- Email: [email protected]
Changes in Antibiotic Resistance Patterns of Conjunctival Flora Due to Repeated Use of Topical Antibiotics after Intravitreal Injection
Changes in Antibiotic Resistance Patterns of Conjunctival Flora Due to Repeated Use of Topical Antibiotics after Intravitreal Injection Eugene Milder, MD, James Vander, MD, Chirag Shah, MD, MPH, Sunir Garg, MD Objective: To determine the effect of repeated intermittent use of topical antibiotics after intravitreal injections on conjunctival bacterial flora and antibiotic resistance. Design: Cross-sectional case-control study. Participants and Controls: A total of 80 eyes of 40 patients were enrolled (40 study eyes, 40 control eyes). Patients were enrolled with unilateral exudative age-related macular degeneration who had received at least 3 prior intravitreal injections with use of postinjection topical antibiotics. Patients had received an average of 7 (range, 3–13) intravitreal injections before enrollment. Methods: At the time of enrollment, the inferior fornix of the treated eye was swept with a culture swab before use of povidone iodine; the inferior fornix of the fellow eye was also cultured and served as a control. The culture and sensitivity data from the study and control eyes were analyzed. Main Outcome Measures: The rate of antibiotic resistance among the conjunctival bacterial flora of the study eyes and control eyes. Results: A total of 80 eyes of 40 patients were enrolled in the study; 29 patients used trimethoprim/ polymyxin B drops, and 11 patients used fluoroquinolone drops after each injection. A total of 58 bacterial colonies were isolated from 50 eyes. There were no significant differences in bacterial species or culture positivity rates between study and control eyes. Coagulase-negative staphylococcus accounted for 41 of the 58 bacterial colonies (71%). There was a 63.6% resistance rate to fluoroquinolones among study eyes compared with 32.1% among control eyes (P ⬍ 0.05). In the subset of 11 study eyes using fluoroquinolone drops for 4 days after injection, there was an 87.5% resistance rate compared with 25.0% in matched control eyes (P ⫽ 0.04). There was no significant difference in trimethoprim resistance rates between study and control eyes: Four of 14 study eyes (28.6%) showed resistance compared with 5 of 18 control eyes (27.7%) (P ⫽ 1.0). Conclusions: Use of fluoroquinolone drops after intravitreal injection leads to increased rates of resistance among conjunctival flora. Repeated use of topical fluoroquinolones after intravitreal injections may have a detrimental effect on eye health by breeding resistance in the bacterial flora. Financial Disclosure(s): The author(s) have no proprietary or commercial interest in any materials discussed in this article. Ophthalmology 2012;119:1420 –1424 © 2012 by the American Academy of Ophthalmology.
The number of intravitreal injections performed over the past decade has increased dramatically. Endophthalmitis, although rare,1,2 is a dreaded complication of this procedure and can be catastrophic.2,3 How best to minimize the risk of endophthalmitis is not precisely known.4 Postprocedure topical antibiotics have long been the standard of care for ocular surgery, and this practice has been carried over to intravitreal injections. However, it is unclear whether postinjection antibiotics have any benefit, and recent evidence suggests that their use does not reduce the risk of endophthalmitis.5 Indiscriminate use of topical antibiotics after these procedures may have the detrimental effect of breeding antibiotic resistance in the conjunctival bacterial flora.6 The purpose of this study was to examine the changes in conjunctival bacterial flora and antibiotic resistance patterns in patients who have had at least 3 intravitreal injections
1420
© 2012 by the American Academy of Ophthalmology Published by Elsevier Inc.
followed by postinjection use of topical antibiotics in 1 eye only, with the fellow eye serving as a control.
Patients and Methods Overview Wills Eye Institutional Review Board approval was obtained for this study, and all subjects gave informed consent at the time of enrollment. This was a single-center, cross-sectional, case-control study. Patients with exudative age-related macular degeneration (AMD) undergoing treatment with intravitreal bevacizumab or ranibizumab (Genentech, South San Francisco, CA) were eligible for this study if they were aged ⱖ50 years, received ⱖ3 intravitreal injections of ranibizumab or bevacizumab for exudative AMD in the study eye only, and had used the same topical antibiotic after each injection, either a fluoroquinolone or trimethoprim/polyISSN 0161-6420/12/$–see front matter doi:10.1016/j.ophtha.2012.01.016
Milder et al 䡠 Antibiotic Resistance Patterns of Conjunctival Flora Table 1. Demographic and Clinical Characteristics of the Study Population Average Age Male Female Average no. of injections Fluoroquinolone users Polymyxin B/trimethoprim users
76.2yrs (Range, 59–87) 27 (67.5%) 13 (32.5%) 7.0 (range, 3–13) 11 (27.5%) 29 (72.5%)
myxin B. Exclusion criteria included use of systemic antibiotics, use of other topical ocular antibiotics in either eye, or any ocular surgery or infection within 6 months of study enrollment. A standard technique for intravitreal injection was followed for all the patients throughout their course of intravitreal treatments. All eyes were prepped with 5% povidone iodine before intravitreal injection, and all eyes were treated with a single drop of fluoroquinolone (ofloxacin or moxifloxacin) immediately afterward. Patients were then instructed to use antibiotic drops on the treated eye 4 times per day for 4 days. The study included patients using fluoroquinolone drops (ofloxacin or moxifloxacin) or trimethoprim/polymyxin B drops. It was verified that the patients had used the same postinjection drops for each of their prior treatments and that there was adherence to the prescribed eyedrop regimen. The age, gender, number of intravitreal injections given, and type of antibiotics used were recorded.
