Risk of heart failure among individuals tested for Borrelia burgdorferi sensu lato antibodies, and serum Borrelia burgdorferi sensu lato seropositive individuals; a nationwide population-based, registry-based matched cohort study.

BACKGROUND: Lyme borreliosis is a tick-borne disease caused by the bacterium Borrelia burgdorferi (Bb) sensu lato complex. Previous studies have suggested an association between Lyme borreliosis and heart failure, which have been suggested to be a possible manifestation of Lyme carditis. We aimed to investigate the risk of heart failure among individuals tested for serum Bb antibodies, and serum Bb seropositive individuals. METHODS: We performed a matched nationwide cohort study (Denmark, 1993-2020) and included 52,200 Bb seropositive individuals, and two age- and sex-matched comparison cohorts: 1) 104,400 Bb seronegative comparison cohort members, and 2) 261,000 population controls. We investigated the risk associated with 1) being tested for serum Bb antibodies, and 2) being Bb seropositive. Outcomes were: 1) a composite of heart failure, cardiomyopathy, and/or myocarditis diagnosis, and 2) redemption of cardiovascular medicine used for treatment of heart failure. We calculated short-term odds ratios (aOR) (within 1 month) and long-term hazard rates (aHR) (after 1 month) adjusted for age, sex, diabetes, pre-existing heart failure, and kidney disease. RESULTS: Compared with the population controls, individuals tested for Bb antibodies, regardless of the test result, had increased short-term risk of heart failure, cardiomyopathy, and myocarditis (aOR 8.3, 95 %CI: 6.7-10.2), and both increased short- and long-term risk of redemption of cardiovascular medicine (aOR 4.3, 95 %CI: 3.8-4.8, aHR 1.13, 95 % CI: 1.11-1.15). The Bb seropositive individuals had no increased short- or long-term risk of any outcome compared with Bb seronegative comparison cohort members. CONCLUSIONS: In conclusion, Bb antibody tests seemed to be performed in the diagnostic work-up of heart failure, but Bb seropositivity was not associated with heart failure.

Lyme arthritis in Western Europe: a multicentre retrospective study.

To characterize Lyme arthritis, with a focus on management, and outcome. Observational retrospective multicentre study in Western France, of all consecutive cases of Lyme arthritis, documented by Borrelia burgdorferi IgG on ELISA serological testing, confirmed by Western blot, with or without positive Borrelia PCR in synovial fluid, with no alternative diagnosis. We enrolled 52 patients (29 males), with a mean age of 43 ± 19.4 years. Most patients had monoarthritis (n = 43, 82.7%), involving the knee (n = 51, 98.1%), with a median delay between symptoms onset and Lyme arthritis diagnosis of 5 months (interquartile range, 1.5-8). Synovial fluid analysis yielded median white cell count of 16,000/mm3 (9230-40,500), and positive PCR in 16 cases (39%), for B. burgdorferi sensu stricto (n = 5), B. garinii (n = 5), B. afzelii (n = 3), and undetermined (n = 3). All patients received antibiotics, for a median duration of 28 days (21-30), with doxycycline (n = 44, 84.6%), ceftriaxone (n = 6, 11.5%), or amoxicillin (n = 2). Twelve patients (23.1%) also received intra-articular injection of glucocorticoids as first-line treatment. Of 47 patients with follow-up, 35 (74.5%) had complete resolution of Lyme arthritis. Lyme arthritis in Western Europe may be due to B. burgdorferi ss, B. afzelii, or B. garinii. Clinical presentation is similar to Lyme arthritis in North America (i.e. chronic knee monoarthritis), with low sensitivity of synovial fluid PCR (39%).

Borrelia Infection in Bank Voles Myodes glareolus Is Associated With Specific DQB Haplotypes Which Affect Allelic Divergence Within Individuals.

