Lyme disease is the most common tick-borne disease in North America, with approximately 35,000 cases reported to the Centers for Disease Control in 2008. The genome of its causative agent, Borrelia burgdorferi, encodes for a set of genes involved in the metabolism and regulatory activities of the second messenger nucleotide, cyclic-di-GMP (c-di-GMP). Rrp1 is a response regulatory-diguanylate cyclase, and its regulatory capability is likely mediated via production of c-di-GMP, as it lacks a DNA-binding domain. One known class of c-di-GMP effector/binding proteins are those that harbor a PIlZ domain. The genome of B. burgdorferi strain 5A4 encodes for one chromosomally-carried PilZ domain, which we have designated PlzA. Additionally, certain B. burgdorferi strains encode for a second PilZ domain-containing protein (PlzB) which is plasmid-carried. Both PlzA and PlzB were found to bind specifically to c-di-GMP, and c-di-GMP binding by PlzA was found to be dependant upon arginine residues in the c-di-GMP binding region. Additionally, expression of PlzA was found to be upregulated by tick feeding and was constitutive in the mammalian host. We next constructed two deletion/allelic exchange mutants – one with the targeted deletion of PlzA, and on ethat replaced PlzA with PlzB in a strain lacking the plzB gene. Our studies demonstrated that ΔplzA was deficient in motility and was also non-infectious in the mouse model of B. burgdorferi infection. Additionally, this strain remained viable in larval Ixodes ticks. Also, B31-plzB KI was deficient in motility, as well as infectivity, demonstrating that PlzB is unable to complement for functions fo PlzA in vitro and in vivo and that it may play other roles in the biology of B. burgdorferi strains carrying the plzB gene. These studies represent the first identification of a c-di-GMP binding protein in any spirochete, but also represent the first demonstration of the importance of PilZ domain proteins in a spirochetal system. We additionally examined the effects of c-di-GMP synthesis and breakdown in the related bacterium, B. hermsii, a causative agent of tick-borne relapsing fever (TBRF). Deletion mutants in Rrp1 (B. hermsii’s sole diguanylate cyclase) and PdeA (B. hermsii’s only EAL domain-containing phosphodiesterase) were created. These strains were analyzed in order to determine: 1) the effect(s) of the losse of Rrp1/PdeA on intracellular spirochete c-di-GMP levels, and 2) the effects of Rrp1/PdeA on the establishment of murine infection and on gross motility/chemotaxis. It was demonstrated that c-di-GMP accumulates intracellularly in the cells lacking PdeA. Additionally, spirochetes were shown to chemotax towards N-acetyl-glucosamine (NAG) and they did not form soft agar swarms. In contrast, cells lacking Rrp1 did not accumulate detectable levels of c-di-GMP, demonstrated a reduced ability to chemotax towards NAG, and swarmed on soft agar in a fashion indistinguishable from wild type. Despite these differences in phenotype, both mutant strains display an attenuated murine infectivity. These results indicate that c-di-GMP is indeed important in the TBRF spirochete, B. hermsii and this vital second messenger plays key roles in virulence, motility, and chemotaxis. These studies also pave the way for future investigation of B. hermsii through use of targeted genetic manipulation.

