Whole genome sequencing of human Borrelia burgdorferi isolates reveals linked blocks of accessory genome elements located on plasmids and associated with human dissemination.

Lyme disease is the most common vector-borne disease in North America and Europe. The clinical manifestations of Lyme disease vary based on the genospecies of the infecting Borrelia burgdorferi spirochete, but the microbial genetic elements underlying these associations are not known. Here, we report the whole genome sequence (WGS) and analysis of 299 B. burgdorferi (Bb) isolates derived from patients in the Eastern and Midwestern US and Central Europe. We develop a WGS-based classification of Bb isolates, confirm and extend the findings of previous single- and multi-locus typing systems, define the plasmid profiles of human-infectious Bb isolates, annotate the core and strain-variable surface lipoproteome, and identify loci associated with disseminated infection. A core genome consisting of ~900 open reading frames and a core set of plasmids consisting of lp17, lp25, lp36, lp28-3, lp28-4, lp54, and cp26 are found in nearly all isolates. Strain-variable (accessory) plasmids and genes correlate strongly with phylogeny. Using genetic association study methods, we identify an accessory genome signature associated with dissemination in humans and define the individual plasmids and genes that make up this signature. Strains within the RST1/WGS A subgroup, particularly a subset marked by the OspC type A genotype, have increased rates of dissemination in humans. OspC type A strains possess a unique set of strongly linked genetic elements including the presence of lp56 and lp28-1 plasmids and a cluster of genes that may contribute to their enhanced virulence compared to other genotypes. These features of OspC type A strains reflect a broader paradigm across Bb isolates, in which near-clonal genotypes are defined by strain-specific clusters of linked genetic elements, particularly those encoding surface-exposed lipoproteins. These clusters of genes are maintained by strain-specific patterns of plasmid occupancy and are associated with the probability of invasive infection.

Assessment of three criteria to establish borrelial infection in suspected lyme neuroborreliosis.

PURPOSE: Diagnosis of (European) Lyme neuroborreliosis has been based on clinical presentation, cerebrospinal fluid (CSF) pleocytosis and demonstration of intrathecal borrelial antibody synthesis (ITBAS) to document Borrelia burgdorferi s. l. INFECTION: It is not known if other criteria to document Borrelia infection may contribute to the diagnosis. METHODS: We compared the sensitivity of three individual criteria (ITBAS, CSF Borrelia culture, and the presence of erythema migrans [EM]) to confirm the diagnosis of early Lyme neuroborreliosis in 280 patients ≥ 15 years of age evaluated at a Lyme borreliosis outpatient clinic in Slovenia. The patients had either radicular pain of new onset or involvement of a cranial nerve but without radicular pain, each in conjunction with CSF pleocytosis. Evaluation was of patients who had each of the three confirmatory criteria assessed, and for whom at least one criterion was positive. RESULTS: Analysis of 280 patients, 120 women and 160 men, median age 57 (range 15-84) years, revealed that ITBAS was the most frequently observed positive criterion (85.4%), followed by EM (52.9%), and by a positive CSF Borrelia culture (9.6%). Of the 280 patients, 154 (55%) met only one criterion (43.2% ITBAS only, 10.7% EM only, and 1.1% positive CSF culture only), whereas 42.1% met two criteria. Only 2.9% of patients were positive by all three criteria. CONCLUSION: Although ITBAS was the most frequent criterion for confirmation for Borrelia infection, the presence of EM alone confirmed an additional 10.7% of patients and a positive CSF Borrelia culture alone added another 1.1%.

Colocalization of Radicular Pain and Erythema Migrans in Patients With Bannwarth Syndrome Suggests a Direct Spread of Borrelia Into the Central Nervous System.

