) than for the pulmonary form (18.18%). The specificity was 92.73%.


CONCLUSIONS
Despite the difficulties in diagnosing TB in children and the low number of cases evaluated in the present study, nPCR in blood samples proved to be a rapid and specific technique, albeit one with low sensitivity. In order to establish its true usefulness in the diagnosis of paucibacillary forms, especially extrapulmonary TB, further studies need to be carried out with a larger sample of children and analyzing biological specimens other than blood.
Lima
Juliana Figueirêdo da Costa
JF
Centro de Pesquisas Aggeu Magalhães, Fundação Oswaldo Cruz, Recife, PE, Brasil. jfcl@cpqam.fiocruz.br
Montenegro
Lílian Maria Lapa
LM
Montenegro
Rosana de Albuquerque
Rde A
Cabral
Marta Maciel Lyra
MM
Lima
Andrea Santos
AS
Abath
Frederico Guilherme Coutinho
FG
Schindler
Haiana Charifker
HC
D000293:Adolescent; D016022:Case-Control Studies; D002648:Child; D005188:False Negative Reactions; D005189:False Positive Reactions; D005260:Female; D006801:Humans; D055985:Latent Tuberculosis / Q000097:blood / Q000175:diagnosis* / Q000382:microbiology; D008297:Male; D009169:Mycobacterium tuberculosis / Q000235:genetics*; D016133:Polymerase Chain Reaction / Q000592:standards*; D011446:Prospective Studies; D012680:Sensitivity and Specificity; D014397:Tuberculosis, Pulmonary / Q000097:blood / Q000175:diagnosis* / Q000382:microbiology