Culture Collection Technique At the time of enrollment, a culture swab (BBL CultureSwab collection and transport system; Becton Dickinson and Company, Sparks, MD) of the inferior fornix was taken from the study eye and the contralateral (control) eye after a single drop of proparacaine was placed in both eyes and before placement of 5% povidone iodine solution in the study eye. Care was taken not to contact the eyelids or eyelashes with the swab. The swabs were sent for bacterial culture and antibiotic sensitivity testing. The cultures were incubated in the Phoenix automated microbiology system (Becton Dickinson and Company). All bacterial isolates were tested for sensitivity to 15 different antibiotics, including moxifloxacin, gatifloxacin, trimethoprim, and vancomycin. On the basis of automated readings of the machine, not every bacterial isolate had a definitive sensitivity result to every antibiotic. Calculations of rates of resistance were based on definitive readings of sensitivity or resistance. Descriptive statistics were calculated for means and rates of case characteristics. Statistical analysis comparing groups of patients was performed using the 2-tailed Fisher exact test (StataCorp LP, College Station, TX).
Conjunctival Culture Results Fifty-eight bacterial cultures were isolated from 50 eyes, and the remaining 30 eyes showed no growth. The no growth rate was evenly split between study and control eyes (16 study eyes, 14 control eyes; P ⫽ 0.8). The most frequent bacteria identified was coagulase-negative staphylococcus (CNS), comprising 41 of 58 (70.7%) of the cultures. Six isolates of Staphylococcus aureus were identified, including one of methicillin-resistant S. aureus. Other bacteria identified included streptococcus mitis and coryneform species. The distribution of bacterial species between study and control eyes was balanced. Eighteen of 41 CNS isolates were from study eyes compared with 23 from control eyes (P ⫽ 0.38).
Antibiotic Resistance Rates Resistance to fluoroquinolones was defined as intermediate or full resistance to both moxifloxacin and gatifloxacin. Moxifloxacin resistance and sensitivity were defined as a minimum inhibitory concentration (MIC) ⱖ2.0 g/ml and ⱕ0.5 g/ml, respectively. Gatifloxacin resistance and sensitivity were defined as an MIC ⱖ8.0 g/ml and ⱕ2.0 g/ml, respectively. Among the control eyes, 9 of 28 eyes (32.1%) showed fluoroquinolone resistance compared with 14 of 22 eyes (63.6%) showing resistance among all study eyes (P ⫽ 0.04). In the subset of study eyes in patients who also used fluoroquinolones for 4 days after injection, 7 of 8 eyes (87.5%) showed fluoroquinolone resistance compared with 2 of 8 (25.0%) in the matched control eyes (P ⫽ 0.04). Figure 1 illustrates the rates of fluoroquinolone resistance among the different sets of eyes. Thirty-two bacterial isolates were grown from trimethoprim/ polymyxin B users: 14 from study eyes and 18 from control eyes. Trimethoprim resistance and sensitivity were defined as an MIC ⱖ16.0 g/ml and ⱕ8.0 g/ml, respectively. There was no significant difference in trimethoprim resistance rates between study and control eyes: Four of 14 study eyes (28.6%) showed resistance compared with 5 of 18 control eyes (27.7%) (P ⫽ 1.0). There was a single CNS isolate from the study eye of a trimethoprim/polymyxin B user that showed resistance to vancomycin. All other bacterial isolates in this study were sensitive to vancomycin. There was no cephalosporin resistance identified among the CNS isolates. The streptococcus mitis isolate was grown from a study eye, and the methicillin-resistant S. aureus colony came from a control eye. Figure 2 summarizes the resistance rates to other antimicrobials for the CNS isolates in both the study and control eyes.
Discussion The objective of intravitreal injection is to deliver pharmaceuticals into the vitreous cavity while minimizing the risk % Resistant
% Sensive
Results 12.5
Cohort Characteristics Eighty eyes (40 patients) were enrolled in the study. The average patient age was 76.2 years. A total of 27 of 40 patients (67.5%) were male, and 13 patients (32.5%) were female. Patients had received an average of 7 intravitreal injections for exudative AMD before enrollment, with a range of 3 to 13. Eleven of 40 patients used fluoroquinolone drops after the injections; 29 patients used trimethoprim/polymyxin B drops. Table 1 summarizes the baseline demographics of the study population.
36.4 67.9
75 87.5
63.6 32.1 All Study Eyes (n=22)
All Control Eyes (n=28)
25 Study Eyes using Fluoroquinolones (n=8)
Control Eyes using Fluoroquinolones (n=8)
Figure 1. Fluoroquinolone resistance rates among the different sets of eyes.