BACKGROUND: There is a general assumption that after deposition into skin, Lyme borreliae disseminate hematogenously to other organs, resulting in extracutaneous manifestations of Lyme borreliosis, including Lyme neuroborreliosis. However, our experience over the past 40 years, along with several published case reports that observed colocalization of radicular pain and erythema migrans (EM) in patients with borrelial meningoradiculoneuritis (Bannwarth syndrome), argues against hematogenous dissemination in Lyme neuroborreliosis. METHODS: We compared the location of EM in 112 patients with Bannwarth syndrome to 12315 EM patients without neurological involvement. Moreover, we assessed the colocalization of EM and radicular pain in patients with Bannwarth syndrome. RESULTS: Compared to >12000 EM patients without neurological involvement, patients with Bannwarth syndrome had a significantly higher frequency of EM on head/neck (6% vs 1%; P=.0005) and trunk (47% vs 24%; P<.0001), similar frequency on arms (16% vs 16%; P=.91), but lower frequency on legs (30% vs 59%; P<.0001). Moreover, in 79% (89/112) of patients the site of EM matched the dermatomes of radicular pain. The odds for a congruent location of EM and radicular pain were highly significant with the highest odds ratios (OR) observed for head (OR=221), followed by neck (OR=159), legs (OR=69), arms (OR=48), and trunk (OR=33). CONCLUSIONS: The greater frequency of EM on head/neck and trunk and the colocalization of EM with radicular pain in patients with Bannwarth syndrome suggest that central nervous system involvement in Lyme neuroborreliosis is due to a retrograde spread of borrelia from skin to the spinal cord via peripheral nerves.

Borrelial Lymphocytoma in Adult Patients.

BACKGROUND: Information on the course and outcome of borrelial lymphocytoma (BL) is limited. METHODS: The study comprised 144 adult patients (75 female, 69 male; median age, 49 years) who had BL diagnosed at a single center between 1986 and 2014 and were followed up for 1 year. RESULTS: BL was located on the breast in 106 patients (73.6%), on the ear lobe in 27 (18.8%), and elsewhere in 11 (7.6%). The median duration of BL before diagnosis was 27 days (interquartile range [IQR], 9-68 days). Concomitant erythema migrans was registered in 104 of 144 patients (72.2%); other objective manifestations of Lyme borreliosis (LB) were present in 11 (7.6%). Immunoglobulin M and/or G borrelial serum antibodies were present in 72 patients (50%). Borreliae were isolated from BL lesions in 14 of 42 patients (33.3%) who had not received antibiotics before skin biopsy. Of 13 typed Borrelia strains, 11 were B. afzelii, 1 was B. garinii, and 1 was B. bissettii The median duration of BL after starting antibiotic treatment was 21 days ([IQR], 10-30 days); the average duration was longer in patients who were older, had longer BL duration before treatment, or had signs of disseminated LB. Treatment failure occurred in 14 of 144 patients (9.7%). Patients with signs or symptoms of disseminated LB before treatment had nearly 4 times higher odds of treatment failure (95% confidence interval, 1.22-13.07) than those without such symptoms. All patients with treatment failure had uneventful outcome after retreatment. CONCLUSIONS: BL is a rare manifestation of early localized LB. Fourteen-day antibiotic treatment, as used for erythema migrans, is effective.

Course and outcome of Early Lyme borreliosis in patients with hematological malignancies.

Patients with erythema migrans and underlying hematological malignancy more often had signs of disseminated Lyme borreliosis and more frequently needed antibiotic retreatment than sex-, age-, and antibiotic treatment-matched immunocompetent persons with erythema migrans. However, the outcome was excellent in both groups.

Microbiologic Findings in a Cohort of Patients with Erythema Migrans.

Erythema migrans (EM) is the initial and the most frequent clinical manifestation of Lyme borreliosis (LB). Herein, we report on the capacity of culture and serology for the demonstration of Borrelia infection in a cohort of 292 patients diagnosed with typical EM at a single medical center. The median duration of EM at diagnosis was 12 days, and the largest diameter was 16 cm; 252 (86.3%) patients presented with solitary EM, whereas 40 (13.7%) had multiple EM. A total of 95/292 (32.5%) patients had positive IgM, and 169 (57.9%) had positive IgG serum antibodies; the Borrelia isolation rate was 182/292 (62.3%). The most frequent species by far was B. afzelii (142/148, 95.9%) while B. garinii (2.7%) and B. burgdorferi s.s. (1.4%) were rare. IgM seropositivity was associated with a younger age, multiple EM and the absence of underlying chronic illness; IgG seropositivity was associated with the duration of EM at diagnosis, the diameter of the EM, having had a previous episode of LB and the absence of symptoms at the site of the EM. Furthermore, the Borrelia isolation rate was statistically significantly lower in patients with positive Borrelia IgM antibodies. Although microbiologic analyses are not needed for the diagnosis of typical EM, they enable insights into the etiology and dynamics of the immune response in the course of early LB.

