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.

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Detection of four Borrelia burgdorferi genospecies and first report of human granulocytic ehrlichiosis agent in Ixodes ricinus ticks collected in central Italy

Epidemiology and Infection ◽

10.1017/s0950268802007057 ◽

2002 ◽

Vol 129 (1) ◽

pp. 93-97 ◽

Cited By ~ 5

Author(s):

I. SANTINO ◽

M. DEL PIANO ◽

R. SESSA ◽

G. FAVIA ◽

A. IORI

Keyword(s):

Borrelia Burgdorferi ◽

Ixodes Ricinus ◽

Protected Area ◽

Central Italy ◽

First Report ◽

Granulocytic Ehrlichiosis ◽

Ehrlichia Phagocytophila ◽

Human Granulocytic Ehrlichiosis ◽

Ixodes Ricinus Ticks ◽

Male Tick

The presence of Borrelia burgdorferi s.l. and of Ehrlichia phagocytophila group was sought by PCR in Ixodes ricinus collected in a protected area of central Italy. Nymphs (n = 1475, gathered in 295 pools of 5 nymphs each) and adult ticks (n = 28) were examined. B. burgdorferi s.l. was detected in 13.8% of the nymph pools; of these, 63.4% were infected by B. valaisiana, 26.8% by B. afzelii, 7.3% by B. garinii, and 2.5% by B. burgdorferi s.s. Only a single adult male tick proved to host B. afzelii. The agent of human granulocytic ehrlichiosis (HGE) was detected in 2.7% of the nymph pools. Two HGE agent-positive nymph pools were also found to be positive for B. garinii and for B. afzelii, respectively. This is the first report from central Italy of the finding of the HGE agent in ticks.

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Identity of Ehrlichial DNA Sequences Derived from Ixodes ricinus Ticks with Those Obtained from Patients with Human Granulocytic Ehrlichiosis in Slovenia

Journal of Clinical Microbiology ◽

10.1128/jcm.37.1.209-210.1999 ◽

1999 ◽

Vol 37 (1) ◽

pp. 209-210 ◽

Cited By ~ 45

Author(s):

M. Petrovec ◽

J. W. Sumner ◽

W. L. Nicholson ◽

J. E. Childs ◽

F. Strle ◽

...

Keyword(s):

Heat Shock ◽

16S Rrna ◽

Ixodes Ricinus ◽

Dna Sequences ◽

Rrna Gene ◽

Granulocytic Ehrlichiosis ◽

Human Granulocytic Ehrlichiosis ◽

Ixodes Ricinus Ticks ◽

Pcr Assays ◽

The 16S Rrna Gene

Adult Ixodes ricinus (Acari: Ixodidae) ticks collected near Ljubljana, Slovenia, were tested for the agent of human granulocytic ehrlichiosis (HGE) by using PCR assays based on the 16S rRNA gene. Three (3.2%) of 93 ticks were found to contain granulocytic ehrlichiae. Nucleotide sequences of portions of the bacterialgroESL heat shock operon amplified from these ticks were identical or nearly (99.8%) identical to those previously determined for human patients with HGE from Slovenia, providing additional evidence that the ticks were infected with the HGE agent. This study identified I. ricinus as the likely vector for these ehrlichial pathogens of humans in this part of Europe.

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Quantitative Real-Time PCR for Detection of Members of the Ehrlichia phagocytophila Genogroup in Host Animals and Ixodes ricinus Ticks

Journal of Clinical Microbiology ◽

10.1128/jcm.37.5.1329-1331.1999 ◽

1999 ◽

Vol 37 (5) ◽

pp. 1329-1331 ◽

Cited By ~ 51

Author(s):

Nicola Pusterla ◽

Jon B. Huder ◽

Christian M. Leutenegger ◽

Ueli Braun ◽

John E. Madigan ◽

...

Keyword(s):

16S Rrna ◽

16S Rrna Gene ◽

Ixodes Ricinus ◽

Nested Pcr ◽

Rrna Gene ◽

Granulocytic Ehrlichiosis ◽

Ehrlichia Phagocytophila ◽

Ixodes Ricinus Ticks ◽

Taqman Pcr ◽

The 16S Rrna Gene

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 ofE. 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.

