ORIGINAL ARTICLE
Is it possible to prevent recurrent vulvovaginitis? The role
of Lactobacillus plantarum I1001 (CECT7504)
S. Palacios1 & J. Espadaler2 & J. M. Fernández-Moya3 & C. Prieto4 & N. Salas1
Received: 16 March 2016 /Accepted: 20 June 2016
# The Author(s) 2016. This article is published with open access at Springerlink.com
Abstract The purpose of this study was to prospectively
evaluate the impact of the use of L. plantarum I1001 applied
vaginally on Vulvovaginal Candidiasis (VVC) time-until-
recurrence after treatment with single-dose vaginal clotrima-
zole. This was a clinical open-label, prospective study of two
non-randomized parallel cohorts with symptomatic acute
VVC: (1) 33 sexually active women 18–50 years old, pre-
scribed a standard single-dose 500 mg vaginal tablet of clotri-
mazole followed by vaginal tablets with L. plantarum I1001
as adjuvant therapy, and (2) 22 women of similar characteris-
tics but prescribed single-dose clotrimazole only. Use of the
probiotic and factors that might influence recurrence risk (age,
recurrent VVC within previous year, antibiotic prior to study
enrolment, diaphragm or IUD contraception, among others)
were included in a multivariate Cox regression model to adjust
for potential between-cohort differences. Probiotic use was
associated with a three-fold reduction in the adjusted risk of
recurrence (HR [95 %CI]: 0.30 [0.10–0.91]; P = 0.033).
Adjusted free-survival recurrence was 72.83 % and 34.88 %
for the probiotic and control groups, respectively. A higher
cumulative recurrence was also observed in cases with use
of antibiotics prior to enrolment (HR [95 %CI]: 10.46 [2.18–
50.12]; P = 0.003). Similar findings were found at six months
after azole treatment in women with RVVC. Overall, good
compliance with the probiotic was reported for 91.3 % of
women. The study suggests that follow-up therapy with vag-
inal tablets with L. plantarum I1001 could increase the effec-
tiveness of single-dose 500 mg clotrimazole at preventing
recurrence of VVC, an effect that was also observed in women
with recurrent vulvovaginal candidiasis (RVVC) after six
months of azole treatment.
Introduction
Vulvovaginal candidiasis (VVC) is one of the most frequent
infections of the female genital tract and is mainly associated
with Candida albicans (followed by Candida glabrata) [1].
Candida species are normally found in the lower genital tract
of 10–20 % of healthy women of childbearing age [2].
Evolution from colonization to symptomatic infection involves
different host factors such as susceptibility and inflammatory
responses and/or the imbalance of vaginal microbiota [3].
Available literature on VVC suggests that 75 % of women
will show a VVC episode [4, 5] through their lifetime, and that
5–10 % of all women will experience recurrent vulvovaginal
candidiasis (RVVC), i.e ≥4 episodes/year [6].
Short-term and single-dose vaginal antifungals are success-
ful in 80–90 % of acute uncomplicated cases [7, 8]. However,
azole resistance rates are above 15 % in women with RVVC
[9, 10]. Antifungal resistance, infection recurrence and side
effects of pharmacological treatments are key issues for
patients and their physicians seeking alternative interventions
for VVC.
Electronic supplementary material The online version of this article
(doi:10.1007/s10096-016-2715-8) contains supplementary material,
which is available to authorized users.
* C. Prieto
cprieto@gynea.com
1
Instituto Palacios de Salud y Medicina de la Mujer, Antonio Acuña
9CP, 28009 Madrid, Spain
2
Autonomous University of Barcelona, AB-Biotics S.A. Eureka
building, 08193 Bellaterra, Barcelona, Spain
3
Instituto de Medicina EGR, Camino de la Zarzuela, 19,
28023 Aravaca, Madrid, Spain
4
Gynea Laboratorios by Kern Pharma, Pol. Ind. Colom II, C/Venus,
72, 08228 Terrasa, Barcelona, Spain
Eur J Clin Microbiol Infect Dis
DOI 10.1007/s10096-016-2715-8. 

Scene 1

ORIGINAL ARTICLE
Is it possible to prevent recurrent vulvovaginitis? The role
of Lactobacillus plantarum I1001 (CECT7504)
S. Palacios1 & J. Espadaler2 & J. M. Fernández-Moya3 & C. Prieto4 & N. Salas1
Received: 16 March 2016 /Accepted: 20 June 2016
# The Author(s) 2016. This article is published with open access at Springerlink.com
Abstract The purpose of this study was to prospectively
evaluate the impact of the use of L. plantarum I1001 applied
vaginally on Vulvovaginal Candidiasis (VVC) time-until-
recurrence after treatment with single-dose vaginal clotrima-
zole. This was a clinical open-label, prospective study of two
non-randomized parallel cohorts with symptomatic acute
VVC: (1) 33 sexually active women 18–50 years old, pre-
scribed a standard single-dose 500 mg vaginal tablet of clotri-
mazole followed by vaginal tablets with L. plantarum I1001
as adjuvant therapy, and (2) 22 women of similar characteris-
tics but prescribed single-dose clotrimazole only. Use of the
probiotic and factors that might influence recurrence risk (age,
recurrent VVC within previous year, antibiotic prior to study
enrolment, diaphragm or IUD contraception, among others)
were included in a multivariate Cox regression model to adjust
for potential between-cohort differences. Probiotic use was
associated with a three-fold reduction in the adjusted risk of
recurrence (HR [95 %CI]: 0.30 [0.10–0.91]; P = 0.033).
Adjusted free-survival recurrence was 72.83 % and 34.88 %
for the probiotic and control groups, respectively. A higher
cumulative recurrence was also observed in cases with use
of antibiotics prior to enrolment (HR [95 %CI]: 10.46 [2.18–
50.12]; P = 0.003). Similar findings were found at six months
after azole treatment in women with RVVC. Overall, good
compliance with the probiotic was reported for 91.3 % of
women. The study suggests that follow-up therapy with vag-
inal tablets with L. plantarum I1001 could increase the effec-
tiveness of single-dose 500 mg clotrimazole at preventing
recurrence of VVC, an effect that was also observed in women
with recurrent vulvovaginal candidiasis (RVVC) after six
months of azole treatment.
Introduction
Vulvovaginal candidiasis (VVC) is one of the most frequent
infections of the female genital tract and is mainly associated
with Candida albicans (followed by Candida glabrata) [1].
Candida species are normally found in the lower genital tract
of 10–20 % of healthy women of childbearing age [2].
Evolution from colonization to symptomatic infection involves
different host factors such as susceptibility and inflammatory
responses and/or the imbalance of vaginal microbiota [3].
Available literature on VVC suggests that 75 % of women
will show a VVC episode [4, 5] through their lifetime, and that
5–10 % of all women will experience recurrent vulvovaginal
candidiasis (RVVC), i.e ≥4 episodes/year [6].
Short-term and single-dose vaginal antifungals are success-
ful in 80–90 % of acute uncomplicated cases [7, 8]. However,
azole resistance rates are above 15 % in women with RVVC
[9, 10]. Antifungal resistance, infection recurrence and side
effects of pharmacological treatments are key issues for
patients and their physicians seeking alternative interventions
for VVC.
Electronic supplementary material The online version of this article
(doi:10.1007/s10096-016-2715-8) contains supplementary material,
which is available to authorized users.
* C. Prieto
cprieto@gynea.com
1
Instituto Palacios de Salud y Medicina de la Mujer, Antonio Acuña
9CP, 28009 Madrid, Spain
2
Autonomous University of Barcelona, AB-Biotics S.A. Eureka
building, 08193 Bellaterra, Barcelona, Spain
3
Instituto de Medicina EGR, Camino de la Zarzuela, 19,
28023 Aravaca, Madrid, Spain
4
Gynea Laboratorios by Kern Pharma, Pol. Ind. Colom II, C/Venus,
72, 08228 Terrasa, Barcelona, Spain
Eur J Clin Microbiol Infect Dis
DOI 10.1007/s10096-016-2715-8

