Studies on microbial flora of the conjunctival sac in Korean
[영문]In recent years, cross infections due to extensive use of chemotherapeutics, particularly of antibiotics and infections due to the decrease of tissue resistance from the abuse of corticosteriod preparations have been serious problems in clinical medicines, and the normal flors have been incriminal as causative agents in these diseased conditions.
Normal flora, resident or transient, are the microorganisms found in a given area of the skin or mucous membrane. They are considered to be nonpathogenic under the normal conditions, but their flourishing, as the result of host-parasite interactions, will depend directly or indirectly not only on biological and
physiological variations of the host but also on the changes in environmental factors that affect the daily activities of human life.
Weinstein (1946) showed that the incidences of development of cross infection due to the wide-spread application of antibiotics were similar to those of occurrence of anaphylactic reactions caused by antibiotic usages, and that major groups of the causative agents of such cross infections were antibiotic resistant organisms such as Pseudomonas, Proteus, Klebsiella pneumoniae, Alkaligenes facalis, Staphylococcus aureus, Candida, Aspergillus, Cryptococcus and so on.
Broad-spectrum antibiotics and corticosteroid preparations have also been widely used in the ophthalmological field. In Korea these drugs are readily available not only through the prescription by ophthalmologist but also through direct purchase
from the drug stores, and this results in the abuse of these medicines. Such abuse of antibiotics and corticosteroid preparations have brought in the problems of cross-infections and the decrease of tissue resistance which may cause emergence of
highly antibiotic-resistant organisms, superinfections and fungal infections, and ocular infections caused by Pseudomonas and fungi have gradually become important clinical entities in the field of ophthalmology.
Eyes, one of the exposed organ of human body, are prone to be contaminated with vaginal flora during birth (Grunberger and Kofler, 1954) and later in life normal flora of the eye will be established the contamination from the environments.
In this regard, studies have been made on the normal bacterial flora of conjunctival sacand eye lids as an effort to identify the causative agents of ocular infecions (Axenfeld, 1908; Lucic, 1927; Khorazo and Thompson, 1935; Bachrach et al., 1953; Barfoed, 1953; Grunberger and Kofler, 1954; Smith, 1954; Cason and
Winkler, 1954; Khristov, 1956; Doden, 1957; Chang, 1957; Yamazaki, 1958; Orfila and Courden, 1961; De Ocampo et al., 1965; Locatcher-Khorazo and Gutierrez, 1956, 1968; Allansmith et al., 1969; Matsuura, 1971). At the same time a number of mycological
studies have also been made on fungal flora of the eye (Fazakas, 1935, 1938, 1953; Mitsui and Hanabusa, 1955; Veirs and Davis, 1958; Hammeke and Ellis, 1960; Janke and Schwab, 1961; Agarwal and Khosla, 1963; Ovsepian and Osipian, 1965; Ainley and
Smith, 1965; Vasquez and Pereiro, 1965; Nama et al., 1966; Sinha and Das, 1968; Williamson et al., 1968; Wilson et al., 1969; White, 1969). Recently, cross infections and fungal infections due to the abuse of antibiotics and corticosteroid preparations have become the subjects of greater concern of many investigators, and increased number of reports have been made on the stuides of fungal infections of the eye.
It has been established that fungal flora of the normal conjunctival sac consisted of 43 genera including 83 species most of which are the inhabitants of soil and vegetables. Among these fungal flora, 25 genera including 38 species have been isolated from the patients with keratomycosis.
These results suggested that members of fungal flora of the conjunctiva could become the causative agents of mycotic infections of this organ. However, studies by Percebois and Raspiller (1968) and Wilson et al. (1969) showed that the
distribution of microbial flora of the normal conjunctiva was not constant and varied significantly in repeated cultivation experiments, and suggested that normal flora of the conjunctiva could affect the establishment of endogenous infections.
The first report on corneal infection by Aspergillus was made by Leber (1879), and Duke-Elder (1965) pointed out that the destruction of normal symbiosis between bacteria and fungi due to wide use of antibiotics and the infections caused by normal (fungal) flora due to the decrease of tissue resistance affected by increasing use of corticosteroid preparations could be responsible for recent gradual increases in fungal infections of the eye.
Mitsui and Hanabusa (1955) made a comparative study of fungal isolations from conjunctival sac among corticosteroid-treated and corticosteroid-untreated (control) groups, and the result showed that the fungi were isolated in 67% of corticosteroid-treated (topical administration) group and 28% of the control (corticosteroid-untreated).
Ley (1956) observed that topical abuse of antibiotics and corticosteroid preparations could result in the increase of mycotic intra-and extra-ocular infections. On the other hand, studies by Fine and Zimmerman (1959), Diamond and Kirk (1962) and Prenner and Theodore (1962) indicated that the fungi residing in
the conjunctival sac as normal flora could be introduced intraocularly either by perforating injury or intraocular surgical procedures and this would result in intraocular infection as a complication.