Lyme disease is a tick-borne infection caused by Borrelia burgdorferi, B. garinii, and B. afzelii. These spirochetes experience environmental fluctuations as they are passed between mammalian and Ixodes tick hosts throughout their enzootic cycle. Recent studies have suggested cyclic diguanylate (c-di-GMP), a ubiquitous secondary messenger, is a key modulator of B. burgdorferi adaptive responses and may play a significant role in cycle progression. In this study, we examined the impact of the sole diguanylate cyclase (Rrp1), c-di-GMP binding proteins (PlzA and PlzB), and HD-GYP-containing phosphodiesterase (PdeB) in disease establishment of both murine and Ixodes tick systems. Strains harboring targeted gene deletions or plasmid-based constitutive gene expression constructs were generated. Rrp1 was required for tick colonization, yet overexpression abolished murine disease, thus implicating the requirement of finely regulated c-di-GMP levels for enzootic cycle progression. Deletion of rrp1 disrupted translational motion and swarming patterns by causing extended cell runs, eliminating stops/flexes, and reducing swarming capabilities. This was attributed to a defect in N-acetyl-D-glucosamine (NAG) metabolism and chemotaxis. NAG is a major source of nutrition for B. burgdorferi within the tick environment; therefore this defect would impede spirochete migration towards feeding ticks, as well as pathogen uptake and survival within the Ixodes vector. In contrast, the downstream c-di-GMP effector, PlzA, was critical for murine disease but nonessential for survival within ticks nor functionally complemented by PlzB. Deletion of plzA altered strain motility and swarming similarly to the rrp1 deletion mutant, yet had a distinct phenotype with significantly slower translational motion and no affect on NAG chemotaxis and metabolism. This indicates B. burgdorferi could possess alternate c-di-GMP effectors or Rrp1 could be directly influencing these cellular processes. Uniquely, PdeB did not abolish murine infection via needle inoculation, but wasrequired for natural transmission from ticks. This defect was linked to the decreased tick colonization efficiency upon pdeB deletion. Together, these analyses indicate that c-di-GMP signaling is an important virulence mechanism of Borrelia burgdorferi and demonstrate the complexity of this signaling pathway in an arthropod-borne pathogen. The data presented here additionally provide significant new insight into the gene regulatory mechanisms of the Lyme disease spirochetes.

Introduction: Lyme disease is caused by the tick-borne spirochete Borrelia burgdorferi. Research has shown that dogs can be used as sentinels for human infection of Lyme disease. The purpose of this 5-year, retrospective study was to determine if there was any evidence that the incidence of canine Lyme disease has increased between 2000 and 2005 in Accomack and Northampton counties. An increased incidence in Lyme disease in dogs may indicate an increased present or future risk of Lyme disease in humans.Methods: Cases of canine Lyme disease were identified via practice invoicing systems and dogs that received doxycycline were entered into the database. Demographic information and the absence or presence of clinical signs such as fever, lameness, articular swelling, lymphadenomegaly, anorexia, general malaise and improvement after antibiotic use were collected. Testing history also was recorded.Results: Cases of canine Lyme disease that met any definition were identified (n=1048). Over the 5-year period the number of positive ELISA test results increased and the frequency of clinical signs decreased. The incidence of disease meeting the practitioner's definition increased until 2004 when the incidence dropped from 105.33 cases per 1,000 dogs to 56.93 cases per 1,000. The incidence of disease based on the study probable definition remained fairly constant with a high in 2002 of 2.94 cases per 1,000 dogs.Discussion: Trends of canine Lyme disease coincided with the introduction and use of the in-house ELISA test. Practitioners could identify more dogs exposed to Borrelia burgdorferi. The areas with the highest frequency of canine cases of Lyme disease also had the highest frequency of human cases reported to the Virginia Department of Health. Further study could identify animals that tested positive and later developed clinical signs. Using dogs as sentinels for human infection allows public health workers to identify endemic areas regardless of human case reports.

In total, 76 birds, mainly breeding females, were captured in years 2007-2009. Birds were inspected for the presence of avian influenza (AI) viruses and the AI antibodies. They were inspected as well for the presence of bacteria Borrelia burgdorferi s. l. and Anaplasma phagocytophilum and protozoan Leucocytozoon simondi in blood. The feces were examined for the oocysts and eggs of intestinal parasites. AI viruses were found in 4 cases, while over 80% of inspected birds had AI antibodies. Bacteria B. burgdorferi s. l. and A. phagocytophilum were found in 2 and 1 case respectively. Protozoan L. simondi was not found in any blood smear. The oocysts from protozoan genera Eimeria and Tyzzeria and eggs from roundworm genera Capillaria, Amidostomum and Echinuria were observed in feces. Statistical analyses were due to the small size of our data set inconclusive.

