J Clin Periodontol 2016; doi: 10.1111/jcpe.

12559

Non-surgical periodontal Raluca Cosgarea1,2,, Raluca Juncar2,

Christian Heumann3, Roxana Tristiu2,
Liana Lascu2, Nicole Arweiler1,

treatment in conjunction with 3 Andreas Stavropoulos4 and

Anton Sculean5
1
Department of Periodontology, Philipps

or 7 days systemic University Marburg, Marburg, Germany;

2
Clinic for Prosthetic Dentistry, University
Iuliu Hatieganu, Cluj-Napoca, Romania;

administration of amoxicillin and

3
Department for Statistics, Ludwig-
Maximilians University, Munich, Germany;
4
Department of Periodontology, Faculty of
dentistry, Malmo University, Malmo , Sweden;

metronidazole in severe 5
Department of Periodontology, School of
Dental Medicine, University of Bern, Bern,
Switzerland

chronic periodontitis patients.

A placebo-controlled randomized
clinical study
Cosgarea R, Juncar R, Heumann C, Tristiu R, Lascu L, Arweiler N, Stavropoulos
A, Sculean A. Non-surgical periodontal treatment in conjunction with 3 or 7 days
systemic administration of amoxicillin and metronidazole in severe chronic
periodontitis patients. A placebo-controlled randomized clinical study. J Clin
Periodontol 2016; doi: 10.1111/jcpe.12559.

Abstract
Aim: To evaluate the effect of 3 or 7 days systemic administration of amoxicillin
(AMX) and metronidazole (MET) or placebo as adjunct to non-surgical peri-
odontal treatment in severe chronic periodontitis patients.
Methods: One hundred and two patients with severe chronic periodontitis [e.g. 1
site with probing pocket depth (PD) 6 mm per quadrant] were randomly
divided into three equally sized groups and treated with either scaling and root
planing within 24 h (SRP) + placebo (Group A) or SRP + AMX + MET (both

500 mg 9 3 times daily) for 3 days (Group B) or SRP + AMX + MET (both Dr. Raluca Cosgarea presented this paper
500 mg 9 3 times daily) for 7 days (Group C). PD, clinical attachment level for the Jaccard-EFP Research Prize
competition at EuroPerio8 in London.
(CAL), bleeding on probing (BOP), full-mouth plaque scores (FMPS) and gingi-
val bleeding index (GBI) were assessed prior to treatment (baseline), and at 3 and Key words: amoxicillin; metronidazole;
6 months post-treatment. The primary outcome variable was the difference (D) in non-surgical periodontal treatment; systemic
the number of sites with PD 6 mm. antibiotics
Results: Ninety-one patients completed the study. At both 3 and 6 months, all
three treatment protocols resulted in statistically significant improvements com- Accepted for publication 26 March 2016
pared to baseline for all evaluated clinical parameters (p < 0.001). At 6 months,

Conflict of interest and sources of funding statement

The authors declare to have no conflict of interest regarding this study. The study has been funded by the authors own
institutions.

2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd 1
2 Cosgarea et al.

a statistically significantly greater reduction in the mean number of sites with

PD 6 mm was observed in group B (28.62  15.32 sites) and group C
(30.45  15.04 sites) compared to the placebo group (17.10  14.68 sites). Fur-
thermore, both the 3- and the 7-day antibiotic regimen resulted in statistically sig-
nificantly higher clinical improvements compared to the placebo group (p < 0.05).
Conclusion: The present findings indicate that in patients with severe chronic peri-
odontitis, non-surgical periodontal therapy in conjunction with a 3 or 7 days sys-
temic administration of AMX + MET may lead to significantly greater clinical
improvements compared to non-surgical therapy alone.