Probiotics are defined as Blive microorganisms, which
when administered in adequate amounts, confer a health ben-
efit on the host^ [11]. Probiotic properties are strain-specific
so their positive health effects cannot be extrapolated to other
strains of the same species or genus [12]. Lactobacilli are the
predominant bacteria in a healthy vaginal ecosystem, and have
been proposed for the treatment and prevention of genitouri-
nary infections including candidiasis [4, 6]. Local use of these
microorganisms results in immunomodulation responses and
the restoration of vaginal microbiota, interfering with the col-
onization and growth of potential pathogens such as Candida
[13]. A Lactobacilli-dominated vaginal microbiota produces
significant levels of lactic acid with strong microbicidal prop-
erties [14].
Evidence of synergy between adjuvant local Lactobacilli
and azole treatment for VVC has been reported [9, 15].
Nevertheless, evidence of the benefit of adding probiotics to
the standard azole on the risk of symptomatic recurrence is
lacking.
Recently, a pilot study for the evaluation of colonization
and tolerability of a vaginal tablet of L. plantarum strain I1001
(deposit code CECT7504) in healthy women, reported that
use of this formulation, three times a week on alternate days,
achieves an adequate vaginal lactobacilli concentration [16].
These positive results led us to prospectively evaluate the
impact of the use of L. plantarum I1001, applied vaginally,
on VVC recurrence after a single-dose vaginal clotrimazole.
Patients and methods
A clinical open-label, prospective study of two non-
randomized parallel cohorts was conducted in the out-patient
gynaecological departments of the Instituto Palacios de Salud
y Medicina de la Mujer and the Instituto de Medicina EGR
(Madrid) from June 2013 until February 2015.
The first cohort (n = 33) corresponded to sexually active
women between 18 and 50 years old with symptomatic acute
VVC, who were prescribed with a standard single-dose
500 mg vaginal tablet of clotrimazole, followed by the
L. plantarum I1001 vaginal probiotic tablet (Melagyn®
Probiotico Vaginal, Gynea by Kern Pharma, Spain; each tablet
contains L. plantarum I1001 at minimum 1 × 108 colony
forming units) as adjuvant therapy (one tablet, three times a
week on alternate days, for two consecutive months, exclud-
ing days with menstruation). The second cohort (n = 22) com-
promised women of similar characteristics but prescribed the
single-dose clotrimazole treatment only.
Exclusion criteria (both cohorts) were pregnancy/breast
feeding, delivery or abortion within the previous trimester,
signs of other vaginal infections, abnormal genital bleeding
within the previous semester, or any concomitant medication
during follow-up that might significantly influence the
evaluation and/or study results (including beta-lactams,
clindamycin and tetracycline). Informed written consent was
obtained from all participants prior to enrolment. All the study
materials were approved by the Ethical Committee of the
Hospital Universitario de la Princesa (Madrid, Spain).
Vaginal probiotic and dose administration
L. plantarum I1001 (CECT7504) is a selected, patented strain
which has been tested in vitro, showing good adherence to
vaginal epithelial cells (VEC), high acidification of simulated
vaginal media, high tolerance to antimicrobial factors of
inflamed vaginal fluid, and intrinsically resistant to high con-
centrations of some typical antibiotics (ATB) and antifungals
used for vaginal affections. This L. plantarum I1001 formula-
tion has been previously tested in healthy women during a
pilot open label clinical trial at Hospital Vall d’Hebron
(Barcelona, Spain) [16], showing successful colonization of
the vagina for at least 48 hours, therefore, allowing its appli-
cation on alternate days.
Data collection and follow-up
The primary outcome was the recurrence-free survival of
signs/symptoms of VVC, as collected in the follow-up visits
at two and three months after the azole treatment, within the
possibility of a six-month visit for women with history of
RVVC.
Specific risk factors (age, diabetes mellitus, RVVC within
the last year, ATB prior to enrolment, diaphragm/intrauterine
device or oral contraception, immunosuppression and history
of other non-candida vulvovaginitis [VV]) were recorded at
baseline. Self-reported presence of pruritus, vulvar soreness/
irritation and burning vulvar pain, and signs of vaginal dis-
charge, vulvar erythema and edema, and malodorousness,
were assessed using a semi-quantitative scale (0 = absent,
1 = light, 2 = moderate and 3 = severe). In the L. plantarum
I1001 cohort, physicians rated the effectiveness and tolerabil-
ity of the product, using a semi-quantitative scale (0 = inade-
quate, 1 = fair, 2 = good, 3 = very good), while women also
reported their opinion on the product and evaluated its tolera-
bility and their satisfaction using the Spanish version of the
Treatment Satisfaction Questionnaire for Medication (TSQM
version 1.4) [17].
Statistical analysis
Baseline characteristics (including known risk factors for
VVC) of both cohorts were compared with Fisher’s exact test
(categorical variables) and Mann-Whitney’s U test (quantita-
tive variables). Values were summarized and presented as per-
centages or mean ± standard deviation (SD). For the main
clinical outcome (VVC recurrence-free survival), use of the
Eur J Clin Microbiol Infect Dis. 

Scene 2

Probiotics are defined as Blive microorganisms, which
when administered in adequate amounts, confer a health ben-
efit on the host^ [11]. Probiotic properties are strain-specific
so their positive health effects cannot be extrapolated to other
strains of the same species or genus [12]. Lactobacilli are the
predominant bacteria in a healthy vaginal ecosystem, and have
been proposed for the treatment and prevention of genitouri-
nary infections including candidiasis [4, 6]. Local use of these
microorganisms results in immunomodulation responses and
the restoration of vaginal microbiota, interfering with the col-
onization and growth of potential pathogens such as Candida
[13]. A Lactobacilli-dominated vaginal microbiota produces
significant levels of lactic acid with strong microbicidal prop-
erties [14].
Evidence of synergy between adjuvant local Lactobacilli
and azole treatment for VVC has been reported [9, 15].
Nevertheless, evidence of the benefit of adding probiotics to
the standard azole on the risk of symptomatic recurrence is
lacking.
Recently, a pilot study for the evaluation of colonization
and tolerability of a vaginal tablet of L. plantarum strain I1001
(deposit code CECT7504) in healthy women, reported that
use of this formulation, three times a week on alternate days,
achieves an adequate vaginal lactobacilli concentration [16].
These positive results led us to prospectively evaluate the
impact of the use of L. plantarum I1001, applied vaginally,
on VVC recurrence after a single-dose vaginal clotrimazole.
Patients and methods
A clinical open-label, prospective study of two non-
randomized parallel cohorts was conducted in the out-patient
gynaecological departments of the Instituto Palacios de Salud
y Medicina de la Mujer and the Instituto de Medicina EGR
(Madrid) from June 2013 until February 2015.
The first cohort (n = 33) corresponded to sexually active
women between 18 and 50 years old with symptomatic acute
VVC, who were prescribed with a standard single-dose
500 mg vaginal tablet of clotrimazole, followed by the
L. plantarum I1001 vaginal probiotic tablet (Melagyn®
Probiotico Vaginal, Gynea by Kern Pharma, Spain; each tablet
contains L. plantarum I1001 at minimum 1 × 108 colony
forming units) as adjuvant therapy (one tablet, three times a
week on alternate days, for two consecutive months, exclud-
ing days with menstruation). The second cohort (n = 22) com-
promised women of similar characteristics but prescribed the
single-dose clotrimazole treatment only.
Exclusion criteria (both cohorts) were pregnancy/breast
feeding, delivery or abortion within the previous trimester,
signs of other vaginal infections, abnormal genital bleeding
within the previous semester, or any concomitant medication
during follow-up that might significantly influence the
evaluation and/or study results (including beta-lactams,
clindamycin and tetracycline). Informed written consent was
obtained from all participants prior to enrolment. All the study
materials were approved by the Ethical Committee of the
Hospital Universitario de la Princesa (Madrid, Spain).
Vaginal probiotic and dose administration
L. plantarum I1001 (CECT7504) is a selected, patented strain
which has been tested in vitro, showing good adherence to
vaginal epithelial cells (VEC), high acidification of simulated
vaginal media, high tolerance to antimicrobial factors of
inflamed vaginal fluid, and intrinsically resistant to high con-
centrations of some typical antibiotics (ATB) and antifungals
used for vaginal affections. This L. plantarum I1001 formula-
tion has been previously tested in healthy women during a
pilot open label clinical trial at Hospital Vall d’Hebron
(Barcelona, Spain) [16], showing successful colonization of
the vagina for at least 48 hours, therefore, allowing its appli-
cation on alternate days.
Data collection and follow-up
The primary outcome was the recurrence-free survival of
signs/symptoms of VVC, as collected in the follow-up visits
at two and three months after the azole treatment, within the
possibility of a six-month visit for women with history of
RVVC.
Specific risk factors (age, diabetes mellitus, RVVC within
the last year, ATB prior to enrolment, diaphragm/intrauterine
device or oral contraception, immunosuppression and history
of other non-candida vulvovaginitis [VV]) were recorded at
baseline. Self-reported presence of pruritus, vulvar soreness/
irritation and burning vulvar pain, and signs of vaginal dis-
charge, vulvar erythema and edema, and malodorousness,
were assessed using a semi-quantitative scale (0 = absent,
1 = light, 2 = moderate and 3 = severe). In the L. plantarum
I1001 cohort, physicians rated the effectiveness and tolerabil-
ity of the product, using a semi-quantitative scale (0 = inade-
quate, 1 = fair, 2 = good, 3 = very good), while women also
reported their opinion on the product and evaluated its tolera-
bility and their satisfaction using the Spanish version of the
Treatment Satisfaction Questionnaire for Medication (TSQM
version 1.4) [17].
Statistical analysis
Baseline characteristics (including known risk factors for
VVC) of both cohorts were compared with Fisher’s exact test
(categorical variables) and Mann-Whitney’s U test (quantita-
tive variables). Values were summarized and presented as per-
centages or mean ± standard deviation (SD). For the main
clinical outcome (VVC recurrence-free survival), use of the
Eur J Clin Microbiol Infect Dis

