Clinicians who are faced with the difficult task of identifying and treating patients
who have tick–bite or tick–exposure-associated illnesses often have difficult searches
for effective laboratory confirmation of clinical impressions. For the emerging tick-borne
infections, Lyme disease and the different forms of ehrlichiosis, this difficult is
due, in part, to the nonspecificity of clinical manifestations and laboratory studies.
The difficulty is confounded even further because of the relative infrequency of tick-borne
diseases in many geographic regions and the potential for severe and fatal outcome
in some cases. One of the major pitfalls in accomplishing laboratory confirmation
has been the limited development and availability of diagnostic tests for these infections,
in part because of the scarcity of “gold-standard” samples for comparison and assay
validation. The maturation of molecular technologies for diagnosis of infectious diseases
has offered great promise, only partly fulfilled, to remedy this situation. For ehrlichiosis,
the major goal of a molecular diagnostic test would be to identify definitively the
infectious agent rapidly and perhaps to assess whether antibiotic therapy has effected
a cure. Because Lyme disease generally presents differently, the major goals for molecular
testing would include (1) the differentiation of annular skin lesions, (2) defining
whether a previously infected (seropositive) or vaccinated patient may have active
infection, (3) differentiating Borrelia burgdorferi meningitis or arthritis from other infectious and noninfectious etiologies, (4) identifying
active infection before the development of antibodies, and (5) use as a test for cure.
Thus, the major objective of this article is to examine molecular techniques that
have been developed and are effective for these uses in the clinical laboratory for
the emerging tick-borne infections, ehrlichiosis and Lyme disease. An important consideration
for application of any laboratory diagnostic test is the predictive value of the test
when applied in any specific circumstance. The positive predictive value is affected
significantly by the prevalence of the infection in a population, or, for a diagnostic
test, the pretest probability of infection. Thus, the clinical value of these molecular
diagnostics is, in part, predicated on excluding patients who are unlikely to be infected
by careful clinical and historic examinations. Accordingly, the overall clinical usefulness
may be characterized, in part, as a function of its sensitivity, specificity, and
the specificity of pretest clinical findings (Table 1).
Table 1PCR diagnostics for Lyme disease, HME, human anaplasmosis, and ehrlichiosis ewingii
| Sample type | Sample preparation | Gene targets | Clinical usefulness |
|---|---|---|---|
| Lyme disease—B burgdorferi sensu lato | |||
| Skin | 2 mm–4 mm punch biopsy | ospA, fla, ssr | ++ |
| Blood plasma | EDTA anticoagulated plasma | ospA, fla, ssr | + |
| Synovial fluid | ospA, fla, ssr | ++ | |
| CSF | ospA, fla, ssr | + | |
| urine | ospA, fla, ssr | 0 | |
| HME—E chaffeensis | |||
| Blood | EDTA anticoagulated whole blood | rrs, gp120, p28, VLPT, groESL | +++ |
| CSF | rrs, gp120, p28, VLPT, groESL | ++ | |
| Human anaplasmosis (formerly HGE)—A phagocytophilum | |||
| Blood | EDTA anticoagulated whole blood | rrs, msp2, ankA, groESL | +++ |
| Ehrlichiosis ewingii—E ewingii | |||
| Blood | EDTA anticoagulated whole blood | rrs, p28, groESL | ++? |
a Major gene targets for PCR diagnosis.
b 0, probably not useful; +, useful infrequently; ++, useful in specific circumstances;
+++, highly useful in appropriate clinical settings (see text for specific applications
and recommendations).
To read this article in full you will need to make a payment
Purchase one-time access:
Academic & Personal: 24 hour online accessCorporate R&D Professionals: 24 hour online accessOne-time access price info
- For academic or personal research use, select 'Academic and Personal'
- For corporate R&D use, select 'Corporate R&D Professionals'
Subscribers receive full online access to your subscription and archive of back issues up to and including 2002.
Content published before 2002 is available via pay-per-view purchase only.