The high polymorphism of Major Histocompatibility Complex (MHC) genes is generally considered to be a result of pathogen-mediated balancing selection. Such selection may operate in the form of heterozygote advantage, and/or through specific MHC allele-pathogen interactions. Specific MHC allele-pathogen interactions may promote polymorphism via negative frequency-dependent selection (NFDS), or selection that varies in time and/or space because of variability in the composition of the pathogen community (fluctuating selection; FS). In addition, divergent allele advantage (DAA) may act on top of these forms of balancing selection, explaining the high sequence divergence between MHC alleles. DAA has primarily been thought of as an extension of heterozygote advantage. However, DAA could also work in concert with NFDS though this is yet to be tested explicitly. To evaluate the importance of DAA in pathogen-mediated balancing selection, we surveyed allelic polymorphism of MHC class II DQB genes in wild bank voles (Myodes glareolus) and tested for associations between DQB haplotypes and infection by Borrelia afzelii, a tick-transmitted bacterium causing Lyme disease in humans. We found two significant associations between DQB haplotypes and infection status: one haplotype was associated with lower risk of infection (resistance), while another was associated with higher risk of infection (susceptibility). Interestingly, allelic divergence within individuals was higher for voles with the resistance haplotype compared to other voles. In contrast, allelic divergence was lower for voles with the susceptibility haplotype than other voles. The pattern of higher allelic divergence in individuals with the resistance haplotype is consistent with NFDS favouring divergent alleles in a natural population, hence selection where DAA works in concert with NFDS.

Infection with Borrelia afzelii and manipulation of the egg surface microbiota have no effect on the fitness of immature Ixodes ricinus ticks.

Arthropod vectors carry vector-borne pathogens that cause infectious disease in vertebrate hosts, and arthropod-associated microbiota, which consists of non-pathogenic microorganisms. Vector-borne pathogens and the microbiota can both influence the fitness of their arthropod vectors, and hence the epidemiology of vector-borne diseases. The bacterium Borrelia afzelii, which causes Lyme borreliosis in Europe, is transmitted among vertebrate reservoir hosts by Ixodes ricinus ticks, which also harbour a diverse microbiota of non-pathogenic bacteria. The purpose of this controlled study was to test whether B. afzelii and the tick-associated microbiota influence the fitness of I. ricinus. Eggs obtained from field-collected adult female ticks were surface sterilized (with bleach and ethanol), which reduced the abundance of the bacterial microbiota in the hatched I. ricinus larvae by 28-fold compared to larvae that hatched from control eggs washed with water. The dysbiosed and control larvae were subsequently fed on B. afzelii-infected or uninfected control mice, and the engorged larvae were left to moult into nymphs under laboratory conditions. I. ricinus larvae that fed on B. afzelii-infected mice had a significantly faster larva-to-nymph moulting time compared to larvae that fed on uninfected control mice, but the effect was small (2.4% reduction) and unlikely to be biologically significant. We found no evidence that B. afzelii infection or reduction of the larval microbiota influenced the four other life history traits of the immature I. ricinus ticks, which included engorged larval weight, unfed nymphal weight, larva-to-nymph moulting success, and immature tick survival. A retrospective power analysis found that our sampling effort had sufficient power (> 80%) to detect small effects (differences of 5% to 10%) of our treatments. Under the environmental conditions of this study, we conclude that B. afzelii and the egg surface microbiota had no meaningful effects on tick fitness and hence on the R0 of Lyme borreliosis.

Cross-reactivity of antibody responses to Borrelia afzelii OspC: Asymmetry and host heterogeneity.

The tick-transmitted bacterium Borrelia afzelii consists of a number of antigenically different strains - often defined by outer surface protein C (OspC) genotype - that coexist at stable frequencies in host populations. To investigate how host antibody responses affect strain coexistence, we measured antibody cross-reactivity to three different OspC types (OspC 2, 3 and 9) in three different strains of laboratory mice (BALB/c, C3H and C57BL/6). The extent of cross-reactivity differed between mouse strains, being higher in C3H than BALB/c and C57BL/6. In one of three pairwise comparisons of OspC types (OspC2 vs OspC9), there was evidence for asymmetry of cross-reactivity, with antibodies to OspC2 cross-reacting more strongly with OspC9 than vice versa. These results indicate that the extent of antibody-mediated competition between OspC types may depend on the composition of the host population, and that such competition may be asymmetric. We discuss the implications of these results for understanding the coexistence of OspC types.

Investigating BB0405 as a novel Borrelia afzelii vaccination candidate in Lyme borreliosis.

