Ratiometric fluorescence immunoassay of SARS-CoV-2 nucleocapsid protein via Si-FITC nanoprobe-based inner filter effect.

The global pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus has necessitated rapid, easy-to-use, and accurate diagnostic methods to monitor the virus infection. Herein, a ratiometric fluorescence enzyme-linked immunosorbent assay (ELISA) was developed using Si-fluorescein isothiocyanate nanoparticles (FITC NPs) for detecting SARS-CoV-2 nucleocapsid (N) protein. Si-FITC NPs were prepared by a one-pot hydrothermal method using 3-aminopropyl triethoxysilane (APTES)-FITC as the Si source. This method did not need post-modification and avoided the reduction in quantum yield and stability. The p-nitrophenyl (pNP) produced by the alkaline phosphatase (ALP)-mediated hydrolysis of p-nitrophenyl phosphate (pNPP) could quench Si fluorescence in Si-FITC NPs via the inner filter effect. In ELISA, an immunocomplex was formed by the recognition of capture antibody/N protein/reporter antibody. ALP-linked secondary antibody bound to the reporter antibody and induced pNPP hydrolysis to specifically quench Si fluorescence in Si-FITC NPs. The change in fluorescence intensity ratio could be used for detecting N protein, with a wide linearity range (0.01-10.0 and 50-300 ng/mL) and low detection limit (0.002 ng/mL). The concentration of spiked SARS-CoV-2 N protein could be determined accurately in human serum. Moreover, this proposed method can accurately distinguish coronavirus disease 2019 (COVID-19) and non-COVID-19 patient samples. Therefore, this simple, sensitive, and accurate method can be applied for the early diagnosis of SARS-CoV-2 virus infection. Electronic Supplementary Material: Supplementary material (characterization of Si-FITC NPs (FTIR spectrum, XRD spectra, and synchronous fluorescence spectra); condition optimization of ALP response (fluorescence intensity ratio change); mechanism investigation of ALP response (fluorescence lifetime decay curves and UV-vis absorption spectra); detection of N protein using commercial ELISA Kit; analytical performance of assays for ALP detection or SARS-CoV-2 N protein detection; and determination results of SARS-CoV-2 N protein in human serum) is available in the online version of this article at 10.1007/s12274-022-4740-5.

Acute Multifocal Placoid Pigment Epitheliopathy-like Presentation following the First Dose of BNT162B2 COVID-19 Vaccination.

PURPOSE: To report a case with acute posterior multifocal placoid pigment epitheliopathy (APMPPE)-like presentation following the first dose of BNT162b2 COVID-19 vaccination. CASE REPORT: An otherwise healthy 45-year-old woman presented with a headache and blurred vision in her right eye 7 days after the administration of first dose of mRNA (BNT162b2) COVID-19 vaccine. Fundus examination of the right eye revealed multiple discrete yellow-white placoid lesions at the level of deep retinal layers throughout the posterior pole, while left fundus was unremarkable at that time. Swept source-optical coherence tomography (SS-OCT) showed subretinal fluid together with an appearance of bacillary layer detachment at the right macula. A detailed systemic evaluation was carried out without any positive finding. Two weeks after the initial eye examination, similar multiple placoid lesions were observed in her left eye. Fundus lesions almost totally resolved without any treatment bilaterally 5 weeks after the onset of initial symptoms. CONCLUSION: To the best of our knowledge, this is the first report of APMPPE-like presentation described after the BNT162b2 COVID-19 vaccination. Previous COVID-19 vaccination should be kept in mind in the differential diagnosis of APMPPE disease spectrum.

Inhibition of SARS-CoV-2 nucleocapsid protein-RNA interaction by guanosine oligomeric RNA.

