Milk casein prevents inactivation effect of black tea galloylated theaflavins on SARS-CoV-2 in vitro (preprint)

Repeated emergence of highly contagious and potentially immune-evading variant SARS-CoV-2 is posing global health and socioeconomical threats. For suppression of the spread of the virus infection among people, a procedure to inactivate virus in saliva may be useful, because saliva of infected persons is the major origin of droplets and aerosols that mediate viral transmission to nearby persons. We previously reported that SARS-CoV-2 is rapidly and remarkably inactivated by treatment in vitro with tea including green tea, roasted green tea, oolong tea and black tea. Tea catechin-derived compounds including theaflavins (TFs) with (a) galloyl moiety(ies) showed this activity. Although black tea is popularly consumed worldwide, a lot of people consume it with sugar, milk, lemon juice, and so on. But it has not been determined whether these ingredients may influence the inactivation effect of black tea against SARS-CoV-2. Moreover, it has not been revealed whether black tea is capable of inactivating variant viruses such as delta variant. Here we examined the effect of black tea on some variants in the presence or absence of sugar, milk, and lemon juice in vitro. Black tea and galloylated TFs remarkably inactivated alpha, gamma, delta and kappa variants. Intriguingly, an addition of milk but not sugar and lemon juice totally prevented black tea from inactivating alpha and delta variant viruses. The suppressive effect was also exerted by milk casein. These results suggest the possibility that intake of black tea without milk by infected persons may result in inactivation of the virus in saliva and attenuation of spread of SARS-CoV-2 to nearby persons through droplets. Clinical studies are required to investigate this possibility.

Rapid inactivation in vitro of SARS-CoV-2 in saliva by black tea and green tea (preprint)

Saliva plays major roles in human-to-human transmission of the SARS-CoV-2. Recently we reported that black, green and oolong tea significantly inactivated SARS-CoV-2 within 1 min. Theaflavin-3,3-di-gallate (TFDG), theasinensin A (TSA) and (-) epigallocatechin gallate (EGCG) were involved in the anti-viral activities. Here we examined how long period is required for the compounds to inactivate the virus. We also assessed whether tea inactivates SARS-CoV-2 diluted in human saliva. Treatment of SARS-CoV-2 with 500 M TFDG or TSA for 10 sec reduced the virus titer to undetectable levels (less than 1/1,000). Black and green tea decreased virus titer to less than 1/100 within 10 sec even in saliva. These findings suggest a possibility that intake of, or gargling with, tea may inactivate SARS-CoV-2 in saliva in infected individuals, which may eventually attenuate spread of COVID-19 within a population, although clinical studies are required to test this hypothesis by determining the intensity and duration of the anti-viral effect of tea in saliva in humans.

Significant inactivation of SARS-CoV-2 by a green tea catechin, a catechin-derivative and galloylated theaflavins in vitro (preprint)

Potential effects of teas and their constituents on SARS-CoV-2 infection were studied in vitro. Infectivity of SARS-CoV-2 was significantly reduced by a treatment with green tea, roasted green tea or oolong tea. Most remarkably, exposure to black tea for 1 min decreased virus titer to an undetectable level (less than 1/1,000 of untreated control). An addition of (-) epigallocatechin gallate (EGCG) significantly inactivated SARS-CoV-2, while theasinensin A (TSA) and galloylated theaflavins including theaflavin 3, 3-di-gallate (TFDG) had more remarkable anti-viral activities. Virus treated with TSA at 500 M or TFDG at 100 M showed less than 1/10,000 infectivity compared with untreated virus. TSA and TFDG significantly inhibited interaction between recombinant ACE2 and RGD of S protein. These results strongly suggest that EGCG, and more remarkably TSA and galloylated theaflavins, inactivate the novel coronavirus.

High affinity modified ACE2 receptors prevent SARS-CoV-2 infection (preprint)

The SARS-CoV-2 spike protein binds to the human angiotensin-converting enzyme 2 (ACE2) receptor via receptor binding domain (RBD) to enter into the cell. Inhibiting this interaction is a main approach to block SARS-CoV-2 infection and it is required to have high affinity to RBD independently of viral mutation for effective protection. To this end, we engineered ACE2 to enhance the affinity with directed evolution in human cells. Three cycles of random mutation and cell sorting achieved more than 100-fold higher affinity to RBD than wild-type ACE2. The extracellular domain of modified ACE2 fused to the Fc region of the human immunoglobulin IgG1 had stable structure and neutralized SARS-CoV-2 pseudotyped lentivirus and authentic virus with more than 100-fold lower concentration than wild-type. Engineering ACE2 decoy receptors with directed evolution is a promising approach to develop a SARS-CoV-2 neutralizing drug that has affinity comparable to monoclonal antibodies yet displaying resistance to escape mutations of virus.

Rapid Inactivation of SARS-CoV-2 by Silicon Nitride, Copper, and Aluminum Nitride (preprint)

IntroductionViral disease spread by contaminated commonly touched surfaces is a global concern. Silicon nitride, an industrial ceramic that is also used as an implant in spine surgery, has known antibacterial activity. The mechanism of antibacterial action relates to the hydrolytic release of surface disinfectants. It is hypothesized that silicon nitride can also inactivate the coronavirus SARS-CoV-2. MethodsSARS-CoV-2 virions were exposed to 15 wt.% aqueous suspensions of silicon nitride, aluminum nitride, and copper particles. The virus was titrated by the TCD50 method using VeroE6/TMPRSS2 cells, while viral RNA was evaluated by real-time RT-PCR. Immunostaining and Raman spectroscopy were used as additional probes to investigate the cellular responses to virions exposed to the respective materials. ResultsAll three tested materials showed >99% viral inactivation at one and ten minutes of exposure. Degradation of viral RNA was also observed with all materials. Immunofluorescence testing showed that silicon nitride-treated virus failed to infect VeroE6/TMPRSS2 cells without damaging them. In contrast, the copper-treated virus suspension severely damaged the cells due to copper ion toxicity. Raman spectroscopy indicated differential biochemical cellular changes due to infection and metal toxicity for two of the three materials tested. ConclusionsSilicon nitride successfully inactivated the SARS-CoV-2 in this study. The mechanism of action was the hydrolysis-mediated surface release of nitrogen-containing disinfectants. Both aluminum nitride and copper were also effective in the inactivation of the virus. However, while the former compound affected the cells, the latter compound had a cytopathic effect. Further studies are needed to validate these findings and investigate whether silicon nitride can be incorporated into personal protective equipment and commonly touched surfaces, as a strategy to discourage viral persistence and disease spread.