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1.  Diagnostic Value of Cytokines and Chemokines in Lyme Neuroborreliosis 
Clinical and Vaccine Immunology : CVI  2013;20(10):1578-1584.
The aims of the present study were to assess the concentrations of different cytokines and chemokines in blood serum and cerebrospinal fluid (CSF) samples of patients with Lyme neuroborreliosis and to identify the possible marker(s) that would enable a distinction between clinically evident and suspected Lyme neuroborreliosis, as well as between Lyme neuroborreliosis and tick-borne encephalitis (TBE). Our additional interest was to evaluate the relationship between cytokine and chemokine concentrations and Borrelia burgdorferi sensu lato isolation from CSF, as well as intrathecal synthesis of specific borrelial antibodies. We found that higher concentrations of CXCL13 and lower concentrations of interleukin 10 (IL-10) in serum were associated with higher odds for clinically evident Lyme neuroborreliosis compared to suspected Lyme neuroborreliosis, as well as to TBE. The concentrations of IL-2, IL-5, IL-6, IL-10, and CXCL13 in the CSF were higher in patients with evident Lyme neuroborreliosis than in those who were only suspected to have the disease. A comparison of CSF cytokine and chemokine levels in patients with and without intrathecal synthesis of specific borrelial antibodies revealed that CXCL13 CSF concentration is significantly associated with intrathecal synthesis of borrelial antibodies. A comparison of the cytokine and chemokine CSF concentrations in patients with clinically evident Lyme neuroborreliosis according to CSF culture results revealed that higher concentrations of gamma interferon (IFN-γ) were associated with lower odds of Borrelia isolation. Although several differences in the blood serum and CSF concentrations of various cytokines and chemokines between the groups were found, the distinctive power of the majority of these findings is low. Further research on well-defined groups of patients is needed to appraise the potential diagnostic usefulness of these concentrations.
doi:10.1128/CVI.00353-13
PMCID: PMC3807194  PMID: 23945160
2.  Clinical and Serological Follow-Up of Patients with Human Granulocytic Ehrlichiosis in Slovenia 
An evaluation of the clinical outcome and the duration of the antibody response of patients with human granulocytic ehrlichiosis (HGE) was undertaken in Slovenia. Adult patients with a febrile illness occurring within 6 weeks of a tick bite were classified as having probable or confirmed HGE based on the outcome of serological or PCR testing. Thirty patients (median age, 44 years) were enrolled, and clinical evaluations and serum collection were undertaken at initial presentation and at 14 days, 6 to 8 weeks, and 3 to 4, 6, 12, 18, and 24 months. An indirect immunofluorescence assay (IFA) was performed, and reciprocal titers of ≥128 were interpreted as positive. Patients presented a median of 4 days after the onset of fever and were febrile for a median of 7.5 days; four (13.3%) received doxycycline. Seroconversion was observed in 3 of 30 (10.0%) patients, and 25 (83.3%) showed >4-fold change in antibody titer. PCR results were positive in 2 of 3 (66.7%) seronegative patients but in none of 27 seropositive patients at the first presentation. IFA antibody titers of ≥128 were found in 14 of 29 (48.3%), 17 of 30 (56.7%), 13 of 30 (43.4%), and 12 of 30 (40.0%) patients 6, 12, 18, and 24 months after presentation, respectively. Patients reporting additional tick bites during the study had significantly higher antibody titers at most time points during follow-up. No long-term clinical consequences were found during follow-up.
doi:10.1128/CDLI.8.5.899-903.2001
PMCID: PMC96168  PMID: 11527800
3.  Identity of Ehrlichial DNA Sequences Derived from Ixodes ricinus Ticks with Those Obtained from Patients with Human Granulocytic Ehrlichiosis in Slovenia 
Journal of Clinical Microbiology  1999;37(1):209-210.
