Retinal toxicity of ceftazidime in the infusion fluid for vitrectomy in the rabbit eye

From the L S U Eye C e n t e r, L o u i s i a n a S t a te University Medical Center School ofMedi c ine, New Orleans, LA. Supported in part by U . S . Public Health Service grants EY0754 I and EY02377 frorn the National Eye l n s t i t u t e , N a t i o n a l I n s t i t u t e s o f H e a l t h , Bethesda, MO a n d b y a n unrestricted departrnental grant frorn Research to Prevent B l indness, New York, NY. Address reprint requests to: Dr. Ismael Caixeta Ri­ beiro. Hospital Santa Lúcia Av. Santos Dumont, 3 34. Centro, Uberaba (MG) CEP 380 1 0-3 70. SUMMARY


INTRODUCfION
Bacterial endophthalmitis is the most dreaded complication in the period after intraocular surgery and after penetrating injuries of the globe1-4• The disease also may be associated with a systemic infection. As an infectious process ofthe internai structures ofthe eye, it commonly leads to a signifi cant decrease in visual acuity and even to loss ofthe eye 4• 5• The prognosis is particularly poor for infections caused by gram-negative pathogens 6• Because topical, subconjunctival, and systemic antibiotics are often ineffective in treating endophthalmitis 4, intravitreal injection of antibiotics with or without vitrectomy can favorably alter the outcome of the disease 7• 8. The use of antimicrobial drugs in the infusion fluid during vitreous surgery . in the treatment or prevention of endophthalmitis has been advocated 3• 9. Before any procedure, vitreous and aqueous taps should be performed to provide samples for culture and antibiotic sensitivity. Because any delay in initiating appropriate therapy can worsen the prognosis, the practice is to Retina/ toxicity of cefiazidime in the infusionfluidfor vitrectomy in the rabbit eye initially treat with broad-spectrum antibiotics. The most com mon regimens are associated with antimicrobials that have good activity against gram-positive and gram-negative bacte ria. The aminoglycosides have very good action against gram negative organisms and are commonly employed, but severe retina! toxic reactions have been reported 1 • For this reason, we are evaluating the toxicity of ceftazidime diluted in the infu sion solution. Ceftazidime is a third-generation cephalosporin with potent bactericida! activity against gram-negative rods which showed no toxic effects when injected intravitreously in the concentration of 2.25 mg in 0.1 mL of balanced salt solution 1•

Antibacterial Agent
Ceftazidime was obtained as a powder from Eli Lilly & Co.

Animais
Twenty-two right eyes of 22 New Zealand white rabbits weighing 2-3 kg were used in the experiment; the left eyes (n = 22) were used as controls. The rabbits were cared for accord ing to the Association for Research in Vision and Ophthalmo logy resolution on the use of animals in research. Indirect ophthalmoscopic examination was performed on ali rabbits after dilation of pupils with tropicamide 1 % (Mydriacyl 1 %, Alcon Laboratories, lnc, Fort Worth, TX), and phenylephrine hydrochloride (Mydfrin 2.5%, Alcon) and ali eyes were found to be normal.

