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. 2015 Oct 9:11:258.
doi: 10.1186/s12917-015-0576-5.

Successful isolation of Leishmania infantum from Rhipicephalus sanguineus sensu lato (Acari: Ixodidae) collected from naturally infected dogs

Viviane Medeiros-Silva  1 , Rodrigo Gurgel-Gonçalves  2   3 , Nadjar Nitz  4 , Lucia Emilia D' Anduraim Morales  5 , Laurício Monteiro Cruz  5 , Isabele Gonçalves Sobral  5 , Mariana Côrtes Boité  6 , Gabriel Eduardo Melim Ferreira  7 , Elisa Cupolillo  8 , Gustavo Adolfo Sierra Romero  9   10
Affiliations

Successful isolation of Leishmania infantum from Rhipicephalus sanguineus sensu lato (Acari: Ixodidae) collected from naturally infected dogs

Viviane Medeiros-Silva et al. BMC Vet Res. .

Abstract

Background: The main transmission route of Leishmania infantum is through the bites of sand flies. However, alternative mechanisms are being investigated, such as through the bites of ticks, which could have epidemiological relevance. The objective of this work was to verify the presence of Leishmania spp. in Rhipicephalus sanguineus sensu lato collected from naturally infected dogs in the Federal District of Brazil.

Methods: Ticks were dissected to remove their intestines and salivary glands for DNA extraction and the subsequent amplification of the conserved region of 120 bp of kDNA and 234 bp of the hsp70 gene of Leishmania spp. The amplified kDNA products were digested with endonucleases HaeIII and BstUI and were submitted to DNA sequencing. Isolated Leishmania parasites from these ticks were analyzed by multilocus enzyme electrophoresis, and the DNA obtained from this culture was subjected to microsatellite analyses.

Results: Overall, 130 specimens of R. sanguineus were collected from 27 dogs. Leishmania spp. were successfully isolated in culture from five pools of salivary glands and the intestines of ticks collected from four dogs. The amplified kDNA products from the dog blood samples and from the tick cultures, when digested by HaeIII and BstUI, revealed the presence of L. braziliensis and L. infantum. One strain was cultivated and characterized as L. infantum by enzyme electrophoresis. The amplified kDNA products from the blood of one dog showed a sequence homology with L. braziliensis; however, the amplified kDNA from the ticks collected from this dog showed a sequence homology to L. infantum.

Conclusion: The results confirm that the specimens of R. sanguineus that feed on dogs naturally infected by L. infantum contain the parasite DNA in their intestines and salivary glands, and viable L. infantum can be successfully isolated from these ectoparasites.

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Figures

Fig. 1
Fig. 1
Sampling processing flow. SG: salivary gland; Int: intestine; Seq: sequencing; Ø: nymph. RFLP: restriction fragment lengh polymorphism. The numbers in the boxes show the number of samples/pools processed
Fig. 2
Fig. 2
Amplified 120pb kDNA products digested by the enzymes Hae III (a) and BstUI (b) in canine blood samples and cultures from the tick contents. Reference strain cultures of L. amazonensis (La), L. braziliensis (Lb), and L. infantum (Li); Cultures from R. sanguineus specimens obtained from dog I (c1), dog III (C3), and dog IV (c4); Blood samples of dog I (s1), dog II (s2), dog III (s3), dog IV (4); MP: molecular weight marker
See this image and copyright information in PMC

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