Skip to main page content

• HOME
• Current Issue
• Archive
• Alerts
• About ASM
• Contact us
• Tech Support
• Journals.ASM.org

This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Derdáková, M. Articles by Fish, D. Search for Related Content PubMed PubMed Citation Articles by Derdáková, M. Articles by Fish, D. Agricola Articles by Derdáková, M. Articles by Fish, D.

 Previous Article  |  Next Article 

Applied and Environmental Microbiology, November 2004, p. 6783-6788, Vol. 70, No. 11
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.11.6783-6788.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Interaction and Transmission of Two Borrelia burgdorferi Sensu Stricto Strains in a Tick-Rodent Maintenance System

Markéta Derdáková,^1,2 Vladimír Dudiòák,² Brandon Brei,^1, John S. Brownstein,¹ Ira Schwartz,³ and Durland Fish^1*

Vector Ecology Laboratory, Department of Epidemiology and Public Health, Yale School of Medicine, New Haven, Connecticut,¹ Parasitological Institute, Slovak Academy of Science, Koice, Slovak Republic,² Department of Microbiology and Immunology, New York Medical College, Valhalla, New York³

Received 20 February 2004/ Accepted 11 June 2004

Top Abstract Introduction Materials and Methods Results Discussion References

 
In the northeastern United States, the Lyme disease agent, Borrelia burgdorferi sensu stricto, is maintained by enzoonotic transmission, cycling between white-footed mice (Peromyscus leucopus) and black-legged ticks (Ixodes scapularis). B. burgdorferi sensu stricto is genetically variable and has been divided into three major genotypes based on 16S-23S ribosomal DNA spacer (RST) analysis. To better understand how genetic differences in B. burgdorferi sensu stricto may influence transmission dynamics in nature, we investigated the interaction between an RST1 and an RST3 strain in a laboratory system with P. leucopus mice and I. scapularis ticks. Two groups of mice were infected with either BL206 (RST1) or B348 (RST3). Two weeks later, experimental mice were challenged with the opposite strain, while control mice were challenged with the same strain as that used for the primary infection. The transmission of BL206 and B348 from infected mice was then determined by xenodiagnosis with uninfected larval ticks at weekly intervals for 42 days. Mice in both experimental groups were permissive for infection with the second strain and were able to transmit both strains to the xenodiagnostic ticks. However, the overall transmission efficiencies of BL206 and B348 were significantly different. BL206 was more efficiently transmitted than B348 to xenodiagnostic ticks. Significantly fewer double infections than expected were detected in xenodiagnostic ticks. The results suggest that some B. burgdorferi sensu stricto strains, such as BL206, may be preferentially maintained in transmission cycles between ticks and white-footed mice. Other strains, such as B348, may be more effectively maintained in different tick-vertebrate transmission cycles.

Top Abstract Introduction Materials and Methods Results Discussion References

 
Borrelia burgdorferi sensu lato spirochetes are maintained in nature by enzoonotic transmission, cycling between vector ticks of the genus Ixodes and a variety of vertebrate reservoir hosts (4, 19, 35). The B. burgdorferi sensu lato complex is highly heterogeneous, with 11 described species (1, 5, 8, 20, 24, 30, 31, 38, 44). B. burgdorferi sensu stricto, Borrelia afzelii, and Borrelia garinii are known to cause Lyme disease in humans and are associated with different clinical manifestations (43). Intraspecific differences within genospecies are probably also important determinants of Lyme disease pathogenesis and ecology (2, 11, 13, 14, 28, 36, 42, 46, 47, 48, 49, 50).

In the northeastern United States, B. burgdorferi sensu stricto is the only agent of Lyme disease. This species is characterized by a high level of genetic variability (11, 27, 28, 48). Based on restriction fragment length polymorphism (RFLP) analysis of the 16S-23S rRNA intergenic spacer, B. burgdorferi sensu stricto isolates from Lyme disease patients have been subdivided into three major rRNA spacer types (RST) (27, 28). There is strong evidence that these distinct genotypes are associated with differing pathologies and dissemination patterns in humans (50). Similarly, isolation success, pathogenicity, and infection kinetics differ significantly between RST1 and RST3 genotypes in C3H/HeJ mice (46, 47). Mixed infections with different B. burgdorferi sensu stricto genotypes occur in questing ticks (11, 45, 48), experimental animals (12), and humans (29). However, interactions between coinfecting spirochetes have not been well studied.

The principal reservoir host in the northeastern United States is the white-footed mouse, Peromyscus leucopus (6, 26). This abundant species can acquire an infection from a single infected nymph and typically remains infective for at least 7 months (6). Other vertebrates such as Eastern chipmunks (Tamias striatus), meadow voles (Microtus pensylvanicus), raccoons (Procyon lotor), Eastern gray squirrels (Sciurus carolinensis), striped skunks (Mephitis mephitis), and American robins (Turdus migratorius) are also competent reservoir hosts (7, 32, 40). Thus, other vertebrate hosts for Ixodes scapularis may also contribute to the enzootic maintenance of B. burgdorferi sensu stricto.