The interaction of the ß-coronavirus severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) nucleocapsid (N) protein with genomic RNA is initiated by specific RNA regions and subsequently induces the formation of a continuous polymer with characteristic structural units for viral formation. We hypothesized that oligomeric RNAs, whose sequences are absent in the 29.9-kb genome sequence of SARS-CoV-2, might affect RNA-N protein interactions. We identified two such hexameric RNAs, In-1 (CCGGCG) and G6 (GGGGGG), and investigated their effects on the small filamentous/droplet-like structures (< a few µm) of N protein-genomic RNA formed by liquid-liquid phase separation. The small N protein structures were sequence-specifically enhanced by In-1, whereas G6 caused them to coalesce into large droplets. Moreover, we found that a guanosine 12-mer (G12, GGGGGGGGGGGG) expelled preexisting genomic RNA from the small N protein structures. The presence of G12 with the genomic RNA suppressed the formation of the small N protein structures, and alternatively apparently altered phase separation to induce the formation of large droplets with unclear phase boundaries. We showed that the N-terminal RNA-binding domain is required for the stability of the small N protein structures. Our results suggest that G12 may be a strong inhibitor of the RNA-N protein interaction.

Routine ophthalmological examinations in adults with sickle cell disease are low and further adversely affected by the COVID-19 Pandemic

Purpose : The American Academy of Ophthalmology (AAO) and the National Heart, Lung, and Blood Institute (NHLBI) suggest sickle cell patients undergo dilated funduscopic exams (DFE) every 1-2 years to screen for sickle cell retinopathy, but there is a paucity of research reporting whether such guidelines are followed in the sickle cell disease (SCD) population. A retrospective study was performed to assess whether adults with SCD at our institution are adhering to the recommendations. Methods : This study is a chart review of 842 adult patients with sickle cell diagnosis, seen from 3/17-3/21 by internists in the Montefiore healthcare system. Data was collected through the Electronic Medical Record (EMR) of patients with a DFE categorized as normal (Retinopathy -, n = 216) or with retinopathy (Retinopathy +, n = 199). A regular DFE was defined as at least one DFE every 2 years. Screening rates were calculated by removing patients with retinopathic disease noted in EMR from total SCD patients. Yearly DFE rates were calculated from March to March, since COVID-19 was declared a global pandemic in March 2020. Two-tailed Student's t-test was used for statistical analysis of continuous data, Fisher's exact test for categorical data, and two-sample proportion test for comparison of yearly exam rates. Results : The Retinopathy-group was younger (p<0.001) and contained less SC genotype (p<0.001) compared to Retinopathy +. 40.3% of the Retinopathy-patients were screened regularly (n = 87), whereas 59.7% had irregular screening (n = 129). There was a significant decrease in the total rate of all patients with DFEs, comparing the average rate of 29.8% pre-COVID (3/17-3/20) to 13.6% during COVID (3/20-3/21) (p<0.001). Similarly, the screening rate for non-retinopathic patients decreased from an average rate of 18.6% pre-COVID to 6.7% during COVID (p<0.001). Conclusions : The results demonstrate that our rate of routine dilated fundoscopic examination for SCD patients is very low. These low rates were even more adversely affected by the onset of the COVID-19 pandemic, as clinics closed and access to physicians decreased. Screening rates need to be increased by patient involvement and education, easier access to ophthalmologists and novel ways of screening for retinopathy (e.g. annual screening fluorescein angiograms) to reduce risk of blindness in these patients.

Reduction of mask-associated dry eye by using surgical tape to secure the top edge of the mask

