Acute Meningoencephalitis Associated with Borrelia miyamotoi, Minnesota, USA.

Borrelia miyamotoi is an emerging tickborne pathogen that has been associated with central nervous system infections in immunocompromised patients, albeit infrequently. We describe a case-patient in Minnesota, USA, who had meningeal symptoms of 1 month duration. B. miyamotoi infection was diagnosed by Gram staining on cerebrospinal fluid and confirmed by sequencing.

First molecular detection of Borrelia sp. in tapirs (Tapirus terrestris).

Borrelia theileri is a tick-borne spirochete causative agent of fever, apathy and reduced food consumption in cattle. Molecular diagnosis has expanded the understanding of Borrelia theileri with new hosts and geographical locations being described. The present study aimed to describe the first molecular detection of B. theileri in wild tapirs (Tapirus terrestris) from South America. Blood DNA samples obtained from 99 tapirs sampled in Pantanal (n = 61) and Cerrado (n = 38) biomes were screened using a qPCR assay based on the 16 S rRNA gene of Borrelia sp. Positive samples in the qPCR assay were subjected to PCR assays to allow characterization of fragments from 16 S rRNA and flaB genes. Two (2/99; 2.0%) animals from Pantanal biome were positive in the qPCR and one sample presented bands of expected size for the flaB protocol. Amplicons from this sample were successfully cloned and sequenced. In the phylogenetic analysis, Borrelia sp. from T. terrestris grouped together with B. theileri sequences previously detected in Rhipicephalus microplus ticks and cattle from Minas Gerais State in Brazil, Rhipicephalus geigyi from Mali, and R. microplus and Haemaphysalis sulcata from Pakistan. This finding contributes to our knowledge regarding susceptible hosts species for B. theileri. More studies are necessary to understand the potential effects of B. theileri on tapir's health.

Tick-borne diseases under the radar in the North Sea Region.

The impact of tick-borne diseases caused by pathogens such as Anaplasma phagocytophilum, Neoehrlichia mikurensis, Borrelia miyamotoi, Rickettsia helvetica and Babesia species on public health is largely unknown. Data on the prevalence of these pathogens in Ixodes ricinus ticks from seven countries within the North Sea Region in Europe as well as the types and availability of diagnostic tests and the main clinical features of their corresponding diseases is reported and discussed. Raised awareness is needed to discover cases of these under-recognized types of tick-borne disease, which should provide valuable insights into these diseases and their clinical significance.

High Prevalence of Borrelia mayonii (Spirochaetales: Spirochaetaceae) in Field-Caught Tamias striatus (Rodentia: Sciuridae) From Northern Wisconsin.

Borrelia mayonii is a recently discovered bacterial spirochete that causes Lyme disease and is transmitted by the blacklegged tick, Ixodes scapularis Say (Acari: Ixodidae). To date, B. mayonii has been isolated from two vertebrate host species in Minnesota: field-caught white-footed mice (Peromyscus leucopus Rafinesque; Rodentia: Cricetidae) and American red squirrel (Tamiasciurus hudsonicus Erxleben). Here, we describe the first detection of B. mayonii in field-caught eastern chipmunks (Tamias striatus L. (Rodentia: Cricetidae)) from northern Wisconsin. During our study, we captured 530 unique small mammals and found an infection prevalence of 23.50% in field-caught eastern chipmunks (4/17) and 1.19% in Peromyscus spp. (5/420). Mean larval and nymphal burdens were determined for captured Blarina brevicauda (0, 0), Glaucomys volans (0.29, 0.14), Myodes gapperi (0.27, 0), Napaeozapus insignis (0, 0.25), Peromyscus spp. (1.88, 0.11), T. striatus (1.06, 0.65), and Sorex cinereus (0.09, 0). The high B. mayonii infection prevalence in eastern chipmunks suggests that the species may be an important reservoir for B. mayonii in the Upper Midwest.

The First Immunocompetent Mouse Model of Strictly Human Pathogen, Borrelia recurrentis.