We used a laboratory system with P. leucopus mice and I. scapularis ticks to study the influence of genetic differences in B. burgdorferi sensu stricto strains on transmission dynamics in nature. We investigated the interaction between two genetically distinct B. burgdorferi sensu stricto strains, representative of two distinct genotypes, in order to better understand how multiple B. burgdorferi sensu stricto strains are maintained in nature.

Top Abstract Introduction Materials and Methods Results Discussion References

 
B. burgdorferi isolates.
Two B. burgdorferi sensu stricto isolates, BL206 and B348, were used. These strains originated from two different Lyme disease patients residing in Westchester County, N.Y. BL206 was isolated from the blood of one patient and B348 was isolated from an erythema migrans lesion from the second patient. BL206 and B348 were genotyped as having RST1 and RST3 genotypes, respectively (27, 28).

Rodents.
Specific-pathogen-free adult P. leucopus mice were obtained from the Peromyscus Genetic Stock Center (Columbia, S.C.). The mice were housed in separate cages maintained at 21 to 24°C and were handled humanely. Prior to tick infestations, mice were anesthetized by an intraperitoneal injection with ketamine (44 mg/kg of body weight).

Ticks.
Uninfected I. scapularis larvae originated from the egg clutches of female ticks collected in the field (Connecticut) and fed on uninfected sheep. Random samples of larvae from the colony were tested for the presence of B. burgdorferi sensu stricto as described below. I. scapularis nymphs infected with the BL206 or B348 strain were reared by a modification of the protocol of Piesman (34). Briefly, specific-pathogen-free C3H mice obtained from Charles River Laboratories (Wilmington, Mass.) were inoculated intraperitoneally with 10⁴ spirochetes. Uninfected larvae were allowed to feed on mice at 14, 21, and 28 days postinfection. More than 100 larvae were collected from each mouse. Replete larvae molted to nymphs and were examined for spirochetes 1 month after molting by a PCR-based assay as described below. Nymphal infection rates for strains BL206 and B348 were 82 and 0%, respectively.

To produce nymphs infected with B348, we inoculated specific-pathogen-free 10-week-old CB-17/scid (severe combined immunodeficient) mice intraperitoneally with 10⁸ spirochetes. Uninfected larvae were allowed to feed on these mice at 30 days postinfection. Replete larvae molted to nymphs and were examined for spirochetes 1 month after molting by PCR as described below. The nymphal infection rate was 97%. Before and after feeding, all ticks in this study were maintained in an environmental chamber at 21°C and 80% relative humidity.

B. burgdorferi transmission and acquisition.
The experimental design for this study is presented in Fig. 1. Four treatment groups were composed of three P. leucopus mice each. The two experimental groups were designated E1 and E3 and the control groups were designated C1 and C3. On day 0, primary infections were established by feeding 12 nymphs infected with BL206 to repletion on each mouse in groups E1 and C1. Twelve nymphs infected with B348 were fed to repletion on each mouse in groups E3 and C3. Infested mice were kept in wire-bottomed cages over water-filled pans. Two weeks later, E1 and E3 mice were challenged with B348- and BL206-infected nymphs, respectively, whereas C1 and C3 mice were challenged with nymphs infected with the same strain as that used for the primary infection (BL206 and B348, respectively).

View larger version (41K): [in this window] [in a new window]

FIG. 1. Experimental design. Mice were divided into four groups as follows: group E1, infected with BL206-infected nymphs (day 0) and challenged with B348-infected nymphs (day 14); group C1, infected and challenged with BL206-infected nymphs (days 0 and 14); group E3, infected with B348-infected nymphs (day 0) and challenged with BL206-infected nymphs (day 14); group C3, infected and challenged with B348-infected nymphs (days 0 and 14). The infection of mice was determined by xenodiagnosis with 100 uninfected larvae per mouse on days 10, 21, 28, and 42.

The infection of mice was determined by xenodiagnosis with 100 uninfected larvae per mouse on days 10, 21, 28, and 42 after the initial infection (Xeno10, Xeno21, Xeno28, and Xeno42 ticks). Engorged larvae were collected daily and allowed to molt into nymphs. Random samples of 12 molted nymphs per mouse were individually tested by PCR for the presence of Borrelia spirochetes. For each PCR-positive tick, the RST genotype(s) of the B. burgdorferi strain(s) was determined by RFLP analysis of the 16S-23S ribosomal DNA spacer (28), which also allowed the detection of mixed infections.

DNA extraction.
DNAs were extracted from tick samples by use of a DNeasy tissue kit (Qiagen, Valencia, Calif.) according to a modified protocol (3). Briefly, each tick was cut with a disposable sterile scalpel, and proteins were degraded overnight at 56°C in 180 µl of buffer ATL (Qiagen) and 20 µl of proteinase K (14 mg/ml) (Boehringer Mannheim, Indianapolis, Ind.). The remaining extraction steps were performed according to the manufacturer's protocol. DNAs were eluted in 50 µl of deionized water and stored at 4°C.

