Development of a PCR for Borrelia burgdorferi sensu lato, targeted on the groEL gene

Chiappa, Giulia; Cafiso, Alessandra; Monza, Elisa; Serra, Valentina; Olivieri, Emanuela; Romeo, Claudia; Bazzocchi, Chiara

doi:10.14411/fp.2020.026

Cited by 5 publications

(5 citation statements)

References 26 publications

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“…In the past few years, several studies provided new or modified PCR assays for detection of Borrelia. 21–27 A fragment of the groEl gene, which encodes a highly conserved 60 kDa heat shock protein of B. burgdorferi s.l. , was used for detection in ticks.…”

Section: Discussionmentioning

confidence: 99%

“…In the past few years, several studies provided new or modified PCR assays for detection of Borrelia. [21][22][23][24][25][26][27] A fragment of the groEl gene, which encodes a highly conserved 60 kDa heat shock protein of B. burgdorferi s.l., was used for detection in ticks. 21 Others focused on identification of Borrelia in blood samples, for example, by way of a quantitative real-time PCR targeting ribosomal genes (16SrRNA), which identified Borrelia infections while IgM serology was still negative 22 or by using specific DNA hybridization to purify B. burgdorferi DNA from blood followed by PCR amplification of flagellin and OspA gene fragments.…”

Section: Discussionmentioning

confidence: 99%

“…[21][22][23][24][25][26][27] A fragment of the groEl gene, which encodes a highly conserved 60 kDa heat shock protein of B. burgdorferi s.l., was used for detection in ticks. 21 Others focused on identification of Borrelia in blood samples, for example, by way of a quantitative real-time PCR targeting ribosomal genes (16SrRNA), which identified Borrelia infections while IgM serology was still negative 22 or by using specific DNA hybridization to purify B. burgdorferi DNA from blood followed by PCR amplification of flagellin and OspA gene fragments. 23 A multiplexed high-definition PCR (HDPCR) Tickborne Panel (TBP) assay (ChromaCode, Carlsbad, CA) attempted to identify several tick-borne pathogens in 1 assay, but it reached only a sensitivity of 44.4% for B. burgdorferi s.l.…”

Section: Discussionmentioning

confidence: 99%

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Diagnosis of Lyme Borreliosis With a Novel, Seminested Real-Time Polymerase Chain Reaction Targeting the 5S-23S Intergenic Spacer Region: Clinical Features, Histopathology, and Immunophenotype in 44 Patients

Podbićanin-Ziburt

Falk

Metze

et al. 2021

The American Journal of Dermatopathology

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:Lyme borreliosis (LB) is the most common tick-borne infection in Europe and North America. Polymerase chain reaction (PCR) is an important tool to confirm the diagnosis, but not always successful, especially when organisms are sparse. We developed a novel, seminested real-time PCR assay [target: 5S-23S intergenic spacer region (IGS)] and compared it with 3 well-established conventional PCR assays (IGS/OspA/real-time IGS) on 596 formalin-fixed, paraffin-embedded routine skin biopsies. The seminested real-time assay identified 46 cases of borreliosis while 25, 27, and 38 were identified by the 3 other assays, respectively (P 0.01, P 0.02, and P 0.42; significance P < 0.05). Clinicopathologic and immunophenotypic analysis of PCR-positive cases revealed 38 erythema migrans (EM), 6 Borrelia lymphocytomas, and 2 acrodermatitis chronica atrophicans (ACA). In the 44 PCR-confirmed cases, plasma cells were present in only a third of EM cases. By contrast, CD123-positive plasmacytoid dendritic cells were common (74%) and therefore are unlikely to be helpful in the differential diagnosis between EM and tumid lupus erythematosus. A loss of CD34 in a third of all LB specimens limits its diagnostic value in the differential diagnosis with morphea. Interstitial macrophages were common in cutaneous LB (42/43) forming interstitial granulomas in a third of all cases, and 3/38 EM, 3/6 Borrelia lymphocytomas, and 1/2 ACA were only identified by the new seminested real-time assay, suggesting that it is especially helpful in confirming the diagnosis of Borrelia lymphocytoma.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

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Diagnosis of Lyme Borreliosis With a Novel, Seminested Real-Time Polymerase Chain Reaction Targeting the 5S-23S Intergenic Spacer Region: Clinical Features, Histopathology, and Immunophenotype in 44 Patients

Podbićanin-Ziburt

Falk

Metze

et al. 2021

The American Journal of Dermatopathology

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“…To further verify the identification, a groEL Borrelia burgdorferi sensu lato specific real-time PCR was designed, with minor modification of the primers described by Chiappa et al . [ 13 ]. The amplified sequence enables differentiation among various Borrelia species.…”

