TREATMENT OF BACTERIAL ULCERS OFTHE CORNEA IN THE RABBIT:

A COMPARISON OF ADMINISTRATION BYEYE DROPS AND SUBCONJUNCTIVAL

INJECTIONS

BY Jules Baum, MD

INTRODUCTION

A BACTERIAL CORNEAL ULCER IS ONE OF THE MOST SERIOUS INFECTIOUS DISEASES

of the eye and must be dealt with swiftly and effectively if vision is to bepreserved. To successfully treat a bacterial corneal ulcer, the clinicianmust not only identify the pathogen and prescribe the appropriate antibi-otic, but also determine the route and frequency of administration thatwill deliver and maintain the drug in therapeutic concentrations in theeye. The purpose of this paper is to examine the various routes of admin-istration available to treat a bacterial corneal ulcer, to review past effortsevaluating topical and subconjunctival drug delivery and to comparedirectly the effectiveness of the topical and subconjunctival routes exper-imentally.

ROUTES OF ANTIBIOTIC ADMINISTRATION: REVIEW OF THE LITERATURE

Antibiotics may be administered in the treatment of bacterial cornealulcers by the following routes: topical (eye drops or ointments), subcon-junctival, and systemic (orally or by intravenous or intramuscular injec-tion). In this section I will discuss and compare the pharmacokinetics andefficacy of these routes. Extrapolation from such comparisons are limitedby differences between the rabbit model and the human eye, and byvariance in experimental protocols among investigators. For instance,Maurice and Ota' found that when fluorescein was given as a subconjunc-tival injection, it streamed out the hole in the rabbit conjunctiva; incontrast, there was very little reflux seen in humans by slit-lamp examina-tion. Moreover, 100 times more fluorescein entered the anterior chamberTR. AM. OPHTH. Soc. vol. LXXX, 1982

Baum

in humans than in rabbits following subconjunctival injections. Since theeye/body size ratio is much higher in rabbits than in humans, an equiva-lent dose administered subconjunctivally produces much higher serum

levels in the rabbit than in humans. Indeed, the serum levels produced inthe rabbit are of a magnitude which would normally be expected usingintramuscular or intravenous therapy.2 They may indirectly contribute tointraocular levels of drug in a way that would not occur in humans. Thesedifferences between rabbits and humans require caution when data fromone species is extrapolated to the other.

TOPICAL ADMINISTRATION

Eye DropsTable I describes the concentration of antibiotics in the rabbit cornea aftertopical administration. It summarizes a representative series in the spe-

cies used most often by those investigating ocular pharmacokinetics. Inmost intances, the concentration of antibiotic was 25 ,ug/g or less. Thelipophilic antibiotics, chloramphenicol and tetracycline, produced some

of the highest corneal concentrations. Although differences in methods

TABLE L CONCENTRATION OF ANTIBIOTICS IN RABBIT CORNEA AFTER TOPICAL APPLICATION*

CORNEAL CON-ANTIBIOTIC REGIMEN CENTRATION (pg/g) REFERENCE

Penicillin G 1.2% corneal bath 28for 3 h

5,000-20,000 u/gm 347-1729 u (57)in cotton pledget

Gentamicin 0.3% drops for 6 h 15 (Lye ulcer) (10)2 (Pseudomonas ulcer)

0.3% drops for 8 h 10 (sterile corneal suture) (60)20 (corneal infection) (60)

Tetracyclines 0.5%-2.5% drops or 3-10corneal bath for1 h

35 (abraded or ulceratedcornea)

0.5%-2% ointment 3-25 (15, 47)Erythromycin 2.5% drops for 1 h 4-9

10 (abraded cornea)Chloramphenicol 0.25% drops or 5% 25 (15, 48)

in propylene gly-col for 30 min.

1% ointment 70

*Modified from Barza46; unless stated otherwise, studies were performed in normal eyes.Data selected from Benson"5 unless otherwise stated.

370

Bacterial Corneal Ulcers

prevent direct comparison of all the studies, the following observationscan be made: (1) higher corneal concentrations of drugs were achieved inabraded or inflamed corneas than in normal ones; (2) larger concentra-tions of administered drug and longer contact times resulted in highertissue concentrations; and (3) a lipophilic antibiotic such as chlorampheni-col, which penetrates corneal epithelium more easily than non-lipophilicantibiotics, administered as an ointment produced higher corneal concen-tration than did the drug given as an eye drop. These data are scanty andthe above observations are only generalizations. However, there areother data supporting these postulates. Davis et a13 demonstrated thatconcentrated eye drops were more effective than less concentrated onesin reducing the bacterial colony count. As an illustration of the effects oflonger contact time, Mishima and Nagataki4 showed that methylcellulosegreatly increased peak concentrations of pilocarpine in iris following topi-cal application; the bioavailability was about three times that of the solu-tion without the methylcellulose. They found no advantage in increasingviscosity of the ophthalmic solution above 10-20 cs, which corresponds tohydroxymethylcellulose concentrations of 0.38% to 0.5%.