vaginal probiotic and additional factors that might influ-
ence the risk of recurrence (see above) were included in a
multivariate Cox (proportional-hazards) regression model;
these results are presented as Hazard ratios (HR) and their
95 % confidence intervals (95 %CI). Drop-outs were cen-
sored at the time of last follow-up. In an attempt to assess
the change in recurrence within cohorts, we compared the
average number of VVC episodes per trimester within
12 months prior to enrolment including the baseline episode
of women with three-months follow-up (obtained from clini-
cal records) to those observed during the follow-up at three
months, by using the non-parametric Wilcoxon signed-rank
test. Only subjects with complete clinical record for the past
12 months and complete follow-up at three months were
considered for this later analysis. Results on the TSQM are
presented as median and interquartile range (IQR), because of
their non-normality. No imputation was performed for missing
values, and significance was set at a level of 5 %, two-tailed.
All statistical tests were performed with SPSS Statistics for
Windows v22.0 (IBM Corp. 2013. Armonk, NY).
Results
Baseline data
A general description of the patients and current VVC episode
is presented in Table 1. Flow of patients throughout the study
is presented in Fig. 1. Drop-out rate was slightly higher in the
clotrimazole-only cohort (18.5 % vs. 5.7 % at 2 months, and
24.1 % vs. 15.4 % at 3 months), although the difference did
not reach statistical significance.
Among the 55 women from both cohorts, 50 (91 %) had a
history of previous VVC anytime during their lifespan
(Table 1), from which 33 (63.5 %) met criteria for VVC within
the 12 months prior to enrolment. The number of VVC epi-
sodes during the previous 12 months was higher in the
L. plantarum I1001 cohort (4.55 ± 2.58 vs. 3.53 ± 1.88), al-
though this difference did not reach statistical signifi-
cance. Conversely, previous non-Candida VVs were re-
ported exclusively in the antifungal-only cohort (n = 3,
P = 0.059, statistical trend).
Current VVC was generally characterized by the specialist
with vulvar erythema, vaginal discharge, edema and burning/
soreness, without differences between cohorts regarding the
number of patients with each symptom.
Patients’ and gynaecologists’ evaluation of the vaginal
probiotic
Overall, patients treated with the vaginal probiotic reported a
compliance of 91.3 %. The TSQM (n = 29) results showed
that patients reported high ratings of efficacy, lack of side
effects, convenience and overall satisfaction (Table 2). Only
one patient reported a side effect (edema and erythema) of
moderate intensity, and rated it as Bsomewhat affecting her
satisfaction with the treatment^ in the TSQM questionnaire
(i.e. a score of 3 in a Likert 1–5 scale). Moreover, gynecolo-
gists considered efficacy of the test product in their patients
(n = 31) as Bgood^/Bvery good^ in 87 % of cases, and tolera-
bility as Bgood^/Bvery good^ in 100 % of them.
Time until symptomatic recurrence
Overall, 10/31 (32.2 %) of women in the probiotic group
and 10/17 (58.8 %) of women in the control group had
experienced one or more symptomatic recurrences during
the 3–month follow-up (P = 0.125).
The Cox regression model on the combined cohorts
(Table 3) showed that adjusted HR for developing a new
VVC within three months of the single-dose clotrimazole,
depending on probiotic use, was 0.30 (95 %CI: 0.10–0.91)
and that this effect was statistically significant (P = 0.033,
Fig. 2). Among other risk factors, ATB prior to enrolment also
showed a significant effect, with an adjusted HR of 10.46
(95 %CI: 2.18–50.12); (P = 0.003, Fig. 3). Thus, comparison
of the recurrence-free survival shows a clear increase in ad-
justed survival at 3 months (73 % vs. 35 %, i.e. a +108.6 %
increase) in women receiving the probiotic, while the ‘previ-
ous ATB’ factor shows marked effect in the opposite direction
(6 % vs. 68 %).
Analysis of the subgroup with history of RVVC (n = 23 of
the clotrimazole + probiotic cohort, and n = 12 from the clo-
trimazole cohort) at 6-months of follow-up, showed a similar
difference in adjusted survival among treatment cohorts (63 %
vs. 22 %, Supplementary material). The adjusted HR for
symptomatic recurrence depending on probiotic use was
0.30 (95 %CI: 0.10–0.89; P = 0.03), likewise for the effect
of ‘previous ATB’ (Supplementary material).
Differences in the number of episodes
In the L. plantarum I1001 cohort the average number of VVC
episodes per trimester during the 3-month follow-up was 0.45
± 0.72, down from 1.14 ± 0.65 episodes/trimester during the
12 months prior to enrolment, thus representing a reduction of
59 % (P = 0.001). Conversely, such difference was not ob-
served in the clotrimazole-only cohort (0.67 ± 0.62 vs. 0.88
± 0.47 episodes/trimester; P > 0.1).
Discussion
In the present study, follow-up therapy with vaginal tab-
lets with L. plantarum I1001 after a single-dose 500 mg
clotrimazole was found to significantly reduce the risk of
Eur J Clin Microbiol Infect Dis. 