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Identification of a Granulocytic Ehrlichia Strain Isolated from a Horse in Switzerland and Comparison with Other Rickettsiae of the Ehrlichia phagocytophilaGenogroup

Journal of Clinical Microbiology ◽

10.1128/jcm.36.7.2035-2037.1998 ◽

1998 ◽

Vol 36 (7) ◽

pp. 2035-2037 ◽

Cited By ~ 21

Author(s):

Nicola Pusterla ◽

Jon B. Huder ◽

Karsten Feige ◽

Hans Lutz

Keyword(s):

Case Report ◽

Nucleotide Sequence ◽

16S Rrna ◽

Clinical Signs ◽

Buffy Coat ◽

Rrna Gene ◽

Granulocytic Ehrlichiosis ◽

Ehrlichia Phagocytophila ◽

Human Granulocytic Ehrlichiosis ◽

Limb Edema

This case report describes a 12-year-old Arabian mare with granulocytic ehrlichiosis. Clinical signs included fever, apathy, anorexia, icterus, limb edema, and reluctance to move. Examination of buffy coat smears revealed Ehrlichia organisms in neutrophils and eosinophils. A band of 1,428 bp was amplified from DNA of leukocytes via nested PCR and was identified as part of theEhrlichia 16S rRNA gene. It differed from the gene sequences of Ehrlichia phagocytophila and E. equi at two and three positions, respectively. Interestingly, the nucleotide sequence of the 16S rRNA was 100% identical to that of the agent of human granulocytic ehrlichiosis.

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Improved Sensitivity of PCR for Diagnosis of Human Granulocytic Ehrlichiosis Using epank1 Genes of Ehrlichia phagocytophila -Group Ehrlichiae

Journal of Clinical Microbiology ◽

10.1128/jcm.38.1.354-356.2000 ◽

2000 ◽

Vol 38 (1) ◽

pp. 354-356

Author(s):

Jennifer J. Walls ◽

Patrizio Caturegli ◽

Johan S. Bakken ◽

Kristin M. Asanovich ◽

J. Stephen Dumler

Keyword(s):

16S Rrna ◽

16S Rrna Gene ◽

Ankyrin Repeat ◽

Rrna Gene ◽

Gene Target ◽

Granulocytic Ehrlichiosis ◽

Ehrlichia Phagocytophila ◽

Human Granulocytic Ehrlichiosis ◽

Group A ◽

Group B

ABSTRACT The agent of human granulocytic ehrlichiosis (HGE), Ehrlichia phagocytophila , and Ehrlichia equi probably comprise variants of a single Ehrlichia species now called the Ehrlichia phagocytophila genogroup. These variants share a unique 153-kDa protein antigen with ankyrin repeat motifs encoded by the epank1 gene. The epank1 gene was investigated as an improved target for PCR diagnosis of HGE compared with the currently used 16S rRNA gene target. Primers for epank1 flanking a region that spans part of the 5′ ankyrin repeat coding region and part of the unique 3′ region were synthesized. Blood samples from 31 patients with suspected HGE who were previously tested by 16S rRNA gene (16S) PCR and indirect immunofluorescent antibody test (IFA) were retrospectively tested with the epank1 primers. Eleven patients were 16S PCR positive and had a seroconversion detected by IFA (group A), 10 patients were 16S PCR negative but had a seroconversion detected by IFA (group B), and 10 patients were 16S PCR negative and seronegative (group C). Ten of the 11 group A patients were epank1 PCR positive, all 10 of the group B patients were epank1 PCR positive, and all of the PCR-negative and seronegative patients (group C) were epank1 PCR negative. The epank1 primers are more sensitive than the previously used 16S rRNA gene primers and therefore may be more useful in diagnostic testing for HGE.

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Prevalence of Borrelia Burgdorferi Sensu Lato Genomospecies and of the Human Granulocytic Ehrlichiosis (HGE) Agent in Ixodes Ricinus Ticks Collected in the Area of Monti Lepini, Italy

International Journal of Immunopathology and Pharmacology ◽

10.1177/039463200301600203 ◽

2003 ◽

Vol 16 (2) ◽

pp. 105-108 ◽

Cited By ~ 10

Author(s):

I. Santino ◽

A. Iori ◽

M. Nicoletti ◽

S. Valletta ◽

C. Cimmino ◽

...

