Establishment of a digital PCR method for detection of Borrelia burgdorferi sensu lato complex DNA in cerebrospinal fluid.

Direct detection of Borrelia burgdorferi sensu lato bacteria in patient samples for diagnosis of Lyme neuroborreliosis (LNB) is hampered by low diagnostic sensitivity, due to few bacteria in cerebrospinal fluids (CSF) samples. Evaluation of novel molecular methods, including digital PCR (dPCR), as future tools in diagnostics of LNB is desirable. This study aimed to establish a dPCR assay and validate pre-PCR procedures for detection of Borrelia in CSF. Synthetic DNA fragments and cultured Borrelia reference strains were used during optimisation experiments. In addition, 59 CSF specimens from patients examined for LNB were included for clinical validation. The results showed that the pre-PCR parameters with the highest impact on Borrelia-specific dPCR method performance were incubation of the PCR-plate at 4 °C for stabilization of droplets, centrifugation for target concentration, quick-spin for dPCR rain reduction, and PCR inhibition by matrix components. Borrelia DNA in CSF was detected in one out of nine patients with LNB. Diagnostic sensitivity was determined to be 11.1% and specificity 100%. In conclusion, this study reports an optimized Borrelia-specific dPCR method for direct detection of Borrelia in CSF samples. The present study does not support the use of Borrelia-specific dPCR as a routine method for diagnosing LNB.

[Modern concepts on the pathogenesis of neuroborreliosis]. / Sovremennye predstavleniya o patogeneze neiroborrelioza.

The analysis of publications on the websites PubMed, ClinicalKey, devoted to the pathogenesis of neuroborreliosis (NB), using keywords for search: «pathogenesis of neuroborreliosis¼, «neuroborreliosis in children¼, «pathogenesis of Lyme disease¼, as well as an analysis of the results of the published research results of the staff of the Research Institute of Pediatric Infections, St-Petersburg, Russia is presented. Syndromes of early and late NB are more often observed among the forms without migrating erythema, and their development can be caused by all representatives of the species B. burgdorferi s.l. (B.b.), but more often - B. garinii, since it most effectively suppresses the factors of innate and adaptive immune response, reducing interferon production, phagocytosis and complement synthesis. The cause of immunosuppression with the development of NB may be simultaneous infection with several genovids and borrelia species or pathogens of other infections transmitted by Ixodes ticks, for example, infection with B.b. and tick-borne encephalitis virus. The ability to move along peripheral nerves, the change of surface antigens of the VlsE protein, as well as the formation of atypical cysts and granular forms allows B.b. to affect different structures of the peripheral and central nervous system, avoid an immune response and persist for a long time, causing chronic neuroinfection. Both the B.b. themselves, capable of being outside and inside glial cells and neurons, and inflammatory reactions developing in response to their introduction and associated with the synthesis of cytokines and chemokines and mimicry, cause damage to the vascular endothelium, vasculitis and impaired blood supply to the brain, demyelination, autoimmune inflammation and degeneration, leading to the development of NB syndromes, the spectrum of which varies depending on the duration of neuroinfection. In the development of NB and its outcomes, the following are also important: early initiation of treatment, the effectiveness of antibacterial drugs, the use of immunotropic agents that optimize the patient's immune response to the fight against neuroinfection, as well as the timely use of pathogenetic drugs, such as Cytoflavin, which have a complex effect on the vascular endothelium.

A novel Borrelia-specific real-time PCR assay is not suitable for diagnosing Lyme neuroborreliosis.