Scene 3

vaginal probiotic and additional factors that might influ-
ence the risk of recurrence (see above) were included in a
multivariate Cox (proportional-hazards) regression model;
these results are presented as Hazard ratios (HR) and their
95 % confidence intervals (95 %CI). Drop-outs were cen-
sored at the time of last follow-up. In an attempt to assess
the change in recurrence within cohorts, we compared the
average number of VVC episodes per trimester within
12 months prior to enrolment including the baseline episode
of women with three-months follow-up (obtained from clini-
cal records) to those observed during the follow-up at three
months, by using the non-parametric Wilcoxon signed-rank
test. Only subjects with complete clinical record for the past
12 months and complete follow-up at three months were
considered for this later analysis. Results on the TSQM are
presented as median and interquartile range (IQR), because of
their non-normality. No imputation was performed for missing
values, and significance was set at a level of 5 %, two-tailed.
All statistical tests were performed with SPSS Statistics for
Windows v22.0 (IBM Corp. 2013. Armonk, NY).
Results
Baseline data
A general description of the patients and current VVC episode
is presented in Table 1. Flow of patients throughout the study
is presented in Fig. 1. Drop-out rate was slightly higher in the
clotrimazole-only cohort (18.5 % vs. 5.7 % at 2 months, and
24.1 % vs. 15.4 % at 3 months), although the difference did
not reach statistical significance.
Among the 55 women from both cohorts, 50 (91 %) had a
history of previous VVC anytime during their lifespan
(Table 1), from which 33 (63.5 %) met criteria for VVC within
the 12 months prior to enrolment. The number of VVC epi-
sodes during the previous 12 months was higher in the
L. plantarum I1001 cohort (4.55 ± 2.58 vs. 3.53 ± 1.88), al-
though this difference did not reach statistical signifi-
cance. Conversely, previous non-Candida VVs were re-
ported exclusively in the antifungal-only cohort (n = 3,
P = 0.059, statistical trend).
Current VVC was generally characterized by the specialist
with vulvar erythema, vaginal discharge, edema and burning/
soreness, without differences between cohorts regarding the
number of patients with each symptom.
Patients’ and gynaecologists’ evaluation of the vaginal
probiotic
Overall, patients treated with the vaginal probiotic reported a
compliance of 91.3 %. The TSQM (n = 29) results showed
that patients reported high ratings of efficacy, lack of side
effects, convenience and overall satisfaction (Table 2). Only
one patient reported a side effect (edema and erythema) of
moderate intensity, and rated it as Bsomewhat affecting her
satisfaction with the treatment^ in the TSQM questionnaire
(i.e. a score of 3 in a Likert 1–5 scale). Moreover, gynecolo-
gists considered efficacy of the test product in their patients
(n = 31) as Bgood^/Bvery good^ in 87 % of cases, and tolera-
bility as Bgood^/Bvery good^ in 100 % of them.
Time until symptomatic recurrence
Overall, 10/31 (32.2 %) of women in the probiotic group
and 10/17 (58.8 %) of women in the control group had
experienced one or more symptomatic recurrences during
the 3–month follow-up (P = 0.125).
The Cox regression model on the combined cohorts
(Table 3) showed that adjusted HR for developing a new
VVC within three months of the single-dose clotrimazole,
depending on probiotic use, was 0.30 (95 %CI: 0.10–0.91)
and that this effect was statistically significant (P = 0.033,
Fig. 2). Among other risk factors, ATB prior to enrolment also
showed a significant effect, with an adjusted HR of 10.46
(95 %CI: 2.18–50.12); (P = 0.003, Fig. 3). Thus, comparison
of the recurrence-free survival shows a clear increase in ad-
justed survival at 3 months (73 % vs. 35 %, i.e. a +108.6 %
increase) in women receiving the probiotic, while the ‘previ-
ous ATB’ factor shows marked effect in the opposite direction
(6 % vs. 68 %).
Analysis of the subgroup with history of RVVC (n = 23 of
the clotrimazole + probiotic cohort, and n = 12 from the clo-
trimazole cohort) at 6-months of follow-up, showed a similar
difference in adjusted survival among treatment cohorts (63 %
vs. 22 %, Supplementary material). The adjusted HR for
symptomatic recurrence depending on probiotic use was
0.30 (95 %CI: 0.10–0.89; P = 0.03), likewise for the effect
of ‘previous ATB’ (Supplementary material).
Differences in the number of episodes
In the L. plantarum I1001 cohort the average number of VVC
episodes per trimester during the 3-month follow-up was 0.45
± 0.72, down from 1.14 ± 0.65 episodes/trimester during the
12 months prior to enrolment, thus representing a reduction of
59 % (P = 0.001). Conversely, such difference was not ob-
served in the clotrimazole-only cohort (0.67 ± 0.62 vs. 0.88
± 0.47 episodes/trimester; P > 0.1).
Discussion
In the present study, follow-up therapy with vaginal tab-
lets with L. plantarum I1001 after a single-dose 500 mg
clotrimazole was found to significantly reduce the risk of
Eur J Clin Microbiol Infect Dis

symptomatic recurrence within 3 months after an acute
VVC, with similar findings in the subset of patients with
RVVC within 6 months of follow-up.
Although most baseline characteristics were comparable,
patient distribution among cohorts was not randomized and
resulted in a larger number of the VVC episodes of women
Table 1 Demographic,
reproductive and gynaecological
characteristics
Characteristic
Total sample
Clotrimazole + L.
plantarum I1001
Clotrimazole
P-value
Total cases
55
33
22
Age (years)
33.36 ± 8.61
33.58 ± 8.23
33.05 ± 9.34
0.830
Marital status
Single
27 (49.1 %)
16 (48.5 %)
11 (50.0 %)
0.190
Divorced
2 (3.6 %)
0 (0.0 %)
2 (9.1 %)
Married/living together
26 (47.3 %)
17 (51.5 %)
9 (40.9 %)
Gynaecological history and reproductive state
Child bearing age
51 (92.7 %)
32 (97.0 %)
19 (86.4 %)
0.290
Perimenopause
4 (7.3 %)
1 (3.0 %)
3 (13.6 %)
Children (yes)
19 (34.5 %)
10 (30.3 %)
9 (40.9 %)
0.564
Relevant medical and/or surgical history
11 (20.4 %)
9 (28.1 %)
2 (9.1 %)
0.167
Risk factors for VV
Antibiotics prior to study enrolmenta
6 (11.5 %)
4 (12.9 %)
2 (9.5 %)
1.000
Diaphragm or IUD contraceptiona
3 (5.8 %)
2 (6.5 %)
1 (4.8 %)
1.000
Oral contraceptiona
11 (21.2 %)
6 (19.4 %)
5 (23.8 %)
0.739
Diabetes mellitusa
0 (0 %)
-
-
-
Immunosuppressiona
1 (1.9 %)
1 (3.2 %)
0 (0.0 %)
1.000
Othersa
4 (7.7 %)
2 (6.5 %)
2 (9.5 %)
1.000
Vulvovaginitis history
Other non-candida VV
3 (5.6 %)
0 (0.0 %)
3 (13.6 %)
0.059
Any previous VVC
50 (90.9 %)
29 (87.9 %)
21 (95.5 %)
0.638
Age at first VVC episodeb
25.15 ± 7.73
25.14 ± 8.07
25.17 ± 7.37
0.742
Total VVC episodes everb
9.54 ± 7.96
9.66 ± 6.04
9.38 ± 10.20
0.242
RVVC evera
35 (67.3 %)
23 (69.7 %)
12 (63.2 %)
0.761
Total VVC in the last 12 monthsc
4.22 ± 2.40
4.55 ± 2.58
3.53 ± 1.88
0.142
Any previous VVC in the last 12 monthsa
33 (63.5 %)
23 (69.7 %)
10 (52.6 %)
0.246
Last VVC episode
Months since last VVCd
4.06 ± 3.73
3.72 ± 3.49
4.62 ± 4.13
0.413
Symptomatic days during the last VVCe
4.36 ± 2.55
4.31 ± 2.71
4.43 ± 2.36
0.719
VVC signs and symptoms
Pruritus
25 (75.8 %)
19 (86.4 %)
0.495
Burning pain/soreness
25 (75.8 %)
20 (90.9 %)
0.284
Vulvar erythema
29 (87.9 %)
21 (95.5 %)
0.638
Vaginal discharge
28 (84.8 %)
20 (90.9 %)
0.689
Vulvar pain/dyspareunia
17 (51.5 %)
12 (54.5 %)
1.000
Edema
27 (81.8 %)
19 (86.4 %)
0.727
Malodourness
11 (33.3 %)
7 (31.8 %)
1.000
VV vulvovaginitis, VVC vulvovaginal candidiasis, IUD intrauterine device, RVVC recurrent vulvovaginal
candidiasis
Values are presented as percentages or mean ± standard deviation (SD). Percentages are calculated based on the
total number of patients available for each item. Comparison among the two intervention groups (P value) was
performed with Fisher's test (categorical variables) or Mann Whitney test (quantitative variables)
a Data available for 52 patientsb Data available for 47 patients c Data available for 46 patients d Data available for
45 patients e Data available for 50 patients
Eur J Clin Microbiol Infect Dis. 

