Shin et al ACKNOWLEDGMENTS 18. Szczotka-Flynn LB, Pearlman E, Ghannoum M. 2010. Microbi We thank Denni Catalano for her technical support in preparation of tamination of contact lenses, lens care solutions, and their access samples for sequencing. We also thank Keith A. Martin teraturereviewEyeContactLens36:116-129.http://dx.doi.org/ 10.1097/CL0b0l3e3l8ld20ca 19. McDermott AM. 2013. Antimicrobial compounds in tears. Exp Eye Res g FUNDING INFORMATION l17:53-61.htp:dx. doi org10.1016/exer2013.0701 20. Stapleton F, willcox MD, Fleming CM, Hickson S, Sweeney DF, This work, including DNA sequencing and the efforts of Jack Dodick and Holden BA. 1995. Changes to the ocular biota with time in extended-and sa Park, was funded by the research to Prevent Blindness Foundation laily-wear disposable contact lens use. Infect Immun 63: 4501-4505 21. Larkin DF, Leeming JP. 1991. Quantitative alterations of the commensal REFERENCES eyebacteriaincontactlenswearEye(lond)5:70-74.http://dx.doiorg/o 1. Willcox MD. 2013. Characterization of the normal microbiota of th 0.1038/eye.1991.14. ocularsurfaceExpEyereS117:99-105.http://dx.doi.org/10.1016/22.ErdoganH,KemalM,TokerMi,TopalkaraA,Bakiciz.2002.efFectof Jexer.2013.06.00 ent-replacement contact lenses on normal conjunctival flora. CLAO 2. Locatelli Cl, Kwitko S, Simonetti AB. 2002. Conjunctival endogenous 28 microbiota in patients submitted to cataract surgery. Braz J Microbiol 23. Leitch EC, Willcox MD. 1999. Lactoferrin increases the susceptibility of s. 34:203-209.http://dx.doiorg/10.1590/s1517-83822003000300004 Graham JE, Moore JE, Jiru X, Goodall EA, Dooley JS, Hayes VE, Dartt DA, dx.doi. org/10. 1076/cer. 9.. ycin. Curr Eye Res 19:12-19. epidermidis biofilms to lysozyme and Downes CS, Moore TC. 2007. Ocular pathogen or commensal: a PCR-based 24. Chen K-I, Hou C-H, Sun M-H, Lai C-c, Sun C-C, Hsiao udy of surface bacterial flora in normal and dry eyes. Invest Ophthalmol Vi dophthalmitis caused by Acinetobacter baumann g Si48:5616-5623.htp/ dxdoiorg/l0.1167/ows07-0588 linMicrobiol46:1148-1150.http://dx.doiorg/10.1128/jcm.01 4. Dong Q, Brule JM, lovieno A, Bates B, Garoutte A, Miller D 25. Willcox MD. 2007. Pseudomonas aeruginosa infection and infl KV, Gao X, Antonopoulos DA, Slepak Vz, Shestopalov during contact lens wear: a review. Optom Vis Sci 84: 273-2 二 Diversity of bacteria at healthy human conjunctiva. Invest dx. doi org/10. 1097/OPX.0b013e3180439c3e. VisSci52:5408-5413.http://dx.doiorg/10.1167/iovs.10-6 6. Lai c-C, Cheng A, Liu W-L, Tan C-K, Huang Y-T, Chung K-P, Lee s M-R, Hsueh P-R 2011. Infections caused by unusual Methylobacterium RL, Burton MI, Weinstock GM, Burr SE. 2014. The conjunctival micro- JClinMicrobiol49:3329-3331.http://dx.doi.org/10.1128/ ome in health and trachomatous disease: a case control study. Genome CM01241-11 Med6:99.http://dx.doi.org/10.1186/s13073-014-0099-x. 27. Costello EK, Lauber CL y M, Fierer N, Gordon JL, 6. Lee SH, Oh DH, Jung JY, Kim JC, Jeon CO. 2012. Comparative ocular 2009. Bacterial communit microbial communities in humans with and without blepharitis. Invest Op and Time. Science 32 1697.http://dx.doi.org/10.1126/ 7. Bonini S, Micera A, lovieno A, Lambiase A, Bonini S. 2005. Expression 28. Langille MG, Zaneveld J, Caporaso JG, McDonald