Ehrlichia phagocytophila genogroup rickettsiae in ixodid ticks from California collected in 1995 and 1996.

A total of 1,246 ixodid ticks collected in 1995 and 1996 from seven California counties were examined for the presence of Ehrlichia phagocytophila genogroup rickettsiae by using a nested PCR technique. Of 1,112 adult Ixodes pacificus Cooley and Kohls ticks tested, nine pools, each containing five ticks, were positive (minimum percentage of ticks harboring detectable ehrlichiae, 0.8%). Positive ticks were limited to four of the seven counties (Sonoma, El Dorado, Santa Cruz, and Orange). In Santa Cruz County, three positive pools were identified at the home of an individual with prior confirmed human granulocytic ehrlichiosis. In El Dorado County, positive ticks were found at sites where cases of granulocytic ehrlichiosis in a horse and a llama had recently occurred. Among 47 nymphal I. pacificus ticks collected in Sonoma County, one positive pool was identified. Fifty-seven adult Dermacentor occidentalis Marx and 30 adult D. variabilis Say ticks, collected chiefly in southern California, were negative. These data, although preliminary, suggest that the prevalence of E. phagocytophila genogroup rickettsiae in ixodid ticks of California may be lower than in cognate vector populations (i.e., I. scapularis Say = I. dammini Spielman, Clifford, Piesman, and Corwin) in the eastern and midwestern United States.     

Prevalence of granulocytic Ehrlichia infection among white-tailed deer in Wisconsin.

Human granulocytic ehrlichiosis (HGE) is caused by an agent that is nearly indistinguishable from the veterinary pathogens Ehrlichia equi and Ehrlichia phagocytophila. The deer tick, Ixodes scapularis, is a vector of the HGE agent, and the white-tailed deer is the primary host for adult Ixodes ticks. We assessed the distribution of granulocytic Ehrlichia infection among deer living within (Wisconsin) and outside (western and southern Iowa) the geographic range of L. scapularis. Whole-blood samples were tested for HGE 16S ribosomal DNA (rDNA) by PCR, and E. equi antibody was detected by indirect immunofluorescence assay (IFA). Antibody titers of > or = 1:64 were defined as positive, and all positive samples were retested with a second lot of substrate antigen. E. equi antibody was present in 14 (8%) of 187 Wisconsin deer and 0 of 60 Iowa specimens (rate ratio undefined; P = 0.025). An additional 30 serum samples from Wisconsin deer were excluded because IFA results were discrepant between substrate lots. The reciprocal antibody titers ranged from 64 to 512 (geometric mean, 141) for positive samples. PCR results were positive for 27 (15%) of 181 Wisconsin deer. The prevalence of infection in northwestern Wisconsin deer was not significantly different from that in central Wisconsin deer, as determined by IFA and PCR. In two samples that were sequenced, the 16S rDNA was nearly identical to that of the granulocytic Ehrlichia species but distinct from that of Anaplasma marginale. The DNA sequences of the samples differed from the published sequences for E. equi, E. phagocytophila, and the HGE agent by 1 or 2 nucleotides (> or = 99.1% homology) at phylogenetically informative sites. Granulocytic Ehrlichia organisms in deer are widely distributed within the geographic range of L. scapularis in Wisconsin. Deer may serve as useful sentinels for areas where HGE transmission to humans may occur.     

Rickettsiae and Borrelia burgdorferi in ixodid ticks.

Nymphs and adults of hard-bodied ticks were collected in Connecticut and tested by direct and indirect immunofluorescence staining methods for rickettsiae and Borrelia burgdorferi. Of the 609 Ixodes dammini ticks examined, 59 (9.7%) harbored rickettsialike microorganisms in hemocytes (blood cells). These bacteria reacted with fluorescein-conjugated antiserum to Ehrlichia canis, the etiologic agent of with fluorescein-conjugated antiserum to Ehrlichia canis, the etiologic agent of canine ehrlichiosis. Prevalence of infection ranged from 6.8 to 12.7% for males and females, respectively. Although the specific identities of the hemocytic rickettsialike organisms are unknown, they share antigens with ehrlichiae. Electron microscopy revealed rickettsiae in ovarian tissues of I. dammini that also had infected hemocytes. Rickettsialike organisms were also observed in the hemocytes of 5 (6.9%) of 73 Dermacentor variabilis ticks. In analyses for B. burgdorferi, 146 (23.7%) of 617 I. dammini ticks harbored these spirochetes in midguts. Hemocytic rickettsialike microorganisms coexisted with B. burgdorferi in 36 (6.7%) of the 537 nymphs and adults of I. dammini examined. I. dammini, with its broad host range, has the potential to acquire multiple microorganisms.     