The aim of the study was to obtain and elaborate information focused on metamorphoses of Borrelia burgdorferi sensu lato (s.l.) spirochetes. The research included detection of various stress conditions for production of dormant forms of Borrelia burgdorferi s.l. spirochetes in vitro. Subsequently, metamorphoses from dormant to motile stages was observed under favourable conditions. Proper PCR method for primers aimed to detect dormant forms of Borrelia was delevoped. The infectious potential of dormant spirochetes of Borrelia burgdorferi sensu stricto (s.s.) complex was observed in vivo. Transformations of spirochetes have also been observed in real time and individual stages have been recorded.

Lyme borreliosis (LB) is the most frequent vectorborne disease in the Northern Hemisphere. It is a complex bacterial zoonosis involving vertebrate hosts and hard ticks of the genus Ixodes. The causative agents, bacteria of the LB group of spirochaetes, form a species complex comprising 17 named species. As is the case for most microbial pathogens, epidemiological and ecological studies require appropriate genotyping. Although the use of single loci may provide rapid results, there are serious disadvantages, in particular when inferring evolutionary relationships or geographic population structure. A novel multilocus sequence analysis (MLSA) system of the LB group spirochaetes has been developed based on housekeeping genes to overcome these problems. Here, the system is optimized and tested using extracted spirochaetal DNA directly from ticks and then utilized to obtain insights into the migration and spread of individual species as well as to investigate the evolutionary origins of the species complex. Species belonging to the LB group of spirochetes display different patterns and levels of host specialisation which makes this an ideal system to study the impact of host associations on spread of zoonotic tickborne diseases. For example, Borrelia valaisiana and B. garinii are transmitted exclusively by birds while B. afzelii is transmitted by rodents. I demonstrate that the migration of the LB species is dependent on, and limited by, the migration of their respective hosts. I also show the presence of B. afzelii strains in England and, through the use of the MLSA scheme, demonstrate that the strains are highly structured. A close evolutionary relationship between B. afzelii and its rodent host species is shown. Furthermore, through phylogenetic analyses, some evidence of a coevolutionary relationship between the LB group species and their major group of vector species, the Ixodes persulcatus species complex, is presented and a Eurasian origin for the species group is suggested.

Made available in DSpace on 2016-04-28T20:15:30Z (GMT). No. of bitstreams: 1 2008- Jania de Resende.pdf: 467062 bytes, checksum: cf6acc7c0e32afbfa397b5b1c07d0d87 (MD5) Previous issue date: 2008-02-22 / Funda??o Carlos Chagas Filho de Amparo a Pesquisa do Estado do Rio de Janeiro / Embryonic cells of tick in vitro constitute an important one tool for culture and study of the biology of B. burgdorferi. Spirochetes Borrelia burgdorferi is the aetiologic agent of borreliose of Lyme in U.S.A. and Europe, where it is transmitted by tick of the Ixodes genus. The aim of this work was in vitro to cultivate B. burgdorferi (American Cepa G39/40) in primary culture of embryonic cells of Rhipicephalus (B.) microplus and the A. cajennense. From the primary culture of embryonic cells of R. (B.) microplus were performed subculture maintained with medium Leibovitz's (L-15) free of antibiotic, that later