D, Knights D, of Toll-like receptors in healthy and allergic conjunctiva. Ophthalmology JA, Clemente JC, Burkepile DE, Vega Thurber RL, Knight R, Beiko RG, 2:1528.e1-1528.e8.http://dx.doi.org/10.1016/j.ophtha.2005.04.009 Huttenhower C. 2013. Predictive functional profiling of microbial com- 8. Clayton-Jeter HD 2010. Looking good: Safe use and care of contact lens munities using 16S rRNA marker gene sequences. Nat Biotechnol 31: May2010).Fda,SilverSpringMd.http://www.fda.gov/downloads/ 4-821.http://dx.doi.org/10.1038/nbt.26 ForHealthProfessionals/Learning Activities/UCM213716pd 29. Caporaso ]G, Lauber CL, Walters WA, Berg-Lyons D, Huntley h, Fiere 9. Vengayil S, Vanathi M, Dada T, Kai S, Panda A. 2008. Filtering bleb N, Owens SM, Betley h, Fraser L, Bauer M, Gormley N, Gilbert JA, induced giant papillary conjunctivitis. Cont Lens Anterior Eye 31: 41-4 Smith G, Knight R. 2012. Ultra-high-throughput microbial community analysis on the Illumina HiSeq and Miseq platforms ISME J6: 1621-1624 10. Schein OD, Glynn RI, Poggio EC, on JM, Kenyon KR. 1989. The http://dx.doi.org/10.1038/ismej.2012.8 relative risk of ulcerative keratitis among users of daily-wear and 30. Salter S), Cox Ml, Turek EM, Calus ST, Cookson wO, Moffatt MF, extended-wear soft contact lenses. A case-control study. Microbial Kera Turner P, Parkhill ], Loman N, Walker Aw. 2014. Reagent and labo- titisStudyGroupNEngljMed321:773-778.http://dx.doi.org/10.1056/ ratory contamination can critically impact sequence-based microbiom NEM198909213211201 analysesBmcBiol12:87.http://dx.doiorg/10.1186/s12915-014-0087-z. 1. Lyczak JB, Cannon CL, Pier GB. 2000. Establishment of Pseudomonas 31. Caporaso JG, Kuczynski J, Stombaugh J, Bittinger K, Bushman FD, t Microbes infect Costello EK, Fierer N, Pena AG, Goodrich JK, Gordon Jl, Huttley Ga 2:1051-1060.http://dx.doiorg/10.1016/s1286-4579(00)01259-4 ley st, Knights D, Koenig JE, Ley re, Lozupone CA, McDonald D, 2. Fleiszig SM, Efron N. 1992. Microbial flora in eyes of current and former M ler J, Sevinsky JR, Turnbaugh Ph, Walters WA, Widmann J, Yatsunenko T, Zaneveld J, Knight R. 2010. QIIME drogelcontactlensesEyeContactLens31:124-126.httpd//dx.doi.org/3methods7:335-336.http://dx.doiorg/10.1038/nmeth..302gdata.nat 13. Iskeleli G, Bahar H, Eroglu E, Torun MM, Ozkan S. 2005. Microbial 10.1097/01.ICL0000141923.63458DF mproves sensitivity and speed of chimera detection. Bioinformatics 27 14.Go GN, Holden BA. 1997. Microbial contamination of hydrogel contact 33. Lozupone C, Hamady M, Knight R. 2006. UniFrac-an online tool for lensesJApplMicrobiol82:653-658.http://dx.doiorg/10.1111/j.1365 comparing microbial community diversity in a phylogenetic context. 2672.1997tb03598x. BmcBioinformatics7:371.http://dxdoiorg/10.1186/1471-2105-7-371 5. HMP Consortium. 2012. Structure, function and diversity of the healthy 34. Anderson M. 2001. A new method for non-parametric multivariate anal umanmicrobiomeNature486:207-214.http://dx.doi.org/10.1038/ ysis of variance. Aust Ecol 26: 32-46. 