Title |
Microbial Lineages in Sarcoidosis. A Metagenomic Analysis Tailored for Low–Microbial Content Samples
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Published in |
American Journal of Respiratory & Critical Care Medicine, August 2017
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DOI | 10.1164/rccm.201705-0891oc |
Pubmed ID | |
Authors |
Erik L Clarke, Abigail P Lauder, Casey E Hofstaedter, Young Hwang, Ayannah S Fitzgerald, Ize Imai, Wojciech Biernat, Bartłomiej Rękawiecki, Hanna Majewska, Anna Dubaniewicz, Leslie A Litzky, Michael D Feldman, Kyle Bittinger, Milton D Rossman, Karen C Patterson, Frederic D Bushman, Ronald G Collman |
Abstract |
The etiology of sarcoidosis is unknown, but microbial agents are suspected as triggers. We sought to identify bacterial, fungal or viral lineages in specimens from sarcoidosis patients enriched relative to controls using metagenomic DNA sequencing. Since DNA from environmental contamination contributes disproportionately to samples with low authentic microbial content, we developed improved methods for filtering environmental contamination. We analyzed specimens from sarcoidosis subjects (n=93), non-sarcoidosis control subjects (n=72) and various environmental controls (n=150). Sarcoidosis specimens consisted of two independent sets of formalin-fixed, paraffin-embedded lymph node biopsies, bronchoalveolar lavage (BAL), Kveim reagent, and fresh granulomatous spleen from a sarcoidosis patient. All specimens were analyzed by bacterial 16S and fungal ITS rRNA gene sequencing. In addition, BAL was analyzed by shotgun sequencing of fractions enriched for viral particles, and Kveim and spleen were subjected to whole-genome shotgun sequencing. In one tissue set, fungi in the Cladosporiaceae family were enriched in sarcoidosis compared to non-sarcoidosis tissues; in the other tissue set, we detected enrichment of several bacterial lineages in sarcoidosis, but not Cladosporiaceae. BAL showed limited enrichment of Aspergillus fungi. Several microbial lineages were detected in Kveim and spleen, including Cladosporium. No microbial lineage was enriched in more than one sample type after correction for multiple comparisons. Metagenomic sequencing revealed enrichment of microbes in single types of sarcoidosis samples, but limited concordance across sample types. Statistical analysis accounting for environmental contamination was essential to avoiding false positives. |
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Geographical breakdown
Country | Count | As % |
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United States | 5 | 31% |
Mexico | 2 | 13% |
Colombia | 1 | 6% |
Ireland | 1 | 6% |
Saudi Arabia | 1 | 6% |
Unknown | 6 | 38% |
Demographic breakdown
Type | Count | As % |
---|---|---|
Members of the public | 8 | 50% |
Scientists | 5 | 31% |
Practitioners (doctors, other healthcare professionals) | 2 | 13% |
Science communicators (journalists, bloggers, editors) | 1 | 6% |
Mendeley readers
Geographical breakdown
Country | Count | As % |
---|---|---|
Unknown | 58 | 100% |
Demographic breakdown
Readers by professional status | Count | As % |
---|---|---|
Researcher | 15 | 26% |
Student > Ph. D. Student | 10 | 17% |
Student > Doctoral Student | 6 | 10% |
Other | 5 | 9% |
Student > Master | 3 | 5% |
Other | 4 | 7% |
Unknown | 15 | 26% |
Readers by discipline | Count | As % |
---|---|---|
Medicine and Dentistry | 13 | 22% |
Biochemistry, Genetics and Molecular Biology | 7 | 12% |
Agricultural and Biological Sciences | 6 | 10% |
Immunology and Microbiology | 6 | 10% |
Chemistry | 2 | 3% |
Other | 5 | 9% |
Unknown | 19 | 33% |