Purpose : During the COVID-19 pandemic, mask-associated dry eye (MADE) has increased worldwide possibly because the breath leaks from the top of the masks changes the ocular surface conditions. We tested the hypothesis that surgical taping of the top edge of the mask to the skin reduces the risk of ocular surface damage. Methods : We enrolled 60 volunteers (30 females, 30 males;mean age, 27.1±5.2 years) who wear face masks over 5 hours a day. We measured the fluorescein tear break-up time (FBUT), ocular surface temperature, and conjunctival blood flow before wearing masks, after wearing masks taped on the top edge, and after wearing masks without tape. We used the Ocular Surface Disease Index (OSDI) to record participants' symptoms of MADE and measured their corneal tactile and pain sensitivity using a Cochet-Bonnet esthesiometer. Results : The FBUT with masks without tape (4.4±2.4 seconds) was significantly shorter than that without masks (6.4±3.1 seconds) and with taped masks (5.8±3.2 seconds) (P<0.01 and P=0.05, respectively, Tukey HSD test). There was no significant difference in the FBUT between use of no masks and taped masks (P>0.05). The differences in the corneal and conjunctival temperatures after wearing masks without tape (0.19±0.28 and 0.13±0.28°C, respectively) were significantly higher than after wearing taped masks (0.05±0.27 and 0.06±0.24°C, respectively, P<0.01, paired t-test). The conjunctival blood flow with masks without tape was significantly higher than that of taped masks (P<0.01). Of the 60 subjects, 13 (21.7%) subjects reported MADE symptoms. In the MADE group, the OSDI (P=0.001, analysis of covariance) was significantly higher and the FBUT of masks without tape (P=0.006) was significantly shorter than in the non-MADE group. Pain sensitivity in the MADE group was significantly higher than in the non-MADE group (P<0.01), indicating that subjects in the MADE group were significantly hypersensitive to corneal pain. Conclusions : Wearing masks decreased FBUT and increased ocular surface temperature and blood flow. Taping the top edge of the mask prevented those changes. Fitting masks tightly to the nose or applying tape over the mask may reduce the MADE risk, which can be associated with ocular surface hypersensitivity.

A comparative study of conjunctivitis in COVID-19 patients between the first and second wave of COVID-19 at a tertiary care center

Background: SARS CoV-2 has become a household name since early 2019 and till very recently has shown many mutations, some of which were predictable in behavior whereas some have been quite bizarre. With all these surreal mutants of the virus, the basic signs and symptoms of the infection also were diversified. A few studies compared the conjunctivitis features of the first and second wave of coronavirus disease 2019 (COVID-19) outbreak. Aims and Objectives: The purpose of this study was to compare the evidence of conjunctivitis between the first and second wave of COVID-19 patients. Materials and Methods: In this retrospective and observational research, clinical and laboratory characteristics of the patients with COVID-19 admitted to a tertiary care hospital in Western Maharashtra. We evaluated the patients including detailed history taken from the medical records and ocular evaluation was carried out by a trained ophthalmologist who was posted in the COVID-19 ward at that time. The patient's symptoms were compared with the severity of COVID-19 disease classified as mild, moderate, and based on oxygen support given to the patient. Results: A total of 150 patients were evaluated in this study, of which 50 patients were from the first wave and 100 were from second wave of COVID-19. Among the total patients, about (80%) of patients with mild severity of COVID-19 disease were from in first wave v/s (61%) patients with mild severity of disease were from the second wave. Furthermore, it was observed that disease severity with moderate patients from the first wave was around (20%) v/s (39%) in the second wave. Among the total patients' data, 66% of patients infected with conjunctivitis were in the first wave of COVID-19 and approximately 56% of patients infected with conjunctivitis were found in the second wave, respectively. About 18% of patients from the first wave and 39% of patients from the second wave with moderate severity required oxygen support. The fluorescein staining, as well as congestion, showed no significant difference in the number of patients between both waves. Conclusion: The present study concludes that both the waves only reported dryness and no clinical signs of conjunctivitis. The present study concludes that increased oxygenation support may act as one of the contributing factor for increased dryness and thereby may lead to various ocular infections. [ FROM AUTHOR] Copyright of Asian Journal of Medical Sciences is the property of Manipal Colleges of Medical Sciences and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full . (Copyright applies to all s.)