it were changed by medium Barbour-Stoener-Kelly (BSK). After the addition of the BSK inoculated B. burgdorferi of and also in Tube of Leighton (TL) with free BSK of cells (control) in a final concentration of 6x106 spirochetes/mL. The embryonic cells of the A. cajennense initially were cultivated in medium L-15 with antibiotic, which was substituted by the BSK. Later, 1,1x107 spirochetes/mL in medium BSK cultivated was inoculated, and also in TL controlled free of cells. All the culture was incubated at 34?C. The development of the culture was observed in microscope of inverted contrast of phase, as well as the countings of B. burgdorferi performed in chamber of neubauer. Cover glass of the TL had been stained with Giemsa. It was evidenced by the observation in microscope of inverted contrast of phase the survival, attach and multiplication of B. burgdorferi, in the embryonic cells of R. (B.) microplus and A. cajennense. It did not have differences in the finale counting of spirochetes cultivated in cells of R. (B.) microplus when compared with the free control of cells, but on with cells of the A. cajennense, the number amount approximately was 1,9x107 spirochetes/mL, and while in the tube it has controlled free of cells was 1x106 spirochetes /mL. The culture of cells of tick R. (B.) microplus and the A. cajennense have potential to be used as substratum for culture of B. burgdorferi, and study of its development. / C?lulas embrion?rias de carrapatos mantidas in vitro constituem uma importante ferramenta para cultivo e estudo da biologia de Borrelia burgdorferi. A espiroquetas B. burgdorferi ? o agente etiol?gico da borreliose de Lyme nos EUA e Europa, onde ? transmitida por carrapatos do g?nero Ixodes. O objetivo deste trabalho foi cultivar in vitro Borrelia burgdorferi (Cepa Americana G39/40) em cultura prim?ria de c?lulas embrion?rias dos carrapatos Rhipicephalus (Boophilus) microplus e Amblyomma cajennense. A partir da cultura prim?ria de c?lulas embrion?rias de R. (Boophilus) microplus foram realizados subcultivos mantidos com meio Leibovitz s L-15 livre de antibi?tico, que posteriomente foi trocado pelo meio Barbour-Stoener-Kelly (BSK). Ap?s a adi??o do BSK na cultura, inoculou-se B. burgdorferi e tamb?m em Tubo de Leighton (TL) com BSK livres de c?lulas (controle), numa concentra??o final de 6x106 espiroquetas/mL. As c?lulas embrion?rias de A. cajennense foram inicialmente cultivadas em meio L-15 com antibi?tico, o qual foi substitu?do pelo BSK. Posteriormente, inoculou-se 1,1x107 espiroquetas/mL cultivadas em meio BSK, e tamb?m em TL controle livre de c?lulas. Todos os cultivos foram incubados em estufa bacteriol?gica a 34?C. O desenvolvimento dos cultivos foram observados em microsc?pio de contraste de fase invertido, assim como as contagens de B. burgdorferi realizados em c?mara de neubauer. As lam?nulas dos TL foram coradas com Giemsa. Foi constatado pela observa??o em microsc?pio de contraste de fase invertido a sobreviv?ncia, ader?ncia e multiplica??o de B. burgdorferi, nas c?lulas embrion?rias de R. (Boophilus) microplus e A. cajennense. N?o houve diferen?as na contagem final de espiroquetas cultivadas em c?lulas de R. (Boophilus) microplus quando comparada ao controle livre de c?lulas, mas sobre c?lulas de A. cajennense o valor total de aproximadamente 1,9x107 espiroquetas/mL, e enquanto no tubo controle livre de c?lulas foi 1x106 espiroquetas/mL. O cultivo de c?lulas do carrapato R. (Boophilus) microplus e A. cajennense t?m potencial para ser utilizado como substrato para cultivo de B. burgdorferi e para estudo de seu desenvolvimento.