Why Is the Duration of Erythema Migrans at Diagnosis Longer in Patients with Lyme Neuroborreliosis Than in Those without Neurologic Involvement?

In prior studies, the skin lesion erythema migrans (EM) was present for a longer time period before diagnosis of concomitant borrelial meningoradiculoneuritis (Bannwarth's syndrome) compared to EM patients without neurologic symptoms. To determine if this observation pertains to other manifestations of Lyme neuroborreliosis (LNB), we compared EM characteristics in patients with borrelial meningoradiculoneuritis (n = 122) to those with aseptic meningitis without radicular pain (n = 72 patients), and to patients with EM but without neurologic involvement (n = 12,384). We also assessed factors that might impact duration. We found that the duration of EM at diagnosis in patients with borrelial meningoradiculoneuritis was not significantly different compared with those with LNB without radicular pain (34 vs. 26 days; p = 0.227). The duration of EM for each of these clinical presentations of LNB, however, was significantly longer than in patients with EM without LNB (10 days; p < 0.001). Contributing factors to this difference might have been that patients with LNB failed to recognize that they had EM or were unaware of the importance of not delaying antibiotic treatment for EM. In conclusion, the duration of the EM skin lesion in EM patients with LNB is longer than in patients with just EM, irrespective of the type of LNB.

Suspected early Lyme neuroborreliosis in patients with erythema migrans.

BACKGROUND: Our objective was to obtain data on patients with erythema migrans (EM) who have symptoms/signs suggesting nervous system involvement and to compare epidemiologic, clinical, and microbiologic findings in patients with and without cerebrospinal fluid (CSF) pleocytosis. METHODS: Adult patients with EM and suspected early Lyme neuroborreliosis were included in this study. RESULTS: Of 161 patients, 31 (19%) had elevated and 130 (81%) had normal CSF cell counts. In contrast to patients with normal CSF cell counts, those with pleocytosis (1) more often reported radicular pain and more often presented with meningeal signs but less frequently complained of malaise; (2) had larger EM skin lesions despite similar duration; (3) more commonly had Borrelia garinii isolated from EM skin lesions (odds ratio for pleocytosis was 31 times higher in patients with established B. garinii skin infection compared to patients with other Borrelia species isolated from their EM skin lesion) and from CSF; and (4) more frequently fulfilled microbiologic criteria for established borrelial infection of the central nervous system. The positive predictive value of pleocytosis for microbiologically proven borrelial infection of the central nervous system (defined by isolation of Borrelia from CSF and/or demonstration of intrathecal synthesis of borrelial antibodies) was 67.9%, whereas normal CSF white cell counts ruled out Lyme neuroborreliosis with a predictive value of 91.9%. CONCLUSIONS: Comparison of European patients with EM who had symptoms/signs suggesting early Lyme neuroborreliosis revealed several differences in the clinical presentation and in microbiologic test results according to CSF findings.

Whole genome sequencing of Borrelia burgdorferi isolates reveals linked clusters of plasmid-borne accessory genome elements associated with virulence.

