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Clinical metagenomics of bone and joint infections: a proof of concept study.

ABSTRACT: Bone and joint infections (BJI) are severe infections that require a tailored and protracted antibiotic treatment. Yet, the diagnostic based on culturing samples lacks sensitivity, especially for hardly culturable bacteria. Metagenomic sequencing could potentially address those limitations. Here, we assessed the performances of metagenomic sequencing on 24 BJI samples for the identification of pathogens and the prediction of their antibiotic susceptibility. For monomicrobial samples in culture (n?=?8), the presence of the pathogen was confirmed by metagenomics in all cases. For polymicrobial samples (n?=?16), 32/55 bacteria (58.2%) were found at the species level (and 41/55 [74.5%] at the genus level). Conversely, 273 bacteria not found in culture were identified, 182 being possible pathogens and 91 contaminants. A correct antibiotic susceptibility could be inferred in 94.1% and 76.5% cases for monomicrobial and polymicrobial samples, respectively. Altogether, we found that clinical metagenomics applied to BJI samples is a potential tool to support conventional culture.

SUBMITTER: Ruppe E

PROVIDER: S-EPMC5552814 | biostudies-literature | 2017 Aug

REPOSITORIES: biostudies-literature

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Clinical metagenomics of bone and joint infections: a proof of concept study.

Ruppé Etienne E Lazarevic Vladimir V Girard Myriam M Mouton William W Ferry Tristan T Laurent Frédéric F Schrenzel Jacques J

Scientific reports 20170810 1

Bone and joint infections (BJI) are severe infections that require a tailored and protracted antibiotic treatment. Yet, the diagnostic based on culturing samples lacks sensitivity, especially for hardly culturable bacteria. Metagenomic sequencing could potentially address those limitations. Here, we assessed the performances of metagenomic sequencing on 24 BJI samples for the identification of pathogens and the prediction of their antibiotic susceptibility. For monomicrobial samples in culture ( ...[more]

PMID: 28798333

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Project description:Clinical metagenomics is actively moving from research to clinical laboratories. It has the potential to change the microbial diagnosis of infectious diseases, especially when detection and identification of pathogens can be challenging, such as in prosthetic joint infection (PJI). The application of metagenomic sequencing to periprosthetic joint tissue (PJT) specimens is often challenged by low bacterial load in addition to high level of inhibitor and contaminant host DNA, limiting pathogen recovery. Shotgun-metagenomics (SMg) performed directly on positive blood culture bottles (BCBs) inoculated with PJT may be a convenient approach to overcome these obstacles. The aim was to test if it is possible to perform SMg on PJT inoculated into BCBs for pathogen identification in PJI diagnosis. Our study was conducted as a laboratory method development. For this purpose, spiked samples (positive controls), negative control and clinical tissue samples (positive BCBs) were included to get a comprehensive overview. We developed a method for preparation of bacterial DNA directly from PJT inoculated in BCBs. Samples were processed using MolYsis5 kit for removal of human DNA and DNA extracted with BiOstic kit. High DNA quantity/quality was obtained, and no inhibition was observed during the library preparation, allowing further sequencing process. DNA sequencing reads obtained from the BCBs, presented a low proportion of human reads (<1%) improving the sensitivity of bacterial detection. We detected a 19-fold increase in the number of reads mapping to human in a sample untreated with MolYsis5. Taxonomic classification of clinical samples identified a median of 96.08% (IQR, 93.85-97.07%; range 85.7-98.6%) bacterial reads. Shotgun-metagenomics results were consistent with the results from a conventional BCB culture method, validating our approach. Overall, we demonstrated a proof of concept that it is possible to perform SMg directly on BCBs inoculated with PJT, with potential of pathogen identification in PJI diagnosis. We consider this a first step in research efforts needed to face the challenges presented in PJI diagnoses.

Project description:IntroductionSubtalar joint (STJ) dysfunction can contribute to movement disturbances. Vibration energy with color Doppler imaging (VECDI) may be useful for detecting STJ stiffness changes.Objectives(1) Support proof-of-concept that VECDI could detect STJ stiffness differences; (2) Establish STJ stiffness range in asymptomatic volunteers; (3) Examine relationships between STJ stiffness and foot mobility; and (4) Assess VECDI precision and reliability for examining STJ stiffness.MethodsAfter establishing cadaveric testing model proof-of-concept, STJ stiffness (threshold units, ΔTU), ankle complex passive range-of-motion (PROM) and midfoot-width-difference (MFWDiff) data were collected in 28 asymptomatic subjects in vivo. Three reliability measurements were collected per variable; Rater-1 collected on all subjects and rater-2 on the first ten subjects. Subjects were classified into three STJ stiffness groups.ResultsCadaveric VECDI measurement intra-rater reliability was 0.80. A significantly lower STJ ΔTU (p = .002) and ankle complex PROM (p < .001) was observed during the screw fixation versus normal condition. A fair correlation (r = 0.660) was observed between cadaveric ΔTU and ankle complex PROM. In vivo VECDI measurements demonstrated good intra-rater (0.76-0.84) versus poor inter-rater (-3.11) reliability. Significant positive correlations were found between STJ stiffness and both dorsum (r = .440) and posterior (r = .390) PROM. MFWDiff exhibited poor relationships with stiffness (r = .103) and either dorsum (r = .256) or posterior (r = .301) PROM. STJ stiffness ranged from 2.33 to 7.50 ΔTUs, categorizing subjects' STJ stiffness as increased (n = 6), normal (n = 15), or decreased (n = 7). Significant ANOVA main effects for classification were found based on ΔTU (p< .001), dorsum PROM (p = .017), and posterior PROM (p = .036). Post-hoc tests revealed significant: (1) ΔTU differences between all stiffness groups (p < .001); (2) dorsum PROM differences between the increased versus normal (p = .044) and decreased (p = .017) stiffness groups; and (3) posterior PROM differences between the increased versus decreased stiffness groups (p = .044). A good relationship was found between STJ stiffness and dorsum PROM in the increased stiffness group (r = .853) versus poor, nonsignificant relationships in the normal (r = -.042) or decreased stiffness (r = -.014) groups.ConclusionPROM may not clinically explain all aspects of joint mobility. Joint VECDI stiffness assessment should be considered as a complimentary measurement technique.

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Clinical metagenomics of bone and joint infections: a proof of concept study.

Publications

Clinical metagenomics of bone and joint infections: a proof of concept study.

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