Infectious endophthalmitis is one of the most serious complications of ocular surgery. Therefore, preventing post-cataract surgery endophthalmitis is a matter of great concern for ophthalmologists. The pathogens causing infectious endophthalmitis primarily originate from the ocular surface, such as bacteria from the conjunctival sac, eyelids, and meibomian glands[12, 13]. These microorganisms can be reduced through disinfection methods. However, even with the most rigorous disinfection protocols currently available, it is not possible to completely eliminate them[14]. Adequate knowledge of the characteristics of the bacterial microbiota in the conjunctival sac in patients is crucial for us to improve measures to prevent postoperative endophthalmitis. In this study, we investigated the composition of conjunctival bacteria in recent middle-aged and elderly patients and analyzed the systemic clinical factors associated with positive conjunctival bacterial cultures.
In recent years, an increasing number of studies have shown that the conjunctival sac of normal individuals harbors a microbiota. These bacteria coexist within the conjunctival sac, maintaining a dynamic equilibrium and inhibiting the growth and invasion of pathogenic bacteria[3]. Staphylococcus epidermidis, Streptococcus spp, Staphylococcus aureus, Propionibacterium acnes, Corynebacterium spp, Streptococcus spp, and Haemophilus influenzae are the most commonly isolated microorganisms in the normal conjunctival microbiota. Among them, earlier studies found that Staphylococcus aureus had the highest isolation rate among numerous studies. Additionally, anaerobic bacteria and fungi may also be occasionally isolated[13, 15, 16].
The bacterial isolation rate in this study was 36.11%. The detection rate is consistent with literature report[3, 17]. A total of 122 strains were detected in the conjunctival sac of 78 patients with positive bacterial culture. A total of 60 Gram-positive rods were detected, accounting for 49.18% of the total, while 55 Gram-positive cocci were detected, constituting 45.08%. Gram-negative bacteria were detected in 6 samples, comprising 4.92% of the total, whereas only 1 fungal species was identified, representing 0.82%. This study revealed that the most abundant genera in the conjunctival sac were Corynebacterium (52%), Staphylococcus (38%), Micrococcus (9.84%), Acinetobacter (4.10%) and Bacillus (3.28%). There are still some discrepancies between our research findings and previous reports. These differences may be associated with variations in sample collection methods, detection techniques as well as the living environment, lifestyle, and physiological factors of the study participants.
As the gold standard for microbial detection, clinical laboratories commonly employ culture-based methods to identify samples at the species level and measure bacterial density. However, the results are often influenced by factors such as culture conditions and incubation time. In this study, 16S rDNA gene sequencing was utilized, which has the capability to identify unculturable microorganisms, providing an efficient, comprehensive, and accurate research approach for microbial community composition[9]. The study by Zhou et al.[18] revealed that Corynebacterium, Streptococcus, Propionibacterium, Bacillus, and Staphylococcus were the top five abundant genera in the healthy conjunctival sac. Huang et al.[4]found that the most abundant genera in the normal conjunctival sac were Corynebacterium (28.22%), Pseudomonas (26.75%), Staphylococcus (5.28%), Acinetobacter (4.74%), and Streptococcus (2.85%). They also employed the method of 16S rDNA gene sequencing, and their findings were similar to ours. These collective research findings suggest that the isolation rate of Corynebacterium in the conjunctival sac may be gradually increasing in recent years, possibly surpassing Staphylococcus as the most abundant genus in the conjunctival sac. As one of the “core resident microbial communities” on the ocular surface, Corynebacterium has the function of enhancing eye immune balance and host defense. Studies have shown that this genus can induce γδT cells in the ocular mucosa to produce commensal-specific interleukin-17. This response is central to local immunity, as it promotes the recruitment of neutrophils and the release of antibiotics in tears, protecting the eyes from pathogenic bacterial infections[19]. Ge et al.'s study[20] confirmed that the decrease in the abundance of Corynebacterium is associated with fungal keratitis.
In this study, the genus Micrococcus also caught our attention, with a significantly increased isolation rate compared to previous studies. Micrococcus is commonly found in the skin, soil, and can also be isolated from food and air. It is considered an opportunistic pathogen, capable of causing local tissue infections such as wounds, but can also lead to severe infections such as endocarditis and brain abscesses[21, 22]. An et al.[23] utilized 16S rDNA sequencing to examine the conjunctival microbial community in 18 healthy adults. They found a relatively high isolation rate of the genus Micrococcus (22.2%), ranking third after Staphylococcus and Corynebacterium. This finding aligns with our results. Wang et al. found that after three days of preoperative use of levofloxacin eye drops, Staphylococcus epidermidis, Kocuria rosea, and Micrococcus luteus were the top three strains with the highest positive culture rates[24]. This suggests that preoperative use of antibiotics does not completely eliminate the presence of the Micrococcus in the conjunctival sac, which may result in opportunistic eye infections after surgery. The increased isolation rate of the Micrococcus in the conjunctival sac should raise concerns among ophthalmologists and prompt further exploration of safer and more effective preoperative disinfection strategies.