Lyme disease is the most common vector-borne disease in North America and Europe. The clinical manifestations of Lyme disease vary based on the genospecies of the infecting Borrelia burgdorferi spirochete, but the microbial genetic elements underlying these associations are not known. Here, we report the whole genome sequence (WGS) and analysis of 299 patient-derived B. burgdorferi sensu stricto ( Bbss ) isolates from patients in the Eastern and Midwestern US and Central Europe. We develop a WGS-based classification of Bbss isolates, confirm and extend the findings of previous single- and multi-locus typing systems, define the plasmid profiles of human-infectious Bbss isolates, annotate the core and strain-variable surface lipoproteome, and identify loci associated with disseminated infection. A core genome consisting of ∻800 open reading frames and a core set of plasmids consisting of lp17, lp25, lp36, lp28-3, lp28-4, lp54, and cp26 are found in nearly all isolates. Strain-variable (accessory) plasmids and genes correlate strongly with phylogeny. Using genetic association study methods, we identify an accessory genome signature associated with dissemination and define the individual plasmids and genes that make up this signature. Strains within the RST1/WGS A subgroup, particularly a subset marked by the OspC type A genotype, are associated with increased rates of dissemination. OspC type A strains possess a unique constellation of strongly linked genetic changes including the presence of lp56 and lp28-1 plasmids and a cluster of genes that may contribute to their enhanced virulence compared to other genotypes. The patterns of OspC type A strains typify a broader paradigm across Bbss isolates, in which genetic structure is defined by correlated groups of strain-variable genes located predominantly on plasmids, particularly for expression of surface-exposed lipoproteins. These clusters of genes are inherited in blocks through strain-specific patterns of plasmid occupancy and are associated with the probability of invasive infection.

Are Differences in Presentation of Early Lyme Borreliosis in Europe and North America a Consequence of a More Frequent Spirochetemia in American Patients?

To assess whether differences in presentation between US and European patients with early Lyme borreliosis are due to the lower rate of spirochetemia in Europe, we compared multiple variables for patients with erythema migrans (EM), restricting the analysis to subjects with a positive blood culture at the time of presentation: 93 US patients infected with Borrelia burgdorferi versus 183 European patients infected with Borrelia afzelii (No = 144) or Borrelia garinii (No = 39). Compared to spirochetemic Slovenian EM patients infected with B. afzelii, US patients with a positive blood culture significantly less often recalled a preceding tick bite at the site of the EM skin lesion, had a shorter duration of EM prior to diagnosis and more often had multiple EM lesions, regional lymphadenopathy, constitutional symptoms, an increased ESR value, a low blood lymphocyte count and detectable borrelia antibodies in acute and convalescent phase blood samples. Similar differences were observed when US patients were compared to Slovenian patients with B. garinii infection, but not all reached statistical significance. The findings are comparable to those previously reported for the corresponding skin culture positive patients and do not support the hypothesis that a higher frequency of spirochetemia at the time of presentation in US patients with EM, compared with European EM patients, is the reason for the observed differences.

Characteristics of spirochetemic patients with a solitary erythema migrans skin lesion in Europe.

Neither pre-treatment characteristics, nor the outcome after antibiotic therapy, have been reported for spirochetemic European patients with Lyme borreliosis. In the present study, patients with a solitary erythema migrans (EM) who had a positive blood culture for either Borrelia afzelii (n = 116) or Borrelia garinii (n = 37) were compared with age- and sex-matched patients who had a negative blood culture, but were culture positive for the corresponding Borrelia species from skin. Collectively, spirochetemic patients significantly more often recalled a tick bite at the site of the EM skin lesion, had a shorter time interval from the bite to the onset of EM, had a shorter duration of the skin lesion prior to diagnosis, and had a smaller EM skin lesion that was more often homogeneous in appearance. Similar results were found for the subset of spirochetemic patients infected with B. afzelii but not for those infected with B. garinii. However, patients with B. garinii bacteremia had faster-spreading and larger EM skin lesions, and more often reported itching at the site of the lesion than patients with B. afzelii bacteremia. Treatment failures were rare (7/306 patients, 2.3%) and were not associated with having spirochetemia or with which Borrelia species was causing the infection.

Course and Outcome of Erythema Migrans in Pregnant Women.