Fine and Zimmerman (1959) reported clinical cases of postoperative mycotic infecions of the eye. These investigators suggested that such infection could have resulted not by the contaminations of surgical instruments but by the infection
with fungal flora of conjunctival sac through the operating wound of the eye and emphasized the necessity of through irrigation of conjunctival sac prior to ocular surgery.
In order to provide basic informations on the causative organisma of ocular infections and to place the emphasis on fungal infections of the eye, studies were made on microbial flora, particularly fungi, of conjunctival sac among Korean, and
the effect of topical antibiotics and steriod treatment on the microbial flora of acute epidemic hemorrhagic conjunctivitis.
Materials and Methods
A. The sources of culture specimens
A total of 1,758 culture specimens were collected from 882 individuals who were without external diseases during one year period from June 1972 to July 1973. These specimens consisted of 387 from out-patients at the department of Ophthalmology,
Severance Hospital (211 specimens in June 1972 and 176 in July 1973), 200 from leprosy patients at the World Vision Special Skin Clinic, 198 from workers at textile industry company, 140 from workers at soy sauce manufacturing company, and 833 from school children and students (273 from primary school children, 282 from
middle school and 278 from high school students), and in order to evaluate the effect of topical administration of antibiotics and steroid preparations, 58 culture speciments were cultured from the patients with acute epidemic hemorrhagic conjunctivitis from the department of Ophthalmology, Severance Hospital in July
1. Collection of the specimens
Culture specimen was obtained with sterile cotton swab through a single gentle transeverse swabbing of exposed lower conjunctival fornix.
2. Isolation and identification of the bacteria
The collected specimen was streaked on blood agar plate and incubated aerobically at 37°. for 24 to 48 hours. The identification of the isolates on blood agar plates was made with colony morphology, Gram staining, and selected appropriate
tests for each organism by Bergey's manual (Breed et al., 1957).
3. Isolation and identification of the fungi.
The conjunctival was streaked on Sabouraud glucose agar plate containing chloramphenicol (100mg/L) and incubated at 25 °. from 1 to 4 weeks and followed by slide culture for microscopic observation. The identification of fungi isolated was made by the procedures of Thom and Raper (1945), Raper and Thom (1949), Lodder and Kreger-van Rij (1952), Barnett (1958), and Raper and Fennell (1965).
In order to establish normal flora of conjunctival sac among Korean, isolation and identification of bacteria and fungi was conducted with a total of 1,758 culture specimens obtained, during the period from June 1972 to July 1973, from 882
individuals without external disease, and to evaluate topical steroid and antibiotics treatment on the microbial flora of the conjunctiva, microbial flora was examined from 58 eyes of acute epidemic hemorrhagic conjunctivitis patients in July 1972. The culture specimens were collected from out-patients at the department of Ophthalmology, Severance Hospital, leprosy patients at the World Vision Special Skin Clinic, the workers at textile industry plant and soy sauce manufacturing company, school children and students (primary, middle and high school) in Seoul
area, and patients with acute epidemic hemorrhagic conjunctivitis who were administered topically more than 15 times a day with chloramphenicol (0.5%) and hydrocortison (0.5%) solutions for more than 2 weeks duration. The results are summarized as follows:
1. Of a total of 1,758 culture specimens collected, 736 specimens (41.3%) yielded bacterial growth and 214 specimens (12.2%) provided fungal growth. Among the specimens yielded positive growth, growth of single species either bacteria of
fungus was obtained from 841 specimens (47.8%), growth of 2 species from 175 specimens (9.9%) and none of them showed the growth of 3 or more species.
2. The incidences of isolation of bacterial species ranged from 32.4%, the lowest, and to 65.4% the highest, i.e., Severance Hospital; 65.4% in 1972 and 43.8% in 1973. leprosy turing company; 43.5%, primary school children; 35.5%, middle school students; 39.7% and high school students; 32.4%. Among the bacterial isolates, Staphylococcus epidermidis was the one with the highest frequency of isolation and followed by Diphtheroid and Staphylococcus aureus in the order of lower frequencies.
3. The incidences of isolation of fungal species ranged from 6.6%, the lowest to 19.9%, the highest, i.e., Severance Hospital; 19.9% in 1972 and 10.2% in 1973, leprosy patients; 10.5%, workers at textile industry plant; 6.6%, workers at soy sauce manufacturing company; 17.9%, primary school children; 12.5%, middle schoool students; 12.1% and high school students; 9.7%. Among the fungal isolates, Aspergillus species was the most frequent one and followed by Penicillium and Alternaria species in the order of lower frequencies.
4. No apparent sex difference was noted in the incidences of isolation of bacteria and fungi from the specimens of primary school children and middle and high school students.
5. Topical administration of chloramphenicol(0.5%) and hydrocortison (0.5%) solutions to patients with acute epidemic hemorrhagic conjunctivitis resulted in the lower incidence of bacteria and significantly higher incidence of fungi.