Establishment of a Duplex Real Time PCR Method for Detection of SARS-COV-2

To quickly and efficiently detect the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and prevent and control the spread of novel coronavirus disease (COVID-19), a highly sensitive duplex real-time PCR (RT-PCR) detection method has been established. In this study, the specificity of primers and probes were designed, respectively, according to the ORF1ab gene and N gene sequence of SARS-COV-2, and fluorescent probes were labeled with carboxyl fluorescein (FAM) and green fluorescent protein (VIC). The duplex RT-PCR method for detecting SARS-COV-2 with TaqMan probe was established, which has a limit of detection of 10 copies/µL, and the linear detection range of ORF1ab and N gene were 1.0 × 101-1.0 × 105 copies/µL and 1.0 × 101-1.0 × 106 copies/µL, respectively, realizing the simultaneous detection of ORF1ab and N genes in simulated SARS-COV-2 samples. The method has high sensitivity, accurate quantification, simple operation, and cost-saving, which can be used for rapid and efficient quantitative detection of SARS-COV-2.

Reactivation of juvenile idiopathic arthritis associated uveitis with posterior segment manifestations following anti-SARS-CoV-2 vaccination.

BACKGROUND/PURPOSE: Juvenile idiopathic arthritis (JIA) is the most common rheumatic disease in the pediatric population and anterior uveitis is its commonest extra-articular manifestation. Typically the uveitis presents as chronic anterior uveitis and there is limited literature of the posterior segment manifestations of the disease. Similar to other vaccines, anti-SARS-CoV-2 vaccination that began as an urgent measure to control the spread of the SARS-CoV-2 pandemic has not been without adverse events. We are reporting a 19-year-old Asian Indian female who was diagnosed and treated for JIA associated anterior uveitis that was unilateral and was under anti-inflammatory control but showed worsening of uveitis with posterior segment inflammation in both eyes following anti-SARS-CoV-2 vaccination. CASE REPORT: A 19-year-old Asian Indian female with a history of juvenile idiopathic arthritis on treatment with methotrexate, presented with right eye chronic anterior uveitis with peripheral subclinical retinal vasculitis and macular edema which was brought under control following administration of adalimumab. She was inflammation free for 6 months until she received anti-SARS-CoV-2 vaccination and developed new onset floaters in both eyes that were initially noted after the first dose and increased after the second dose. Clinical examination revealed presence of keratic precipitates and grade 1+ anterior chamber inflammation along with vitiritis in both eyes. Fundus fluorescein angiography revealed angiographically active retinal vasculitis without the presence of macular edema in both eyes. This was managed with a short course of topical difluprednate and continuation of systemic immunosuppressive therapy with adalimumab and methotrexate. CONCLUSION: JIA associated uveitis results from an autoimmune process which can be controlled with timely immunosuppressive treatment. It is important to be aware of the potential risk of flare up of uveitis with posterior segment manifestations following anti- SARS-CoV-2 vaccination.

The Microbial Activity in PM2.5 in Indoor Air: As an Index of Air Quality Level

Bioaerosols are a major source of pollution in indoor environments, where people spend approximately 90% of their time, and the microorganisms adhered to PM2.5 adversely affect human health. However, most research has focused on the concentration of these aerosols and the factors that influence it rather than the correlation between microbial activity and air quality. Thus, this study used a modified technique of fluorescein diacetate (FDA) hydrolysis to evaluate the activity of microorganisms in the PM2.5 during three seasons (summer, autumn and winter) in Beijing. 0.155–5.388 ng m–3 and 0.091–5.740 ng m–3 of sodium fluorescein, a marker of microbial activity, were measured indoors and outdoors, respectively;thus, no significant difference in concentration between the two environments was detected, but the indoor activity was affected by outdoor conditions to an extent. The most active season was autumn, followed by winter and summer. Furthermore, the highest activity in summer and autumn was observed during conditions of excellent air quality, and in summer, the activity during conditions of good air quality also obviously exceeded that during conditions of slight pollution. Additionally, the microbial activity in a room varied according to the room’s ventilation (or lack thereof), suggesting a strong association between these parameters. In general, when the air quality was excellent, 20 minutes of ventilation achieved the optimal air exchange, but this duration should be reduced during polluted conditions. Our results provide new insights into evaluating the indoor air quality based on the microbial activity.

