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Influence of some climatic factors on Ixodes ricinus ticks studied along altitudinal gradients in two geographic regions in Switzerland
In the context of climate change, the seasonal activity of questing Ixodes ricinus and their infection with Borrelia burgdorferi sensu lato (s.l.) were examined in relation to some climatic data along altitudinal gradients in Switzerland. The first study took place in an Alpine area (Valais) from 750 to 1020 m above sea level. The other gradient was located on a mountain in the foothills of the Jura chain (Neuchâtel) from 620 to 1070 m above sea level. In the Alpine area, the highest questing tick density was observed at the highest altitude. At the lowest altitudes (750 and 880 m), very high saturation deficits, >10 mmHg, were present during most of the tick activity season and they seem to have impaired the thriving of tick populations. The second study in Neuchâtel (2003–2005) was a follow-up of a previous study (1999–2001) in which it was observed that tick density decreased with increasing altitude. During the follow-up study, substantial differences in questing tick density and phenology of ticks were observed: At high elevations, questing tick densities were 2.25 and 3.5 times higher for nymphs and adults, respectively, than during 1999–2001. As observed during 1999–2001, questing tick density decreased with increasing altitude in this site in 2003–2005. Tick questing density remained higher at the lowest altitude. Increased temperatures during summer months, more favorable for ticks, reaching values similar to those registered in the first study at the lowest elevations are probably responsible for the higher tick questing density at high altitudes. B. burgdorferi s.l. infection prevalence in ticks decreased with increasing altitudes along both altitudinal gradients. Long-lasting high saturation deficit values may limit the development of tick populations as too high a moisture stress has a negative effect on tick survival. This factor may have a permanent impact, as it is probably the case at the lowest altitudes in the Alpine area or a more transient effect like in the Neuchâtel gradient.
Ixodes (Pholeoixodes) hexagonus, an efficient vector of Borrelia burgdorferi in the laboratory
Borrelia burgdorferi Johnson et al. was first isolated from the midgut of Ixodes dammini Spielman et al. in the U.S.A. and from the midgut of Lricinus (L.) in Europe. I.ricinus was considered to be the only tick vector of this borrelia, in Europe, until I.hexagonus Leach, the hedgehog tick, was found to harbour spirochaetes. This paper reports an evaluation of the vector competence of I. hexagonus for the spirochaete B. burgdorferi. Transovarial and trans-stadial survival were demonstrated and the spirochaete was transmitted to laboratory mice via the bites of trans-stadially infected I.hexagonus females.
Longitudinal study of Lyme borreliosis in a high risk population in Switzerland
Des coureurs d'orientation venant de toutes les régions de Suisse (n = 416) ont participé à une étude prospective de la borréliose de Lyme. Au printemps 1986, la séroprévalence était de 28,1 %. Au début de l'étude, 84,3 % des coureurs d'orientation ont signalé avoir été piqués par des tiques. Durant la première période (printemps 1986-automne 1986), la deuxième (automne 1986-printemps 1987) et la troisième (printemps 1987-automne 1987), les taux de séroconversion ont été respectivement de 0,6 %, 2,7 %, et 2,1 %. Lors des première et deuxième périodes, l'incidence clinique a été respectivement égale à 1,0 % et 0,25 %. Aucun cas clinique n'a été observé durant la troisième période. Parmi les coureurs ayant présenté une séroconversion durant cette étude (n = 16), deux seulement ont développé des symptômes cliniques. Il semble donc que l'infection à Borrelia burgdorferi soit souvent asymptomatique., Orienteers from all parts of Switzerland (n = 416) were included in a longitudinal study for Lyme borreliosis. In spring 1986, the seroprevalence was 28.1 %. At the beginning of the study, 84.3 % of orienteers reported a history of tick bite, and 3.8 % reported a past history of Lyme borreliosis. During the first (spring 1986-autumn 1986), second (autumn 1986-spring 1987) and third (spring 1987-autumn 1987) period, rates of seroconversion were 0.6 %, 2.7 % and 2.1 % respectively. During the first and second period, clinical incidence were 1.0 % and 0.25 % respectively. No active Lyme borreliosis was detected during the third period. Among orienteers who seroconverted during the study (n = 16), only two developed clinical symptoms. Hence, Borrelia burgdorferi infection is often asymptomatic.
