Development of a PCR-based method for diagnosis of Leishmania in blood samples.
Visceral Leishmaniasis (VL) cases reported in Mediterranean countries and in Northern Europe are becoming increasingly frequent. The past few years several studies have shown Polymerase Chain Reaction to be more effective than the classical methods for the diagnosis of VL in clinical samples. The purpose of this study was the development of a simple, specific and sensitive PCR-based assay for the detection of Leishmania in blood samples. A specific pair of oligonucleotides was designed using conserved sequences of the ssu-rRNA Leishmania infantum gene. Of the 53 blood samples of patients suspected for leishmaniasis that were processed with the newly designed oligonucleotides, 13 were successfully diagnosed positive. The results were confirmed with sequencing and restriction fragment length polymorphism. The lower detection limit of the reported assay was 10 parasites per ml in all seeded samples tested and considered highly satisfactory for diagnosis of Leishmaniasis in blood samples.
Spanakos
Gregory
G
Department of Public and Administrative Health, National School of Public Health, 196 Alexandras Ave, Athens, Greece. grspan@yahoo.com
Patsoula
Eleni
E
Kremastinou
Tzeni
T
Saroglou
George
G
Vakalis
Nicholas
N
D000818:Animals; D017124:Conserved Sequence; D015342:DNA Probes; D016054:DNA, Protozoan / Q000097:blood; D006801:Humans; D018314:Leishmania infantum / Q000235:genetics / Q000302:isolation & purification*; D007896:Leishmaniasis / Q000175:diagnosis; D016133:Polymerase Chain Reaction / Q000379:methods*; D012335:RNA, Ribosomal / Q000235:genetics; D012680:Sensitivity and Specificity
D015342:DNA Probes; D016054:DNA, Protozoan; D012335:RNA, Ribosomal