Periodontal treatment aims at reduc-
tion in the subgingival oral biofilm,
including pathogenic species, to
achieve homeostasis. This is mainly
accomplished by mechanical removal
of the bacterial deposits from the
tooth surfaces, i.e. scaling and root
planing (SRP). However, bacteria
residing in root concavities, dentin
tubules or within soft tissues are not
easily reached with mechanical
instrumentation, and therefore SRP
rarely leads to complete elimination
of the subgingival biofilm (Cugini
et al. 2000, Carvalho et al. 2005).
Furthermore, inefficient reduction in
the subgingival microbial load is
associated with unsuccessful treat-
ment outcomes. For example,
persistence of Aggregatibacter actino-
mycetemcomitans (A. actinomycetem-
comitans), Porphyromonas gingivalis
(P. gingivalis) and other bacterial
species after periodontal treatment
has been associated with additional
tissue destruction (Renvert et al.
1990, Mombelli et al. 1994, 2000),
whereas improved clinical outcomes
have been observed when such peri-
odontopathogenic bacteria were not
detected (Grossi et al. 1994, Haffajee
& Socransky 1994, Dahlen et al.
1996).
Given these limitations, the
adjunctive use of antibacterial agents
(antibiotics and antiseptics) may be
considered. The rationale is that
after mechanical disruption of the
biofilm through instrumentation,
bacteria become more susceptible to
such agents (Marsh 2005). A large
variety of antibiotics (AB) has
indeed been investigated as possible
adjunct to SRP: amoxicillin (with or
without clavulanic acid), metronida-
zole, clindamycin, doxycycline,
azithromycin, moxifloxacin, tetracy-
cline, spiramycin and combinations
thereof (Slots 2004, Guentsch et al.
2008, Herrera et al. 2008, 2012,
Heitz-Mayfield 2009, Flemmig et al.
2011, Griffiths et al. 2011). The most
frequent regime includes a combina-
tion of amoxicillin (AMX) and
metronidazole (MET) and several
studies have shown statistically sig-
nificant reduction in periodontal
pathogens and inflammatory cytoki-
nes after SRP with adjunctive use of
AMX + MET compared to SRP
alone (Pavicic et al. 1994, Goutoudi
et al. 2004, Cionca et al. 2009, 2010,
Ribeiro Edel et al. 2009, Mestnik
et al. 2010, 2012, Yek et al. 2010,
Griffiths et al. 2011, Heller et al.
2011, Silva et al. 2011, Varela et al.
2011, Feres et al. 2012, de Lima Oli-
veira et al. 2012). Several clinical
studies have provided evidence for
significantly better clinical outcomes,
i.e. probing pocket depth (PD)
reduction and clinical attachment
level (CAL) gain, following the
adjunctive use of AMX + MET to
SRP compared to the treatment with
SRP alone (van Winkelhoff et al.
1989, Pavicic et al. 1994, Winkel
et al. 2001, Matarazzo et al. 2008,
Cionca et al. 2009, 2010, Ribeiro
Edel et al. 2009, Mestnik et al. 2010,
2012, Yek et al. 2010, Griffiths et al.
2011, Heller et al. 2011, Silva et al.
2011, Varela et al. 2011, Feres et al.
2012). A recent meta-analysis on the
adjunctive systemic use of antibiotics
in the non-surgical treatment of peri-
odontitis showed a statistically sig-
nificant additional PD reduction
(0.55  0.37 mm, on average) for
the use of AMX and MET as com-
pared to what was achieved with
SRP alone after 12 months (Keestra
et al. 2015). These results are in line
with data reported in two previously
published meta-analyses: Zandbergen
et al. (2013) obtained a full-mouth
2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd
weighted mean PD improvement of
1.41 mm and a CAL gain of
0.94 mm, whereas Sgolastra et al.
(2012) obtained a PD reduction of
0.43 mm and a CAL gain
of 0.21 mm for the adjunctive use of
AMX and MET versus SRP alone,
respectively.
However, dosage and duration of
the prescribed antibiotic regime vary
greatly in the literature. Several stud-
ies have evaluated the effect of
375 mg AMX and 250/500 mg
MET, three times daily (TID), for
78 days as adjuncts to SRP (van
Winkelhoff et al. 1989, Pavicic et al.
1994, Flemmig et al. 1998, Ehmke
et al. 2005, Cionca et al. 2009, 2010,
Ribeiro Edel et al. 2009); other stud-
ies have used a dosage of 500 mg 3
TID for 7 days (Guerrero et al.
2005, Yek et al. 2010, Griffiths et al.
2011), or 500 mg AMX and 400/
250 mg MET, 3 TID for 1014 days
(Matarazzo et al. 2008, Mestnik
et al. 2010, 2012, Heller et al. 2011,
Silva et al. 2011, Feres et al. 2012,
Rodrigues et al. 2012). In perspec-
tive, irrespective dosage and dura-
tion, it is important that AB should
be taken in a minimum bactericidal
concentration and for a minimum
duration (Vogelman & Craig 1986)
to limit the risk for side effects, espe-
cially the development of microbial
antibiotic resistance. Antibiotic resis-
tance leads to the development of
dangerous resistant strains with an
impact on the success of antimicro-
bial therapy of life-threatening infec-
tions (Lassmann et al. 2007, Fair &
Tor 2014), and its prevalence contin-
ues to increase globally (Roberts
et al. 2006, Snydman et al. 2010,
Nagy et al. 2011, Karlowsky et al.
2012). Resistant strains can appear
de novo during a single long-term
infection in association with