PCR amplification.
B. burgdorferi sensu stricto was detected by amplifying a 941-bp fragment of the 16S-23S ribosomal DNA intergenic spacer by nested PCR as described previously (29). A MasterTaq DNA Polymerase kit (Eppendorf, Westbury, N.Y.) was used for both rounds of PCR amplification. For each 5.0-µl DNA sample, the PCR mixture contained 20.8 µl of deionized water, 1x TaqMaster PCR enhancer, 1x Taq buffer (with 15 mM Mg²⁺), 1.5 mM Mg(OAc)₂, 1 U of Taq DNA polymerase, 10 mM deoxynucleoside triphosphate mix (200 µM each) (Eppendorf), and 25 pmol of each primer (Invitrogen, Frederick, Md.). Both the first- and second-round reactions were run for 35 cycles of denaturation at 94°C for 30 s, annealing at 52°C for 30 s, and extension at 72°C for 30 s. Prior to RFLP analysis, amplified PCR products were purified by use of a QIAquick PCR purification kit (Qiagen), eluted in 25 µl of deionized water, and stored at 4°C.

RFLP analysis.
RFLP analysis of positive samples from groups E1 and E3 was performed as described elsewhere (28). Ten microliters of a nested amplification product was digested at 37°C for 9 h in a solution containing 0.2 µl of Hinf1 (10,000 U/ml) (New England Biolabs, Beverly, Mass.), 1.5 µl of 10x NE buffer 2 (New England Biolabs), and 3.3 µl of deionized water. The enzyme was heat inactivated at 85°C for 20 min. Electrophoresis was carried out in 1.5% agarose gels containing 0.5 µg of ethidium bromide per ml. Bands were visualized on a UV transilluminator.

Statistical analysis.
Differences in the prevalence of BL206 and B348 and the dependence of time and strain were compared by a multiple analysis of covariance. A posthoc analysis of the Tukey honestly significant difference test was used to compare the prevalence of BL206 and B348 in different groups of mice (E1, E3, C1, and C3). Differences between the expected and actual prevalence of multiple infections were compared by the chi-square test.

Top Abstract Introduction Materials and Methods Results Discussion References

 
Tick-mediated transmission and acquisition of B. burgdorferi.
All mice that were exposed initially to BL206-infected ticks (groups E1 and C1) acquired infections and together transmitted spirochetes to 58.3% of xenodiagnostic larvae after 10 days (Xeno10) (Table 1). After a challenge with B348-infected nymphs, the overall percentage of infected ticks decreased to 69.4, 58.3, and 38.9% after an additional 7, 14, and 28 days (Xeno21, Xeno28, and Xeno42, respectively) (Table 1; Fig. 2). A further evaluation by RFLP analysis revealed that the original BL206 infections were maintained in 50, 58.3, and 30.5% of Xeno21, Xeno28, and Xeno42 ticks, respectively. In contrast, B348 infections were detected in 19.4, 0, and 8.3% of Xeno21, Xeno28, and Xeno42 ticks (Table 1; Fig. 2). No mixed infections were detected in E1 ticks. Overall, during the three consecutive xenodiagnostic infestations (days 21, 28, and 42), 46.3% of the xenodiagnostic ticks from mice with mixed infections acquired BL206 and 9.3% acquired B348 (Table 2).

View this table: [in this window] [in a new window]

TABLE 1. Infection of xenodiagnostic I. scapularis ticks with B. burgdorferi sensu stricto strains BL206 and B348 in different groups of P. leucopus

View larger version (22K): [in this window] [in a new window]

FIG. 2. Progress of infection of xenodiagnostic ticks with BL206 (blue), B348 (red), and a mixture of B348 and BL206 (green). Percentages of B. burgdorferi sensu stricto strains in four groups of mice were determined after 10, 21, 28, and 42 days. E1, infected with BL206 and challenged with B348; E3, infected with B348 and challenged with BL206; C1, infected and challenged with BL206; C3, infected and challenged with B348.

View this table: [in this window] [in a new window]

TABLE 2. Overall infection of xenodiagnostic I. scapularis ticks fed upon mice infected with B. burgdorferi sensu stricto strains BL206 and/or B348 during three consecutive infestations

All mice that were exposed initially to B348-infected ticks (group E3 and C3) became infected and together transmitted spirochetes to 83.3% of xenodiagnostic ticks on day 10 (Table 1). After a challenge with BL206, the overall percentages of B. burgdorferi infection were maintained at 50, 75, and 66.7% in Xeno21, Xeno28, and Xeno42 ticks, respectively (Table 1; Fig. 2). RFLP analysis revealed that BL206 infected 16.7, 61.1, and 55.6% of Xeno21, Xeno28, and Xeno42 ticks, respectively, whereas the original B348 infection was maintained in only 33.3% of Xeno21, 11.1% of Xeno28, and 11.1% of Xeno42 ticks (Table 1; Fig. 2). A single xenodiagnostic tick acquired dual infections. Overall, during the three consecutive xenodiagnostic infestations (days 21, 28, and 42), 45.4% of the xenodiagnostic ticks from mice with mixed infections acquired BL206 and 19.4% acquired B348 (Table 2).