Tear dynamics play a role in determining bioavailability of antibiotictopically applied to cornea. The average size of an eye drop emanatingfrom a squeeze-type bottle is 40-50 ,l. A recent study showed thatincreasing drop size of fluorescein from 5 to 20 ,lI resulted in only aminimal increase in the initial tear concentration of fluorescein,5 appar-ently because the larger eye drop drained more rapidly, allowing littletime for mixing of fluorescein with tears. In the same study, variousvolumes of pilocarpine solution, ie, 5, 10, 20 and 50 RI were instilled inthe eye and pupillary responses were measured. Increasing the drop sizeover 20 ,ul did little to decrease pupil size. Both the fluorescein andpilocarpine studies demonstrate that the reflex stimulation of tear flowand poor mixing of drug and tears induced by large drop administrationproduce a rapid exit of fluid through the lacrimal outflow tract. These datasuggest that drop sizes larger than 20 RI are no more effective thansmaller ones. Manufacturers should consider reducing drop size for com-mercial topical medications. Since the maximum volume the eye can holdis 30 pI,6 administration of two successive drops merely increases sys-temic absorption in some instances without significantly enhancing drugeffect. Similar principles apply to the instillation of two different consecu-tive eye drops. One should wait at least five minutes before instilling asecond eye drop solution, since reflex stimulation persists for approxi-mately five minutes following instillation of an eye drop as small as 1 p'.6The tonicity of the eye drops also affects bioavailability. Instillation of

371

concentrated antibiotic eye drops, fabricated from parenteral medi-cations, is generally advised for the treatment of a bacterial corneal ulcer.The more concentrated the solution, the greater the tonicity. Maurice7has shown, after instillation of eye drops containing fluorescein to thehuman eye, that increasing tonicity of the drops above that of body fluidsleads to their immediate dilution by osmosis from the conjunctiva with noincreased penetration of fluorescein into the eye. This study suggests thatconcentrated antibiotic eye drops, in addition to inducing reflex tearing,are further diluted by osmosis. Hence, corneal tissue concentrationswould not be directly proportional to the increased concentration of theeye drop. The discrepancy between these findings and those of Davis etal3 (which demonstrated increased effectiveness of concentrated antibi-otic eye drops in the guinea pig) may relate less to the model and drugthan to the effect of the preservative on drug penetration. Burnstein8 hasshown that preservatives applied topically at concentrations commonlyemployed in commercial eye drop solutions decrease the permeability ofthe corneal epithelium. Recently Sidikaro and Jones,9 using human vol-unteers, measured the kinetics of gentamicin following installation ofsolutions with concentrations of 3 and 13.6 mg/ml. They detected higherconcentrations of gentamicin in the tears over a one hour interval follow-ing instillation of the more concentrated eye drops. Thus it would seemthat one can increase the concentration in the tear film, and presumablyin the cornea, by the use of concentrated eye drops.

In summary, these data suggest that more concentrated antibiotic eyedrops are more effective than those now commercially available andshould be employed for the treatment of bacterial corneal ulcers. Drugcompanies should be encouraged to manufacture more highly concentrat-ed drops for use in special instances.

Increased frequency of administration of antibiotic eye drops also helpsachieve higher corneal levels. We now believe that in treating bacterialcorneal ulcers, drops should be given every 15 to 30 minutes10-12 toinduce and maintain therapeutic concentrations in the cornea.A soft contact lens is often required to protect the corneal surface and

aid in re-epithelialization. Antibiotic eye drops are frequently employedconcomitantly either therapeutically or prophylactically. Once a lens issaturated with drug, it serves as a depot and aids in increasing theantibiotic concentration in the cornea.4'1315

OintmentsAntibiotic ointments are rarely, if ever, used initially in the treatment ofbacterial corneal ulcers. This practice seems to derive more from theoret-

372 Baum

Bacterial Corneal Ulcers

ical objections than from clinical or experimental data. Ointments, com-pared to eye drops, prolong the contact time of antibiotic, thus achievinga higher tissue concentration.16'17 This is especially true for the morelipid-soluble antibiotics such as chloramphenicol and the tetracyclines.One objection to the use of antibiotic ointments in treating bacterialcorneal ulcers is the impairment to corneal penetration of an eye dropused concomitantly. A second is that corneal tissue concentration may beelevated by increasing the concentration and frequency of administrationof eye drops, whereas antibiotic ointments do not lend themselves to suchmodifications.