BACKGROUND: Diagnosing Lyme neuroborreliosis (LNB) is complicated by a lack of adequate test systems and by the complex culturing conditions required to grow the causative pathogens in the Borrelia sensu lato complex. Improved testing methods are urgently needed. Here, we evaluate the applicability of a novel commercially available Borrelia-specific real-time PCR assay to diagnose LNB. MATERIALS AND METHODS: The specificity and sensitivity of the novel alphaCube Borrelia real-time PCR assay (Mikrogen) and the well-tested Micro-Dx™ real-time PCR assay (Molzym) were evaluated in cerebrospinal fluid (CSF) spiked with known amounts of Borrelia garinii and CSF from 19 patients with definite or possible LNB. CSF from patients diagnosed with neurosyphilis or enterovirus meningitis served as controls. RESULTS: The alphaCube assay specifically identified Borrelia down to 93 B garinii cells/mL in spiked CSF samples. The Micro-Dx™ real-time PCR assay was able to identify the presence of bacteria down to 9300 cells/mL in spiked samples. In CSF from patients diagnosed with LNB the sensitivity of the alphaCube assay was 0.00 and 0.00 for the Micro-DX. CONCLUSION: Although the alphaCube Borrelia assay was able to identify down to 93 cells/mL in spiked CSF samples, the inability to identify Borrelia in CSF samples from patients with LNB suggests that this type of infection carries a bacterial load in CSF below this detection level. Based on these results, neither the alphaCube Borrelia real-time PCR assay nor the Micro-Dx™ real-time PCR assay can be recommended for routine diagnostics of LNB using CSF samples.

Comparison of the Euroimmun Borrelia 'antibody index' with Virotech immunoblot-based detection of intrathecal Borrelia antibody production for the diagnosis of Lyme neuroborreliosis.

For diagnosis of neuroborreliosis, calculation of the antibody index, based on Euroimmun Anti-Borrelia plus VlsE ELISA was compared to Virotech Borrelia Europe plus TpN17 immunoblot-based detection of Borrelia-specific intrathecal antibody production. CXCL13 results in cerebrospinal fluid were used to evaluate discordant results. A total of 64 serum/CSF pairs were analysed. Patients were classified according to European Federation of Neurological Societies criteria incorporating Virotech results. For the Euroimmun assay, a sensitivity of 100% and specificity of 94% was found. Agreement between the both tests was almost perfect (κ 0.81). Both methods are appropriate for the detection of Borrelia-specific intrathecal antibody production.

Acute facial nerve palsy in children in a Lyme disease-endemic area in the Netherlands.

We assessed the prevalence of Lyme neuroborreliosis in children with acute facial nerve palsy in a Lyme-endemic region and patient characteristics associated with this. All children visiting one of three participating hospitals between January 2010 and December 2016 were included in the study. Of 104 children referred to the hospital with facial nerve palsy, 43% had Lyme neuroborreliosis and 57% idiopathic facial palsy. Characteristics significantly associated with Lyme neuroborreliosis were headache (55% versus 18%), meningeal irritation (21% versus 5%), presentation in summer months (69% versus 37%), and a previous tick bite (33% versus 7%).

Lyme neuroborreliosis in Swedish children-PCR as a complementary diagnostic method for detection of Borrelia burgdorferi sensu lato in cerebrospinal fluid.

The aim of this study was to evaluate polymerase chain reaction (PCR) as a diagnostic method for the detection of Borrelia burgdorferi s.l. in CSF of Swedish children with LNB. This study was performed retrospectively on CSF and serum samples collected from children evaluated for LNB (n = 233) and controls with other specific neurological disorders (n = 59) in a Swedish Lyme endemic area. For anti-Borrelia antibody index, the IDEIA Lyme Neuroborreliosis kit (Oxoid) was used. Two in-house real-time PCR assays targeting the 16S rRNA gene were evaluated (TaqMan® and LUX™). Among patients classified as LNB cases (n = 102), five children (5%) were Borrelia PCR-positive in CSF with the TaqMan® assay. In the Non-LNB group (n = 131), one patient was Borrelia PCR positive with the TaqMan® assay. Among controls (n = 59), all CSF samples were PCR negative. When amplifying and sequencing ospA, we found B. garinii (n = 2), B. afzelii (n = 2), B. bavariensis (n = 1), and one untypable (n = 1). With the LUX™ technology, all CSF samples were PCR negative. The TaqMan® assay could detect only few cases (n = 6) of B. burgdorferi s.l. in CSF among children with LNB and the sensitivity was very low (5%). However, using larger CSF volumes and centrifugation of samples, the PCR technique could still be useful as a complementary diagnostic method when evaluating LNB. Furthermore, detection of spirochete DNA in clinical matrices, including CSF, is the method of choice for studying epidemiological aspects of LNB, a tick-borne emerging disease.