The spirochetal bacterium Borrelia recurrentis causes louse-borne relapsing fever (LBRF). B. recurrentis is unique because, as opposed to other Borrelia spirochetes, this strictly human pathogen is transmitted by lice. Despite the high mortality and historically proven epidemic potential and current outbreaks in African countries and Western Europe, research on LBRF has been obstructed by the lack of suitable animal models. The previously used grivet monkey model is associated with ethical concerns, among other issues. An existing immunodeficient mouse model does not limit bacteremia due to its impaired immune system. In this study, we used genetically diverse Collaborative Cross (CC) lines to develop the first LBRF immunocompetent mouse model. Out of 12 CC lines tested, CC046 mice consistently developed B. recurrentis-induced spirochetemia during the first 3 days postchallenge as concordantly detected by dark-field microscopy, culture, and quantitative PCR. However, spirochetemia was not detected from day 4 through day 10 postchallenge. The high-level spirochetemia (>107 cells/ml of blood) observed in CC046 mice was similar to that recorded in LBRF patients as well as immunocompetent mouse strains experimentally infected by tick-borne relapsing fever (RF) spirochetes, Borrelia hermsii and Borrelia persica. In contrast to the Old World and New World RF spirochetes, which develop multiple relapses (n = 3 to 9), B. recurrentis produced only single culture-detectable spirochetemia in CC046 mice. The lack of relapses may not be surprising, as LBRF patients and the grivet monkey model usually develop no or only 1 to 2 spirochetemic relapses. The novel model will now allow scientists to study B. recurrentis in the context of intact immunity.

Retrospective Survey of Borrelia spp. From Rodents and Ticks in Thailand.

Borrelia is a genus of spirochetal bacteria with several species known to cause disease in humans. The distribution of Borrelia has rarely been studied in Thailand. In this study, a retrospective survey of Borrelia was conducted in ticks and wild rodents to better characterize the prevalence, diversity, and distribution of Borrelia across Thailand. Several pools of DNA from tick samples were positive for Borrelia spp. (36/258, 13.9%). Borrelia theileri/B. lonestari was found in 17 tick samples (16 pools of Haemaphysalis bandicota and 1 pool of Rhipicephalus sp.), and Borrelia yangtzensis was found in 8 tick samples (2 pools of H. bandicota and 6 pools of Ixodes granulatus). Borrelia spp. were detected at low prevalence levels in rodent tissue samples (24/2001, 1.2%), with 19 identified as B. theileri or B. lonestari and 5 identified as B. miyamotoi. Several geographic and species-specific infection trends were apparent, with Ixodes ticks infected with B. yangtzensis and Haemaphysalis and Rhipicephalus ticks infected with both B. yangtzensis and B. theileri/B. lonestari. Notably, B. yangtzensis showed a similar geographic distribution to B. miyamotoi, which was identified in new areas of Thailand in this study. The flagellin gene sequence from B. miyamotoi was more similar to European (99.3-99.9%) than Japanese (96.9-97.6%) genotypes. This study greatly expands the knowledge of Borrelia in Thailand and identified several Borrelia species for the first time. It also found several ticks and rodents infected with the pathogen that were not previously known to carry Borrelia.

Genetic Manipulation of Borrelia.

Genetic studies in Borrelia require special consideration of the highly segmented genome, complex growth requirements and evolutionary distance of spirochetes from other genetically tractable bacteria. Despite these challenges, a robust molecular genetic toolbox has been constructed to investigate the biology and pathogenic potential of these important human pathogens. In this review we summarize the tools and techniques that are currently available for the genetic manipulation of Borrelia, including the relapsing fever spirochetes, viewing them in the context of their utility and shortcomings. Our primary objective is to help researchers discern what is feasible and what is not practical when thinking about potential genetic experiments in Borrelia. We have summarized published methods and highlighted their critical elements, but we are not providing detailed protocols. Although many advances have been made since B. burgdorferi was first transformed over 25 years ago, some standard genetic tools remain elusive for Borrelia. We mention these limitations and why they persist, if known. We hope to encourage investigators to explore what might be possible, in addition to optimizing what currently can be achieved, through genetic manipulation of Borrelia.