SUBCONJUNCTIVAL INJECTIONS

Subconjunctival injections are, in almost all clinical instances, anteriorsub-Tenon's injections. However, there is probably an insignificant ther-apeutic difference between the two routes.20-22 Despite the inaccuracy ofdefinition, I will use the accepted jargon and refer to this type of peri-ocular injection as a subconjunctival injection. Antibiotic levels in con-junctiva, cornea and aqueous are higher following a subconjunctival injec-tion than following a retrobulbar (posterior sub-Tenon's) injection.23 Acomparison of subconjunctival injection with other routes of antibioticadministration will be considered in this section. Table II summarizes arepresentative series of studies in rabbits. The peak corneal concentra-tions displayed in Table II are all strikingly high. However, such injec-tions are routinely administered at 12 to 24 hour intervals and during thisinterval corneal concentrations fall off rapidly.2,10,24-28A subconjunctival injection leaves a hole in the conjunctiva through

which drug may exit, decreasing the bolus and increasing drug concentra-tion in the tears. It has been shown in rabbits that less than 10% of asubconjunctival dose is lost to the tears.29 The rest enters the oculartissues directly or is carried away in the systemic circulation, or it dissectsposteriorly in the sub-Tenon's space.

Large topographical differences in corneal concentrations exist follow-ing subconjunctival injections; the highest concentration is nearest theinjection site.27'30 This unevenness of distribution suggests that most drugenters the cornea by direct diffusion through tissue, rather than throughthe pre-corneal tear film. It might be expected that ocular inflammationwould induce higher concentrations of drug in the cornea compared tothe non-inflamed eye; however, no such trend is seen,30 probably be-cause of a larger flow of drug away from the cornea through the engorgedconjunctival vessels in the inflamed eye.29

373

Baum

TABLE II: PEAK CONCENTRATION OF ANTIBIOTICS IN RABBIT CORNEA AFTER SUBCONJUNCTIVALADMINISTRATION*

ANTIBIOTIC AND NORMAL EYES INFLAMED EYESDOSE (1gg) (1gg) REFERENCES

Penicillin G100 mg

50 mgCarbenicillin

100 mg

50 mgMethicillin

100 mg

Oxacillin100 mg

Cefazolin100 mg25 mg12.5 mg

Cefamandole12.5 mg

Streptomycin50 mg

Gentamicin20 mg

10 mg5 mg

Chloramphenicol150 mg50 mg

Clindamycin34 mg

3000(-> 1500)

300

> 300(-> 42)15-35

> 1000

> 300(-> 25)

700(-> 40)

2000(-> 500)

600(-> 100)

150(-> 40)

140(-> 40)

1000

30

50500

100

200(-> 25)

1010

(26)

(27)

(24)

(49)

(50)

(50)

(50)(51)(50)

(25)

(26)

(2)

(12)(30)

(19)(27)

(52)

*Modified from Barza.'Parentheses contain concentrations 3 to 6 h after administration.

SYSTEMIC ADMINISTRATION

Table III depicts corneal concentrations of antibiotic after systemic ther-apy. In general, higher serum and, hence, higher corneal tissue levels areachieved after intravenous than after intramuscular or subcutaneous in-jections. Ocular inflammation enhances the penetration of antibioticsfrom the systemic circulation into corneal tissues. 20'31 However, com-paring the corneal concentrations achieved by systemic administration to

374

Bacterial Corneal Ulcers

TABLE III: CONCENTRATION OF ANTIBIOTICS IN RABBIT CORNEA AT PEAKAFTER SYSTEMIC ADMINISTRATION*

CORNEAL CONCENTRATION (pg/g)

ANTIBIOTIC REGIMEN NORMAL INFLAMED REFERENCESEYES EYES

Penicillin G 62 mg/kg SC 4 6 (28)40 mg/kg IV 9 (26)

Ampicillin 50 mg/kg IV 3 (53)Amoxicillin 100 mg/kg 2 (54)

POCarbenicillin 100 mg/kg 11 85 (24)

IVStreptomycin 15 mg/kg IV 13 (26)Gentamicin 1.7 mg/kg 0.5 3.4 (55)

IM1.6 mg/kg/h 10 (56)IV

Chloramphen-icol 50 mg/kg IV 5 (16)

Clindamycin 100 mg/kg trace (58)

