Quantitative Detection of
            <i>Borrelia burgdorferi</i>
            in 2-Millimeter Skin Samples of Erythema Migrans Lesions: Correlation of Results with Clinical and Laboratory Findings

Liveris, Dionysios; Wang, Guiqing; Girao, Gary; Byrne, Daniel W.; Nowakowski, John; McKenna, Donna; Nadelman, Robert B.; Wormser, Gary P.; Schwartz, Ira

doi:10.1128/jcm.40.4.1249-1253.2002

Cited by 91 publications

(98 citation statements)

References 28 publications

(19 reference statements)

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“…As shown in this study, comparable numbers of spirochetes were detected in ticks infected with RST1 and RST3 genotypes of B. burgdorferi (with means of 1,980 and 2,212, respectively; P Ͼ 0.05). In contrast, the spirochete load, which correlates directly with the severity of disease, was at least twofold higher in the skin biopsy specimens of patients and in heart and joint tissues of mice infected with RST1 isolates than in those infected with RST3 isolates (19,36,37).…”

Section: Discussionmentioning

confidence: 79%

“…The technique has recently been employed to detect the presence of B. burgdorferi DNA in I. ricinus ticks from Switzerland (15), to quantify the spirochete loads in experimentally infected animal tissues (23,24,35), and to differentiate the three B. burgdorferi sensu lato species that are pathogenic to humans in Europe (22,28,30). By targeting the B. burgdorferi-specific recA gene, we have successfully developed a real-time PCR protocol to quantify the spirochetes in infected animal tissues and in skin biopsy specimens of patients with erythema migrans (19,36,37). The present study evaluates a modified real-time, quantitative PCR (qPCR) protocol for simultaneous detection and quantification of B. burgdorferi DNA in field-collected I. scapularis ticks from the northeastern United States.…”

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confidence: 99%

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Real-Time PCR for Simultaneous Detection and Quantification of Borrelia burgdorferi in Field-Collected Ixodes scapularis Ticks from the Northeastern United States

Wang¹,

Liveris²,

Brei

et al. 2003

Appl Environ Microbiol

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The density of spirochetes in field-collected or experimentally infected ticks is estimated mainly by assays based on microscopy. In this study, A high degree of spirochete aggregation among infected ticks (variance-to-mean ratio of 24,877; moment estimate of k ‫؍‬ 0.279) was observed. From the frequency distribution data and previously published transmission studies, we estimated that a minimum of 300 organisms may be required in a host-seeking nymphal tick to be able to transmit infection to mice while feeding on mice. These data indicate that real-time qPCR is a reliable approach for simultaneous detection and quantification of B. burgdorferi infection in field-collected ticks and can be used for ecological and epidemiological surveillance of Lyme disease spirochetes.Hard ticks (Ixodidae) in the Ixodes persulcatus complex, such as Ixodes scapularis and Ixodes pacificus in North America, Ixodes ricinus in Europe, and Ixodes persulcatus in far eastern Russia and Asia, are the principal vectors of the Lyme disease spirochete, Borrelia burgdorferi, and several other tick-borne pathogens (2, 25). The incidence of human Lyme disease in areas where it is endemic is correlated with the abundance and prevalence of Ixodes ticks infected with B. burgdorferi (34). Moreover, studies of laboratory animals have suggested that the efficiency of host infection is determined in part by the number of spirochetes inoculated using a needle or deposited by the tick at the time of feeding (21, 27). Thus, monitoring B. burgdorferi density in host-seeking ticks will provide more accurate data for assessment of population risk of Lyme disease and the potential variability of disease manifestations after tick bites.Currently, the number of spirochetes in field-collected or experimentally infected Ixodes ticks is estimated mainly by microscopy-based assays in which the spirochetes are counted directly on tick midgut smears stained with fluorescently labeled specific antibodies (8, 9, 27) or on silver-stained histological sections of ticks (11,14). For specific detection of B. burgdorferi in field-collected adult I. scapularis ticks and monitoring of the spirochete kinetics during the tick life cycle, an OspA antigen-capture enzyme-linked immunosorbent assay (ELISA) was also developed (3-5). These methods are laborintensive, which limits the number of ticks that can be analyzed. Moreover, the sensitivities of these methods are relatively low, e.g., a lower detection limit of approximately 150 spirochetes was reported for the antigen-capture ELISA (5).Real-time PCR has features that allow for rapid detection of infectious pathogens in environmental or experimentally infected animal tissues and clinical specimens (38). The technique has recently been employed to detect the presence of B. burgdorferi DNA in I. ricinus ticks from Switzerland (15), to quantify the spirochete loads in experimentally infected animal tissues (23,24,35), and to differentiate the three B. burgdorferi sensu lato species that are pathogenic to humans in Europe (22,2...