Acquisition of coinfection and simultaneous transmission of Borrelia burgdorferi and Ehrlichia phagocytophila by Ixodes scapularis ticks

The agents of Lyme disease (Borrelia burgdorferi) and human granulocytic ehrlichiosis (Ehrlichia phagocytophila) are both transmitted by the tick Ixodes scapularis. In nature, ticks are often infected with both agents simultaneously. We studied whether previous infection with either Borrelia or Ehrlichia in ticks would affect acquisition and transmission of a second pathogen. Ehrlichia-infected I. scapularis nymphs were fed upon Borrelia-infected mice, and Borrelia-infected I. scapularis nymphs were fed upon Ehrlichia-infected mice. The efficiency with which previously infected nymphal ticks acquired a second pathogen from infected hosts was compared to that of uninfected ticks. An average of 51% +/- 15% of ticks acquired Ehrlichia from infected mice regardless of their prior infection status with Borrelia. An average of 85% +/- 10% of ticks acquired Borrelia from infected mice regardless of their prior infection status with Ehrlichia. Also, we assessed the efficiency with which individual nymphs could transmit either agent alone, or both agents simultaneously, to individual susceptible hosts. An average of 76% +/- 9% of Borrelia-infected ticks and 84% +/- 10% of Ehrlichia-infected ticks transmitted these agents to mice regardless of the presence of the other pathogen. There was no evidence of interaction between the agents of Lyme disease and human granulocytic ehrlichiosis in I. scapularis ticks. The presence of either agent in the ticks did not affect acquisition of the other agent from an infected host. Transmission of the agents of Lyme disease and human granulocytic ehrlichiosis by individual ticks was equally efficient and independent. Dually infected ticks transmitted each pathogen to susceptible hosts as efficiently as ticks infected with only one pathogen.     

Evidence of the human granulocytic ehrlichiosis agent in Ixodes ricinus ticks in Switzerland

A total of 1,667 Ixodes ricinus ticks were collected from five regions in Switzerland where there have been sporadic occurrences of granulocytic ehrlichiosis in dogs and horses. The ticks were examined for rickettsiae of the Ehrlichia phagocytophila group via nested PCR. Twenty-one ticks (1.3%) were positive; 3 (0.5%) were nymphs, 6 (1.3%) were adult males, and 12 (1.9%) were adult females. The number of positive ticks varied with the stage of development and with the geographical origin. Nucleotide sequencing of the isolated PCR products identified these products as part of the 16S rRNA gene of Ehrlichia. In addition, these products had 100% homology with the agent of human granulocytic ehrlichiosis. The occurrence of this agent in I. ricinus in Switzerland presents a potential danger of transmission of granulocytic ehrlichiosis to dogs, horses, and humans.     

Isolation of the equine granulocytic ehrlichiosis agent, Ehrlichia equi, in tick cell culture.

The equine granulocytic ehrlichiosis agent, Ehrlichia equi, is closely related or identical to the human granulocytic ehrlichiosis (HGE) agent. Both are suspected of being transmitted by ticks. We have successfully isolated E. equi in a cell line, IDE8, derived from a putative vector, the tick Ixodes scapularis. Peripheral blood leukocytes from an experimentally infected horse were inoculated onto IDE8 monolayers. Cultures were incubated in a candle jar at 34 degrees C in tick cell culture medium with NaHCO3 and an organic buffer [3-(N-morpholino)-propanesulfonic acid] (MOPS). Within 2 weeks, infected cells were detected in Giemsa-stained culture samples, and the organisms subsequently spread to uninfected cells in the cultures. E. equi was passaged serially by transferring a portion of an infected culture to new cell layers every 2 to 3 weeks. The identity of the organisms was confirmed by PCR using oligonucleotide primers specific for E. equi and the HGE agent and by immunocytology. Homologous equine antibodies and human anti-HGE convalescent serum recognized E. equi grown in tick cell culture. Electron microscopy revealed electron-lucent and -dense ehrlichia-like forms developing within host cell endosomes. E. equi passaged twice in tick cell culture retained infectivity and pathogenicity for the equine host, as demonstrated by intravenous inoculation of a suspension of infected tick cells and subsequent reisolation from peripheral blood, in fulfillment of Koch's postulates. The horse developed severe clinical signs, i.e., fever, inappetence, thrombocytopenia, icterus, and limb edema, typical of granulocytic equine ehrlichiosis, within 1 week.     