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Ophthalmology Volume 119, Number 7, July 2012 100% 90% 80% 70% 60% 50% 40%
Sensive
30%
Resistant
20% 10% 0%
Figure 2. Antibiotic resistance rates. Resistance rates to other antimicrobials among study eyes and control eyes. CE ⫽ control eyes; SE ⫽ study eyes.
of complications, including bleeding, pain, vitreous or drug reflux, intraocular pressure elevation, and endophthalmitis.7–9 Techniques and recommendations for minimizing the risk of endophthalmitis vary, and recent evidence suggests that subtle differences in intravitreal injection technique do not alter the inherent risk of endophthalmitis with this procedure.2 There is widespread use of postinjection topical antibiotics with the assumption that their use reduces the risk of infection; however, there is evidence disputing this assumption.5 Some of the commonly used antibiotic drops, including moxifloxacin, ofloxacin, and trimethoprim/polymyxin B, which we studied, are approved by the Food and Drug Administration for the treatment of ocular surface infections. None have been shown to reduce the incidence of postprocedural endophthalmitis. When the currently available newer-generation fluoroquinolones became commercially available for ocular use, there was hope that their use would aid in the prevention of postprocedure endophthalmitis.10 Fluoroquinolones offer broad-spectrum antimicrobial coverage and good ocular penetration when used topically.11 However, studies have shown that endophthalmitis still occurs despite use of these agents.12,13 Of note, many patients receive topical antibiotic drops preoperatively for cataract surgery, but for many patients this is the only intraocular procedure they will ever have. In contrast, patients with wet exudative AMD receive treatment for years on an ongoing basis at regular intervals.14,15 Repeated, short, 4-day courses of antibiotics have the potential for selection of resistant bacterial strains. The short course might be enough to create a selection bias for resistance, whereas the duration might not be long enough for eradication. The repetition of antibiotic use every 4 to 6 weeks enhances this selection force further. This could partly explain why fluoroquinolone resistance is an emerging problem in ocular microbiology.16,17 Our patients demonstrated a baseline fluoroquinolone resistance rate of 32.1% in the control eye. This is comparable to results from similar studies of ocular bacterial flora.17–21 Remarkably, after an average of 7 intravitreal injections, the fluoroquinolone resistance rate was 63.6% among all study eyes and 87.5% among the subset of eyes
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exposed to topical fluoroquinolones for 4 days after each injection. This suggests that repeated exposure to fluoroquinolones, even just a single drop on the eye after each injection, can lead to increased rates of resistance. These results are similar to those found by Kim and Toma.6 Of note, there is a definite trend toward cross-resistance among fluoroquinolones. This phenomenon has been described by Hooper22 and was also found on ocular surface bacteria by Kim and Toma6 and among staphylococcal endophthalmitis isolates by Miller et al.23 Our patients were exposed only to ofloxacin and moxifloxacin, and yet high rates of resistance to both moxifloxacin and gatifloxacin were found. As described by Hooper,22 among CNS strains, a single mutation may be sufficient to confer resistance to the entire fluoroquinolone class of antimicrobials. The other topical antibiotic we examined was trimethoprim/polymyxin B. Resistance to trimethoprim among gram-positive specimens is less well understood, especially from ocular sources. Although frequently used for ocular surface infections and as endophthalmitis prophylaxis, there are little data on the use of these drugs.24,25 Results from studies on CNS strains isolated from cases of cystitis show a trend toward resistance, with increasing rates of resistance emerging after widespread use of trimethoprim.26,27 Our study did not show a similar trend toward resistance to trimethoprim in patients using ocular trimethoprim/polymyxin B. The overall resistance rate of 26% that we found is identical to that found by Kim and Toma19 in conjunctival isolates tested for resistance to trimethoprim/sulfamethoxazole.
Study Limitations Our study has several limitations. It was performed on a relatively small group of patients at a single center. The influence of community-acquired, drug-resistant bacterial strains is therefore possible. We did not exclude patients who have had other intraocular surgeries and who likely used topical antibiotics at that time. Patients were excluded if any such procedure had been performed within 6 months of study enrollment, but lingering changes to the ocular flora beyond 6 months after such a procedure is certainly possible. The same can be said for systemic antibiotic use, and the same 6-month rule was used for exclusion. The study eyes were also repeatedly exposed to 5% povidone iodine, which could alter the characteristics of the ocular surface flora. Our study group consisted of patients who had received an average of 7 intravitreal injections in 1 eye only. This enabled us to use the fellow eye as a control. This assumes that there is no significant transfer of conjunctival bacterial flora between eyes in a given subject. This assumption could be called into question because direct transfer of flora through eye-rubbing or gradual transfer via the continuous mucous membrane of the nasolacrimal system, nasal cavity, and nasopharynx is certainly possible. However, such a transfer of flora would have only served to undermine our results by reducing the difference in resistance rates between study and control eyes. Either this transfer of flora was not significant in our study population or, if it did occur,
Milder et al 䡠 Antibiotic Resistance Patterns of Conjunctival Flora the power of the selection bias to create resistant strains on the study eye was enough to overcome it. The strength of our study is that it examines a crosssection of patients being treated for exudative AMD in a clinical setting. We use the subject’s fellow eye as a control, isolating the repeated povidone iodine preparation, intravitreal injections, and use of topical antibiotics as the possible factors responsible for changes in ocular bacterial flora. By including patients who use both fluoroquinolone and trimethoprim/polymyxin B drops, we uncovered 3 separate sets of eyes in terms of fluoroquinolone exposure: the control eyes with no exposure, the set of eyes using trimethoprim/ polymyxin B drops that only get exposed to a single drop of fluoroquinolone immediately after each injection, and the set of eyes exposed to fluoroquinolones for 4 days after each injection. Our results showed a definite linear, statistically significant increase in resistance, suggesting a cause and effect relationship between fluoroquinolone exposure and the development of high rates of resistance. In conclusion, the results of our study demonstrate that a significant proportion of patients exposed to repeated short courses of topical fluoroquinolones after intravitreal injections for exudative AMD develop ocular surface bacteria that are resistant to fluoroquinolones. Additional studies are needed to provide more definitive conclusions, but this work raises concerns about the routine use of topical antibiotics for endophthalmitis prophylaxis after repeated intravitreal injections. Such a strategy may not reduce the risk of infection and could produce changes in the conjunctival flora that are potentially harmful by selecting for more antibiotic-resistant organisms.
