Validation of cultivation and PCR methods for diagnosis of Lyme neuroborreliosis.

Borrelial infection may manifest with a wide range of clinical signs, and in many cases, microbiological findings are essential for a proper diagnosis. This study included 48 patients with a working clinical diagnosis of Lyme neuroborreliosis, 45 patients with a working clinical diagnosis of suspected Lyme neuroborreliosis, and a control group comprising 42 patients with tick-borne encephalitis and 21 neurosurgical patients. The aim of the study was to analyze and compare findings of two PCR methods and Borrelia burgdorferi sensu lato culture results by examination of prospectively collected cerebrospinal fluid (CSF) and blood specimens from patients with clinical features of Lyme neuroborreliosis. Borrelial DNA was detected with at least one of the PCR approaches in 16/135 (11.9%) blood samples and 24/156 (15.4%) CSF samples. Using MseI restriction of PCR products of the amplified rrf-rrl region, we identified the majority of strains as Borrelia afzelii. Borreliae were isolated from 1/135 (0.7%) blood samples and from 5/156 (3.2%) CSF specimens. Using MluI restriction for characterization of isolated strains, Borrelia garinii was identified in all CSF isolates. Our study revealed that different approaches for direct demonstration of borrelial infection give distinct results, that there is an urgent need for standardization of the methods for direct detection of borrelial infection, and that the design of studies evaluating the validation of such methods should include appropriate control group(s) to enable assessment of both sensitivity and specificity.

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.

Comparison of immunofluorescence assay (IFA) and LIAISON in patients with different clinical manifestations of Lyme borreliosis.

Serological tests for detection of borrelial antibodies are frequently used in laboratory diagnostics of Lyme borreliosis. Unfortunately these tests are not standardized and the results obtained with different assays may not be concordant. The aim of the present study was to compare two different serological tests, IFA and LIAISON, for detection of Borrelia burgdorferi sensu lato IgM and IgG antibody. We analyzed the serological immune response in 383 patients with different clinical manifestations of Lyme borreliosis and in 49 healthy blood donors. LIAISON detected IgM and IgG antibodies more often than IFA in all groups of patients except those with chronic Lyme borreliosis. The differences were significant for IgM and IgG antibodies in patients with solitary erythema migrans and in those with early disseminated Lyme borreliosis. There was no significant difference in the specificity of the two tests.

Clinical and epidemiological findings for patients with erythema migrans. Comparison of cohorts from the years 1993 and 2000.

OBJECTIVES: i) To present clinical and epidemiological findings on adult patients diagnosed with typical erythema migrans (EM) at our institution during the year 2000 and ii) To compare the findings with data obtained by the same approach for an identical group of patients from 1993, with the aim of testing the hypothesis that, because of increased knowledge of Lyme borreliosis in 2000 compared with 1993, patients will visit us earlier and consequently present with smaller skin lesions. PATIENTS AND METHODS: Patients who qualified for inclusion in the study were adults diagnosed with typical EM in 2000 and 1993 at the Lyme borreliosis Outpatients' Clinic, Ljubljana, Slovenia. Epidemiological and clinical data were obtained from a questionnaire. RESULTS: In 2000, 535 patients had typical EM: 309 (57.8%) females and 226 (42.2%) males, aged 15-80 (median 47) years. Tick bite at the site of later EM was recalled by 311 (58.1%) patients a median of 14 days before the onset of the skin lesion, which was localised on the legs in 55.7% of patients. The median duration of EM prior to examination was 8.5 days, the largest diameter of EM was 12 (4-87) cm, and the average area of involved skin was 75 cm2. Thirty-six of 535 (6.7%) patients had more than one skin lesion (maximum number 21), 281/535 (52.5%) patients reported local symptoms (itching 45.2%, burning 12.9%, and pain 7.1%), and 191/535 (35.7%) reported systemic symptoms (fatigue 17.4%, headache 17.2%, myalgia 12.6%, arthralgia 11.2%, vertigo 3.6%, fever 2.5%, and chills 1.5%). In 1993, 892 patients with typical EM were diagnosed. They had similar characteristics to those in 2000 but were younger (44 versus 47 years; p = 0.025), more often remembered a tick bite (654/892 versus 311/535; p = 0.009), had a smaller diameter (10 versus 12 cm; p < 0.001) and surface area of EM (50.2 versus 75.0 cm2), presented more often with homogeneous skin lesions (378/892 versus 191/535; p = 0.010), and more frequently had some of the associated symptoms including nausea (43/892 versus 12/535; p = 0.021) and malaise (124/892 versus 51/535; p = 0.019). CONCLUSIONS: The majority of findings for the cohorts of EM patients seen in 1993 and 2000 were comparable but--contrary to expectations--in 2000 the duration of EM prior to attending our clinic was not shorter, and the skin lesions were larger and less often homogeneous. These findings indicate that knowledge of Lyme borreliosis did not increase during the period from 1993 to 2000.