Quantitative real-time PCR for detection of members of the Ehrlichia phagocytophila genogroup in host animals and Ixodes ricinus ticks

A TaqMan PCR was established for identification and quantitation of members of the Ehrlichia phagocytophila group in experimentally infected cows and in Ixodes ricinus ticks. The TaqMan PCR identified a 106-bp section of the 16S rRNA gene by use of a specific fluorogenic probe and two primers. This technique was specific for members of the E. phagocytophila group, which include E. phagocytophila, Ehrlichia equi, and the agent of human granulocytic ehrlichiosis. The TaqMan system identified 10 copies of a cloned section of the 16S rRNA gene of E. phagocytophila. The sensitivity and specificity of the TaqMan PCR were similar to those of conventional nested PCR. The numbers of ehrlichiae in leukocytes of the two cows experimentally infected with E. phagocytophila were measured daily by TaqMan PCR and had a course similar to that of the percentages of infected leukocytes determined daily by light microscopy. The prevalence of infected free-living ticks, which were collected from areas where bovine ehrlichiosis is endemic and from regions with sporadic occurrences of granulocytic ehrlichiosis in dogs and horses, was identical as determined by nested PCR and TaqMan PCR.     

PCR amplification and comparison of nucleotide sequences from the groESL heat shock operon of Ehrlichia species.

Degenerate PCR primers derived from conserved regions of the eubacterial groESL heat shock operon were used to amplify groESL sequences of Ehrlichia equi, Ehrlichia phagocytophila, the agent of human granulocytic ehrlichiosis (HGE), Ehrlichia canis, Bartonella henselae, and Rickettsia rickettsii. The groESL nucleotide sequences were less conserved than the previously determined 16S rRNA gene sequences of these bacteria. A phylogenetic tree derived from deduced GroEL amino acid sequences was similar to trees based on 16S rRNA gene sequences. Nucleotide sequences obtained from clinical samples containing E. equi, E. phagocytophila, or the HGE agent were very similar (99.9 to 99.0% identity), and the deduced amino acid sequences were identical. Some divergence was evident between nucleotide sequences amplified from samples originating from the United States (E. equi and the HGE agent) and sequences from the European species, E. phagocytophila. A single pair of PCR primers derived from these sequences was used to detect E. chaffeensis and HGE agent DNA in blood samples from human patients with ehrlichiosis.     

Detection of Rickettsia rickettsii and Bartonella henselae in Rhipicephalus sanguineus ticks from California

Sixty-two questing adult Rhipicephalus sanguineus (Latreille) ticks were collected by direct removal from blades of turfgrass and adjacent concrete walkways at a suburban home in Riverside County, CA, and tested for the presence of Rickettsia, Bartonella, and Ehrlichia DNA. Polymerase chain reaction (PCR) was used to amplify fragments of the 17-kDa antigen gene and the rOmpA gene of the spotted fever group rickettsiae. One male tick contained R. rickettsii DNA; its genotype differed from R. rickettsii isolates found in Montana and Arizona that cause Rocky Mountain spotted fever and from Hlp#2 and 364D serotypes. One male tick and one female tick contained B. henselae DNA. No Ehrlichia platys or Ehrlichia canis DNAs were detected using nested PCR for their 16S rRNA genes. These findings extend the area where Rickettsia rickettsii may be vectored by Rh. sanguineus. Rh. sanguineus also may be infected with Bartonella henselae, a human pathogen that is typically associated with fleas and causes cat scratch disease.     

Serologic and molecular detection of granulocytic ehrlichiosis in Rhode Island.

A new indirect fluorescent-antibody (IFA) assay with antigen produced in vitro in the human promyelocytic leukemia cell line HL60 was used to identify the first recognized case of human granulocytic ehrlichiosis in Rhode Island. This IFA assay was used to detect granulocytic ehrlichiae in white-footed mice and in a dog inhabiting the area surrounding the patient's residence. Host-seeking Ixodes scapularis ticks found in the same habitat also were infected. I. scapularis ticks collected from other locations were fed on dogs and New Zealand White rabbits to assess the competency of these species as hosts of granulocytotropic Ehrlichia. Tick-induced infections of dogs were confirmed by serologic testing, tissue culture isolation, and PCR amplification, whereas several rabbits seroconverted but were PCR and culture negative. PCR amplification of the 16S rRNA gene and DNA sequencing of the PCR products or culture isolation was used to confirm granulocytic Ehrlichia infections in humans, dogs, white-footed mice, and ticks.     

