Circulation of SARS-CoV-Related Coronaviruses and Alphacoronaviruses in Bats from Croatia.

Bats are natural hosts of various coronaviruses (CoVs), including human CoVs, via an assumed direct zoonotic spillover or intermediate animal host. The present study aimed to investigate the circulation of CoVs in a bat colony in the Mediterranean region of Croatia. Guano and individual droppings from four bat species were sampled and tested with the E-gene sarbecovirus RT-qPCR, the pan-CoV semi-nested RT-PCR targeting the RdRp gene and NGS. Furthermore, bat blood samples were investigated for the presence of sarbecovirus-specific antibodies with the surrogate virus neutralization test (sVNT). The initial testing showed E-gene Sarebeco RT-qPCR reactivity in 26% of guano samples while the bat droppings tested negative. The application of RdRp semi-nested RT-PCR and NGS revealed the circulation of bat alpha- and betaCoVs. Phylogenetic analysis confirmed the clustering of betaCoV sequence with SARS-CoV-related bat sarbecoviruses and alpha-CoV sequences with representatives of the Minunacovirus subgenus. The results of sVNT show that 29% of bat sera originated from all four species that tested positive. Our results are the first evidence of the circulation of SARS-CoV-related coronaviruses in bats from Croatia.

Virus transmission by ultrasonic scaler and its prevention by antiviral agent: An in vitro study.

BACKGROUND: It is well recognized that dental procedures represent a potential way of infection transmission. With the COVID-19 pandemic, the focus of dental procedure associated transmission has rapidly changed from bacteria to viruses. The aim was to develop an experimental setup for testing the spread of viruses by ultrasonic scaler (USS) generated dental spray and evaluate its mitigation by antiviral coolants. METHODS: In a virus transmission tunnel, the dental spray was generated by USS with saline coolant and suspension of Equine Arteritis Virus (EAV) delivered to the USS tip. Virus transmission by settled droplets was evaluated with adherent RK13 cell lines culture monolayer. The suspended droplets were collected by a cyclone aero-sampler. Antiviral activity of 0.25% NaOCl and electrolyzed oxidizing water (EOW) was tested using a suspension test. Antiviral agents' transmission prevention ability was evaluated by using them as a coolant. RESULTS: In the suspension test with 0.25% NaOCl or EOW, the TCID50/mL was below the detection limit after 5 seconds. With saline coolant, the EAV-induced cytopathic effect on RK13 cells was found up to the distance of 45 cm, with the number of infected cells decreasing with distance. By aero-sampler, viral particles were detected in concentration ≤4.2 TCID50/mL. With both antiviral agents used as coolants, no EAV-associated RK-13 cell infection was found. CONCLUSION: We managed to predictably demonstrate EAV spread by droplets because of USS action. More importantly, we managed to mitigate the spread by a simple substitution of the USS coolant with NaOCl or EOW.

A Throat Lozenge with Fixed Combination of Cetylpyridinium Chloride and Benzydamine Hydrochloride Has Direct Virucidal Effect on SARS-CoV-2

Viruses are the most common causative agents of inflammation in the oral cavity and throat region. Most respiratory tract infections are self-limiting and require no specific treatment. However, patients often use different self-medication therapies that can treat both the symptoms and the cause. Throat lozenges with a fixed combination of benzydamine hydrochloride and cetypiridinium chloride (BH/CPC) have been shown to provide effective symptomatic relief for sore throat, but their effect on viruses has not been investigated to date. The antiseptic, cetylpyridinium chloride (CPC), has already been described as a successful bactericide. In addition, there are some studies suggesting its efficacy against certain enveloped viruses. Thus, the aim of our study was to examine the virucidal activity of CPC and a combination of BH/CPC as a free active substance or as lozenge on SARS-CoV-2 in vitro. Under in-laboratory simulated conditions of lozenge administration, we incubated SARS-CoV-2 with three different concentrations of each of the active substances, CPC, free BH/CPC or BH/CPC, as a lozenge suspension for 1 min, 5 min and 15 min of contact time. Infective viral particles were detected in cell cultures and the viral titre was calculated accordingly. Our results show that all active substances in high-concentration suspensions, as well as a medium concentration of the BH/CPC combination, exhibited a 4-log reduction in viral titre. Additionally, the highest concentration of BH/CPC as a lozenge had a faster virucidal effect compared to CPC as a free active substance alone, since a contact time as short as 1 min reduced the initial virus concentration by more than 4-log. This study demonstrates the effective strong virucidal effect of the lozenge, with the possibility of viral load reduction in the oral cavity and, consequently, reduced risk of viral transmission.