Antimicrobial susceptibility of canine normal conjunctival flora in Shiraz, Iran

Sarchahi, A.A.1, Haghkhah, M.2, and Molazem, M.3

1Departments of Clinical Studies, and 2 Pathobiology
3Graduated from School of Veterinary Medicine
School of Veterinary Medicine
Shiraz University
P.O.Box: 71345-1731
Shiraz, Iran
Fax: +98 711 6280707
1E. mail: sarchahi@shirazu.ac.ir

Summary

Bacteria were isolated from the conjunctival sacs of 39 (65%) of 60 clinically normal dogs. Organisms isolated included staphylococci, Bacillus spp, streptococci, Escherichia coli, Pseudomonas spp, Lactobacillus spp, Neisseria spp, Alcaligenes spp, Pasteurella spp, Klebsiella spp, and Francisella spp.
Susceptibility testing on the isolated bacteria to routine antibiotics (including: bacitracin, chloramphenicol, gentamicin, kanamycin, neomycin, oxytetracycline, penicillin G, sulfadiazine, tetracycline and tylosin) indicated that the most potent of these compounds was tylosin and the less one was bacitracin.

Key words: Bacteria, Dog, Eye, Normal flora and Antimicrobial susceptibility.

Introduction

The ideal basis for selection of an ocular antibiotic is an identification of the responsible organism and its antibiotic sensitivity. Often, however, obtaining this information cannot be justified either because of expense or because treatment must be instituted before the results are available. Therefore, a knowledge of the most likely organisms, their sensitivity and the most likely effective antibiotics is necessary. Because of the wide variety of organisms present, treating infections on such an empirical basis, although practical and unavoidable, dose not always lead to a satisfactory result (Slatter, 1981).
Knowledge of the most commonly isolated microorganisms in the canine eye in a geographic area and their antibiotic susceptibility is important to provide the most efficacious antimicrobial treatment (Gerding et al., 1988), because significant geographic differences occur in the types of organisms present (Slatter, 1981; Gerding and Kakoma, 1990). Thus the purpose of the study reported here was to determine the antimicrobial susceptibility of most bacteria isolated from the normal flora of dogs that were presented for check up examination and vaccination to Veterinary Clinic of Shiraz University.

Materials and Methods

Ocular specimens from both eyes of 60 dogs were collected over a 5-month period (May, 2003, to September, 2003). Samples were obtained from dogs presented to Veterinary Clinic of the School of Veterinary Medicine, Shiraz University, Iran, for Check up examination, vaccination, ovariohystrectomy and castration. The conjunctival sacs of sampled animals were normal on inspection and none of the dogs was receiving antimicrobial treatment. Specimens were obtained by rotating a sterile cotton swab (for each eye) directly into the conjunctival sac, then removing it while taking care to avoid contact with the lashes or skin of the lids. Topical anesthetics were not used prior to culturing, because of their potential toxic effect on microorganisms (Slatter, 1981). The swabs were transferred immediately to the laboratory and inoculated onto blood agar and MacConkey agar. After 24 hours of incubation at 37C, cultures were examined and subcultured if necessary. Multiple biochemical tests were used to identify the organism (Haghkhah et al., 2004). For susceptibility testing two colonies of isolated bacteria inoculated in nutrient broth and incubated at 37C for 3 hours to become the turbidity of that as 0.5 McFarland (Baron and Finegold, 1990). Then 0.1 ml of nutrient broth was transferred onto Muller Hinton agar and spread by a sterile swab. Antibiotic discs were placed with appropriate distance on the culture. The results of antibiogram was observed and recorded after 24 hours of incubation at 37C.

Results

Bacteria were isolated from conjunctival sac of 39 (65%) of the 60 dogs. The staphylococci including S. aureus 19 (30.16%) and coagulase-negative S. epidermidis 4 (6.35%)were isolated from 23 (36.51%) of 60 dogs. Bacillus spp was the next most frequent organisms isolated (26.98%). ? and ? hemolytic streptococci were isolated from 7.93% of the cases. Next in the order of isolation were Escherichia coli, Pseudomonas aeruginosa, Lactobacillus spp, Neisseria spp, Alcaligenes faecalis, Pasteurella canis, Klebsiella spp and Francisella tularensis.
The results of antibiogram testing on isolated bacteria are given in table 1. The results of susceptibility testing indicated that the relative potency of these compounds were tylosin> gentamicin> oxytetracycline> neomycin> chloramphenicol> tetracycline> kanamycin> sulfadiazine> penicillin> bacitracin.

Results of susceptibility testing of the most commonly isolated bacteria indicated that for Staphylococcus spp, most efficacious antimicrobial agents were tylosin, chloramphenicol, oxytetracycline, and gentamicin. For Streptococcus spp, most effective antimicrobial agent was chloramphenicol. For Neisseria spp, tylosin, oxytetracycline, sulfadiazine, gentamicin, chloramphenicol, and tetracycline were the most effective antimicrobial agents. Bacillus spp had susceptibility to sulfadiazine, tylosin, oxytetracycline, tetracycline, gentamicin, neomycin, chloramphenicol and kanamycin. Lactobacillus spp had the greatest susceptibility to bacitracin, and tylosin; Pseudomonas spp, to tylosin, and sulfadiazine; Klebsiella spp, to tylosin, oxytetracycline, gentamicin; E. coli to tylosin; and Pasteurella canis and Francisella tularensis to all used antimicrobial agents.