16. Bouslimani A, Porto C, Rath CM, Wang M, Guo Y, Gonzalez A, 35. Segata N, Izard J, Waldron L, Gevers D, Miropolsky L, Garrett wS, aturel1234 Huttenhower C 2011. Metagenomic biomarker discovery and expla Berg-Lyon D, Ackermann G, Moeller Christensen Gh, Nakatsuji T, tionGenomeBiol12:R60.http://dxdoi.org/10.1186/gb-2011-12-6-r60. hang L, Borkowski Aw, Meehan M], Dorrestein K, Gallo RL, Ban- 36. Kanehisa M, Goto S, Sato Y, Kawashima M, Furumichi M, Tanabe M. night R, Alexandrov T, Dorrestein PC. 2015. Molecula 2014. Data, information, knowledge and principle: back to metabolism in of the human skin surface in 3D. Proc Natl Acad sciU S Kegg.NucleicAcidsRes42:D199-d205.http://dx.doiorg/10.1093/nar/ 17. Keay L Stapleton E, Schein oorgo, 073/pnas. 142440911 gy of contact lens- 37. Knights D, Kuczynski J, Charlson ES, Zaneveld J, Mozer MC, Colima related inflammation and microbial keratitis: a 20-year perspective. RG, Bushman FD, Knight R, Kelley ST. 2011. Bayesian community-wid Lens33:346-353.http://dx.doi.org/10.1097/ culture-independent microbial source tracking. Nat Methods 8: 761-763. ICL. 0b013e318157c49d http://dxdoiorg/10.1038/nmeth.1650 6 mBio mbio. asm. org March/April 2016 Volume 7 Issue 2 800198-16ACKNOWLEDGMENTS We thank Denni Catalano for her technical support in preparation of samples for sequencing. We also thank Keith A. Martinez II for language editing. FUNDING INFORMATION This work, including DNA sequencing and the efforts of Jack Dodick and Lisa Park, was funded by the Research to Prevent Blindness Foundation. REFERENCES 1. Willcox MD. 2013. Characterization of the normal microbiota of the ocular surface. Exp Eye Res 117:99 –105. http://dx.doi.org/10.1016/ j.exer.2013.06.003. 2. Locatelli CI, Kwitko S, Simonetti AB. 2002. Conjunctival endogenous microbiota in patients submitted to cataract surgery. Braz J Microbiol 34:203–209. http://dx.doi.org/10.1590/S1517-83822003000300004. 3. Graham JE, Moore JE, Jiru X, Goodall EA, Dooley JS, Hayes VE, Dartt DA, Downes CS, Moore TC. 2007. Ocular pathogen or commensal: a PCR-based study of surface bacterial flora in normal and dry eyes. Invest Ophthalmol Vis Sci 48:5616–5623. http://dx.doi.org/10.1167/iovs.07-0588. 4. Dong Q, Brulc JM, Iovieno A, Bates B, Garoutte A, Miller D, Revanna KV, Gao X, Antonopoulos DA, Slepak VZ, Shestopalov VI. 2011. Diversity of bacteria at healthy human conjunctiva. Invest Ophthalmol Vis Sci 52:5408 –5413. http://dx.doi.org/10.1167/iovs.10-6939. 5. Zhou Y, Holland MJ, Makalo P, Joof H, Roberts CH, Mabey DC, Bailey RL, Burton MJ, Weinstock GM, Burr SE. 2014. The conjunctival micro￾biome in health and trachomatous disease: a case control study. Genome Med 6:99. http://dx.doi.org/10.1186/s13073-014-0099-x. 6. Lee SH, Oh DH, Jung JY, Kim JC, Jeon CO. 2012. Comparative ocular microbial communities in humans with and without blepharitis. Invest Oph￾thalmol Vis Sci 53:5585–5593. http://dx.doi.org/10.1167/iovs.12-9922. 7. Bonini S, Micera A, Iovieno A, Lambiase A, Bonini S. 2005. Expression of Toll-like receptors in healthy and allergic conjunctiva. Ophthalmology 112:1528.e1–1528.e8. http://dx.doi.org/10.1016/j.ophtha.2005.04.009. 8. Clayton-Jeter HD. 2010. Looking good: Safe use and care of contact lenses (May 2010). FDA, Silver Spring, MD. http://www.fda.gov/downloads/ ForHealthProfessionals/LearningActivities/UCM213716.pdf. 9. Vengayil S, Vanathi M, Dada T, Kai S, Panda A. 2008. Filtering bleb￾induced giant papillary conjunctivitis. Cont Lens Anterior Eye 31:41– 43. http://dx.doi.org/10.1016/j.clae.2007.07.005. 