Cerebrospinal fluid findings in adult patients with multiple erythema migrans.

This prospective study was performed at the Department of Infectious Diseases, University Medical Centre Ljubljana, Slovenia, in the period from 1991 to 2000. We included all adult patients with multiple erythema migrans who gave consent to lumbar puncture, had routine blood and CSF tests performed, and borrelial antibody titres in CSF and blood determined. In the majority of these patients skin, blood, and CSF specimens were cultured in MKP medium for the presence of Borrelia. Of 332 patients with multiple erythema migrans, 200 (115 females, 85 males, aged 15-80 years) fulfilled inclusion criteria. The median number of skin lesion was three (2-60). Sixty-three (31.5%) patients had no associated symptoms, whereas 137 (68.5%) patients (including two with arthritis, six with radicular pain, a patient with facial palsy, another patient with foot palsy and a patient with transitory diplopia) reported local and/or constitutional symptoms. Routine CSF examination revealed abnormal results in 62/200 (31%) patients: lymphocytic pleocytosis (6-1119 x 10(6)/L leukocytes) was found in 15 (7.5%) patients (six were clinically without systemic symptoms, six had mild systemic symptoms, three reported radicular pains) and elevated CSF protein concentration was present in 52 (26%) patients (nine also had elevated CSF cell counts). Intrathecal borrelial antibody production was demonstrated in eight (4%) patients (only three of them had elevated CSF cell counts) and B. burgdorferi sensu lato was isolated from skin lesions, blood, and CSF in 77/191 (40.3%), 3/154' (1.95%), and 2/200 (1%) patients, respectively. B. afzelii predominated among the isolates. In patients with multiple erythema migrans abnormal CSF findings are not rare and may be present without any clinical sign of central nervous system involvement.

Electrocardiographic findings in patients with erythema migrans.

OBJECTIVE: To identify asymptomatic heart involvement early in the course of Lyme borreliosis by analysis of electrocardiograms (ECGs) of patients with solitary erythema migrans (EM). METHODS: We enrolled in this prospective study 220 consecutive previously healthy patients, receiving no medication, who were diagnosed with typical solitary EM at our Lyme borreliosis Outpatient Clinic in 1998. Their ECG findings were compared with the ECG results in 165 age and sex-matched healthy persons. RESULTS: Three patients with EM but none in the control group had first degree AV block (p = 0.319). Second and third degree AV blocks, electrocardiographic signs of pericarditis, myocarditis or rhythm disturbances were not found in any of the ECGs. Comparison of findings in patients with EM and the control group revealed statistically significant differences in frequency (66.2 +/- 1.47 beats/min in patients with EM versus 68.4 +/- 1.52 beats/min in controls; p = 0.043), duration of Q (0.013 +/- 0.001 s in patients with EM versus 0.015 +/- 0.001 s in controls; p < 0.001), and in depth of S (0.135 +/- 0.016 mV in patients with EM versus 0.104 +/- 0.012 mV in controls; p < 0.05), but not for several other ECG parameters. These differences were found in females as well as in males. Comparison of ECG findings in subgroups of patients with EM (grouped according to EM characteristics such as length of incubation, duration of skin lesion and the presence of associated systemic symptoms) did not reveal any significant difference. CONCLUSIONS: AV blocks, the most typical heart manifestation of early disseminated Lyme borreliosis, are a rare finding in patients with solitary EM, and in our study their frequency did not differ from that in the control group. We do not have a reliable explanation for the shorter duration of Q and deeper S wave in patients with EM compared with age and sex-matched controls.

