Survival of Borrelia burgdorferi in antibiotically treated patients with lyme borreliosis

Summary The persistence ofBorrelia burgdorferi in patients treated with antibiotics is described. The diagnosis of Lyme disease is based on clinical symptoms, epidemiology and specific IgG and IgM antibody titers toB. burgdorferi in serum. Antibiotic therapy may abrogate the antibody response to the infection as shown in our patients.B. burgdorferi may persist as shown by positive culture in MKP-medium; patients may have subclinical or clinical disease without diagnostic antibody titers toB. burgdorferi. We conclude that early stage of the disease as well as chronic Lyme disease with persistence ofB. burgdorferi after antibiotic therapy cannot be excluded when the serum is negative for antibodies againstB. burgdorferi.

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Persistenz der Borrelia burgdorferi bei negativer Serologie und Behandlung mit Antibiotika

Zusammenfassung Es wird über die Persistenz vonBorrelia burgdorferi bei sechs Patienten berichtet. Nach dem Zecken- bzw. Insektenstich und Erythema migrans konnteB. burgdorferi noch Wochen nach der Antibiotikatherapie nachgewiesen werden. Serologische Befunde waren außer bei einem Patienten negativ. Diese Ergebnisse bestätigen unsere früheren Beobachtungen und sprechen dafür, dß die Antibiotikabehandlung die Antikörperbildung gegenB. burgdorferi beeinflussen kann. Ferner zeigen diese Ergebnisse und Beobachtungen, daß nicht nur im Frühstadium der Lyme Borreliose, sondern auch in chronischen Stadien bzw. bei Persistenz des Erregers der Nachweis von Antikörpern negativ bleiben kann.

     

Prevalence of antibodies to Borrelia burgdorferi sensu stricto in humans from a Cuban village

Lyme disease has not been officially reported in Cuba. However, clinical cases have been serologically reported. Seroprevalence survey of Borrelia burgdorferi sensu stricto antibodies in humans in the country has not been conducted.ObjectiveTo estimate the prevalence of borrelial antibodies in inhabitants of a village with historically high level of tick infestation.MethodsSerum specimens from 247 persons randomly selected from the population of the village were examined by IgG Western blot using B31 strain for estimating the prevalence of antibodies profile.ResultsA seroprevalence value interval (95% CI) of 0.6%–7.2% was estimated for the studied population. The prevalent borrelial protein bands on immunoblots were 41, 72, 90/93, 34, 47, 60, 58, 56, 65/66 and 31 kDa in a decreasing order of significance.ConclusionThese results support the previous serological findings, suggesting the presence of this borreliosis in Cuba.     

Terrestrial distribution of the Lyme borreliosis agent Borrelia burgdorferi sensu lato in East Asia

Lyme borreliosis is the most prevalent tick-borne zoonosis and an important emerging infection in Europe, North America, and Far Eastern countries. The geographical distribution of Borrelia spp. and the relationship between Borrelia spp. and tick spp. in East Asian countries have been studied. In Northern Asian countries, Ixodes persulcatus carries Eurasian-type Borrelia garinii (20047 type), Borrelia afzelii, and Asian-type B. garinii (variant NT29), whereas Borrelia burgdorferi sensu stricto has not been isolated. In contrast, Asian-type B. garinii has not been found in the European vector tick Ixodes ricinus. These Borrelia spp. cause Lyme borreliosis in their respective countries. The specificity between Borrelia spp. and tick spp. has been confirmed from studies in the Moscow region, which is a sympatric region for the tick spp. I. persulcatus and I. ricinus. In Southeast Asian countries including the southernmost island of Okinawa, the Borrelia valaisiana-related genomic group is carried by Ixodes granulatus. In Japan, a similar borrelia strain Am501, is transmitted by Ixodes columnae. Ixodes ovatus transmits Borrelia japonica but not other species. On other hand, in central China and Nepal, the ticks harbor Borrelia sinica. It is believed that these molecular epidemiological surveys will provide useful information for the diagnoses and prevention of Lyme borreliosis in these countries.     