Conserved lysine residues in decorin binding proteins of Borrelia garinii are critical in adhesion to human brain microvascular endothelial cells.

Lyme borreliosis is a tick-borne disease caused by Borrelia burgdorferi sensu lato spirochetes (Lyme borreliae). When the disease affects the central nervous system, it is referred to as neuroborreliosis. In Europe, neuroborreliosis is most often caused by Borrelia garinii. Although it is known that in the host Lyme borreliae spread from the tick bite site to distant tissues via the blood vasculature, the adherence of Lyme borreliae to human brain microvascular endothelial cells has not been studied before. Decorin binding proteins are adhesins expressed on Lyme borreliae. They mediate the adhesion of Lyme borreliae to decorin and biglycan, and the lysine residues located in the binding site of decorin binding proteins are important to the binding activity. In this study, we show that lysine residues located in the canonical binding site can also be found in decorin binding proteins of Borrelia garinii, and that these lysines contribute to biglycan and decorin binding. Most importantly, we show that the lysine residues are crucial for the binding of Lyme borreliae to decorin and biglycan expressing human brain microvascular endothelial cells, which in turn suggests that they are involved in the pathogenesis of neuroborreliosis.

Cerebrospinal fluid oligoclonal bands in Neuroborreliosis are specific for Borrelia burgdorferi.

BACKGROUND: Cerebrospinal fluid (CSF) oligoclonal bands (OCB) occur in chronic or post-acute phase of inflammatory diseases of the central nervous system. OBJECTIVE: To determine whether CSF OCB in patients with neuroborreliosis (NB) are specific for borrelia burgdorferi senso lato. METHODS: We performed isoelectric focusing followed by immunoblotting in CSF of 10 NB patients and 11 controls (7 patients with multiple sclerosis, 2 patients with neuromyelitis optica spectrum disease, 1 patient with dementia and 1 patient with monoclonal gammopathy). Immunoblotting was performed using an uncoated as well as a borrelia antigen pre-coated nitrocellulose membrane (NCM). OCB were counted by visual inspection and photometric analysis. OCB were compared between uncoated und pre-coated NCM both in the NB and control group. For validation purposes inter-assay precision was determined by calculating the coefficient of variation (CV). RESULTS: Borrelia-specific OCB were found in the CSF of 9 NB patients and in none of the control subjects resulting in a sensitivity of 90% and a specificity of 100%. Number of NB specific OCB were 11±7 bands by photometric analyses compared to 9±5 bands by visual inspection. Validation experiments revealed an inconsistent inter-assay precision between visual and photometric analyses (NB uncoated: visual 28% versus photometric 14%, control subject uncoated: visual 16% versus photometric 24%). CONCLUSIONS: In CSF samples with positive OCB, Borrelia-specific bands were detected in almost all NB patients and in none of the control subjects. Inconsistent inter-assay precision may be explained by a poor comparability of visual and photometric approach.

Neutrophils and Neutrophil Extracellular Traps Regulate Immune Responses in Health and Disease.

Neutrophils are first responders of antimicrobial host defense and sterile inflammation, and therefore, play important roles during health and disease [...].

[A story of neuroborreliosis†: case report]. / Histoire de neuroborréliose†: étude d'un cas.