This study found that the isolation rate of bacteria in the conjunctival sac showed an increasing trend with age. The population aged 80 years and above had the highest bacterial isolation rate (70.0%). In other words, elderly patients are at a higher risk of positive conjunctival sac cultures before cataract surgery. Our study also demonstrated that certain clinical factors, such as being female, having meibomian gland dysfunction (MGD), diabetes, and recent history of infections, were associated with a higher bacterial load in the conjunctival sac. This has been confirmed in previous studies as well [2, 3, 24].
Due to the ongoing COVID-19 period, wearing masks has become a necessity in hospitals and public places. Another important finding in our study was that individuals who did not have the habit of regularly changing their masks had a significantly higher isolation rate of bacteria in the conjunctival sac compared to those who had this habit. Therefore, we speculate that mask-wearing habits can influence the microbial composition in the conjunctival sac. Barbara B.B et al.[8] conducted a retrospective study on the impact of facial masks during the COVID-19 period on the ocular surface. Prolonged mask-wearing indeed increases the likelihood of ocular irritation and discomfort symptoms, leading to more ocular surface diseases such as dry eye and corneal epithelial damage. Previous studies have found that the microbial subgroups on both the inner and outer surfaces of masks can lead to changes in facial and gut microbiota [25, 26]. During the pandemic, there was a significant accumulation of lactobacilli and Corynebacterium in the gut, and the abundance of Bacteroides gradually increased, resulting in significant differences in bacterial species before and during the outbreak. This study also found a significant increase in the isolation rate of Corynebacterium in the conjunctival sac of the current population compared to before. Additionally, facial microbiota diversity decreased [27], but the pathogenicity (functionality) of facial microbiota significantly increased, leading to more skin issues. Alterations in gut microbiota (ecological disruption) can contribute to various eye diseases [28] such as uveitis and dry eye disease. Since there exists a gut-eye or gut-eye-lacrimal gland microbiota axis, disruptions in gut microbiota may also lead to changes in ocular surface microbiota[29].
In addition to the aforementioned factors, the impact of masks on the microbial composition in the conjunctival sac may also involve the following factors. Firstly, during mask-wearing, exhalation generates an upward airflow [8]. This upward airflow disrupts the lipid layer of the tear film, accelerating the evaporation of the ocular surface tear film. As tears serve as a natural barrier on the ocular surface, the disruption of this barrier makes it easier for bacteria to adhere. Moreover, tears contain abundant antimicrobial substances such as lysozyme, cationic antimicrobial peptides, and surfactant protein D [30]. When tear film stability decreases, the antimicrobial abilities of these molecules may be compromised, leading to changes in the bacterial population in the conjunctival sac. Secondly, the upward airflow contains a high concentration of carbon dioxide (4–5% in exhaled gas) compared to inhaled air (0.4%), which can lead to decreased corneal nerve sensitivity [31]. Additionally, the upward airflow generated during exhalation can lead to changes in the temperature around the eyes. Kapelushnik et al. [32] investigated the changes in ocular surface temperature during each respiratory cycle while wearing surgical masks and observed a significant increase of approximately 0.5°C, particularly at the eyelid margin. This temperature elevation persists over a prolonged duration, leading to a decrease in tear film stability, barrier disruption, and increased susceptibility to bacterial colonization. Although there is no direct research on the impact of temperature on ocular surface microbiota, Juan Sepulveda et al. [33] found that temperature alterations increased gut microbiota diversity in animals. This suggests that ocular microbiota may also adapt to specific temperature conditions, and temperature changes may affect the adaptability of resident microbiota to the host, leading to disruption of the existing microbiome and the emergence of new resident microbial populations.
There are limitations to this study. Firstly, our sample size was still limited, and individuals’ microbial compositions can vary significantly. More samples from larger regions, different seasons, and various age groups are needed. Secondly, the influence of occupational exposure and living environment on the conjunctival sac microbiota was not further investigated. Thirdly, the differences between microbial compositions inside masks and in the conjunctival sac were not examined. Finally, further research is needed to explore the drug sensitivity characteristics of the conjunctival sac microbiota at the current stage, providing insights for optimizing clinical preoperative antimicrobial strategies.