Scene 4

symptomatic recurrence within 3 months after an acute
VVC, with similar findings in the subset of patients with
RVVC within 6 months of follow-up.
Although most baseline characteristics were comparable,
patient distribution among cohorts was not randomized and
resulted in a larger number of the VVC episodes of women
Table 1 Demographic,
reproductive and gynaecological
characteristics
Characteristic
Total sample
Clotrimazole + L.
plantarum I1001
Clotrimazole
P-value
Total cases
55
33
22
Age (years)
33.36 ± 8.61
33.58 ± 8.23
33.05 ± 9.34
0.830
Marital status
Single
27 (49.1 %)
16 (48.5 %)
11 (50.0 %)
0.190
Divorced
2 (3.6 %)
0 (0.0 %)
2 (9.1 %)
Married/living together
26 (47.3 %)
17 (51.5 %)
9 (40.9 %)
Gynaecological history and reproductive state
Child bearing age
51 (92.7 %)
32 (97.0 %)
19 (86.4 %)
0.290
Perimenopause
4 (7.3 %)
1 (3.0 %)
3 (13.6 %)
Children (yes)
19 (34.5 %)
10 (30.3 %)
9 (40.9 %)
0.564
Relevant medical and/or surgical history
11 (20.4 %)
9 (28.1 %)
2 (9.1 %)
0.167
Risk factors for VV
Antibiotics prior to study enrolmenta
6 (11.5 %)
4 (12.9 %)
2 (9.5 %)
1.000
Diaphragm or IUD contraceptiona
3 (5.8 %)
2 (6.5 %)
1 (4.8 %)
1.000
Oral contraceptiona
11 (21.2 %)
6 (19.4 %)
5 (23.8 %)
0.739
Diabetes mellitusa
0 (0 %)
-
-
-
Immunosuppressiona
1 (1.9 %)
1 (3.2 %)
0 (0.0 %)
1.000
Othersa
4 (7.7 %)
2 (6.5 %)
2 (9.5 %)
1.000
Vulvovaginitis history
Other non-candida VV
3 (5.6 %)
0 (0.0 %)
3 (13.6 %)
0.059
Any previous VVC
50 (90.9 %)
29 (87.9 %)
21 (95.5 %)
0.638
Age at first VVC episodeb
25.15 ± 7.73
25.14 ± 8.07
25.17 ± 7.37
0.742
Total VVC episodes everb
9.54 ± 7.96
9.66 ± 6.04
9.38 ± 10.20
0.242
RVVC evera
35 (67.3 %)
23 (69.7 %)
12 (63.2 %)
0.761
Total VVC in the last 12 monthsc
4.22 ± 2.40
4.55 ± 2.58
3.53 ± 1.88
0.142
Any previous VVC in the last 12 monthsa
33 (63.5 %)
23 (69.7 %)
10 (52.6 %)
0.246
Last VVC episode
Months since last VVCd
4.06 ± 3.73
3.72 ± 3.49
4.62 ± 4.13
0.413
Symptomatic days during the last VVCe
4.36 ± 2.55
4.31 ± 2.71
4.43 ± 2.36
0.719
VVC signs and symptoms
Pruritus
25 (75.8 %)
19 (86.4 %)
0.495
Burning pain/soreness
25 (75.8 %)
20 (90.9 %)
0.284
Vulvar erythema
29 (87.9 %)
21 (95.5 %)
0.638
Vaginal discharge
28 (84.8 %)
20 (90.9 %)
0.689
Vulvar pain/dyspareunia
17 (51.5 %)
12 (54.5 %)
1.000
Edema
27 (81.8 %)
19 (86.4 %)
0.727
Malodourness
11 (33.3 %)
7 (31.8 %)
1.000
VV vulvovaginitis, VVC vulvovaginal candidiasis, IUD intrauterine device, RVVC recurrent vulvovaginal
candidiasis
Values are presented as percentages or mean ± standard deviation (SD). Percentages are calculated based on the
total number of patients available for each item. Comparison among the two intervention groups (P value) was
performed with Fisher's test (categorical variables) or Mann Whitney test (quantitative variables)
a Data available for 52 patientsb Data available for 47 patients c Data available for 46 patients d Data available for
45 patients e Data available for 50 patients
Eur J Clin Microbiol Infect Dis

with any previous VVC in the last 12 months and a dif-
ferent history of non-Candida VV in the probiotic cohort.
Taking into account the lack of randomization and those
differences between cohorts, we considered appropriate to
use a multivariate survival test to study the primary out-
come (the recurrence-free survival). After adjustment for
potential confounding factors by Cox multivariate analy-
sis, the effect of adjuvant probiotic was significant in the
full study population (at 3-months) and in women with
RVVC within a 6-month follow-up. Nevertheless, we
were surprised to observe that the hazard ratio was virtu-
ally the same in the full population at 3 months and in the
subpopulation with RVVC at 6 months. However, this
lack of difference may be due to the small study sample.
In future studies, it would be interesting to reproduce the
same analysis using a larger sample in order to clear up
whether there are any differences between the evaluation
at 3- and 6-months of follow-up regarding the use of
probiotics. Also, the difference in the fraction of women
experiencing at least one symptomatic recurrence between
the two study cohorts (32.2 % vs 58.8 %) did not reach
statistical significance. However, this analysis at study
endpoint does not take into account the baseline
differences between the two cohorts, nor captures the
time-to-event effect as the Cox analysis does. The latter
may be of special relevance in the study population, as it
was particularly prone to recurrence (67.3 % had a history
of RVVC at any time of their lives and 63.5 % reported at
least one previous VVC episode within the 12 months
prior to enrolment).
Variability among candidiasis and the selection of treat-
ment are issues that have aroused considerable discussion.
Recent reports suggest the possibility of two types of VVC:
(a) the typical, characterized by dense vaginal discharge with
occasional recurrences, and (b) the cyclic-recurrent, with hy-
persensitive reactions even in presence of small quantities of
Candida [4]. In both cases, a strong vaginal ecosystem is de-
sirable to fend off Candida strains and reduce the risk of VVC
recurrence. Several authors suggest that maintenance ther-
apy needs to be given frequently enough to prevent vag-
inal regrowth or recolonization, and transformation to a
symptomatic state in cases with Bhost intolerance^; how-
ever, once the azole is suspended, the risk of a new epi-
sode is particularly high [8].
Witt et al. [18] found no advantages in treating RVVC with
monthly 200 mg itraconazole, despite the adjuvant use of local
L. gasseri lyophilisates for 6 days. Thus, in our case, the pro-
biotic intervention was maintained over 2 months to ensure
proper concentrations of lactobacilli.
Probiotics are being used in the gynaecological field as an
alternative intervention for VVC and its prevention after ATB
therapy, and so for bacterial vaginosis. Evidence suggests that
the coaggregation of lactobacilli prevents Candida from bind-
ing to VEC [19], therefore improving the efficacy of antifun-
gals [20, 21]. A retrospective comparative study by De Seta
et al. [21] showed a better subjective resolution of typical
VVC symptoms after treatment with clotrimazole 2 % vaginal
cream for 3 days followed by vaginal application of
Fig. 1 Patients flow-chart across study visits
Table 2
Patients satisfaction with L. plantarum I1001 vaginal tablets
(TSQM questionnaire)
Parameter
Total number of patients
Median
IQR
Efficacy
29
83.3
66.7–83.3
Side effects (lack of)
29
100
100–100
Convenience
29
83.3
66.7–83.3
Overall satisfaction
29
76.4
52.8–76.4
IQR interquartile range
Table 3
Cox proportional-hazards regression model for recurrence of
VVC at 3 months
Variable
Hazard
ratio
95 % CI
P-value
Lower
Upper
Age
0.96
0.90
1.03
0.228
Any previous VVC within the last
12 months
1.69
0.49
5.79
0.406
Antibiotics prior to enrolment
10.46
2.18
50.12
0.003
Diaphragm or IUD contraception
1.55
0.30
7.91
0.600
Immunosuppression
0.42
0.03
5.15
0.493
Oral contraception
0.97
0.22
4.32
0.965
History of other non-candida VV
0.34
0.03
3.52
0.365
Use of L. plantarum I1001
0.30
0.10
0.91
0.033
VVC vulvovaginal candidiasis, RVVC recurrent VVC, VV vulvovaginitis,
IUD intrauterine device, CI confidence intervals
Eur J Clin Microbiol Infect Dis. 

