Journal
CLINICAL INFECTIOUS DISEASES
Volume 70, Issue 11, Pages 2405-2412Publisher
OXFORD UNIV PRESS INC
DOI: 10.1093/cid/ciz655
Keywords
blood culture-negative endocarditis; laser capture microdissection; human RNA bait-depletion; Moraxella osloensis
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Funding
- IHU Mediterranee Infection, Agence Nationale de la Recherche [ANR-13JSV6-0004]
- French government under the Investissements d'avenir program [10-IAHU-03]
- Region Provence-Alpes-Cote d'Azur
- European funding FEDER PRIMI
- French National Research Agency through the Investments for the Future program (France-BioImaging) [ANR-10-INBS-04]
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Background. Etiological diagnosis is a key to therapeutic adaptation and improved prognosis, particularly for infections such as endocarditis. In blood culture-negative endocarditis (BCNE), 22% of cases remain undiagnosed despite an updated comprehensive syndromic approach. This prompted us to develop a new diagnostic approach. Methods. Eleven valves from 10 BCNE patients were analyzed using a method that combines human RNA bait-depletion with phi29 DNA polymerase-based multiple displacement amplification and shotgun DNA sequencing. An additional case in which a microbe was serendipitously visualized by immunofluorescence was analyzed using the same method, but after laser capture microdissection. Results. Background DNA prevented any diagnosis in cases analyzed without microdissection because the majority of sequences were contaminants. Moraxella sequences were dramatically enriched in the stained microdissected region of the additional case. A consensus genome sequence of 2.4 Mbp covering more than 94% of the Moraxella osloensis KSH reference genome was reconstructed with 234X average coverage. Several antibiotic-resistance genes were observed. Etiological diagnosis was confirmed using Western blot and specific polymerase chain reaction with sequencing on a different valve sample. Conclusions. Microdissection could be a key to the metagenomic diagnosis of infectious diseases when a microbe is visualized but remains unidentified despite an updated optimal approach. Moraxella osloensis should be tested in blood culture-negative endocarditis.
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