Section: Case Presentationmentioning

confidence: 99%

Mycotic abdominal aortic aneurysm caused by Borrelia afzelii: a case report

et al. 2022

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Background Inflammatory aneurysms and mycotic aneurysms make up a minority of abdominal aortic aneurysms. Mainly autoimmune mechanisms are proposed in the pathogenesis of inflammatory aneurysms, and it is not routine to check for infectious agents as disease culprits. Case presentation A 58-year-old European male with complaints of abdominal and back pain for 8 weeks was admitted after a semi-urgent computed tomography scan revealed an 85 mm inflammatory abdominal aortic aneurysm. The patient had normal vital signs, slightly elevated inflammatory markers, and mild anemia on admission. Clinical examination revealed a tender pulsating mass in his abdomen. His clinical condition was interpreted as impending rupture and urgent repair of the aneurysm was deemed necessary. Due to the patient’s relatively young age and aneurysm neck morphology, open aortic repair was preferred. Preoperatively, the aneurysm appeared inflamed, with fibrous wall thickening and perianeurysmal adhesions. Aneurysm wall biopsies were sent to histopathological and microbiological diagnostics. Routine cultures were negative, but 16S rRNA gene real-time polymerase chain reaction was positive and Borrelia afzelii was identified by DNA sequencing of the polymerase chain reaction product. B. afzelii was also identified by sequencing the polymerase chain reaction product of a Borrelia-specific groEL target. Immunoglobulin G and M anti-Borrelia antibodies were present on serological analysis. Histopathological analysis displayed loss of normal aortic wall structure and diffuse infiltration of lymphocytes and plasma cells. The patient had an uneventful recovery and was discharged after 1 week to a regional rehabilitation facility. Though the patient fares clinically well and inflammatory markers had normalized, antimicrobial treatment with doxycycline continues at 3 months follow-up due to remaining radiologic signs of inflammation. Conclusions Borrelia infection in the setting of acute aortic pathology is a rare entity. To our knowledge, this is the first case report to demonstrate a mycotic abdominal aortic aneurysm as a rare manifestation of Lyme disease. Aortic wall biopsies and real-time polymerase chain reaction analysis of the specimen were essential for accurate diagnosis. This finding may contribute to the understanding of the etiology of inflammatory aneurysmal disease and abdominal aneurysms in general.

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“…bacteria at the species level have been developed. Some of these methods include qualitative PCR protocols based on multi-locus sequence typing (MLST; e.g., [23]), nested PCR (e.g., [24][25][26]), amplicon sequencing for species determination (e.g., [27]), and qPCR systems based on species-specific probes [28][29][30][31]. However, the described approaches are often characterized by low sensitivity and/or low specificity, leading to possible false negative or false positive results [15,32,33].…”

Section: Introductionmentioning

confidence: 99%

A Novel High Discriminatory Protocol for the Detection of Borrelia afzelii, Borrelia burgdorferi Sensu Stricto and Borrelia garinii in Ticks

et al. 2022

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Bacteria of the Borrelia burgdorferi sensu lato complex are the causative agents of Lyme borreliosis (LB). Even if the conventional diagnosis of LB does not rely on the species itself, an accurate species identification within the complex will provide a deepened epidemiological scenario, a better diagnosis leading to a more targeted therapeutic approach, as well as promote the general public’s awareness. A comparative genomics approach based on the 210 Borrelia spp. genomes available in 2019 were used to set up three species-specific PCR protocols, able to detect and provide species typing of Borrelia afzelii, Borrelia burgdorferi sensu stricto (s.s.) and Borrelia garinii, the three most common and important human pathogenic Lyme Borrelia species in Europe. The species-specificity of these protocols was confirmed on previously identified B. afzelii, B. burgdorferi s.s. and B. garinii specimens detected in Ixodes ricinus samples. In addition, the protocols were validated on 120 DNA samples from ticks collected in Sweden, showing 88% accuracy, 100% precision, 72% sensitivity and 100% specificity. The proposed approach represents an innovative tool in epidemiological studies focused on B. burgdorferi s.l. occurrence in ticks, and future studies could suggest its helpfulness in routine diagnostic tests for health care.

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Development of a PCR for Borrelia burgdorferi sensu lato, targeted on the groEL gene

Cited by 5 publications

References 26 publications

Diagnosis of Lyme Borreliosis With a Novel, Seminested Real-Time Polymerase Chain Reaction Targeting the 5S-23S Intergenic Spacer Region: Clinical Features, Histopathology, and Immunophenotype in 44 Patients

Diagnosis of Lyme Borreliosis With a Novel, Seminested Real-Time Polymerase Chain Reaction Targeting the 5S-23S Intergenic Spacer Region: Clinical Features, Histopathology, and Immunophenotype in 44 Patients

Mycotic abdominal aortic aneurysm caused by Borrelia afzelii: a case report

A Novel High Discriminatory Protocol for the Detection of Borrelia afzelii, Borrelia burgdorferi Sensu Stricto and Borrelia garinii in Ticks

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