Before enrolment in the study, we will perform a thorough clinical examination of each
patient, which will involve assessing several periodontal parameters at six sites around
each tooth on all teeth, excluding third molars: plaque index (PlI), probing depth (PD),
gingival recession (GR), clinical attachment loss (CAL), bleeding on probing (BOP),
furcation involvement, and tooth mobility. All clinical parameters will be measured using
a William's periodontal probe (Hu-Friedy, USA, PQW6). Local X-rays of sites affected by
periodontitis will be taken. The performed clinical examination will not differ in any
way from the examination conducted on all patients, even those not included in our study.
The population of included subjects will consist of individuals of both sexes, aged 25-70
years, smokers and non-smokers, with moderate or advanced stage of chronic periodontitis
(stage III, IV). They will be selected based on the following inclusion criteria:
clinical attachment loss ≥ 5 mm on at least two teeth in two different jaw quadrants;
presence of stable occlusion and at least 16 teeth, among which at least 12 teeth are
suitable for evaluation (excluding wisdom teeth, teeth with orthodontic wires, bridges,
crowns, and implants). We will exclude individuals who: suffer from chronic systemic
diseases (diabetes, cancer, HIV infection, metabolic bone diseases, and diseases that
interfere with wound healing processes); are undergoing radiation or chemotherapy; are
taking immunosuppressants, antiepileptic drugs, calcium antagonists, nonsteroidal
anti-inflammatory drugs; have been treated with antibiotics in the past 12 months; have a
known allergy to CHX; have undergone scaling and root planing or surgical periodontal
treatment in the past year; are pregnant or breastfeeding. Patients will be informed
about the purpose of the research in writing. Prior to the commencement of the study, all
patients will fill out an informed consent form.
Subjects will then be divided into three groups of 20; the first group will use a
mouthwash consisting of EOS (active ingredient) for one month after the non-surgical
treatment; the second group will use a mouthwash containing CHX (positive control); and
the third group will use distilled water (negative control/placebo). Randomization will
be performed using a computer program. Only one member of the research group, who will
also prepare the mouthwashes, will know the content of the randomization table or the
type of mouthwash used by each individual. Identification codes will remain hidden from
all other researchers and subjects until the final follow-up examinations. The packaging
of the mouthwashes will be identical, and the active ingredients will not be
distinguishable by color.
Samples of subgingival fluid will be collected at the beginning of the test period for
the purpose of microbiological characterization using sterile paper points from the
buccal surface of four teeth (sites with the greatest probing depths in each quadrant)
and transported to the laboratory of the Institute of Microbiology and Immunology using a
transport medium (RTF). The microbiological analysis will be performed using Quantitative
Polymerase Chain Reaction (qPCR).
Subsequently, all patients will undergo a non-surgical periodontal treatment, which will
include instruction and motivation for proper oral hygiene, removal of hard and soft
plaque deposits using a piezoelectric device and ultrasonic tip, and root planing and
scaling under local anesthesia. Each subject will then receive a package with an
identification number and the label A, B, or C on it, according to the computer-generated
randomization scheme. One-third of the patients will receive EOS in the package,
one-third will receive CHX, and one-third will receive distilled water. Subjects will
then rinse their oral cavities with 15 ml of their allocated mouthwash twice daily (in
the morning and evening) for 30 seconds for four consecutive weeks.
After four weeks, subjects will be invited for their first follow-up examination. They
will report any side effects and return the packaging of the for compliance control. They
will also be asked about the organoleptic properties of the mouthwash they used. We will
perform a thorough clinical examination again and collect samples of subgingival fluid in
the same manner as during the initial examination. The second follow-up examination will
be conducted three months after the oral hygiene phase and will include a clinical
assessment and collection of subgingival fluid samples.
The primary outcome variable will be the number of residual diseased sites (PD > 4mm +
BOP). Secondary outcome variables will include plaque index assessment, BOP, PD, and
quantity of periodontopathogens. Since teeth are "nested" within patients and probing
sites are "nested" within teeth, the collected data is interdependent. A multilevel
multiple logistic regression model will therefore be used to examine associations between
variables.