Prevalence of single and coinfections of human pathogens in Ixodes ticks from five geographical regions in the United States, 2013-2019.

As the geographic distributions of medically important ticks and tick-borne pathogens continue to expand in the United States, the burden of tick-borne diseases continues to increase along with a growing risk of coinfections. Coinfection with multiple tick-borne pathogens may amplify severity of disease and complicate diagnosis and treatment. By testing 13,400 Ixodes ticks from 17 US states spanning five geographical regions for etiological agents of Lyme disease (Borrelia burgdorferi sensu stricto [s.s.] and Borrelia mayonii), Borrelia miyamotoi disease (Borrelia miyamotoi), anaplasmosis (Anaplasma phagocytophilum), and babesiosis (Babesia microti) we show that B. burgdorferi s.s. was the most prevalent and widespread pathogen. Borrelia miyamotoi, A. phagocytophilum, and B. microti were widespread but less prevalent than B. burgdorferi s.s. Coinfections with B. burgdorferi s.s. and A. phagocytophilum or B. microti were most common in the Northeast and occurred at rates higher than expected based on rates of single infections in that region.

Immune Response to Borrelia: Lessons from Lyme Disease Spirochetes.

The mammalian host responds to infection with Borrelia spirochetes through a highly orchestrated immune defense involving innate and adaptive effector functions aimed toward limiting pathogen burdens, minimizing tissue injury, and preventing subsequent reinfection. The evolutionary adaptation of Borrelia spirochetes to their reservoir mammalian hosts may allow for its persistence despite this immune defense. This review summarizes our current understanding of the host immune response to B. burgdorferi sensu lato, the most widely studied Borrelia spp. and etiologic agent of Lyme borreliosis. Pertinent literature will be reviewed with emphasis on in vitro, ex vivo and animal studies that influenced our understanding of both the earliest responses to B. burgdorferi as it enters the mammalian host and those that evolve as spirochetes disseminate and establish infection in multiple tissues. Our focus is on the immune response of inbred mice, the most commonly studied animal model of B. burgdorferi infection and surrogate for one of this pathogen's principle natural reservoir hosts, the white-footed deer mouse. Comparison will be made to the immune responses of humans with Lyme borreliosis. Our goal is to provide an understanding of the dynamics of the mammalian immune response during infection with B. burgdorferi and its relation to the outcomes in reservoir (mouse) and non-reservoir (human) hosts.

Characterization of a Borrelia miyamotoi membrane antigen (BmaA) for serodiagnosis of Borrelia miyamotoi disease.

Borrelia miyamotoi is a tick-borne pathogen that causes Borrelia miyamotoi disease (BMD), an emerging infectious disease of increasing public health significance. B. miyamotoi is transmitted by the same tick vector (Ixodes spp.) as B. burgdorferi sensu lato (s.l.), the causative agent of Lyme disease, therefore laboratory assays to differentiate BMD from Lyme disease are needed to avoid misdiagnoses and for disease confirmation. We previously performed a global immunoproteomic analysis of the murine host antibody response against B. miyamotoi infection to discover antigens that could serologically distinguish the two infections. An initial assessment identified a putative lipoprotein antigen, here termed BmaA, as a promising candidate to augment current research-based serological assays. In this study, we show that BmaA is an outer surface-associated protein by its susceptibility to protease digestion. Synthesis of BmaA in culture was independent of temperature at either 23 °C or 34 °C. The BmaA gene is present in two identical loci harbored on separate plasmids in North American strains LB-2001 and CT13-2396. bmaA-like sequences are present in other B. miyamotoi strains and relapsing fever borrelia as multicopy genes and as paralogous or orthologous gene families. IgM and IgG antibodies in pooled serum from BMD patients reacted with native BmaA fractionated by 2-dimensional gel electrophoresis and identified by mass spectrometry. IgG against recombinant BmaA was detected in 4 of 5 BMD patient serum samples as compared with 1 of 23 serum samples collected from patients with various stages of Lyme disease. Human anti-B. turicatae serum did not seroreact with recombinant BmaA suggesting a role as a species-specific diagnostic antigen. These results demonstrated that BmaA elicits a human host antibody response during B. miyamotoi infection but not in a tested group of B. burgdorferi-infected Lyme disease patients, thereby providing a potentially useful addition for developing BMD serodiagnostic tests.