Information on Lyme borreliosis (LB) during pregnancy is limited. In the present study, the course and outcome of erythema migrans (EM) in 304 pregnant women, diagnosed in the period 1990-2015, was assessed and compared with that in age-matched non-pregnant women. The frequency of unfavorable outcome of pregnancies was also evaluated. The pregnant women reported constitutional symptoms less frequently than the non-pregnant women (22.4% vs. 37.2%, p < 0.001). Pregnant women diagnosed with EM later during pregnancy had a lower probability of reporting constitutional symptoms (odds ratio = 0.97 for 1-week difference in gestation week at diagnosis of EM, 95% CI: 0.94-0.99, p = 0.02). The outcome of pregnancy was unfavorable in 42/304 (13.8%) patients: preterm birth in 22/42 (52.4%), fetal/perinatal death in 10/42 (23.8%), and/or anomalies in 15/42 (35.7%). Several patients had potential explanation(s) for the unfavorable outcome. In conclusion, the course of early LB during pregnancy is milder than in age-matched non-pregnant women. The outcome of pregnancy with the treatment approach used in the present study (i.v. ceftriaxone 2 g once daily for 14 days) is favorable.

Early Lyme Borreliosis in Patients Treated with Tumour Necrosis Factor-Alfa Inhibitors.

The study evaluated the course and outcome of erythema migrans in patients receiving tumour necrosis factor-alpha (TNF-α) inhibitors. Among 4157 adults diagnosed with erythema migrans in the period 2009-2018, 16 (2.6%) patients were receiving TNF-α inhibitors (adalimumab, infliximab, etarnecept, golimumab), often in combination with other immunosuppressants, for rheumatic (13 patients) or inflammatory bowel (three patients) disease. Findings in this group were compared with those in 32 sex- and age-matched immunocompetent patients diagnosed with erythema migrans in the same years. In comparison with the control group, the immunocompromised patients had a shorter incubation period (7 vs. 14 days; p = 0.0153), smaller diameter of erythema migrans (10.5 vs. 15.5 cm; p = 0.0014), and more frequent comorbidities other than immune-mediated diseases (62.5% vs. 25%, p = 0.0269), symptoms/signs of disseminated Lyme borreliosis (18.8% vs. 0%, p = 0.0324), and treatment failure (25% vs. 0%, p = 0.0094). After retreatment with an antibiotic, the clinical course of Lyme borreliosis resolved. Continuing TNF inhibitor treatment during concomitant borrelial infection while using identical approaches for antibiotic treatment as in immunocompetent patients resulted in more frequent failure of erythema migrans treatment in patients receiving TNF inhibitors. However, the majority of treatment failures were mild, and the course and outcome of Lyme borreliosis after retreatment with antibiotics was favourable.

Acrodermatitis chronica atrophicans in children: Report on two cases and review of the literature.

Acrodermatitis chronica atrophicans is a late manifestation of European Lyme borreliosis and is characterized by high levels of borrelial IgG antibodies, slowly expanding skin redness usually beginning on distal parts of extremities, and corresponding histologic findings. It very rarely develops in children. The main prerequisite for the diagnosis is clinical suspicion. In the present article we report on two children with acrodermatitis chronica atrophicans and on the findings of a PubMed literature search on acrodermatitis chronica atrophicans in childhood, published in the past three decades.

Erythema Migrans: Course and Outcome in Patients Treated With Rituximab.

BACKGROUND: Information on Lyme borreliosis (LB) in patients treated with rituximab is limited to individual case reports. METHODS: We reviewed data on adult patients diagnosed with typical erythema migrans (EM) at the LB outpatient clinic of the University Medical Center Ljubljana, Slovenia, in the 10-year period 2008-2017. For all patients, clinical and laboratory information was acquired prospectively using a standardized questionnaire. RESULTS: Among 4230 adult patients with a diagnosis of EM, 7 patients (0.17%), 5 women and 2 men with a median age of 65 years (range, 55-66 years), were receiving rituximab for an underlying medical condition. In these 7 patients, signs of disseminated LB (43%) and the isolation rates of borreliae from blood before antibiotic treatment (40%) were unusually high compared with corresponding findings in immunocompetent patients who had EM diagnosed at the same institution (8% vs <2%, respectively). The rates of LB-associated constitutional symptoms and borrelial antibodies in serum were lower than expected (14% and 29%, respectively, in patients receiving rituximab vs 25% and 65% in immunocompetent patients). One of the 7 patients (14%) experienced treatment failure; nevertheless, the outcome of early LB 1 year after antibiotic treatment, as used for immunocompetent patients with EM, was excellent in all 7 patients. CONCLUSIONS: Findings in 7 patients with EM who were receiving rituximab for underlying disease suggest that although early LB in these patients is more often disseminated than in immunocompetent patients, the outcome 1 year after antibiotic treatment, as used for immunocompetent patients, is excellent.