Comparison of an Air-Fed Mask System with Hospital-Issued Personal Protection Equipments for Dental Aerosol Protection During the COVID-19 Pandemic

Aim: A pilot study was conducted with the aim of developing a system to protect the eyes, nose, and mouth from the aerosol generated from a high-speed dental handpiece during the COVID-19 pandemic. Background: The SARS-CoV-2 virus is known to be present in the saliva of an infected individual during the contagious viral shedding phase of the disease. The use of rotary dental instruments places oral health practitioners at risk of contracting COVID-19 from infected individuals. In particular, it is very difficult to protect the mucous membranes of the face against the extremely fine aerosol produced from a high-speed dental handpiece. Objectives: This study aimed to develop and test a novel PPE system for use during the COVID-19 pandemic. An air-fed spray-painting mask was used under a plastic hood to protect against the aerosol from a high-speed dental handpiece. This was found to be superior compared to hospital-issued N-95 masks and eye protection in our test model. Methods: Subjects donned various forms of PPE whilst using a high-speed dental handpiece in a confined cubicle. The efficacy of each form of PPE was evaluated by adding fluorescein to the water coolant supply line of a high-speed dental handpiece before checking for facial contamination with an ophthalmology slit lamp. Results: Under our test conditions, the N-95 mask did not prevent nasal and mouth contaminations, but the combination of an air-fed mask with a sealed hood prevented these contaminations. Although goggles worn tightly did prevent contamination, the air-fed mask system was far more comfortable and did not fog up. Discussion: Under the rigorous test conditions of our model, we found hospital-issued PPE ineffective. We also found the single strategy of using positive airflow into a face mask ineffective, even with extremely high levels of airflow. Complete protection was only achieved reliably by the combination of physically sealing off the face from the surrounding airspace and using the air-fed system to provide an external source of air to breathe. We effectively made the clinical equivalent of a dive bell helmet. The air-fed mask is supplied by a standard dental air compressor and is simple to install for someone familiar with the technical aspects of compressors. The compressor does not rely on a filter and proves effective with cheap and easily accessible disposable items. Conclusion: Under rigorous testing conditions, the developed air-fed mask system with a sealed hood on low flow performed better than hospital-issued PPE against high-speed dental aerosol protection. The developed system protects the operators from the air of the room contaminated with aerosol and brings in safe air from the outside for them to breathe.

Polyamidoamine-Remdesivir Conjugate: Physical Stability and Cellular Uptake Enhancement

Nucleoside analogue antiviral remdesivir works by inhibiting the RNA-dependent RNA polymerase enzyme and terminating the viral replication. Currently, remdesivir is under a clinical trial for its activity against SARS-CoV-2. In the blood, remdesivir will undergo an enzymatic reaction to become monophosphate analogue form which is difficult to penetrate into the cell membrane. PAMAM (polyamidoamine) dendrimer is a good carrier to encapsulate remdesivir as a water-insoluble drug (0,339 mg/mL). Entrapment of remdesivir in the PAMAM cavity avoided remdesivir molecules to not undergo the enzymatic reactions. This study aimed to synthesize, characterize and evaluate cellular uptake of PAMAM-Remdesivir conjugate. PAMAM-Remdesivir was prepared with various stirring times (3, 6, 12, 24, and 48 hours). The conjugates were characterized to observe the size and particle distribution using Particle Size Analyzer, encapsulating efficiency using UV-Vis Spectroscopy, interaction between PAMAM and remdesivir particle using Fourier Transform Infrared Spectroscopy and cellular uptake of PAMAM-RDV using Fluorescence Microscope. The optimized stirring time of PAMAM-Remdesivir conjugate was 24 hours wich resulted the particles charge of + 23,07 mV of zeta potential, 1008 nm of particle size, 0,730 of PDI, and 69% entrapment efficiency. In addition, the FTIR analysis showed that remdesivir molecules successfully conjugated to PAMAM. Thus, through strring optimization time, the remdesivir molecules were successfully entrapped to PAMAM cavity. The cellular uptake in Vero Cell of PAMAM-RDV conjugated fluorescein isothiocyanate was observed with fluorescence microscope and had a stronger intensity than remdesivir only solution.