long-term AB intake, especially
when low drug doses are prescribed,
which in turn may lead to poor drug
compliance and/or incomplete pene-
tration of the drug in all tissues
(Lipsitch & Levin 1997, Musher
et al. 2002, Lieberman et al. 2011).
With regard to periodontal therapy,
some studies have shown a decline in
resistance frequencies of oral biofilm
species after AB removal (Seppala
et al. 1997, Barbosa & Levy 2000,
Feres et al. 2002). For example,
Feres et al. (2002) observed that the
percentage of antibiotic-resistant
strains 90 days after antibiotic treat-
ment was similar to that at prior to
AB intake, which indicates that any
possible increase in the percentage of
resistant subgingival species during
AB intake appears to be only tran-
sient (Feres et al. 2002).
Nevertheless, the theoretical pos-
sibility of development of microbial
antibiotic resistance, in general, can-
not be completely excluded. Consid-
ering this, as well as other undesired
effects associated with prolonged AB
regimes e.g. taste disturbance,
headache, severe diarrhoea, nausea/
vomiting, hypersensitivity, renal and
liver toxicity, problems with compli-
ance regarding intake and high costs
and in light of recently advocated
AB regimens as adjunct to SRP over
longer periods (i.e. 2 weeks) (Matar-
azzo et al. 2008, Mestnik et al. 2010,
2012, Heller et al. 2011, Silva et al.
2011, Feres et al. 2012, Rodrigues
et al. 2012), it appears relevant to
attempt optimizing the duration of
the AB intake. Indeed, in many
areas of general medicine, shorter,
high-dosage regimens compared to
older regimens have been success-
fully employed (Green & Rothrock
1993, Gooch et al. 1996, Barnett
et al. 1997, de Boer et al. 1997,
Schrag et al. 2001, Chastre et al.
2003, Dunbar et al. 2003, Milo et al.
2005, Joshi 2011).
At the time being, it remains
unknown whether a short-term (e.g.
3 days) administration of
AMX + MET as adjunct to non-sur-
gical periodontal therapy may pro-
vide additional benefits to scaling
and root planing (SRP) alone. Thus,
the aim of this study was to evaluate
the potential effect of 3- or 7-day
systemic administration of AMX
and MET as adjunct to SRP over
SRP alone during non-surgical
2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd
SRP with a 3 or 7-day antibiotic regimen
periodontal therapy in patients with
severe chronic periodontitis (ChP).
Material and Methods
Trial design, patient population, inclusion
and exclusion criteria
The hypothesis to be tested in this
prospective, randomized, placebo-
controlled, double-masked clinical
trial was that the systemic use of
AMX and MET administered for 3
or 7 days as adjunct to SRP leads to
superior clinical results compared
with SRP alone.
Sample size calculation was per-
formed before starting the study, con-
sidering the possibility to detect a
difference in five sites (Silva et al.
2011, Feres et al. 2012) with a
PD 6 mm (Matuliene et al. 2008)
and a standard deviation of 6 sites
(Cionca et al. 2009, Feres et al. 2012)
between AB and control groups at
6 months, as clinically important. A
study power of 90% at a statistical
significance level of 0.05 was reached
with 30 subjects per treatment group.
Taking into account a possible attri-
tion of 13%, 34 subjects per
treatment group were included.
One hundred and two subjects
were recruited from the patients
seeking dental treatment at the
University Clinic of Cluj-Napoca
(University Iuliu Hatieganu, Cluj-
Napoca, Romania) between January
2012 and January 2014. To be
included in the study, subjects had
to fulfil following criteria:
>30 years of age
12 natural teeth present in the
oral cavity distributed in all four
quadrants
clinical (i.e. 1 site per quadrant
with PD 6 mm) and radio-
graphic signs of generalized severe
chronic periodontitis (Armitage
1999)
full-mouth plaque scores (FMPS)
25% (OLeary et al. 1972)
systemically healthy, i.e. absence
of a known condition that may
influence the severity or progres-
sion of periodontal disease (e.g.
Down Syndrome, HIV, Diabetes
Mellitus types 1 and 2)
no head and neck radiation
therapy
no infectious or heart diseases
that need prophylactic admin-
3
istration of AB before dental
treatment
no liver disease.
Exclusion criteria were as follows:
non-surgical periodontal therapy
within the previous 12 months
systemic or local use of AB
within the previous 3 months
medication that could interact
with AMX or MET (e.g. cou-
marin derivates, containing alco-
hol derivates, 5-fluor-uracyl/
disulfiram derivates, amprenavir
oral solutions, lopinavir/ritonavir
oral solution)
medication with a possible influence
on the periodontium (e.g. Ciclos-
porin A, compounds of Phenytoin,
calcium channel blockers).
pregnancy or lactation.
The study was conducted accord-
ing to the Declaration of Helsinki
(1964, revision 2008) and approved
by the Ethical Committee of the
Faculty of Medicine and Pharmacy
of Cluj-Napoca (Application #514/
09.01.2012). Prior to participation,
all eligible subjects were informed of
the purpose of the study, and the
possible risks and benefits of their
participation. Informed written con-
sent to participate in this study was
obtained from all subjects prior to
inclusion in this study. The study
was registered in the ISRCTN reg-
istry (study ID ISRCTN17605083).
Treatment protocol
This study was a prospective, ran-
domized, placebo-controlled, double-
masked clinical trial.
All subjects enrolled in the study
were allocated according to a com-
puter-generated list to one of the fol-
lowing three treatment groups:
SRP + placebo (for 7 days; control
group A, n = 34), SRP + AMX +
MET (both 500 mg TID 9 3 days
afterwards placebo TID 9 4 days;
group B, n = 34) and SRP + AMX +
MET (both 500 mg TID 9 7 days;
group C, n = 34). Treatment alloca-
tion was performed according to a
block randomization procedure and
the randomization list was concealed
to all examiners, except for the person
in charge for delivering the pills to the
patients (RT).
At baseline (i.e. prior to SRP), at
3 and 6 months after SRP,
4 Cosgarea et al.
assessment of the medical and smok-
ing history, oral hygiene and clinical
periodontal parameters was per-
