[Emerging Infectious Diseases * Volume 4 * Number 1 * January - March 1998]

Dispatches

Prevalence of Tick-Borne Pathogens in _Ixodes scapularis_ in a Rural New
Jersey County

Shobha Varde,* John Beckley, and Ira Schwartz*
*New York Medical College, Valhalla, New York, USA; and Department of
Health, County of Hunterdon, Flemington, New Jersey, USA
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      To assess the potential risk for other tick-borne diseases, we
      collected 100 adult _Ixodes scapularis_ in Hunterdon County, a
      rapidly developing rural county in Lyme diseaseendemic western
      New Jersey. We tested the ticks by polymerase chain reaction
      for _Borrelia burgdorferi_, _Babesia microti_, and the
      rickettsial agent of human granulocytic ehrlichiosis (HGE).
      Fifty-five ticks were infected with at least one of the three
      pathogens: 43 with _B. burgdorferi_, five with _B. microti_,
      and 17 with the HGE agent. Ten ticks were coinfected with two
      of the pathogens. The results suggest that county residents are
      at considerable risk for infection by a tick-borne pathogen
      after an _I. scapularis_ bite.

The vector-borne diseases Lyme disease, human babesiosis, and human
granulocytic ehrlichiosis (HGE) are emerging in the Northeast and upper
Midwest regions of the United States (1,2). The etiologic agents for these
diseases (_Borrelia burgdorferi_, _Babesia microti_, and the HGE agent,
respectively) appear to share the same vertebrate reservoir host
(_Peromyscus leucopus_) and tick vector (_Ixodes scapularis_) (3-6).
Immunologic evidence of human coinfection with these pathogens has been
reported (7-10), and a culture-confirmed case of coinfection with _B.
burgdorferi_ and the HGE agent has recently been described (11). Thus, in
Lyme diseaseendemic areas, there may be a substantial risk for coinfection
with _B. burgdorferi_ and either _B. microti_ or the HGE-causing
rickettsia.

Hunterdon County (population 118,000) is a rural, but rapidly developing,
county in western New Jersey (Figure), with many homes in wooded settings.
In 1996, the county had the third highest case rate of Lyme disease (524
per 100,000) of all counties in the United States (Centers for Disease
Control and Prevention case reports). The county has also had many
suspected, but no serologically confirmed, cases of HGE.

 [fig]
 Figure. Map of New Jersey showing Hunterdon County. Black dots indicate
 tick collection sites.

The risk of acquiring a tick-borne pathogen in a specific geographic
location depends largely on tick density and the prevalence rate of the
agent in the tick population. Although cultivation of a microbial agent is
considered optimal, molecular detection is more practical since it permits
the rapid assay of large numbers of individual specimens. We report here
the prevalence rate of _B. burgdorferi_, _B. microti_, and the agent of HGE
in _I. scapularis_ in Hunterdon County by species-specific polymerase chain
reaction (PCR).

One hundred adult _I. scapularis_ were collected by drag cloth or from
personal clothing at 10 sites in the county during the fall of 1996
(Figure). Ticks were stored at room temperature in 70% ethanol until
analysis. DNA was isolated from individual ticks with the Isoquick DNA
extraction kit (ORCA Research, Bothell, WA) (12). The final DNA pellets
were resuspended in 50 l of sterile water, and a 10-l aliquot was used
for each PCR test. _B. burgdorferi_specific PCR used primers IS1 and IS2
(13). The HGE agent was detected by PCR using primers GER3 and GER4, as
described by Munderloh et al. (14), and _B. microti_ DNA was detected
according to methods used by Persing et al. (15). PCR products were
electrophoresed on 2% agarose gels stained with ethidium bromide. For _B.
burgdorferi_ and _B. microti_, hybridization with specific probes was
carried out after transfer to nylon membranes (13). In each PCR experiment
appropriate negative controls were used; these included master mix controls
lacking template and reactions containing _B. burgdorferi_ DNA, HGE agent
DNA, or _Escherichia coli_ DNA as additional controls for HGE agent, _B.
burgdorferi_, or _B. microti_ PCR, respectively. To prevent
cross-contamination, specimen preparation, PCR amplification, and post-PCR
analysis were performed in separate laboratories.

