key: cord-0813229-ka0d3xqp
authors: LIU, Shasha; YEUNG, Tiffany Lok Man; TSO, Eugene Yuk Keung; FUNG, Kitty Sau Chung; TANG, Emily Wai Ho; LI, Kenneth Kai Wang
title: Study of Conjunctival Carriage of SARS-CoV2 using Serial Sampling: Risk factors and Protective factors
date: 2021-01-13
journal: Can J Ophthalmol
DOI: 10.1016/j.jcjo.2021.01.003
sha: c749ff626ff58e45ee7a316aa3b9d1da19e6811f
doc_id: 813229
cord_uid: ka0d3xqp

OBJECTIVE: The conflicting results of conjunctival carriage of SARS-CoV2 in current literature may be due to variabilities in sampling technique and sampling time window. In this study, we set out to standardize the sampling times and sampling technique in order to minimize the risk of false negative results. DESIGN: A prospective observational study METHOD: Thirty-nine consecutive COVID19 patients admitted to a regional hospital in Hong Kong underwent 2 serial conjunctival swabs for RT-PCR analysis (one on admission and another prior to discharge). Using clear instructional video, standardized technique for conjunctival sampling was adopted,. Patient's medical history, ocular symptoms, nasopharyngeal SARS-CoV2 results and personal hygiene habits were prospectively collected and analyzed. RESULTS: The majority had mild to moderate COVID19 disease (89.74% of study subjects). 15.38% reported symptoms suggestive of ocular surface inflammation, including epiphora (10.25%), itchiness (7.69%) and foreign body sensation (5.13%). Only 1 (2.56%) case had bilateral overt conjunctivitis. There was no correlation between the COVID19 severity and ocular symptoms. Despite obtaining adequate samples, SARS-CoV2 was not detected in all 78 conjunctival samples. CONCLUSION: Even after adoption of standardized sampling technique, our study could not detect conjunctival carriage of SARS-CoV2 in mild to moderate COVID patients. However, our group of patients practiced good personal hygiene, with universal masking, frequent hand washing and mobile phone sanitation. Further studies to investigate the contribution of personal hygiene on conjunctiva transmission as well as self-inoculation is warranted.

To date, there remains many unknowns regarding ocular involvement in COVID19. Various studies have shown conflicting rates of conjunctival carriage of SARS-CoV2 (Table 1) . A cross-sectional study on 1,099 patients in China showed only 0.8% of COVID19 patients developed conjunctival congestion. 1 While a recent meta-analysis showed pooled prevalence of ocular manifestations among COVID-19 patients was 5.5%, and the sensitivity of detecting SARS-CoV-2 in ocular fluids was merely 0.6%. 2 The low detection rate may either suggest a low incidence of viral infiltration into ocular surface or due to variations in sampling technique, sampling time window and under-representation from critical cases. In order to provide an answer, we set out to standardize the sampling technique and sampling time in order to minimize the chance of false negative results and to gauge whether SARS-CoV2 can really infect the ocular surface.

We conducted a prospective study by adopting standardized technique for conjunctiva sampling via clear protocol and instructional video (see supplementary video). Adequate tissue sampling was ensured by sweeping lower tarsal conjunctiva twice. In case of unilateral eye redness, only the involved eye was sampled to avoid transmitting any microorganisms to the uninvolved eye.

Two serial eyeswabs were taken, one on admission (presentation sample) and another prior to hospital discharge (convalescent sample) in order to gauge whether viral shedding is present on the conjunctiva at the beginning and end of the COVID19 disease course. A total of 78 eyeswab samples were obtained from 39 consecutive COVID19 patients admitted to a regional hospital in Hong Kong.

Patient's medical history, ocular symptoms, nasopharyngeal SARS-CoV2 results and personal hygiene habits were prospectively collected and analyzed. The eyeswab specimens were tested for rRT-PCR using the E gene assay, where a 76 bp long fragment from a conserved region in the E gene would be detected by 

All 39 cases had proven COVID19 disease by respiratory samples for SARS-CoV2 RT-PCR ( Table 2 ). The mean age was 40 (ranged 19 to 72), with equal gender representation. Of note, most patients in our case series have mild to moderate COVID19 disease (89.74%).

15.38% (6/39) reported symptoms suggestive of ocular surface inflammation, including epiphora (10.25%), itchiness (7.69%) and foreign body sensation (5.13%). However only 1 (2.56%) case had bilateral overt conjunctivitis, which began 2 days after onset of mild URTI symptoms and spontaneously resolved after 1 week. There was no correlation between the COVID19 severity and ocular symptoms (p=0.308).