Ticks, fleas and rodent-hosts analyzed for the presence of Borrelia miyamotoi in Slovakia: the first record of Borrelia miyamotoi in a Haemaphysalis inermis tick.

In Slovakia, little knowledge is available on the occurrence, hosts and vectors of Borrelia miyamotoi of the relapsing fever group. In the current study, 2160 questing and rodent-attached ticks of six species (Ixodes ricinus, Ixodes trianguliceps, Dermacentor marginatus, Dermacentor reticulatus, Haemaphysalis concinna and Haemaphysalis inermis), 279 fleas belonging to 9 species (Ctenophthalmus agyrtes, Ctenophthalmus solutus, Ctenophthalmus assimilis, Megabothris turbidus, Amalareus penicilliger, Hystrichopsylla orientalis, Ctenophthalmus uncinatus, Doratopsylla dasycnema and Nosopsyllus fasciatus) and skin biopsies from 245 small mammals belonging to eight species (Apodemus agrarius, Apodemus flavicollis, Apodemus uralensis, Myodes glareolus, Crocidura leucodon, Micromys minutus, Microtus arvalis, Microtus subterraneus) were screened for the presence of B. miyamotoi DNA. The overall prevalence of B. miyamotoi found in questing and rodent-attached ticks was 1.8% (23 positive/1260 examined) and 3.4% (31 positive/900 examined), respectively. Borrelia miyamotoi was detected in questing I. ricinus, rodent-attached I. ricinus and H. inermis ticks, and in one male of the common vole (M. arvalis) in different habitats (mainly rural) in eastern Slovakia. However, B. miyamotoi was not found in any of the tested fleas. Our findings indicate that rural habitats with different species of tick vectors and hosts are appropriate for the occurrence of B. miyamotoi.

Rapid clearance of Borrelia burgdorferi from the blood circulation.

BACKGROUND: Borrelia burgdorferi is a tick-borne spirochete that causes Lyme borreliosis (LB). After an initial tick bite, it spreads from the deposition site in the dermis to distant tissues of the host. It is generally believed that this spirochete disseminates via the hematogenous route. Borrelia persica causes relapsing fever and is able to replicate in the blood stream. Currently the exact dissemination pathway of LB pathogens in the host is not known and controversially discussed. METHODS: In this study, we established a strict intravenous infection murine model using host-adapted spirochetes. Survival capacity and infectivity of host-adapted B. burgdorferi sensu stricto (Bbss) were compared to those of B. persica (Bp) after either intradermal (ID) injection into the dorsal skin of immunocompetent mice or strict intravenous (IV) inoculation via the jugular vein. By in vitro culture and PCR, viable spirochetes and their DNA load in peripheral blood were periodically monitored during a 49/50-day course post-injection, as well as in various tissue samples collected at day 49/50. Specific antibodies in individual plasma/serum samples were detected with serological methods. RESULTS: Regardless of ID or IV injection, DNA of Bp was present in blood samples up to day 24 post-challenge, while no Bbss was detectable in the blood circulation during the complete observation period. In contrast to the brain tropism of Bp, Bbss spirochetes were found in ear, skin, joint, bladder, and heart tissue samples of only ID-inoculated mice. All tested tissues collected from IV-challenged mice were negative for traces of Bbss. ELISA testing of serum samples showed that Bp induced gradually increasing antibody levels after ID or IV inoculation, while Bbss did so only after ID injection but not after IV inoculation. CONCLUSIONS: This study allows us to draw the following conclusions: (i) Bp survives in the blood and disseminates to the host's brain via the hematogenous route; and (ii) Bbss, in contrast, is cleared rapidly from the blood stream and is a tissue-bound spirochete.