The propensity of different Borrelia burgdorferi sensu stricto genotypes to cause disseminated infections in humans

Lineages of Borrelia burgdorferi, the bacterium that causes Lyme disease, can be characterized by distinct alleles at the outer surface protein C (ospC) locus. The lineages marked by ospC genotypes have been shown to be differentially invasive in different species of mammals, including humans; genotypes A, B, I, and K effectively disseminate to human blood and cerebrospinal fluid. In this report, we extend the sample of genotypes isolated from human blood to include genotypes N, H, C, M, and D, and rank each by their probability of disseminating from ticks to the blood of humans. Our results demonstrate that only some genotypes of B. burgdorferi present in ticks have a high propensity to disseminate in humans.     

Investigations on the mode and dynamics of transmission and infectivity of Borrelia burgdorferi sensu stricto and Borrelia afzelii in Ixodes ricinus ticks

Borrelia burgdorferi sensu lato (sl), the agent of Lyme disease, is transmitted to the host during the blood meal of Ixodes ticks. In most unfed ticks, spirochetes are present in the midgut and migrate during blood feeding to the salivary glands, from which they are transmitted to the host via saliva. In the present study, the efficiency of Ixodes ricinus ticks to transmit B. afzelii and B. burgdorferi sensu stricto (ss) and their infectivity for mice were examined in relation to the duration of the blood meal. In addition, we investigated whether these two Borrelia species can penetrate intact skin. Three modes of infection of mice were studied: tick-bite infection, inoculation of tick homogenates, and transcutaneous infection by topical application of tick homogenates on mouse skin. Transmission of B. burgdorferi sl from I. ricinus nymphs to mouse increased with duration of tick attachment. B. afzelii-infected ticks start to transmit infection earlier (< or = 48 h) than B. burgdorferi ss-infected ticks. As previously shown for B. burgdorferi ss in Ixodes scapularis, B. burgdorferi ss and B. afzelii in unfed I. ricinus were noninfectious for mice when tick homogenates were inoculated. However, the inoculation of homogenates of ticks fed for 24 h readily produced infection in mice. Therefore, B. burgdorferi ss and B. afzelii spirochetes are potentially infectious in the tick before natural transmission can occur. None of the mice (n = 33) became infected by transcutaneous transmission when tick homogenates were applied on mouse skin, showing that B. burgdorferi ss and B. afzelii are unable to penetrate intact skin, in contrast to relapsing fever spirochetes. This study also shows that B. afzelii is transmitted by I. ricinus to the host earlier than B. burgdorferi ss and that I. ricinus seems to be a more efficient vector of B. afzelii than B. burgdorferi ss.     

Persistence of borrelia burgdorferi in ligamentous tissue from a patient with chronic lyme borreliosis

Objective. To document the persistence of Borrelia burgdorferi in ligamentous tissue samples obtained from a woman with chronic Lyme borreliosis. Methods. Spirochetes were isolated from samples of ligamentous tissue, and the spirochetes were characterized antigenetically and by molecular biology techniques. The ligamentous tissue was examined by electron microscopy. Humoral and cellular immune responses were analyzed. Results. Choroiditis was the first recognized manifestation of Lyme disease in this patient. Despite antibiotic therapy, there was progression to a chronic stage, with multisystem manifestations. The initially significant immune system activation was followed by a loss of the specific humoral immune response and a decrease in the cellular immune response to B burgdorferi over the course of the disease. “Trigger finger” developed, and a portion of the flexor retinaculum obtained at surgery was cultured. Viable spirochetes were identified. Ultramorphologically, the spirochetes were situated between collagen fibers and along fibroblasts, some of which were deeply invaginated by these organisms. The cultured bacteria were identified as B burgdorferi by reactions with specific immune sera and monoclonal antibodies, and by polymerase chain reaction amplification and Southern blot hybridization techniques. Conclusion. To our knowledge, this is the first report of the isolation of B burgdorferi from ligamentous tissue. This suggests that tendon tissues serve as a specific site of spirochete residence in human hosts.     

