Highly sensitive real-time PCR for specific detection and quantification of Coxiella burnetii

Klee, Silke R.; Tyczka, Judith; Ellerbrok, Heinz; Franz, Tatjana; Linke, Sonja; Baljer, G.; Appel, Bernd

doi:10.1186/1471-2180-6-2

Cited by 235 publications

(120 citation statements)

References 27 publications

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“…C. burnetii was reported to productively proliferate in human alveolar macrophages as well (18). Due to the limited number of bovine alveolar macrophages available for this study and the laborious replating assay, only quantitation of genome equivalents (GE) was applied to assess C. burnetii replication in this cell type (37). While NMI-inoculated alveolar macrophages showed a clear increase in GE numbers from day 1 to day 14 of culture, GE numbers in NMII-inoculated cultures increased slightly between days 3 and 7 (P ϭ 0.041).…”

Section: Discussionmentioning

confidence: 99%

“…For quantification by PCR, cell suspension was additionally inactivated by boiling (95°C, 20 min). Replication rates were calculated by quantitation of the icd gene by absolute quantitative real-time PCR (37). C T values (where C T is threshold cycle) of technical duplicates varied by less than 0.51 and were used to calculate genome equivalents (GE) considering values obtained with an entrained icd-harboring plasmid standard.…”

Section: Methodsmentioning

confidence: 99%

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Coxiella burnetii Infects Primary Bovine Macrophages and Limits Their Host Cell Response

et al. 2016

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Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Coxiella burnetii Infects Primary Bovine Macrophages and Limits Their Host Cell Response

et al. 2016

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“…The majority of PCR assays for C. burnetii have targeted the insertion sequence element IS1111 (6)(7)(8)(9)(10). As a mobile genetic element, there is the potential for IS1111 to move from one organism type to another, theoretically resulting in loss of specificity (11).…”

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confidence: 99%

A Case of Q Fever Prosthetic Joint Infection and Description of an Assay for Detection of Coxiella burnetii

et al. 2013

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We present the first published case of Coxiella burnetii prosthetic joint infection. Diagnosis was established with PCR and culture of periprosthetic tissue and synovial fluid (and serology). A novel PCR assay is described herein. Q fever should be considered in patients with prosthetic joint infection without an identified pathogen. J oint replacement is a life-enhancing procedure performed on hundreds of thousands of patients in the United States each year. While prosthetic joint infection (PJI) occurs in only approximately 2% of patients following knee arthroplasty (1), it is a devastating complication. Accurate microbiologic diagnosis of PJI is important for directing appropriate management. There is no identified pathogen in approximately 7% of PJI cases, frequently as a result of antecedent antimicrobial therapy, but occasionally due to an inability to detect a pathogen by standard laboratory methods (2). Coxiella burnetii is not an organism that would traditionally be considered in a case of culture-negative PJI. We describe here the first published case of PJI caused by C. burnetii.The causative agent of the zoonosis Q fever, C. burnetii, is a small, obligate intracellular Gram-negative bacterium. Acute and chronic types of Q fever are classically diagnosed based on compatible clinical illnesses, supported by serologic results (3). Isolation of C. burnetii in cell culture is laborious, requires specially trained staff, and poses a safety risk to staff (4, 5), confining C. burnetii culture to a small number of research and public health laboratories and limiting medical practitioners' access to this technique for diagnostics.Q fever PCR offers a safe alternative to culture for direct detection of C. burnetii in clinical specimens. The majority of PCR assays for C. burnetii have targeted the insertion sequence element IS1111 (6-10). As a mobile genetic element, there is the potential for IS1111 to move from one organism type to another, theoretically resulting in loss of specificity (11). The diagnosis of C. burnetii PJI in the case reported here was made using a novel real-time PCR assay (described below) targeting the single-copy housekeeping gene target, shikimate dehydrogenase (aroE) of C. burnetii. CASE REPORTA 56-year-old male presented to our institution with new increasing right knee pain and swelling around a previously revised total knee arthroplasty (TKA).He first developed right knee pain after patellectomy for a comminuted patellar fracture at 27 years of age. He underwent repeated arthroscopies without improvement; right TKA was performed at age 44. This was complicated by early postoperative deep vein thrombosis (DVT). He developed increasing right knee pain, swelling, and stiffness approximately 12 weeks after arthroplasty. Subcutaneous corticosteroid and lidocaine injections were administered around the branches of the saphenous nerve on a monthly basis for presumed neuropathic pain.At the age of 53, he presented to our facility with a 5-day history of general malaise, mild nonproductive c...

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“…1 allowed for the sensitivity of the assay to be increased, because this a multi-copy gene (7-110 copies) 25 . DNA sequences generated in the present study confirm that C. burnetii is circulating in goats and sheep from some herds of Valledupar, Colombia.…”

Section: Discussionmentioning

confidence: 99%

Coxiella burnetii infection in sheep and goats: a public risk health, Colombia

Contreras¹,

González²,

Álvarez³

et al. 2018

Infect

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Objective. The aim of this study was to provide molecular evidence of C. burnetii in sheep and goats from some herds of Valledupar, Cesar, Colombia. Materials and methods. Fifteen herds of sheep and goats were chosen by convenience to investigate the infection by C. burnetii, during March and April of 2013. 328 female goats and 66 sheep from 15 herds were included in this study. Milk from ewes and vaginal mucus samples from goats were analyzed by Polymerase Chain Reaction for DNA detection of transposase gene (IS1111) of C. burnetii. Results. DNA of C. burnetii in 6% (4/66) of sheep's milk and 0.6% (2/328) vaginal mucus from goats was found. 13% (2/15) of the herds had at least one infected animal. Discussion. Our findings suggest the circulation of C. burnetii in sheep and goats from some herds of Valledupar, Colombia, and it highlights the possibility of occurrence of infections in humans and animals. Conclusions. The detection of C. burnetii in sheep milk could represent a public health risk factor for people who consuming raw milk, cheeses or people associated to agriculture and livestock handling. Further studies are necessary to evaluate other routes such as tick's bite, feces, milk from goats and vaginal mucus from sheep of this region of Colombia.

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Highly sensitive real-time PCR for specific detection and quantification of Coxiella burnetii

Cited by 235 publications

References 27 publications

Coxiella burnetii Infects Primary Bovine Macrophages and Limits Their Host Cell Response

Coxiella burnetii Infects Primary Bovine Macrophages and Limits Their Host Cell Response

A Case of Q Fever Prosthetic Joint Infection and Description of an Assay for Detection of Coxiella burnetii

Coxiella burnetii infection in sheep and goats: a public risk health, Colombia

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