Procalcitonin levels in patients with Lyme borreliosis.

BACKGROUND: Serum and cerebrospinal fluid (CSF) procalcitonin levels were assessed and compared for different groups of patients with Lyme borreliosis. PATIENTS AND METHODS: 50 adult patients with Lyme borreliosis, referred to our department from March to June 2001, were included in this prospective study. Patients were divided into three groups. The first group consisted of 20 consecutive patients with typical solitary erythema migrans, representing early localised Lyme borreliosis, the second group comprised 20 patients with early disseminated Lyme borreliosis (10 with multiple erythema migrans and 10 with neuroborreliosis), and 10 patients with acrodermatitis chronica athrophicans represented the group with chronic Lyme borreliosis. Blood specimens were taken from all patients included in the study, but CSF samples were restricted to those with disseminated and chronic Lyme borreliosis. The serum and CSF procalcitonin levels were determined utilizing the LUMI PCT (an immunoluminometric assay using two antigen-specific monoclonal antibodies). RESULTS: Serum and CSF procalcitonin levels were in normal range in the large majority of patients. The levels of serum procalcitonin did not differ in the three groups of patients with Lyme borreliosis (p = 0.5006). The corresponding values for patients with solitary erythema migrans (early localised Lyme borreliosis), early disseminated Lyme borreliosis, and chronic Lyme borreliosis were 0.26 (0.11-0.43), 0.22 (0.10-0.67), and 0.28 (0.13-0.66) microgram/ml, respectively. Moreover, procalcitonin levels in CSF were also low and comparable for patients with multiple erythema migrans (median 0.38, range 0.24-0.54 microgram/ml), neuroborreliosis (median 0.16, range 0.10-0.47 microgram/ml), and acrodermatitis chronica athrophicans (median 0.30, range 0.15-0.45 microgram/ml). The differences were not statistically significant (p = 0.7579). CONCLUSIONS: In the large majority of patients with Lyme borreliosis procalcitonin values are within normal range. Serum and CSF procalcitonin levels are of no value for differentiation between early localised, early disseminated and chronic Lyme borreliosis.

Cerebrospinal fluid findings in patients with symptoms suggesting chronic Lyme borreliosis.

BACKGROUND: Patients with non-specific long-lasting symptoms such as headache, concentration disturbances, and vertigo and who have positive serum borrelial antibody titres are often assumed to have chronic Lyme borreliosis. Because of the possibility that they may have central nervous system involvement they are frequently treated with courses of i.v. ceftriaxone. We assessed central nervous system involvement by examining cerebrospinal fluid samples in a group of such patients. PATIENTS AND METHODS: Adult patients who qualified for the study had non-specific symptoms suggesting central nervous system involvement for longer than six months (but without overt clinical signs of such involvement) and positive serum borrelial antibody titres and/or erythema migrans prior to the onset of symptoms. Cerebrospinal fluid was examined in all patients. RESULTS: None of the 77 patients included in the study (median duration of symptoms 18 months) had pleocytosis and there was no isolation of Borrelia burgdorferi sensu lato from cerebrospinal fluid. Mildly elevated protein concentration and intrathecal borrelial IgG antibody synthesis were demonstrated in 16 (21%) and 7 (9.1%) patients, respectively. CONCLUSIONS: In patients with non-specific long-lasting symptoms attributed to Lyme borreliosis but with the absence of overt clinical signs suggesting central nervous system involvement, the findings of cerebrospinal fluid examination are usually in the normal range. Routine treatment of such patients with i.v. ceftriaxone is not to be encouraged.

Evaluation of immunofluorescence test (IFT) and immuno (western) blot (WB) test in patients with erythema migrans.