Borrelia burgdorferi, the causative agent of Lyme disease, exists in a defined enzootic cycle involving Ixodes scapularis ticks and various vertebrates. Humans can serve as an accidental host, if a tick colonized with B. burgdorferi happens to feed on a human. B. burgdorferi are also accidental pathogens: they do not make toxins, or destroy host tissue by other mechanisms. They merely transmit between vector and host to survive. In order to do this, they must effectively sense their current environment, and appropriately alter cellular processes. Understanding the regulatory mechanisms of how B. burgdorferi manages to do this has been a focus of the Stevenson lab for many years. Previous work identified SpoVG as a DNA-binding protein. Although a homologue of this protein had been implicated to serve a regulatory role in other bacteria, the Stevenson lab was the first to demonstrate a function for the protein, both for B. burgdorferi and two other bacteria. Studies contained in this body of work aim to provide insight into regulation of SpoVG by B. burgdorferi as well the impact that it has on gene regulation. By using genetic mutants, we determined that SpoVG is regulated at the levels of transcription and translation in culture by growth rate, temperature, and other regulatory factors. Additionally, we provide evidence that SpoVG regulates its own expression. Numerous genes are under control of SpoVG. Biochemical analyses revealed that SpoVG specifically interacts with DNAs and RNAs associated with genes found to be under its regulatory control. Finally, we provide evidence for SpoVG acting in concert with other known regulatory factors such as other DNA-binding proteins and the cyclic di-nucleotide second messengers cyclic-di-GMP and cyclic-di-AMP. All together, these studies provide insight into how B. burgdorferi broadly regulates cellular processes during different stages of the enzootic cycle. We hypothesize that SpoVG does this through globally manipulating the three-dimensional structure of the bacterial chromosome, and that exactly how SpoVG acts at any given point will be dependent on the other regulatory factors that are also present in the cell.

In der vorliegenden Arbeit sollten die unterschiedlichen Manifestationsformen der Borreliose anhand verschiedener Parameter verglichen und der Verlauf einer Borrelioseerkrankung analysiert werden. Dabei konnte gezeigt werden, dass es sich um eine ganzjährige Erkrankung handelt, die jedoch - je nach Manifestationsform - verschiedene jahreszeitliche Gipfel aufweist. So kommen die Neuroborreliose und die Frühborreliose gehäuft im Frühling und im Sommer vor, während die Lyme Arthritis durchweg ganzjährig auftritt. Da die Borrelien erst nach ca. 12 Stunden Latenz auf den Wirt übertragen werden, sind es vor allem die unbemerkten Zeckenstiche, die eine Infektion verursachen. Deshalb ist es nicht verwunderlich, dass sich nur rund 55 % der Patienten an einen zurück liegenden Zeckenstich erinnern konnten. Bei der Untersuchung der verschiedenen Laborparameter ergaben sich keine wesentlichen Unterschiede zwischen den Manifestationsformen bezüglich Hämoglobingehalt, Blutsenkungsgeschwindigkeit, Thrombozyten und Leukozyten im Serum. Desweiteren ergaben sich keine Hinweise auf eine Beteiligung des Rheumafaktors oder antinukleärer Antikörper an den immunologischen Prozessen während einer Lyme Arthritis Erkrankung. IL-17 konnte mittels ELISA weder in Serum/Plasma noch in Synovialflüssigkeit signifikant erhöht nachgewiesen werden. Es bleibt jedoch unklar, ob dies aufgrund einer langen Lagerung der Proben der Fall war oder ob IL-17 tatsächlich nicht signifikant erhöht vorlag. Aus der Literatur kann man entnehmen, dass IL-17 bei den Abläufen einer Infektion mit B. burgdorferi eine wichtige Rolle zu spielen scheint. Dies könnte einen Ansatz für neue Behandlungsmethoden der Lyme Arthritis darstellen, weshalb die Durchführung weitere Untersuchungen wichtig ist. Durch die Analyse der initialen Serologie konnte die Antikörperreaktion während einer Infektion mit B. burgdorferi veranschaulicht werden, die mit der Bildung von IgM- Antikörpern beginnt und dann einen Shift zu IgG-Antikörpern vollzieht. Die Antigene p19, p31/34, p39 und p65 scheinen hierbei vor allem bei der Lyme Arthritis im Vordergrund zu stehen. Weiterhin konnte bei dieser Analyse beobachtet werden, dass teilweise zahlreiche serologische Untersuchungen bei ein und demselbem Patienten statt fanden, was darauf schließen lässt, dass diese Untersuchung zur Verlaufskontrolle