The B. burgdorferi infection rate for mice that were initially infected with BL206 and subsequently challenged on day 14 with BL206 nymphs infected with the same strain (group C1) gradually increased from 66.7% on day 21 to 83.3% by day 42 (Fig. 2). Overall, 75% of all xenodiagnostic ticks placed on mice in this group acquired a BL206 infection (Table 2). In contrast, for mice that were challenged in a similar manner with B348-infected nymphs (group C3), the percentage of infected ticks decreased from 54.2 to 4.1% (Table 1; Fig. 2), and only 34.7% of all diagnostic ticks acquired infection (Table 2).

Strain and group comparisons.
A multiple analysis of covariance showed significant differences in the infection rates of ticks with BL206 and B348 over time (F = 3.18, P < 0.05, df = 2) and with the strain (F = 26.19, P < 0.001, df = 3). A posthoc analysis with the Tukey honestly significant difference test showed significant differences between the infection rates of BL206 and B348 for xenodiagnostic ticks from both experimental groups (for group E1, P < 0.001; for group E3, P < 0.001). Infection rates of ticks with BL206 significantly differed between the experimental and control groups (for E1 versus C1, P < 0.01; for E3 versus C1, P < 0.01). Infection rates of ticks with B348 also significantly differed between the experimental and control groups (for E1 versus C3, P < 0.01; for E3 versus C3, P < 0.001). Overall, the infection rates of ticks also significantly differed between the control groups (C1 versus C3, P < 0.001). Neither strain's infection rates significantly differed between the experimental groups, E1 and E3 (for BL206, P = 0.979; for B348, P = 0.721). The frequency of mixed infections in xenodiagnostic ticks was significantly lower than expected (P < 0.001, ² = 3.84) according to a chi-square analysis.

Top Abstract Introduction Materials and Methods Results Discussion References

 
Several studies have recently reported considerable B. burgdorferi sensu stricto genetic heterogeneity in the United States. Wang et al. (46, 47) observed that in experimentally infected C3H mice, spirochete dissemination and the severity of symptoms depend on the infecting B. burgdorferi sensu stricto strain. Thus, genetic variability within the genospecies is probably an important factor in the pathogenesis of Lyme disease (46, 47). Specific associations between reservoir hosts and B. burgdorferi sensu lato genetic variants in Europe and Asia have been reported (9, 14, 15, 16, 17, 18, 21, 22, 23). However, the ecology and transmission of different B. burgdorferi sensu stricto strains among reservoir hosts and vector ticks are not well studied for North America or Eurasia. The main purpose of this study was to determine how B. burgdorferi sensu stricto genetic heterogeneity might influence transmission dynamics in nature.

Infected P. leucopus mice in both experimental groups (group E1 was infected with BL206 and challenged with B348, and group E3 was infected with B348 and challenged with BL206) were permissive for infection with a second strain of B. burgdorferi sensu stricto. The mice established mixed infections and were able to transmit both strains to xenodiagnostic ticks (Table 1; Fig. 2). Previously, Hofmeister et al. (12) described experimental infections of C3H mice with heterologous B. burgdorferi sensu stricto clones in North America by coinfection and sequential infection. Additionally, many authors have reported mixed infections of different Borrelia genospecies in reservoir hosts from Eurasia (15, 16, 33, 37). Despite the presence of coinfections with both BL206 and B348 spirochetes in experimentally infected mice, only one dual infection was detected among xenodiagnostic ticks. This was a significantly lower frequency (P < 0.001, ² = 3.84) than would be expected by independent assortment of the strains, which suggests a possible interference interaction between these two strains. Such interference has been demonstrated between Anaplasma phagocytophilum and B. burgdorferi (25). Alternatively, genetically distinct Borrelia populations may be preferentially selected for in the tick while it feeds on the host (41).

Although mice were able to establish multistrain infections, the overall efficiency of transmission of BL206 to xenodiagnostic ticks was significantly higher than that of B348. The results suggest that BL206 is more efficiently transmitted from P. leucopus mice to ticks. Similar findings of the dependence of transmission on genotype were obtained in England, where B. burgdorferi sensu stricto and B. garinii were detected in wild rodents, but only B. burgdorferi sensu stricto was transmitted to xenodiagnostic ticks (21).

P. leucopus was susceptible to infection with both strains (BL206 and B348), but the abilities of mice to maintain and/or transmit the two strains to uninfected larvae were different. In mice sequentially infected with BL206, infections were efficiently maintained and transmitted to xenodiagnostic ticks at a consistently high level (Table 1; Fig. 2). In contrast, in mice sequentially infected with isolate B348, the percentage of ticks infected with B348 started decreasing after day 28 (Table 1; Fig. 2). Since both infected nymphs and uninfected xenodiagnostic larvae were placed on P. leucopus ears, xenodiagnostic larvae probably became infected with B348 mostly by means of localized "extended cofeeding" before systemic infections of mice could develop (10, 39).