Natural infection of small mammal species in Minnesota with the agent of human granulocytic ehrlichiosis.

The natural reservoirs for the agent of human granulocytic ehrlichiosis (HGE) are suspected to be the small mammals that host immature stages of Ixodes scapularis ticks. To determine if such small mammals are naturally infected, we collected blood and serum samples from small mammal species in rural and suburban areas of Minneapolis and St. Paul, Minn. Samples were collected from white-footed mice (Peromyscus leucopus), eastern chipmunks (Tamias striatus), southern red-backed voles (Clethrionomys gapperi), and insectivorous shrews (Blarina brevicauda and Sorex cinereus). Blood samples were tested by PCR for active infection with the HGE agent, and sera from P. leucopus mice were tested for serologic evidence of infection by indirect immunofluorescence. PCR analyses revealed the presence of HGE agent DNA in 20 of the 190 samples (10.5%) tested. Of the 119 P. leucopus mouse serum samples that were analyzed, 12 (10.1%) contained Ehrlichia equi antibodies. In 3 of 119 (2.5%) P. leucopus mice from which both blood and serum were collected. HGE agent DNA and antibodies against E. equi were present. Animals with evidence of infection with the HGE agent are widely distributed around the Minneapolis-St. Paul area in regions with known I. scapularis tick activity. Small mammals that are frequent hosts for larval I. scapularis ticks and that are found in areas where HGE occurs are likely to be a major reservoir from which infected ticks that bite humans are derived.     

Human disease in Europe caused by a granulocytic Ehrlichia species.

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.     

Rickettsia rickettsii (Rickettsiales: Rickettsiaceae) in Amblyomma americanum (Acari: Ixodidae) from Kansas

The role of lone star ticks as vectors for Rocky Mountain spotted fever (RMSF) remains poorly described. We compared the entomological inoculation rates (EIRs) for Rickettsia spp. for representative sites in Missouri and Kansas, states that frequently report RMSF each year. Host-seeking ticks were collected during 2006 and pooled tick homogenates analyzed by polymerase chain reaction to detect probable R. rickettsii, with confirmation for multiple gene targets performed on individual ticks from pools that screened positive. Of 870 adult and nymphal lone star ticks, Amblyomma americanum (L.), 0.46% contained DNA of Rickettsia rickettsii. Interestingly, two of these positive ticks were concurrently infected by R. amblyommii. More than 90% of lone star tick pools contained R. amblyommii DNA. Of 169 dog ticks that were analyzed, none were infected by R. rickettsii. The entomological inoculation rate for spotted fever group (SFG) rickettsiae within lone star ticks was an order of magnitude greater than that for dog ticks. We conclude that lone star ticks may be epidemiologically significant vectors of Rocky Mountain spotted fever and of spotted fever group rickettsiae.     

Prevalence of Borrelia burgdorferi and Granulocytic and Monocytic Ehrlichiae in Ixodes ricinus Ticks from Southern Germany

A total of 287 adult Ixodes ricinus ticks, collected in two regions of southern Germany (Frankonia and Baden-Württemberg) where Borrelia burgdorferi infections are known to be endemic, were examined for the presence of 16S ribosomal DNA specific for the Ehrlichia phagocytophila genogroup, E. chaffeensis, E. canis, and B. burgdorferi by nested PCR. Totals of 2.2% (6 of 275) and 21.8% (65 of 275) of the ticks were positive for the E. phagocytophila genogroup and B. burgdorferi, respectively. Two ticks (0.7%) were coinfected with both bacteria. Of 12 engorged I. ricinus ticks collected from two deer, 8 (67%) were positive for the E. phagocytophila genogroup and one (8%) was positive for B. burgdorferi. There was no evidence of infection with E. canis or E. chaffeensis in the investigated tick population. The nucleotide sequences of the 546-bp Ehrlichia PCR products differed at one or two positions from the original sequence of the human granulocytic ehrlichiosis (HGE) agent (S.-M. Chen, J. S. Dumler, J. S. Bakken, and D. H. Walker, J. Clin. Microbiol. 32:589-595, 1994). Three groups of sequence variants were detected; two of these were known to occur in other areas in Europe or the United States, whereas one has not been reported before. Thus, in the German I. ricinus tick population closely related granulocytic ehrlichiae are prevalent, which might represent variants of E. phagocytophila or the HGE agent.     