Molecular detection of reptile-associated Borrelia in Boa constrictor (Squamata: Boidae) from Veracruz, Mexico.

The reptile-associated Borrelia represent a monophyletic group of bacteria transmitted by several species of hard ticks, which has been reported to only infect amphibians and reptiles in Eurasia and Middle East, however, this bacterial group has not been studied in North America. The aim of this study was to assess the presence of Borrelia spirochetes in blood samples of native reptiles of Mexico. Blood samples were directly obtained from individuals, DNA extractions were performed using Chelex-100. The Borrelia detection was performed by conventional PCR. From 102 reptiles tested, only five individuals of Boa constrictor were positive for the presence of DNA of the reptile-associated Borrelia group. Supported by phylogenetic analysis, this study presents the first record of these spirochetes group in Mexico, and initial evidence of B. constrictor as a host of this group.

Controversies in bacterial taxonomy: The example of the genus Borrelia.

In this paper we survey key issues in bacterial taxonomy and review the literature regarding the recent genus separation proposed for the genus Borrelia. We discuss how information on members of the genus Borrelia is increasing but detailed knowledge on the relevant features is available only for a small subset of species. The data accumulated here show that there is considerable overlap in ecology, clinical aspects and molecular features between clades that argue against splitting of the genus Borrelia.

Genomes, expression profiles, and diversity of mitochondria of the White-footed Deermouse Peromyscus leucopus, reservoir of Lyme disease and other zoonoses.

The cricetine rodents Peromyscus leucopus and P. maniculatus are key reservoirs for several zoonotic diseases in North America. We determined the complete circular mitochondrial genome sequences of representatives of 3 different stock colonies of P. leucopus, one stock colony of P. maniculatus and two wild populations of P. leucopus. The genomes were syntenic with that of the murids Mus musculus and Rattus norvegicus. Phylogenetic analysis confirmed that these two Peromyscus species are sister taxa in a clade with P. polionotus and also uncovered a distinction between P. leucopus populations in the eastern and the central United States. In one P. leucopus lineage four extended regions of mitochondrial pseudogenes were identified in the nuclear genome. RNA-seq analysis revealed transcription of the entire genome and differences from controls in the expression profiles of mitochondrial genes in the blood, but not in liver or brain, of animals infected with the zoonotic pathogen Borrelia hermsii. PCR and sequencing of the D-loop of the mitochondrion identified 32 different haplotypes among 118 wild P. leucopus at a Connecticut field site. These findings help to further establish P. leucopus as a model organism for studies of emerging infectious diseases, ecology, and in other disciplines.

Two Cases of Borrelia miyamotoi Meningitis, Sweden, 2018.

We report 2 human cases of Borrelia miyamotoi disease diagnosed in Sweden, including 1 case of meningitis in an apparently immunocompetent patient. The diagnoses were confirmed by 3 different independent PCR assays and DNA sequencing from cerebrospinal fluid, supplemented by serologic analyses.

Borrelia spp. in ticks and birds from a protected urban area in Buenos Aires city, Argentina.