Scene 5

with any previous VVC in the last 12 months and a dif-
ferent history of non-Candida VV in the probiotic cohort.
Taking into account the lack of randomization and those
differences between cohorts, we considered appropriate to
use a multivariate survival test to study the primary out-
come (the recurrence-free survival). After adjustment for
potential confounding factors by Cox multivariate analy-
sis, the effect of adjuvant probiotic was significant in the
full study population (at 3-months) and in women with
RVVC within a 6-month follow-up. Nevertheless, we
were surprised to observe that the hazard ratio was virtu-
ally the same in the full population at 3 months and in the
subpopulation with RVVC at 6 months. However, this
lack of difference may be due to the small study sample.
In future studies, it would be interesting to reproduce the
same analysis using a larger sample in order to clear up
whether there are any differences between the evaluation
at 3- and 6-months of follow-up regarding the use of
probiotics. Also, the difference in the fraction of women
experiencing at least one symptomatic recurrence between
the two study cohorts (32.2 % vs 58.8 %) did not reach
statistical significance. However, this analysis at study
endpoint does not take into account the baseline
differences between the two cohorts, nor captures the
time-to-event effect as the Cox analysis does. The latter
may be of special relevance in the study population, as it
was particularly prone to recurrence (67.3 % had a history
of RVVC at any time of their lives and 63.5 % reported at
least one previous VVC episode within the 12 months
prior to enrolment).
Variability among candidiasis and the selection of treat-
ment are issues that have aroused considerable discussion.
Recent reports suggest the possibility of two types of VVC:
(a) the typical, characterized by dense vaginal discharge with
occasional recurrences, and (b) the cyclic-recurrent, with hy-
persensitive reactions even in presence of small quantities of
Candida [4]. In both cases, a strong vaginal ecosystem is de-
sirable to fend off Candida strains and reduce the risk of VVC
recurrence. Several authors suggest that maintenance ther-
apy needs to be given frequently enough to prevent vag-
inal regrowth or recolonization, and transformation to a
symptomatic state in cases with Bhost intolerance^; how-
ever, once the azole is suspended, the risk of a new epi-
sode is particularly high [8].
Witt et al. [18] found no advantages in treating RVVC with
monthly 200 mg itraconazole, despite the adjuvant use of local
L. gasseri lyophilisates for 6 days. Thus, in our case, the pro-
biotic intervention was maintained over 2 months to ensure
proper concentrations of lactobacilli.
Probiotics are being used in the gynaecological field as an
alternative intervention for VVC and its prevention after ATB
therapy, and so for bacterial vaginosis. Evidence suggests that
the coaggregation of lactobacilli prevents Candida from bind-
ing to VEC [19], therefore improving the efficacy of antifun-
gals [20, 21]. A retrospective comparative study by De Seta
et al. [21] showed a better subjective resolution of typical
VVC symptoms after treatment with clotrimazole 2 % vaginal
cream for 3 days followed by vaginal application of
Fig. 1 Patients flow-chart across study visits
Table 2
Patients satisfaction with L. plantarum I1001 vaginal tablets
(TSQM questionnaire)
Parameter
Total number of patients
Median
IQR
Efficacy
29
83.3
66.7–83.3
Side effects (lack of)
29
100
100–100
Convenience
29
83.3
66.7–83.3
Overall satisfaction
29
76.4
52.8–76.4
IQR interquartile range
Table 3
Cox proportional-hazards regression model for recurrence of
VVC at 3 months
Variable
Hazard
ratio
95 % CI
P-value
Lower
Upper
Age
0.96
0.90
1.03
0.228
Any previous VVC within the last
12 months
1.69
0.49
5.79
0.406
Antibiotics prior to enrolment
10.46
2.18
50.12
0.003
Diaphragm or IUD contraception
1.55
0.30
7.91
0.600
Immunosuppression
0.42
0.03
5.15
0.493
Oral contraception
0.97
0.22
4.32
0.965
History of other non-candida VV
0.34
0.03
3.52
0.365
Use of L. plantarum I1001
0.30
0.10
0.91
0.033
VVC vulvovaginal candidiasis, RVVC recurrent VVC, VV vulvovaginitis,
IUD intrauterine device, CI confidence intervals
Eur J Clin Microbiol Infect Dis

L. plantarum P17630 daily for 6 days, plus a weekly applica-
tion for 4 weeks, all compared with azole cream alone.
However, they excluded patients with known risk factors of
VVC (RVVC, prior ATB, etc.). In our study, on the contrary,
no exclusion of them was considered since these women are
more likely to require supplementary therapies to standard
treatments.
Despite the similarities between the published evidence
and our results, it should be stressed that most reports have
assessed multidose azole treatment [20, 21]; hence, the effects
of the L. plantarum I1001 strain may be of particular interest
since use of these adjuvant vaginal tablets was associated with
a significant 3-fold reduction in the HR of recurrence, during a
3-month follow-up. To our knowledge, this is the first study to
report a significant effect of a probiotic in the risk of symp-
toms recurrence in women with high prevalence of RVVC. It
is noteworthy that ATB prior to enrolment significantly in-
creased the VVC recurrence risk, consistent with previous
reports such as those from Pirotta et al. [22] and Spinillo
et al. [23], who also concluded that this factor could entail
even a first-episode of VVC. Their findings and those present-
ed herein enhance the role of vaginal probiotics as a preven-
tive strategy (complementing pharmacological treatment), and
a primary intervention for reducing the risk of vaginal infec-
tions. Since ‘Prior ATB’ per se seems to significantly increase
probabilities of VVC and of recurrences, L. plantarum I1001
Fig. 2 Recurrence-free survival
curve 3 months after single-dose
clotrimazole treatment.
Comparison of the overall
recurrence-free survival between
patients treated with clotrimazole
500 mg single dose-only vs. clo-
trimazole + L. plantarum I1001.
The difference on adjusted sur-
vival between the two cohorts is
significant, with a higher
recurrence-free survival in pa-
tients receiving adjuvant vaginal
probiotic (HR [0.30 [95 %CI:
0.10–0.91]; P = 0.033, multivari-
ate Cox regression model)
Fig. 3 Recurrence-free survival
curve 3 months after single-dose
clotrimazole treatment. Patients
with history of antibiotic treat-
ment (ATB) prior to enrolment.
Comparison of the overall
recurrence-free survival depend-
ing on the use of antibiotic treat-
ment prior to enrolment. The dif-
ference on adjusted survival be-
tween the two cohorts achieves
significance with a markedly
lower recurrence-free survival in
patients with such risk factor (HR:
10.46 [95 %CI: 2.18–50.12]; P =
0.003, multivariate Cox regres-
sion model)
Eur J Clin Microbiol Infect Dis. 

Scene 6

L. plantarum P17630 daily for 6 days, plus a weekly applica-
tion for 4 weeks, all compared with azole cream alone.
However, they excluded patients with known risk factors of
VVC (RVVC, prior ATB, etc.). In our study, on the contrary,
no exclusion of them was considered since these women are
more likely to require supplementary therapies to standard
treatments.
Despite the similarities between the published evidence
and our results, it should be stressed that most reports have
assessed multidose azole treatment [20, 21]; hence, the effects
of the L. plantarum I1001 strain may be of particular interest
since use of these adjuvant vaginal tablets was associated with
a significant 3-fold reduction in the HR of recurrence, during a
3-month follow-up. To our knowledge, this is the first study to
report a significant effect of a probiotic in the risk of symp-
toms recurrence in women with high prevalence of RVVC. It
is noteworthy that ATB prior to enrolment significantly in-
creased the VVC recurrence risk, consistent with previous
reports such as those from Pirotta et al. [22] and Spinillo
et al. [23], who also concluded that this factor could entail
even a first-episode of VVC. Their findings and those present-
ed herein enhance the role of vaginal probiotics as a preven-
tive strategy (complementing pharmacological treatment), and
a primary intervention for reducing the risk of vaginal infec-
tions. Since ‘Prior ATB’ per se seems to significantly increase
probabilities of VVC and of recurrences, L. plantarum I1001
Fig. 2 Recurrence-free survival
curve 3 months after single-dose
clotrimazole treatment.
Comparison of the overall
recurrence-free survival between
patients treated with clotrimazole
500 mg single dose-only vs. clo-
trimazole + L. plantarum I1001.
The difference on adjusted sur-
vival between the two cohorts is
significant, with a higher
recurrence-free survival in pa-
tients receiving adjuvant vaginal
probiotic (HR [0.30 [95 %CI:
0.10–0.91]; P = 0.033, multivari-
ate Cox regression model)
Fig. 3 Recurrence-free survival
curve 3 months after single-dose
clotrimazole treatment. Patients
with history of antibiotic treat-
ment (ATB) prior to enrolment.
Comparison of the overall
recurrence-free survival depend-
ing on the use of antibiotic treat-
ment prior to enrolment. The dif-
ference on adjusted survival be-
tween the two cohorts achieves
significance with a markedly
lower recurrence-free survival in
patients with such risk factor (HR:
10.46 [95 %CI: 2.18–50.12]; P =
0.003, multivariate Cox regres-
sion model)
Eur J Clin Microbiol Infect Dis