BB0405 is a surface exposed Borrelia burgdorferi protein and its vaccination protected mice against B. burgdorferi infection. As BB0405 is highly conserved across different B. burgdorferi sensu lato species, we investigated whether vaccination with recombinant BB0405 or through intradermal bb0405 DNA tattoo vaccination could provide protection against different Borrelia species, specifically against Borrelia afzelii, the predominant B. burgdorferi sensu lato genospecies causing Lyme borreliosis across Eurasia. We immunized C3H/HeN mice with recombinant BB0405 or with a codon-optimized bb0405 DNA vaccine using the pVAC plasmid and immunized corresponding control groups mice with only adjuvant or empty vectors. We subsequently subjected these immunized mice to a tick challenge with B. afzelii CB43-infected Ixodes ricinus nymphs. Upon vaccination, recombinant BB0405 induced a high total IgG response, but bb0405 DNA vaccination did not elicit antibody responses. Both vaccine formulations did not provide protection against Borrelia afzelii strain CB43 after tick challenge. In an attempt to understand the lack of protection of the recombinant vaccine, we determined expression of BB0405 and showed that B. afzelii CB43 spirochetes significantly and drastically downregulate the expression of BB0405 protein at 37 °C compared to 33 °C, where as in B. burgdorferi B31 spirochetes expression levels remain unaltered. Vaccination with recombinant BB0405 was previously shown to protect against B. burgdorferi sensu stricto. Here we show that vaccination with either recombinant BB0405 (or non-immunogenic bb0405 DNA), despite being highly conserved among B. burgdorferi sl genospecies, does not provide cross-protection against B. afzelii, mostly likely due to downregulation of this protein in B. afzelii in the mammalian host.

Immunoserological Diagnosis of Human Borrelioses: Current Knowledge and Perspectives.

Spirochetes of the genus Borrelia are divided into relapsing fever borreliae and Lyme disease borreliae. Immunoserological assays have been poorly developed for relapsing fever borreliae, where direct detection methods are more adapted to the pathophysiology of these infections presenting with massive bacteraemia. However, emergence of the novel agent of relapsing fever B. miyamotoi has renewed interest in serology in this context. In Lyme disease, because direct detection methods show low sensitivity, serology plays a central role in the diagnostic strategy. This diagnostic strategy is based on a two-tier methodology involving a first test (ELISA) with high sensitivity and acceptable specificity and a second, more specific test (western blot) for diagnostic confirmation. The most frequent limitations and pitfalls of serology are cross reactions, false IgM positivity, a seronegative window period at the early time of the infection, and serologic scars with a suspicion of reinfection. International guidelines have thus been proposed to avoid these difficulties with interpretation. Finally, unconventional diagnostic tests have been developed recently in the context of a highly publicized disease, with widely varying results, some of which have no available evidence-based data. New two-tier testing strategies using two ELISA tests (C6 and WCS for example) to replace immunoblot are currently proposed by some authors and guidelines, and promising new tests such as CXCL-13 in CSF are promising tools for the improvement of the diagnosis of Lyme borreliosis.

History of Lyme Disease as a Predictor of Atrial Fibrillation.

In many cases, atrial fibrillation (AF) is associated with a history of cardiac inflammation. One of the potential pathogens responsible for atrial inflammation might be Borrelia burgdorferi - a pathogen involved in Lyme carditis. This study aimed to assess whether the serological history of Borrelia infection was associated with the risk of AF. The study included 113 AF patients and 109 patients in sinus rhythm. All patients underwent a clinical evaluation, echocardiography and had their blood taken for the assessment of anti-Borrelia IgG antibodies. Patients with AF compared with the non-AF group had more often serological signs of Borrelia infection (34.5% vs 6.4%; p <0.0001). The multivariate analysis showed that positive results for anti-Borrelia IgG antibodies were a strong independent predictor of AF (odds ratio 8.21; 95% confidence interval 3.08 to 21.88; p < 0.0001). In conclusion, presented data show that exposure to Borrelia spp. infection is associated with an increased risk of AF. Whether the early treatment of Lyme disease lowers the risk of AF development remains to be explored.

Serological diagnostics of Lyme borreliosis: comparison of assays in twelve clinical laboratories in Northern Europe.