Wang et al. observed two- to three-fold larger spirochete burdens in the skin, heart, and joint tissues of C3H/HeJ mice that were experimentally infected with BL206 than those in mice infected with B348 (47). Furthermore, the culture positivity rate from the blood was higher for mice infected with BL206 than for mice infected with B348. This may be another explanation for the differences in acquisition of strains BL206 and B348 by xenodiagnostic larvae. Alternatively, B348 and BL206 may have different tissue tropisms that affect their ease of transmission to feeding ticks. This was reported from Japan, where different borrelial genospecies were isolated from tissues of Apodemus speciosus mice (33).

The persistent infection rate of BL206 in xenodiagnostic ticks observed in this study suggests that this strain is well tolerated by P. leucopus and that this mouse species is an efficient reservoir host for this strain. The decreasing percentage of infected ticks with B348 from the group with sequential homologous infections (C3) indicates that P. leucopus is not an efficient reservoir for the B348 strain. The BL206 and B348 strains may therefore be maintained by different reservoir host species.

It should be noted that this study was undertaken with two B. burgdorferi isolates representing two different RSTs. Whether the differences in maintenance and transmission observed here are broadly reflective of all members of these RSTs must await confirmation by further investigations with additional isolates.

B348, as well as other B. burgdorferi genotypes belonging to the same restriction type (RST3), has been isolated from patients. B. burgdorferi genotypes belonging to the RST3 genotype were also detected in host-seeking ticks in the northeastern United States (45). Since our study suggests that P. leucopus is not the most suitable reservoir host for this human pathogen, the competence of other possible reservoir hosts warrants further investigation. Our study supports the hypothesis that the genetic heterogeneity of B. burgdorferi sensu stricto is an important determinant of Lyme disease ecology. We propose that, like the case in Europe, different reservoir-pathogen associations maintain Borrelia genetic variants. Field experiments are needed to further evaluate this hypothesis.

 
This research was sponsored by a Fulbright Fellowship and grant SPVV 2003 SP 51/028-09-08 to Marketa Derdáková, by NIH grant SROI-AR41511 to Ira Schwartz, and by USDA-ARS (cooperative agreement 58-1265-7-002) and a Mathers Foundation grant to Durland Fish.

We thank Lorenza Beati, Jean Tsao, Dionysios Liveris, and Guiqing Wang for their helpful comments.

 
* Corresponding author. Mailing address: Department of Epidemiology and Public Health, Yale School of Medicine, P.O. Box 208034, New Haven, CT 06520-8034. Phone: (203) 785-3525. Fax: (203) 785-3604. E-mail: durland.fish{at}yale.edu.

Deceased.

Top Abstract Introduction Materials and Methods Results Discussion References

 