References 1. Fintak DR, Shah GK, Blinder KJ, et al. Incidence of endophthalmitis related to intravitreal injection of bevacizumab and ranibizumab. Retina 2008;28:1395–9. 2. Shah CP, Garg SJ, Vander JF, et al, Post-Injection Endophthalmitis (PIE) Study Team. Outcomes and risk factors associated with endophthalmitis after intravitreal injection of antivascular endothelial growth factor agents. Ophthalmology 2011;118:2028 –34. 3. Jager RD, Aiello LP, Patel SC, Cunningham ET Jr. Risks of intravitreous injection: a comprehensive review. Retina 2004; 24:676 –98. 4. Pilli S, Kotsolis A, Spaide RF, et al. Endophthalmitis associated with intravitreal anti-vascular endothelial growth factor therapy injections in an office setting. Am J Ophthalmol 2008;145:879 – 82. 5. Bhavsar AR, Googe JM Jr, Stockdale CR, et al, Diabetic Retinopathy Clinical Research Network. Risk of endophthalmitis after intravitreal drug injection when topical antibiotics are not required: the Diabetic Retinopathy Clinical Research Network laser-ranibizumab-triamcinolone clinical trials. Arch Ophthalmol 2009;127:1581–3. 6. Kim SJ, Toma HS. Ophthalmic antibiotics and antimicrobial resistance: a randomized, controlled study of patients undergoing intravitreal injections. Ophthalmology 2011; 118:1358 – 63. 7. Knecht PB, Michels S, Sturm V, et al. Tunnelled versus straight intravitreal injection: intraocular pressure changes, vitreous reflux, and patient discomfort. Retina 2009;29:1175– 81.
8. Rodrigues EB, Grumann A Jr, Penha FM, et al. Effect of needle type and injection technique on pain level and vitreal reflux in intravitreal injection. J Ocul Pharmacol Ther 2011;27:197–203. 9. Maurice D. Review: practical issues in intravitreal drug delivery. J Ocul Pharmacol Ther 2001;17:393– 401. 10. Chang DF, Braga-Mele R, Mamalis N, et al, ASCRS Cataract Clinical Committee. Prophylaxis of postoperative endophthalmitis after cataract surgery: results of the 2007 ASCRS member survey. J Cataract Refract Surg 2007;33:1801–5. 11. Hariprasad SM, Blinder KJ, Shah GK, et al. Penetration pharmacokinetics of topically administered 0.5% moxifloxacin ophthalmic solution in human aqueous and vitreous. Arch Ophthalmol 2005;123:39 – 44. 12. Moshirfar M, Feiz V, Vitale AT, et al. Endophthalmitis after uncomplicated cataract surgery with the use of fourth-generation fluoroquinolones: a retrospective observational case series. Ophthalmology 2007;114:686–91. 13. Deramo VA, Lai JC, Fastenberg DM, Udell IJ. Acute endophthalmitis in eyes treated prophylactically with gatifloxacin and moxifloxacin. Am J Ophthalmol 2006;142:721–5. 14. Brown DM, Michels M, Kaiser PK, et al, ANCHOR Study Group. Ranibizumab versus verteporfin photodynamic therapy for neovascular age-related macular degeneration: two-year results of the ANCHOR Study. Ophthalmology 2009;116:57–65. 15. Rosenfeld PJ, Brown DM, Heier JS, et al, MARINA Study Group. Ranibizumab for neovascular age-related macular degeneration. N Engl J Med 2006;355:1419 –31. 16. Fintelmann RE, Hoskins EN, Lietman TM. Topical fluoroquinolone use as a risk factor for in vitro fluoroquinolone resistance in ocular cultures. Arch Ophthalmol 2011;129:399–402. 17. Park SH, Lim JA, Choi JS, et al. The resistance patterns of normal ocular bacterial flora to 4 fluoroquinolone antibiotics. Cornea 2009;28:68 –72. 18. Ta CN, Chang RT, Singh K, et al. Antibiotic resistance patterns of ocular bacterial flora: a prospective study of patients undergoing anterior segment surgery. Ophthalmology 2003; 110:1946 –51. 19. Kim SJ, Toma HS, Midha NK, et al. Antibiotic Resistance of Conjunctiva and Nasopharynx Evaluation Study: a prospective study of patients undergoing intravitreal injections. Ophthalmology 2010;117:2372– 8. 20. Chung JL, Seo KY, Yong DE, et al. Antibiotic susceptibility of conjunctival bacterial isolates from refractive surgery patients. Ophthalmology 2009;116:1067–74. 21. Moss JM, Sanislo SR, Ta CN. Antibiotic susceptibility patterns of ocular bacterial flora in patients undergoing intravitreal injections. Ophthalmology 2010;117:2141–5. 22. Hooper DC. Emerging mechanisms of fluoroquinolone resistance. Emerg Infect Dis 2001;7:337– 41. 23. Miller D, Flynn PM, Scott IU, et al. In vitro fluoroquinolone resistance in staphylococcal endophthalmitis isolates. Arch Ophthalmol 2006;124:479 – 83. 24. Asbell PA, Colby KA, Deng S, et al. Ocular TRUST: nationwide antimicrobial susceptibility patterns in ocular isolates. Am J Ophthalmol 2008;145:951– 8. 25. Price FW Jr, Dobbins K, Zeh W. Penetration of topically administered ofloxacin and trimethoprim into aqueous humor. J Ocul Pharmacol Ther 2002;18:445–53. 26. Araújo SM, Mourão TC, Oliveira JL, et al. Antimicrobial resistance of uropathogens in women with acute uncomplicated cystitis from primary care settings. Int Urol Nephrol 2011;43:461– 6. 27. Gupta K, Scholes D, Stamm WE. Increasing prevalence of antimicrobial resistance among uropathogens causing acute uncomplicated cystitis in women. JAMA 1999;281:736–8.