Lyme borreliosis (LB), caused by spirochetes belonging to the Borrelia burgdorferi sensu lato complex, is the most common tick-borne infection in Europe. Laboratory diagnosis of LB is mainly based on the patients' medical history, clinical signs and symptoms in combination with detection of Borrelia-specific antibodies where indirect enzyme-linked-immunosorbent assay (ELISA) is the most widely used technique. The objective of the study was to evaluate and compare the diagnostic accuracy (sensitivities and specificities) of serological tests that are currently in use for diagnosis of LB in clinical laboratories in Northern Europe, by use of a large serum panel. The panel consisted of 195 serum samples from well-characterized and classified patients under investigation for clinically suspected LB (n = 59) including patients with Lyme neuroborreliosis, Lyme arthritis, acrodermatitis chronica atrophicans, erythema migrans or other diseases (n = 112). A total of 201 serum samples from healthy blood donors were also included. The panel (396 serum samples altogether) was sent to 12 clinical laboratories (using five different ELISA methods) as blinded for group affiliation and the laboratories were asked to perform serological analysis according to their routine procedure. The results from the study demonstrated high diagnostic concordance between the laboratories using the same diagnostic assay and lower diagnostic concordance between laboratories using different diagnostic assays. For IgG, the results were in general rather homogenous and showed an average sensitivity of 88% (range 85-91%) compared to IgM which showed lower average sensitivity of 59% (range 50-67%) and more heterogeneous results between assays and laboratories.

Neutrophil extracellular traps in the central nervous system hinder bacterial clearance during pneumococcal meningitis.

Neutrophils are crucial mediators of host defense that are recruited to the central nervous system (CNS) in large numbers during acute bacterial meningitis caused by Streptococcus pneumoniae. Neutrophils release neutrophil extracellular traps (NETs) during infections to trap and kill bacteria. Intact NETs are fibrous structures composed of decondensed DNA and neutrophil-derived antimicrobial proteins. Here we show NETs in the cerebrospinal fluid (CSF) of patients with pneumococcal meningitis, and their absence in other forms of meningitis with neutrophil influx into the CSF caused by viruses, Borrelia and subarachnoid hemorrhage. In a rat model of meningitis, a clinical strain of pneumococci induced NET formation in the CSF. Disrupting NETs using DNase I significantly reduces bacterial load, demonstrating that NETs contribute to pneumococcal meningitis pathogenesis in vivo. We conclude that NETs in the CNS reduce bacterial clearance and degrading NETs using DNase I may have significant therapeutic implications.

Investigation of the performance of serological assays used for Lyme disease testing in Australia.

Spirochaetes of the Borrelia burgdorferi sensu lato complex, which includes those that cause Lyme disease, have not been identified in Australia. Nevertheless, Australian patients exist, some of whom have not left the country, who have symptoms consistent with so-called "chronic Lyme disease". Blood specimens from these individuals may be tested in Australian laboratories and in specialist laboratories outside Australia and sometimes conflicting results are obtained. Such discrepancies cause the patients to question the results from the Australian laboratories and seek assistance from the Australian Government in clarifying why the discrepancies occur. The aim of this study was to determine the level of agreement in results between commonly used B. burgdorferi serology assays in specimens of known status, and between results reported by different laboratories when they use the same serology assay. Five immunoassays and five immunoblots used in Australia and elsewhere were examined for the detection of IgG antibodies to Borrelia burgdorferi sensu lato. Predominantly, archived specimens previously tested for Lyme disease were used for the study and included 639 contributed by seven clinical laboratories located either in Australia or in areas endemic for Lyme disease. Also included were 308 prospectively collected Australian blood donor specimens. All clinical specimens were tested in all 10 assays whereas blood donor specimens were tested in all immunoassays and a subset was tested on immunoblots. With the exception of one immunoblot, the results between the assays agreed with each other in a known positive specimen population ≥ 77% of the time and in a known negative population, 88% of the time or greater. The test results obtained during the study were different from the participating laboratory's less than 2% of the time when the same assay was used. These findings suggest that discordance in results between laboratories is more likely due to variation in algorithms or in the use of assays with different sensitivities or specificities rather than conflicting results being reported from the same assay in different laboratories. In the known negative population, specificities of the immunoassays ranged between 87.7% and 99.7%. In Australia's low prevalence population, this would translate to a positive predictive value of < 4%.

Values of diagnostic tests for the various species of spirochetes.