Subscribe:
Subscribe to Clinics in Laboratory MedicineAlready a print subscriber? Claim online access
Already an online subscriber? Sign in
Register: Create an account
Institutional Access: Sign in to ScienceDirect
References
- Lyme arthritis: an epidemic of oligoarticular arthritis in children and adults in three Connecticut communities.Arthritis Rheum. 1977; 20: 7-17
- Lyme disease.N Engl J Med. 2001; 345: 115-125
- Clinical characteristics and treatment outcome of early Lyme disease in patients with microbiologically confirmed erythema migrans.Ann Intern Med. 2002; 136: 421-428
- Two controlled trials of antibiotic treatment in patients with persistent symptoms and a history of Lyme disease.N Engl J Med. 2001; 345: 85-92
- Evolution of the serologic response to Borrelia burgdorferi in treated patients with culture-confirmed erythema migrans.J Clin Microbiol. 1996; 34: 1-9
- Lyme disease—a tick-borne spirochetosis?.Science. 1982; 216: 1317-1319
- Yield of large-volume blood cultures in patients with early Lyme disease.J Infect Dis. 2001; 184: 1070-1072
- Genomic sequence of a Lyme disease spirochaete, Borrelia burgdorferi.Nature. 1997; 390: 580-586
- Antigenic and genetic heterogeneity of Borrelia burgdorferi populations transmitted by ticks.Proc Natl Acad Sci USA. 2001; 98: 670-675
- Changes in temporal and spatial patterns of outer surface lipoprotein expression generate population heterogeneity and antigenic diversity in the Lyme disease spirochete, Borrelia burgdorferi.Infect Immun. 2002; 70: 3468-3478
- Borrelia burgdorferi-induced inflammation facilitates spirochete adaptation and variable major protein-like sequence locus recombination.J Immun. 2001; 167: 3383-3390
- Evidence that the variable regions of the central domain of VlsE are antigenic during infection with lyme disease spirochetes.Infect Immun. 2002; 70: 4196-4203
- Differential binding of host complement inhibitor factor H by Borrelia burgdorferi erp surface proteins: a possible mechanism underlying the expansive host range of Lyme disease spirochetes.Infect Immun. 2002; 70: 491-497
- Comparison of in vitro culture, immunohistochemical staining and PCR for detection of Borrelia burgdorferi in tissue from experimentally infected animals.J Clin Microbiol. 1995; 33: 2328-2333
- Amplification of Borrelia burgdorferi DNA in skin biopsies from patients with Lyme disease.J Clin Microbiol. 1991; 29: 2401-2406
- Molecular detection of persistent Borrelia burgdorferi in the urine of patients with active Lyme disease.Infect Immun. 1991; 59: 269-278
- Demonstration of Borrelia burgdorferi DNA in urine samples from healthy humans whose sera contain B burgdorferi-specific antibodies.J Clin Microbiol. 1994; 32: 2312-2314
- Diagnosis of early Lyme disease by polymerase chain reaction amplification and culture of skin biopsies from erythema migrans lesions.J Clin Microbiol. 1992; 30: 3082-3088
- PCR detection of Borrelia burgdorferi DNA in cerebrospinal fluid of Lyme neuroborreliosis patients.Neurology. 1992; 42: 32-42
- Invasion of the central nervous system by Borrelia burgdorferi in acute disseminated infection.JAMA. 1992; 267: 1364-1367
- Bloodstream invasion in early Lyme disease: results from a prospective, controlled, blinded study using polymerase chain reaction.Am J Med. 1995; 99: 6-12
- Detection of Borrelia burgdorferi DNA by polymerase chain reaction in synovial fluid from patients with Lyme arthritis.N Engl J Med. 1994; 330: 229-234
- Detection of Borrelia burgdorferi infection in Ixodes dammini ticks with the polymerase chain reaction.J Clin Microbiol. 1990; 28: 566-572
- Laboratory diagnostic techniques for patients with early Lyme disease associated with erythema migrans: a comparison of different techniques.Clin Infect Dis. 2001; 33: 2023-2027
- Practice guidelines for the treatment of Lyme disease. The Infectious Diseases Society of America.Clin Infect Dis. 2000; 31: 1-14