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Footnotes and Financial Disclosures Originally received: August 14, 2011. Final revision: December 9, 2011. Accepted: January 9, 2012. Available online: March 13, 2012.
Manuscript no. 2011-1222.
Presented at: the American Society of Retina Specialists, 29th Annual Meeting, August 2011, Boston, Massachusetts.
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Financial Disclosure(s): The author(s) have no proprietary or commercial interest in any materials discussed in this article. Funding: Retina Endowment Fund, Philadelphia, Pennsylvania. The funding organization had no role in the design or conduct of this research. Correspondence: James Vander, MD, 840 Walnut St, Suite 1020, Philadelphia, PA 19107. E-mail: [email protected].
The number of intravitreal injections performed over the past decade has increased dramatically. Endophthalmitis, although rare,1,2 is a dreaded complication of this procedure and can be catastrophic.2,3 How best to minimize the risk of endophthalmitis is not precisely known.4 Postprocedure topical antibiotics have long been the standard of care for ocular surgery, and this practice has been carried over to intravitreal injections. However, it is unclear whether postinjection antibiotics have any benefit, and recent evidence suggests that their use does not reduce the risk of endophthalmitis.5 Indiscriminate use of topical antibiotics after these procedures may have the detrimental effect of breeding antibiotic resistance in the conjunctival bacterial flora.6 The purpose of this study was to examine the changes in conjunctival bacterial flora and antibiotic resistance patterns in patients who have had at least 3 intravitreal injections
1420
© 2012 by the American Academy of Ophthalmology Published by Elsevier Inc.
followed by postinjection use of topical antibiotics in 1 eye only, with the fellow eye serving as a control.
Patients and Methods Overview Wills Eye Institutional Review Board approval was obtained for this study, and all subjects gave informed consent at the time of enrollment. This was a single-center, cross-sectional, case-control study. Patients with exudative age-related macular degeneration (AMD) undergoing treatment with intravitreal bevacizumab or ranibizumab (Genentech, South San Francisco, CA) were eligible for this study if they were aged ⱖ50 years, received ⱖ3 intravitreal injections of ranibizumab or bevacizumab for exudative AMD in the study eye only, and had used the same topical antibiotic after each injection, either a fluoroquinolone or trimethoprim/polyISSN 0161-6420/12/$–see front matter doi:10.1016/j.ophtha.2012.01.016
Milder et al 䡠 Antibiotic Resistance Patterns of Conjunctival Flora Table 1. Demographic and Clinical Characteristics of the Study Population Average Age Male Female Average no. of injections Fluoroquinolone users Polymyxin B/trimethoprim users
76.2yrs (Range, 59–87) 27 (67.5%) 13 (32.5%) 7.0 (range, 3–13) 11 (27.5%) 29 (72.5%)
myxin B. Exclusion criteria included use of systemic antibiotics, use of other topical ocular antibiotics in either eye, or any ocular surgery or infection within 6 months of study enrollment. A standard technique for intravitreal injection was followed for all the patients throughout their course of intravitreal treatments. All eyes were prepped with 5% povidone iodine before intravitreal injection, and all eyes were treated with a single drop of fluoroquinolone (ofloxacin or moxifloxacin) immediately afterward. Patients were then instructed to use antibiotic drops on the treated eye 4 times per day for 4 days. The study included patients using fluoroquinolone drops (ofloxacin or moxifloxacin) or trimethoprim/polymyxin B drops. It was verified that the patients had used the same postinjection drops for each of their prior treatments and that there was adherence to the prescribed eyedrop regimen. The age, gender, number of intravitreal injections given, and type of antibiotics used were recorded.
Culture Collection Technique At the time of enrollment, a culture swab (BBL CultureSwab collection and transport system; Becton Dickinson and Company, Sparks, MD) of the inferior fornix was taken from the study eye and the contralateral (control) eye after a single drop of proparacaine was placed in both eyes and before placement of 5% povidone iodine solution in the study eye. Care was taken not to contact the eyelids or eyelashes with the swab. The swabs were sent for bacterial culture and antibiotic sensitivity testing. The cultures were incubated in the Phoenix automated microbiology system (Becton Dickinson and Company). All bacterial isolates were tested for sensitivity to 15 different antibiotics, including moxifloxacin, gatifloxacin, trimethoprim, and vancomycin. On the basis of automated readings of the machine, not every bacterial isolate had a definitive sensitivity result to every antibiotic. Calculations of rates of resistance were based on definitive readings of sensitivity or resistance. Descriptive statistics were calculated for means and rates of case characteristics. Statistical analysis comparing groups of patients was performed using the 2-tailed Fisher exact test (StataCorp LP, College Station, TX).