Comparison of Clinical Course and Treatment Outcome for Patients With Early Disseminated or Early Localized Lyme Borreliosis.

Importance: Multiple erythema migrans (MEM) has been suggested as a risk factor for unfavorable antibiotic treatment outcome compared with solitary erythema migrans (EM). However, no direct comparison of early Lyme borreliosis manifested as MEM with solitary EM has been undertaken. Objective: To investigate the potential differences in clinical course and treatment outcome between MEM and solitary EM. Design, Setting, and Participants: This prospective cohort study was conducted from June 1, 2010, to October 31, 2015, at the University Medical Center Ljubljana, Slovenia. Data were analyzed from June 1, 2017, to January 3, 2018. Of the 778 consecutive adult patients with early Lyme borreliosis evaluated, 200 patients with MEM and 403 patients with solitary EM were enrolled. Patients were asked to refer a family member or a friend of similar age (±5 years) without a history of Lyme borreliosis to serve as a control participant. Clinical course and posttreatment outcome of MEM were compared with those of solitary EM. Outcome was assessed at 14 days and at 2, 6, and 12 months after enrollment. At each visit, patients completed a written questionnaire about their symptoms; controls completed the same questionnaire. Nonspecific symptoms reported by patients and controls without a history of Lyme borreliosis were compared. Main Outcomes and Measures: The proportion of patients with incomplete response at 12 months after enrollment and the associated 2-sided 95% CI for the difference between MEM and solitary EM were estimated using the normal approximation with continuity correction. Results: A total of 200 patients with MEM and 403 patients with solitary EM were included. Among the 200 patients with MEM, 94 (47.0%) were males and 106 (53.0%) were females, with a median (interquartile range [IQR]) age of 47 (35-58) years. Among the 403 patients with solitary EM, 182 (45.2%) were males and 221 (54.8%) were females, with a median (IQR) age of 55 (42-62) years. Patients with MEM reported Lyme borreliosis-associated constitutional symptoms at enrollment more often than those with solitary EM (93 [46.5%]; 95% CI, 39.4-53.7 vs 96 [23.8%]; 95% CI, 19.7-28.3; P < .001). During the initial 6 months after treatment, the proportion of patients with incomplete response was higher in the MEM group than in the solitary EM group (14 days: 62 of 193 [32.1%] vs 72 of 391 [18.4%]; P < .001; 2 months: 38 of 193 [19.7%] vs 55 of 394 [14.0%]; P = .28; 6 months: 29 of 182 [15.9%] vs 31 of 359 [8.6%]; P = .02). However, at the 12-month visit, the outcome was comparable: 10 of 170 (5.9%) patients with MEM vs 20 of 308 (6.5%) patients with solitary EM showed incomplete response (-0.6; 95% CI, -5.5 to 4.3; P = .95). The frequency of nonspecific symptoms in patients was similar to that in controls. Conclusions and Relevance: The long-term outcome at 12 months after treatment was comparable, regardless of dissemination. Follow-up of at least 12 months after treatment is thus recommended for future studies that investigate post-Lyme borreliosis symptoms.

Comparison of MKP and BSK-H media for the cultivation and isolation of Borrelia burgdorferi sensu lato.