Adult Ixodes ricinus (Acari: Ixodidae) ticks collected near Ljubljana, Slovenia, were tested for the agent of human granulocytic ehrlichiosis (HGE) by using PCR assays based on the 16S rRNA gene. Three (3.2%) of 93 ticks were found to contain granulocytic ehrlichiae. Nucleotide sequences of portions of the bacterial groESL heat shock operon amplified from these ticks were identical or nearly (99.8%) identical to those previously determined for human patients with HGE from Slovenia, providing additional evidence that the ticks were infected with the HGE agent. This study identified I. ricinus as the likely vector for these ehrlichial pathogens of humans in this part of Europe.
PMCID: PMC84210  PMID: 9854093
4.  A two year prospective study to compare culture and polymerase chain reaction amplification for the detection and diagnosis of Lyme borreliosis. 
Molecular Pathology  1997;50(4):186-193.
AIM: To compare polymerase chain reaction (PCR) amplification of borrelial DNA and culture isolation of spirochaetes for the diagnosis of Lyme borreliosis by direct detection of Borrelia burgdorferi sensu lato in patients with erythema migrans and acrodermatitis chronica atrophicans lesions. METHODS: Skin biopsy specimens from erythema migrans and acrodermatitis chronica atrophicans lesions were subdivided and tested by PCR amplification assay and culture using two artificial growth media, Barbour-Stoenner-Kelly II (BSK II) and modified Kelly-Pettenkofer (MKP). Five classes of lesions were studied: typical erythema migrans, spontaneously resolved erythema migrans, atypical/partially treated erythema migrans, typical acrodermatitis chronica atrophicans, and atypical/partially treated acrodermatitis chronica atrophicans. RESULTS: For both erythema migrans and acrodermatitis chronica atrophicans lesions, the most sensitive detection method was MKP culture. PCR was less sensitive than MKP culture, but more sensitive than BSK II culture. Results for 758 typical erythema migrans specimens showed positivity rates of 36% for MKP, 25% for PCR, and 24% for BSK II. Differences were statistically significant. The overall positivity rate for all three methods combined was 54%, but few specimens (6%) were positive by all three methods. Examination of multiple erythema migrans lesions from the same patient increased the diagnostic yield. These findings, and similar results for acrodermatitis chronica atrophicans lesions, suggest that the distribution of spirochaetes in skin biopsies is not homogeneous. CONCLUSIONS: Although possessing the potential to provide a rapid diagnosis, PCR is not more sensitive than culture for the direct detection of borrelia. Spirochaetes appear to be unevenly distributed throughout biopsy specimens, suggesting that diagnosis of Lyme borreliosis by direct detection of the causative agent in skin lesions in vulnerable to sample bias.
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PMCID: PMC379624  PMID: 9350301
5.  Human disease in Europe caused by a granulocytic Ehrlichia species. 
Journal of Clinical Microbiology  1997;35(6):1556-1559.
Human granulocytic ehrlichiosis (HGE) was recently described in North America. It is caused by an Ehrlichia species closely related to Ehrlichia phagocytophila and Ehrlichia equi, recognized to infect mostly ruminants and horses, respectively. The vector in North America is the tick Ixodes scapularis, which is also the vector of the Lyme disease agent, Borrelia burgdorferi. Previous serologic studies in patients with a diagnosis of Lyme borreliosis indicate that HGE may exist in Europe. We report the first documented case of HGE in Europe. The diagnosis was established by seroconversion to E. equi and the HGE agent and by PCR with sequence analysis of the gene encoding the HGE agent 16S rRNA. Interestingly, the patient presented with a self-limited but moderately severe illness. Thus, European physicians need to be aware that HGE exists in Europe and that the diagnosis should be considered in febrile patients with tick bites in areas where Lyme disease is endemic.
PMCID: PMC229786  PMID: 9163481
6.  Genotypic and phenotypic characterization of Borrelia burgdorferi isolated from ticks and small animals in Illinois. 
Journal of Clinical Microbiology  1995;33(9):2304-2315.