10. Schein OD, Glynn RJ, Poggio EC, Seddon JM, Kenyon KR. 1989. The relative risk of ulcerative keratitis among users of daily-wear and extended-wear soft contact lenses. A case-control study. Microbial Kera￾titis Study Group. N Engl J Med 321:773–778. http://dx.doi.org/10.1056/ NEJM198909213211201. 11. Lyczak JB, Cannon CL, Pier GB. 2000. Establishment of Pseudomonas aeruginosa infection: lessons from a versatile opportunist. Microbes Infect 2:1051–1060. http://dx.doi.org/10.1016/S1286-4579(00)01259-4. 12. Fleiszig SM, Efron N. 1992. Microbial flora in eyes of current and former contact lens wearers. J Clin Microbiol 30:1156 –1161. 13. Iskeleli G, Bahar H, Eroglu E, Torun MM, Ozkan S. 2005. Microbial changes in conjunctival flora with 30-day continuous-wear silicone hy￾drogel contact lenses. Eye Contact Lens 31:124 –126. http://dx.doi.org/ 10.1097/01.ICL.0000141923.63458.DF. 14. Gopinathan U, Stapleton F, Sharma S, Willcox MD, Sweeney DF, Rao GN, Holden BA. 1997. Microbial contamination of hydrogel contact lenses. J Appl Microbiol 82:653– 658. http://dx.doi.org/10.1111/j.1365 -2672.1997.tb03598.x. 15. HMP Consortium. 2012. Structure, function and diversity of the healthy human microbiome. Nature 486:207–214. http://dx.doi.org/10.1038/ nature11234. 16. Bouslimani A, Porto C, Rath CM, Wang M, Guo Y, Gonzalez A, Berg-Lyon D, Ackermann G, Moeller Christensen GJ, Nakatsuji T, Zhang L, Borkowski AW, Meehan MJ, Dorrestein K, Gallo RL, Ban￾deira N, Knight R, Alexandrov T, Dorrestein PC. 2015. Molecular cartography of the human skin surface in 3D. Proc Natl Acad SciUSA 112:E2120 –E2129. http://dx.doi.org/10.1073/pnas.1424409112. 17. Keay L, Stapleton F, Schein O. 2007. Epidemiology of contact lens￾related inflammation and microbial keratitis: a 20-year perspective. Eye Contact Lens 33:346 –353. http://dx.doi.org/10.1097/ ICL.0b013e318157c49d. 18. Szczotka-Flynn LB, Pearlman E, Ghannoum M. 2010. Microbial con￾tamination of contact lenses, lens care solutions, and their accessories: a literature review. Eye Contact Lens 36:116 –129. http://dx.doi.org/ 10.1097/ICL.0b013e3181d20cae. 19. McDermott AM. 2013. Antimicrobial compounds in tears. Exp Eye Res 117:53– 61. http://dx.doi.org/10.1016/j.exer.2013.07.014. 20. Stapleton F, Willcox MD, Fleming CM, Hickson S, Sweeney DF, Holden BA. 1995. Changes to the ocular biota with time in extended- and daily-wear disposable contact lens use. Infect Immun 63:4501– 4505. 21. Larkin DF, Leeming JP. 1991. Quantitative alterations of the commensal eye bacteria in contact lens wear. Eye (Lond) 5:70 –74. http://dx.doi.org/ 10.1038/eye.1991.14. 22. Erdog˘an H, Kemal M, Toker MI, Topalkara A, Bakici Z. 2002. Effect of frequent-replacement contact lenses on normal conjunctival flora. CLAO J 28:94 –95. 23. Leitch EC, Willcox MD. 1999. Lactoferrin increases the susceptibility of S. epidermidis biofilms to lysozyme and vancomycin. Curr Eye Res 19:12–19. http://dx.doi.org/10.1076/ceyr.19.1.12.5342. 24. Chen K-J, Hou C-H, Sun M-H, Lai C-C, Sun C-C, Hsiao C-H. 2008. Endophthalmitis caused by Acinetobacter baumannii: report of two cases. J Clin Microbiol 46:1148–1150. http://dx.doi.org/10.1128/JCM.01604-07. 