The isolation of B. burgdorferi sensu lato requires the use of complex cultivation media. The aim of the study was to compare the usefulness of BSK-H (a commercial medium produced by HiMedia, India) and MKP medium. MKP and BSK-H media were prepared in accordance with the relevant protocols. Borrelia strains and skin culture biopsies were simultaneously inoculated into both media, incubated and checked for growth. Borrelial growth characteristics, isolation rates and characteristics of the isolated borreliae were analysed and compared. Initially, numbers of spirochaetes were higher in BSK-H than in MKP; however, in comparison with MKP, the strains subcultured in BSK-H medium were more frequently irregular, thin and non-motile, and rapidly died. In addition, the borrelial isolation rate from erythema migrans skin samples was higher in MKP than in BSK-H medium (108/171, 63.2% versus 70/171, 40.9%; p<0.0001). The far most frequently isolated species was Borrelia afzelii (92.9% and 97.2% strains isolated from BSK-H and MKP, respectively). Comparison of strains cultured from individual patients in both media showed differences in plasmid contents in 9/46 (19.6%) strain pairs, and protein profiles differed in 30/43 (69.8%) strain pairs, most often in the expression of OspC (in 27/28 patients OspC was expressed only in strains growing in MKP). BSK-H medium supports the growth of borrelial strains but MKP is superior with regard to the isolation rate, morphology and motility of strains. BSK-H medium supports fast initial growth of borreliae but this is followed by rapid deformation and death of the spirochaetes.

Pathogenetic implications of the age at time of diagnosis and skin location for acrodermatitis chronica atrophicans.

BACKGROUND: The pathogenesis of acrodermatitis chronica atrophicans (ACA) is not well understood. OBJECTIVE: The purpose of this study was to gain a better understanding of ACA by utilizing a large data set of adult Slovenian patients with Lyme borreliosis. METHODS: The age of 590 ACA patients was compared with that of patients with other manifestations of Lyme borreliosis. The location of the ACA lesion on the body was compared with that of erythema migrans (EM). RESULTS: Patients diagnosed with ACA were on average 14.3 years older than patients with EM (p<0.001). ACA patients were also significantly older than patients with Lyme neuroborreliosis or Lyme arthritis (p<0.001). The average delay in diagnosis of ACA was 1.6 years (range 0.1-20 years). For 572 (96.9%) of the ACA patients, the site of the skin lesion(s) was confined to an extremity vs. 79.6% for patients with EM, p<0.001. For the 20 ACA patients who reported a preceding untreated EM lesion at the same body site, the mean time between the development of the EM and the onset of ACA was 3.0±4.4 (median 1.3, range 0.1-15.0) years. CONCLUSIONS: ACA is more likely to be diagnosed in older individuals than any other manifestation of Lyme borreliosis. ACA is more likely than EM to be localized anatomically to the extremities. Available data favor the hypothesis that ACA occurs most often on the extremities of older individuals because of predisposing age-related anatomic or physiologic changes, but more data are needed to define the latency period and other aspects of the pathogenesis of this skin condition.

Erythema migrans in solid-organ transplant recipients.

Six adult solid-organ transplant recipients who had chronic drug-induced immunosuppression and who presented with solitary erythema migrans were treated with antibiotics administered at the same dosage and for the same duration used for the treatment of early, localized Lyme borreliosis in immunocompetent patients. The patients had a smooth course of illness and a favorable outcome but did not develop a measurable borrelial serum antibody response.

Susceptibility of Borrelia afzelii strains to antimicrobial agents.

The aim of the present study was to determine the susceptibility of Borrelia afzelii strains to antibiotics, and to test the hypothesis that persistence of borrelia in skin, after therapy, is a consequence of resistance to the antibiotic used for treatment. Ten B. afzelii strains isolated from skin of seven adult patients (two with acrodermatitis chronica atrophicans, five with erythema migrans) were studied. In three patients B. afzelii was isolated from erythema migrans lesion before antibiotic therapy and 2-3 months after treatment with cefuroxime axetil (two patients) or with ceftriaxone (one patient). MICs and MBCs for amoxicillin, azithromycin, ceftriaxone, cefuroxime, doxycycline and amikacin were measured. There was total resistance to amikacin but isolates were susceptible to all other antibiotics except one isolate that was resistant to cefuroxime, MIC > 4 mg/L. Comparison of MBC values after 3 and 6 weeks' incubation revealed comparable results for azithromycin and ceftriaxone while for amoxicillin, cefuroxime and doxycycline, some differences were found. In one of the patients from whom there were borrelia isolated before and after treatment with cefuroxime axetil, both isolates were resistant to cefuroxime. In the other two patients, the paired isolates were susceptible to the antibiotic used for therapy.