- Molecular tools for the diagnosis of Lyme disease: review and meta-analysis.Mol Diagn. 2001; 6: 1-11
- Four clones of Borrelia burgdorferi sensu stricto cause invasive infection in humans.Infect Immun. 1999; 67: 3518-3524
- Yield of large-volume blood cultures in patients with early Lyme disease.J Infect Dis. 2001; 184: 1070-1072
- Musculoskeletal manifestations of Lyme disease.Am J Med. 1995; 98: 44S-48S
- Lack of Borrelia burgdorferi DNA in synovial samples from patients with antibiotic treatment-resistant Lyme arthritis.Arthritis Rheum. 1999; 42: 2705-2709
- Lyme arthritis: lessons learned and to be learned.Arthritis Rheum. 1999; 42: 1809-1812
- Lyme arthritis and post-Lyme disease syndrome.Curr Opin Rheumatol. 2002; 14: 383-387
- Target imbalance: disparity of Borrelia burgdorferi genetic material in synovial fluid from Lyme arthritis patients.J Infect Dis. 1994; 169: 668-672
- Neurologic aspects of Lyme disease.Med Clin North Am. 2002; 86: 261-284
- Tick bites and Lyme disease in an endemic setting: problematic use of serologic testing and prophylactic antibiotic therapy.JAMA. 1998; 279: 206-210
- Neuroborreliosis: central nervous system involvement.Semin Neurol. 1997; 17: 19-24
- Evaluation of the intrathecal antibody response to Borrelia burgdorferi as a diagnostic test for Lyme neuroborreliosis.J Infect Dis. 1990; 161: 1203-1209
- Central nervous system abnormalities in Lyme neuroborreliosis.Neurology. 1991; 41: 1571-1582
- Laboratory evaluation in the diagnosis of Lyme disease. Clinical guideline, part 2.Ann Intern Med. 1997; 127: 1109-1123
- Diagnostic value of PCR for detection of Borrelia burgdorferi DNA in clinical specimens from patients with erythema migrans and Lyme neuroborreliosis.Mol Diagn. 2000; 5: 139-150
- Detection of Borrelia burgdorferi in cerebrospinal fluid by the polymerase chain reaction.J Med Microbiol. 1991; 35: 98-102
- Detection of Borrelia burgdorferi DNA in urine samples and in cerebrospinal fluid samples from patients with early and late Lyme neuroborreliosis by polymerase chain reaction.J Clin Microbiol. 1992; 30: 1646-1653
- Polymerase chain reaction with the 30-kb circular plasmid of Borrelia burgdorferi B31 as a target for detection Lyme borreliosis agents in cerebrospinal fluid.Res Microbiol. 1993; 144: 211-219
- Detection of Borrelia burgdorferi by nest polymerase chain reaction in cerebrospinal fluid and urine of children with neuroborreliosis.Eur J Pediatr. 1993; 152: 414-417
- The polymerase chain reaction in the diagnosis of Lyme neuroborreliosis.Ann Neurol. 1993; 34: 544-550
- Comparison of two methods for detecting intrathecal synthesis of Borrelia burgdorferi—specific antibodies and PCR for diagnosis of Lyme neuroborreliosis.J Clin Microbiol. 1994; 32: 1795-1798
- Evaluation of the polymerase chain reaction for the detection of Borrelia burgdorferi in cerebrospinal fluid of children with acute peripheral facial palsy.Eur J Pediatr. 1995; 154: 374-377
- Value of clinical symptoms, intrathecal specific antibody production and PCR in CSF in the diagnosis of childhood Lyme neuroborreliosis.Klin Pediatr. 1996; 208: 106-109
- Detection of Borrelia burgdorferi DNA by polymerase chain reaction in cerebrospinal fluid in Lyme neuroborreliosis.J Infect Dis. 1996; 174: 623-627
- Diagnostic detection and direct genotyping of Borrelia burgdorferi by polymerase chain reactions in cerebrospinal fluid in Lyme neuroborreliosis.Mol Diagn. 1998; 3: 131-141
- Effective use of polymerase chain reaction for diagnosis of central nervous system infections.Clin Infect Dis. 1999; 29: 803-806
- Evaluation of the detection of Borrelia burgdorferi DNA in urine samples by polymerase chain reaction.Infection. 1995; 23: 173-179
- Borrelia burgdorferi DNA in the urine of treated patients with chronic Lyme disease symptoms. A PCR study of 97 cases.Infection. 1996; 24: 347-353
