Oral administration of S-nitroso-L-glutathione (GSNO) provides anti-inflammatory and cytoprotective effects during ocular bacterial infections.

Bacterial endophthalmitis is a severe complication of eye surgeries that can lead to vision loss. Current treatment involves intravitreal antibiotic injections that control bacterial growth but not inflammation. To identify newer therapeutic targets to promote inflammation resolution in endophthalmitis, we recently employed an untargeted metabolomics approach. This led to the discovery that the levels of S-nitroso-L-glutathione (GSNO) were significantly reduced in an experimental murine Staphylococcus aureus (SA) endophthalmitis model. In this study, we tested the hypothesis whether GSNO supplementation via different routes (oral, intravitreal) provides protection during bacterial endophthalmitis. Our results show that prophylactic administration of GSNO via intravitreal injections ameliorated SA endophthalmitis. Therapeutically, oral administration of GSNO was found to be most effective in reducing intraocular inflammation and bacterial burden. Moreover, oral GSNO treatment synergized with intravitreal antibiotic injections in reducing the severity of endophthalmitis. Furthermore, in vitro experiments using cultured human retinal Muller glia and retinal pigment epithelial (RPE) cells showed that GSNO treatment reduced SA-induced inflammatory mediators and cell death. Notably, both in-vivo and ex-vivo data showed that GSNO strengthened the outer blood-retinal barrier during endophthalmitis. Collectively, our study demonstrates GSNO as a potential therapeutic agent for the treatment of intraocular infections due to its dual anti-inflammatory and cytoprotective properties.

Aspergillus Endophthalmitis: Epidemiology, Pathobiology, and Current Treatments.

Fungal endophthalmitis is one of the leading causes of vision loss worldwide. Post-operative and traumatic injuries are major contributing factors resulting in ocular fungal infections in healthy and, more importantly, immunocompromised individuals. Among the fungal pathogens, the Aspergillus species, Aspergillus fumigatus, continues to be more prevalent in fungal endophthalmitis patients. However, due to overlapping clinical symptoms with other endophthalmitis etiology, fungal endophthalmitis pose a challenge in its diagnosis and treatment. Hence, it is critical to understand its pathobiology to develop and deploy proper therapeutic interventions for combating Aspergillus infections. This review highlights the different modes of Aspergillus transmission and the host immune response during endophthalmitis. Additionally, we discuss recent advancements in the diagnosis of fungal endophthalmitis. Finally, we comprehensively summarize various antifungal regimens and surgical options for the treatment of Aspergillus endophthalmitis.

SARS-CoV-2 and its beta variant of concern infect human conjunctival epithelial cells and induce differential antiviral innate immune response.

PURPOSE: SARS-CoV-2 RNA has been detected in ocular tissues, but their susceptibility to SARS-CoV-2 infection is unclear. Here, we tested whether SARS-CoV-2 can infect human conjunctival epithelial cells (hCECs) and induce innate immune response. METHODS: Conjunctival tissue from COVID-19 donors was used to detect SARS-CoV-2 spike and envelope proteins. Primary hCECs isolated from cadaver eyes were infected with the parental SARS-CoV-2 and its beta variant of concern (VOC). Viral genome copy number, and expression of viral entry receptors, TLRs, interferons, and innate immune response genes were determined by qPCR. Viral entry receptors were examined in hCECs and tissue sections by immunostaining. Spike protein was detected in the cell culture supernatant by dot blot. RESULTS: Spike and envelope proteins were found in conjunctiva from COVID-19 patients. SARS-CoV-2 infected hCECs showed high viral copy numbers at 24-72h post-infection; spike protein levels were the highest at 24hpi. Viral entry receptors ACE2, TMPRSS2, CD147, Axl, and NRP1 were detected in conjunctival tissue and hCECs. SARS-CoV-2 infection-induced receptor gene expression peaked at early time points post-infection, but gene expression of most TLRs peaked at 48 or 72hpi. SARS-CoV-2 infected hCECs showed higher expression of genes regulating antiviral response, RIG-I, interferons (α, ß, & λ), ISG15 & OAS2, cytokines (IL6, IL1ß, TNFα), and chemokines (CXCL10, CCL5). Compared to the parental strain, beta VOC induced increased viral copy number and innate response in hCECs. CONCLUSIONS: Conjunctival epithelial cells are susceptible to SARS-CoV-2 infection. Beta VOC is more infectious than the parental strain and evokes a higher antiviral and inflammatory response.

Prevalence of SARS-CoV-2 in human post-mortem ocular tissues.

