Researchers know that the virus that causes COVID-19 has been found in the saliva (spit) of individuals who exhibit signs of the disease. Investigators would like to test the ability of three mouthwashes to reduce the levels of this virus in participants' mouths. Investigators will ask participants to use a liquid to swish around in the mouth for 30 seconds and spit it into a collection cup. Investigators will also collect spit from participants before and after participants use the mouthwash. Although participants will have no direct benefits from the study, investigators will gain a wealth of information that would benefit patients who are at risk for COVID-19.
Project aims:
Aim 1: To examine the salivary carriage of COVID-19 in individuals before and after use of
mouthwashes known to reduce oral microbiota, using real time reverse transcriptase
quantitative PCR (Polymerase Chain Reaction) to quantify viral load.
RESEARCH STRATEGY:
A. Recruitment and baseline data:
This is a cross-sectional interventional 4 arm study. 60 subjects between 18-80 years of age
will be recruited from those who are being treated for COVID-19 at the Ohio State University
Medical Center. There is no data upon which to base sample size estimates, and investigators
have landed on this number in an effort to obtain a representative sample of Ohio's
population. If the potential subject is interested in participating, the study coordinator
will collect contact information from the candidate and the consent form will be presented to
these individuals. Subjects will be allowed sufficient time to read and ask questions, and
once sign these forms, a detailed history including information about ethnicity, education,
income, age, sex, race, medical status (including pregnancy) and dental history will be
elicited. Subjects will self-collect saliva in a 5 ml collection tube, following which they
will be asked to use one of 3 mouthwashes (chlorhexidine, povidone iodine, hydrogen peroxide
or control(saline) for thirty seconds and spit them into collection tubes. Investigators will
use Peridex, Hydrogen Peroxide and Betadine, which are commercially available formulations of
chlorhexidine, hydrogen peroxide and betadine respectively, to reduce formulation bias.
Investigators have attached the product labels as appendices to this proposal. The exact
mouthwash will be decided by random chance using a random number generator. Investigators
have generated the following randomization schedule using Microsoft excel: Each subject will
be assigned a number as they are recruited (E.g. first patient to sign the consent will be
patient 1, and the second will be patient 2 and so on). Patients 1 and 2 will receive saline
and 3 will receive chlorhexidine etc. base on the above randomization schedule. Subjects will
then collect saliva in separate collection tubes 15 and 30 minutes after rinsing.
B. Sample collection Saliva will be collected using a methodology described by previous
investigators. Briefly, subjects will be asked to collect saliva in their mouth for 3 minutes
and then continuously drool into a tube for 3 minutes. This method will allow investigators
to collect unstimulated saliva. Saliva will be collected using kits containing RNA
(Ribonucleic Acid) stabilizer (Oragen RNA kit).
C. Measurement / Instrumentation
Total RNA will be isolated from saliva using the mirVana miRNA (Messenger Ribonucleic Acid)
isolation kit (Applied Biosystems). Microbial cells will be lysed and RNA will be extracted
by Acid-Phenol:Chloroform and ethanol precipitation and eluted in nuclease-free water.
Investigators will use well validated primers to quantify copy numbers of COVID-19 spike
protein mRNA (Messenger Ribonucleic Acid) from the saliva. For an absolute quantification of
COVID-19, investigators will use one step RT-PCR (Reverse Transcriptase) assay by using an
appropriate commercially available kit. In this step investigators will convert mRNA to cDNA
(complementary Deoxyribonucleic Acid) and then quantify the cDNA by using targeted primers
and then copy numbers of the COVID-19 will be calculated by comparing ct (cycle
threshold)Values samples against standard curve. In order to generate standard curves for
absolute quantification, target genes will be amplified with PCR. The PCR products will be
cloned into suitable vector. Then plasmids will be extracted, serially diluted, and used as
templates in qPCR (quantitative polymerase chain reaction) for generating standard curves.
D. Internal Validity Subjects will be recruited randomly through voluntary participation.
This will eliminate presentation bias.
E. Data Analysis
Descriptive statistics will be provided as estimates of salivary carriage of COVID-19.
Frequency of salivary carriage of the virus will be expressed as a percent of the total
sample population. Reduction in COVID-19 following mouthwash use will be expressed as fold
change and compared between the three mouthwash groups using parametric tests.
Drug: 0.5% Povidone Iodine
Commercially Available Mouth Rinses and Over the Counter Standard Formulations
Other Name: Betadine Mouthwash
Drug: 0.12% Chlorhexidine Gluconate Mouth Rinse
Commercially Available Mouth Rinses
Other Name: Peridex
Drug: 1% Hydrogen Peroxide
Commercially Available Over the Counter Standard Formulations USP (United States Pharmacopeia)
Other Name: Hydrogen Peroxide
Drug: 0.9% Normal Saline
Commercially Available Over the Counter Standard Formulations USP (United States Pharmacopeia)
Other Name: Salt Water
Inclusion Criteria:
1. adults age 18-80 admitted to The Ohio State University Medical Center,
2. diagnosis of COVID-19 confirmed by Polymerase Chain Reaction (PCR),
3. salivary carriage of COVID-19 virus
4. ability to perform mouthwash rinses and to expectorate.
Exclusion Criteria:
1. allergy to any study mouth rinse,
2. active uncontrolled thyroid disease,
3. pregnancy
4. patients undergoing radioactive iodine therapy.
The Ohio State University Wexner Medical Center
Columbus, Ohio, United States
Investigator: David L Hall, DDS
Contact: 614-292-2622
hall.611@osu.edu
David L Hall, DDS
614-292-2622
hall.611@osu.edu
Purnima Kumar, PhD,DDS
614-804-2112
kumar.83@osu.edu
David L Hall, DDS, Study Director
The Ohio State University College of Dentistry