SARS-CoV-2 Short-Time Infection Produces Relevant Cytopathic Effects in Vero E6 Cell Line

Zupin, Luisa; Fontana, Francesco; Gratton, Rossella; Milani, Margherita; Clemente, Libera; Pascolo, Lorella; Ruscio, Maurizio; Crovella, Sérgio

doi:10.3390/ijerph18179020

Cited by 11 publications

(14 citation statements)

References 20 publications

(24 reference statements)

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“…High concentration of ACE-2 receptors was found to correlate with high rates of cell infection. Cell lines such as apical plasma membrane of polarized respiratory epithelial cells, colon carcinoma cell line (Caco-2) ( Pascoal et al., 2021 ), a lung carcinoma cell line (Calu-3) ( Kanimozhi et al., 2021 ) and Vero E6 cells ( Zupin et al., 2021 ) have been put forward as candidates for models in COVID19 research. Human cancer cell lines were screened for the SARS-CoV-2 cellular entry factors ACE2 and TMPRSS2 based on RNA-seq data of the Cancer Cell Line Encyclopedia (CCLE).…”

Section: In Vitro Models Of Sars-cov-2 Infectionmentioning

confidence: 99%

“…This model was also used to aid in a SARS-CoV-2 pathogenesis study in which the accumulation of lipids was seen in infected in vitro Vero E6 cells as well as lungs of COVID19 patients, suggesting that lipids are involved in SARS-CoV-2 pathogenesis ( Nardacci et al., 2021 ). Vero E6 cells were recently used in a study determining the in vitro minimal exposure time and viral concentration needed to establish persistent SARS-CoV-2 infection ( Zupin et al., 2021 ). Vero E6 cell lines have also been used in multiple studies of SARS-CoV-2 repurposed antivirals ( Canal et al., 2021 , Leneva et al., 2021 ), protease inhibitors ( Cui and Jia, 2021 ; Han et al., 2021 ) as well as inhibiting peptides ( Kelesidis et al., 2021 ).…”

Section: In Vitro Models Of Sars-cov-2 Infectionmentioning

confidence: 99%

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Experimental Models of COVID-19

Caldera-Crespo

Paidas

Roy

et al. 2022

Front. Cell. Infect. Microbiol.

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COVID-19 is the most consequential pandemic of the 21st century. Since the earliest stage of the 2019-2020 epidemic, animal models have been useful in understanding the etiopathogenesis of SARS-CoV-2 infection and rapid development of vaccines/drugs to prevent, treat or eradicate SARS-CoV-2 infection. Early SARS-CoV-1 research using immortalized in-vitro cell lines have aided in understanding different cells and receptors needed for SARS-CoV-2 infection and, due to their ability to be easily manipulated, continue to broaden our understanding of COVID-19 disease in in-vivo models. The scientific community determined animal models as the most useful models which could demonstrate viral infection, replication, transmission, and spectrum of illness as seen in human populations. Until now, there have not been well-described animal models of SARS-CoV-2 infection although transgenic mouse models (i.e. mice with humanized ACE2 receptors with humanized receptors) have been proposed. Additionally, there are only limited facilities (Biosafety level 3 laboratories) available to contribute research to aid in eventually exterminating SARS-CoV-2 infection around the world. This review summarizes the most successful animal models of SARS-CoV-2 infection including studies in Non-Human Primates (NHPs) which were found to be susceptible to infection and transmitted the virus similarly to humans (e.g., Rhesus macaques, Cynomolgus, and African Green Monkeys), and animal models that do not require Biosafety level 3 laboratories (e.g., Mouse Hepatitis Virus models of COVID-19, Ferret model, Syrian Hamster model). Balancing safety, mimicking human COVID-19 and robustness of the animal model, the Murine Hepatitis Virus-1 Murine model currently represents the most optimal model for SARS-CoV-2/COVID19 research. Exploring future animal models will aid researchers/scientists in discovering the mechanisms of SARS-CoV-2 infection and in identifying therapies to prevent or treat COVID-19.

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Section: In Vitro Models Of Sars-cov-2 Infectionmentioning

confidence: 99%

Section: In Vitro Models Of Sars-cov-2 Infectionmentioning

confidence: 99%

Experimental Models of COVID-19

Caldera-Crespo

Paidas

Roy

et al. 2022

Front. Cell. Infect. Microbiol.

