Detection and discrimination of Loa loa, Mansonella perstans and Wuchereria bancrofti by PCR-RFLP and nested-PCR of ribosomal DNA ITS1 region.
The ribosomal deoxyribonucleic acid (DNA) internal transcribed spacer region (ITS1) of two filarial nematodes, Loa loa and Mansonella perstans, was amplified and further sequenced to develop an species-specific polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) protocol for the differentiation of both species from Wuchereria bancrofti, three filarial nematodes with blood circulating microfilariae. The ITS1-PCR product digested with the restriction endonuclease Ase I generated an specific diagnostic pattern for each of the three species. Moreover, three new specific nested-PCRs, targeting the ITS1 region, for differential detection of L. loa, M. perstans and W. bancrofti were developed and used when the ITS1-PCR products were insufficient for the Ase I enzymatic digestion. These filarial species-specific molecular protocols were evaluated in forty blood samples from African adult immigrants attending in the Hospital Insular of Gran Canaria, Canarias, Spain.
Jiménez
Maribel
M
Servicio de Parasitología, Centro Nacional de Microbiología, Instituto de Salud Carlos III, 28220 Majadahonda, Madrid, Spain. mjimenez@isciii.es
González
Luis Miguel
LM
Carranza
Cristina
C
Bailo
Begoña
B
Pérez-Ayala
Ana
A
Muro
Antonio
A
Pérez-Arellano
José Luis
JL
Gárate
Teresa
T
D000818:Animals; D003001:Cloning, Molecular; D018509:DNA, Helminth / Q000097:blood / Q000737:chemistry; D004275:DNA, Ribosomal / Q000737:chemistry; D003937:Diagnosis, Differential; D004605:Elephantiasis, Filarial / Q000175:diagnosis* / Q000469:parasitology; D006801:Humans; D008117:Loa / Q000235:genetics / Q000302:isolation & purification*; D008118:Loiasis / Q000175:diagnosis* / Q000469:parasitology; D008366:Mansonella / Q000235:genetics / Q000302:isolation & purification*; D008368:Mansonelliasis / Q000175:diagnosis* / Q000469:parasitology; D016133:Polymerase Chain Reaction / Q000379:methods; D012150:Polymorphism, Restriction Fragment Length; D018510:RNA, Helminth / Q000235:genetics; D012335:RNA, Ribosomal / Q000235:genetics; D016415:Sequence Alignment; D013045:Species Specificity; D014958:Wuchereria bancrofti / Q000235:genetics / Q000302:isolation & purification*
D018509:DNA, Helminth; D004275:DNA, Ribosomal; D018510:RNA, Helminth; D012335:RNA, Ribosomal