Keyword(s):

Borrelia Burgdorferi ◽

Pcr Amplification ◽

Sensu Stricto ◽

16S Rrna Genes ◽

Rrna Genes ◽

Borrelia Burgdorferi Sensu Lato ◽

Granulocytic Ehrlichiosis ◽

Ehrlichia Phagocytophila ◽

Human Granulocytic Ehrlichiosis ◽

Ixodes Ricinus Ticks

Ticks are obligate hematophagous arthropods that are parasites in every class of vertebrates in most regions of the world. They are also considered to be important vectors for the transmission of human infectious diseases. In the present study we used polymer chain reaction (PCR) amplification analysis to determine the prevalence of Borrelia burgdorferi and Ehrlichia phagocytophila, the agents of, respectively, Lyme borreliosis and human granulocytic ehrlichiosis, among ticks inhabiting the area of Monti Lepini, a wild area located in the Latium Region of Italy. A total of 141 I. ricinus ticks (125 nymphs and 16 adults) were collected in the studied area. Total DNAs were extracted from I. ricinus nymphs (pooled in groups of five) and from individual adults. The DNA samples were examined for the presence of B. burgdorferi sensu lato and E. phagocytophila by PCR using two specific pairs of oligonucleotides that specifically amplify distinct DNA regions of the 16S rRNA genes of the two species. The prevalence of vectors infected with B. burgdorferi s. 1. was 16% in pooled nymphs samples, and 12.5% in adult ticks, while E. phagocytophila was found only in pooled nymphs samples (8%). Three genomospecies were identified, namely Borrelia afzelii, Borrelia garinii, and Borrelia valaisiana, in samples found positive for B. burgdorferi s. 1. No sample was found positive for Borrelia burgdorferi sensu stricto.

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

Journal of Clinical Microbiology ◽

10.1128/jcm.37.11.3448-3451.1999 ◽

1999 ◽

Vol 37 (11) ◽

pp. 3448-3451 ◽

Cited By ~ 52

Author(s):

Birgit U. Baumgarten ◽

Martin Röllinghoff ◽

Christian Bogdan

Keyword(s):

Borrelia Burgdorferi ◽

The United States ◽

Tick Population ◽

Original Sequence ◽

Southern Germany ◽

Granulocytic Ehrlichiosis ◽

Ehrlichia Phagocytophila ◽

16S Ribosomal Dna ◽

Human Granulocytic Ehrlichiosis ◽

Pcr Products

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. burgdorferiby 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 andB. burgdorferi, respectively. Two ticks (0.7%) were coinfected with both bacteria. Of 12 engorged I. ricinusticks 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-bpEhrlichia 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.

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PCR Detection of Granulocytic Ehrlichiae in Ixodes ricinus Ticks and Wild Small Mammals in Western Switzerland

Journal of Clinical Microbiology ◽

10.1128/jcm.38.3.1002-1007.2000 ◽

2000 ◽

Vol 38 (3) ◽

pp. 1002-1007 ◽

Cited By ~ 126

Author(s):

Jorge S. Liz ◽

Laurence Anderes ◽

John W. Sumner ◽

Robert F. Massung ◽

Lise Gern ◽

...

Keyword(s):

Ixodes Ricinus ◽

Small Mammals ◽

Common Shrew ◽

Wood Mouse ◽

Clethrionomys Glareolus ◽

Rrna Gene ◽

Questing Ticks ◽

Ixodes Ricinus Ticks ◽

High Degree ◽

Dual Infections

The presence of granulocytic ehrlichiae was demonstrated by PCR inIxodes ricinus ticks and wild small mammals in Switzerland in two areas of endemicity for bovine ehrlichiosis. Six ticks (three females and three nymphs) (1.4%) of 417 I. ricinus ticks collected by flagging vegetation contained ehrlichial DNA. A total of 201 small mammals from five species, wood mouse (Apodemus sylvaticus), yellow-necked mouse (Apodemus flavicollis), earth vole (Pitymys subterraneus), bank vole (Clethrionomys glareolus), and common shrew (Sorex araneus), were trapped. The analysis of I. ricinus mammals collected on 116 small mammals showed that nine C. glareolus voles and two A. sylvaticus mice hosted infected tick larvae. In these rodents, granulocytic ehrlichia infection was also detected in blood, spleen, liver, and ear samples. Further examinations of 190 small mammals without ticks or with noninfected ticks showed the presence of ehrlichial DNA in spleen and other tissues from six additional C. glareolus, three A. flavicollis, and one S. araneus mammals. This study suggests thatA. sylvaticus, A. flavicollis, S. araneus, and particularly C. glareolus are likely to be natural reservoirs for granulocytic ehrlichiae. Partial 16S rRNA gene sequences of granulocytic ehrlichiae from ticks and rodents showed a high degree of homology (99 to 100%) with granulocytic ehrlichiae isolated from humans. In contrast, groESL heat shock operon sequence analysis showed a strong divergence (approximately 5%) between the sequences in samples derived from rodents and those derived from samples from questing ticks or from other published ehrlichia sequences. Dual infections with granulocytic ehrlichia andBorrelia burgdorferi were found in ticks and small mammals.