formed by one calibrated examiner
(RJ), blinded to the treatment group.
Patients were asked about their
smoking habits; those smoking 10
cigarettes per day for the last 5 years
were defined as smokers (Ammen-
heuser et al. 1997). Oral hygiene sta-
tus and gingival inflammation were
assessed by FMPS (OLeary et al.
1972) and full-mouth bleeding scores
[gingival bleeding index; GBI
(Ainamo & Bay 1975)] at four sites
per tooth as percentage values. Prob-
ing PD and vertical CAL were
recorded at six sites per tooth to the
nearest millimetre using a periodon-
tal probe (PCPUNC 15, Hu Friedy,
Chicago, IL, USA). The reference
point for the CAL measurements
was considered the cementenamel
junction (CEJ); if the CEJ had been
modified by restorative treatment,
then the reference point was consid-
ered the most apically located mar-
gin of the restoration. Bleeding on
probing (BOP) was assessed as the
percentage of sites showing bleeding
30 s after probing.
Oral hygiene instruction sessions,
including professional prophylaxis,
were performed until each patient
had a FMPS 25%. Once the
desired FMPS was reached, SRP was
performed within 24 h by one experi-
enced periodontist (RC) blinded to
the treatment allocation. In particu-
lar, all pockets with PD 4 mm were
scaled and root planed to the bottom
of the pocket with ultrasonic instru-
ments (Kavo Sonicflex Scaler; Kavo
Dental GmbH, Biberach, Germany)
and Gracey curets (Hu Friedy, Chi-
cago, IL, USA), under local anaesthe-
sia; subsequently, the pockets were
thoroughly rinsed with 0.2%
chlorhexidine digluconate solution
(Corsodyl, GlaxoSmithKline, Brent-
ford, London, UK). The patients
were then instructed to rinse twice
daily for 2 min. with a 0.2%
chlorhexidine digluconate solution
(Corsodyl, GlaxoSmithKline), and
to brush their teeth with 0.2%
chlorhexidine digluconate tooth paste
(Elugel, Pierre Fabre, Paris, France)
for 14 days. At the end of the SRP
sessions, one clinician (RT) allocated
the patients to one of the three treat-
ment groups according to the ran-
domization protocol, gave their
medications and instructed them on
how to take them. Each patient
received four identical bottles: bottles
No. 1 and 2 were with pills for the
first 3 days after SRP; bottles No. 3
and 4 were distributed for the follow-
ing 4 days. All subjects had to take
one pill from each bottle every 8 h.
The placebo group (group A) had
only placebo pills in all bottles;
Group B had AMX and MET in bot-
tles No. 1 and 2, and placebo pills in
bottles No. 3 and 4. Group C had
AMX and MET in all four bottles.
All bottles were prepared by the phar-
macy of the University of Medicine
and Pharmacy Iuliu Hatieganu
Cluj-Napoca. All capsules were taste-
less with identical appearance.
Patients were recalled at 2 weeks,
3 months and 6 months after comple-
tion of SRP. At these appointments,
information about any adverse
events, compliance regarding intake
of the study medication and intake of
any additional concomitant medica-
tion was collected by asking the
patients. Furthermore, at 3 and
6 months, GBI, FMPS, periodontal
clinical parameters and BOP were
recorded by one experienced clinician
(RJ), who was blinded to treatment
allocation. Finally, at the 3- and 6-
month appointments supragingival
calculus was removed, but residual
periodontal pockets, i.e. pockets with
PD = 4 mm + BOP, or PD 5 mm,
were not re-instrumented.
Intra-examiner reproducibility
Ten patients, each with 10 teeth (sin-
gle and multi-rooted) with
PD 6 mm on at least one aspect
of each tooth, were used to calibrate
the examiner. The examiner evalu-
ated the patients on two separate
occasions 48 h apart, prior to com-
mencing the study. Calibration was
accepted if the two sets of measure-
ments of PD and CAL did not differ
1 mm in >90% of the cases. The
mean intra-examiner reliability was
0.87 for PD and 0.76 for CAL
(Cohens Kappa Analyses).
Statistical analyses
All sampled data were introduced
into a database by one person (LI),
and checked for errors by compar-
ison with the original patients charts
by two additional independent
2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd
persons (LL, EB). The data were
then transferred into a statistical
software program (SPSS statistics
21, IBM, Armonk, NY, USA), and
an experienced professional statisti-
cian (CH) performed all analyses.
The statistical unit was the patient,
and all sites with PD 4 mm at
baseline were considered for the sta-
tistical analyses. The primary out-
come variable was the difference (D)
in the number of sites per patient
with PD 6 mm between baseline
and 6 months. Secondary variables
were average changes in BOP, GBI,
FMPS, PD, CAL, number of sites
with PD 6 mm, number of sites
with PD 5 mm, the total number
of sites with PD = 4 mm and BOP+
or with PD 5 mm and their D. p
values < 0.05 were considered to be
statistically significant.
Comparisons within the groups
between the various time points were
performed using Paired t-test and
Wilcoxon Signed Ranks Test.
Adjusting for baseline values and
smoking, comparisons among the
groups at the various time points
were performed by means of ANCOVA
using the Bonferroni adjustment.
By mean of a Poisson regression
analysis, the relationship between the
number of sites with PD 6 mm at
6 months and the following vari-
ables: AB for 3 days, AB for 7 days,
smoking, gender, baseline BOP,
FMPS, GBI, baseline mean PD,
baseline mean CAL, severe disease
(i.e. >25 sites with PD 6 mm at
baseline) was determined.
Results
Patients
Mean age of the 102 included patients
(65 females, 64%) was
43.37  9.85 years, and 35 (34%)
were smokers. Non-surgical peri-
odontal therapy was performed
between April 2012 and April 2014.
Eleven subjects were lost during fol-
low-up due to the following reasons:
additional AB intake for other medi-
cal reasons (n = 4), moved to another
town (n = 2) and non-compliance
with the appointments (n = 5). Thus,
per protocol analyses included only
the 91 patients [mean age
43.05  9.84 years, 57 females
(62%), 33 smokers (36%)] who com-
pleted the study (Figure 1, Table 1).
 SRP with a 3 or 7-day antibiotic regimen 5