Reduced abundance of Ixodes scapularis (Acari: Ixodidae) and the tick parasitoid Ixodiphagus hookeri (Hymenoptera: Encyrtidae) with reduction of white-tailed deer

The principal vector for the pathogens of Lyme disease, human granulocytic ehrlichiosis, and human babesiosis is the tick Ixodes scapularis Say. A chalcid wasp, Ixodiphagus hookeri, in the family Encyrtidae parasitizes populations of the tick on several islands or other geographically isolated sites in New England with high densities of these ticks and white-tailed deer (Odocoileus virginianus), the principal host for adult I. scapularis. Deer densities were reduced at a forested tract in Bridgeport and the Bluff Point Coastal Reserve in Groton, Connecticut, from levels exceeding 90 animals per km2 in 1992 (Bridgeport) and 1994 (Bluff Point) to 17 and 10 animals per km2, respectively, by fall 2001. Tick densities declined with sustained reductions in the population of white-tailed deer. Similarly, prevalence of tick parasitism by Ixodes hookeri declined at both sites from 30 to 25% to <1.0% and was significantly correlated with previous year's deer density at both sites (r(s) = 0.933 and r(s) = 0.867, P < or = 0.0001) and with nymphal tick densities at Bridgeport (r(s) = 0.867, P < or = 0.0001), but was not as well correlated with tick densities in Groton. The virtual disappearance of I. hookeri in this study corresponds with a lack of I. hookeri in mainland I. scapularis at comparable deer and tick densities, suggesting that there is a threshold deer density of approximatley 10-20/km2, with corresponding tick densities necessary for I. hookeri to successfully parasitize I. scapularis.     

Spotted fever group rickettsiae or Borrelia burgdorferi in Ixodes cookei (Ixodidae) in Connecticut.

Immatures and females of Ixodes cookei, a hard-bodied tick, were collected from woodchucks and other mammals in the northeastern United States and examined for spotted fever group rickettsiae and Borrelia burgdorferi. Of the 93 nymphs analyzed by a hemolymph test, 4 (4.3%) harbored rickettsiae. Six (15%) of 40 females were also infected. All infected ticks were collected from woodchucks in Connecticut. Indirect fluorescent antibody staining of midgut tissues from 128 nymphs revealed B. burgdorferi in two (1.6%) ticks, whereas larval and female ticks were negative. Further consideration should be given to I. cookei as a possible vector of spotted fever group rickettsiae or spirochetes that cause Lyme borreliosis.     

Geographic, clinical, serologic, and molecular evidence of granulocytic ehrlichiosis, a likely zoonotic disease, in Minnesota and Wisconsin dogs.

Seventeen Minnesota and Wisconsin dogs with granulocytic ehrlichosis were studied. The diagnoses were made by finding ehrlichia morulae in peripheral blood neutrophils. Eight dogs were studied retrospectively, and nine dogs were studied prospectively. The medical records of all dogs were reviewed. Eighty-eight percent of the dogs were purebred and 76% were spayed females. The median age was 8 years. Sixty-five percent of the cases were diagnosed in October and November. Fever and lethargy were the most common clinical signs. The most frequent laboratory findings were lymphopenia, thrombocytopenia, elevated activities of serum alkaline phosphatase and amylase, and hypoalbuminemia. No dogs seroreacted to Ehrlichia canis or Ehrlichia chaffeensis antigens, which are cross-reactive. Seventy-five percent of the dogs tested during the acute phase of disease and 100% of the dogs tested during convalescence were seropositive for E. equi antigens. Granulocytic ehrlichial 16S rRNA gene DNAs from six dogs were amplified by PCR. Sequence analysis of a 919-bp sequence of the ehrlichial 16S rRNA gene amplified by PCR from the blood of two dogs revealed the agent to be identical to the agent of human granulocytic ehrlichiosis in Minnesota and Wisconsin and to be very similar to E. equi and Ehrlichia phagocytophila and less similar to E. canis, Ehrlichia ewingii, and E. chaffeensis. The geographic, clinical, serologic, and molecular evidence indicates that granulocytic ehrlichiosis in Minnesota and Wisconsin dogs is not caused by E. ewingii, but suggests that it is a zoonotic disease caused by an agent closely related to E. equi and that dogs likely contribute to the enzootic cycle and human infection.     

