Detection and species identification of Leishmania DNA from filter paper lesion impressions for patients with American cutaneous leishmaniasis.
BACKGROUND
Traditional detection of Leishmania from ulcers involves collection of invasive specimens that cause discomfort, require technical expertise, and carry risks of invasive procedures. We compared traditional diagnostic methods with a molecular noninvasive filter paper-based method for the diagnosis of cutaneous leishmaniasis.


METHODS
Consecutive patients presenting to the Leishmania Clinic at Hospital Nacional Cayetano Heredia were enrolled. Polymerase chain reaction (PCR) was performed on lesion scrapings, aspirates, and filter paper impressions. The reference standard was any 2 of 5 tests positive: smear, aspirate culture, invasive-specimen PCR (scrapings and aspirates), filter paper PCR, and leishmanin skin test. Outcome measures were sensitivity and specificity. Leishmania speciation was performed by PCR-restriction fragment length polymorphism (RFLP) of positive specimens.


RESULTS
Forty-five patients with 66 lesions were enrolled. Of 52 lesions diagnosed as cutaneous leishmaniasis, 50 were positive by PCR of invasive specimens versus 48 by PCR of filter papers (P=.930). Sensitivity and specificity of PCR on invasively obtained specimens were 94.2% (95% confidence interval [CI], 87.9%-100%) and 92.9% (95% CI, 79.4%-100%). Sensitivity and specificity of filter paper PCR were 92.3% (95% CI, 85.1%-99.5%) and 100%. Culture, smear, and leishmanin skin test all had inferior sensitivities, compared with PCR of invasive or noninvasive specimens (P<.001). Of 50 specimens positive by PCR, 19 had sufficient DNA for PCR-RFLP analysis.


CONCLUSIONS
Filter paper PCR constitutes a sensitive and specific alternative to traditional diagnostic assays. This novel, rapid, well-tolerated method has the potential for widespread use in the field and in pediatric populations where traditional specimen collection is most difficult to perform, and can potentially be used for rapid species identification.
Boggild
Andrea K
AK
Tropical Disease Unit, Division of Infectious Diseases, Toronto General Hospital, Toronto, Canada. andrea.boggild@utoronto.ca
Valencia
Braulio Mark
BM
Espinosa
Diego
D
Veland
Nicolas
N
Ramos
Ana Pilar
AP
Arevalo
Jorge
J
Llanos-Cuentas
Alejandro
A
Low
Donald E
DE
D000293:Adolescent; D000368:Aged; D002648:Child; D002675:Child, Preschool; D004247:DNA / Q000032:analysis* / Q000235:genetics; D005260:Female; D006801:Humans; D007891:Leishmania / Q000145:classification* / Q000235:genetics; D016773:Leishmaniasis, Cutaneous / Q000175:diagnosis / Q000469:parasitology*; D008297:Male; D008875:Middle Aged; D010568:Peru; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D012867:Skin / Q000469:parasitology; D013045:Species Specificity; D013048:Specimen Handling / Q000379:methods
D004247:DNA