benutzt wurde. Grundsätzlich ist die Serologie hierzu jedoch nicht geeignet. Vielmehr sollte sie lediglich zur Diagnosestellung dienen. In dieser Funktion ist sie bei korrekter Interpretation ein hervorragendenes diagnostisches Mittel. Leider werden serologische Untersuchungen jedoch häufig falsch interpretiert und angewendet, was dazu führt, dass Patienten häufiger oder auch länger als nötig antibiotisch behandelt werden. So erhielten auch rund 8 % der Lyme Arthritis Patienten mehr als 2 Zyklen antibiotischer Therapie. Die Nebenwirkungen dieser langwierigen antibiotischen Behandlung sind nicht zu vernachlässigen. Die aktuellen Leitlinien sehen im Gegensatz dazu nämlich nach dem zweiten antibiotischen Zyklus den Beginn einer Therapie mit DMARD ́s vor, selten kommen auch intraartikuläre Steroide zum Einsatz. Diese beobachtete „Überdiagnostik und Übertherapie“ der Borrelioseerkrankung spiegelt vermutlich die Angst vor chronischen Verlaufsformen in der Bevölkerung und teilweise auch unter Ärzten wider. Bereits in anderen Studien konnte die Existenz einer therapieresistenten Verlaufsform der Borreliose - insbesondere der Lyme Arthritis – nachgewiesen werden. In der vorliegenden Arbeit litten 24 % der Lyme Arthritis Patienten nach einer adäquaten antibiotischen Behandlung laut eigenen Angaben zum Zeitpunkt der Umfrage noch unter Gelenkbeschwerden. Ob es sich hierbei nun um eine schwerwiegendere Verlaufsform in Europa handelt, bleibt zunächst offen. Zur Objektivierung der Beschwerden, zum Ausschluss einer Zweitinfektion oder aber auch einer Fibromyalgie als mögliche Differentialdiagnose sind prospektive Studien mit klinischer Untersuchung der Patienten im Verlauf nötig. Eindeutig ist jedoch, dass durch eine frühzeitige antibiotische Behandlung nach Infektion das Fortschreiten der Erkrankung effektiv verhindert werden kann. Chronische Beschwerden gaben hingegen auch die Patienten mit zurückliegender Neuroborreliose an. Rund 20% der Patienten litten nach eigener Angabe unter Allgemeinsymptomen, die ihre Lebensqualität subjektiv wesentlich einschränkten. Der spezifische Zusammenhang zwischen diesen Beschwerden und einer Neuroborreliose bzw. einer Borrelioseerkrankung im Allgemeinen ließ sich jedoch nicht endgültig klären. Es wäre hierfür der Vergleich mit einer gesunden Kohorte und die Anwendung spezieller neuropsychologischer Untersuchungen nötig. / This paper should compare different manifestations of borreliosis based on various parameters as well as analyze the development of a borreliosis infection. It could be shown that it is a year-round disease reaching different seasonal peaks depending on its manifestation. As an example, neuro borreliosis and early borreliosis occur frequently in spring and summer, whereas lyme arthritis infections occur during the whole year. As borrelia are transmitted to the host only after 12 hours of latency, especially tick bites that go unnoticed cause infections. That is why it does not come as a surprise that only 55% of the patients were able to remember a tick bite in the past. Examining the different laboratory parameters, no substantial differences between the different types of manifestation regarding haemoglobin content, erythrocyte sedimentation rate, thrombocytes and leucocytes in the serum were discovered. In addition, there were no hints as to a contribution of the rheumatic factor or antinuclear antibodies to immunologic processes during a lyme arthritis infection. Using ELISA, IL-17 could neither be detected in serum/plasma nor in synovial liquid in significantly increased concentrations. However, it remains unclear whether this was due to the long storage of the samples or whether IL-17 was actually not significantly increased. According to literature on this topic, IL-17 seems to play a major role during the course of an infection with B. burgdorferi. This could be a starting point for new ways of treating lyme arthritis and makes further research necessary. By analyzing initial serology, the antibody reaction during an infection with B. burgdorferi could be shown. It starts with the creation of IgM antibodies and then shifts to IgG antibodies. The antigenes p19, p31/34, p39 and p65 seem to be especially important when it comes to lyme arthritis. The analysis also made it clear that in some cases a single patient was exposed to numerous serological examinations what makes it probable that this examination was used to control the course of the infection. Basically, serology is not a suitable instrument to do