Serological responses to Ehrlichia equi, Ehrlichia chaffeensis, and Borrelia burgdorferi in patients from New York State.

Serological testing at the New York State Department of Health for human granulocytic ehrlichiosis in the residents of Westchester County, N.Y., was performed with specimens from 176 patients by the indirect fluorescent-antibody (IFA) technique with Ehrlichia equi MRK-infected neutrophils. To understand whether human monocytotropic ehrlichiosis also occurs in this northeastern geographic region, specimens were also tested for antibodies to Ehrlichia chaffeensis Arkansas. Screening tests and immunoblots for Lyme disease (Borrelia burgdorferi infection) were also performed. Thirty-two patients had antibodies only to E. equi and 21 patients had antibodies to both E. equi and E. chaffeensis, whereas 12 patients had only E. chaffeensis antibodies by the IFA technique. The remaining patients did not have antibodies to either ehrlichia. Eighteen serum samples from 13 of these patients were coded and sent to the Ehrlichia Research Laboratory (Baltimore, Md.) for repeat analysis by the IFA test and for E. equi and E. chaffeensis immunoblots. Immunoblot analysis for E. equi in samples with positive IFA test results confirmed the results for eight of the nine specimens. Immunoblot analyses for E. chaffeensis were negative for all 18 serum samples. Borrelia-reactive antibodies were found in sera both from patients with granulocytic ehrlichiosis and from patients with monocytotropic ehrlichiosis from New York State. Our results suggest that E. equi antigen is an appropriate substrate for identifying human granulocytic ehrlichiosis. E. chaffeensis antigen lacks appropriate sensitivity to serve as a surrogate substrate for the detection of human granulocytic ehrlichiosis and should be used solely for the diagnosis of human monocytotropic ehrlichiosis. Heat shock proteins may, in some cases, cause cross-reactivity between B. burgdorferi and ehrlichiae.     

Reassessment of a midwestern Lyme disease focus for Borrelia burgdorferi and the human granulocytic ehrlichiosis agent

Previous studies from the late 1980s defined the risk of human Lyme disease by determining the prevalence of Borrelia burgdorferi infection in Ixodes scapularis ticks and Peromyscus sp. mice captured from areas around La Crosse, Wis. High percentages of B. burgdorferi-infected I. scapularis ticks and P. leucopus mice were common in areas located north of Interstate 90 but were not detected in areas south of this major east-west thoroughfare. In this study, we reevaluated the extent of B. burgdorferi infection. High percentages of mice captured from sites north of the interstate were still infected with B. burgdorferi. In addition, B. burgdorferi was recovered from 12 (67%) of 18 mice captured from a site well south of the highway. However, none of 104 mice or 713 I. scapularis ticks captured from the study sites were infected with Ehrlichia spp. The results confirmed the continued high risk for humans to contract infection with B. burgdorferi and the significant southward expansion of the area in which Lyme disease is endemic. In contrast, the risk of acquiring human granulocytic ehrlichiosis remains minimal despite the abundance of appropriate vector ticks and reservoir rodents.     

New Ehrlichia species closely related to Ehrlichia chaffeensis isolated from Ixodes ovatus ticks in Japan

Seven Ehrlichia strains (six HF strains and one Anan strain) that were obtained from laboratory mice by intraperitoneally inoculating homogenates of adult Ixodes ovatus collected in Japan were characterized. 16S rRNA sequences of all six HF strains were identical, and the sequences were 99.7, 98.2, and 97.7% identical to those of Anan strain, Ehrlichia chaffeensis (human monocytic ehrlichiosis agent), and E. muris, respectively. Partial GroEL amino acid sequencing also revealed that the six HF strains had identical sequences, which were 99.0, 98.5, and 97.3% identical to those of E. chaffeensis, the Anan strain, and E. canis, respectively. All HF strains were lethal to mice at higher dosages and intraperitoneal inoculation, whereas the Anan or E. muris strain induced only mild clinical signs. Light and electron microscopy of moribund mice inoculated with one of the HF strains revealed severe liver necrosis and the presence of numerous ehrlichial inclusions (morulae) in various organs. The study revealed that members of E. canis genogroup are naturally present in Ixodes ticks. HF strains that can cause severe illness in immunocompetent laboratory mice would be valuable in studying the pathogenesis and the roles of both cellular and humoral immune responses in ehrlichiosis caused by E. canis genogroup.