Bacteria of the Borrelia burgdorferi sensu lato complex, responsible for Lyme disease, are members of the spirochetes phylum. Diagnostic difficulties of Lyme disease are partly due to the characteristics of spirochetes as their culture is tedious or even impossible for some of them. We performed a literature review to assess the value of the various diagnostic tests of spirochetes infections of medical interest such as Lyme borreliosis, relapsing fever borreliae, syphilis, and leptospirosis. We were able to draw similarities between these four infections. Real-time PCR now plays an important role in the direct diagnosis of these infections. However, direct diagnosis remains difficult because of a persistent lack of sensitivity. Serological testing is therefore crucial in the diagnostic process. All currently available diagnostic tools are imperfect, with a potential risk of false positive and false negative results depending on the clinical context. Physicians should always take into consideration the clinical and epidemiological context when Lyme disease, relapsing fever borreliae, syphilis, and leptospirosis are suspected.

Polymorphic factor H-binding activity of CspA protects Lyme borreliae from the host complement in feeding ticks to facilitate tick-to-host transmission.

Borrelia burgdorferi sensu lato (Bbsl), the causative agent of Lyme disease, establishes an initial infection in the host's skin following a tick bite, and then disseminates to distant organs, leading to multisystem manifestations. Tick-to-vertebrate host transmission requires that Bbsl survives during blood feeding. Complement is an important innate host defense in blood and interstitial fluid. Bbsl produces a polymorphic surface protein, CspA, that binds to a complement regulator, Factor H (FH) to block complement activation in vitro. However, the role that CspA plays in the Bbsl enzootic cycle remains unclear. In this study, we demonstrated that different CspA variants promote spirochete binding to FH to inactivate complement and promote serum resistance in a host-specific manner. Utilizing a tick-to-mouse transmission model, we observed that a cspA-knockout B. burgdorferi is eliminated from nymphal ticks in the first 24 hours of feeding and is unable to be transmitted to naïve mice. Conversely, ectopically producing CspA derived from B. burgdorferi or B. afzelii, but not B. garinii in a cspA-knockout strain restored spirochete survival in fed nymphs and tick-to-mouse transmission. Furthermore, a CspA point mutant, CspA-L246D that was defective in FH-binding, failed to survive in fed nymphs and at the inoculation site or bloodstream in mice. We also allowed those spirochete-infected nymphs to feed on C3-/- mice that lacked functional complement. The cspA-knockout B. burgdorferi or this mutant strain complemented with cspA variants or cspA-L246D was found at similar levels as wild type B. burgdorferi in the fed nymphs and mouse tissues. These novel findings suggest that the FH-binding activity of CspA protects spirochetes from complement-mediated killing in fed nymphal ticks, which ultimately allows Bbsl transmission to mammalian hosts.

Predictive value of Borrelia burgdorferi IgG antibody levels in patients referred to a tertiary Lyme centre.

A two-step testing strategy is recommended in serological testing for Lyme borreliosis; positive and indeterminate enzyme-linked immunosorbent assay (ELISA) results are confirmed with immunoblots. Several ELISAs quantify the concentration of antibodies tested, however, no recommendation exists for an upper cut-off value at which an IgG ELISA is sufficient and the immunoblot can be omitted. The study objective was to determine at which IgG antibody level an immunoblot does not have any additional predictive value compared to ELISA results. Data of adult patients who visited a tertiary Lyme centre between 2008 and 2014 were analysed. Both an ELISA (Enzygnost Lyme link VlsE IgG) and immunoblot (recomLine blot Borrelia) were performed. Clinical data were extracted from the patient's digital medical record. Positive predictive values (PPVs) for either previous or active infection with Borrelia burgdorferi s.l. were calculated for different cut-off ELISA IgG antibody levels where the immunoblot was regarded as reference test. In total, 1454 patients were included. According to the two-step test strategy, 486 (33%), 69 (5%) and 899 (62%) patients had positive, indeterminate and negative Borrelia IgG serology, respectively. At IgG levels of 500 IU/ml and higher, all immunoblots were positive, resulting in a 100% PPV (95% CI: 97.0-100). At IgG levels of 200 IU/ml and higher, the PPV was 99.3% (95% CI: 97.4-99.8). In conclusion, at IgG levels of 200 IU/ml and higher, an ELISA was sufficient to detect antibodies to Borrelia burgdorferi s.l. At those IgG levels, a confirmatory immunoblot may be omitted in patients referred to a tertiary Lyme centre. Before these results can be implemented in routine diagnosis of Lyme borreliosis, confirmation of the results is necessary in other patient populations and using other quantitative ELISAs and immunoblots.

A serological study of antibodies to Anaplasma phagocytophilum and Borrelia burgdorferi sensu lato in the sera of healthy individuals collected two decades apart.