The diagnosis of Lyme borreliosis is based on the recognition of typical clinical signs and is assisted by laboratory confirmation of borrelial infection. The aim of the present study was to assess the value of an immunofluorescence test (IFT) and an immuno (western) blot (WB) test for the detection of Borrelia burgdorferi sensu lato antibodies in patients with erythema migrans residing in Slovenia. We determined specific IgM and IgG antibodies in 117 patients with erythema migrans and 96 healthy persons using an IFT (in-house test) and a commercial WB test. Skin biopsies of erythema migrans lesions were cultured, and isolated strains were identified with PFGE. There were 66/117 (56.4%) culture-positive and 51/117 (43.6%) culture-negative patients. B. afzelii was found in 52/62 (84%) and B. garinii in 10/62 (16%) biopsies. IFT-IgM antibodies were established in 2/117 (1.7%) erythema migrans patients and in none of the control group, while WB-IgM antibodies were present in 56/117 (48%) patients with erythema migrans and 21/96 (22%) members of the control group (p = 0.002). IFT-IgG antibodies were demonstrated in 3/117 (2.2%) erythema migrans patients and 2/96 (4%) persons of the control group, while corresponding values for WB-IgG were 36/117 (31%) and 26/96 (27%), respectively (non-significant differences). IgM antibodies directed against p41 and OspC, and IgG antibodies directed against p41, p18 and OspC were frequently found in both erythema migrans patients and the control group. The only significant difference between erythema migrans patients and the control group in the WB test was in the reaction of IgM antibodies with OspC antigen, which was found in 54/117 (46%) erythema migrans patients and 18/96 (18.8%) healthy persons (p < 0.0001). The immune response in patients with erythema migrans was very similar to that of the control group determined with either the IFT or WB test.

Double infection with tick borne encephalitis virus and Borrelia burgdorferi sensu lato.

The limited information on co-infection with Borrelia burgdorferi sensu lato and tick-borne encephalitis (TBE) virus was a stimulus for presentation of two patients with well-defined double infection of the central nervous system. TBE virus and B. burgdorferi sensu lato infections are searched for in all patients with lymphocytic meningitis and/or meningoencephalitis admitted to our department. During the last ten years we identified two patients who had ELISA IgM and IgG antibodies to TBE virus in serum and a positive PCR result for TBE virus in cerebrospinal fluid as well as B. burgdorferi sensu lato isolated from cerebrospinal fluid. Intrathecal production of borrelial antibodies was not proven in either of the two patients. These findings show that in patients with acute lymphocytic meningitis originating in regions endemic for Lyme borreliosis and TBE, the possibility of concomitant infection should be considered.

Solitary borrelial lymphocytoma in adult patients.

During the period from 1986 to 2000, 85 adult patients with solitary borrelial lymphocytoma were diagnosed at the Department of Infectious Diseases, University Medical Centre Ljubljana, Slovenia. There were 36 (42.4%) females and 49 (57.6%) males with a median age of 49 (15-74) years. Borrelial lymphocytoma was located on the breast (nipple--areola mammae region) in 68 (80%) patients, on the ear lobe in eight (9.4%), and in other locations in nine (10.6%). A concomitant erythema migrans enabling clinical diagnosis of Lyme borreliosis was registered or reported in 67 (78.8%) patients. Fifteen (17.6%) patients had no accompanying symptoms, 34 (40%) reported local and constitutional symptoms, 23 (27.1%) recounted only local symptoms, and 13 (15.3%) patients had solely constitutional symptoms. Clinical findings indicating early disseminated borrelial infection were observed at the first visit in 12 (14.1%) patients: six (7.1%) had multiple erythema migrans, one had meningitis, one meningoradiculitis and arthritis, one radiculoneuritis and arthritis, one peripheral facial palsy and concomitant meningitis, and two arthritis. In addition, one of the patients with borrelial lymphocytoma on the breast had acrodermatitis chronica atrophicans. A seropositive response to borrelial antigens was found in 30 (35.3%) patients at the initial examination. In 11/46 (23.9%) patients, infection with Borrelia burgdorferi sensu lato was confirmed by isolation of the agent from lymphocytoma tissue. Eight out of nine (88.9%) typed borrelial strains were found to be B. afzelii, and one (11.1%) B. bissettii. Patients were treated with doxycycline, azithromycin, amoxycillin, cefuroxime-axetil, phenoxymethylpenicillin, or ceftriaxone. Median time to complete disappearance of lymphocytoma was 28 days (range 7-270 days) after the institution of antibiotic treatment; disappearance took longer in patients with prolonged duration of the skin lesion prior to treatment. Treatment failure was registered in 11 (12.9%) patients who were later re-treated. The outcome of borrelial infection assessed at the end of a follow-up period of one year was favourable.