could be of relevance as a resource for women needing to
replenish their vaginal microbiota after taking ATB.
It has to be noted that, in our study, VVC relapse was
considered on the basis of any self-reported sign/symptom
suggesting a new episode. Despite the subjective nature of
these criteria, it could probably represent a more realistic and
practical criteria in daily practice, considering that the pure
presence of symptoms can entail a medical consultation and/
or use of over-the-counter medication. Beyond the clinical
impact of the probiotic, high rates of satisfaction, tolerability
and effectiveness were reported by both patients and
gynaecologists.
Our study has certain limitations that must be pointed out,
such as the previously commented lack of randomization and
the open-label nature of the trial, thus a placebo effect cannot
be ruled out. However, it is worth pointing out that the high
number of women fulfilling RVVC criteria suggests they may
be well aware of the symptoms of a true VVC episode. And
finally, the sample size available was limited, and drop-outs
during the study follow-up were not uncommon. Thus, addi-
tional studies should be conducted to evaluate the effective-
ness of L. plantarum I1001, using double blind randomized
designs and larger sample sizes. Also, it would be interesting
to evaluate the efficacy of L. plantarum I1001 as add-on or
follow-up therapy of other antifungal regimes, as well as for
preventing the recurrence of bacterial vaginosis.
Acknowledgments
The authors thank all the gynaecologists of the
Instituto Palacios de Salud y Medicina de la Mujer and the Instituto de
Medicina EGR Madrid who actively participated in the study and to
Dr. Cindy L. Larios from the Medical Department of Clever
Instruments S.L. (Barcelona) for the statistical analysis and editorial
assistance for the manuscript.
Compliance with ethical standards
Funding
Gynea by Kern Pharma funded the study.
Conflict of interest
J. Espadaler is a full-time employee of AB-Biotics
S.A; C. Prieto is a full-time employee of Gynea by Kern Pharma.
Ethical approval
All the study materials were reviewed and approved
by the Ethical Committee of the Hospital Universitario de la Princesa
(Madrid, Spain).
Informed consent
Informed written consent was obtained from all
participants prior to enrolment.
Open Access This article is distributed under the terms of the Creative
Commons Attribution 4.0 International License (http://
creativecommons.org/licenses/by/4.0/), which permits unrestricted use,
distribution, and reproduction in any medium, provided you give
appropriate credit to the original author(s) and the source, provide a link
to the Creative Commons license, and indicate if changes were made.
References
1.
Richter SS, Galask RP, Messer SA, Hollis RJ, Diekema DJ, Pfaller
MA (2005) Antifungal susceptibilities of Candida species causing
vulvovaginitis and epidemiology of recurrent cases. J Clin
Microbiol 43:2155–2162
2.
Tibaldi C, Cappello N, Latino MA et al (2009) Vaginal and
endocervical microorganisms in symptomatic and asymptomatic
non-pregnant females: risk factors and rates of occurrence. Clin
Microbiol Infect 15:670–679
3.
Peters BM, Yano J, Noverr MC, Fidel PL (2014) Candida vaginitis:
when opportunism knocks, the host responds. PLoS Pathog 10:
e1003965
4.
Murina F, Graziottin A, Felice R et al (2011) The recurrent
vulvovaginal candidiasis: proposal of a personalized therapeutic
protocol. ISRN Obstet Gynecol 2011:806065
5.
Marchaim D, Lemanek L, Bheemreddy S et al (2012) Fluconazole-
resistant Candida albicans vulvovaginitis. Obstet Gynecol 120:
1407–1414
6.
Sobel JD (2016) Recurrent vulvovaginal candidiasis. Am J Obstet
Gynecol 214:15–21
7.
Mendling W, Brasch J, Cornely OA, Effendy I, Friese K, Ginter-
Hanselmayer G et al (2015) Guideline: vulvovaginal candidosis
(AWMF 015/072). S2k (excluding chronic mucocutaneous
candidosis). Mycoses 58:1–15
8.
Dovnik A, Golle A, Novak D, Arko D, Takac I (2015) Treatment of
vulvovaginal candidiasis: a review of literature. Acta
Dermatovenerol Alp Pannonica Adriat 24:5–7
9.
Kovachev SM (2015) Local probiotic therapy for vaginal Candida
albicans infections. Probiotics Antimicrob Proteins 7:38–44
10.
Sobel JD, Zervos M, Reed BD, Hooton T, Soper D, Nyirjesy P et al
(2003) Fluconazole susceptibility of vaginal isolates obtained from
women with complicated Candida vaginitis: clinical implications.
Antimicrob Agents Chemother 47:34–38
11.
Food and Agriculture Organization of the United Nations and
World Health Organization. 2001, posting date. Regulatory and
clinical aspects of dairy probiotics. Food and Agriculture
Organization of the United Nations and World Health
Organization Expert Consultation Report. Food and Agriculture
Organization of the United Nations and World Health
Organization Working Group Report. http://www.who.
int/foodsafety/fs_management/en/probiotic_guidelines.pdf.
Accessed 30 June 2016
12.
Hill C, Guarner F, Reid G et al (2014) Expert consensus document.
The International Scientific Association for Probiotics and
Prebiotics consensus statement on the scope and appropriate use
of the term probiotic. Nat Rev Gastroenterol Hepatol 11:506–514
13.
Borges S, Silva J, Teixeira P (2014) The role of lactobacilli and
probiotics in maintaining vaginal health. Arch Gynecol Obstet
289:479–489
14.
O’Hanlon DE, Moench TR, Cone RA (2013) Vaginal pH and
microbiocidal lactic acid when lactobacilli dominate the microbio-
ta. PLoS One 8:e80074
15.
Martinez RC, Franceschini SA, Patta MC, Quintana SM, Candido
RC, Ferreira JC et al (2009) Improved treatment of vulvovaginal
candidiasis with fluconazole plus probiotic Lactobacillus
rhamnosus GR-1 and Lactobacillus reuteri RC-14. Lett Appl
Microbiol 48:269–274
16.
Prieto C (2016) Lactobacillus plantarum I1001 en la prevención de
la candidiasis vulvovaginal recurrente. Oral communication at the
Workshop SEPYP (Sociedad Española de Probióticos y
Prebióticos). Seville, Spain
17.
Atkinson MJ, Sinha A, Hass SL et al (2004) Validation of a general
measure of treatment satisfaction, the Treatment Satisfaction
Eur J Clin Microbiol Infect Dis. 

