Infectious Disease

Woodward AM, Mauris J, Argüeso P. Binding of transmembrane mucins to galectin-3 limits herpesvirus 1 infection of human corneal keratinocytes. J Virol 2013;87(10):5841-7.Abstract
Epithelial cells lining mucosal surfaces impose multiple barriers to viral infection. At the ocular surface, the carbohydrate-binding protein galectin-3 maintains barrier function by cross-linking transmembrane mucins on the apical glycocalyx. Despite these defense mechanisms, many viruses have evolved to exploit fundamental cellular processes on host cells. Here, we use affinity assays to show that herpes simplex virus type 1 (HSV-1), but not HSV-2, binds human galectin-3. Knockdown of galectin-3 in human corneal keratinocytes by small interfering RNA significantly impaired HSV-1 infection, but not expression of nectin-1, indicating that galectin-3 is a herpesvirus entry mediator. Interestingly, exposure of epithelial cell cultures to transmembrane mucin isolates decreased viral infectivity. Moreover, HSV-1 failed to elute the biological counterreceptor MUC16 from galectin-3 affinity columns, suggesting that association of transmembrane mucins to galectin-3 provides protection against viral infection. Together, these results indicate that HSV-1 exploits galectin-3 to enhance virus attachment to host cells and support a protective role for transmembrane mucins under physiological conditions by masking viral entry mediators on the epithelial glycocalyx.
Wurster JI, Bispo PJM, Van Tyne D, Cadorette JJ, Boody R, Gilmore MS. Staphylococcus aureus from ocular and otolaryngology infections are frequently resistant to clinically important antibiotics and are associated with lineages of community and hospital origins. PLoS One 2018;13(12):e0208518.Abstract
Staphylococcus aureus is an important human pathogen that causes serious antibiotic-resistant infections. Its population structure is marked by the appearance and dissemination of successful lineages across different settings. To begin understanding the population structure of S. aureus causing ocular and otolaryngology infections, we characterized 262 isolates by antimicrobial sensitivity testing and multilocus sequence typing (MLST). Methicillin-resistant S. aureus were subjected to SCCmec typing and Panton-Valentine leukocidin (PVL) screening. Although we detected a high level of genetic diversity among methicillin-sensitive (MSSA) isolates, (63 sequence types-STs), the population was dominated by five lineages: ST30, ST5, ST8, ST15 and ST97. Resistance to penicillin, erythromycin and clindamycin was common among the major MSSA lineages, with fluctuations markedly impacted by genetic background. Isolates belonging to the predominant lineage, ST30, displayed high rates of resistance to penicillin (100%), erythromycin (71%), and clindamycin (63%). Overall, 21% of the isolates were methicillin-resistant (MRSA), with an apparent enrichment among otitis and orbital cellulitis isolates (>40%). MRSA isolates belonged to 14 STs grouped in 5 clonal complexes (CC), however, CC5 (56.1%) and CC8 (38.6%) dominated the population. Most CC5 strains were SCCmec type II, and resembled the hospital-adapted USA100 clone. CC8 strains were SCCmec type IV, and 86% were positive for the PVL toxin, common features of the community-acquired clone USA300. CC5 strains harboring a SCCmec type IV, typical for the USA800 clone, comprised 15.5% of the population. USA100 strains were highly resistant to clindamycin, erythromycin and levofloxacin (100%), while USA300 strains were frequently resistant to erythromycin (89%) but displayed lower rates of resistance to levofloxacin (39%) and clindamycin (17%). Our data demonstrate that the ocular and otolaryngology S. aureus populations are composed of strains that are commonly resistant to clinically relevant antibiotics, and are associated with the major epidemic clonal complexes of both community and hospital origins.
Wurster JI, Saavedra JT, Gilmore MS. Impact of Antibiotic Use on the Evolution of Enterococcus faecium. J Infect Dis 2016;213(12):1862-5.
Yu Z, Zeng Z, Zhang J, Pan Y, Chen M, Guo Y, Yu N, Chodosh J, Fu N, Che X, Zhang Q. Fatal Community-acquired Pneumonia in Children Caused by Re-emergent Human Adenovirus 7d Associated with Higher Severity of Illness and Fatality Rate. Sci Rep 2016;6:37216.Abstract

Human adenoviruses (HAdVs) are highly contagious pathogens causing acute respiratory disease (ARD), such as community-acquired pneumonia. HAdV-7d, a re-emergent genomic variant, has been recently reported in Asia and the United States after a several-decade absence. However, whether HAdV-7d is associated with higher severity than other types is currently unclear. In this study, the clinical and epidemiological investigation showed that fever, cough, and sore throat were the three most common respiratory symptoms of HAdV infections. HAdV-7 caused longer duration of fever, higher morbidity of tachypnea/dyspnea, pleural effusion, diarrhea, hepatosplenomegaly, consciousness alteration, as well as higher rates of pneumonia, mechanical ventilation and higher fatality rate (28.6%) than other types, particularly HAdV-3 and HAdV-2. The genomes of seven HAdV-7d isolates from mild, severe, and fatal cases were sequenced and highly similar with each other. Surprisingly, two isolates (2011, 2012) had 100% identical genomes with an earlier strain from a fatal ARD outbreak in China (2009), which elucidates the virus origin and confirms the unexpected HAdV genomic conservation and stability. Phylogenetic analysis indicated that L1 52/55-kDa DNA packaging protein may be associated with the higher severity of illness and fatality rate of HAdV-7. Clinicians need to be aware of HAdVs in children with ARD.

