Identification And Quantitation Of Conjunctival Aerobic And Anaerobic Bacterial Flora In Microecosystem From Healthy People

Background and Aims Conjunctival sac, a semi-open lacouna, is parasitized with microflora all our lifetime as we were born , and this microbiota which do not always cause abnormal senses or sickness were called conjunctival normal flora in microecology. Normal flora play an important role in keeping our ocular surface healthy. Disease will be caused when the normal flora lose restriction each other or appear overbalance with the host. Therefore, overall and penetrating understanding on conjunctival flora will be significantly important in the field of infectious diseases of ocular surface, especially in the origin, prevention and cure of chronic infections. Normal flora are always changing and developing successionally, while show relative balance and stabilization with periods of different physiological stages. At present, the researches of conjunctival normal flora were mainly focused on prophylaxis of postoperative endophthalmitis using antibiotics topically in patients undergoing intraocular surgery, and the enumeration of conjunctival flora was mianly on the level of semi-quantitation. However, it is not accurate and lack of comparability to enumerate by the way of semi-quantitation. Therefore, we had no overall understanding of conjunctival normal flora because of lacking methods to quantitate them.This study is designed to identify conjunctival aerobic and anaerobic bacterial flora from the lower fornix of both eyes of 33 children, 31youths and 30 elders by collecting samples and carrying out bacterial culture.The conjunctival flora will be quantitated by counting the number of CFUs and weighing the samples, and then to investigate the species and quantity of normal conjunctival flora of people at different ages drawing assistance from statistical analysis from the viewpoint of microecology.In order to provide experimental data for the research of normal conjunctival flora and for the application and development of microecology in the field of ophthalmology. Methods1. 94 volunteers including 33 children (9.85±0.57 years old), 31youths (21.23±0.88 years old), and 30 elders (72.97±4.94 years old) were selected.2. Aerobic bacteria detection: Samples taken with moist swabs from the lower fornix of both eyes of volunteers were inoculated onto columbia blood agar (CBA) and then incubated in a atmosphere containing 5%CO2 at 37℃.Positive ones were identified and colony forming units (CFUs) were counted. Aerobic bacterial quantitation was expressed as the number of CFUs divided by the weight of each sample which was measured before and after collection.3. Anaerobic bacteria detection: Samples taken with moist swabs from the lower fornix of both eyes of volunteers were put into anaerobic glove box to inoculate samples onto brain-heart infusion (BHI) agar and then incubated in the box. Positive ones were done oxygen tolerance test, then obligate anaerobes were identified and colony forming units (CFUs) were counted. Anaerobic bacterial quantitation was expressed the same as arobic bacteria.Results1. Conjunctival aerobic bacteria: Staphylococcus epidermidis isolated from the elders (94.64%) and the youths (83.87%) all had a greater percentage than children (33.33%, P<0.05); Corynebacterium diphtheroides isolated from the elders (67.86%) had a higher isolation frequency than the youth (45.16%) and children (30.00%, P<0.05).2. The quantitation of conjunctival aerobic bacteria ranged from 1.14×102 to 3.63×105 CFU/g; Corynebacterium diphtheroides was the most numerous isolate from 188 eyes, and the elders had a larger number of which than the youths (P<0.05), the youths showed a grater number than children (P<0.05); Staphylococcus epidermidis was the second numerous bacteria, and the elders group had a grater number (P<0.01).3. Conjunctival anaerobic bacteria: Propionibacterium acnes isolated from the elders (79.31%) and youths (72.22%) all had a greater percentage than children (30.00%,P<0.05); Peptococcus niger isolated from the elders (44.83%) had a higher percentage than children (10.00%,P<0.05); Bacteroides fragilis isolated from the elders (13.79%) had a lower percentage than children (50.00%,P<0.05). 4. The quantitation of conjunctival anaerobic bacteria ranged from 1.89×102 to 8.67×104 CFU/g; Propionibacterium acnes was the most numerous isolate from 178 eyes, and the elders and the youths had a larger number of which than children (P<0.01); Peptococcus niger was the second numerous bacteria,and the elders group had a grater number (P<0.05).5. The isolation frequency of aerobic and anaerobic bacterial flora and the number of species isolated per eye from the elders were all larger than the youths and children (P<0.05); The isolation frequency and quantity of normal flora from different sex or eyes had no statistical difference (P>0.05).6. The quantity of whole aerobic and anaerobic bacterial flora showed no statistical difference (P>0.05); aerobic bacterial isolation frequency in 3 groups were all grater than that of anaerobic bacteria (P<0.05); and the number of aerobic species per eye from 3 groups were all larger than that of anaerobic ones (P<0.05).Conclusions1. The quantitation of conjunctival aerobic bacterial flora ranged from 102～105 CFU/g. With increasing age Staphylococcus epidermidis and Corynebacterium diphtheroides are found more frequently and in larger quantity.2. The quantitation of conjunctival anaerobic bacterial flora ranged from 102～104 CFU/g.With increasing age Propionibacterium acnes and Peptococcus niger are found more frequently and in larger quantity, while Bacteroides fragilis are demonstrated less frequently.3. The quantitation of conjunctival anaerobic bacterial flora was not higher than that of aerobic ones.While aerobic bacteria were found more frequently and showed more species.4. Characteristic changes appear in the species and quantitation of conjunctival flora at different stages of life.This conclusion may by helpful to illuminate the pathogenesy of ocular bacterial infectious diseases when we do reasch on the prevention and cure of them.