so. It should rather be used to diagnose. Interpreting it correctly, it is in that function an excellent means of diagnosing. Unfortunately, serological examinations are often interpreted and applied in the wrong way, what leads to a more frequent or unnecessarily long antibiotics treatment of patients. Around 8% of lyme arthritis patients received more than two cycles of antibiotics treatment. The side effects of this longsome antibiotics treatment are not to be neglected. In contrast, current guidelines recommend a DMARDs therapy after the second cycle of antibiotics treatment, in rare cases intraarticular steroids are used. This observed “overdiagnostics and overtherapy” of borreliosis infections presumably reflects the fear of chronic infections among the population and partly among doctors. There have already been other studies proving the existence of a form of borreliosis infection resistant to therapy – especially regarding lyme arthritis. According to this paper, 24% of lyme arthritis patients said, when they were interrogated, that they were still suffering from arthralgia after an adequate antibiotics treatment. It remains open whether this is just a more serious progression in Europe. To come to an objective judgement of the pains, to exclude a second infection or a fibromyalgia as a potential differential diagnosis, prospective studies invoving clinical examinations of the patients are necessary. However, it is beyond any doubt that an early antibiotics treatment after the infection can effectively prevent the progress of the infection. Nevertheless, also patients with an overcome neuroborreliosis reported chronic pains. About 20% of the patients reported that they were suffering from general syndromes limiting their quality of life subjectively essentially. The specific connection between these pains and a neuroborreliosis respectively a borreliosis infection in general could, however, not be proven in the end. For this, it would be necessary to make a comparison with a healthy cohort and to apply special neuropsychological studies.

<p>This thesis examines seasonal, diel and vertical distribution patterns of activity of host-seeking <i>Ixodes ricinus</i> (L.) ticks at three localities in south-central Sweden. In addition, by examining the prevalence of infection in ticks with Lyme borreliosis (LB) spirochetes, <i>Borrelia burgdorferi</i> s.l, information for estimating relative LB risk in humans and the effect of control measures directed against this tick vector is provided.</p><p>The seasonal activity pattern of <i>I. ricinus</i> was, in general, bimodal with peaks of activity in May-June and August-September. Tick densities were generally high at Torö and low at Kungshamn-Morga. The greatest variation in tick density occurred at the sample site level, which indicated a patchy distribution of ticks. The diel activity of adult <i>I. ricinus</i> sampled at Bogesund showed a distinct nocturnal activity peak while nymphal ticks exhibited no particular diel variation. At the meadow site, there was a strong negative association between activity of each tick stage and ambient air temperature, and larval ticks also showed a nocturnal activity peak. <i>I. ricinus</i> of all stages were present in the vegetation up to at least 140 cm above ground level. At Torö, host-seeking larvae were found at significantly lower levels (below 20 cm) in the vegetation compared to nymphs and adults (50-59 and 60-79 cm, respectively). Vegetation structure is likely to be the main factor governing tick vertical distribution at this locality. The northern limit of the geographical distribution of <i>I. ricinus</i> in Sweden corresponds with the southern boundary of the taiga zone, as well as with several other climatic or vegetational isoclines primarily associated with the vegetation period.</p><p>The prevalence rates of <i>Borrelia</i> spirochetes, recorded by phase-contrast microscopy in host-seeking <i>I. ricinus</i>, were 0% in larvae, 5.8-13.1% in nymphs and 14.5-28.6% in adult ticks. The human LB risk, estimated by the number of <i>Borrelia</i>-infected nymphs per hectare, was greater at Torö than at Kungshamn-Morga and greater in woodland than in open areas. The risk also possessed a bimodal seasonal pattern similar to that of subadult host-seeking activity. Controlling the number of infected nymphs through de-ticking of reservoir hosts seems not to be an effective control measure in Sweden due to the ubiquitous availability of alternative reservoir hosts.</p>