Sensitivity of culture and polymerase chain reaction for the etiologic diagnosis of erythema migrans.

Skin biopsy samples from 150 patients with typical cutaneous manifestation of Lyme borreliosis, erythema migrans, were cultivated for the presence of Borrelia burgdorferi sensu lato in modified Kelly Pettenkofer (MKP) medium and analysed with two different polymerase chain reactions using either flagellin or nested OspA primers. Cultivation was successful in 75 of 150 (50%) skin samples. Out of 70 strains that were typed using PFGE, B. afzelii was identified in 60 (86%), B. garinii in 10 (14%) specimens, while no B. burgdorferi sensu stricto strains were found. B. burgdorferi sensu lato DNA was detected with polymerase chain reaction in 28% and 61% of skin samples, using flagellin and nested OspA primers, respectively. Concordant results in all three procedures employed in the present study were found in 62 (41%) specimens: 25/150 (17%) were positive with all three methods and 37/150 (25%) samples were completely negative.

Are patients with erythema migrans who have leukopenia and/or thrombocytopenia coinfected with Anaplasma phagocytophilum or tick-borne encephalitis virus?

Lyme borreliosis (LB), tick-borne encephalitis (TBE) and human granulocytic anaplasmosis (HGA) are endemic in central part of Slovenia. We tested the hypothesis that patients with erythema migrans (EM) from this region, who have leukopenia and/or thrombocytopenia (typical findings in HGA and in the initial phase of TBE but not in patients with LB) are coinfected with Anaplasma phagocytophilum and/or with TBE virus, i.e. that cytopenia is a result of concomitant HGA or the initial phase of TBE. Comparison of clinical and laboratory findings for 67 patients with EM who disclosed leukopenia/thrombocytopenia with the corresponding results in sex- and age-matched patients with EM and normal blood cell counts revealed no differences. In addition, patients with typical EM and leukopenia and/or thrombocytopenia tested negative for the presence of IgM and IgG antibodies to TBE virus by ELISA as well as for the presence of specific IgG antibodies to A. phagocytophilum antigens by IFA in acute and convalescent serum samples. Thus, none of 67 patients (95% CI: 0 to 5.3%) with typical EM (the presence of this skin lesion attests for early Lyme borreliosis and is the evidence for a recent tick bite) was found to be coinfected with A. phagocytophilum or had a recent primary infection with TBE virus. The findings in the present study indicate that in Slovenia, and probably in other European countries endemic for LB, TBE and HGA, patients with early LB are rarely coinfected with the other tick-transmitted agents.

Clinical characteristics associated with Borrelia burgdorferi sensu lato skin culture results in patients with erythema migrans.

Clinical characteristics associated with isolation of Borrelia burgdorferi sensu lato from skin have not been fully evaluated. To gain insight into predictors for a positive EM skin culture, we compared basic demographic, epidemiologic, and clinical data in 608 culture-proven and 501 culture-negative adult patients with solitary EM. A positive Borrelia spp. skin culture was associated with older age, a time interval of >2 days between tick bite and onset of the skin lesion, EM ≥ 5 cm in diameter, and location of the lesion on the extremities, whereas several other characteristics used as clinical case definition criteria for the diagnosis of EM (such as tick bite at the site of later EM, information on expansion of the skin lesion, central clearing) were not. A patient with a 15-cm EM lesion had almost 3-fold greater odds for a positive skin culture than patients with a 5-cm lesion. Patients with a free time interval between the tick bite and onset of EM had the same probability of a positive skin culture as those who did not recall a tick bite (OR=1.02); however, the two groups had >3-fold greater odds for EM positivity than patients who reported a tick bite with no interval between the bite and onset of the lesion. In conclusion, several yet not all clinical characteristics used in EM case definitions were associated with positive Borrelia spp. skin culture. The findings are limited to European patients with solitary EM caused predominantly by B. afzelii but may not be valid for other situations.