Conjunctival Culture Results Fifty-eight bacterial cultures were isolated from 50 eyes, and the remaining 30 eyes showed no growth. The no growth rate was evenly split between study and control eyes (16 study eyes, 14 control eyes; P ⫽ 0.8). The most frequent bacteria identified was coagulase-negative staphylococcus (CNS), comprising 41 of 58 (70.7%) of the cultures. Six isolates of Staphylococcus aureus were identified, including one of methicillin-resistant S. aureus. Other bacteria identified included streptococcus mitis and coryneform species. The distribution of bacterial species between study and control eyes was balanced. Eighteen of 41 CNS isolates were from study eyes compared with 23 from control eyes (P ⫽ 0.38).
Antibiotic Resistance Rates Resistance to fluoroquinolones was defined as intermediate or full resistance to both moxifloxacin and gatifloxacin. Moxifloxacin resistance and sensitivity were defined as a minimum inhibitory concentration (MIC) ⱖ2.0 g/ml and ⱕ0.5 g/ml, respectively. Gatifloxacin resistance and sensitivity were defined as an MIC ⱖ8.0 g/ml and ⱕ2.0 g/ml, respectively. Among the control eyes, 9 of 28 eyes (32.1%) showed fluoroquinolone resistance compared with 14 of 22 eyes (63.6%) showing resistance among all study eyes (P ⫽ 0.04). In the subset of study eyes in patients who also used fluoroquinolones for 4 days after injection, 7 of 8 eyes (87.5%) showed fluoroquinolone resistance compared with 2 of 8 (25.0%) in the matched control eyes (P ⫽ 0.04). Figure 1 illustrates the rates of fluoroquinolone resistance among the different sets of eyes. Thirty-two bacterial isolates were grown from trimethoprim/ polymyxin B users: 14 from study eyes and 18 from control eyes. Trimethoprim resistance and sensitivity were defined as an MIC ⱖ16.0 g/ml and ⱕ8.0 g/ml, respectively. There was no significant difference in trimethoprim resistance rates between study and control eyes: Four of 14 study eyes (28.6%) showed resistance compared with 5 of 18 control eyes (27.7%) (P ⫽ 1.0). There was a single CNS isolate from the study eye of a trimethoprim/polymyxin B user that showed resistance to vancomycin. All other bacterial isolates in this study were sensitive to vancomycin. There was no cephalosporin resistance identified among the CNS isolates. The streptococcus mitis isolate was grown from a study eye, and the methicillin-resistant S. aureus colony came from a control eye. Figure 2 summarizes the resistance rates to other antimicrobials for the CNS isolates in both the study and control eyes.
Discussion The objective of intravitreal injection is to deliver pharmaceuticals into the vitreous cavity while minimizing the risk % Resistant
% Sensive
Results 12.5
Cohort Characteristics Eighty eyes (40 patients) were enrolled in the study. The average patient age was 76.2 years. A total of 27 of 40 patients (67.5%) were male, and 13 patients (32.5%) were female. Patients had received an average of 7 intravitreal injections for exudative AMD before enrollment, with a range of 3 to 13. Eleven of 40 patients used fluoroquinolone drops after the injections; 29 patients used trimethoprim/polymyxin B drops. Table 1 summarizes the baseline demographics of the study population.
36.4 67.9
75 87.5
63.6 32.1 All Study Eyes (n=22)
All Control Eyes (n=28)
25 Study Eyes using Fluoroquinolones (n=8)
Control Eyes using Fluoroquinolones (n=8)
Figure 1. Fluoroquinolone resistance rates among the different sets of eyes.
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Ophthalmology Volume 119, Number 7, July 2012 100% 90% 80% 70% 60% 50% 40%
Sensive
30%
Resistant
20% 10% 0%
Figure 2. Antibiotic resistance rates. Resistance rates to other antimicrobials among study eyes and control eyes. CE ⫽ control eyes; SE ⫽ study eyes.
of complications, including bleeding, pain, vitreous or drug reflux, intraocular pressure elevation, and endophthalmitis.7–9 Techniques and recommendations for minimizing the risk of endophthalmitis vary, and recent evidence suggests that subtle differences in intravitreal injection technique do not alter the inherent risk of endophthalmitis with this procedure.2 There is widespread use of postinjection topical antibiotics with the assumption that their use reduces the risk of infection; however, there is evidence disputing this assumption.5 Some of the commonly used antibiotic drops, including moxifloxacin, ofloxacin, and trimethoprim/polymyxin B, which we studied, are approved by the Food and Drug Administration for the treatment of ocular surface infections. None have been shown to reduce the incidence of postprocedural endophthalmitis. When the currently available newer-generation fluoroquinolones became commercially available for ocular use, there was hope that their use would aid in the prevention of postprocedure endophthalmitis.10 Fluoroquinolones offer broad-spectrum antimicrobial coverage and good ocular penetration when used topically.11 However, studies have shown that endophthalmitis still occurs despite use of these agents.12,13 Of note, many patients receive topical antibiotic drops preoperatively for cataract surgery, but for many patients this is the only intraocular procedure they will ever have. In contrast, patients with wet exudative AMD receive treatment for years on an ongoing basis at regular intervals.14,15 Repeated, short, 4-day courses of antibiotics have the potential for selection of resistant bacterial strains. The short course might be enough to create a selection bias for resistance, whereas the duration might not be long enough for eradication. The repetition of antibiotic use every 4 to 6 weeks enhances this selection force further. This could partly explain why fluoroquinolone resistance is an emerging problem in ocular microbiology.16,17 Our patients demonstrated a baseline fluoroquinolone resistance rate of 32.1% in the control eye. This is comparable to results from similar studies of ocular bacterial flora.17–21 Remarkably, after an average of 7 intravitreal injections, the fluoroquinolone resistance rate was 63.6% among all study eyes and 87.5% among the subset of eyes