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Antimicrobial Susceptibility ofEhrlichia phagocytophila

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.45.3.786-788.2001 ◽

2001 ◽

Vol 45 (3) ◽

pp. 786-788 ◽

Cited By ~ 45

Author(s):

Harold W. Horowitz ◽

T.-C. Hsieh ◽

Maria E. Aguero-Rosenfeld ◽

Fatemeh Kalantarpour ◽

Ishraq Chowdhury ◽

...

Keyword(s):

New York ◽

Culture System ◽

Minimum Bactericidal Concentration ◽

New York State ◽

Cell Culture System ◽

Obligate Intracellular ◽

Granulocytic Ehrlichiosis ◽

Ehrlichia Phagocytophila ◽

Human Granulocytic Ehrlichiosis ◽

Quinolone Antibiotics

ABSTRACT Human granulocytic ehrlichiosis is a recently described disease caused by an obligate intracellular gram-negative organism recently named Ehrlichia phagocytophila. To expand our knowledge of the susceptibility of E. phagocytophila, we tested six New York State isolates for susceptibility to 12 antimicrobials using an HL-60 cell culture system. All of the isolates were susceptible to doxycycline (MIC, ≤0.125 μg/ml; minimum bactericidal concentration [MBC], 0.125 to 0.5 μg/ml), rifampin (MIC, ≤0.125 μg/ml; MBC, ≤0.125 μg/ml), ofloxacin (MIC, ≤2 μg/ml; MBC, ≤2 μg/ml), levofloxacin (MIC, ≤1 μg/ml; MBC, ≤1 μg/ml), and trovafloxacin (MIC, ≤0.032 μg/ml; MBC, ≤0.032 μg/ml). Isolates were uniformly resistant to amoxicillin, ceftriaxone, erythromycin, azithromycin, clarithromycin, and amikacin. For one strain, the MBC of chloramphenicol was ≤8 μg/ml. These data suggest that quinolone antibiotics and rifampin may be alternative agents for patients with intolerance to tetracyclines.

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Natural Ehrlichia phagocytophila transmission coefficients from sheep ‘carriers’ to Ixodes ricinus ticks vary with the numbers of feeding ticks

Parasitology ◽

10.1017/s003118200100107x ◽

2002 ◽

Vol 124 (2) ◽

pp. 127-136 ◽

Cited By ~ 28

Author(s):

N. H. OGDEN ◽

A. N. J. CASEY ◽

N. P. FRENCH ◽

K. J. BOWN ◽

J. D. W. ADAMS ◽

...

Keyword(s):

Ixodes Ricinus ◽

Tick Infestation ◽

Infection Intensity ◽

Adult Tick ◽

Transmission Efficiency ◽

Infection Prevalence ◽

Direct Effects ◽

Transmission Coefficients ◽

Ehrlichia Phagocytophila ◽

Ixodes Ricinus Ticks

In a longitudinal study in a UK upland site, 38% of adult sheep were detected as infected with the tick-borne bacterium Ehrlichia phagocytophila by PCR of blood samples. Infection prevalence declined significantly with sheep age but varied significantly and non-linearly with the number of adult Ixodes ricinus ticks feeding per sheep. These findings suggested that under conditions of natural repeated tick-borne challenge sheep remain partially susceptible to re-infections, but the likelihood of re-infection depended on the numbers of feeding ticks. Transmission efficiency from sheep to immature ticks also varied significantly and non-linearly with the number of adult ticks feeding per sheep: transmission efficiency was almost zero in sheep with low adult tick infestations rising to 30% at certain levels of adult tick infestation. Infection intensity in infected engorged immature ticks also varied with the number of adult ticks feeding per sheep, but neither prevalence nor intensity of infection in engorged ticks were related to sheep blood PCR result. These findings suggest that variation in the numbers of ticks feeding per sheep may influence E. phagocytophila transmission by direct effects on transmission at the tick–host interface.

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