*Modified from Barza.4SC = Subcutaneous.

those found after subconjunctival injection (Table II), it is obvious thatsystemic administration is an inefficient route of drug delivery to thecornea.32 Not only are topical administration and subconjunctival injec-tions more effective, but they pose less risk of serious side effects thandoes systemic administration, especially with antibiotics whose side-ef-fects are dose-related (eg, aminoglycosides). Thus, there is no valid rea-son, in my opinion, to administer systemic antibiotics in the treatment ofa non-perforating bacterial corneal ulcer, especially since bacteria havenever been demonstrated intraocularly prior to a corneal perforation.However, systemic antibiotics are indicated if the cornea perforates, ifthere is extension to sclera, if the pathogen is Neisseria gonorrhoeae, or incases of Pseudomonas3 or Haemophilus influenzae in infants, in whichtwo instances systemic spread of the infection may ensue.

THE PROS AND CONS OF TOPICAL VERSUS SUBCONJUNCTIVAL ADMINISTRATION

A bacterial corneal ulcer is a sight-threatening condition which demandsprompt sterilization. In the last decade, we have witnessed a dramaticimprovement in our ability to sterilize such ulcers, decrease morbidityand preserve vision, largely due to the use ofnew antibiotics and recogni-tion of the need to achieve higher and more sustained corneal concentra-tion of drug. Augmented corneal levels have been achieved by increasing

375

the concentration and frequency of application of topical antibiotic eyedrops and by the use of subconjunctival injections.Although subconjunctival injections are in common use today to treat

severe corneal infections, invariably in combination with topical therapy,there are no data to indicate that subconjunctival or the combination ofroutes is more effective than the topical therapy alone. Furthermore,what data are available concerning the relative efficacy of these two routeshave generally been derived by (1) measuring tissue concentration of thedrug following administration, and by (2) evaluating the effectiveness ofan antibiotic in eradicating an infection, ie, the number of bacteria killed.Curiously, to date no investigators have employed the two techniquessimultaneously.The data in Table II demonstrate that peak corneal levels of antibiotic

are strikingly high following subconjunctival injections. The data docu-menting fall-off in levels hours after a subconjunctival injection suggestthat, in the majority of instances, concentrations decline rapidly by threeto six hours after injection (Table II). Such levels are probably therapeuticin most instances. Similarly, a number of studies suggest that subconjunc-tival injections are effective in "killing" bacteria following the productionof experimental bacterial keratitis.34-38The data in Table I, compared to those in Table II, indicate that peak

values are not as high following topical administration as following sub-conjunctival injections. However, drops can be instilled every 15 to 30minutes, thereby maintaining tissue levels of drug throughout the day.Moreover, in virtually all instances, with epithelium absent or derangedover a bacterial corneal ulcer, drug levels are higher than in a normal eyecovered with epithelium. As stated above, more important than deter-mining tissue concentrations is the effectiveness of an antibiotic in eradi-cating bacteria in the infected cornea. Davis, Hyndiuk and co-workersperformed a comprehensive series of experiments, mainly in guinea pigs,in part to determine whether the topical, subconjunctival or parenteralroute was most effective in eradicating organisms from bacterial cornealulcers. They found the topical route most effective,39-43 especially withconcentrated antibiotic eye drops,3 frequent applications,39 with the cor-neal epithelium removed,3 and when treatment was started early.3 Kup-ferman and Leibowitz44'45 also demonstrated the efficacy of topical ther-apy in eradicating organisms.

Curiously, Leibowitz et al45 could not demonstrate a statistically sig-nificant reduction in the number of bacteria in the cornea when theantibiotic was administered by the subconjunctival route. It is difficult toreconcile these efficacy studies which demonstrate the superiority of

376 Baum

Bacterial Corneal Ulcers

topical over subconjunctival administration with those studies showingmassive concentrations of antibiotic in cornea after subconjunctival injec-tions (Table II).

It is exactly for these reasons that the present investigation was under-taken. Moreover, it is clinically important to know the relative value ofeach of the two main routes of antibiotic administration in the treatmentof a bacterial corneal ulcer.

MATERIALS AND METHODS

All studies were performed in Dutch belted (pigmented) rabbits, weigh-ing approximately 2 kg.