Development and Assessment of an Inexpensive Smartphone-Based Respiratory Droplet Simulation Model.

Background. Droplet simulation often requires expensive and inaccessible equipment. Herein, we develop and assess a low-cost droplet simulation model using easily accessible materials, open-source software, and a smartphone-based cobalt blue light. Methods. The simulation model was developed using commercial-grade materials and fluorescein dye. A clear face shield was assessed ten times following a simulated cough using fluorescein dye. A conventional ultraviolet Woods lamp was compared to a smartphone-based cobalt blue light to detect fluorescein illumination. Results. The simulation platform and smartphone-based cobalt blue light cost $20.18. A Wilcoxon signed rank test revealed that the median droplet area of fluorescence under the UV Wood's lamp was not significantly different than that of the smartphone-based cobalt blue light (2.89 vs 2.94, P = .386). Conclusions. This simulation model is inexpensive and easily reproducible. The smartphone application may be a convenient alternative to standard ultraviolet lights. This model has great potential for use in financially restricted academic centers during the COVID-19 pandemic and beyond.

Aerosolised fluorescein can quantify FFP mask faceseal leakage: a cost-effective adaptation to the existing point of care fit test.

BACKGROUND: A qualitative fit test using bitter-tasting aerosols is the commonest way to determine filtering face-piece (FFP) mask leakage. This taste test is subjective and biased by placebo. We propose a cheap, quantitative modification of the taste test by measuring the amount of fluorescein stained filter paper behind the mask using image analysis. METHODS: A bitter-tasting fluorescein solution was aerosolised during mask fit tests, with filter paper placed on masks' inner surfaces. Participants reported whether they could taste bitterness to determine taste test 'pass' or 'fail' results. Filter paper photographs were digitally analysed to quantify total fluorescence (TF). RESULTS: Fifty-six healthcare professionals were fit tested; 32 (57%) 'passed' the taste test. TF between the taste test 'pass' and 'fail' groups was significantly different (p<0.001). A cut-off (TF = 5.0 × 106 units) was determined at precision (78%) and recall (84%), resulting in 5/56 participants (9%) reclassified from 'pass' to 'fail' by the fluorescein test. Seven out of 56 (12%) reclassified from 'fail' to 'pass'. CONCLUSION: Fluorescein is detectable and sensitive at identifying FFP mask leaks. These low-cost adaptations can enhance exiting fit testing to determine 'pass' and 'fail' groups, protecting those who 'passed' the taste test but have high fluorescein leak, and reassuring those who 'failed' the taste test despite having little fluorescein leak.

Comparative performances of seven quantitative Reverse-Transcription Polymerase Chain Reaction assays (RT-qPCR) for detecting SARS-CoV-2 infection in samples from individuals suspected of COVID-19 in São Paulo, Brazil.

Introduction: Brazil is the second largest country with COVID-19 positive cases worldwide. Due to the potent spread of the virus and the scarcity of kits and supplies, the Brazilian Ministry of Health has granted authorization for the use of kits available during this emergency, without an accurate evaluation of their performance. This study compared the performance and cost-effectiveness of seven molecular assays/kits available in São Paulo, Brazil, for SARS-CoV-2 diagnosis. Materials and methods: A total of 205 nasopharyngeal/oropharyngeal samples from suspected cases of COVID-19, were tested using the following assays: (i) GeneFinder COVID-19 plus RealAmp kit; (ii) 2019-nCoV RNA PCR-Fluorescence Probing, Da An Gene Co.; (iii) in-house RT-qPCR SARS-CoV-2 IAL; (iv) 2019-nCoV kit, IDT; (v) molecular SARS-CoV-2 (E) kit, Bio-Manguinhos; (vi) Allplex 2019-nCoV modified Assay, Seegene Inc, and (vii) Biomol one-step COVID-19 kit, IBMP. The criteria for determining a SARS-CoV-2 true positive result included the cycle threshold cut-off values, the characteristics of exponential/linear curves, the gene target diversity, and a positive result in at least two assays. Results: The overall sensitivity of the assays listed were GeneFinder 83.6%, Da An Gene 100.0%, IAL 90.4%, IDT 94.6%, Bio-Manguinhos 87.7%, Allplex 97.3%, and IBMP 87.7%. The minor sensitive gene target was RdRP. Although all assays had a Cohen's Kappa index ≥0.893, the best tests used multiplex assays identifying N-gene and/or E-gene targets. Conclusion: All assays tested accurate for diagnosis, but considering cost-effectiveness (cost, time consumption, number of samples tested, and performance), the in-house IAL assay was ideal for COVID-19 diagnosis in São Paulo, Brazil.

