Rapid diagnosis and genotyping of Leishmania isolates from cutaneous and visceral leishmaniasis by microcapillary cultivation and polymerase chain reaction-restriction fragment length polymorphism of miniexon region.
We have performed a combination of microcapillary cultivation method and restriction fragment length polymorphism (RFLP) analysis of amplified products by 1 single PCR of miniexon region of Leishmania for molecular diagnosis and genotyping of different Leishmania species isolated from cutaneous leishmaniasis (CL) and visceral leishmaniasis. We have analyzed 10 microcapillary cultivated isolates from cutaneous cases and 5 microcapillary cultivated isolates from visceral cases (totally 15) by polymerase chain reaction-RFLP (PCR-RFLP). Of 10 isolates, 3 (30%) were genotyped as Leishmania infantum and 7 (70%) of 10 isolates were genotyped as Leishmania tropica from the microcapillary cultivated isolates of cutaneous cases. On the other hand, all 5 isolates (100%) were genotyped as L. infantum from microcapillary cultivated visceral cases. Our most interesting finding is the presence of 3 L. infantum isolates in CL cases without kala-azar history. Therefore, we suggest that further investigations must be done about this subject. On the other hand, we suggest the combination of microcapillary culture method and PCR-RFLP of miniexon region of leishmaniae can be used in routine laboratory experimentation because of their simple, cheap, and rapid benefits (within a week), whereas other different approaches offer a multitude of valid taxonomic characters for species identification.
Serin
Mehmet S
MS
Department of Microbiology, Faculty of Pharmacy, Mersin University, 33343, Turkey. serinmss@cu.edu.tr
Daglioglu
Kenan
K
Bagirova
Melahat
M
Allahverdiyev
Adil
A
Uzun
Soner
S
Vural
Zeynep
Z
Kayar
Begum
B
Tezcan
Seda
S
Yetkin
Mesut
M
Aslan
Gonul
G
Emekdas
Gurol
G
Koksal
Fatih
F
D000818:Animals; D003470:Culture Media; D015252:Deoxyribonucleases, Type II Site-Specific / Q000378:metabolism; D005091:Exons / Q000235:genetics; D006801:Humans; D007891:Leishmania / Q000145:classification* / Q000235:genetics* / Q000302:isolation & purification; D016773:Leishmaniasis, Cutaneous / Q000175:diagnosis* / Q000469:parasitology; D007898:Leishmaniasis, Visceral / Q000175:diagnosis* / Q000469:parasitology; D010274:Parasitology / Q000295:instrumentation / Q000379:methods; D016133:Polymerase Chain Reaction / Q000379:methods*; D012150:Polymorphism, Restriction Fragment Length; D013997:Time Factors
D003470:Culture Media; D015252:Deoxyribonucleases, Type II Site-Specific; C037794:YGGCCR-specific type II deoxyribonucleases