INFECTION

Two bacteria were employed to cause infection. The first, Staphylococcusaureus 209P, was grown overnight at 37 C in trypticase-soy broth (TSB)and diluted with phosphate-buffered saline (PBS) to yield an anticipated 2x 105 colony-forming units per ml (CFU/ml); the second, Pseudomwnasaeruginosa 107 (kindly supplied by Starkey Davis, MD), was grownovernight at 37 C in TSB and was diluted with PBS to yield an anticipated2 x 104 CFU/ml. The animals were anesthetized with intramuscularketamine and chlorpromazine (Thorazine) whenever necessary. Ten mi-croliters of the bacterial suspension was injected into the stroma of thecentral cornea using a 30-gauge needle on a microsyringe. Simultaneouscolony counts (see Sampling and Assays, below), showed the inoculacontained from 56 to 274 CFU of Staphylococcus aureus or 227 to 400CFU of Pseudomonas aeruginosa per injection. Drug therapy was initiat-ed 24 hours after infecting a cornea.Two antibiotics, cefazolin and gentamicin, administered either in eye

drop form or as a subconjunctival injection, were used to treat the infect-ed corneas. The staphylococcal ulcers were treated with gentamicin orcefazolin; the Pseudomonas ulcers were treated with gentamicin. Cefazo-lin was prepared by adding 4.5 ml PBS to 1 gram of cefazolin (finalconcentration 200 mg/ml); 0.5 ml (100 mg) was given as a subconjunctivalinjection into the superior quadrant using a 30-gauge needle. Cefazolineye drops were prepared by adding 10 ml PBS to 2 ml of the subconjunc-tival solution for a final concentration of 33.3 mg/ml. Gentamicin eyedrops, either 3 mg/ml or 14 mg/ml were prepared from commercialinjectable gentamicin and diluted in PBS. The solution for subconjuncti-val injection was prepared by adding powdered gentamicin to commercialgentamicin to a concentration of 80 mg/ml; 0.5 ml was injected subcon-

377

junctivally as above. Eye drops were administered to both eyes, subcon-junctival injections to the left eye only. Only one mode of therapy wasused per animal. Control animals received sterile PBS eye drops in theright eye and a subconjunctival injection of 0.5 ml sterile PBS to the lefteye.The treatment schedule is summarized in Table IV. Subconjunctival

injections were administered at 0 and 8 hours. Following each subcon-junctival injection, the cornea was irrigated for 1.5 minutes with 30 mlsterile PBS. Eye drops, 100 ,ul, were administered every hour, either for8 or 16 hours. These details in the protocol were chosen to duplicate thoseemployed by Leibowitz et al.45

SAMPLING AND ASSAYS

Serum samples were taken 30 minutes following the subconjunctivalinjection and immediately following the last eye drop. Animals weresacrificed with pentobarbital one hour after the 8th or 16th topical admin-istration and one or nine hours after the last subconjunctival injection (asdid Leibowitz et al).45 An aqueous sample was removed with a 25-gaugeneedle on a tuberculin syringe. The corneas were removed and a 7.5 mmbutton was trephined around the central ulcer. The buttons wereweighed, minced briefly and placed in a Wheaton microgrinder in an icebath. PBS, 0.3 ml for cefazolin and 0.4 ml for gentamicin, was added. Thecorneas were ground for - 60 strokes and the supernatant was removedwith a Pasteur pipette. Since the initial corneas weighed 25 to 40 mg, thisprocedure resulted in a dilution factor of about 10 to 20-fold.

In order to determine the number of viable bacteria remaining in eachcornea, a sample (0.05 ml) of supernatant was serially diluted from 10'1to 10-5. Fifty microliter drops of each dilution were plated on agar andthe number of colonies which formed after 24 hours was counted.Another sample of the corneal supernatant fluid was assayed for antibi-

otic content using an agar-diffusion bioassay with standards prepared inPBS. Because the initial corneal specimens were diluted in about 10 to20-fold their weight of PBS during the procedure of grinding, the lowerlimit of sensitivity of the assay was approximately 2 to 8 ,ug per gram ofcornea for gentamicin and 4 to 8 ,ug per gram of cornea for cefazolin.Aqueous humor and serum were assayed in a similar manner using PBSand normal rabbit serum, respectively, as diluent for the standards.Because these samples did not require dilution, the lower limit of sensi-tivity of the assay was 0.2 to 0.4 ,ug/ml for gentamicin and 0.4 to 0.8,ug/ml for cefazolin. As a fiurther verification of the accuracy of the genta-

378 Baum

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micin bioassay, some samples of corneal supernatant were studied simul-taneously by radioimmunoassay (RIANEN, New England Nuclear, Bos-ton, MA). The results were very similar to those measured by bioassayand will not be presented.

For the purposes of statistical analysis, the concentrations of antibioticin specimens which contained no detectable antibiotics were consideredto be one-half of the lowest detectable concentration.