1
1. Baranton, G., D. Postic, I. Saint Girons, P. Boerlin, J. C. Piffaretti, M. Assous, and P. A. Grimont. 1992. Delineation of Borrelia burgdorferi sensu stricto, Borrelia garinii sp. nov., and group VS461 associated with Lyme borreliosis. Int. J. Syst. Bacteriol. 42:378-383.[Abstract/Free Full Text]
2
2. Baranton, G., G. Seinost, G. Theodore, D. Postic, and D. Dykhuizen. 2001. Distinct levels of genetic diversity of Borrelia burgdorferi are associated with different aspects of pathogenicity. Res. Microbiol. 152:149-156.[Medline]
3
3. Beati, L., and J. E. Keirans. 2001. Analysis of the systematic relationships among ticks of the genera Rhipicephalus and Boophilus (Acari: Ixodidae) based on mitochondrial 12S ribosomal DNA gene sequences and morphological characters. J. Parasitol. 87:32-48.[CrossRef][Medline]
4
4. Burgdorfer, W., A. G. Barbour, S. F. Hayes, J. L. Benach, E. Grunwaldt, and J. P. Davis. 1982. Lyme disease—a tick-borne spirochetosis? Science 216:1317-1319.[Abstract/Free Full Text]
5
5. Canica, M. M., F. Nato, L. du Merle, J. C. Mazie, G. Baranton, and D. Postic. 1993. Monoclonal antibodies for identification of Borrelia afzelii sp. nov. associated with late cutaneous manifestations of Lyme borreliosis. Scand. J. Infect. Dis. 25:441-448.[Medline]
6
6. Donahue, J. G., J. Piesman, and A. Spielman. 1987. Reservoir competence of white-footed mice for Lyme disease spirochetes. Am. J. Trop. Med. Hyg. 36:92-96.
7
7. Fish, D., and T. J. Daniels. 1990. The role of medium-sized mammals as reservoirs of Borrelia burgdorferi in southern New York. J. Wildl. Dis. 26:339-345.[Abstract]
8
8. Fukunaga, M., A. Hamase, K. Okada, and M. Nakao. 1996. Borrelia tanukii sp. nov. and Borrelia turdae sp. nov. found from ixodid ticks in Japan: rapid species identification by 16S rRNA gene-targeted PCR analysis. Microbiol. Immunol. 40:877-881.[Medline]
9
9. Gern, L., A. Estrada-Pea, F. Frandsen, J. S. Gray, T. G. T. Jaenson, F. Jongejan, O. Kahl, E. Korenberg, R. Mehl, and P. A. Nuttall. 1998. European reservoir hosts of Borrelia burgdorferi sensu lato. Zentbl. Bakteriol. 287:196-204.
10
10. Gern, L., and O. Rais. 1996. Efficient transmission of Borrelia burgdorferi between cofeeding Ixodes ricinus ticks (Acari: Ixodidae). J. Med. Entomol. 33:189-192.[Medline]
11
11. Guttman, D. S., P. W. Wang, I. N. Wang, E. M. Bosler, B. J. Luft, and D. E. Dykhuizen. 1996. Multiple infections of Ixodes scapularis ticks by Borrelia burgdorferi as revealed by single-strand conformation polymorphism analysis. J. Clin. Microbiol. 34:652-656.[Abstract]
12
12. Hofmeister, E. K., G. E. Glass, J. E. Childs, and D. H. Persing. 1999. Population dynamic of naturally occurring heterogeneous mixture of Borrelia burgdorferi clones. Infect. Immun. 67:5709-5716.[Abstract/Free Full Text]
13
13. Hu, C. M., B. Wilske, V. Fingerle, Y. Lobet, and L. Gern. 2001. Transmission of Borrelia garinii OspA serotype 4 to BALB/c mice by Ixodes ricinus ticks collected in the field. J. Clin. Microbiol. 39:1169-1171.[Abstract/Free Full Text]
14
14. Huegli, D., C. M. Hu, P.-F. Humair, B. Wilske, and L. Gern. 2002. Apodemus species mice are reservoir hosts of Borrelia garinii OspA serotype 4 in Switzerland. J. Clin. Microbiol. 40:4735-4737.[Abstract/Free Full Text]
15
15. Humair, P.-F., and L. Gern. 1998. Relationship between Borrelia burgdorferi sensu lato species, red squirrels (Sciurus vulgaris) and Ixodes ricinus in enzootic areas in Switzerland. Acta Trop. 69:213-227.[CrossRef][Medline]
16
16. Humair, P.-F., D. Postic, R. Wallich, and L. Gern. 1998. An avian reservoir (Turdus merula) of the Lyme disease spirochetes. Zentbl. Bakteriol. 287:521-538.
17
17. Humair, P.-F., O. Rais, and L. Gern. 1999. Transmission of Borrelia afzelii from Apodemus mice and Clethrionomys voles to Ixodes ricinus ticks: differential transmission pattern and overwintering maintenance. Parasitology 118:33-42.
18
18. Humair, P.-F., and L. Gern. 2000. The wild hidden face of Lyme borreliosis in Europe. Microbes Infect. 2:915-922.[CrossRef][Medline]
19
19. Johnson, R. C., G. P. Schmid, F. W. Hyde, A. G. Steigerwalt, and D. J. Brenner. 1984. Borrelia burgdorferi sp. nov.: etiologic agent of Lyme disease. Int. J. Syst. Bacteriol. 34:496-497.
20
20. Kawabata, H., T. Masuzawa, and Y. Yanagihara. 1993. Genomic analysis of Borrelia japonica sp. nov. isolated from Ixodes ovatus in Japan. Microbiol. Immunol. 37:843-848.[Medline]
21
21. Kurtenbach, K., M. Peacey, S. G. T. Rijpkema, A. N. Hoodless, P. A. Nuttall, and S. E. Randolph. 1998. Differential transmission of the genospecies of Borrelia burgdorferi sensu lato by game birds and small rodents in England. Appl. Environ. Microbiol. 64:1169-1174.[Abstract/Free Full Text]
22
22. Kurtenbach, K., H. S. Sewell, N. H. Ogden, S. E. Randolph, and P. A. Nuttall. 1998. Serum complement sensitivity as a key factor in Lyme disease ecology. Infect. Immun. 66:1248-1251.[Abstract/Free Full Text]
23
23. Kurtenbach, K., S. DeMichelis, S. Etti, S. M. Schäfer, H.-S. Sewell, V. Brade, and P. Kraiczy. 2002. Host association of Borrelia burgdorferi sensu lato—the key role of host complement. Trends Microbiol. 10:74-79.[CrossRef][Medline]
24
24. Le Fleche, A., D. Postic, K. Girardet, O. Péter, and G. Baranton. 1997. Characterization of Borrelia lusitaniae sp. nov. by 16S ribosomal sequence analysis. Int. J. Syst. Bacteriol. 47:921-925.[Abstract/Free Full Text]
25
25. Levin, M., and D. Fish. 2001. Interference between the agents of Lyme disease and human granulocytic ehrlichiosis in a natural reservoir host. Vector Borne Zoonotic Dis. 1:139-148.[CrossRef][Medline]
26
26. Levine, J. F., M. L. Wilson, and A. Spielman. 1985. Mice as reservoirs of the Lyme disease spirochete. Am. J. Trop. Med. Hyg. 34:355-360.
27
27. Liveris, D., A. Gazumyan, and I. Schwartz. 1995. Molecular typing of Borrelia burgdorferi sensu lato by PCR-restriction fragment length polymorphism analysis. J. Clin. Microbiol. 36:589-595.
28