Results of PCR analysis for each of the pathogens on individual ticks are
presented in the Table. At least one of the three pathogens was present in
55% of the ticks. The highest prevalence rate was that for _B. burgdorferi_
(43/100), followed by the agent of HGE (17/100) and _B. microti_ (5/100).
Furthermore, these pathogens were widely distributed. At least one tick at
each site was infected with _B. burgdorferi_, nine of 10 sites had HGE
agentpositive ticks, and _B. microti_positive ticks were found at five
different collection sites. Ten ticks were coinfected with two of the three
agents; no ticks were infected with all three pathogens. The _B. microti_
PCR amplification products were gel purified and subjected to automated DNA
sequencing. The 238 bp products yielded sequences identical to that
reported for _B. microti_ 18S ribosomal DNA (15).

Table. Prevalence rate of _Borrelia burgdorferi_, human granulocytic
ehrlichiosis (HGE) agent, and _Babesia microti_ in 100 adult _Ixodes
scapularis_

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Pathogen                                     No. infected ticks

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_B. burgdorferi_                                       43

 HGE agent                                             17

_B. microti_                                            5

_B. burgdorferi_ and HGE agent                          6

_B. burgdorferi_ and _B. microti_                       2

HGE agent and _B. microti_                              2

_B. burgdorferi_, HGE agent, and _B. microti_           0

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The prevalence rates for the three pathogens reported here are consistent
with earlier studies at selected sites in the northeastern United States.
Telford et al. reported prevalence rates of 36%, 11%, and 9% for _B.
burgdorferi_, the HGE agent, and _B. microti_, respectively, in adult _I.
scapularis_ on Nantucket Island, Massachusetts; coprevalence of _B.
burgdorferi_ and the HGE agent was 4%, and no simultaneous infection with
_B. burgdorferi_ and _B. microti_ was observed (4). We have reported
prevalence rates of 52% and 53% for _B. burgdorferi_ and the HGE pathogen,
and coprevalence of 26% at a site in Westchester County, New York (5).
Given the simultaneous infection of _I. scapularis_ with these pathogens,
it is not surprising that numerous serosurveys of Lyme disease patients
indicate the coincident presence of antibodies to _B. burgdorferi_ and _B.
microti_ and human granulocytic ehrlichia (7-10). These findings do not
establish concurrent, active infection, nor do they demonstrate
simultaneous transmission by a single tick bite; however, they suggest that
persons living in disease-endemic areas are exposed to these agents,
presumably as a result of tick bites.

Human infection by any of the three tick-borne agents alone generally
results in similar acute manifestations, including fever, headache, and
myalgia (1,10,16). Reported cases of Lyme disease greatly outnumber those
of babesiosis or HGE. This, in conjunction with seroepidemiologic data,
suggests that many infections with _B. microti_ and granulocytic ehrlichia
are subclinical. Dual infection with _B. burgdorferi_ and _B. microti_
often results in more severe illness (10); whether this is also true for
concurrent infection by the agents of Lyme disease and HGE remains to be
established. Human coinfection by multiple tick-borne agents may account
for the variable nature of the clinical manifestations of Lyme disease.

This study demonstrates that the agents of Lyme disease, human babesiosis,
and HGE coexist in the tick population of a previously identified Lyme
diseaseendemic region of New Jersey. The study provides further evidence
that these three pathogens may be prevalent throughout the range of _I.
scapularis._ Infection with any of these three tick-borne pathogens should
be considered for residents or visitors of a disease-endemic area who have
flulike symptoms and a history of a tick bite.

Acknowledgments

We thank Gary Wormser and Dennis White for helpful comments, Chris Kolbert
and David Persing for providing advice and positive controls for _B.
microti_ PCR, and the Hunterdon County Tickborne Disease Research Group and
Hunterdon Medical Center for their support. This work was supported in part
by NIH grant AR41511.

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Emerging Infectious Diseases
National Center for Infectious Diseases
Centers for Disease Control and Prevention
Atlanta, GA

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