Gern, Lise

2007, Cadenas, Francisca Moran, Schneider, Helene, Lommano, Elena, Burri, Caroline, Moret, Jacqueline, Gern, Lise

We tested two approaches to extract Borrelia DNA from live Ixodes ricinus ticks before polymerase chain reaction (PCR) and reverse line blotting (RLB): DNA extraction of one half of the tick after incubation in BSK medium and DNA extraction of the other half of the tick directly, using ammonium hydroxide. Among 2079 ticks, 31.2% (n = 649) were found to be Borrelia-infected by PCR-RLB test using at least one of the DNA extraction methods. Five hundred four ticks (24.2%) were found infected after incubation in BSK and 481 (23.1%) after direct DNA extraction from the tick. The difference was not significant. However, these prevalences were significantly lower when only one method was applied (23.1% and 24.2%) compared to the prevalence obtained by the use of both methods (31.2%). In 313 infected ticks discordant results were obtained, i.e., one half of the tick was found to be infected whereas the other half was uninfected. Among these ticks, B. garinii and B. burgdorferi sensu stricto (ss) were significantly more frequently identified in the half tick incubated in BSK. No significant differences were observed for B. burgdorferi ss, B. valaisiana, and for undetermined Borrelia species.

2007, Humair, Pierre-François, Douet, Véronique, Cadenas, Francisca Moran, Schouls, Leo M, Van de Pol, Ingrid, Gern, Lise

We developed an efficient molecular method for the identification of the bloodmeal sources in the tick Ixodes ricinus (L.), the European vector of the agents of Lyme borreliosis and tick-borne encephalitis. A approximate to 145-bp orthologous fragment of the vertebrate mitochondrial 12S rDNA was used as a molecular marker to discriminate host vertebrate species. The method consists of a single run polymerase chain reaction amplification of the 12S rDNA molecular marker by using nondegenerate primers followed by a reverse line blot hybridization assay by using specific oligonucleotide probes. The palette of probes allowed us to distinguish major groups of host vertebrates (e.g., mammals, small rodents, artiodactyls, birds, lizards) and to identify the bloodmeal sources at the genus or species level. External primers were designed and used to sequence the 12S rDNA molecular marker of a broad range of known or potential host vertebrate species (n = 60), including mammal (n = 28), bird (n = 31), and reptile (n = 1) species. The use of this technique coupled with known methods for identification of tick-borne pathogens (e.g., Borrelia burgdorferi sensu lato) allowed us to determine the source of infective bloodmeal and to identify reservoir species. The present method was successfully used to identify the source of bloodmeals in all feeding I. ricinus ticks and in half of questing field-collected L ricinus ticks. Moreover, the bloodmeal source was identified in 65% of ticks infected with B. burgdorferi sensu lato. Further development of this technique may be envisaged for the detection of other vector-borne patbogens and their reservoir hosts.