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“…The SARS-CoV-2 selected variant isolated, amplified and quantified on Vero E6 cell line, is employed in the aerosolization [ 30 , 31 , 32 ]. The virus is diluted at 10 5 and 10 4 PFU/mL for aerosol generation in the infection medium composed by MEM supplemented with 2 mM glutamine, 2% fetal bovine serum and 100 U/mL penicillin/streptomycin.…”

Section: Methodsmentioning

confidence: 99%

“…The infectivity of the bio-aerosol samples is determined in vitro on Vero E cells (epithelial kidney standard cell line derived from Cercopithecus aethiops ). Vero E6 cells still represent a gold standard for viral infection experiments and are highly susceptible to SARS-CoV-2 infection [ 31 ].…”

Section: Methodsmentioning

confidence: 99%

Operative Protocol for Testing the Efficacy of Nasal Filters in Preventing Airborne Transmission of SARS-CoV-2

Semeraro

Gaetano

Zupin

et al. 2022

IJERPH

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Background: Standardized methods for testing Viral Filtration Efficiency (VFE) of tissues and devices are lacking and few studies are available on aerosolizing, sampling and assessing infectivity of SARS-CoV-2 in controlled laboratory settings. NanoAg-coated endonasal filters appear a promising aid for lowering viable virus inhalation in both adult and younger populations (e.g., adolescents). Objective: to provide an adequate method for testing SARS-CoV-2 bioaerosol VFE of bio-gel Ag nanoparticles endonasal filters, by a model system, assessing residual infectivity as cytopathic effect and viral proliferation on in vitro cell cultures. Methods: A SARS-CoV-2 aerosol transmission chamber fed by a BLAM aerosol generator produces challenges (from very high viral loads (105 PFU/mL) to lower ones) for endonasal filters positioned in a Y shape sampling port connected to a Biosampler. An aerosol generator, chamber and sampler are contained in a class II cabinet in a BSL3 facility. Residual infectivity is assessed from aliquots of liquid collecting bioaerosol, sampled without and with endonasal filters. Cytopathic effect as plaque formation and viral proliferation assessed by qRT-PCR on Vero E6 cells are determined up to 7 days post inoculum. Results: Each experimental setting is replicated three times and basic statistics are calculated. Efficiency of aerosolization is determined as difference between viral load in the nebulizer and in the Biosampler at the first day of experiment. Efficiency of virus filtration is calculated as RNA viral load ratio in collected bioaerosol with and without endonasal filters at the day of the experiment. Presence of infectious virus is assessed by plaque forming unit assay and RNA viral load variations. Conclusions: A procedure and apparatus for assessing SARS-CoV-2 VFE for endonasal filters is proposed. The apparatus can be implemented for more sophisticated studies on contaminated aerosols.

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“…SARS-CoV-2, previously isolated in the BSL3 facility of the San Polo hospital (ASUGI, Monfalcone, GO, Italy) from the supernatant of infected Vero E6 cell line (epithelial kidney normal cell line from Cercopithecus aethiops , ATCC CRL-1586), was employed in the aerosolization ( Supplementary Information File S1 ) [ 50 , 51 ]. The virus was quantified by using a standard plaque forming unit (PFU) assay [ 52 ].…”

Section: Methodsmentioning

confidence: 99%

Evaluation of Residual Infectivity after SARS-CoV-2 Aerosol Transmission in a Controlled Laboratory Setting

Zupin

Licen

Milani

et al. 2021

IJERPH

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Severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) is mainly transmitted through respiratory droplets, aerosols, or direct contact with fomites from an infected subject. It has been reported that SARS-CoV-2 is stable and viable in aerosol up to 16 h in controlled laboratory conditions. However, the aerosolization conditions varied a lot between the studies. In this work, an experimental laboratory model of SARS-CoV-2 aerosolization was established, employing an impinger nebulizer, a cylindrical chamber for aerosol travel, and a SKC biosampler for the collection of particles. The efficiency of the system was assessed based on the molecular determination of the viral load in the nebulizer after the aerosolization and in the aerosol collected at the end of the travel. Moreover, the residual infectivity was tested in vitro on the Vero E6 cell line, through the observation of the cytopathic effect (CPE), and the quantification of the viral load in the supernatants at 7 days post inoculation (dpi). A high RNA viral load was found in the SKC biosampler after aerosolization, indicating that it was possible to transport a high virus titer through the 30-cm chamber with all the dilutions (initial 105, 104, 103 plaque forming unit—PFU/mL). At the 7 dpi, an increment of the RNA viral load was determined for the dilutions 105 and 104 PFU/mL tested, while only the initial 105 PFU/mL resulted in visible CPE. Our findings allowed us to achieve the resilience of SARS-CoV-2 in aerosol form, at a concentration comparable to those reported for clinical samples. This mode of transmission should be considered for the mitigation and preventive measures to counteract SARS-CoV-2 spreading.

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SARS-CoV-2 Short-Time Infection Produces Relevant Cytopathic Effects in Vero E6 Cell Line

Cited by 11 publications

References 20 publications

Experimental Models of COVID-19

Experimental Models of COVID-19

Operative Protocol for Testing the Efficacy of Nasal Filters in Preventing Airborne Transmission of SARS-CoV-2

Evaluation of Residual Infectivity after SARS-CoV-2 Aerosol Transmission in a Controlled Laboratory Setting

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