Association between human leukocyte antigen class II alleles and genotype of Borrelia burgdorferi in patients with early lyme disease

BACKGROUND: On the basis of a polymerase chain reaction-restriction fragment-length polymorphism analysis of the 16S-23S ribosomal DNA intergenic spacer, clinical isolates of Borrelia burgdorferi can be classified into 3 genotypes designated as RST1, RST2, and RST3. RST1 strains are the most pathogenic, and RST3 strains are the least pathogenic. METHODS: Human leukocyte antigen (HLA) class II alleles were determined for a group of culture-positive patients with Lyme disease-associated erythema migrans and were evaluated for an association with the genotype of the infecting B. burgdorferi strain. RESULTS: The DRB1*0101 allele carriage rate was higher in patients infected with RST3 strains (9/25 [36.0%]) than in patients infected with RST1 strains (2/28 [7.1%]) or RST2 strains (7/36 [19.4%]) (P=.010). The same relationship was found for carriage of the DRB1*0101-DQB1*0501 haplotype (P=.018), because of tight linkage disequilibrium. Similar associations could not be demonstrated for any of the other DRB1 and DQB1 alleles or haplotypes that were assessed. CONCLUSION: The DRB1*0101 allele and the DRB1*0101-DQB1*0501 haplotype may be relevant to the development of infection with strains from the least invasive genotypes of B. burgdorferi.     

Lyme borreliosis incidence in two French departments: correlation with infection of Ixodes ricinus ticks by Borrelia burgdorferi sensu lato

We conducted a prospective study to estimate the Lyme borreliosis incidence in two rural French departments, Meuse and Puy-de-D?me. Concurrently, we investigated the prevalence of ticks infected with Borrelia burgdorferi sensu lato (sl) and Anaplasma phagocytophilum. The incidence of Lyme borreliosis decreased from 156 to 109/100,000 inhabitants in Meuse and from 117 to 76/100,000 inhabitants in Puy-de-D?me in 2004 and 2005, respectively, corresponding to a decrease in the density of Ixodes ricinus nymphs infected with B. burgdorferi sl. During the same period, the density of adult ticks increased. Interestingly, B. valaisiana, a nonpathogenic species, infected adult ticks more often than nymphs. These results confirmed the correlation between the Lyme borreliosis incidence and the density of infected nymphs, a stage preferentially infected with B. afzelii. In contrast, we found a low rate of infection by A. phagocytophilum, ranging from 0% to 0.4% in Puy-de-D?me and from 0.8% to 1.4% in Meuse, suggesting a low risk for humans.     

Kill kinetics ofBorrelia burgdorferi and bacterial findings in relation to the treatment of lyme borreliosis

Summary For a better understanding of the persistence ofBorrelia burgdorferi sensu lato (s.l.) after antibiotic therapy the kinetics of killingB. burgdorferi s.l. under amoxicillin, doxycycline, cefotaxime, ceftriaxone, azithromycin and penicillin G were determined. The killing effect was investigated in MKP medium and human serum during a 72 h exposure to antibiotics. Twenty clinical isolates were used, including ten strains ofBorrelia afzelii and ten strains ofBorrelia garinii. The results show that the kinetics of killing borreliae differ from antibiotic to antibiotic. The killing rate of a given antibiotic is less dependent on the concentration of the antibiotic than on the reaction time. Furthermore, the data show that the strains ofB. afzelii andB. garinii have a different reaction to antibiotics used in the treatment of Lyme borreliosis and that different reactions to given antibiotics also exist within one species. TheB. garinii strains appear to be more sensitive to antibiotics used in therapy. Furthermore, the persistence ofB. burgdorferi s.l. and clinical recurrences in patients despite seemingly adequate antibiotic treatment is described. The patients had clinical disease with or without diagnostic antibody titers toB. burgdorferi.

