Illuminating uveitis: metagenomic deep sequencing identifies common and rare pathogens
Thuy Doan, Michael R. Wilson, Emily D. Crawford, Eric D. Chow, Lillian M. Khan, Kristeene A. Knopp, Brian D. O ’ Donovan, Dongxiang Xia, Jill K. Hacker, Jay M. Stewart, John A. Gonzales, Nisha R. Acharya and Joseph L. DeRisi

Genome Medicine, 2016

Abstract: Background: Ocular infections remain a major cause of blindness and morbidity worldwide. While prognosis is dependent on the timing and accuracy of diagnosis, the etiology remains elusive in ~ 50% of presumed infectious uveitis cases. The objective of this study is to determine if unbiased metagenomic deep sequencing (MDS) can accurately detect pathogens in intraocular fluid samples of patients with uveitis. Methods: This is a proof-of-concept study, in which intraocular fluid samples were obtained from 5 subjects with known diagnoses, and one subject with bilateral chronic uveitis without a known etiology. Samples were subjected to MDS, and results were compared with conventional diagnostic tests. Pathogens were identified using a rapid computational pipeline to analyze the non-host sequences obtained from MDS. Results: Unbiased MDS of intraocular fluid produced results concordant with known diagnoses in subjects with (n=4) and without (n=1) uveitis. Samples known to be positive for Cryptococcus neoformans, Toxoplasma gondii, and herpes simplex virus 1 (HSV-1) by tests performed at a Clinical Laboratory Improvement Amendments (CLIA)-certified laboratory, were correctly identified with MDS. Rubella virus (RV) was identified in one case of chronic bilateral idiopathic uveitis. The subject’s strain was most closely related to a German RV strain isolated in 1992, one year before he developed a fever and rash while living in Germany. The pattern and the number of viral identified mutations present in the patient’s strain were consistent with long-term viral replication in the eye. Conclusions: MDS can identify fungi, parasites, and DNA and RNA viruses in minute volumes of intraocular fluid samples. The identification of chronic intraocular RV infection highlights the eye’s role as a long-term pathogen reservoir, which has implications for virus eradication and emerging global epidemics.