Abstract
Leprosy is an infectious disease caused by Mycobacterium leprae. The polymerase chain reaction (PCR) has been applied to detect M. leprae in different clinical samples and urine seems to be attractive for this purpose. PCR was used to improve the sensitivity for diagnosing leprosy by amplifying a 151-bp PCR fragment of the M. leprae pra gene (PCR-Pra) in urine samples. Seventy-three leprosy patients (39 males and 34 females, 14 to 78 years old) were selected for leprosy diagnosis at a reference laboratory in Maringá, PR, Brazil. Of these, 36 were under anti-leprosy multidrug therapy with dapsone and rifampicin for tuberculoid (TT) and dapsone, rifampicin and clofazimine for borderline (BB) and lepromatous (LL) forms. The control group contained 50 healthy individuals without any clinical history of leprosy. DNA isolated from leprosy patients' urine samples was successfully amplified by PCR-Pra in 46.6% (34/73) of the cases. The positivity of PCR-Pra for patients with the TT form was 75% for both patients under treatment and non-treated patients (P = 0.1306). In patients with the LL form, PCR-Pra positivity was 52 and 30% for patients under treatment and non-treated patients, respectively (P = 0.2386). PCR-Pra showed a statistically significant difference in detecting M. leprae between the TT and LL forms of leprosy in patients under treatment (P = 0.0033). Although the current study showed that the proposed PCR-Pra has some limitations in the detection of M. leprae, this method has the potential to be a useful tool for leprosy diagnosis mainly in TT leprosy where the AFB slit-skin smear is always negative.
Highlights
The polymerase chain reaction (PCR) has been applied to detect Mycobacterium leprae in different clinical samples, such as slit-skin smear [1], blood [2], nasal cavity [3,4], skin tissues [2,5], and urine [6] for the improvement of the laboratory diagnosis of leprosy
Parkash et al [6] detected M. leprae DNA in the urine of leprosy patients when they used primers proposed by Hartskeerl et al [7], which amplified a large fragment of DNA (530 bp) of the pra encoding gene, even though the sample studied had a limited number of individuals
As small fragments of bacterial DNA may be found in patient urine samples [10], the use of this kind of clinical samples and primer set to detect a smaller fragment of M. leprae should be better evaluated since the amplification of a smaller DNA fragment may improve the sensitivity of the PCR assay, as pointed out by Goulart et al [12]
Summary
The polymerase chain reaction (PCR) has been applied to detect Mycobacterium leprae in different clinical samples, such as slit-skin smear [1], blood [2], nasal cavity [3,4], skin tissues [2,5], and urine [6] for the improvement of the laboratory diagnosis of leprosy. Some studies have reported the presence of mycobacterial DNA in urine [6,11], a fact that makes this clinical specimen attractive for the detection of M. leprae DNA. This is due to the fact that urine samples are collected and enhance patient adherence when compared to the distressing and traumatic slit-skin smear. Parkash et al [6] detected M. leprae DNA in the urine of leprosy patients when they used primers proposed by Hartskeerl et al [7], which amplified a large fragment of DNA (530 bp) of the pra encoding gene, even though the sample studied had a limited number of individuals. As small fragments of bacterial DNA may be found in patient urine samples [10], the use of this kind of clinical samples and primer set to detect a smaller fragment of M. leprae should be better evaluated since the amplification of a smaller DNA fragment may improve the sensitivity of the PCR assay, as pointed out by Goulart et al [12]
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