Detection and identification of Borrelia burgdorferi sensu lato genospecies in ticks from three different regions in Slovakia
Lyme-Borreliose ist eine der häufigsten von Zecken vermittelten Erkrankungen, die in der Slowakei auftreten. In dieser Studie wurde Borrelia burgdorferi sensu lato in Wirt-suchenden Zecken, die aus drei Regionen der Slowakei gesammelt worden sind, nachgewiesen und kultiviert. Zur Identifizierung der Genospezies wurden zwei Methoden eingesetzt, die RFLP (restriction fragment length polymorphism) und der "reverse line blot". Ebenso wurde die Prävalenz von B. burgdorferi s.l. in den Zecken ermittelt, welche rund 32% betrug. Von den vier identifizierten Genospezies, B. afzelii, B. burgdorferi sensu stricto, B. garinii und B. valaisiana, wurde B. garinii am häufigsten nachgewiesen., Lyme borreliosis is one of the most common tick-borne diseases that occur in Slovakia. In this study, Borrelia burgdorferi sensu lato was detected and cultivated from questing ticks collected in three areas of Slovakia. Two methods, restriction fragment length polymorphism and reverse line blot, were used for identification of isolates and determination of the prevalence of B. burgdorferi s.l. in the ticks. The prevalence of B. burgdorferi s.l. in I. ricinus detected by reverse line blot was 31.9%. Four genospecies, namely B. garinii, B. valaisiana, B. afzelii and B. burgdorferi sensu stricto were found. B. garinii was the most prevalent genospecies.
Changes in the protein profile and antigenicity of different Borrelia burgdorferi strains after reintroduction to Ixodes ricinus ticks
Eight Swiss strains of Borrelia burgdorferi, with various protein profiles and the North-American strain B31 were artificially introduced into Ixodes ricinus ticks and reisolated 10 days later. All isolates were subsequently examined by SDS-PAGE analysis. Comparing initial isolates with the reisolates, we observed that 7 out of 9 strains changed their protein pattern with respect to the major proteins OspA, OspB and the 22 kDa protein after passage in the tick. The strains NE2, NE4 and NE83 with the initial phenotype of OspA and 22 kDa proteins changed to the phenotype of OspA and OspB, the strains B2 and NE202 with the initial phenotype of OspA acquired an additional protein of 22 kDa and the strain NE58 with the initial phenotype of OspA also acquired a protein of 22 kDa. Examination of these isolates by Western blot analysis demonstrated that the reaction with the monoclonal antibody H5332 and a monospecific polyclonal antibody PoAb/anti-22 kDa differed between the initial isolates and the reisolates.
Infection of field-collected Ixodes ricinus (Acari: Ixodidae) larvae with Borrelia burgdorferi in Switzerland
Free-living larvae from natural areas in five regions in Switzerland were collected to determine the infection rate of Ixodes ricinus L. larvae by Borrelia burgdorferi Johnson. Larvae were examined for the spirochetes using direct fluorescent antibody assay. Of 652 larvae examined, spirochetes were detected in 20 (3.1%). No differences in infection rate among sites were detected. The low prevalence of B. burgdorferi in larvae, compared with higher infection rate in nymphs (12.8%) and adults (14.5%), suggests that transovarial transmission is inefficient. These results suggest that I. ricinus can serve as a reservoir for B. burgdorferi in nature. Further study is needed on the relative importance of ticks, compared with mammals and birds, as reservoirs for B. burgdorferi.
Quantitative analysis of Borrelia burgdorferi gene expression in naturally (tick) infected mouse strains
Adaptation of Borrelia burgdorferi in the vector and vertebrate host is mediated by mechanisms that regulate differential expression of outer surface lipoproteins (Osps). In this study, real time PCR was applied to quantify tissue-specific expression of four linear plasmid (lp54)-encoded (ospA, zs7.a36, zs7.a66 zs7.a68) and one circular plasmid (cp26)-encoded (ospC) gene from B. burgdorferi sensu stricto, in a natural setting of tick-infected immunodeficient (C.B-17 SCID) and immunocompetent (BALB/c and AKR/OlaHsd) mice for up to 120 days post-infection (p.i.). Early during infection (day 30 p.i.) high numbers of spirochetes were found in the heart and joint, but not the ear and spleen tissues of disease-susceptible SCID mice. In disease-susceptible AKR mice spirochetes colonized the ear and joint tissues, but were undetectable in tissues of disease-resistant BALB/c mice. Later in infection (day 120 p.i.), spirochetes had expanded (~1,000-fold) in all SCID tissues tested but were undetectable in AKR and BALB/c mice. Of the five genes analyzed, only zs7.a36 transcripts were detected in various tissues of all infected mouse strains, though at differing levels, whereas ospC transcripts were only found in tissue specimens of SCID mice. Furthermore, gene expression of ospC and zs7.a36 appears to be differentially regulated in distinct organs of individual mice. In contrast, transcripts for ospA, zs7.a66, and zs7.a68 were not detected in any of the mouse strains, independent of their immune status and/or the severity of their infection/inflammatory responses. Late during infection (day 120 p.i.), transcription of zs7.a36 and ospC was down-regulated in the tissues of SCID mice despite expansion of spirochetes. This type of quantitative analysis may be helpful to further disclose principles of pathogenesis of Lyme borreliosis and to design strategies for its therapeutic treatment.