Comparison of droplet spread in standard and laminar flow operating theatres: SPRAY study group.

BACKGROUND: Reducing COVID-19 transmission relies on controlling droplet and aerosol spread. Fluorescein staining reveals microscopic droplets. AIM: To compare the droplet spread in non-laminar and laminar air flow operating theatres. METHODS: A 'cough-generator' was fixed to a theatre trolley at 45°. Fluorescein-stained 'secretions' were projected on to a series of calibrated targets. These were photographed under UV light and 'source detection' software measured droplet splatter size and distance. FINDINGS: The smallest droplet detected was ∼120 µm and the largest ∼24,000 µm. An average of 25,862 spots was detected in the non-laminar theatre, compared with 11,430 in the laminar theatre (56% reduction). The laminar air flow mainly affected the smaller droplets (<1000 µm). The surface area covered with droplets was: 6% at 50 cm, 1% at 2 m, and 0.5% at 3 m in the non-laminar air flow; and 3%, 0.5%, and 0.2% in the laminar air flow, respectively. CONCLUSION: Accurate mapping of droplet spread in clinical environments is possible using fluorescein staining and image analysis. The laminar air flow affected the smaller droplets but had limited effect on larger droplets in our 'aerosol-generating procedure' cough model. Our results indicate that the laminar air flow theatre requires similar post-surgery cleaning to the non-laminar, and staff should consider full personal protective equipment for medium- and high-risk patients.

Traumatic Nonmissile Penetrating Transnasal Anterior Skull Base Fracture and Brain Injury with Cerebrospinal Fluid Leak: Intraoperative Leak Detection and an Effective Reconstruction Procedure for a Localized Skull Base Defect Especially After Coronavirus Disease 2019 Outbreak.

BACKGROUND: Cerebrospinal fluid (CSF) leakage after penetrating skull base injury is relatively rare compared with close head injuries involving skull base fractures. CASE DESCRIPTION: We report the case of a 65-year-old man who had presented with epistaxis and serous rhinorrhea. When he had fallen to the ground near his bee boxes, a garden pole had poked into his right nostril. He had instantly removed the pole from his nostril himself. However, immediately after removal of the pole, he had developed nasal bleeding and serous rhinorrhea. He then drove to our emergency room. Computed tomography showed pneumocephalus with a minor cerebral contusion in the left frontal lobe and a penetrating injury in the left anterior skull base. His CSF leakage had not resolve spontaneously within 1 week after the injury with strict bed rest. We repaired the CSF leakage using a fat (adipose tissue)-on-fascia autograft plug and caulked the defect in the anterior skull base with the fat-on-fascia graft (FFG) plug through the left nostril with endoscopic guidance. The CSF rhinorrhea was successfully controlled. Intranasal local application of fluorescein aided in the detection of the direction of flow of the CSF leakage. CONCLUSIONS: Endonasal endoscopic caulking of a skull base defect using an FFG plug can be useful to treat CSF leakage due to the localized skull base defect, especially in the coronavirus disease 2019 pandemic. It is simple, inexpensive, and timesaving. It requires no special skills nor sophisticated instruments that can cause aerosolization, reducing the risk of infection during the surgery.