This study was aimed to know epidemiological aspects of Borrelia spp. in a protected urban area of Buenos Aires city, Argentina, where thousands of people visit this area for recreational purposes. Ticks were collected from vegetation, birds and dogs. Three hundred and forty birds belonging to 43 species, 41 genera, 18 families and six orders were captured (90.3% corresponded to the order Passeriformes). One hundred and twenty ticks were collected from 47 birds (13.8%) belonging to 10 species (23.2%), all of them from to the order Passeriformes (Emberizidae, Furnariidae, Parulidae, Thraupidae, Troglodytidae, Turdidae). Ticks were identified as Ixodes auritulus (56 larvae, 44 nymphs and 8 females) and Amblyomma aureolatum (1 larva and 11 nymphs). One thousand and ninety-one ticks collected from vegetation, 100 ticks collected from birds, and 89 ticks from dogs were tested for Borrelia infection by PCR trials targeting the flagellin (fla) and 16S rRNA genes. In addition, 101 blood and 168 tissue samples from birds were analyzed. Nine nymphs of A. aureolatum (2.1%) and four nymphs of I. auritulus (0.7%) collected from vegetation were positive. Five nymphs of A. aureolatum (45.4%), and five pools of larvae (minimum infection rate 13.5%), 18 nymphs (40.9%) and one female (14.3%) of I. auritulus collected on birds were also positive. The remaining samples were negative. The phylogenetic tree generated with fla sequences shows that seven of the eight different haplotypes of Borrelia detected in I. auritulus conform an independent lineage within the Borrelia burgdorferi sensu lato complex together with sequences of Borrelia sp. detected in I. auritulus from Canada and Uruguay. The fla sequences of Borrelia obtained from A. aureolatum and one sequence of a single specimen of I. auritulus conform a phylogenetic group with Borrelia turcica, Borrelia sp. isolated from a tortoise in Zambia, Borrelia spp. detected in Amblyomma maculatum from USA and Amblyomma longirostre from Brazil. The epidemiological risk that implies the infection with Borrelia genospecies associated with I. auritulus seems to be low because this tick is not aggressive to humans, but it helps to maintain borrelial spirochetes in the enzootic transmission cycles. The pathogenicity to humans of the Borrelia found in A. aureolatum is unknown; however, adults of this tick species are known to bite humans. This is the first report of the presence of Borrelia in A. aureolatum. Further investigations are necessary to know the risk of transmission of borreliosis by hard ticks in the study area.

The presence of Borrelia theileri in Argentina.

The presence of Borrelia theileri in Argentina is confirmed after recording the spirochete from a bovine in northern Argentina. The analysis of sequences of the flagellin gene (fla) and length of Borrelia spp. specimens on thick blood films shows that the local isolate clusters within a well-supported clade with B. theileri isolates from different geographical origins, confirming the presence of B. theileri in Argentina. The mean length of 30 specimens of B. theileri was 12.89 µm (standard deviation 2.88 µm, range 9.35-20.16 µm). The only known vector of Borrelia theileri in northern Argentina is the cattle tick Rhipicephalus microplus, therefore Borrelia infection should be regarded as a potential complication of other cattle tick-borne diseases such as babesiosis, especially on cattle introduced from areas free of R. microplus. The possibility of serologic cross-reaction with B. theileri must not be minimized in studies of other spirochaetes in the R. microplus infested region of Argentina.

A new Borrelia on the block: Borrelia miyamotoi - a human health risk?

BackgroundBorrelia miyamotoi clusters phylogenetically among relapsing fever borreliae, but is transmitted by hard ticks. Recent recognition as a human pathogen has intensified research into its ecology and pathogenic potential.AimsWe aimed to provide a timely critical integrative evaluation of our knowledge on B. miyamotoi, to assess its public health relevance and guide future research.MethodsThis narrative review used peer-reviewed literature in English from January 1994 to December 2018.ResultsBorrelia miyamotoi occurs in the world's northern hemisphere where it co-circulates with B. burgdorferi sensu lato, which causes Lyme disease. The two borreliae have overlapping vertebrate and tick hosts. While ticks serve as vectors for both species, they are also reservoirs for B. miyamotoi. Three B. miyamotoi genotypes are described, but further diversity is being recognised. The lack of sufficient cultivable isolates and vertebrate models compromise investigation of human infection and its consequences. Our understanding mainly originates from limited case series. In these, human infections mostly present as influenza-like illness, with relapsing fever in sporadic cases and neurological disease reported in immunocompromised patients. Unspecific clinical presentation, also occasionally resulting from Lyme- or other co-infections, complicates diagnosis, likely contributing to under-reporting. Diagnostics mainly employ PCR and serology. Borrelia miyamotoi infections are treated with antimicrobials according to regimes used for Lyme disease.ConclusionsWith co-infection of tick-borne pathogens being commonplace, diagnostic improvements remain important. Developing in vivo models might allow more insight into human pathogenesis. Continued ecological and human case studies are key to better epidemiological understanding, guiding intervention strategies.