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Fluorescence In Situ Hybridization (FISH) and Peptide Nucleic Acid Probe-Based FISH for Diagnosis of Q Fever Endocarditis and Vascular Infections.

ABSTRACT: Endocarditis and vascular infections are common manifestations of persistent localized infection due to Coxiella burnetii, and recently, fluorescence in situ hybridization (FISH) was proposed as an alternative tool for their diagnosis. In this study, we evaluated the efficiency of FISH in a series of valve and vascular samples infected by C. burnetii We tested 23 C. burnetii-positive valves and thrombus samples obtained from patients with Q fever endocarditis. Seven aneurysms and thrombus specimens were retrieved from patients with Q fever vascular infections. Samples were analyzed by culture, immunochemistry, and FISH with oligonucleotide and PNA probes targeting C. burnetii-specific 16S rRNA sequences. The immunohistochemical analysis was positive for five (17%) samples with significantly more copies of C. burnetii DNA than the negative ones (P = 0.02). FISH was positive for 13 (43%) samples and presented 43% and 40% sensitivity compared to that for quantitative PCR (qPCR) and culture, respectively. PNA FISH detected C. burnetii in 18 (60%) samples and presented 60% and 55% sensitivity compared to that for qPCR and culture, respectively. Immunohistochemistry had 38% and 28% sensitivity compared to that for FISH and PNA FISH, respectively. Samples found positive by both immunohistochemistry and PNA FISH contained significantly more copies of C. burnetii DNA than the negative ones (P = 0.03). Finally, PNA FISH was more sensitive than FISH (60% versus 43%, respectively) for the detection of C. burnetii We provide evidence that PNA FISH and FISH are important assays for the diagnosis of C. burnetii endocarditis and vascular infections.

SUBMITTER: Prudent E

PROVIDER: S-EPMC6113452 | biostudies-literature | 2018 Sep

REPOSITORIES: biostudies-literature

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Fluorescence In Situ Hybridization (FISH) and Peptide Nucleic Acid Probe-Based FISH for Diagnosis of Q Fever Endocarditis and Vascular Infections.

Prudent Elsa E Lepidi Hubert H Angelakis Emmanouil E Raoult Didier D

Journal of clinical microbiology 20180827 9

Endocarditis and vascular infections are common manifestations of persistent localized infection due to Coxiella burnetii, and recently, fluorescence in situ hybridization (FISH) was proposed as an alternative tool for their diagnosis. In this study, we evaluated the efficiency of FISH in a series of valve and vascular samples infected by C. burnetii We tested 23 C. burnetii-positive valves and thrombus samples obtained from patients with Q fever endocarditis. Seven a ...[more]

PMID: 29899006

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Project description:BackgroundOur current understanding of biofilms indicates that these structures are typically composed of many different microbial species. However, the lack of reliable techniques for the discrimination of each population has meant that studies focusing on multi-species biofilms are scarce and typically generate qualitative rather than quantitative data.Methodology/principal findingsWe employ peptide nucleic acid fluorescence in situ hybridization (PNA FISH) methods to quantify and visualize mixed biofilm populations. As a case study, we present the characterization of Salmonella enterica/Listeria monocytogenes/Escherichia coli single, dual and tri-species biofilms in seven different support materials. Ex-situ, we were able to monitor quantitatively the populations of ∼56 mixed species biofilms up to 48 h, regardless of the support material. In situ, a correct quantification remained more elusive, but a qualitative understanding of biofilm structure and composition is clearly possible by confocal laser scanning microscopy (CLSM) at least up to 192 h. Combining the data obtained from PNA FISH/CLSM with data from other established techniques and from calculated microbial parameters, we were able to develop a model for this tri-species biofilm. The higher growth rate and exopolymer production ability of E. coli probably led this microorganism to outcompete the other two [average cell numbers (cells/cm(2)) for 48 h biofilm: E. coli 2,1 × 10(8) (± 2,4 × 10(7)); L. monocytogenes 6,8 × 10(7) (± 9,4 × 10(6)); and S. enterica 1,4 × 10(6) (± 4,1 × 10(5))]. This overgrowth was confirmed by CSLM, with two well-defined layers being easily identified: the top one with E. coli, and the bottom one with mixed regions of L. monocytogenes and S. enterica.SignificanceWhile PNA FISH has been described previously for the qualitative study of biofilm populations, the present investigation demonstrates that it can also be used for the accurate quantification and spatial distribution of species in polymicrobial communities. Thus, it facilitates the understanding of interspecies interactions and how these are affected by changes in the surrounding environment.

Dataset Information

Fluorescence In Situ Hybridization (FISH) and Peptide Nucleic Acid Probe-Based FISH for Diagnosis of Q Fever Endocarditis and Vascular Infections.

Publications

Fluorescence <i>In Situ</i> Hybridization (FISH) and Peptide Nucleic Acid Probe-Based FISH for Diagnosis of Q Fever Endocarditis and Vascular Infections.

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