Sera collected from healthy individuals from the general population in the Czech Republic during repeated cross-sectional surveys were analyzed. Samples collected in the same six districts in two time periods, 1978-1989 and 2001, were compared. The study subjects were divided into six age categories between 10 and 59 years. Overall, 434 samples from 1978-1989 and 270 samples from 2001 were screened for Anaplasma phagocytophilum (AP) and Borrelia burgdorferi sensu lato (BB). The anti-AP positivity rates were 13.1% and 11.5% in the first and second period, respectively, and did not differ significantly between the periods (P = 0.559). The anti-BB antibodies were detected in 33.9% and 14.8% of study subjects, respectively. The positivity rates were significantly lower in the second period (P<0.001). No considerable changes were observed in the sex distribution of positive findings between the two periods. The highest positivity rates of anti-AP antibodies were found in the 10-14 year age group: 16.0% in 1978-1989 and 16.7% in 2001. The age distribution of the anti-AP antibody positivity rates did not change substantially (P = 0.872). In 1978-1989, the lowest anti-BB antibody positivity rate (26.7%) was found in the 10-14 year age group, with a gradual increase with age to 41.1% in 50-59 year-olds. In 2001, the positivity rate in the 10-14 year age group was 26.2% and was not significantly different from that in the first period (P = 0.955). However, the positivity rates in the older age groups 15-59 years decreased significantly (P<0.001) and varied between 8.3% and 15.1%.

Cutaneous Lyme borreliosis: Guideline of the German Dermatology Society.

This guideline of the German Dermatology Society primarily focuses on the diagnosis and treatment of cutaneous manifestations of Lyme borreliosis. It has received consensus from 22 German medical societies and 2 German patient organisations. It is the first part of an AWMF (Arbeitsgemeinschaft der Wissenschaftlichen Medizinischen Fachgesellschaften e.V.) interdisciplinary guideline: "Lyme Borreliosis - Diagnosis and Treatment, development stage S3". The guideline is directed at physicians in private practices and clinics who treat Lyme borreliosis. Objectives of this guideline are recommendations for confirming a clinical diagnosis, recommendations for a stage-related laboratory diagnosis (serological detection of IgM and IgG Borrelia antibodies using the 2-tiered ELISA/immunoblot process, sensible use of molecular diagnostic and culture procedures) and recommendations for the treatment of the localised, early-stage infection (erythema migrans, erythema chronicum migrans, and borrelial lymphocytoma), the disseminated early-stage infection (multiple erythemata migrantia, flu-like symptoms) and treatment of the late-stage infection (acrodermatitis chronica atrophicans with and without neurological manifestations). In addition, an information sheet for patients containing recommendations for the prevention of Lyme borreliosis is attached to the guideline.

The recomBead Borrelia antibody index, CXCL13 and total IgM index for laboratory diagnosis of Lyme neuroborreliosis in children.

For laboratory diagnostics of Lyme neuroborreliosis (LNB), the recomBead Borrelia antibody index (AI) assay has shown promising results in a mixed age population, but has not previously been evaluated with specific focus on paediatric patients. The aim of the study was to evaluate the recomBead Borrelia AI assay in cerebrospinal fluid (CSF) for the laboratory diagnosis of LNB in children. We also wanted to explore whether early markers, such as CXCL13 in CSF and/or total IgM index could be useful as complementary diagnostic tools. Children being evaluated for LNB in a Swedish Lyme endemic area were included in the study (n = 146). Serum and CSF were collected on admission. Patients with other specific diagnoses were controls (n = 15). The recomBead Borrelia AI assay and the recomBead CXCL13 assay (Mikrogen) were applied together with total IgM index. The overall sensitivity for recomBead Borrelia AI (IgM and IgG together) was 74% and the specificity was 97%. However, the highest sensitivity (91%) at an acceptable level of specificity (90%) was obtained by recomBead Borrelia AI together with CXCL13 and total IgM index, showing a positive predictive value of 84% and a negative predictive value of 95%. Thus, the recomBead Borrelia AI assay performs with moderate sensitivity and high specificity in paediatric LNB patients. The major advantage seems to be increased sensitivity in the possible LNB group compared to the IDEIA assay. The diagnostic sensitivity may be further increased by using a combination of early markers, such as CXCL13 in CSF and total IgM index.