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New real-time PCR-based method for Kingella kingae DNA detection: application to samples collected from 89 children with acute arthritis.

ABSTRACT: Inoculation of blood culture vials with joint fluid samples has revealed the important pathogenic role of Kingella kingae in pediatric arthritis. However, recent studies based on broad-range 16S ribosomal DNA PCR and real-time PCR without a probe suggest that conventional methods remain suboptimal. We developed a new real-time PCR method with a probe that is highly specific for K. kingae and applied it to joint fluid samples collected from 89 children with suspected arthritis admitted to our institution during a 2-year period. Real-time PCR was also applied to blood samples obtained before surgery and to joint drainage fluid samples obtained during several days after surgery. Thirty-six (40%) of the 89 cases of suspected septic arthritis had positive culture. Staphylococcus aureus was the main isolate (n = 19/36, 53%), followed by K. kingae (n = 7/36, 19%). Specific real-time PCR identified K. kingae in 24 of the 53 culture-negative cases. Thus, K. kingae was present in 31 (52%) of the 60 documented cases, making it the leading pathogen. Real-time PCR on all 15 blood DNA extracts from patients with K. kingae infection was negative, demonstrating that joint fluid positivity did not result from DNA circulating in blood. Real-time PCR amplification of drainage fluid samples showed that the pathogen could be detected for up to 6 days after antibiotic initiation. K. kingae real-time PCR applied to DNA extracted from joint fluid samples, but not from blood samples, markedly improved the etiological diagnosis of septic arthritis in children. Retrospective diagnosis is feasible for up to 6 days after treatment initiation.

SUBMITTER: Ilharreborde B

PROVIDER: S-EPMC2691089 | biostudies-literature | 2009 Jun

REPOSITORIES: biostudies-literature

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New real-time PCR-based method for Kingella kingae DNA detection: application to samples collected from 89 children with acute arthritis.

Ilharreborde Brice B Bidet Philippe P Lorrot Mathie M Even Julien J Mariani-Kurkdjian Patricia P Liguori Sandrine S Vitoux Christine C Lefevre Yann Y Doit Catherine C Fitoussi Franck F Penneçot Georges G Bingen Edouard E Mazda Keyvan K Bonacorsi Stéphane S

Journal of clinical microbiology 20090415 6

Inoculation of blood culture vials with joint fluid samples has revealed the important pathogenic role of Kingella kingae in pediatric arthritis. However, recent studies based on broad-range 16S ribosomal DNA PCR and real-time PCR without a probe suggest that conventional methods remain suboptimal. We developed a new real-time PCR method with a probe that is highly specific for K. kingae and applied it to joint fluid samples collected from 89 children with suspected arthritis admitted to our ins ...[more]

PMID: 19369442

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Project description:Kingella kingae is an important cause of invasive infections in young children from Western countries. Although increasing reports indicate that this organism is the leading agent of bone and joint infections in early childhood, data on K. kingae infections from resource-limited settings are scarce, and none has yet been reported in Africa. We herein report the diagnostic and epidemiological investigations of the first case of K. kingae arthritis identified in a child from sub-Saharan Africa. A 5-year-old Cameroonian boy presented with a sudden painful limp which appeared in the course of a mild rhinopharyngitis. He lived in Cameroon where he had been vaccinated with BCG at birth and moved to France for holidays 4 days before consultation. There was no history of trauma and he did not have any underlying medical condition. Upon admission, he had a temperature of 36.7°C, and clinical examination revealed right-sided knee tenderness and effusion that was confirmed by ultrasound imaging. Laboratory results showed a white blood cell count of 5,700 cells/mm3, C-reactive protein level of 174 mg/L, and platelet count of 495,000 cells/mm3. He underwent an arthrocentesis and was immediately given intravenous amoxicillin-clavulanate. Conventional cultures from blood samples and synovial fluids were negative. Polymerase chain reaction (PCR) assay targeting the broad-range 16S rRNA gene and real-time quantitative PCR assays targeting Mycobacterium species were negative. Surprisingly, real-time PCR assays targeting the cpn60, rtxA, and rtxB genes of K. kingae were positive. Multicolor fluorescence in situ hybridization specific for K. kingae identified the presence of numerous coccobacilli located within the synovial fluid. Finally, multilocus sequence typing analysis performed on deoxyribonucleic acid directly extracted from joint fluid disclosed a novel K. kingae sequence-type complex. This case report demonstrates that K. kingae may be considered as a potential cause of septic arthritis in children living in sub-Saharan Africa, and hence the burden of K. kingae infection may be not limited to the Western countries. Further studies are required to determine the prevalence of K. kingae infection and carriage in Africa.

Dataset Information

New real-time PCR-based method for Kingella kingae DNA detection: application to samples collected from 89 children with acute arthritis.

Publications

New real-time PCR-based method for Kingella kingae DNA detection: application to samples collected from 89 children with acute arthritis.

Similar Datasets

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