Clinical findings

The clinical parameters registered at

baseline, 3 months and 6 months are
presented in Table 2. No statistical
differences were observed among the
groups at baseline for any of the
evaluated parameters (p > 0.05). At
3 and 6 months, all investigated
parameters demonstrated statistically
highly significant improvements
compared to baseline in all treatment
groups (p < 0.001, paired t-test and
Wilcoxon Signed Ranks Test). At
both 3 and 6 months, statistically
significantly greater improvements
(p < 0.05) in the two antibiotic
groups compared to the placebo
group were observed regarding sev-
eral of the evaluated parameters
(Table 2).
In particular, at 6 months, aver-
age PD reduction in the placebo
group (1.90  0.69 mm) was signifi-
cantly smaller compared to the 3-
day (2.54  0.62 mm) or the 7-day
AB regime (2.76  0.65 mm)
groups. Significantly larger CAL
gains were also recorded in groups B
and C (1.68  0.55 mm and
1.77  0.53 mm, respectively) com-
pared to the placebo group
(1.13  0.74 mm) at 6 months
Fig 1. Flowchart of the study. PCR, plaque control record; SRP, scaling and root (Table 2). When looking separately
planning. at moderately deep (46 mm) and
deep (7 mm) sites, statistically sig-
There was no statistical significant Adverse events
nificantly greater PD reductions
difference between the groups regard- Adverse effects including headaches, (p < 0.05) were obtained both at 3
ing gender, smoking status (Table 1) musculoskeletal pain, taste disorders and 6 months in the two AB groups
and baseline clinical parameters (e.g. metallic taste), vertigo, gastroin- as compared with the placebo group
(Table 2). Patient compliance with testinal disorders (i.e. nausea, diar- (Table 2). Also, statistically signifi-
AB intake was excellent; all patients rhoea), fever and shivering were cantly higher CAL gains (p < 0.05)
reported taking all the given pills, and reported by one or two patients in were obtained in the two AB groups
the only deviation from schedule was each treatment group, with no appar- compared with the placebo group at
that a few patients took occasionally ent differences among groups 6 months (Table 2).
the pills with a few hours difference (Table S1). None of the subjects pre- The total number of sites with
from the advised 8 h interval. sented allergies or candida infections. PD 6 mm in group B, decreased
from 921 sites at baseline, to 76 and
Table 1. Demographic characteristics 52 sites at 3 and 6 months, respec-
tively; in group C, it was reduced
Variables Group A Group B Group C
from 1100 sites at baseline, to 176
Gender (Female/male %) 21/12 22/11 20/14 and 156 sites at 3 and 6 months,
Age (years) 42.5  9.64 42.3  9.73 44.3  10.3 respectively. In the placebo group,
Smoker (%) 11 11 13 the decrease in the total number of
GBI (%) sites with PD 6 mm from baseline
Baseline 7.92  7.79 8.03  11.04 13.50  18.82 (767 sites) to 3 (211 sites) and
3 months 5.37  8.35 3.08  4.09 4.05  5.95 6 months (254 sites) was smaller com-
6 months 4.12  5.59 2.03  2.45 3.55  5.11
pared to the two AB groups. On aver-
FMPS (%)
Baseline 19.86  6.11 17.29  6.04 19.51  5.72 age, 28.62  15.32 less sites per
3 months 33.13  22.46 32.04  20.07 37.06  17.85 patient in group B and 30.45  15.04
6 months 37.28  17.92 31.49  19.24 28.75  19.59 less sites per patient in group C, com-
pared to baseline, presented at
GBI, gingival bleeding index (Ainamo & Bay 1975); FMPS, full-mouth plaque score after 6 months a PD 6 mm. The
OLeary et al. (1972).

2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd
6 Cosgarea et al.

Table 2. Mean values and group comparisons (ANCOVA adjusted for baseline values and smoking) of PD, CAL, BoP, GBI, FMPS, number
(No.) of sites with PD 6 mm, with PD 5 mm, with PD = 4 BOP+ and PD 5 and their differences (D) between the follow-ups and
baseline (mean  SD)
Variables Group A Group B Group C Group comparisons Smoker
Non-smoker
(SRP + Placebo) (SRP + AB 3 days) (SRP + AB 7 days) p value A-B p value A-C p value
N = 30 N = 30 N = 31

PD (mm)
Baseline 5.45  0.61 5.46  0.50 5.79  0.58
3 months 3.47  0.42 3.03  0.39 3.16  0.62 0.001* 0.007* 0.10
6 months 3.52  0.48 2.90  0.45 2.97  0.45 <0.001* <0.001* 0.04*
baseline3 month 1.97  0.58 2.43  0.53 2.63  0.80
baseline-6 month 1.90  0.69 2.54  0.62 2.76  0.65
CAL (mm)
Baseline 5.92  1.24 5.80  1.16 6.14  1.18
3 months 4.68  1.19 4.40  1.15 4.54  1.29 0.403 0.089 0.151
6 months 4.76  1.22 4.12  1.16 4.29  1.14 0.001* <0.001* 0.055
baseline3 months 1.24  0.64 1.44  0.51 1.60  0.68
baseline6 months 1.13  0.74 1.68  0.55 1.77  0.53
PD 46 mm (mm)
Baseline 4.88  0.22 4.90  0.25 4.94  0.21
3 months 3.20  0.37 2.83  0.36 2.87  0.48 0.001* 0.004* 0.021*
6 months 3.26  0.46 2.72  0.39 2.70  0.37 <0.001* <0.001* 0.044*
baseline3 months 1.68  0.44 2.07  0.40 2.07  0.55
baseline6 months 1.62  0.53 2.17  0.46 2.22  0.39
CAL of PD 46 mm (mm)
Baseline 5.48  1.09 5.32  1.14 5.36  0.98
3 months 4.42  1.14* 4.10  1.16* 4.16  1.17* 0.66 0.82 0.43
6 months 4.53  1.19* 3.87  1.14* 3.96  1.07* 0.004* 0.007* 0.287
baseline3 months 1.06  0.66 1.21  0.43 1.20  0.56
baseline6 months 0.94  0.72 1.40  0.47 1.36  0.44
PD >7 mm (mm)
Baseline 7.73  0.58 7.63  0.62 7.97  0.92
3 months 4.77  1.10 3.73  0.76 4.06  1.08 <0.001* 0.015* 0.079
6 months 4.94  1.14 3.64  0.91 3.85  0.92 <0.001* <0.001* 0.003*
baseline3 months 2.86  1.24 3.89  0.89 3.91  1.44
baseline6 months 2.66  1.31 3.96  0.98 4.08  1.36
CAL of PD >7 mm (mm)
Baseline 7.72  1.01 7.97  1.49 8.06  1.48
3 months 5.92  1.18 5.56  1.53 5.74  1.74 0.105 0.27 0.204
6 months 6.00  1.17 5.15  1.64 5.46  1.54 0.001* 0.01* 0.052
baseline3 months 1.79  1.20 2.41  1.11 2.31  0.99
baseline6 months 1.67  1.30 2.82  1.16 2.52  0.94
BoP (%)
Baseline 64.93  24.66 64.06  34.33 71.27  26.55
3 months 16.05  10.67 11.62  9.10 9.92  7.59 0.19 0.018* 0.99
6 months 13.07  9.02 9.47  5.34 7.75  6.64 0.23 0.013* 0.82
GBI (%)
Baseline 7.92  7.79 8.03  11.04 13.50  18.82
3 months 5.37  8.35 3.08  4.09 4.05  5.95 0.56 1.00 0.57
6 months 4.12  5.59 2.03  2.45 3.55  5.11 0.40 1.00 0.022*
FMPS (%)
Baseline 19.86  6.11 17.29  6.04 19.51  5.72
3 months 33.13  22.46 32.04  20.07 37.06  17.85 1.00 1.00 0.76
6 months 37.28  17.92 31.49  19.24 28.75  19.59 0.95 0.33 0.078
No. PD 6
Baseline 25.32  16.86 30.70  14.93 36.89  16.91
3 months 7.54  5.62 2.53  3.59 6.29  6.72 <0.001* 0.148 0.282
6 months 8.47  6.14 1.93  3.28 5.03  5.77 <0.001* <0.001* 0.023*
baseline3 months 17.78  14.73 28.16  14.41 30.60  16.10
baseline6 months 17.10  14.68 28.62  15.32 30.45  15.04
No. PD 5
Baseline 41.71  19.62 48.30  19.85 54.43  19.80
3 months 11.54  7.13 6.40  6.75 10.54  9.41 0.004* 0.367 0.289
6 months 12.60  8.40 5.41  6.45 8.71  8.52 <0.001* 0.004* 0.046*
baseline3 months 30.17  16.22 41.90  18.73 43.89  19.31
baseline6 months 29.76  16.66 42.55  20.31 44.25  16.56