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exposed to topical fluoroquinolones for 4 days after each injection. This suggests that repeated exposure to fluoroquinolones, even just a single drop on the eye after each injection, can lead to increased rates of resistance. These results are similar to those found by Kim and Toma.6 Of note, there is a definite trend toward cross-resistance among fluoroquinolones. This phenomenon has been described by Hooper22 and was also found on ocular surface bacteria by Kim and Toma6 and among staphylococcal endophthalmitis isolates by Miller et al.23 Our patients were exposed only to ofloxacin and moxifloxacin, and yet high rates of resistance to both moxifloxacin and gatifloxacin were found. As described by Hooper,22 among CNS strains, a single mutation may be sufficient to confer resistance to the entire fluoroquinolone class of antimicrobials. The other topical antibiotic we examined was trimethoprim/polymyxin B. Resistance to trimethoprim among gram-positive specimens is less well understood, especially from ocular sources. Although frequently used for ocular surface infections and as endophthalmitis prophylaxis, there are little data on the use of these drugs.24,25 Results from studies on CNS strains isolated from cases of cystitis show a trend toward resistance, with increasing rates of resistance emerging after widespread use of trimethoprim.26,27 Our study did not show a similar trend toward resistance to trimethoprim in patients using ocular trimethoprim/polymyxin B. The overall resistance rate of 26% that we found is identical to that found by Kim and Toma19 in conjunctival isolates tested for resistance to trimethoprim/sulfamethoxazole.
Study Limitations Our study has several limitations. It was performed on a relatively small group of patients at a single center. The influence of community-acquired, drug-resistant bacterial strains is therefore possible. We did not exclude patients who have had other intraocular surgeries and who likely used topical antibiotics at that time. Patients were excluded if any such procedure had been performed within 6 months of study enrollment, but lingering changes to the ocular flora beyond 6 months after such a procedure is certainly possible. The same can be said for systemic antibiotic use, and the same 6-month rule was used for exclusion. The study eyes were also repeatedly exposed to 5% povidone iodine, which could alter the characteristics of the ocular surface flora. Our study group consisted of patients who had received an average of 7 intravitreal injections in 1 eye only. This enabled us to use the fellow eye as a control. This assumes that there is no significant transfer of conjunctival bacterial flora between eyes in a given subject. This assumption could be called into question because direct transfer of flora through eye-rubbing or gradual transfer via the continuous mucous membrane of the nasolacrimal system, nasal cavity, and nasopharynx is certainly possible. However, such a transfer of flora would have only served to undermine our results by reducing the difference in resistance rates between study and control eyes. Either this transfer of flora was not significant in our study population or, if it did occur,
Milder et al 䡠 Antibiotic Resistance Patterns of Conjunctival Flora the power of the selection bias to create resistant strains on the study eye was enough to overcome it. The strength of our study is that it examines a crosssection of patients being treated for exudative AMD in a clinical setting. We use the subject’s fellow eye as a control, isolating the repeated povidone iodine preparation, intravitreal injections, and use of topical antibiotics as the possible factors responsible for changes in ocular bacterial flora. By including patients who use both fluoroquinolone and trimethoprim/polymyxin B drops, we uncovered 3 separate sets of eyes in terms of fluoroquinolone exposure: the control eyes with no exposure, the set of eyes using trimethoprim/ polymyxin B drops that only get exposed to a single drop of fluoroquinolone immediately after each injection, and the set of eyes exposed to fluoroquinolones for 4 days after each injection. Our results showed a definite linear, statistically significant increase in resistance, suggesting a cause and effect relationship between fluoroquinolone exposure and the development of high rates of resistance. In conclusion, the results of our study demonstrate that a significant proportion of patients exposed to repeated short courses of topical fluoroquinolones after intravitreal injections for exudative AMD develop ocular surface bacteria that are resistant to fluoroquinolones. Additional studies are needed to provide more definitive conclusions, but this work raises concerns about the routine use of topical antibiotics for endophthalmitis prophylaxis after repeated intravitreal injections. Such a strategy may not reduce the risk of infection and could produce changes in the conjunctival flora that are potentially harmful by selecting for more antibiotic-resistant organisms.
References 1. Fintak DR, Shah GK, Blinder KJ, et al. Incidence of endophthalmitis related to intravitreal injection of bevacizumab and ranibizumab. Retina 2008;28:1395–9. 2. Shah CP, Garg SJ, Vander JF, et al, Post-Injection Endophthalmitis (PIE) Study Team. Outcomes and risk factors associated with endophthalmitis after intravitreal injection of antivascular endothelial growth factor agents. Ophthalmology 2011;118:2028 –34. 3. Jager RD, Aiello LP, Patel SC, Cunningham ET Jr. Risks of intravitreous injection: a comprehensive review. Retina 2004; 24:676 –98. 4. Pilli S, Kotsolis A, Spaide RF, et al. Endophthalmitis associated with intravitreal anti-vascular endothelial growth factor therapy injections in an office setting. Am J Ophthalmol 2008;145:879 – 82. 5. Bhavsar AR, Googe JM Jr, Stockdale CR, et al, Diabetic Retinopathy Clinical Research Network. Risk of endophthalmitis after intravitreal drug injection when topical antibiotics are not required: the Diabetic Retinopathy Clinical Research Network laser-ranibizumab-triamcinolone clinical trials. Arch Ophthalmol 2009;127:1581–3. 6. Kim SJ, Toma HS. Ophthalmic antibiotics and antimicrobial resistance: a randomized, controlled study of patients undergoing intravitreal injections. Ophthalmology 2011; 118:1358 – 63. 7. Knecht PB, Michels S, Sturm V, et al. Tunnelled versus straight intravitreal injection: intraocular pressure changes, vitreous reflux, and patient discomfort. Retina 2009;29:1175– 81.