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Abtötungskinetik derBorrelia burgdorferi s.l. — bakteriologische und klinische Befunde

Zusammenfassung Die bakterizide Aktivität von Amoxicillin, Doxycyclin, Cefotaxim, Ceftriaxon, Azithromycin und Penicillin G gegenBorrelia burgdorferi s.l. Stämme wurde in MKP-Medium und Humanserum während der 72 Stunden Einwirkungszeit untersucht. Die Antiborrelienwirkung der Antibiotika wurde an 20 Patientenisolaten, zehnBorrelia afzelii und zehnBorrelia garinii Stämmen, getestet. Der Abtötungseffekt der einzelnen Antibiotika auf getestete Stämme ist sehr unterschiedlich. Die Unterschiede hinsichtlich der Keimreduktion bestehen zwischen den Stämmen derB. afzelii undB. garinii species als auch zwischen den Stämmen innerhalb einer Spezies. Der Anteil der abgetöteten Borrelien ist weniger abhängig von der Konzentration des Antibiotikums als von der Einwirkungszeit. DieB. garinii Stämme sind offensichtlich empfindlicher gegen die in der Therapie eingesetzten Antibiotika. Die Persistenz vonB. burgdorferi s.l. und Rezidive der Erkrankung nach der Antibiotikatherapie werden diskutiert.

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This article is the expanded version of a lecture delivered in Portoroz on 14 May 1995. See also page 59.     

Seasonality of Ixodes ricinus ticks on vegetation and on rodents and Borrelia burgdorferi sensu lato genospecies diversity in two lyme borreliosis-endemic areas in Switzerland

We compared Ixodes ricinus questing density, the infestation of rodents by immature stages, and the diversity of Borrelia burgdorferi sensu lato (sl) in questing ticks and ticks collected from rodents in two Lyme borreliosis (LB)-endemic areas in Switzerland (Portes-Rouges [PR] and Staatswald [SW]) from 2003 to 2005. There were variations in the seasonal pattern of questing tick densities among years. Questing nymphs were globally more abundant at PR than at SW, but the proportion of rodents infested by immature ticks was similar (59.4% and 61%, respectively). Questing tick activity lasted from February to November with a strong decline in June. The seasonal pattern of ticks infesting rodents was different. Ticks infested rodents without decline in summer, suggesting that the risk of being bitten by ticks remains high during the summer. Rodents from SW showed the highest infestation levels (10±21.6 for larvae and 0.54±1.65 for nymphs). The proportion of rodents infested simultaneously by larvae and nymphs (co-feeding ticks) was higher at SW (28%) than at PR (11%). Apodemus flavicollis was the species the most frequently infested by co-feeding ticks, and Myodes glareolus was the most infective rodent species as measured by xenodiagnosis. At PR, the prevalence of B. burgdorferi sl in questing ticks was higher (17.8% for nymphs and 32.4% for adults) than at SW (10.4% for nymphs and 24.8% for adults), with B. afzelii as the dominant species, but B. garinii, B. burgdorferi sensu stricto, and B. valaisiana were also detected. Rodents transmitted only B. afzelii (at PR and at SW) and B. bavariensis (at SW) to ticks, and no mixed infection by additional genospecies was observed in co-feeding ticks. This implies that co-feeding transmission does not contribute to genospecies diversity. However, persistent infections in rodents and co-feeding transmission contribute to the perpetuation of B. afzelii in nature.     

Three major Lyme Borrelia genospecies (Borrelia burgdorferi sensu stricto,B. afzelii and B. garinii) identified by PCR in cerebrospinal fluid from patients with neuroborreliosis in Sweden

The Lyme Borrelia genospecies Borrelia afzelii and B. garinii have previously been isolated using a culture method in Swedish patients with Lyme borreliosis (LB). There are reports suggesting that the genospecies distribution in human tissue specimens as determined by molecular methods is different from that obtained by culture. In the present study, we developed a nested PCR for detection of Lyme Borrelia-specific DNA in cerebrospinal fluid from Swedish patients with LB. The genospecies were subsequently identified by sequence analysis in a total of 7 PCR-positive patients. Two sequences were identified as B. burgdorferi sensu stricto (s. s.), 1 as B. afzelii and 4 as B. garinii. These are the first reported cases in which B. burgdorferi s. s. has been shown to be the causative agent of human LB in Sweden. The results of our study confirm that the use of direct molecular analytical methods for Borrelia genospecies identification in clinical specimens can provide epidemiological information additional to that obtained by culture.     

