Australasian Dentist Issue 89

Category AustrAlAsiAn Dentist 75 in silico studies, compounds that could be of interest in the oral viral load fight against sArs-CoV-2, such as hydrogen peroxide, ß-cyclodextrin, flavonoids, essential oils, cetylpyridinium chloride or povidone-iodine, were noticed. 12,14,15 the concept put forward is that some commercially available mouthwash formulations may play a role in decreasing the risk of transmission of sArs-CoV-2. 12–14 However given the limited number of in vitro and in silico studies that have been conducted, there is no scientific evidence to recommend anti-sArs-CoV-2 mouthwashes to control viral load in the oral cavity. At this time, no randomized controlled trial (rCt) studies have been conducted or completed. the research hypothesis is that commercially available mouthwashes with ß-cyclodextrin and citrox (bioflavonoid) antivirals could decrease the sArs-CoV-2 viral load and thus represent an additional barrier measure to fight against the COViD-19 pandemic. the specific question, formulated according to PiCO principles, was as follows: in COViD-19 outpatients (population), does ß-cyclodextrin-citrox rinsing as a posttreatment protocol (intervention) have a beneficial effect on sArs-CoV-2 saliva load parameters (outcome) compared to rinsing with placebo solution (control)? Study design this trial was a multicenter, double-blind rCt with two parallel arms (1:1 ratio). the clinical trial was conducted with the collaboration of four French hospital centers: emile roux Hospital Center (le Puy-en-Velay, France), Protestant infirmary (lyon, France), st Joseph st luc Hospital (lyon, France) and Hospital Center (Mont de Marsan, France). Participants were enrolled at these centers, and monitoring occurred at home. Written informed consent was obtained from each participant before enrollment. the “Committee for the Protection of Persons south Mediterranean iii” (university Hospital Center of nîmes, France) reviewed and approved the clinical trial protocol. the study followed COnsOrt guidelines for trials. the study was conducted in compliance with the principles of the Declaration of Helsinki, the Good Clinical Practice guidelines, and local regulatory requirements. Participants the study’s planned population consisted of sArs-CoV-2 PCr-positive adult outpatients, asymptomatic patients and patients with mild clinical symptoms. Asymptomatic patients were defined as individuals without clinical signs, whereas patients with mild cases were defined as outpatients and patients with mild clinical symptoms or lower or upper respiratory tract infections. 16 eligibility was restricted to adults aged 18-85 years old with a clinical diagnosis of COViD-19 infection, clinical signs that had been present for less than 8 days, virological confirmation, an understanding and acceptance of the trial and written agreement to participate in the trial. the exclusion criteria were pregnancy, breastfeeding, an inability to comply with the protocol, a lack of written agreement, mouthwash use on a regular basis (more than once a week), an inability to answer questions and a lack of cooperation. Randomization and masking eligible patients were randomly assigned (1:1) to either the ß-cyclodextrin-citrox mouthwash (CDCM) group or the placebo group. randomization was stratified by center. randomization was integrated into the inclusion process by generating a random number after automatic analysis of the center’s prior randomizations. the randomization codes for each allocation schedule cohort were generated individually. the randomization code was allocated to each participant in sequence in the order of registration, and then participants received experimental materials labeled with the same code. A randomization sequence was prepared by a clinical senior investigator not involved in the trial using e-CrF Voozalyon 1.3 (Voozanoo, Caluire, France). Once enrolled, participants each received three 200 ml medication vials. the contents of the medication vials were unknown to participants, investigators and staff. each vial contained either a mouthwash containing the antiviral components (ß-cyclodextrin (0·1%) and citrox (0·01%)) or placebo with similar appearance and content without the above-mentioned antiviral components; the labels on the vials were identical. All participants, investigators, statisticians, and laboratory staff were masked to treatment allocation. Procedures Participants were instructed to use three mouthwashes per day (at 9 a.m., 2 p.m., and 7 p.m.), with either 30 ml of CDCM or placebo, both provided by Curaden AG (Kriens, switzerland) for 1 min (Appendix Fig. s1). Participants were instructed to collect their saliva by trained nurses using the “saliva Collection system” kit (Greiner Bio-one, Kremsmünster, Austria). saliva sampling was performed on the first day at t1 (9 a.m., before the first mouthwash) and then at t2 (1 p.m.) and t3 (6 p.m.). On the following 6 days, only one sample was taken at 3 p.m. All samples were refrigerated and shipped by similar means of transport to laboratories of the national reference Center for respiratory Viruses (Hospices Civils de lyon, lyon, France) with PCr testing equipment, where procedures for sArs-CoV2 rnA detection and quantification were performed. rnA extraction was realized using the nuclisens easyMAG instrument (bioMérieux, Marcy-l’etoile, France) following the manufacturer’s instructions. rdrp-iP2 and rdrp-iP4 quantitative rt- PCr was used to detect sArs-CoV-2 with the invitrogen superscripttM iii Platinum One-step qrt-PCr system (invitrogen, illkirch, France). When a sample was positive for rdrp- iP4, the quantification of the number of rnA copies was performed according to a scale ranging from 102 to 106 copies per μl. the viral load in saliva was calculated as the number of rnA copies per ml of saliva. Primer and probe sequences (eurofins, Genomics, Germany; Appendix table s1) correspond to the rdrp-iP2, the rdrp-iP4 assays designed at the institut Pasteur to target a section of the rdrp gene based on the sequences of sArs- CoV-2 made available on the Global initiative on sharing All influenza Data database on January 11, 2020. For further details see the appendix (pp 3-4). Outcomes the general objective of the study was to describe the evolution of salivary sArs- CoV-2 viral load (copies/ml) in COViD-19 outpatients receiving mouthwashes with or without antivirals. Primary outcome measures included changes from baseline sArs-CoV-2 in salivary samples at two time points, 4 and 9 hours, within 1 day after the first intake. secondary outcome measures included changes from baseline sArs-CoV-2 in salivary samples at 6 days after the first dose. exploratory outcomes were i) the virological efficacy of the experimental therapy (CDCM) compared to the placebo as evaluated by the quantity of sArs- CoV-2 in salivary samples at t1 and 4 hours after the first and second intakes and ii) the virological efficacy of the experimental therapy compared to the placebo as evaluated by the quantity of sArs-CoV-2 in salivary samples on day 1 at t2 and t3 and day 7. Statistical analysis sample-size calculations were estimated using the freeware stPlAn (Version lInICal