The antibiogram results in this study revealed that the most effective antibiotics on the isolated bacteria are tylosin and gentamicin and the less effective antibiotics are bacitracin and penicillin G.

Discussion

A combination of topical neomycin, polymyxin B and bacitracin is often used in dogs of noncomplicated external ocular infection when gram-positive cocci are identified in the scraping of conjunctiva for cytologic evaluations (Bistner, 1980; Peiffer et al., 1984), while in the present study most gram-positive cocci (staphylococci and streptococci) were not susceptible to neomycin and bacitracin but to chloramphenicol, tylosin and oxytetracycline. Thus in external ocular infections while waiting for culture results, chloramphenicol is recommended. Chloramphenicol has a broader spectrum and is notable for its ability to penetrate the ocular tissues (Peiffer et al., 1984). Because of these properties chloramphenicol is widely used in veterinary practice. However in simple disease processes, it is more suitable to use one of the bactericidal broad-spectrum mixtures (Slatter, 1981).
In dogs with gram-negative rods, polymyxin B and chloramphenicol (Peiffer et al., 1984), gentamicin, tobramycin or carbenicillin (Bistner, 1980) have been recommended. In the present study gram-negative bacteria were the most susceptible to tylosin, oxytetracycline, gentamicin and some to chloramphenicol. Thus, because there are not ophthalmic preparations of tylosin, in gram-negative infections, while waiting for culture results, oxytetracycline, gentamicin and chloramphenicol may be the preferred drugs.
Susceptibility testing on the most commonly isolated bacteria from the study by Gerding et al. indicated that for -hemolytic Streptococcus spp, the most efficacious antimicrobial agents were erythromycin, cephalothin, chloramphenicol, and carbenicillin. For -hemolytic Streptococcus spp, chloramphenicol, erythromycin, carbenicillin, and cephalothin were the most effective antimicrobial agents. Coagulase-positive Staphylococcus spp were most susceptible to cephalothin, gentamicin, bacitracin, and tobramycin, whereas coagulase-negative Staphylococcus spp had susceptibility to bacitracin, cephalothin, and gentamicin. Pseudomonas spp had the greatest susceptibility to tobramycin, gentamicin, polymyxin B, and amikacin (Gerding et al., 1988).
The antibiogram results in this study, were different from that reported by Gerding et al. The reasons for these differences are not known but may be due to different resistances of isolated bacteria in different geographical locations.
Because the most bacteria isolated in the present study were more susceptible to tylosin, gentamicin and oxytetracycline than the other antibiotics, before obtaining microbial susceptibility results, these antimicrobial drugs are recommended, although the choice of antimicrobial therapy before obtaining microbial susceptibility results is made on the basis of clinical signs, gram stains, and a history of previous antimicrobial treatment and the response to therapy (Gerding et al., 1988).

References

Baron, EJ and Finegold, SM (1990). Bailey and Scott’s Diagnostic Microbiology. 8th Ed. Mosby Co. pp: 172-173.
Bistner, S (1980). Ocular Therapeutics. In: Kirk, RW, (Ed), Current Veterinary Therapy. (7th ed.,), Philadelphia. WB Saunders Co. pp: 517-527.
Carter, GR and Cole, JR (1990). Diagnostic Procedures in Veterinary Bacteriology and Mycology. 5th Ed. Academic Press. pp: 485
Gerding, PA and Kakoma, I (1990). Microbiology of the canine and feline eye. Vet. Clin. North Am.: Small Anim. Pract. 20: 615-625.
Gerding, PA; McLaughlin, SA and Thoop, MW (1988). Pathogenic bacteria and fungi associated with external ocular diseases in dogs: 131 cases (1981-1986). J. Am. Vet. Med. Assoc. 193: 242-244.
Haghkhah, M; Sarchahi, AA; and Molazem, M (2004). Conjunctival flora of clinically normal dogs in Shiraz, Iran (Submitted).
Peiffer, RL; Cook, CS, and Muller, I (1984). Therapeutic strategies involving antimicrobial treatment of ophthalmic disease in small animals. J. Am. Vet. Med. Assoc. 185: 1172-1175.
Slatter, DH (1981). Ocular Pharmacology and Therapeutics. In: Slatter, DH (Ed), Fundamentals of Veterinary Ophthalmology. Philadelphia. W B Saunders Co. pp: 63, 80-81.
Table 1: Antimicrobial susceptibility of bacteria isolates from eyes of 60 healthy dogs*

Bacteria

AntibioticStaphylococcus sppBacillus sppStreptococcus sppE. coliPseudomonas aeruginosaLactobacillus sppNeisseria sppKlebsiella sppPasteurella canisFrancisella tularensisBacitracinRRRRRSIRSSChloramphenicolSSSIRRSISSGentamicinSSRIIISSSSKanamycinISRRRRIRSSNeomycinISRRRRIISSOxytetracyclineSSIIIRSSSSPenicillin GRRRRRRIRSSSulfadiazineRSRRSRSRSSTetracyclineRSIRRRSRSSTylosinSSISSSSSSSS, Sensitive; I, Intermediate sensitive; R, Resistant
* Based on “zone diameter interpretive standards” adapted from: Carter, GR and Cole, JR (1990)