2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd
 SRP with a 3 or 7-day antibiotic regimen 7

Table 2. (continued)
Variables Group A Group B Group C Group comparisons Smoker
Non-smoker
(SRP + Placebo) (SRP + AB 3 days) (SRP + AB 7 days) p value A-B p value A-C p value
N = 30 N = 30 N = 31

No. PD = 4 BOP+ and PD 5

Baseline 54.79  23.45 62.83  22.31 68.75  23.06
3 months 14.21  8.14 8.07  7.57 12.07  9.85 0.002* 0.159 0.335
6 months 14.63  9.47 6.67  7.08 10.03  9.60 <0.001* 0.004* 0.038*
baseline3 months 40.57  19.67 54.76  21.61 56.67  21.98
baseline6 months 41.76  19.05 55.77  23.48 57.29  19.79

PD, pocket depth; CAL, clinical attachment level; BOP, bleeding on probing; GBI, gingival bleeding index (Ainamo & Bay 1975); FMPS,
full-mouth plaque score after OLeary et al. (1972).
*Statistical significant p values.

Table 3. Poisson regression analyses for the number of residual sites with PD 6 mm at difference in the number of pockets
6 months after non-surgical periodontal therapy as related to placebo group (group A) with PD 6 mm between baseline
and 6 months after treatment. This
Variables Exp. 95% CI p value
coefficient
is a relevant outcome variable after
periodontal therapy, as it has been
Group B (AB for 3 days) 0.24 0.170.32 <0.001* previously observed that patients
Group C (AB for 7 days) 0.33 0.250.43 <0.001* with a large number of residual
Female gender 0.75 0.590.93 0.010* pockets 6 mm show a statistically
Smoker 1.35 1.071.69 0.010* significantly higher risk for further
25 sites with PD 6 mm 1.26 0.951.65 0.103 attachment loss, and thus require
at baseline
further treatment usually surgical
BOP baseline 1.00 0.991.01 0.16
GBI baseline 0.98 0.970.99 0.027*
therapy (Matuliene et al. 2008,
FMPS baseline 0.99 0.991.00 0.66 2010). Likewise, several studies have
Mean PD baseline 1.42 1.141.79 0.002* previously investigated the effect of
Mean CAL baseline 1.30 0.911.15 0.60 AB as adjunct to non-surgical peri-
odontal treatment in various treat-
PD, pocket depth; CAL, clinical attachment level; BOP, bleeding on probing; GBI, gingival ment protocols by using as endpoint
bleeding index (Ainamo & Bay 1975); FMPS, full-mouth plaque score after OLeary et al.
the number of sites of specific depths
(1972).
*Statistically significant p values. Reference categories are as follows: Group A, Male, Non- (Cionca et al. 2009, Feres et al.
Smoker (Exp. Coefficient equals). 2012, Arweiler et al. 2014). Other
studies have chosen mean full-mouth
PD or CAL as endpoint (Winkel
reduction in the number of sites with smoking, baseline GBI and baseline et al. 2001, Ehmke et al. 2005,
PD 6 mm from baseline to mean PD influenced statistically sig- Matarazzo et al. 2008, Silva et al.
6 months in group A (17.10  14.68 nificant the residual number of sites 2011, Feres et al. 2012) for evaluat-
sites) was statistically significantly less with PD 6 mm: groups B and C ing the efficacy of various antibiotic
compared to that in the two AB showed 4.19 and 3.00 times less sites regimens. However, using mean val-
groups. with PD 6 mm, respectively, com- ues instead of frequencies is prob-
The numbers of sites with pared to group A (Table 3). Smok- lematic; shallow sites showing
PD 6 mm, PD 5 mm and the ers had 1.35 times more chances to limited improvements even after
total number of sites with show sites with PD 6 mm at treatment compared to deeper sites
PD = 4 mm and BOP+ or with 6 months after treatment. Female in terms of numerical values (Winkel
PD 5 mm in the placebo group patients had 0.74 times less chances et al. 2001, Ehmke et al. 2005,
(group A) were statistically signifi- to show sites with PD 6 mm at Cionca et al. 2009, Silva et al. 2011,
cantly higher as compared to the 3- 6 months after treatment (Table 3). Feres et al. 2012) are likely to signifi-
day AB group (group B) at both cantly dilute the effect observed in
recall time points; when compared to deeper sites, which are actually in
Discussion
the 7-day AB group (group C), need for further treatment.
group A exhibited statistically signif- This study has evaluated the clinical The results of this study showed
icantly higher number of residual outcomes following non-surgical that both, a 3- or 7-day antibiotic
pockets with PD 6 mm, with periodontal therapy in conjunction regimen as adjunct to SRP exhibited
PD 5 mm and a total number of with adjunctive AMX and MET superior clinical results than those
sites with PD = 4 mm and BOP+ or administered systemically for 3 or obtained by SRP alone. Specifically,
with PD 5 mm at 6 months for 7 days compared with SRP alone the use of AB as adjunct to SRP
(Table 2). in patients with generalized severe resulted in statistically significantly
The regression analysis showed chronic periodontitis. The main out- higher clinical improvements than
that, at 6 months, AB, gender, come variable in this study was the those obtained in the placebo group
2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd
8 Cosgarea et al.
regarding the number of sites with
PD 6 mm, the number of sites
with PD 5 mm and their differ-
ences before and after treatment (),
the total number of sites with
PD = 4 and BOP+ and mean PD
reduction and CAL gain (Tables 2
and 3).
Comparable improvements after
SRP combined with systemic
AMX + MET have been reported by
Feres et al. (Feres et al. 2012); for
example, at 1 year after non-surgical
treatment 6.9  10 sites with
PD 6 mm in the placebo group
versus 1.2  2.2 sites in the
AMX + MET group were obtained
(Feres et al. 2012). However, the
antibiotic regimen in that study was
substantially different from herein,
i.e. MET (400 mg TID) and AMX
(500 mg TID) for 14 days. Interest-
ingly, their results after 1 year are
comparable with the results obtained