STATISTICAL ANALYSIS

Initially, a one-way analysis of variance was performed in order to deter-mine whether or not a significant difference existed within each experi-mental group between the means of the various subgroups with respect tothe following dependent variables: (a) the number ofCFU per cornea, (b)the concentration of antibiotic in the cornea, (c) the concentration ofantibiotic in the aqueous. Logarithmic transformations were performedon each of the dependent variables in order to induce homoskedasticity.If the results of the analysis of variance procedure indicated that a signifi-cant difference did exist between the means, Scheffe's method for mul-tiple comparisons was used to make pairwise comparisons between se-lected pairs of means. If there were only two subgroups available forconsideration within an experimental group, a two-sample t-test wasemployed to test for equality of the means. The 0.05 level of significancewas used throughout the analysis.

RESULTS

The major results are displayed in Figures 1 to 3 and Table V. Bothtopical and subconjunctival antibiotic therapy, administered as either twosubconjunctival injections every eight hours or as hourly eye drops over aperiod of 17 hours, significantly (P < 0.05) reduced bacterial CFU fromexperimentally produced corneal ulcers as compared to control eyes.These differences occurred with each drug-organism combination at both9 and 17 hours with one exception. There was no significant difference incolony counts between staphylococcal ulcers treated with gentamicin bysubconjunctival injection and controls after nine hours of therapy.

Corneal concentrations of antibiotic were significantly higher in allinstances one hour after the second subconjunctival injection than whenantibiotic was administered topically. Conversely, by the 17th hour oftherapy, corneal concentrations of antibiotic were significantly lowerwhen antibiotic was administered subconjunctivally (nine hours after the

380 Baum

Bacterial Corneal Ulcers

STAPHYLOCOCCAL CORNEAL ULCERS TREATED WITH CEFAZOLIN

THERAPY FOR 9 HOURSDROPS HOURLY FOR 8 HOURS

ORS/C AT 0 AND 8 HOURS

S/C ICn*aFIGURE 1

THERAPY FOR I7 HOURSDROPS HOURLY FOR 16 HOURS

ORS/C AT 0 AND 8 HOURS

STAPHYLOCOCCAL CORNEAL ULCERS TREATED WITH GENTAMICIN*

THERAPY FOR 9 HOURS THERAPY FOR 17 HOURSDROPS HOURLY FOR 8 HOURS DROPS HOURLY FOR 16 HOURS

OR ORS/C AT 0 AND 8 HOURS S/C AT 0 AND 8 HOURS

Colony Forming Units Gentamidin Conc.per Cornea (logo) in Cornea (ug/g)

NOT

Centr Tr iul S/C

*3 mg/ml topically.

--

-

--

----

-

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-Nar _00"g _ _

Colony Forming Units Gentomicln Conc.per Cornea (log,0) in Conea (ugig)

4

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NOT V-

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FIGURE 2

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Baum

PSEUDOMONAL CORNEAL ULCERS TREATED WITH GENTAMICIN*

THERAPY FOR 9 HOURS THERAPY FOR 17 HOURSDROPS HOURLY FOR 8 HOURS DROPS HOURLY FOR 16 HOURS

OR ORS/C AT 0 AND 8 HOURS S/C AT 0 AND 8 HOURS

Colony Forming Units Gentamicin Cone. Colony Forming Units Gentamicin Conc.per Cornea (logO) in Corneo (ug/g) per Cornea (logo) In Cornea (ug/g)

'I

St7ST

Mt

ILS0

MrDOME

A^W S/C

*3 mg/ml topically.

Tq/W S/C

FIGURE 3

second subconjunctival injection) than after topical therapy. With one

exception, there was never a significant difference in viable bacterialcounts between the topical and subconjunctival routes with any drug-or-ganism combination at either treatment interval. The sole exception was

seen in the Pseudomonal ulcer group treated with gentamicin for ninehours, at which time treatment by the subconjunctival route was signifi-cantly more effective than topical therapy. This difference was no longerapparent after 17 hours of treatment.When gentamicin eye drops were administered at the higher concen-

tration of 14 mg/ml for nine hours, it was significantly more effective inreducing the colony count in the Pseudomonas ulcer group, but was no

more effective than gentamicin at the lower concentration at 3 mg/ml inthe staphylococcal ulcer group.

Antibiotic could not be detected in serum following topical administra-tion of drug. One-half hour following subconjunctival injection,serum levels of cefazolin and gentamicin ranged from 20.7 to 41.9 pug/mland 6.5 to 29.9 jxg/ml respectively. Aqueous levels of antibiotic are

displayed in Table VI.