Detection of Borrelia burgdorferi by DNA amplification in synovial tissue samples from patients with lyme arthritis

Objective. To compare the detection rates of chromosomal flagellin gene from Borrelia burgdorfen in synovial tissue (ST) and synovial fluid (SF) using polymerase chain reaction (PCR) techniques. Methods. B burgdorferi DNA was sought in SF and ST from 12 consecutive patients with Lyme arthritis and from 29 patients with noninfectious diseases (controls). Results. No DNA amplification was observed in samples obtained from the 29 control patients, whereas B burgdorferi DNA was detected in all ST and/or SF samples from the 12 patients with Lyme arthritis. Results from 1 ST sample were not interpretable because of PCR inhibitors. Among the 11 remaining patients, 10 had positive ST samples, whereas only 4 had positive SF samples (P < 0.05). Conclusion. These data suggest that detection of chromosomal B burgdorferi DNA may be more efficient in ST than SF.     

Molecular and serological evidence of Borrelia burgdorferi sensu lato in wild rodents in the Czech Republic

The aim of the present study was to determine the frequency and spatial distribution of the Borrelia species in wild rodents in the Czech Republic. In total, 293 muscle tissue samples and 106 sera from 293 wild rodents captured in North Bohemia and North-East and South Moravia were examined for the presence of Borrelia spp. and antibodies. Muscle samples were investigated with real-time polymerase chain reaction (PCR) with a recA primer set, with DNA quantification and melting curve analysis, and with restriction fragment length polymorphism (RFLP) analysis of the 5S-23S rDNA intergenic spacer. Infection with Borrelia burgdorferi sensu lato was found in 16.4% of the muscle samples. The most abundant genospecies was Borrelia afzelii (11.3%), followed by Borrelia burgdorferi sensu stricto (4.8%) and Borrelia garinii (0.7%). Borrelia infection was more frequently observed in Clethrionomys glareolus than in Apodemus spp. Sera were analyzed with an enzyme-linked immunosorbent assay (ELISA) test, yielding the total seropositivity rates of 24.5% for anti-Borrelia IgM antibodies and 25.5% for IgG antibodies. Total seroprevalence was higher in Apodemus spp. than in C. glareolus. In conclusion, our data indicate that in the Czech Republic small wild rodents can serve as hosts for B. burgdorferi s. s. as well as for B. afzelii.     

Significance of red deer (Cervus elaphus) in the ecology of Borrelia burgdorferi sensu lato

BACKGROUND: Red deer (Cervus elaphus) is one of the most important host of the adult tick (Ixodes ricinus) which is the basic vector of the Lyme disease causative agent--Borrelia burgdorferi sensu lato in Europe. The aim of the present study was to establish the role of red deer in the transmission of B. burgdorferi s.1. Material and methods. Tissues from 74 red deers were evaluated and the presence of B. burgdorferi s.1 DNA was identified using nested PCR technique based on fla gene. The identification of species belonging to B. burgdorferi s.1 complex was performed after restriction digestion of nested PCR product with Ddel enzyme and sequencing of nested PCR product. The study included also 55 isolates of I. ricinus females removed from red deer and 466 ticks (73 adult and 393 nymphs) collected from the vegetation in the area where the red deer lives. RESULTS: There were no DNA of B. burgdorferi s.1 complex in the red deer tissues and in ticks removed from deer, however in one tick removed from deer the DNA of other Borrelia species--B. miyamotoi was identified. In ticks collected from vegetation 3 species belonging to B. burgdorferi s.1. complex were identified: B. garinii (3.2% ticks studied), B. afzelii (6.9%) and B. valaisiana (3.6%), however DNA of B. miyamotoi was absent. These results confirm inability of survival of B. burgdorferi s.1. species in tick I. ricinus feeding on red deer blood. However there is a possibility of survival of B. miyamotoi in presence of deer blood at least in ticks feeding on red deer. The main role of red deer in keeping the constant infection level of B. burgdorferi s.1. in the whole population of I. ricinus ticks does not concern B. miyamotoi.