In vitro susceptibility of European human Borrelia burgdorferi sensu stricto strains to antimicrobial agents.

Broth microdilution and macrodilution assays were used to determine minimum inhibitory concentrations (MICs) and minimum bactericidal concentrations (MBCs) of six antimicrobial agents (ceftriaxone, cefuroxime sodium, azithromycin, amoxicillin, doxycycline and amikacin) for nine European human isolates of Borrelia burgdorferi sensu stricto (s.s.). Strains were obtained from patients diagnosed with Lyme borreliosis in Slovenia. Modified Kelly-Pettenkofer medium with a final inoculum of 105 Borrelia cells/mL and incubation periods of 72 h and of 3 weeks and 6 weeks were used in the determination of MICs and MBCs, respectively. Observed MICs indicated that all isolates were susceptible to all the tested antimicrobial agents with the exception of amikacin. Cefuroxime sodium (MIC90=0.063 mg/L), azithromycin (MIC90=0.22 mg/L) and ceftriaxone (MIC90=0.25 mg/L) displayed the lowest MICs, followed by amoxicillin (MIC90=1 mg/L) and doxycycline (MIC90=2 mg/L); no strain was susceptible to amikacin (MIC90=256 mg/L). MBCs after incubation for 3 weeks and 6 weeks were determined for amoxicillin (MBC90=32 mg/L), doxycycline (MBC90)=32 mg/L) and amikacin (MBC90=1024 mg/L) and were found to be high (but not defined) for azithromycin (MBC90>0.88 mg/L), cefuroxime sodium (MBC90>4 mg/L) and ceftriaxone (MBC90>4 mg/L). In determination of borrelial susceptibility to antimicrobial agents, intrinsic low susceptibility or methodological factors could result in low in vitro susceptibility of individual strains. This study is the first report on the antibiotic susceptibility of a series of European human isolates of B. burgdorferi s.s.

Gender disparity between cutaneous and non-cutaneous manifestations of Lyme borreliosis.

Cutaneous manifestations of Lyme borreliosis in Europe include erythema migrans (EM) and acrodermatitis chronica atrophicans (ACA); the most common non-cutaneous manifestations are Lyme neuroborreliosis (LNB) and Lyme arthritis. The purpose of this study was to evaluate the gender distribution of patients with these clinical manifestations of Lyme borreliosis. Data on gender were obtained from the clinical records of patients with Lyme borreliosis aged ≥15 years who had been evaluated at the University Medical Center Ljubljana, Ljubljana, Slovenia. Among 10,539 patients diagnosed with EM, 6,245 (59.3%) were female and among 506 ACA patients 347 (68.6%) were female. In contrast, among the 60 patients with Lyme arthritis only 15 (25%) were female (p<0.0001 for the comparison of gender with EM or ACA) and among the 130 patients with LNB only 51 (39.2%) were females (p<0.0001for the comparison of gender with EM or ACA). Although the proportion that was female in the LNB group was greater than that of patients with Lyme arthritis, this difference did not reach statistical significance (p = 0.10). Although older individuals are more likely to be female in the general Slovenian population, the age of patients with cutaneous versus non-cutaneous manifestations was not the explanation for the observed differences in gender. In conclusion, patients with cutaneous manifestations of Lyme borreliosis were predominantly female, whereas those with non-cutaneous manifestations were predominantly male. This provocative finding is unexplained but may have direct relevance to the pathogenesis of Lyme borreliosis.

Comparison of erythema migrans caused by Borrelia burgdorferi and Borrelia garinii.

