RESEARCH ARTICLE MCITY mBIO Changes in the Eye Microbiota Associated with Contact Lens Wearing (D Hakdong Shin,a Kenneth Price, a, b Luong Albert, a Jack Dodick, Lisa Park, b Maria Gloria Dominguez-Belloa Division of Translational Medicine and Department of Ophthalmology. New York University School of Medicine, New York, New York, USA lentified as a risk factor for the development of eye conditions such as giant papil lary conjunctivitis and keratitis. We hypothesized that wearing contact lenses is associated with changes in the ocular microbi- ota. We compared the bacterial communities of the conjunctiva and skin under the eye from 58 subjects and analyzed samples from 20 subjects(9 lens wearers and 11 non-lens wearers)taken at 3 time points using a 16S rRNA gene-based sequencing tech- nique(V4 region; Illumina MiSeq). We found that using anesthetic eye drops before sampling decreases the detected ocular mi- crobiota diversity Compared to those from non-lens wearers, dry conjunctival swabs from lens wearers had more variable and skin-like bacterial community structures(Uni Frac; P value <0.001), with higher abundances of Methylobacterium, Lactoba cillus, Acinetobacter, and Pseudomonas and lower abundances of Haemophilus, Streptococcus, Staphylococcus, and Corynebacte rium(linear discriminant analysis[LDA] score = >3.0). The results indicate that wearing contact lenses alters the microbial g structure of the ocular conjunctiva, making it more similar to that of the skin microbiota. Further research is needed to deter mine whether the microbiome structure provides less protection from ocular infections. IMPORTANCE As in other body sites(i.e, the gut, skin, and mouth), the eye has a normal community of bacteria which are ex pected to confer resistance that provides protection from invaders. However, the eye microbiome has been largely neglected and t is relevant to eye health and understanding eye diseases and to discovery of its functions. This report of a baseline study shows differences in the eye microbiome of contact lens wearers in relation to those of non-lens wearers and has the potential to help future studies explore novel insights into a possible role of the microbiome in the increased risk for eye infections in contact lens wearers. eceived 9 February 2016 Accepted 25 February 2016 Published 22 March 2016 Citation Shin H, Price K, Albert L, Dodick J, Park L Dominguez-Bello MG 2016. Changes in the eye microbiota associated with contact lens wearing. mBio 7(2): e00198-16 doi 10.128/mBo0019816. Editor Janet K Jansson, Pacific Northwest National Laboratory Copyright o 2016 Shin et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license Address correspondence to Maria G Dominguez-Bello, Maria. Dominguez-Belloenyumcorg This article is a direct contribution from a Fellow of the American Academy of Microbiology. External solicited reviewers: Jonathan Eisen, UC Davis Genome Center, Anthony St eger, NIH/NEE Jack A Gilbert, Argonne National Laborator n culture-dependent studies, over 50% of swabs from the con- supporting the hypothesis of microbial involvement and interaction junctiva showed growth of skin-like bacteria(mostly coagulase- with host conjunctival epithelium. negative staphylococci, Propionibacterium, and Corynebacterium While several studies have used culture-dependent approaches (1). Ocular bacterial communities have been studied using to demonstrate bacterial contamination of contact lenses(13, 14) culture-dependent methods(2, 3)and, more recently, with 16s little is known about the impact of contact lenses on the structure rRNA gene sequencing in healthy subjects(4)and in people with and function of the microbiota on the ocular surface. In this work, such as Pseudomonas, Bradyrhizobium, Acinetobacter, Brevundi. we compare the microbiota of the ocular surface of lens wearers eye diseases(5, 6). Using sequencing methods, additional bacteria, we using 16s rRNA gene sequencing monas, Aquabacterium, Sphingomonas, Streptococcus, Strepto- surveys phyta, Methylobacterium, Enhydrobacter, Bacillus, and Ralstonia spp, were detected(4-6). As in other body sites, the ocular mi- RESULTS robiota is expected to play a defensive role against colonization of We obtained 7,010,096 sequences(paired end; Phred = Q20)with pathogens in the eye(7). Despite being important in ophthalmol- an average of 21, 569 reads per sample, yielding 11,750 operating ogy, the eye microbiome has been largely neglected, and its func- taxonomic units(OTUs)(11, 700 OTUs without singletons)(see tions remain unknown Table SI in the supplemental material) In the United States, over 30 million people wear contact lenses, The bacterial alpha diversity of the conjunctiva was signifi nearly one-third of the -100 million worldwide(8). Wearing contact cantly higher in subjects sampled at the laboratory without anes- lenses has been identified as a risk factor for eye conditions such as thetic than was seen in the ophthalmologic practice, which used an giant papillary conjunctivitis(9)and keratitis(10-12). The conjunc- anesthetic eye drop (whole-tree phylogenetic diversity [PD] tiva of patients with vernal keratoconjunctivitis(inflammation of the number of observed species; P value =<0.001 [nonparametric eye that involves both the cornea and conjunctiva)showed increased Students t test))(see Fig. SIA in the supplemental material). US- Toll-like receptor 4(TLR-4)levels in relation to those of controls(7), ing an anesthetic eye drop significantly altered microbial commu March/April 2016 Volume 7 Issue 2 e00198-16 msio° mbio. asm. orgChanges in the Eye Microbiota Associated with Contact Lens Wearing Hakdong Shin,a