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Section: Discussionmentioning

confidence: 79%

mentioning

confidence: 99%

Real-Time PCR for Simultaneous Detection and Quantification of Borrelia burgdorferi in Field-Collected Ixodes scapularis Ticks from the Northeastern United States

Wang¹,

Liveris²,

Brei

et al. 2003

Appl Environ Microbiol

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“…There are some reports confirming the utility of PCR assays in the detection of Borrelia spp. in cerebrospinal fluid (CSF), synovial fluid, skin bioptates and urine samples [18][19][20]. Taking into account that the etiologic agent of LD, Borrelia burgdorferi sensu lato, was recovered first from the tick Ixodes dammini in the year 1982 [2] and then from a skin biopsy, CSF and blood specimens of patients with LB around the world [2,[21][22][23][24][25], the main aim of this study was to discuss the utility of introduction of a well-optimized, sensitive and highly specific quantitative real-time PCR assay into the diagnostics of Borrelia burgdorferi sensu lato infections.…”

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confidence: 99%

Lyme Borreliosis - the Utility of Improved Real-Time PCR Assay in Detection of Borrelia burgdorferi Infections

Bil‐Lula¹,

Matuszek²,

Woźniak³

2015

Adv Clin Exp Med

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Background. Infections of Borrelia burgdorferi sensu lato reveal clinical manifestations affecting numerous organs and tissues. The standard diagnostic procedure of these infections is quite simple if a positive history of tick exposure or typical erythema migrans appears. Lack of unequivocal clinical symptoms creates the necessity for further evaluation with laboratory tests. Objectives. This study discusses the utility of a novel, improved, well-optimized, sensitive and highly specific quantitative real-time PCR assay for the diagnostics of infections caused by Borrelia burgdorferi sensu lato. Material and Methods. We designed an improved, specific, highly sensitive real-time quantitative polymerase chain reaction (RQ-PCR) assay for the detection and quantification of all Borrelia burgdorferi genotypes. A wide validation effort was undertaken to ensure confidence in the highly sensitive and specific detection of B. burgdorferi. Results. Due to high sensitivity and great specificity, as low as 1.6 × 10² copies of Borrelia per mL of whole blood could be detected. As much as 12 (3%) negative ELISA IgM results, 14 (2.8%) negative results of Line blot IgM, 11 (3.1%) and 7 (2.7%) of negative ELISA IgG and Line blot IgG results, respectively, were positive in real-time PCR. Conclusions. The data in this study confirms the high positive predictive value of real-time PCR test in the detection of Borrelia infections (Adv Clin Exp Med 2015, 24, 4, 663-670).

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“…When coupled with culture results, levels of diagnostic sensitivity of Ͼ90% were obtained. However, the burgdorferi DNA in skin biopsies (9). While the real-time quantitative PCR on skin biopsies did not achieve that high level of sensitivity in our hands, it was still a sensitive early marker in almost 60% of persons with EM who were tested by PCR.…”

Section: Discussionmentioning

confidence: 91%

“…However, when accepted signs and/or symptoms are unsatisfactory or inconclusive, laboratory testing may be needed for improving diagnostic certainty. Serologic testing is the mainstay for laboratory-based diagnosis, although modern technologies have provided other tools that variably contribute, including PCR and culture (5,9,10,15). The utility of specific laboratory diagnostics, culture in particular, has also been emphasized given the need for gold standards for evaluation of new diagnostic tests and for use in laboratory authentication of Lyme disease in investigations of new treatments and vaccines (10)(11)(12).…”

Section: Discussionmentioning

confidence: 99%

Two-Year Evaluation of Borrelia burgdorferi Culture and Supplemental Tests for Definitive Diagnosis of Lyme Disease

et al. 2005

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Lyme disease is usually diagnosed and treated based on clinical manifestations. However, laboratory testing is useful for patients with confusing presentations and for validation of disease in clinical studies. Although cultivation of Borrelia burgdorferi is definitive, prior investigations have shown that no single test is optimal for Lyme disease diagnosis. We applied high-volume blood culture, skin biopsy culture, PCR, and serodiagnosis to a cohort of patients with suspected Lyme disease acquired in Maryland and southern Pennsylvania. The study was performed to confirm the relative utility of culture and to identify laboratory testing algorithms that will supplement clinical diagnosis. Overall, 30 of 86 patients (35%) were culture positive, whereas an additional 15 of 84 (18%) were seropositive only (51% total sero-and culture positive), and PCR on skin biopsy identified 4 additional patients who were neither culture nor seropositive. Among 49 laboratory test-positive patients, the highest sensitivity (100%) for diagnosis was obtained when culture, skin PCR, and serologic tests were used, although serologic testing with skin PCR was almost as sensitive (92%). Plasma PCR was infrequently positive and provided no additional diagnostic value. Although culture is definitive and has a relatively high sensitivity, the results required a mean of 3.5 weeks to recovery. The combination of acute-phase serology and skin PCR was 75% sensitive, offering a practical and relatively rapid alternative for confirming clinical impression. The full battery of tests could be useful for patients with confusing clinical signs or for providing strong laboratory support for clinical studies of Lyme disease.

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Quantitative Detection of Borrelia burgdorferi in 2-Millimeter Skin Samples of Erythema Migrans Lesions: Correlation of Results with Clinical and Laboratory Findings

Cited by 91 publications

References 28 publications

Real-Time PCR for Simultaneous Detection and Quantification of Borrelia burgdorferi in Field-Collected Ixodes scapularis Ticks from the Northeastern United States

Real-Time PCR for Simultaneous Detection and Quantification of Borrelia burgdorferi in Field-Collected Ixodes scapularis Ticks from the Northeastern United States

Lyme Borreliosis - the Utility of Improved Real-Time PCR Assay in Detection of Borrelia burgdorferi Infections

Two-Year Evaluation of Borrelia burgdorferi Culture and Supplemental Tests for Definitive Diagnosis of Lyme Disease

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