Scene 7

could be of relevance as a resource for women needing to
replenish their vaginal microbiota after taking ATB.
It has to be noted that, in our study, VVC relapse was
considered on the basis of any self-reported sign/symptom
suggesting a new episode. Despite the subjective nature of
these criteria, it could probably represent a more realistic and
practical criteria in daily practice, considering that the pure
presence of symptoms can entail a medical consultation and/
or use of over-the-counter medication. Beyond the clinical
impact of the probiotic, high rates of satisfaction, tolerability
and effectiveness were reported by both patients and
gynaecologists.
Our study has certain limitations that must be pointed out,
such as the previously commented lack of randomization and
the open-label nature of the trial, thus a placebo effect cannot
be ruled out. However, it is worth pointing out that the high
number of women fulfilling RVVC criteria suggests they may
be well aware of the symptoms of a true VVC episode. And
finally, the sample size available was limited, and drop-outs
during the study follow-up were not uncommon. Thus, addi-
tional studies should be conducted to evaluate the effective-
ness of L. plantarum I1001, using double blind randomized
designs and larger sample sizes. Also, it would be interesting
to evaluate the efficacy of L. plantarum I1001 as add-on or
follow-up therapy of other antifungal regimes, as well as for
preventing the recurrence of bacterial vaginosis.
Acknowledgments
The authors thank all the gynaecologists of the
Instituto Palacios de Salud y Medicina de la Mujer and the Instituto de
Medicina EGR Madrid who actively participated in the study and to
Dr. Cindy L. Larios from the Medical Department of Clever
Instruments S.L. (Barcelona) for the statistical analysis and editorial
assistance for the manuscript.
Compliance with ethical standards
Funding
Gynea by Kern Pharma funded the study.
Conflict of interest
J. Espadaler is a full-time employee of AB-Biotics
S.A; C. Prieto is a full-time employee of Gynea by Kern Pharma.
Ethical approval
All the study materials were reviewed and approved
by the Ethical Committee of the Hospital Universitario de la Princesa
(Madrid, Spain).
Informed consent
Informed written consent was obtained from all
participants prior to enrolment.
Open Access This article is distributed under the terms of the Creative
Commons Attribution 4.0 International License (http://
creativecommons.org/licenses/by/4.0/), which permits unrestricted use,
distribution, and reproduction in any medium, provided you give
appropriate credit to the original author(s) and the source, provide a link
to the Creative Commons license, and indicate if changes were made.
References
1.
Richter SS, Galask RP, Messer SA, Hollis RJ, Diekema DJ, Pfaller
MA (2005) Antifungal susceptibilities of Candida species causing
vulvovaginitis and epidemiology of recurrent cases. J Clin
Microbiol 43:2155–2162
2.
Tibaldi C, Cappello N, Latino MA et al (2009) Vaginal and
endocervical microorganisms in symptomatic and asymptomatic
non-pregnant females: risk factors and rates of occurrence. Clin
Microbiol Infect 15:670–679
3.
Peters BM, Yano J, Noverr MC, Fidel PL (2014) Candida vaginitis:
when opportunism knocks, the host responds. PLoS Pathog 10:
e1003965
4.
Murina F, Graziottin A, Felice R et al (2011) The recurrent
vulvovaginal candidiasis: proposal of a personalized therapeutic
protocol. ISRN Obstet Gynecol 2011:806065
5.
Marchaim D, Lemanek L, Bheemreddy S et al (2012) Fluconazole-
resistant Candida albicans vulvovaginitis. Obstet Gynecol 120:
1407–1414
6.
Sobel JD (2016) Recurrent vulvovaginal candidiasis. Am J Obstet
Gynecol 214:15–21
7.
Mendling W, Brasch J, Cornely OA, Effendy I, Friese K, Ginter-
Hanselmayer G et al (2015) Guideline: vulvovaginal candidosis
(AWMF 015/072). S2k (excluding chronic mucocutaneous
candidosis). Mycoses 58:1–15
8.
Dovnik A, Golle A, Novak D, Arko D, Takac I (2015) Treatment of
vulvovaginal candidiasis: a review of literature. Acta
Dermatovenerol Alp Pannonica Adriat 24:5–7
9.
Kovachev SM (2015) Local probiotic therapy for vaginal Candida
albicans infections. Probiotics Antimicrob Proteins 7:38–44
10.
Sobel JD, Zervos M, Reed BD, Hooton T, Soper D, Nyirjesy P et al
(2003) Fluconazole susceptibility of vaginal isolates obtained from
women with complicated Candida vaginitis: clinical implications.
Antimicrob Agents Chemother 47:34–38
11.
Food and Agriculture Organization of the United Nations and
World Health Organization. 2001, posting date. Regulatory and
clinical aspects of dairy probiotics. Food and Agriculture
Organization of the United Nations and World Health
Organization Expert Consultation Report. Food and Agriculture
Organization of the United Nations and World Health
Organization Working Group Report. http://www.who.
int/foodsafety/fs_management/en/probiotic_guidelines.pdf.
Accessed 30 June 2016
12.
Hill C, Guarner F, Reid G et al (2014) Expert consensus document.
The International Scientific Association for Probiotics and
Prebiotics consensus statement on the scope and appropriate use
of the term probiotic. Nat Rev Gastroenterol Hepatol 11:506–514
13.
Borges S, Silva J, Teixeira P (2014) The role of lactobacilli and
probiotics in maintaining vaginal health. Arch Gynecol Obstet
289:479–489
14.
O’Hanlon DE, Moench TR, Cone RA (2013) Vaginal pH and
microbiocidal lactic acid when lactobacilli dominate the microbio-
ta. PLoS One 8:e80074
15.
Martinez RC, Franceschini SA, Patta MC, Quintana SM, Candido
RC, Ferreira JC et al (2009) Improved treatment of vulvovaginal
candidiasis with fluconazole plus probiotic Lactobacillus
rhamnosus GR-1 and Lactobacillus reuteri RC-14. Lett Appl
Microbiol 48:269–274
16.
Prieto C (2016) Lactobacillus plantarum I1001 en la prevención de
la candidiasis vulvovaginal recurrente. Oral communication at the
Workshop SEPYP (Sociedad Española de Probióticos y
Prebióticos). Seville, Spain
17.
Atkinson MJ, Sinha A, Hass SL et al (2004) Validation of a general
measure of treatment satisfaction, the Treatment Satisfaction
Eur J Clin Microbiol Infect Dis

Questionnaire for Medication (TSQM), using a national panel study
of chronic disease. Health Qual Life Outcomes 2:12
18.
Witt A, Kaufmann U, Bitschnau M, Tempfer C, Ozbal A,
Haytouglu E et al (2009) Monthly itraconazole versus classic ho-
meopathy for the treatment of recurrent vulvovaginal candidiasis: a
randomised trial. BJOG 116:1499–1505
19.
Falagas ME, Betsi GI, Athanasiou S (2006) Probiotics for preven-
tion of recurrent vulvovaginal candidiasis: a review. J Antimicrob
Chemother 58:266–272
20.
Ehrstrom S, Daraczy K, Rylander E et al (2010) Lactic acid bacteria
colonization and clinical outcome after probiotic supplementation
in conventionally treated bacterial vaginosis and vulvovaginal can-
didiasis. Microbes Infect 12:691–699
21.
De Seta F, Parazzini F, De Leo R, Banco R, Maso GP, De
Santo D et al (2014) Lactobacillus plantarum P17630 for
preventing Candida vaginitis recurrence: a retrospective com-
parative study. Eur J Obstet Gynecol Reprod Biol 182:136–
139
22.
Pirotta M, Gunn J, Chondros P, Grover S, O'Malley P, Hurley S,
Garland S (2004) Effect of lactobacillus in preventing post-
antibiotic vulvovaginal candidiasis: a randomised controlled trial.
BMJ 329:548
23.
Spinillo A, Capuzzo E, Acciano S, De Santolo A, Zara F
(1999) Effect of antibiotic use on the prevalence of symp-
tomatic vulvovaginal candidiasis. Am J Obstet Gynecol
180:14–17
Eur J Clin Microbiol Infect Dis. 

Scene 8

Questionnaire for Medication (TSQM), using a national panel study
of chronic disease. Health Qual Life Outcomes 2:12
18.
Witt A, Kaufmann U, Bitschnau M, Tempfer C, Ozbal A,
Haytouglu E et al (2009) Monthly itraconazole versus classic ho-
meopathy for the treatment of recurrent vulvovaginal candidiasis: a
randomised trial. BJOG 116:1499–1505
19.
Falagas ME, Betsi GI, Athanasiou S (2006) Probiotics for preven-
tion of recurrent vulvovaginal candidiasis: a review. J Antimicrob
Chemother 58:266–272
20.
Ehrstrom S, Daraczy K, Rylander E et al (2010) Lactic acid bacteria
colonization and clinical outcome after probiotic supplementation
in conventionally treated bacterial vaginosis and vulvovaginal can-
didiasis. Microbes Infect 12:691–699
21.
De Seta F, Parazzini F, De Leo R, Banco R, Maso GP, De
Santo D et al (2014) Lactobacillus plantarum P17630 for
preventing Candida vaginitis recurrence: a retrospective com-
parative study. Eur J Obstet Gynecol Reprod Biol 182:136–
139
22.
Pirotta M, Gunn J, Chondros P, Grover S, O'Malley P, Hurley S,
Garland S (2004) Effect of lactobacillus in preventing post-
antibiotic vulvovaginal candidiasis: a randomised controlled trial.
BMJ 329:548
23.
Spinillo A, Capuzzo E, Acciano S, De Santolo A, Zara F
(1999) Effect of antibiotic use on the prevalence of symp-
tomatic vulvovaginal candidiasis. Am J Obstet Gynecol
180:14–17
Eur J Clin Microbiol Infect Dis

Is it possible to prevent recurrent vulvovaginitis? The role of Lactobacillus plantarum I1001 (CECT7504)