Neuroborreliosis affects approximately 15 % of people infected with Borrelia burgdorferi. The symptoms are very varied, which can sometimes delay the diagnosis. We can diagnose a neuroborreliosis in front of a compatible clinic and laboratory examinations, in particular a lumbar puncture showing a pleocytosis, an intrathecal synthesis of antibodies against B. burgdorferi, and an increased level of chemokine CKCL13. We present the case of a patient in whom the diagnosis was delayed in connection with non-specific symptoms and we review the latest recommendations in terms of treatment of neuroborreliosis.

La neuroborréliose touche entre 10 et 15 % des personnes infectées par Borrelia burgdorferi. Les symptômes sont très variés, ce qui peut parfois retarder le diagnostic. Celui-ci est posé en présence d'une clinique compatible et des examens de laboratoire, notamment une ponction lombaire montrant une pléocytose, une synthèse intrathécale d'anticorps contre B. burgdorferi et une augmentation de la chimiokine CXCL13. Nous présentons le cas d'un patient chez qui le diagnostic a mis du temps à être posé en lien avec des symptômes peu spécifiques et revoyons les dernières recommandations en termes de traitement de la neuroborréliose.

Spatial and temporal patterns of Lyme Neuroborreliosis on Funen, Denmark from 1995-2014.

In Europe, Lyme neuroborreliosis (LNB) is the most severe manifestation of Lyme borreliosis and has recently been added to the communicable disease surveillance list for EU/EEA by the European Commission. In Northern Europe, LNB is primarily caused by the spirochete Borrelia garinii and transmitted by the tick Ixodes ricinus. This Danish observational epidemiologic case-control study includes every identified LNB patient (n = 401) on Funen, Denmark, from 1995-2014. We display spatial and temporal LNB incidence variation, seasonal distribution of cases and local spatial case clustering. Seasonal patterns show LNB symptom-onset peaking in July and a significant seasonal difference in number of cases (p < 0.01). We found no significant change in seasonality patterns over time when dividing the study period into 5-year intervals. We identified a significant local geographical hot-spot of cases with a relative risk of 2.44 (p = 0.013). Analysis revealed a significantly shorter distance to nearest forest for cases compared with controls (p < 0.001). We present a novel map of the focal geographical distribution of LNB cases in a high endemic borreliosis area. Continued studies of case clustering in the epidemiology of LNB are of key importance in guiding intervention strategies.

[Epidemiological and clinical aspects of chronic neuroborreliosis with parkinsonism in the Yaroslavl region]. / Epidemiologicheskie i klinicheskie aspekty khronicheskogo neiroborrelioza s sindromom parkinsonizma v Yaroslavskoi oblasti.

OBJECTIVE: To study clinical and epidemiological features of chronic neuroborreliosis (CB) with parkinsonism (PS) in the Yaroslavl region. MATERIALS AND METHODS: The study included the main group of patients (n=5) with CB and PS of the average age of 61±3/4, the comparison group (n=6) with Parkinson's disease (PD) of the average age of 54.7±8.3 and a group of 6 healthy people. Diagnostic criteria of Lyme disease based on the recommendation of the US Centers for Disease Control and Prevention and criteria for the diagnosis of PS were used. PD was diagnosed by the criteria of the Parkinson's UK Brain Bank. Serological diagnosis of CB was carried out using immunoenzyme assay and immunoblotting in dynamics. The following scales were administered: HOEHN and YAHR, MMSE, MFI-20, CGI. All patients underwent MRI of the brain and spinal cord. RESULTS AND CONCLUSION: PS in patients with CB in the Yaroslavl region was observed in 2.17% of cases among patients with CB and amounted to 0.25% of all cases of the revealed PS. The features that complicate the diagnosis of PS within chronic borreliosis were: the absence of erythema migrans in the history of 80% of patients and in more than half of the cases of the acute period of the disease, the presence in most patients (60%) of asymmetric onset of the PS with rest tremor in 40% cases and a significant reduction in the severity of PS as a result of therapy with levodopa. The onset of complete regression of the clinical manifestations of PS and reduction of the titer of antibodies and obtaining the negative results during dynamic serological study in response to prolonged antibiotic therapy provided a basis to verify the diagnosis of borreliosis with PS in these patients.