Zabaleta N, Dai W, Bhatt U, Hérate C, Maisonnasse P, Chichester JA, Sanmiguel J, Estelien R, Michalson KT, Diop C, Maciorowski D, Dereuddre-Bosquet N, Cavarelli M, Gallouët A-S, Naninck T, Kahlaoui N, Lemaitre J, Qi W, Hudspeth E, Cucalon A, Dyer CD, Pampena BM, Knox JJ, LaRocque RC, Charles RC, Li D, Kim M, Sheridan A, Storm N, Johnson RI, Feldman J, Hauser BM, Contreras V, Marlin R, Tsong Fang RH, Chapon C, van der Werf S, Zinn E, Ryan A, Kobayashi DT, Chauhan R, McGlynn M, Ryan ET, Schmidt AG, Price B, Honko A, Griffiths A, Yaghmour S, Hodge R, Betts MR, Freeman MW, Wilson JM, Le Grand R, Vandenberghe LH. An AAV-based, room-temperature-stable, single-dose COVID-19 vaccine provides durable immunogenicity and protection in non-human primates. Cell Host Microbe 2021;29(9):1437-1453.e8.Abstract
The SARS-CoV-2 pandemic has affected more than 185 million people worldwide resulting in over 4 million deaths. To contain the pandemic, there is a continued need for safe vaccines that provide durable protection at low and scalable doses and can be deployed easily. Here, AAVCOVID-1, an adeno-associated viral (AAV), spike-gene-based vaccine candidate demonstrates potent immunogenicity in mouse and non-human primates following a single injection and confers complete protection from SARS-CoV-2 challenge in macaques. Peak neutralizing antibody titers are sustained at 1 year and complemented by functional memory T cell responses. The AAVCOVID vector has no relevant pre-existing immunity in humans and does not elicit cross-reactivity to common AAVs used in gene therapy. Vector genome persistence and expression wanes following injection. The single low-dose requirement, high-yield manufacturability, and 1-month stability for storage at room temperature may make this technology well suited to support effective immunization campaigns for emerging pathogens on a global scale.
Zhai H, Bispo PJM, Kobashi H, Jacobs DS, Gilmore MS, Ciolino JB. Resolution of fluoroquinolone-resistant keratitis with a PROSE device for enhanced targeted antibiotic delivery. Am J Ophthalmol Case Rep 2018;12:73-75.Abstract
Purpose: To report the resolution of a fluoroquinolone-resistant keratitis with use of a prosthetic replacement of the ocular surface ecosystem (PROSE) device for enhanced targeted delivery of moxifloxiacin. Observations: A 62-year-old female presented with a 3-day history of pain, photophobia, and declining vision in left eye. The patient had a 2-year history of binocular PROSE treatment for ocular chronic graft-vs-host disease (cGVHD). A corneal ulcer was diagnosed and treated with topical 0.5% moxifloxacin solution 6 times per day, with continued wear of the PROSE device. After 4 days, worsening symptoms led to an increase in application of moxifloxicin to every 2 hours while awake. The drug was administered by removal of the device, cleaning and replenishing the reservoir with sterile saline, and adding one drop of the drug to the reservoir prior to reinsertion. Four days later, the corneal surface was epithelialized with only small subepithelial infiltrate remaining. The corneal culture grew an isolate carrying multiple mutations in the topoisomerase genes. These mutations were correlated with varying levels of resistance to ciprofloxacin (256 μg/mL), levofloxacin (8 μg/mL), and moxifloxacin (16 μg/mL). Conclusions and Importance: Although the infecting strain exhibited resistance to fluoroquinolones, the infection resolved when moxifloxacin was combined with PROSE therapy. Frequent dosing to the PROSE reservoir is likely to increase fluoroquinolone bioavailability and may represent a valuable approach to overcome antibiotic resistance.
Zhang S, Lebreton F, Mansfield MJ, Miyashita S-I, Zhang J, Schwartzman JA, Tao L, Masuyer G, Martínez-Carranza M, Stenmark Pål, Gilmore MS, Doxey AC, Dong M. Identification of a Botulinum Neurotoxin-like Toxin in a Commensal Strain of Enterococcus faecium. Cell Host Microbe 2018;23(2):169-176.e6.Abstract
Botulinum neurotoxins (BoNTs), produced by various Clostridium strains, are a family of potent bacterial toxins and potential bioterrorism agents. Here we report that an Enterococcus faecium strain isolated from cow feces carries a BoNT-like toxin, designated BoNT/En. It cleaves both VAMP2 and SNAP-25, proteins that mediate synaptic vesicle exocytosis in neurons, at sites distinct from known BoNT cleavage sites on these two proteins. Comparative genomic analysis determines that the E. faecium strain carrying BoNT/En is a commensal type and that the BoNT/En gene is located within a typical BoNT gene cluster on a 206 kb putatively conjugative plasmid. Although the host species targeted by BoNT/En remains to be determined, these findings establish an extended member of BoNTs and demonstrate the capability of E. faecium, a commensal organism ubiquitous in humans and animals and a leading cause of hospital-acquired multi-drug-resistant (MDR) infections, to horizontally acquire, and possibly disseminate, a unique BoNT gene cluster.
Zhou X, Ramke M, Chintakuntlawar AV, Lee JY, Rajaiya J, Chodosh J. Role of MyD88 in adenovirus keratitis. Immunol Cell Biol 2017;95(1):108-116.Abstract