- Detection of Borrelia burgdorferi DNA by polymerase chain reaction in urine specimens of patients with erythema migrans lesions.Mol Cell Probes. 1997; 11: 89-94
- Diagnostic value of PCR for detection of Borrelia burgdorferi in skin biopsy and urine samples from patients with skin borreliosis.J Clin Microbiol. 1998; 36: 2658-2665
- Detection of Borrelia burgdorferi DNA in urine of patients with ocular Lyme borreliosis.Br J Ophthalmol. 2001; 85: 552-555
- Intralaboratory reliability of serologic and urine testing for Lyme disease.Am J Med. 2001; 110: 217-219
- Human infection with Ehrlichia canis, a leukocytic rickettsia.N Engl J Med. 1987; 316: 853-856
- Identification of a granulocytotropic Ehrlichia species as the etiologic agent of human disease.J Clin Microbiol. 1994; 32: 589-595
- Ehrlichia ewingii, a newly recognized agent of human ehrlichiosis.N Engl J Med. 1999; 341: 148-155
- Tick-borne ehrlichioses: More of them, higher incidences, and greater clinical diversity.Lancet Infect Dis. 2001; 0: 21-28
- Human ehrlichioses.Med Clin North Am. 2002; 86: 375-392
- Reorganization of genera in the families Rickettsiaceae and Anaplasmataceae in the order Rickettsiales: unification of some species of Ehrlichia with Anaplasma, Cowdria with Ehrlichia and Ehrlichia with Neorickettsia, descriptions of six new species combinations and designation of Ehrlichia equi and “HGE agent” as subjective synonyms of Ehrlichia phagocytophila.Int J Syst Evol Microbiol. 2001; 51 ([published correction appears in Int J System Evol Microbiol 2002;52:5–6]): 2145-2165
- Human granulocytic ehrlichiosis (HGE): epidemiologic, clinical and laboratory characteristics for 41 patients from Minnesota and Wisconsin.JAMA. 1996; 275: 199-205
- Clinical and laboratory spectrum of culture-proven human granulocytic ehrlichiosis: comparison with culture-negative cases.Clin Infect Dis. 1998; 27: 1314-1317
- Primary isolation of Ehrlichia chaffeensis from patients with febrile illnesses: clinical and molecular characteristics.J Infect Dis. 2000; 181: 1082-1088
- Diagnosing human ehrlichioses: current status and recommendations.ASM News. 2000; 66 (Available at:) (Accessed November 4, 2003): 287-291
- Outcome of diagnostic tests using samples from patients with culture-proven human monocytic ehrlichiosis: implications for surveillance.J Clin Microbiol. 1999; 37: 2997-3000
- Improved sensitivity of PCR for diagnosis of human granulocytic ehrlichiosis using epank1 genes of Ehrlichia phagocytophila-group ehrlichiae.J Clin Microbiol. 2000; 38: 354-356
- Human ehrlichiosis in the United States, 1985 to 1990.Ann Intern Med. 1994; 120: 736-743
- Susceptibility of dogs to infection with Ehrlichia chaffeensis, the causative agent of human ehrlichiosis.Am J Vet Res. 1992; 53: 1322-1327
- Susceptibility of white-tailed deer (Odocoileus virginianus) to infection with Ehrlichia chaffeensis, the etiologic agent of human ehrlichiosis.J Clin Microbiol. 1994; 32: 2725-2728
- Identification of a p28 gene in Ehrlichia ewingii: evaluation of gene for use as a target for a species-specific PCR diagnostic assay.J Clin Microbiol. 2001; 39: 3871-3876
- The incidence of ehrlichial and rickettsial infection in patients with unexplained fever and recent history of tick bite in central North Carolina.J Infect Dis. 1999; 180: 900-903
- Detection of the etiologic agent of human ehrlichiosis by polymerase chain reaction.J Clin Microbiol. 1992; 30: 775-780
- Human ehrlichiosis in adults after tick exposure. Diagnosis using polymerase chain reaction.Ann Intern Med. 1994; 120: 730-735
- Infections with Ehrlichia chaffeensis and Ehrlichia ewingii in persons coinfected with human immunodeficiency virus.Clin Infect Dis. 2001; 33: 1586-1594
- PCR amplification and comparison of nucleotide sequences from the groESL heat shock operon of Ehrlichia species.J Clin Microbiol. 1997; 35: 2087-2092