Guidelines for diagnosis and treatment in neurology - Lyme neuroborreliosis.

Lyme borreliosis is the most common tick-borne infectious disease in Europe. A neurological manifestation occurs in 3-15% of infections and can manifest as polyradiculitis, meningitis and (rarely) encephalomyelitis. This S3 guideline is directed at physicians in private practices and clinics who treat Lyme neuroborreliosis in children and adults. Twenty AWMF member societies, the Robert Koch Institute, the German Borreliosis Society and three patient organisations participated in its development. A systematic review and assessment of the literature was conducted by the German Cochrane Centre, Freiburg (Cochrane Germany). The main objectives of this guideline are to define the disease and to give recommendations for the confirmation of a clinically suspected diagnosis by laboratory testing, antibiotic therapy, differential diagnostic testing and prevention.

Lyme neuroborreliosis in adults: A nationwide prospective cohort study.

The goal of this paper is to characterize the clinical presentation, serological results, current antibiotic treatment practice, including compliance with current European guidelines, and outcome in adults with Lyme neuroborreliosis (LNB) diagnosed at departments of infectious diseases in Denmark. Using a nationwide prospective cohort of patients with central nervous system infections, we identified all adults (≥ 18 years of age) treated for LNB at departments of infectious diseases in Denmark from 2015 through 2017. The database contains information on baseline demographics, history of tick bite, erythema migrans, clinical presentation, laboratory results of blood samples, and cerebrospinal (CSF) biochemistry (e.g. specific Borrelia burgdorferi sensu lato (s.l.) antibodies in serum, B. burgdorferi s.l. intrathecal antibody index) as well as antibiotic therapy. Outcome was assessed by the Glasgow Outcome Scale (GOS) and the presence of residual symptoms at follow-up one month after discharge. We included 194 LNB patients with a median age of 59 years (range 18-85 years, interquartile range [IQR] 47-69 years). The female-to-male ratio was 0.8. A total of 177 of 191 (93 %) of patients had early (second stage) LNB. A history of tick bite or erythema migrans was registered in 75 (39 %) and 49 (25 %) patients, respectively. The median duration of neurological symptoms before first hospital contact was 21 days (range 0-600 days, IQR 10-42 days). Predominant symptoms consisted of radicular pain in 135 of 194 (70 %), cranial nerve paresis in 88 of 194 (45 %), headache in 71 of 185 (38 %), and extremity paresis in 33 of 194 (17 %) patients. Serum-B. burgdorferi s.l. IgM and/or IgG antibodies were detectable in 166 of 181 (92 %) patients at the time of first CSF investigation. Median duration of antibiotic treatment was 14 days (range 10-35 days, IQR 14-21 days) and 59 (39 %) of the patients received intravenous ceftriaxone and/or benzylpenicillin G throughout treatment. At the 1-month follow-up, GOS was unfavorable (< 5) in 54 of 193 (28 %) patients. An unfavorable GOS score was more often registered in patients with ≥ 45 days of symptom duration (20 of 45 (44 %) vs. 34 of 145 (23 %); P = 0.006). In conclusion, a European cohort of adult patients with LNB diagnosed between 2015-2017 presented with classic symptoms and CSF findings. However, a substantial diagnostic delay was still observed. In disagreement with current guidelines, a substantial part of LNB patients were treated with antibiotics longer than 14 days and/or intravenously as route of administration.

Cytokines and chemokines in cerebrospinal fluid in relation to diagnosis, clinical presentation and recovery in children being evaluated for Lyme neuroborreliosis.