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383

Baum

TABLE VI: AQUEOUS CONCENTRATIONS OF ANTIBIOTIC

AT 9 HOURS

Staphylococcus!cefazolinTopicalS/CContralateral

Staphylococcus!gentamicinTopical

3 mg/ml14 mg/ml

S/CContralateral

Pseudomonas/gentamicinTopical

3 mg/ml14 mg/ml

S/CContralateral

17.7 ± 10.0 (4)9.6 ± 2.9 (5)1.2 ± 0.4 (6)

5.3 ± 1.7 (6)11.0 ± 2.2 (4)46.4 ± 24.3 (6)4.9 ± 1.3 (6)

8.3 ± 0.6 (16)44.2 ± 6.4 (5)78.7 ± 18.3 (6)5.3 ± 0.9 (8)

AT 17 HOURS

6.5 ± 2.6 (11)0.8 ± 0.3 (8)

ND

3.3 ± 0.9 (11)

3.8 ± 1.0 (9)0.5 ± 0.1 (9)

8.8 ± 1.6 (9)

1.1 ± 0.3 (7)ND

ND = Not detectable.Values are means + SE (number of eyes).S/C = Subconjunctival.

Significant differences (P < 0.05)Table V were found:

Log1o CFU/corneaIA a not equal to b, c, d

e not equal to f, g, hIIA a not equal to b, b'

b not equal to de not equal to f, g, h

IIIA a not equal to b, b', cb not equal to b', c, db' not equal to de not equal to f, gf not equal to h

between the following groups in

Log1o antibiotic in corneaIB b not equal to c

f not equal to gIIB b not equal to b', c

f not equal to hIIIB b not equal to b', c

f not equal to g

Comparisons between the subconjunctival group and the contralateralgroup were not made.

DISCUSSION

TOPICAL VS CONTROL AND SUBCONJUNCTIVAL CONTROL

Both topical and subconjunctival administration of cefazolin and gentami-cin effectively reduce or eliminate sensitive bacterial pathogens from

384

Bacterial Corneal Ulcers

infected corneal tissue 9 and 17 hours after administration of drug com-pared to untreated animals. Past studies have documented a similarantibacterial effect of topical antibiotics on corneal pathogens.3"1844,59However, staphylococcal ulcers treated with subconjunctival gentamicinrequire 17 hours to show an effect of antibiotic. Although not shown inthis study, it would be reasonable to believe that administering antibioticby either of these routes over a longer duration would further reduce andprobably sterilize experimental bacterial corneal ulcers, providing thepathogen was sensitive to the antibiotic. We did not pursue this further,as bacterial counts were approaching zero after 17 hours.The antibiotic concentrations in this study demonstrate transient high

peaks and low troughs for both cefazolin and gentamicin after subconjunc-tival injection. Although drug concentration in the cornea following topi-cal administration never approaches the peak obtained after subconjunc-tival injection, it appears that, within the limits of the study, such peaksare not needed to eliminate bacteria effectively from a bacterial cornealulcer. Indeed, the cumulative effect of the repeated eye drop administra-tion was evident in the fact that levels by this route were slightly butsignificantly higher (P < 0.05) than with the subconjunctival route after17 hours of therapy for cefazolin in staphylococcal ulcers and gentamicinin Pseudomonal ulcers.

TOPICAL VS SUBCONJUNCTIVAL ADMINISTRATION

The principle conclusion of the present study is that for both cefazolinand gentamicin, the subconjunctival and topical routes are of similarefficacy in eliminating Staphylococcus aureus and Pseudomonas aerugi-nosa from a corneal ulcer in a rabbit model. The sole exception was seenin the Pseudomonal ulcer group treated with gentamicin for nine hours,at which time treatment by the subconjunctival route was significantlymore effective than topical therapy. This difference was no longer appar-ent after 17 hours of treatment. The findings in this study differ fromthose of Leibowitz et al,45 who reported no therapeutic effect from sub-conjunctival injections to rabbit eyes. In that study, six antibiotics wereemployed to treat bacterial keratitis induced by Staphylococcus aureus orPseudomonas aeruginosa. The present study used a similar protocol in-cluding frequency and technique of drug administration, and immediateirrigation of the ocular surface and conjunctival fornices with PBS follow-ing each subconjunctival injection of antibiotic. Drug dosage was alsosimilar wherever applicable. I am at a loss to explain why bacteria wereeffectively eliminated from the cornea after subconjunctival injection inthe present study while no effect was found in the study by Leibowitz et

385

al.45 The present study also documents the high corneal levels (< 100,ug/gm) of drug in the cornea while the infected cornea is being treated.Leibowitz et al did not study drug levels.