- Sequence and characterization of an Ehrlichia chaffeensis gene encoding 314 amino acids highly homologous to the NAD A enzyme.FEMS Microbiol Lett. 1997; 154: 53-58
- Isolation and characterization of Ehrlichia chaffeensis strains from patients with fatal ehrlichiosis.J Clin Microbiol. 1997; 35: 2496-2502
- The clinical spectrum of disease due to Ehrlichia chaffeensis: report of two cases and review.Infect Dis Clin Prac. 1998; 7: 252-256
- Human granulocytic ehrlichiosis in the upper midwest United States. A new species emerging?.JAMA. 1994; 272: 212-218
- Transmission and passage in horses of the agent of human granulocytic ehrlichiosis.J Infect Dis. 1995; 172: 1141-1144
- The human ehrlichioses in the United States.Emerg Infect Dis. 1999; 5: 635-642
- Seroprevalence of human granulocytic ehrlichiosis among permanent residents of northwestern Wisconsin.Clin Infect Dis. 1998; 27: 1491-1496
- Human monocytic and granulocytic ehrlichioses. Discovery and diagnosis of emerging tick-borne infections and the critical role of the pathologist.Arch Pathol Lab Med. 1997; 121: 785-791
- Human granulocytic ehrlichiosis (HGE): a series from a single medical center in New York State.Ann Intern Med. 1996; 125: 904-908
- Serial measurements of hematologic counts during the active phase of human granulocytic ehrlichiosis.Clin Infect Dis. 2001; 32: 862-870
- Quantitative evaluation of ehrlichial burden in horses after experimental transmission of human granulocytic Ehrlichia agent by intravenous inoculation with infected leukocytes and by infected ticks.J Clin Microbiol. 1999; 37: 4042-4044
- Experimental inoculation with human granulocytic Ehrlichia agent derived from high- and low-passage cell culture in horses.J Clin Microbiol. 2000; 38: 1276-1278
- Evaluation of an improved PCR diagnostic assay for human granulocytic ehrlichiosis.Mol Diagn. 1996; 1: 41-49
- The emergence of another tickborne infection in the 12-town area around Lyme, Connecticut: human granulocytic ehrlichiosis.J Infect Dis. 2000; 181: 1388-1393
- Reinfection with the agent of human granulocytic ehrlichiosis.Ann Intern Med. 1998; 129: 461-463
- Direct cultivation of the causative agent from patients with human granulocytic ehrlichiosis.N Engl J Med. 1996; 334: 209-215
- Major antigenic proteins of the agent of human granulocytic ehrlichiosis are encoded by members of a multigene family.Infect Immun. 1998; 66: 3711-3718
- Cloning of the gene encoding the 44-kilodalton antigen of the agent of human granulocytic ehrlichiosis and characterization of the humoral response.Infect Immun. 1998; 66: 3264-3269
- ankA: an Ehrlichia phagocytophila group gene encoding a cytoplasmic protein antigen with ankyrin repeats.Infect Immun. 2000; 68: 5277-5283
- Genetic variability and stability of Anaplasma phagocytophila msp2 (p44).Infect Immun. 2002; 70: 1230-1234
- Cloning and expression of the 44-kilodalton major outer membrane protein gene of the human granulocytic ehrlichiosis agent and application of the recombinant protein to serodiagnosis.J Clin Microbiol. 1998; 36: 1666-1673
- Molecular cloning and sequencing of three granulocytic Ehrlichia genes encoding high-molecular weight immunoreactive proteins.Infect Immun. 1998; 66: 1356-1363
- Experimental transmission of a granulocytic form of the tribe Ehrlichieae by Dermacentor variabilis and Amblyomma americanum to dogs.Am J Vet Res. 1990; 51: 929-931
- Ehrlichia ewingii infection in white-tailed deer (Odocoileus virginianus).Emerg Infect Dis. 2002; 8: 668-671
- Granulocytic ehrlichiosis in dogs from North Carolina and Virginia.J Vet Intern Med. 1998; 12: 61-70
- A molecular and serologic survey of Ehrlichia canis, E chaffeensis, and E ewingii in dogs and ticks from Oklahoma.Vet Parasitol. 1998; 79: 325-339
Article info
Identification
Copyright
© 2003 Elsevier Inc. Published by Elsevier Inc. All rights reserved.