In Lyme neuroborrelios (LNB), the immune response has been in focus, but the association between different cytokines/chemokines and clinical manifestations in LNB patients has not been fully investigated. The aim of this study was to evaluate a large number of cytokines and chemokines in cerebrospinal fluid (CSF) in relation to diagnosis, clinical presentation and recovery in children being evaluated for LNB. MATERIALS AND METHODS: Pediatric patients (n = 105) were recruited at seven Swedish pediatric departments during 2010-14. Serum and CSF samples were drawn on admission, before start of antibiotic treatment. Patients diagnosed as Definite LNB or Possible LNB were categorized as LNBtot patients, all LNBtot patients presented with pleocytosis in CSF. Patients diagnosed as Non-LNB or Other diagnosis were categorized as Controlstot, all controlstot presented without pleocytosis in CSF. Multiplex bead array (Luminex) kits were used for analyses of 41 different cytokines/chemokines in CSF (Millipore). RESULTS: Twenty-eight cytokines/chemokines were detectable in CSF and the levels of 26 of these mediators were significantly higher in LNBtot patients than in Controlstot. In a discriminant analysis, a combination of four cytokines/chemokines (CXCL1, GM-CSF, IL-7 and IL-10) were shown to independently separate relevant patient groups. Furthermore, an IL-10/CXCL1 ratio was created and shown to have an improved diagnostic performance in distinguishing LNBtot vs Non-LNB patients, as compared to CXCL13 in CSF. No immune mediator differed significantly, when comparing LNBtot patients with different clinical presentation on admission or when comparing patients with or without recovery within 2 months of admission. CONCLUSION: A discriminant analysis was shown to be useful to distinguish the independently most important cytokines/chemokines (CXCL1, GM-CSF, IL-7 and IL-10) in CSF, in order to discriminate LNBtot patients from Non-LNB patients. An IL-10/CXCL1 ratio was shown to have a promising diagnostic profile with a better performance than the chemokine CXCL13 in CSF. However, further evaluation is required to address future possible usefulness of these cytokines and chemokines in laboratory diagnostics in LNB, including control groups with neuro-inflammation. No significant associations were found between CSF immune mediator levels and clinical presentation or recovery in pediatric LNB patients.

Atypical papillitis: An isolated manifestation of Lyme disease.

Lyme disease is a rare condition caused by the bacterium Borrelia burgdorferi. Despite typical symptoms including fever, headache, fatigue, and a characteristic skin rash, sometimes we cannot find those due to the lack of physician consultation in those early stages. If this disease is left untreated, infection could spread to the nervous system causing neuroborreliosis, an atypical and complicated manifestation of this disease. We present the case of an atypical papillitis, probably caused by this bacterium. We suspected this because of the results on the indirect test bloods and the improvement of the symptoms after treatment. This entity should be considered as a possible diagnosis of atypical optical neuropathies, particularly if it occurs in an endemic area.

Comparison of Neuroborreliosis Between Children and Adults.

BACKGROUND: Lyme neuroborreliosis (NB) is a tick-borne infectious disorder of the nervous system caused by Borrelia burgdorferi spirochetes. There are not many data available regarding the differences in the course of NB in children and adults. The aim of our study was to compare the clinical course of NB between children and adults. METHODS: Retrospective analysis of medical documentation of 181 patients with NB was performed. The patients were divided into 2 groups: Group I: 57 children with NB; Group II: 124 adults with NB. Medical data, such as patients' age, sex, place of living (residence), time from a tick bite, subjective complaints, general examination results, laboratory parameters and treatment were analyzed. RESULTS: In children, the most common symptoms were headache (89.5%), neck stiffness (64.9%) and nausea and vomiting (56.1%). In adults, the most common symptoms were headache (77.4%), facial nerve palsy (59.7%), neck stiffness (59.7%), vertigo (41.9%) and lumbosacral region pain (37.1%). Bannwarth's syndrome was observed in 10.5% of children and 36.3% of adults. In cerebrospinal fluid (CSF) in children, the pleocytosis at admission was higher than in adults, but protein concentration was significantly lower. There were no differences in percentage of mononuclear cells in CSF smear between the groups. In CSF examination after treatment, a decrease in pleocytosis and protein concentration was observed in both groups. Analysis of effectiveness of treatment mostly with third generation cephalosporins (defined as complete recovery) between the groups, calculated by the Kaplan-Meier method and compared with the use of the log-rank test, showed no significant differences between children and adults (log-rank P = 0.619). CONCLUSIONS: In children, NB more frequently presented as meningitis, and in adults in the form of Bannwarth's syndrome. CSF pleocytosis in children with NB was higher than in adults, while the protein concentration in children was lower. Outcomes in children and adults were favorable and did not differ after standard NB treatment.

