Etiology and antibacterial susceptibility pattern of community-acquired bacterial ocular infections in a tertiary eye care hospital in south India

Aims: To identify the etiology, incidence and prevalence of ocular bacterial infections, and to assess the in vitro susceptibility of these ocular bacterial isolates to commonly used antibiotics. Materials and Methods: Retrospective analysis of consecutive samples submitted for microbiological evaluation from patients who were clinically diagnosed with ocular infections and were treated at a tertiary eye care referral center in South India between January 2002 and December 2007. Results: A total of 4417 ocular samples was submitted for microbiological evaluation, of which 2599 (58.8%) had bacterial growth, 456 (10.3%) had fungal growth, 15 (0.34%) had acanthamoebic growth, 14 (0.32%) had mixed microbial growth and the remaining 1333 (30.2%) had negative growth. The rate of culture-positivity was found to be 88% (P < 0.001) in eyelids’ infection, 70% in conjunctival, 69% in lacrimal apparatus, 67.4% in corneal, 51.6% in intraocular tissues, 42.9% in orbital and 39.2% in scleral infections. The most common bacterial species isolated were Staphylococcus aureus (26.69%) followed by Streptococcus pneumoniae (22.14%). Sta. aureus was more prevalent more in eyelid infections (51.22%; P = 0.001) coagulase-negative staphylococci in endophthalmitis (53.1%; P = 0.001), Str. pneumoniae in lacrimal apparatus and corneal infections (64.19%; P = 0.001), Corynebacterium species in blepharitis and conjunctivitis (71%; P = 0.001), Pseudomonas aeruginosa in keratitis and dacryocystitis (66.5%; P = 0.001), Haemophilus species in dacryocystitis and conjunctivitis (66.7%; P = 0.001), Moraxella lacunata in blepharitis (54.17%; P = 0.001) and Moraxella catarrhalis in dacryocystitis (63.83%; P = 0.001). The largest number of gram-positive isolates was susceptible to moxifloxacin (98.7%) and vancomycin (97.9%), and gram-negative isolates to amikacin (93.5%) and gatifloxacin (92.7%). Conclusions: Gram-positive cocci were the most frequent bacteria isolated from ocular infections and were sensitive to moxifloxacin and vancomycin, while gram-negative isolates were more sensitive to amikacin and gatifloxacin.

suppurative scleritis, canaliculitis, keratitis, dacryocystitis, preseptal cellulitis, endophthalmitis and panophthalmitis, and treated at a tertiary eye care referral center located at Tirunelveli district, Tamil Nadu, South India, between January 2002 and December 2007. All the patients were examined on the slit-lamp biomicroscope and infective diseases included in this study were diagnosed clinically by a group of ophthalmologists. [4,5] After detailed ocular examinations using standard techniques, [15,16] specimens for culture and smear were obtained by scraping the eyelid margin using a sterile blade (#15) on a Bard-Parker handle and by swabbing the lid margins with sterile broth-moistened cotton swabs in cases of blepharitis. Similarly, specimens were also obtained from the corneal ulcers by scraping. For cases of suppurative scleritis, specimens were collected by scraping and swabbing the area of the suppurative abscess. Conjuctival cultures were obtained by wiping a brothmoistened swab across the lower conjunctival cul-de-sac in conjunctivitis cases, and thick, tenacious purulent punctal discharge was collected from the canaliculus by pressure applied over the area of the eyelid that overlies the canaliculus in cases of canaliculitis. In cases of external and internal hordeolum, the abscesses were incised and the drained pus was obtained. From the cases of dacryocystitis, purulent material was collected from everted punta by applying pressure over the lacrimal sac area, and from the surgically excised lacrimal sac. Specimens from cases of preseptal cellulitis were obtained after stab incision or through an open wound or drainage site, if present. In patients in whom infectious endophthalmitis and panophthalmitis are suspected, lid and conjunctival specimens along with anterior chamber and vitreous fluids were obtained.

Results
Of 2599 eyes with bacterial growth alone, 2587 (99.54%) had infection with single species of bacteria and the remaining 12 (0.46%) had infection with two species of bacteria, and thus, a total of 2611 (2587 + 24) bacterial isolates was recovered [ Table 3]. The predominant bacterial species isolated was Sta. aureus (26.69%; 697 of 2611), followed by Str. pneumoniae  Table 4].
Significantly more number of Sta. aureus was recovered from eyes with eyelid infections (51.22%; 357 of 697 total Sta. aureus isolates) than from eyes with other ocular infections Bharathi, et al.: Bacteriology of ocular infections