BACKGROUND: SARS-CoV-2 is found in conjunctival swabs and tears of COVID-19 patients. However, the presence of SARS-CoV-2 has not been detected in the human eye to date. We undertook this study to analyze the prevalence of SARS-CoV-2 in human post-mortem ocular tissues. METHODS: The expression of SARS-CoV-2 RNA was assessed by RT-PCR in corneal and scleral tissues from 33 surgical-intended donors who were eliminated from a surgical use per Eye Bank Association of America (EBAA) donor screening guidelines or medical director review or positive COVID-19 test. Ocular levels of SARS-CoV-2 RNA (RT-PCR), Envelope and Spike proteins (immunohistochemistry) and anti-SARS-CoV-2 IgG and IgM antibodies (ELISA) in blood were evaluated in additional 10 research-intent COVID-19 positive donors. FINDINGS: Of 132 ocular tissues from 33 surgical-intended donors, the positivity rate for SARS-CoV-2 RNA was ~13% (17/132). Of 10 COVID-19 donors, six had PCR positive post-mortem nasopharyngeal swabs whereas eight exhibited positive post-mortem anti-SARS-CoV-2 IgG levels. Among 20 eyes recovered from 10 COVID-19 donors: three conjunctival, one anterior corneal, five posterior corneal, and three vitreous swabs tested positive for SARS-CoV-2 RNA. SARS-CoV-2 spike and envelope proteins were detected in epithelial layer of the corneas that were procured without Povidone-Iodine (PVP-I) disinfection. INTERPRETATIONS: Our study showed a small but noteworthy prevalence of SARS-CoV-2 in ocular tissues from COVID-19 donors. These findings underscore the criticality of donor screening guidelines, post-mortem nasopharyngeal PCR testing and PVP-I disinfection protocol to eliminate any tissue harboring SARS-CoV-2 being used for corneal transplantation.

Aging, But Not Sex and Genetic Diversity, Impacts the Pathobiology of Bacterial Endophthalmitis.

Purpose: Age, sex, and genetics are important biological variables in determining an individual's susceptibility or response to infectious agents; however, their role has not been evaluated in intraocular infections. In this study, we comprehensively examined the impact of these host biological factors in the pathogenesis of experimental bacterial endophthalmitis. Methods: Endophthalmitis was induced by intravitreal injection of bacteria (Staphylococcus aureus) in the eyes of male and female C57BL/6 mice of different ages: group I (young, 6-8 weeks), group II (mid-age, 18-20 weeks), and group III (old, 1 year). Highly heterogeneous outbred J:DO mice were used for genetic diversity analysis. Eyes were subjected to clinical examination, retinal function testing using electroretinography (ERG), histopathological analysis (hematoxylin and eosin staining), and bacterial burden estimation. The levels of inflammatory mediators were measured using qPCR and ELISA, and the infiltration of neutrophils was determined by flow cytometry. Results: Both inbred C57BL/6 and diversity outbred (J:DO) mice were equally susceptible to S. aureus endophthalmitis, as evidenced by a time-dependent increase in clinical scores, bacterial burden, intraocular inflammation, and retinal tissue damage, in addition to decreased retinal function. However, no significant differences were observed in disease severity and innate responses in male versus female mice. Older mice (group III) exhibited higher clinical scores coinciding with increased bacterial proliferation and intraocular inflammation, resulting in enhanced disease severity. Moreover, bone-marrow-derived macrophages from old mice exhibited reduced phagocytic activity but increased inflammatory response toward S. aureus challenge. Conclusions: Age, but not sex, is an important biological variable in bacterial endophthalmitis. Identification of pathways underlying altered innate immunity and impaired bacterial clearance in aging eyes could provide new insights into the pathobiology of intraocular infections in elderly patients.

Prevalence of SARS-CoV-2 in human post-mortem ocular tissues.

BACKGROUND: SARS-CoV-2 is found in conjunctival swabs and tears of COVID-19 patients. However, the presence of SARS-CoV-2 has not been detected in the human eye to date. We undertook this study to analyze the prevalence of SARS-CoV-2 in human post-mortem ocular tissues. METHODS: The expression of SARS-CoV-2 RNA was assessed by RT-PCR in corneal and scleral tissues from 33 surgical-intended donors who were eliminated from a surgical use per Eye Bank Association of America (EBAA) donor screening guidelines or medical director review or positive COVID-19 test. Ocular levels of SARS-CoV-2 RNA (RT-PCR), Envelope and Spike proteins (immunohistochemistry) and anti-SARS-CoV-2 IgG and IgM antibodies (ELISA) in blood were evaluated in 10 COVID-19 donors. FINDINGS: Of 132 ocular tissues from 33 surgical-intended donors, the positivity rate for SARS-CoV-2 RNA was ~13% (17/132). Of 10 COVID-19 donors, six had PCR positive post-mortem nasopharyngeal swabs whereas eight exhibited positive post-mortem anti-SARS-CoV-2 IgG levels. Among 20 eyes recovered from 10 COVID-19 donors: three conjunctival, one anterior corneal, five posterior corneal, and three vitreous swabs tested positive for SARS-CoV-2 RNA. SARS-CoV-2 spike and envelope proteins were detected in epithelial layer of the corneas that were procured without Povidone-Iodine (PVP-I) disinfection. INTERPRETATIONS: Our study showed a small but noteworthy prevalence of SARS-CoV-2 in ocular tissues from COVID-19 donors. These findings underscore the criticality of donor screening guidelines, post-mortem nasopharyngeal PCR testing and PVP-I disinfection protocol to eliminate any tissue harboring SARS-CoV-2 being used for corneal transplantation.