We have characterized 33 isolates of Borrelia burgdorferi from northern Illinois (32 isolates) and Wisconsin (1 isolate) representing the largest series of midwestern isolates investigated to date. The techniques used for molecular analysis of strains included (i) genospecies typing with species-specific PCR primers, (ii) plasmid profiling by pulsed-field gel electrophoresis of total genomic DNA, (iii) large-restriction-fragment pattern (LRFP) analysis by pulsed-field gel electrophoresis of MluI-digested genomic DNA (J. Belfaiza, D. Postic, E. Bellenger, G. Baranton, and I. Saint Girons, J. Clin. Microbiol. 31:2873-2877, 1993), (iv) sodium dodecyl sulfate-polyacrylamide gel electrophoresis of total proteins, (v) microsequencing of high-performance liquid chromatography-purified peptides derived from proteins showing high levels of expression, (vi) amino acid composition analysis of proteins, and (vii) immunological analysis of proteins with a polyclonal antiserum of human origin. Five reference strains as well as two atypical tick isolates from California (DN127) and New York (25015) were included for comparison. All of the Illinois and Wisconsin isolates were typed as B. burgdorferi sensu stricto with genospecies-specific PCR primers. The isolates were found to be heterogeneous with regard to their plasmid and protein profiles. One isolate from Illinois possessed two large-molecular-size plasmids instead of the usual 49-kb plasmid. Fragment patterns resulting from MluI digestion of genomic DNA from the 33 isolates and strains DN127 and 25015 were separable into six distinct LRFPs, five of which have not previously been described. Strain 25015 and an isolate from Illinois (CT39) shared an unusual LRFP that is not typical of other B. burgdorferi sensu stricto strains, suggesting that they may represent a fifth species of B. burgdorferi sensu lato. Five of the 33 isolates and strains DN127 and 25015 showed high-level expression of proteins with molecular masses of approximately 22 kDa. Investigation of these proteins by microsequencing of individual peptides and total amino acid composition analysis indicated that the 22-kDa proteins expressed by the seven strains were polymorphic OspC proteins. By using a polyclonal serum of human origin, expression of OspC could be detected in all 33 Illinois and Wisconsin isolates.
PMCID: PMC228401  PMID: 7494019
7.  In vitro susceptibility of Borrelia burgdorferi to 11 antimicrobial agents. 
The in vitro susceptibility of Borrelia burgdorferi to 11 antimicrobial agents was investigated. The antimicrobial agents evaluated included ceftizoxime, FK037, cefotaxime, dirithromycin, clarithromycin and its metabolite 14-hydroxy-clarithromycin, erythromycin, doxycycline, amoxicillin, ciprofloxacin, and ofloxacin. Isolates of B. burgdorferi tested included two reference strains (B31 and ATCC 53899), six isolates from the midwestern United States, and three from Europe. A broth macrodilution method was used to determine MICs and MBCs. B. burgdorferi was inhibited by < or = 0.5 micrograms of each of the agents except the quinolones per ml. The MBCs for 90% of strains tested of ceftizoxime, FK037, clarithromycin, 14-OH clarithromycin, and dirithromycin (< or = 1.0 microgram of each per ml) were superior to those of amoxicillin (2.0 micrograms/ml) and doxycycline (4.0 micrograms/ml). Further in vivo studies are warranted to determine whether these agents may be efficacious in the treatment of Lyme borreliosis.
PMCID: PMC187991  PMID: 8363373
8.  Influence of ceftriaxone on emergence of Clostridium difficile. 
Antimicrobial Agents and Chemotherapy  1992;36(12):2850-2851.
The influence of long-term ceftriaxone administration on the emergence of Clostridium difficile was studied with 80 patients receiving ceftriaxone for 14 days. In five patients (6.3%) C. difficile was cultured. The appearance of gastrointestinal disturbances during treatment with ceftriaxone was common, but the rate of emergence of C. difficile in immunocompetent patients was not high.
PMCID: PMC245559  PMID: 1482159

Results 1-8 (8)