8. Rodrigues EB, Grumann A Jr, Penha FM, et al. Effect of needle type and injection technique on pain level and vitreal reflux in intravitreal injection. J Ocul Pharmacol Ther 2011;27:197–203. 9. Maurice D. Review: practical issues in intravitreal drug delivery. J Ocul Pharmacol Ther 2001;17:393– 401. 10. Chang DF, Braga-Mele R, Mamalis N, et al, ASCRS Cataract Clinical Committee. Prophylaxis of postoperative endophthalmitis after cataract surgery: results of the 2007 ASCRS member survey. J Cataract Refract Surg 2007;33:1801–5. 11. Hariprasad SM, Blinder KJ, Shah GK, et al. Penetration pharmacokinetics of topically administered 0.5% moxifloxacin ophthalmic solution in human aqueous and vitreous. Arch Ophthalmol 2005;123:39 – 44. 12. Moshirfar M, Feiz V, Vitale AT, et al. Endophthalmitis after uncomplicated cataract surgery with the use of fourth-generation fluoroquinolones: a retrospective observational case series. Ophthalmology 2007;114:686–91. 13. Deramo VA, Lai JC, Fastenberg DM, Udell IJ. Acute endophthalmitis in eyes treated prophylactically with gatifloxacin and moxifloxacin. Am J Ophthalmol 2006;142:721–5. 14. Brown DM, Michels M, Kaiser PK, et al, ANCHOR Study Group. Ranibizumab versus verteporfin photodynamic therapy for neovascular age-related macular degeneration: two-year results of the ANCHOR Study. Ophthalmology 2009;116:57–65. 15. Rosenfeld PJ, Brown DM, Heier JS, et al, MARINA Study Group. Ranibizumab for neovascular age-related macular degeneration. N Engl J Med 2006;355:1419 –31. 16. Fintelmann RE, Hoskins EN, Lietman TM. Topical fluoroquinolone use as a risk factor for in vitro fluoroquinolone resistance in ocular cultures. Arch Ophthalmol 2011;129:399–402. 17. Park SH, Lim JA, Choi JS, et al. The resistance patterns of normal ocular bacterial flora to 4 fluoroquinolone antibiotics. Cornea 2009;28:68 –72. 18. Ta CN, Chang RT, Singh K, et al. Antibiotic resistance patterns of ocular bacterial flora: a prospective study of patients undergoing anterior segment surgery. Ophthalmology 2003; 110:1946 –51. 19. Kim SJ, Toma HS, Midha NK, et al. Antibiotic Resistance of Conjunctiva and Nasopharynx Evaluation Study: a prospective study of patients undergoing intravitreal injections. Ophthalmology 2010;117:2372– 8. 20. Chung JL, Seo KY, Yong DE, et al. Antibiotic susceptibility of conjunctival bacterial isolates from refractive surgery patients. Ophthalmology 2009;116:1067–74. 21. Moss JM, Sanislo SR, Ta CN. Antibiotic susceptibility patterns of ocular bacterial flora in patients undergoing intravitreal injections. Ophthalmology 2010;117:2141–5. 22. Hooper DC. Emerging mechanisms of fluoroquinolone resistance. Emerg Infect Dis 2001;7:337– 41. 23. Miller D, Flynn PM, Scott IU, et al. In vitro fluoroquinolone resistance in staphylococcal endophthalmitis isolates. Arch Ophthalmol 2006;124:479 – 83. 24. Asbell PA, Colby KA, Deng S, et al. Ocular TRUST: nationwide antimicrobial susceptibility patterns in ocular isolates. Am J Ophthalmol 2008;145:951– 8. 25. Price FW Jr, Dobbins K, Zeh W. Penetration of topically administered ofloxacin and trimethoprim into aqueous humor. J Ocul Pharmacol Ther 2002;18:445–53. 26. Araújo SM, Mourão TC, Oliveira JL, et al. Antimicrobial resistance of uropathogens in women with acute uncomplicated cystitis from primary care settings. Int Urol Nephrol 2011;43:461– 6. 27. Gupta K, Scholes D, Stamm WE. Increasing prevalence of antimicrobial resistance among uropathogens causing acute uncomplicated cystitis in women. JAMA 1999;281:736–8.
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Ophthalmology Volume 119, Number 7, July 2012
Footnotes and Financial Disclosures Originally received: August 14, 2011. Final revision: December 9, 2011. Accepted: January 9, 2012. Available online: March 13, 2012.
Manuscript no. 2011-1222.
Presented at: the American Society of Retina Specialists, 29th Annual Meeting, August 2011, Boston, Massachusetts.
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Financial Disclosure(s): The author(s) have no proprietary or commercial interest in any materials discussed in this article. Funding: Retina Endowment Fund, Philadelphia, Pennsylvania. The funding organization had no role in the design or conduct of this research. Correspondence: James Vander, MD, 840 Walnut St, Suite 1020, Philadelphia, PA 19107. E-mail: [email protected].