Pattern recognition receptors (PRRs) are critical to the early detection and innate immune responses to pathogens. In particular, the toll-like receptor (TLR) system and its associated adaptor proteins have essential roles in early host responses to infection. Epidemic keratoconjunctivitis, caused by the human adenovirus, is a severe ocular surface infection associated with corneal inflammation (stromal keratitis). We previously showed that adenovirus capsid was a key molecular pattern in adenovirus keratitis, with viral DNA having a lesser role. We have now investigated the role of the adaptor molecule MyD88 in a mouse model of adenovirus keratitis in which there is no viral replication. In MyD88(-/-) mice infected with human adenovirus type 37, clinical keratitis was markedly reduced, along with infiltration of CD45(+) cells, and expression of inflammatory cytokines. Reduction of inflammatory cytokines was also observed in infected primary human corneal fibroblasts pretreated with a MyD88 inhibitory peptide. Keratitis similar to wild type mice was observed in TLR2, TLR9 and IL-1R knockout mice, but was reduced in TLR2/9 double knockout mice, consistent with synergy of TLR2 and TLR9 in the response to adenovirus infection. MyD88 co-immunoprecipitated with Src kinase in mice corneas and in human corneal fibroblasts infected with adenovirus, and MyD88 inhibitory peptide reduced Src phosphorylation, linking MyD88 activation to inflammatory gene expression through a signaling cascade previously shown to be directed by Src. Our findings reveal a critical role for the PRRs TLR2 and 9, and their adaptor protein MyD88, in corneal inflammation upon adenovirus infection.

Zhou X, Robinson CM, Rajaiya J, Dehghan S, Seto D, Jones MS, Dyer DW, Chodosh J. Analysis of human adenovirus type 19 associated with epidemic keratoconjunctivitis and its reclassification as adenovirus type 64. Invest Ophthalmol Vis Sci 2012;53(6):2804-11.Abstract
PURPOSE: Human adenovirus species D type 19 (HAdV-D19) has been associated with epidemic keratoconjunctivitis (EKC), a highly inflammatory infection of the ocular surface. Confusion exists regarding the origins of HAdV-D19. The prototype virus (HAdV-D19p) does not cause EKC, while a virus identified later with the identical serologic determinant is a significant ocular pathogen. METHODS: High throughput genome sequencing and bioinformatics analysis were performed on HAdV-D19p and three HAdV-D19 EKC strains, and compared to the previously sequenced clinical isolate, HAdV-D19 (C) and HAdV-D37. Corneas of C57BL/6J mice were injected with HAdV-D19p, HAdV-D19 (C), or virus-free buffer, and inflammation assessed by clinical examination, flow cytometry, and cytokine ELISA. Confocal microscopy and real-time PCR of infected corneal cell cultures were used to test viral entry. RESULTS: HAdV-D19 (C) and the other clinical EKC isolates showed nearly 100% sequence identity. EKC strains diverged from HAdV-D19p in the penton base, E3, and fiber transcription units. Simplot analysis showed recombination between EKC-associated HAdV-D19 with HAdV-D37, HAdV-D22, and HAdV-D19p, the latter contributing only the hexon gene, the principal serum neutralization determinant. HAdV-D19p induced stromal keratitis in the C57BL/6J mouse, but failed to infect productively human corneal epithelial cells. These data led to retyping of the clinical EKC isolates with a HAdV-D19 hexon gene as HAdV-D64. CONCLUSIONS: HAdV-D19 associated with EKC (HAdV-D64) originated from a recombination between HAdV-D19p, HAdV-D37, and HAdV-D22, and was mischaracterized because of a shared hexon gene. HAdV-D19p is not infectious for corneal epithelial cells, thus explaining the lack of any association with keratitis.