28. Liveris, D., G. P. Wormser, J. Nowakowski, R. Nadelman, S. Bittker, D. Cooper, S. Varde, F. H. Moy, G. Forseter, C. S. Pavia, and I. Schwartz. 1996. Molecular typing of Borrelia burgdorferi from Lyme disease patients by PCR-restriction fragment length polymorphism analysis. J. Clin. Microbiol. 34:1306-1309.[Abstract]
29
29. Liveris, D., S. Varde, R. Iyer, S. Koenig, S. Bittker, D. Cooper, D. McKenna, J. Nowakowski, R. B. Nadelman, G. P. Wormser, and I. Schwartz. 1999. Genetic diversity of Borrelia burgdorferi in Lyme disease patients as determined by culture versus direct PCR with clinical specimen. J. Clin. Microbiol. 37:565-569.[Abstract/Free Full Text]
30
30. Marconi, R. T., D. Liveris, and I. Schwartz. 1995. Identification of novel insertion elements, restriction fragment length polymorphism patterns, and discontinuous 23S rRNA in Lyme disease spirochetes: phylogenetic analyses of rRNA genes and their intergenic spacers in Borrelia japonica sp. nov. and genomic group 21038 (Borrelia andersonii sp. nov.) isolates. J. Clin. Microbiol. 33:2427-2434.[Abstract]
31
31. Masuzawa, T., N. Takada, M. Kudeken, T. Fukui, Y. Yano, F. Ishiguro, Y. Kawamura, Y. Imai, and T. Ezaki. 2001. Borrelia sinica sp. nov., a Lyme disease-related Borrelia species isolated in China. Int. J. Syst. Evol. Microbiol. 51:1817-1824.[Abstract]
32
32. Mather, T. N., M. L. Wilson, S. I. Moore, J. M. Ribeiro, and A. Spielman. 1989. Comparing the relative potential of rodents as reservoirs of the Lyme disease spirochete (Borrelia burgdorferi). Am. J. Epidemiol. 130:143-150.[Abstract/Free Full Text]
33
33. Nakao, M., and K. Miyamoto. 1995. Mixed infection of different Borrelia species among Apodemus speciosus mice in Hokkaido, Japan. J. Clin. Microbiol. 33:490-492.[Abstract]
34
34. Piesman, J. 1993. Standard system for infecting ticks (Acari: Ixodidae) with the Lyme disease spirochete Borrelia burgdorferi. J. Med. Entomol. 30:199-203.[Medline]
35
35. Piesman, J., and C. M. Happ. 2001. The efficacy of co-feeding as a means of maintaining Borrelia burgdorferi: a North American model system. J. Vector Ecol. 26:216-220.[Medline]
36
36. Postic, D., M. V. Assous, P. A. D. Grimont, and G. Baranton. 1994. Diversity of Borrelia burgdorferi sensu lato evidenced by restriction fragment length polymorphism of rrf (5S)-rrl (23S) intergenic spacer amplicons. Int. J. Syst. Bacteriol. 44:743-752.[Abstract/Free Full Text]
37
37. Postic, D., E. Korenberg, N. Gorelova, Y. V. Kovalevski, E. Bellenger, and G. Baranton. 1997. Borrelia burgdorferi sensu lato in Russia and neighboring countries: high incidence of mixed isolates. Res. Microbiol. 148:691-702.[Medline]
38
38. Postic, D., N. M. Ras, R. S. Lane, M. Hendson, and G. Baranton. 1998. Expanded diversity among Californian Borrelia isolates and description of Borrelia bissetii sp. nov. (formerly Borrelia group DN127). J. Clin. Microbiol. 36:3497-3504.[Abstract/Free Full Text]
39
39. Randolph, S. E., L. Gern, and P. A. Nuttall. 1996. Co-feeding ticks: epidemiological significance for tick-borne pathogen transmission. Parasitol. Today 12:472-479.[CrossRef][Medline]
40
40. Richter, D., A. Spielman, N. Komar, and F.-R. Matuschka. 2000. Competence of American robins as reservoir hosts for Lyme disease spirochetes. Emerg. Infect. Dis. 6:133-138.[Medline]
41
41. Ryan, J. R., J. F. Levine, C. S. Apperson, L. Lubke, R. A. Wirtz, P. A. Spears, and P. E. Orndorff. 1998. An experimental chain of infection reveals that distinct Borrelia burgdorferi populations are selected in arthropod and mammalian hosts. Mol. Microbiol. 30:365-379.[CrossRef][Medline]
42
42. Seinost, G., D. E. Dykhuizen, R. J. Dattwyler, W. T. Golde, J. J. Dunn, I. N. Wang, G. P. Wormser, M. E. Schriefer, and B. J. Luft. 1999. Four clones of Borrelia burgdorferi sensu stricto cause invasive infection in humans. Infect. Immun. 67:3518-3524.[Abstract/Free Full Text]
43
43. van Dam, A. P., H. Kuiper, K. Vos, A. Widjojokusumo, B. M. de Jongh, L. Spanjaard, A. C. Ramselaar, M. D. Kramer, and J. Dankert. 1993. Different genospecies of Borrelia burgdorferi are associated with distinct clinical manifestations of Lyme borreliosis. Clin. Infect. Dis. 17:708-717.[Medline]
44
44. Wang, G., A. P. van Dam, A. Le Fleche, D. Postic, O. Péter, G. Baranton, R. de Boer, L. Spanjaard, and D. Dankert. 1997. Genetic and phenotypic analysis of Borrelia valaisiana sp. nov. (Borrelia genomic groups VS116 and M19). Int. J. Syst. Bacteriol. 47:926-932.[Abstract/Free Full Text]
45
45. Wang, G., D. Liveris, B. Brei, H. Wu, R. C. Falco, D. Fish, and I. Schwartz. 2003. Real-time PCR for simultaneous detection and quantification of Borrelia burgdorferi in field collected Ixodes scapularis ticks from the Northeastern United States. Appl. Environ. Microbiol. 69:4561-4565.[Abstract/Free Full Text]
46
46. Wang, G., C. Ojaimi, R. Iyer, V. Saksenberg, D. Liveris, S. A. McClain, G. P. Wormser, and I. Schwartz. 2001. Impact of genotypic variation of Borrelia burgdorferi sensu stricto on kinetics of dissemination and severity on disease in C3H/HeJ mice. Infect. Immun. 69:4303-4312.[Abstract/Free Full Text]
47
47. Wang, G., C. Ojaimi, H. Wu, V. Saksenberg, R. Iyer, D. Liveris, S. A. McClain, G. P. Wormser, and I. Schwartz. 2002. Disease severity in a murine model of Lyme borreliosis is associated with the genotype of the infecting Borrelia burgdorferi sensu stricto strain. J. Infect. Dis. 186:782-791.[CrossRef][Medline]
48
48. Wang, I.-N., D. E. Dykhauzen, W. Qiu, J. J. Dunn, E. M. Bosler, and B. J. Luft. 1999. Genetic diversity of ospC in a local population of Borrelia burgdorferi sensu stricto. Genetics 151:15-30.[Abstract/Free Full Text]
49
49. Wilske, B., V. Preaç-Mursic, U. B. Göbel, B. Graf, S. Jauris, E. Soutschek, E. Schwab, and G. Zumstein. 1993. An OspA serotyping system for Borrelia burgdorferi based on reactivity with monoclonal antibodies and OspA sequence analysis. J. Clin. Microbiol. 31:340-350.[Abstract/Free Full Text]
50
50. Wormser, G. P., D. Liveris, J. Nowakowski, R. N. Nadelman, L. F. Cavaliere, D. McKenna, D. Holmgren, and I. Schwartz. 1999. Association of specific subtypes of Borrelia burgdorferi with hematogenous dissemination in early Lyme disease. J. Infect. Dis. 180:720-725.[CrossRef][Medline]