Discussion
A combination of mechanical, anatomic, immunologic and microbiologic factors prevents ocular infections and do not allow the survival of pathogenic species in the eye. [18,19] However, in certain circumstances, they gain access to the Bharathi, et al.: Bacteriology of ocular infections eye and cause a variety of infections. Prompt and specific therapy can be instituted if the microbes can be isolated and their susceptibility to the antimicrobials is known. However, the ability to isolate the causative organism depends on a variety of factors including the amount of inoculum, [20] the site from which it is taken, the media used for culture (whether enriched media are used or not) [21] and also on the empirical treatment received before collection of the samples. [22] Hence, the culture-positivity varies from center to center. In this study, the overall culture-positivity was 69.8%. We found the highest rate of culture-positivity among the samples collected from eyelid infections probably due to two reasons, that is, being the outermost defense mechanism it harbors a large number of microorganisms and the amount of inoculum is also sufficient to inoculate the various media.
In this study, bacteria (58.8%) were the most common pathogens and were involved in infections of all the tissues of the eye, whereas fungi (10.3%) caused keratitis and endophtalmitis and Acanthamoeba (0.34%) caused only keratitis. The most common bacteria isolated from ocular specimens were Sta. aureus (26.69%) followed by Str. pneumoniae (22.14%). Sta. aureus caused infections of the eyelids (52.57%), conjunctiva (40.45%) and canaliculus (33.85%), whereas Str. pneumoniae caused lacrimal sac (31.35%) and corneal infections (37%) and CoNS, postoperative (68.27%) and posttraumatic endophthalmitis (33%). Though Staphylococci and Streptococci along with other bacteria like Corynebacterium, Haemophilus, Moraxella and Neisseria are part of the normal flora of the conjunctiva, under appropriate conditions they cause infections. [2,23,24] Sta. aureus is commonly involved in primary pyoderma and acts as a secondary invader on diseased skin. It produces coagulase, a factor capable of clotting the plasma which may play a role in the development of staphylococcal abscess by producing local fibrin thrombi that protect organisms and concentrate toxic factors. [25] CoNS elaborate a surface slime that facilitates adherence to the surface and may play a role in the pathogenesis of endophthalmitis. The surface slime protects the organism from phagocytosis and the action of antimicrobial agents. CoNS, especially Staphylococcus epidermidis is the commonest cause for postoperative endophthalmitis. [6,7] Being a Vol. 58 No. 6 normal inhabitant of the upper respiratory tract, Str. pneumoniae is frequently found in the lacrimal apparatus and conjunctiva. [2,3] Any minor corneal epithelial disruption facilitates invasion of the bacteria, hence causing corneal ulcer.
Among gram-negative bacilli, the most common pathogen was Pseudomonas spp. (8.4%), followed by Haemophilus spp. been attributed to the action of proteases and glycocalyx that allow the organisms that adhere to the host cells forming micro colonies that resist phagocytosis. [26] Natural pathogenicity of Haemophilus appears to be directly related to the capsule formation which renders resistance to complement-mediated immunity. [27] The gram-negative bacilli, Klebsiella spp., Enterobacter spp., Citrobacter spp., Proteus spp., Serratia spp. etc., are found in soil and sewage and are opportunistic pathogens causing conjunctivitis, keratitis, dacryocystitis, orbital cellulitis and endophthalmitis when the host defenses are low. [4,5] Among the gram-negative coccobacilli, the predominant isolate, M. catarrhalis demonstrated 3.6% of incidence and was more frequently present in dacryocystitis (63.8%; 60 of 94), Bharathi, et al.: Bacteriology of ocular infections      whereas M. lacunata was prevalent more in blepharitis (54.2%; 26 of 48), Neisseria in conjunctivitis (57%; 8 of 14). M. cararrhalis, a constant inhabitant of the respiratory tract, tends to cause dacryocystitis and less frequently meibomitis, conjunctivitis, keratitis and rarely postoperative endophthalmitis. [28] M. lacunata is commonly found in hot and dry areas of the world and causes angular conjunctivitis in alcoholics and debilitated patients. [29] Neisseria spp. infects mucosa of genitourinary tract and conjunctiva of neonates, adolescents and adults. [28] The filamentous bacteria, Nocardia, accounted for 2% of the incidence and its prevalence was 49% in keratitis and 20.7% in canaliculitis. In comparison, the incidence of gram-positive bacilli was 9%, of which Corynebacterium spp. accounted a higher rate of prevalence in blepharitis (35.7%; 74 of 207) and conjunctivitis (35.3%; 73 of 207), whereas Bacillus species was present in conjunctivitis (40.7%; 11 of 27) and in post-traumatic endophthalmitis (26%; 7 of 27). Nocardia infection usually occurs following trauma with objects contaminated with soil, and there have been sporadic reports of conjunctivitis, dacryocystitis, canaliculitis, scleritis, keratits, episcleral granuloma and endophthalmitis. [30] Corynebacterium spp. are almost constant saprophytes in the conjunctiva, however, Corynebacterium diphtheriae causes severe membranous conjunctivitis associated with pharyngeal diphtheria. [31] Bacillus spp. are ubiquitous in nature and are known to cause severe endophthalmitis following penetrating injury with metallic or vegetative foreign bodies and also by endogenous spread in drug abusers. [32] Resistance and sensitivity based on in vitro testing may not reflect true clinical resistance and response to an antibiotic because of the host factors and penetration of the drug. In this study, moxifloxacin and vancomycin revealed a higher efficacy against gram-positive isolates compared with other antibacterial agents. Vancomycin is a glycopeptide; it inhibits early stages in cell wall mucopeptide synthesis and it exhibited greatest potency against ocular gram-positive isolates. Moxifloxacin was specifically developed with methoxy group in the C-8 position and bicyclic side-chain in the C-7 position, which was specifically engineered to increase the potency and further inhibit bacterial resistance by hindering the cell's efflux pump mechanism, increasing the drug's length of stay within bacterial cells. Recent studies have also shown the excellent gram-positive coverage of moxifloxacin in ocular infections. [33] However, moxifloxacin has incomplete coverage against gramnegative isolates. We found greatest coverage of gatifloxacin and amikacin against gram-negative isolates. Ciprofloxacin and ofloxacin were introduced earlier and have been widely used since 1990, whereas gatifloxacin's usage has started in recent years. In addition to methoxy side chain at the C-8 position, gatifloxacin carries a methyl group on the piperazinyl ring. There was a slight decrease in all pathogens' susceptibilities to ciprofloxacin and ofloxacin, with a subsequent increase in the efficacy of gatifloxacin. [33] The relationship between antibiotic use and resistance is complex. Improper selection of antibiotics, inadequate dosing and poor compliance to therapy may play as important a role in increasing resistance as their overuse. This report documents the prevalence of bacterial species causing ocular infections in South India. The information provided in this article would aid the clinician in formulating rationalebased decisions in the antibiotic treatment of bacterial ocular infections that cause major public health problems.