Prospective comparison of two enzyme-linked immunosorbent spot assays for the diagnosis of Lyme neuroborreliosis.

Commercial cellular tests are used to diagnose Lyme borreliosis (LB), but studies on their clinical validation are lacking. This study evaluated the utility of an in-house and a commercial enzyme-linked immunosorbent spot (ELISpot) assay for the diagnosis of Lyme neuroborreliosis (LNB). Prospectively, peripheral blood mononuclear cells (PBMCs) were isolated from patients and controls and analysed using an in-house Borrelia ELISpot assay and the commercial LymeSpot assay. B. burgdorferi B31 whole cell lysate and a mixture of outer surface proteins were used to stimulate the PBMCs and the numbers of interferon-gamma-secreting T cells were measured. Results were evaluated using receiver operating characteristic (ROC) curve analysis. Eighteen active and 12 treated LNB patients, 10 healthy individuals treated for an early (mostly cutaneous) manifestation of LB in the past and 47 untreated healthy individuals were included. Both assays showed a poor diagnostic performance with sensitivities, specificities, positive and negative predictive values ranging from 44.4-66.7%, 42.0-72.5%, 21.8-33.3% and 80.5-87.0%, respectively. The LymeSpot assay performed equally poorly when the calculation method of the manufacturer was used. Both the in-house and the LymeSpot assay are unable to diagnose active LNB or to monitor antibiotic treatment success.

Lyme Neuroborreliosis in Children: Etiology and Comparison of Clinical Findings of Lyme Neuroborreliosis Caused by Borrelia garinii and Borrelia afzelii.

BACKGROUND: Information on the etiology of Lyme neuroborreliosis (LNB) in children in Europe and the influence of Borrelia burgdorferi sensu lato species isolated from cerebrospinal fluid (CSF) on clinical presentation of LNB in children are limited. METHODS: The study was monocentric. During its 17-year period, children younger than 15 years with presentation suggestive of LNB or confirmed Lyme borreliosis that had B. burgdorferi sensu lato isolated from CSF and had species of B. burgdorferi sensu lato identified by pulsed-field gel electrophoresis were included. Demographic and medical data were compared for children infected with Borrelia garinii to those infected with Borrelia afzelii. RESULTS: One hundred and fifty-three children had B. burgdorferi sensu lato isolated from CSF. In 71/113 (62.8%) and 42/113 (37.2%) patients, B. garinii and B. afzelii, respectively, were identified. Patients infected with B. garinii did not report symptoms suggestive of central nervous system (CNS) involvement or any other symptoms more often than patients infected with B. afzelii. Compared with children infected with B. afzelii, children infected with B. garinii had erythema migrans less often (18.3% vs. 45.2%) but had positive meningeal signs (69.0% vs. 38.1%), CSF lymphocytic predominance (97.1% vs. 75.0%), and elevated albumin CSF/serum quotient (80.6% vs. 50.0%) more often. CONCLUSIONS: In Slovenia, LNB in children is more often caused by B. garinii, followed by B. afzelii. The clinical picture of LNB in children caused by B. garinii is not more often suggestive of CNS involvement, but CNS inflammation is more pronounced in children infected with B. garinii, compared with children infected with B. afzelii.