after 6 months in the 3 or 7 days
AB regimens of this study. In
another study evaluating the out-
come of SRP and AMX + MET, a
lower number of sites with
PD 5 mm was obtained after
6 months comparing to that
observed herein (Cionca et al. 2009),
i.e. 3.0  4.3 sites in the placebo
group and 0.4  0.8 in the AB
group. The lower number of deep
sites observed in Cionca et al.
(Cionca et al. 2009) compared to
that obtained herein, may well be
due to the fact that only patients
with moderate-to-severe ChP (i.e.
presenting at least four teeth with
PD > 4 mm and CAL 2 mm) were
included in the former study,
whereas only generalized severe ChP
patients with at least 1 tooth per
quadrant with PD 6 mm were
included in this study. Indeed, in
our study double as much number
of sites with PD 5 mm were pre-
sent in each treatment group com-
pared to those in the study by
Cionca et al. (Cionca et al. 2009).
Similarly, these differences regarding
the severity of baseline clinical peri-
odontal conditions of the patients
included in the above-mentioned
studies and those included herein
may also explain the larger average
clinical improvements (PD reduction
and CAL gain) observed herein in
comparison to what observed Cionca
et al. and Feres et al. (Cionca et al.
2009, Feres et al. 2012). Indeed,
several studies have shown that deep
pockets profit more by AB adminis-
tration as compared to shallow sites
(Winkel et al. 2001, Ehmke et al.
2005, Cionca et al. 2009, Silva et al.
2011, Feres et al. 2012). In general,
the average additional PD reduction
obtained from the adjunct use of AB
in the moderately deep pockets in
this study (ca. 0.5 mm) is compara-
ble to that reported in the recent
meta-analysis of Keestra et al.
(2015) mentioned earlier. However,
when interpreting the results of this
study, it is important to point out
that due to its design (i.e. the study
was set up as a superiority trial of
the two AB regimes over SRP alone)
direct comparisons between the two
AB regimes are not appropriate as
such comparisons would be under-
powered and might lead to erro-
neous conclusions.
Furthermore, by means of a Pois-
son regression model, AB, female
gender, smoking, initial mean PD
and initial mean GBI were shown to
statistically influence the number of
residual pockets with PD 6 mm
after treatment. The finding that AB
had a major positive influence on the
clinical outcomes is in agreement with
the results of the logistic regression
analyses performed in the studies by
Cionca et al. (2009) and Feres et al.
(2012) where AB was the only vari-
able that significantly influenced the
primary outcome variable. In con-
trast to Cionca et al., where smoking
had no statistically significant influ-
ence, the results of this study indicate
that smokers had a 0.74 (1/1.35)
higher number of residual sites with
PD 6 mm compared to non-smo-
kers, irrespective of treatment group.
This is also recognizable when look-
ing at the smoking-adjusted analyses
for the number of sites with
PD 6 mm, PD 5 mm and
PD 4 mm and BOP+, where smok-
ers had significantly more (p < 0.05)
of these specific sites at 6 months as
compared to non-smokers (Table 2).
This is in agreement with previous
findings, which have shown that
smoking is strongly associated with
poorer clinical and microbiological
responses after conventional or
regenerative periodontal therapy
(Winkel et al. 2001, Van der Velden
et al. 2003, Hughes et al. 2006).
In perspective, development of
microbial strains resistant to AB is
2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd
an increasing problem globally. It
appears thus relevant to attempt
optimizing AB intake if this should
be considered as standard adjunct to
periodontal therapy; of course it is
important that the AB is given in an
adequate (minimum) bactericidal
dose for an adequate (minimum)
length of time (Vogelman & Craig
1986). Various dosages of AB have
previously been employed in the var-
ious studies evaluating the effect of
SRP in combination with AB, but
the optimal dose (single drug or
combination of AB) for the treat-
ment of periodontitis has not yet
been clearly defined. For example,
many studies evaluating AMX as
adjunct to SRP have used dose
ranges 375500 mg TID for 7
14 days (van Winkelhoff et al. 1989,
Guerrero et al. 2005, Matarazzo
et al. 2008, Cionca et al. 2009, Yek
et al. 2010, Griffiths et al. 2011,
Silva et al. 2011, Feres et al. 2012).
Studies evaluating MET have used
ranges 250400 mg TID (Lindhe
et al. 1983, van Winkelhoff et al.
1989, Loesche et al. 1992, Winkel
et al. 2001, Ehmke et al. 2005,
Matarazzo et al. 2008, Silva et al.
2011). More important is that the
antibiotic should be given in an ade-
quate bactericidal dose for an ade-
quate length of time. MET reaches
an effective serum concentration in
an adult for a concentration of 20
25 mg/kg body weight (van Winkel-
hoff et al. 1999). Therefore, for a
70 kg patient, the daily needed MET
dose would be 14001750 mg (Win-
kel et al. 1997, van Winkelhoff et al.
1999). A more recent study (Cionca
et al. 2009) used a MET dosage of
500 mg TID for 7 days in combina-
tion with AMX 375 mg TID for
7 days, showing significant clinical
and microbiological results for the
AB group (Cionca et al. 2009, 2010).
In this study the AB protocol con-
sisted of MET 500 mg TID and
AMX 500 mg TID, both for 3 and
7 days. Thus, the AB regimen used
herein falls within the effective daily
dose for the average adult patient.
Regarding the aspect of AB
administration duration, several stud-
ies have shown that resistant micro-
bial strains can appear de novo when
low-drug doses are prescribed for
long-term infections (Lipsitch &
Levin 1997, Musher et al. 2002,
Lieberman et al. 2011). Therefore, in