Kenneth Price,a,b Luong Albert,a Jack Dodick,b Lisa Park,b Maria Gloria Dominguez-Belloa Division of Translational Medicinea and Department of Ophthalmology,b New York University School of Medicine, New York, New York, USA ABSTRACT Wearing contact lenses has been identified as a risk factor for the development of eye conditions such as giant papil￾lary conjunctivitis and keratitis. We hypothesized that wearing contact lenses is associated with changes in the ocular microbi￾ota. We compared the bacterial communities of the conjunctiva and skin under the eye from 58 subjects and analyzed samples from 20 subjects (9 lens wearers and 11 non-lens wearers) taken at 3 time points using a 16S rRNA gene-based sequencing tech￾nique (V4 region; Illumina MiSeq). We found that using anesthetic eye drops before sampling decreases the detected ocular mi￾crobiota diversity. Compared to those from non-lens wearers, dry conjunctival swabs from lens wearers had more variable and skin-like bacterial community structures (UniFrac; P value <0.001), with higher abundances of Methylobacterium, Lactoba￾cillus, Acinetobacter, and Pseudomonas and lower abundances of Haemophilus, Streptococcus, Staphylococcus, and Corynebacte￾rium (linear discriminant analysis [LDA] score >3.0). The results indicate that wearing contact lenses alters the microbial structure of the ocular conjunctiva, making it more similar to that of the skin microbiota. Further research is needed to deter￾mine whether the microbiome structure provides less protection from ocular infections. IMPORTANCE As in other body sites (i.e., the gut, skin, and mouth), the eye has a normal community of bacteria which are ex￾pected to confer resistance that provides protection from invaders. However, the eye microbiome has been largely neglected and is relevant to eye health and understanding eye diseases and to discovery of its functions. This report of a baseline study shows differences in the eye microbiome of contact lens wearers in relation to those of non-lens wearers and has the potential to help future studies explore novel insights into a possible role of the microbiome in the increased risk for eye infections in contact lens wearers. Received 9 February 2016 Accepted 25 February 2016 Published 22 March 2016 Citation Shin H, Price K, Albert L, Dodick J, Park L, Dominguez-Bello MG. 2016. Changes in the eye microbiota associated with contact lens wearing. mBio 7(2):e00198-16. doi: 10.1128/mBio.00198-16. Editor Janet K. Jansson, Pacific Northwest National Laboratory Copyright © 2016 Shin et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license. Address correspondence to Maria G. Dominguez-Bello, Maria.Dominguez-Bello@nyumc.org. This article is a direct contribution from a Fellow of the American Academy of Microbiology. External solicited reviewers: Jonathan Eisen, UC Davis Genome Center; Anthony St. Leger, NIH/NEI; Jack A. Gilbert, Argonne National Laboratory. I n culture-dependent studies, over 50% of swabs from the con￾junctiva showed growth of skin-like bacteria (mostly coagulase￾negative staphylococci, Propionibacterium, and Corynebacterium) (1). Ocular bacterial communities have been studied using culture-dependent methods (2, 3) and, more recently, with 16S rRNA gene sequencing in healthy subjects (4) and in people with eye diseases (5, 6). Using sequencing methods, additional bacteria, such as Pseudomonas, Bradyrhizobium, Acinetobacter, Brevundi￾monas, Aquabacterium, Sphingomonas, Streptococcus, Strepto￾phyta, Methylobacterium, Enhydrobacter, Bacillus, and Ralstonia spp., were detected (4–6). As in other body sites, the ocular mi￾crobiota is expected to play a defensive role against colonization of pathogens in the eye (7). Despite being important in ophthalmol￾ogy, the eye microbiome has been largely neglected, and its func￾tions remain unknown. In the United States, over 30 million people wear contact lenses, nearly one-third of the ~100 million worldwide (8). Wearing contact lenses has been identified as a risk factor for eye conditions such as giant papillary conjunctivitis (9) and keratitis (10–12). The conjunc￾tiva of patients with vernal keratoconjunctivitis (inflammation of the eye that involves both the cornea and conjunctiva) showed increased Toll-like receptor 4 (TLR-4) levels in relation to those of controls (7), supporting the hypothesis of microbial involvement and interaction with host conjunctival epithelium. While several studies have used culture-dependent approaches to demonstrate bacterial contamination of contact lenses (13, 14), little is known about the impact of contact lenses on the structure and function of the microbiota on the ocular surface. In this work, we compare the microbiota of the ocular surface of lens wearers with that of non-lens wearers using 16S rRNA gene sequencing surveys. RESULTS We obtained 7,010,096 sequences (paired end; Phred Q20) with an average of 21,569 reads per sample, yielding 11,750 operating taxonomic units (OTUs) (11,700 OTUs without singletons) (see Table S1 in the supplemental material). The bacterial alpha diversity of the conjunctiva was signifi- cantly higher in subjects sampled at the laboratory without anes￾thetic than was seen in the ophthalmologic practice, which used an anesthetic eye drop (whole-tree phylogenetic diversity [PD], number of observed species; P value 0.001 [nonparametric Student’s t test]) (see Fig. S1A in the supplemental material). Us￾ing an anesthetic eye drop significantly altered microbial commu￾RESEARCH ARTICLE crossmark March/April 2016 Volume 7 Issue 2 e00198-16 ® mbio.asm.org 1 mbio.asm.org on June 29, 2016 - Published by mbio.asm.org Downloaded from