Identification of blood meals in field captured sand flies by a PCR-RFLP approach based on cytochrome b gene.
Leishmaniasis is a vector-borne disease transmitted by phlebotomine sand flies. Information about blood meal preferences in sand flies is essential to understand the epidemiology of the disease to adopt control measures. In previous studies, a polymerase chain reaction (PCR) of 359bp fragment of the conserved gene cytochrome b (cyt b) and further sequencing were applied in the study of blood meal sources in sand flies collected in the area of a leishmaniasis outbreak in southwest Madrid, Spain, providing significant information about blood meal preferences in the focus. In this work, a PCR-restriction fragment length polymorphism (RFLP) targeting a fragment of 359bp of vertebrate cyt b gene was developed. Restriction endonucleases HaeIII and HinfI generated specific patterns consistent with the blood meal sources found in sand flies. The protocol has been validated with twenty six engorged females collected in the field with CDC traps. Blood meals from nine vertebrates were identified based on PCR-cyt b and sequencing-human, dog, cat, horse, hare, rabbit, sheep, goat and chicken - and mixed blood meals (sheep/human; sheep/goat) - and successfully distinguished by PCR-RFLP. Therefore, this approach is an efficient and reliable alternative method to be applied in entomological surveys.
Unidad de Entomología Médica, Servicio de Parasitología, Centro Nacional de Microbiología, Instituto de Salud Carlos III, Ctra. Majadahonda-Pozuelo s/n, 28220 Majadahonda, Madrid, Spain.
Estela
E
González
Laboratori de Parasitologia, Facultat de Farmàcia, Universitat de Barcelona (UB), Spain; ISGlobal, Barcelona Ctr. Int. Health Res. (CRESIB), Hospital Clínic-Universitat de Barcelona, Barcelona, Spain.
Montserrat
M
Gállego
Unidad de Entomología Médica, Servicio de Parasitología, Centro Nacional de Microbiología, Instituto de Salud Carlos III, Ctra. Majadahonda-Pozuelo s/n, 28220 Majadahonda, Madrid, Spain.
Ricardo
R
Molina
Laboratori de Parasitologia, Facultat de Farmàcia, Universitat de Barcelona (UB), Spain; ISGlobal, Barcelona Ctr. Int. Health Res. (CRESIB), Hospital Clínic-Universitat de Barcelona, Barcelona, Spain.
Alba
A
Abras
Laboratori de Parasitologia, Facultat de Farmàcia, Universitat de Barcelona (UB), Spain; ISGlobal, Barcelona Ctr. Int. Health Res. (CRESIB), Hospital Clínic-Universitat de Barcelona, Barcelona, Spain.
M Magdalena
MM
Alcover
Laboratori de Parasitologia, Facultat de Farmàcia, Universitat de Barcelona (UB), Spain; ISGlobal, Barcelona Ctr. Int. Health Res. (CRESIB), Hospital Clínic-Universitat de Barcelona, Barcelona, Spain.
Cristina
C
Ballart
Laboratori de Parasitologia, Facultat de Farmàcia, Universitat de Barcelona (UB), Spain.
Anna
A
Fernández
Unidad de Entomología Médica, Servicio de Parasitología, Centro Nacional de Microbiología, Instituto de Salud Carlos III, Ctra. Majadahonda-Pozuelo s/n, 28220 Majadahonda, Madrid, Spain. Electronic address: mjimenez@isciii.es.
Maribel
M
Jiménez
D000818:Animals; D001483:Base Sequence; D045303:Cytochromes b / Q000235:genetics*; D005260:Female; D006801:Humans; D007896:Leishmaniasis / Q000635:transmission*; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods*; D012150:Polymorphism, Restriction Fragment Length; D011576:Psychodidae / Q000235:genetics / Q000502:physiology*
D045303:Cytochromes b
PCR-RFLP; blood meal; cytochrome b; sand fly