LOW VS HIGH CONCENTRATION OF ANTIBIOTICS

We administered gentamicin eye drops either as the 3 mg/ml commer-cially available concentration or in more concentrated form, 14 mg/ml.Cefazolin was administered at a single concentration of 33.3 mg/ml. Thehigh dose (14 mg/ml) gentamicin eye drops were significantly more effec-tivein reducing colony counts than the low dose (3 mg/ml) commerciallyavailable gentamicin eye drops over the nine hour treatment period forPseudomonal ulcer group, but not for the staphylococcal ulcer group.Others have found similar differences between low and high concentra-tions of antibiotic eye drops.3'9 The comparison was not extended to the17 hour treatment period because the low dose gentamicin eye drops,administered for 17 hours, successfully reduced the colony count to unde-tectable or near undetectable levels in both the staphylococcal and Pseu-domonal ulcer groups. Nonetheless, there was no significant differencebetween those ulcers treated with high dose eye drops and those treatedby subconjunctival injection. Therefore, since both routes are highlyeffective, the choice of routes should be based on other factors.

LOCAL VS SYSTEMIC ADMINISTRATION OF ANTIBIOTIC

Although a direct comparison between systemic (oral or parenteral) andlocal (topical or subconjunctival) administration of antibiotic was not per-formed in this study, our measurement of corneal drug levels allow anindirect comparison with data generated from other studies. As can beseen in Table III, administration of antibiotic by systemic routes producesa lower concentration of drug in the cornea than when the antibiotic isgiven either by eye drops or subconjunctival injection, and a lesser effi-cacy would be anticipated. Furthermore, since much less drug is given bythe topical and subconjunctival routes than by systemic administration,the incidence of systemic toxicity should be substantially decreased.

CLINICAL SIGNIFICANCE

I would like to extend the conclusions of the present study to disease inhumans, recognizing that such extrapolations must be made cautiously. Ifantibiotics administered frequently as eye drops are as effective in steril-izing a bacterial corneal ulcer as a subconjunctival injection, what shoulddictate the route of administration? In fact, in the treatment of bacterialcorneal ulcers, antibiotics are usually given as an eye drop and as a

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subconjunctival injection concomitantly. However, based on the resultsof this study, fortified antibiotic eye drops instilled every 15 to 30 minutesmight be employed alone in cases with mild, incipient corneal ulcers ifcompliance were assured. On the one hand, the subconjunctival injectioncauses more pain and apprehension than a regimen of eye drops. It is alsomore expensive. Subconjunctival injections, in contrast to eye drops,must be administered by a physician. On the other hand, compliance maybe a significant problem when either the patient or patient's family mustbe relied upon to instill the eye drops, especially when they must beadministered every 15 to 30 minutes around the clock during the first fewdays of therapy. Even a nurse is taxed to administer drug that often. It isdifficult and of questionable effectiveness to administer eye drops to asquirming, crying child. Tears may dilute the eye drops to an ineffectivedrug concentration and forceful closure of the eye lids reduces the admin-istered volume. It may be more effective to place the child under generalanesthesia and give a subconjunctival injection, once a da,y for four days.General anesthesia has risks. However, in the hands of a competentanesthesiologist, the risk of general anesthesia is minimal compared tothe greater risk of undertreating a bacterial corneal ulcer. The use ofketamine may be a reasonable alternative to general anesthesia.

SUMMARY

The various routes of antibiotic administration available to treat a bacteri-al corneal ulcer were reviewed and compared, and the pharmacokineticsand efficacy of each route analyzed in the rabbit. I then evaluated theefficacy of eye drops and subconjunctival injections in the treatment ofbacterial corneal ulcers for each of the following drug-organism combina-tions: staphylococcal ulcers/cefazolin, staphylococcal ulcers/gentamicinand Pseudomonas ulcers/gentamicin. Both topical and subconjunctivaladministration of each drug effectively reduced or eliminated the bacteriafrom infected corneal tissue. For both cefazolin and gentamicin, thesubconjunctival and the topical route were of similar efficacy in the treat-ment of Staphylococcal and Pseudomonal ulcers. Subconjunctival injec-tions produced high but transient peaks followed by persistent lowtroughs for both drugs. In contrast, eye drops produced moderate butsustained concentrations throughout the treatment period. Under thecircumstances studied, it appears that transient high peaks are notneeded to eliminate bacteria effectively from a bacterial corneal ulcer.Since both routes are equally effective, other factors should determinethe preferred route.

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