25. Willcox MD. 2007. Pseudomonas aeruginosa infection and inflammation during contact lens wear: a review. Optom Vis Sci 84:273–278. http:// dx.doi.org/10.1097/OPX.0b013e3180439c3e. 26. Lai C-C, Cheng A, Liu W-L, Tan C-K, Huang Y-T, Chung K-P, Lee M-R, Hsueh P-R. 2011. Infections caused by unusual Methylobacterium species. J Clin Microbiol 49:3329 –3331. http://dx.doi.org/10.1128/ JCM.01241-11. 27. Costello EK, Lauber CL, Hamady M, Fierer N, Gordon JI, Knight R. 2009. Bacterial community variation in human body habitats across space and Time. Science 326:1694 –1697. http://dx.doi.org/10.1126/ science.1177486. 28. Langille MG, Zaneveld J, Caporaso JG, McDonald D, Knights D, Reyes JA, Clemente JC, Burkepile DE, Vega Thurber RL, Knight R, Beiko RG, Huttenhower C. 2013. Predictive functional profiling of microbial com￾munities using 16S rRNA marker gene sequences. Nat Biotechnol 31: 814 – 821. http://dx.doi.org/10.1038/nbt.2676. 29. Caporaso JG, Lauber CL, Walters WA, Berg-Lyons D, Huntley J, Fierer N, Owens SM, Betley J, Fraser L, Bauer M, Gormley N, Gilbert JA, Smith G, Knight R. 2012. Ultra-high-throughput microbial community analysis on the Illumina HiSeq and MiSeq platforms. ISME J 6:1621–1624. http://dx.doi.org/10.1038/ismej.2012.8. 30. Salter SJ, Cox MJ, Turek EM, Calus ST, Cookson WO, Moffatt MF, Turner P, Parkhill J, Loman NJ, Walker AW. 2014. Reagent and labo￾ratory contamination can critically impact sequence-based microbiome analyses. BMC Biol 12:87. http://dx.doi.org/10.1186/s12915-014-0087-z. 31. Caporaso JG, Kuczynski J, Stombaugh J, Bittinger K, Bushman FD, Costello EK, Fierer N, Peña AG, Goodrich JK, Gordon JI, Huttley GA, Kelley ST, Knights D, Koenig JE, Ley RE, Lozupone CA, McDonald D, Muegge BD, Pirrung M, Reeder J, Sevinsky JR, Turnbaugh PJ, Walters WA, Widmann J, Yatsunenko T, Zaneveld J, Knight R. 2010. QIIME allows analysis of high-throughput community sequencing data. Nat Methods 7:335–336. http://dx.doi.org/10.1038/nmeth.f.303. 32. Edgar RC, Haas BJ, Clemente JC, Quince C, Knight R. 2011. UCHIME improves sensitivity and speed of chimera detection. Bioinformatics 27: 2194 –2200. http://dx.doi.org/10.1093/bioinformatics/btr381. 33. Lozupone C, Hamady M, Knight R. 2006. UniFrac—an online tool for comparing microbial community diversity in a phylogenetic context. BMC Bioinformatics 7:371. http://dx.doi.org/10.1186/1471-2105-7-371. 34. Anderson M. 2001. A new method for non-parametric multivariate anal￾ysis of variance. Aust Ecol 26:32– 46. 35. Segata N, Izard J, Waldron L, Gevers D, Miropolsky L, Garrett WS, Huttenhower C. 2011. Metagenomic biomarker discovery and explana￾tion. Genome Biol 12:R60. http://dx.doi.org/10.1186/gb-2011-12-6-r60. 36. Kanehisa M, Goto S, Sato Y, Kawashima M, Furumichi M, Tanabe M. 2014. Data, information, knowledge and principle: back to metabolism in KEGG. Nucleic Acids Res 42:D199–D205. http://dx.doi.org/10.1093/nar/ gkt1076. 37. Knights D, Kuczynski J, Charlson ES, Zaneveld J, Mozer MC, Collman RG, Bushman FD, Knight R, Kelley ST. 2011. Bayesian community-wide culture-independent microbial source tracking. Nat Methods 8:761–763. http://dx.doi.org/10.1038/nmeth.1650. Shin et al. 6 ® mbio.asm.org March/April 2016 Volume 7 Issue 2 e00198-16 mbio.asm.org on June 29, 2016 - Published by mbio.asm.org Downloaded from