Rickettsia-like microorganisms in the ovarian primordial of molting Ixodes ricinus (acari : ixodidae) larvae and nymphs
Un grand nombre de micro-organismes de type rickettsien ont été observés dans les cellules des ébauches ovariennes de larves et de nymphes d'Ixodes ricinus, lors de la mue. La morphologie de ces organismes est identique chez toutes les tiques examinées. Ils mesurent 0,3-0,4 µm de diamètre et 0,6-1,3 µm, parfois jusqu’à 2,5 µm, de long. Ils possèdent une paroi externe membraneuse fortement ondulée et une membrane plasmique interne. L’épaisseur de chacune de ces 2 structures est de 6-10 nm. Les microorganismes sont toujours inclus, soit seuls soit en groupe, dans des vacuoles contenues dans le cytoplasme de la cellule hôte, dans des mitochondries vacuolées, ou dans des vacuoles dont les parois sont constituées en partie de restes de mitochondrie. Plus les ovaires des nymphes sont développés, plus le nombre de micro-organismes associés aux mitochondries augmente. La relation entre les microorganismes et les mitochondries est discutée, la morphologie des gonades chez les tiques immatures est décrite., A large number of coccoid, or oval to spindle-shaped rickettsialike microorganisms are detected intracellularly within the primordial ovaries of all examined molting larvae and nymphs, descendants of a female Ixodes ricinus collected in a forest near Neuchâtel, Switzerland, and within all molting nymphal I. ricinus captured as starved ticks in the same area. The morphology of the microorganisms is identical in all ticks. They measure 0.3-0.4 µm in diameter and 0.6-1.3 µm, occasionally up to about 2.5 µm in length. The microorganisms possess a highly rippled membranous outer cell wall and an inner plasma membrane. Both cellular boundaries show a thickness of approximately 6-10 nm. The microorganisms are always enclosed, singly or in groups, within membrane limited vacuoles of host cells, within vacuolated mitochondria, or within vacuoles with their wall consisting partially of a deformed mitochondrion and partially of a plasma membrane of host cells. With the advance of the development of the nymphal ovaries, the groups of microorganisms associated with mitochondria become larger and increase in number, and they are dominant within the posterior part of the organs in the nymphs examined at day 21 after repletion. The relationship between the microorganisms and the mitochondria is discussed. The morphology of the gonads of the examined immature ticks is also described.
Protective immunization with plasmid DNA containing the outer surface lipoprotein A gene of Borrelia burgdorferi is independent of an eukaryotic promoter
Plasmid DNA encoding the outer surface lipoprotein A (OspA) of Borrelia burgdorferi under the control of either strong eukaryotic/viral or its own bacterial promoter was injected intramuscularly (m. tibialis anterior) or intradermally into BALB/c and AKR/N mice. OspA-specific antibodies and OspA-reactive T helper 1 cells (Th1) were induced only with those plasmids containing the ospA structural gene including its own regulatory control region immediately upstream. In the absence of the ospA promoter, no or only marginal immune responses to ospA were obtained, even when strong eukaryotic promoter/enhancer elements were present. Together with the finding that the ospA promoter is active in a mouse B- lymphoma line, the data suggest that spirochetes are able to express at least part of their genes in the mammalian environment. Mice previously vaccinated with the relevant ospA plasmid DNA were protected against subsequent experimental challenge with a virulent strain of B. burgdorferi, as measured by the appearance of antibodies to a prominent protective epitope (LA-2) and the failure to re- isolate spirochetes from ear biopsies. In addition, C.B-17 severe-combined immunodeficient mice could be protected against infection by passive transfer of immune sera from ospA plasmid DNA-inoculated normal mice. Protective LA-2- related antibody titers obtained after repeated immunization persisted for 200 days and longer. This simple procedure of immunization using plasmid DNA consisting of a prokaryotic gene under the control of its own promoter holds great promise for the development of alternative subunit vaccines against bacterial infections, including Lyme disease. In addition, the availability of this novel prokaryotic promoter element now allows the study of the basis for the differential expression of bacterial genes in prokaryotic and eukaryotic environments.