All cases had conjunctival sampling for SARS-CoV-2 on Day 1 post admission and before hospital discharge (Day 4-40). On average, the first swab was taken on Day 6 since the onset of COVID19 symptoms. The corresponding cycle threshold in matching nasopharyngeal samples was 29.81 on admission, which is indicative of intermediate to low viral load 3 . In all 78 conjunctival samples, SARS-CoV2 was not detected by RT-PCR. This was true across the spectrum of COVID19 disease severity.

There are two possible explanations for viral non-detection in the conjunctiva in our study.

Hypothetically, the various means of personal hygiene ( Table 2 ) observed in our patients could partially account for the low transmission rate onto the conjunctiva surface both prior to or during the COVID19 disease course (e.g. via self-inoculation). Due to the high level of vigilance in Hong Kong, the local residents have adopted stringent personal hygiene since early reports of COVID19 in January 2020. This was reflected in our study subjects, who had adopted universal face masking (100%), 4 times or more handwashings per day (100%), and daily mobile phone disinfection with alcohol wipes (76.92%). Other ocular protection included 76.92% wore spectacles while 7.69% more worn goggles. Regarding risk factors of ocular transmission, 28.95% of study subjects wore contact lenses, including the patient with conjunctivitis, who worn daily disposable contact lenses. We believe that the simpler hypothesis of the ocular surface being infected by the virus (either via droplet transmission or via self-inoculation) is more likely. The risk of self-inoculation exists. As shown by a cross-sectional study of 534 patients, COVID19 seldom presents initially with conjunctivitis (0.56%). In contrast, 4.12% developed conjunctivitis after systemic symptoms. 6 A COVID19 case was also reported to develop follicular conjunctivitis with positive conjunctival carriage not at the beginning but late in the disease (Day 13), amidst antiviral treatments. 7 This raises the possibility that the virus is subsequently introduced into patients' eyes via self-inoculation. Therefore, good personal hygiene throughout the disease course would theoretically lower the chance of self-inoculation.

In summary, our study found that conjunctival carriage of SARS-CoV2 is low even in place of standardised sampling technique. This is probably good news for ophthalmic healthcare workers. Nevertheless, adequate eye protection and stringent personal hygiene are still important as they may contribute to further reduce ocular COVID19 transmission to minimal.

The current study is limited by the lack of a control group to assess the protective and risk factors of viral transmission of COVID19 via the conjunctiva. Future case-controlled studies and the use of a validated personal hygiene questionnaire 8 would yield further fruitful findings. As there are reports of positive conjunctiva swab for SARS-CoV2 in the absence of ocular symptoms (Table 1 ) and vice versa. The relationship between the presence of SARS-CoV2 on the conjunctiva and ocular symptoms remains to be elucidated. In addition, molecular study also suggested the possibility of the cornea being an ocular site of infection instead of conjunctiva. 9 Future larger-scale studies aimed at investigating these issues could provide more insight.

No conflicting relationship exists for any author Table 1 . Summary of existing studies on the presence of SARS-CoV2 in ocular secretions 

Clinical characteristics of coronavirus disease 2019 in China

The prevalence of ophthalmic manifestations in COVID-19 and the diagnostic value of ocular tissue/fluid

Direct RT-qPCR detection of SARS-CoV2 RNA from patient nasopharyngeal swabs without an RNA extraction step 2020

Is COVID-19-related Guillain-Barré syndrome different

Ocular manifestations and clinical characteristics of 534 cases of COVID-19 in China: A cross-sectional study

Ocular manifestations of a hospitalised patient with confirmed 2019 novel coronavirus disease

Do various personal hygiene habits protect us against influenza-like illness?

Expression of SARS-CoV-2 receptor ACE2 and TMPRSS2 in human primary conjunctival and pterygium cell lines and in mouse cornea

Evaluation of coronavirus in tears and conjunctival secretions of patients with SARS-CoV2 infection

Characteristics of Ocular Findings of Patients With Coronavirus Disease

Ocular Findings and Proportion with Conjunctival SARS-COV-2 in COVID-19 Patients

Assessing Viral Shedding and Infectivity of Tears in Coronavirus Disease 2019 (COVID-19) Patients

Ophthalmologic evidence against the interpersonal transmission of 2019 novel coronavirus through conjunctiva

Tropism, replication competence, and innate immune responses of the coronavirus SARS-CoV-2 in human respiratory tract and conjunctiva: An analysis in ex-vivo and in-vitro cultures

Detection of SARS-CoV-2 in conjunctival secretions from patients without ocular symptoms