---

Applied and Environmental Microbiology, November 2004, p. 6783-6788, Vol. 70, No. 11
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.11.6783-6788.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

This article has been cited by other articles:

• Gatewood, A. G., Liebman, K. A., Vourc'h, G., Bunikis, J., Hamer, S. A., Cortinas, R., Melton, F., Cislo, P., Kitron, U., Tsao, J., Barbour, A. G., Fish, D., Diuk-Wasser, M. A. (2009). Climate and Tick Seasonality Are Predictors of Borrelia burgdorferi Genotype Distribution. Appl. Environ. Microbiol. 75: 2476-2483 [Abstract] [Full Text]  
• Dykhuizen, D. E., Brisson, D., Sandigursky, S., Wormser, G. P., Nowakowski, J., Nadelman, R. B., Schwartz, I. (2008). The Propensity of Different Borrelia burgdorferi sensu stricto Genotypes to Cause Disseminated Infections in Humans. Am J Trop Med Hyg 78: 806-810 [Abstract] [Full Text]  
• Brisson, D., Dykhuizen, D. E, Ostfeld, R. S (2008). Conspicuous impacts of inconspicuous hosts on the Lyme disease epidemic. Proc R Soc B 275: 227-235 [Abstract] [Full Text]  
• Hanincova, K., Ogden, N. H., Diuk-Wasser, M., Pappas, C. J., Iyer, R., Fish, D., Schwartz, I., Kurtenbach, K. (2008). Fitness Variation of Borrelia burgdorferi Sensu Stricto Strains in Mice. Appl. Environ. Microbiol. 74: 153-157 [Abstract] [Full Text]  
• Swanson, S. J., Neitzel, D., Reed, K. D., Belongia, E. A. (2006). Coinfections Acquired from Ixodes Ticks. Clin. Microbiol. Rev. 19: 708-727 [Abstract] [Full Text]  
• Bockenstedt, L. K., Liu, N., Schwartz, I., Fish, D. (2006). MyD88 Deficiency Enhances Acquisition and Transmission of Borrelia burgdorferi by Ixodes scapularis Ticks. Infect. Immun. 74: 2154-2160 [Abstract] [Full Text]  

This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Derdáková, M. Articles by Fish, D. Search for Related Content PubMed PubMed Citation Articles by Derdáková, M. Articles by Fish, D. Agricola Articles by Derdáková, M. Articles by Fish, D.