several areas of general medicine AB
administration in high dosage and for
shorter intervals has been evaluated
for various infections (Green &
Rothrock 1993, Gooch et al. 1996,
Barnett et al. 1997, de Boer et al.
1997, Schrag et al. 2001, Chastre
et al. 2003, Dunbar et al. 2003, Milo
et al. 2005, Joshi 2011). For example,
in two recent reviews on short-term
AB administration in acute strepto-
coccal pharyngitis in children (Alta-
mimi et al. 2009, 2012), it was
concluded that a 3-day oral AB regi-
men yielded comparable clinical and
microbiological efficacy to that of
standard treatment of 10 days. Fur-
thermore, the short duration treat-
ment had shorter periods of clinical
symptoms (fever, throat soreness)
and a lower risk of early clinical treat-
ment failure. Similar findings favour-
ing shorter AB administration have
been reported for acute urinary tract
infections (Michael et al. 2003), intra-
abdominal infections (Sawyer et al.
2015) and infectious episodes in gen-
eral surgery units (Hedrick et al.
2006). Further data that are in sup-
port for our decision to evaluate the
3- and the 7 day AB therapy are those
obtained by Feres et al. (2002): the
transient increase in resistant bacte-
rial species was at 3 days of AMX
and MET administration lesser than
that obtained for the 7 days regimen.
Thus, a 3-day AB regimen appeared
as a logical duration to be also evalu-
ated, but neither final conclusions on
the clinical relevance of the 3-day reg-
imen nor one on the possibility that
similar results might have been
obtained with even shorter AB
administration can be made.
In this study, a trend for better
clinical outcomes after 7 days of AB
was observed. Nevertheless, a direct
comparison regarding the clinical
findings between the two AB regimes
would not be appropriate, as any
comparisons would be underpowered
and might lead to erroneous conclu-
sions. A valid comparison between
the two AB groups would require a
non-inferiority trial design and not
superiority trial of the two AB
regimes over SRP alone, as herein.
This study was sized with 30 subjects
per treatment group, based on
already published data and consider-
ing as important the possibility to be
able to disclose a difference in five
sites with PD 6 mm (Matuliene
2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd
SRP with a 3 or 7-day antibiotic regimen
et al. 2008, Cionca et al. 2009, Silva
et al. 2011, Feres et al. 2012) and a
standard deviation of 6 (Cionca et al.
2009, Feres et al. 2012) between the
treatment groups at 6 months, with a
study power set at 90% and a signifi-
cance level of 0.05. When considering
the results obtained herein (i.e. differ-
ence in the number of sites with
PD 6 mm between baseline and
6 months, a standard deviation of
approximately 15, 30 subjects per
group and a desired power of 90%
with a statistical significance level at
0.05), the minimum detectable differ-
ence between the two AB groups is
approximately 11 sites. If a difference
in five sites with an assumed SD of 15
should have been detected with
80% power and the significance level
set at 0.05142 patients would have
been needed in each group; to detect
a difference of two sites, 884 patients
would have been needed per group.
Such high numbers are unfortunately
almost impossible to achieve under
usual settings. However, post hoc
power calculation revealed a power
for the overall F-Test of 92% power,
whereas when adjusting for pair-wise
comparisons related to the placebo
group, the power was 85% (Tukey
Test) and 84% (Bonferroni Test). In
context, to determine the equivalence
or non-inferiority of the 3-day
adjunctive AMX and MET therapy
over the 7-day AB administration,
further studies with the proper design
are necessary.
In conclusion, these findings have
shown that in patients with severe
chronic periodontitis SRP in con-
junction with a 3- or 7-day systemic
administration of AMX and MET
may lead to significantly greater clin-
ical improvements compared to SRP
alone.
Acknowledgements
The authors would like to thank the
study nurses Livia Indolean and
Erika Batiz for data entries and
error checking.
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11
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Supporting Information
Additional Supporting Information
may be found in the online version
of this article:
Table S1. Adverse events.
Address:
Raluca Cosgarea
Department of Periodontology
University of Marburg
Georg-Voigt Str. 3, 35039 Marburg
Germany
E-mail: ralucacosgarea@gmail.com
Practical implications: In patients
with severe chronic periodontitis,
non-surgical periodontal therapy in
conjunction with a 3- or 7-day
antibiotic protocol with
AMX + MET may yield greater
clinical improvements than non-
surgical therapy alone.