BACKGROUND: A comparison of patients with erythema migrans due to Borrelia garinii versus Borrelia burgdorferi has not been reported. PATIENTS AND METHODS: One hundred nineteen patients from New York State with erythema migrans caused by B. burgdorferi were compared with 116 patients from Slovenia with erythema migrans due to B. garinii infection. RESULTS: Patients with B. garinii infection were older, more often reported a tick bite, and developed larger lesions (median largest diameter: 18 and 14 cm, respectively; p = 0.01) that more often had central clearing (61.2% compared with 35.3%; p < 0.0001). Patients infected with B. burgdorferi, however, more often had systemic symptoms (68.9% vs. 37.1%; p < 0.0001), including fatigue (p < 0.0001), arthralgia (p = 0.0003), myalgia (p < 0.0001), headache (p = 0.0008), fever and/or chills (p < 0.0001), and stiff neck (p < 0.0001), and more often had abnormal physical findings (57.1% compared with 11.2%; p < 0.0001), such as fever (p = 0.0002) or regional lymphadenopathy (p < 0.0001). There was a trend for more patients with B. burgdorferi infection to have multiple erythema migrans skin lesions (13.4% compared with 5.2%; p = 0.051), and among patients with multiple erythema migrans the number of lesions was greater in B. burgdorferi-infected patients (median: 5.5 compared with 2.0; p = 0.006). CONCLUSIONS: The results of the present study indicate that in patients with erythema migrans the clinical features vary according to whether infection is caused by B. garinii or B. burgdorferi.

Comparison of isolation rate of Borrelia burgdorferi sensu lato in MKP and BSK-II medium.

Different media have been utilized for borrelial cultivation. The aim of the present study was to evaluate the isolation rate of Borrelia burgdorferi sensu lato from two commonly used media, i.e. modified Kelly-Pettenkofer (MKP) and Barbour-Stoenner-Kelly II (BSK-II) medium, and to compare the isolated strains with regard to their phenotypic and genotypic characteristics. Skin biopsy specimens of 2 x 2 x 4 mm were taken from the peripheral site of human solitary erythema lesions and were divided in two pieces, one of which was inoculated into MKP and the other one into BSK-II medium. Species analysis of the obtained strains was performed and their plasmid and protein profiles were determined. Borrelia strains were isolated from 48/96 patients (50%) with erythema migrans. We obtained in 26/48 patients (54%) from MKP as well as from BSK-II, in 11 patients (23%) only from MKP, and in another 11 (23%) only from BSK-II medium a positive result. B. afzelii was isolated from 43 patients (23 were positive in both media, nine in MKP only and 11 in BSK-II medium only), while B. garinii was isolated from five patients (in three from both media, in two from MKP only). All strains of the obtained strain pairs were identical according to species and the type within the species. Plasmid profiles were identical in 17/21 B. afzelii strain pairs (81%) and in 1/3 B. garinii strain pairs; in 6/24 strain pairs, distinctions in the number of plasmids or in their molecular mass were present. Differences in the protein profile were found in 7/24 strain pairs (29%). The distinctions were uniform and were limited to the expression of OspC. In conclusion, our study showed comparable Borrelia isolation rates from MKP and BSK-II medium. The results of the present study indicate that human patients with Lyme borreliosis may simultaneously harbor heterogeneous B. burgdorferi s.l. strains.

Intravenous ceftriaxone compared with oral doxycycline for the treatment of Lyme neuroborreliosis.

This prospective, open-label, non-randomized trial at the University Departments of Infectious Diseases in Ljubljana, Slovenia, and Göteborg, Sweden, was conducted to compare the kinetics of the cerebrospinal fluid (CSF) mononuclear cell count after 10-14 d of ceftriaxone or doxycycline for treatment of Lyme neuroborreliosis. 29 patients were treated with intravenous ceftriaxone 2 g daily in Ljubljana and 36 patients with oral doxycycline 400 mg daily in Göteborg. The study protocol included lumbar puncture before and 6-8 weeks after treatment initiation. There was a marked decrease (1.2 log10 x 10(6)/l) of the median CSF mononuclear cell count following treatment. With the assumption of a linear regression of the logarithmic mononuclear cell counts between the 2 lumbar punctures, no significant difference between the 2 antibiotic treatments could be found. All patients were clinically much improved after treatment. At 6 months follow-up 23 (79%) of the ceftriaxone- and 26 (72%) of the doxycycline-treated patients were completely recovered. Intravenous ceftriaxone or oral doxycycline was found to be effective, safe, and convenient for treatment of Lyme neuroborreliosis.