Electroretinography
For electroretinography (ERG), the rabbits were dark adapted for 30 min and then anesthetized with a mixture of ketamine (50 mg/kg) and xylazine (5 mg/kg). Pupils were di lated and ERGs were recorded using gold contact tenses and stainless steel needle electrodes positioned behind the ears (reference) and on the forehead (ground). Scotopic conditions involved a 10 µsec Ganzfeld flash (1 per 10 sec) ofblue light (Wratten filter Nº 47) at low intensity (events 1 and 2). The low-intensity exposure was repeated without the blue filter (events 3 and 4). The light intensity was increased to medium in events 5 and 6 and to high for events 7 and 8, using strobe generator Model 1539-A (General Radio, Concord, MA).
Vitrectomy infusion solutions were prepared using cefta zidime in BSS in the concentrations mentioned above. Con centrations of 20 , u g/mL were used in 2 eyes, 40 µg/mL in 4 eyes, 80 µg/mL in 5 eyes, 100 µg/mL in 2 eyes, 150 µg/mL in 2 eyes, 200 µg/mL in 2 eyes and 300 µg/mL in 5 eyes. Ali left eyes served as controls, receiving only BSS as infusion fluid. We used more eyes in the concentrations of 40 µg/mL and 80 µg/mL because we thought that could be the ideal concentra tions to use, and 300 µg/mL to better evaluate any signs of toxicity that could appear with this concentration.
Complete vitrectomy was performed via the pars plana. A 360º peritomy and three sclerotomies were made with a microvitreoretinal (MVR) blade, 4 mm posterior to the limbus. The infusion cannula was located superiorly and the other two sclerotomies located inferiorly were used for the insertion of the fiberoptic light pipe and the vitrectomy probe. The procedures lasted 15-20 min and a minimum of 120 mL of infusion solution containing the drug was used during each operation. The infusion cannula was fixed with 5-0 Dacron sutures and the sclerotomies were closed with 6-0 polyglactin aft er the instruments were withdrawn. The conjunctiva was also repaired with 6-0 polyglactin sutures.
The rabbits were examined daily for two weeks after surgery by means of externai examination and indirect ophthalmoscopy. Two weeks after surgery, ERGs were performed in the sarne manner and the rabbits were sacrificed by intravenous injection of 1 mL sodium pentobarbital. The eyes were enucleated and processed either for light or electron microscopy.

RESULT
Ali animais showed mild to moderate eyelid and conjuncti val edema. Three presented with discrete posterior cataracts caused by surgical trauma (80 µg/mL and 300 µg/mL); two presented with small and localized vitreous hemorrhage at the time ofsclerotomy closure in the inferior quadrant (80 µg/mL). None ofthe complications impaired visualization ofthe retina, which appeared normal by indirect ophthalmoscopy at ali concentrations used. Vitreous hemorrhage had almost cleared by the second week.
Preoperative and postoperative ERGs were comp ared for ali animals. Because ofnormal variation in amplitude in rabbit ERGs, variations of less than 25% of the preoperative ERG were considered insignificant. Variations of25% to 50% were rated significant and more than 50% reduction in amplitude was rated severe. No significant changes in the ERG were found in eyes given infusion fluid containing concentrations of 20 µg/mL and 40 µg/mL. With the use of 80 µg/mL, 1 of 5 eyes developed changes classified as significant (35% of the preoperative ERG). ln the concentration of 100 and 150 µg/mL, there were no significant alterations in the ERG, but with 200 and 300 µg/mL, the postoperative ERG had a significant de crease in amplitude, especially in the first two events (sco topic). ln the control group, the alterations of ERG were con sidered insignificant.

Light Microscopy
None ofthe treated eyes showed significant alterations in    Table 1. glycoside-sensitive P. aeruginosa of ceftazidime is 8 mg/l, and the MIC9 0 for strains resistant to one or more amino glycosides is 128 mg/l. The MIC9 0 for Pseudomonas aeru ginosa isolates highly resistant to ali three aminoglycosides (MIC9 0 > 128 mg/l) is 32 mg/l for ceftazidime. A 2.25 mg intravitreal dose of ceftazidime has been proven nontoxic to the retina 'º. We tested the toxicity of ceftazidime in the infusion fluid and found that at concentrations of20, 40, 80, 100 and 150 µg/mL, there were no signs of retina! toxicity, either by ERG or histology. At concentrations of200 and 300 µg/mL (300 µg/mL is the sarne as 45 mg into 150 mi ofBSS ), the ERGs changed significantly and histological examination showed small alterations in the outer segments ofthe photore ceptors 16• (Figs. 1 and 2).
We found that 150 µg/mL of ceftazidime is safe for ocular tissue and still effective in the treatment of gram-negative endophthalmitis. We believe that 200 µg/ml is a borderline dosage and further studies should be done, specifically in primates, because in previous studies on ceftazidime toxicity, the macula was shown to be the site ofthe retina that was most affected by high doses (macular cyst or hole formation).
As ceftazidime has a low levei of activity against Staphylo coccus aureus and S. ep idermidis, it should always be used in combination with a drug that has good activity against gram positive bacteria for the treatment or prevention of endophthalmitis.