Experimental infection of Phlebotomus perniciosus by bioluminescent Leishmania infantum using murine model and artificial feeder

Leishmaniasis is a vector-borne disease that is transmitted by sandflies and caused by obligate intracellular protozoa of the genus Leishmania. In the present study, we carried out a screening on the experimental infection of Phlebotomus pernioucus by bioluminescent Leishmania infantum using murine model and artificial feeder. We developed a real-time polymerase chain reaction (RT-PCR)-based method to determine individually the number of Leishmania promastigotes fed by infected flies. Among 1840 new emerged female sand flies, 428 were fed on the infected mice. After their death, they were analysed individually by RT-PCR. Our results demonstrated just a single Leishmania positive female at sixth day post meal. A total of 1070 female sand flies were exposed in contact with artificial feeder containing the human blood with two different quantities of Leishmania parasites: 2.106/mL and 1.107/mL. A blood meal including 1.107/mL LUC-promastigotes was proposed to 270 females and 75 (28%) flies were engorged. Among them, 44 (59%) were positive by RT-PCR analysis, with a relative average of 50551 Leishmania parasites. In case of blood feeding of females with 2.106/mL promastigotes, 57 out of 800 (7%) females succeed to feed from artificial feeder which 22 (39%) were positive with a relative average of 6487 parasites.

How to increase the population of a Phlebotomus perniciosus (Diptera: Psychodidae) colony: a new method

The sandfly Phlebotomus perniciosus is the most widespread vector of Leishmania infantum in Spain. Laboratory colonisation represents the most feasible source of information on the biology of these insects, but in conducting any study, the density of individuals in the colony may drop to such an extent that it is sometimes difficult to recover the initial population levels. A new technique was tested for the recovery of sandfly eggs in three different colonies; the recovery rate was studied by comparing the standard method of mass rearing with this new method of colony management. The results demonstrate a mean increase of 18.4 percent in adult production, a growth in colony productivity that justifies the inclusion of this process in the routine maintenance of any colony of sandflies.

Molecular detection of Leishmania infantum in naturally infected Phlebotomus perniciosus from Algarve Region, Portugal.

Background & objectives: In Portugal, Phlebotomus perniciosus and P. ariasi, (Subgenus Larroussius; Diptera: Psychodidae) are the proven vectors of leishmaniasis caused by Leishmania infantum. The Algarve Region in southern Portugal has been considered an endemic focus of leishmaniasis since 1980s. The main objective of the present study was to validate a molecular approach to detect Leishmania infection in phlebotomines based on DNA extraction from the female sandfly whole body, minus genitalia, followed by PCR for application on epidemiological surveys. Methods: In Algarve Region, from early May until early November 2006, sandflies were captured by CDC miniature light-traps. kDNA-PCR and ITS1-PCR were used to screen the presence of Leishmania DNA in female sandflies after species identification by entomological keys. Results: A total of 474 sandflies were collected in 108 biotopes. One female of P. perniciosus, the predominant species, was found infected with L. infantum reflecting an overall infection rate of 0.47%. Interpretation & conclusion: PCR associated with morphological characterization of the sandflies will be a powerful epidemiological tool for the determination of the number of phlebotomines infected with Leishmania spp in nature. In addition, the simultaneous occurrence of dogs and P. perniciosus infected with L. infantum shows that Algarve continues to be an endemic focus of canine leishmaniasis. Furthermore, as P. sergenti and P. papatasi which transmit L. tropica and L. major, respectively were present, the future introduction of these two Leishmania species in southern region of Portugal should not be neglected.