Original article

The serum levels of calcium, magnesium, iron and zinc in patients

with recurrent vulvovaginal candidosis during attack, remission
and in healthy controls

Jiri Spacek,1 Petr Jilek,2 Vladimir Buchta,2,3 Miroslav Fo}rstl,3 Miloslav Hronek2 and Magdalena
Holeckova4
1
Department of Obstetrics and Gynecology, University Hospital, 2Department of Biological and Medical sciences, Faculty of Pharmacy, 3Department of Clinical
Microbiology, University Hospital, and 4Department of Clinical Chemistry and Diagnostics, University Hospital, Hradec Kralove, Charles University, Prague,
Czech Republic

Summary The real cause of recurrent vulvovaginal candidosis (RVVC) is concealed and the
etiopathogenesis of this disease remains to be determined. In a cohort study,
concentrations of metals in 44 patients with RVVC and 30 healthy age-matched
women were measured and compared. The concentrations of serum calcium (Ca),
magnesium (Mg) and iron (Fe) were measured photometrically, the zinc (Zn) levels
were determined using flame atomic absorption spectrometry. For statistical analysis
were used the Student’s t-tests (paired analysis for attack vs. remission; non-paired
analysis for patient vs. control). Although all measured metals were within normal
ranges the patients with RVVC had in contrast to the healthy controls significantly
lower levels of serum Ca, Mg and Zn and insignificantly higher levels of Fe. These
relative changes may contribute to the development of attacks in patients with RVVC.

Key words: recurrent vulvovaginal candidosis, attack, remission, metals.

episodes of RVVC are largely manifested as a vulvovaginal

Introduction
Vulvovaginal candidosis (VVC) is considered recurrent
(RVVC) when at least four episodes occur in one year or,
at least, three episodes unrelated to antibiotic therapy
occur during the last 12 months.1 The acute sporadic
form of VVC affects approximately 75% of all women. Up
to 5% of women with a primary sporadic episode of VVC
will subsequently develop RVVC.2 Majority of the RVVC
patients relapses 1 month after antimycotic treatment
and, in fact, almost all relapse within 3 months.3
Unfortunately, the real reason of this disease is unknown
and the ethiopathogenesis of RVVC is still not resolved.
The acute sporadic VVC can be considered as a local
inflammation of yeast origin, while the individual
Correspondence: Jiri Spacek, Department of Obstetrics and Gynecology,
University Hospital, Sokolska 581, Hradec Kralove, 500 05, Czech Republic.
Tel.: +420495834341. Fax: +420495832032.
E-mail: spacekj@fnhk.cz
Accepted for publication 27 July 2005
discomfort with minimal inflammatory changes. There is
frequently striking discrepancy between severe symp-
toms and signs which are often minimal or absent in
culture positive attacks in RVVC patients.3 While the
presence of the yeast in RVVC patients usually coincides
with the symptoms, the finding of Candida in vagina of
healthy women often represents asymptomatic coloniza-
tion as well. There is a number of potential causes of VVC.
Hormonal dependence is generally accepted and VVC
occurs almost exclusively in the childbearing period,
while premenarcheal girls and postmenopausal women
rarely suffer from it. Other factors to be taken into
consideration, include the disturbance of host-microbes
relationships or the alteration of mucosal immunological
mechanisms of vaginal compartment which is relatively
independent of the immune reactions mediated by blood
or lymph cells. RVVC is believed to be a disease charac-
terized by recurrent episodes due to immunological
vaginal problems.4
The main problem with the investigation of RVVC in
most patients is the inability to find and define any

2005 Blackwell Publishing Ltd • Mycoses, 48, 391–395 391

Jiri Spacek et al.

predisposing condition. However there is no doubt, that
RVVC attack represents a certain form of the impair-
ment of the delicate balance between the host and the
commensal yeast in the vagina. The question is, which
of these conditions can be considered as trigger factors
and which as underlying ones. Among the latter are
hereditary predispositions, hormonal status, latent
(local) immunodeficiency, distress etc.
This study deals with pathophysiological aspects of
RVVC with focus on the serum levels of several metals
[calcium (Ca), magnesium (Mg), iron (Fe) and zinc (Zn)].
The reports about this are lacking or provide contra-
dictory data.5,6
Methods
Patients
This study involved 44 patients aged between 17 and
51 (mean 31.7). All women were attending the special
outpatient department of the gynecological clinic. The
basic inclusion criterion was the presence of four
culture-documented attacks of VVC during the previous
12 months. The patients were followed up during attack
and remission periods. Assessment included a detailed
history, gynecological examination including visualiza-
tion of the vulva, introitus and vagina and determin-
ation of pH in the vagina. Microscopic examination of a
vaginal smear (native and KOH mounting) was per-
formed in all cases.
Control group was consisted of 30 healthy women
with no history of VVC and the negative findings of
Candida in culture during investigation of the levels of
Ca, Mg, Fe and Zn. The age structure of control group
was similar to patients (range 21–44; mean 33.7).
There was no pregnant woman and no patient with
proven immunoalteration in this study.
The blood samples were taken in RVVC patients
twice. During culture positive attacks and remission. In
remission in RVVC patients and in healthy controls
between 2lst and 23rd day of the cycle, during culture
positive attack in these patients regardless of the cycle.
The blood samples were processed the same day with no
delay.
The serum levels of Ca, Mg and Fe were measured
photometrically after reaction with o-cresolphthalein,
xylidin blue and ferrozine, respectively using Hita-
chi 917 (Roche Cambridge, UK), zinc was determined
using flame atomic absorption spectrometry Solaar 959
(Unicam Mannheim, Germany).
The results of patients and controls were statistically
evaluated. For statistical analysis, the Student’s t-tests
were used (paired analysis for attack vs. remission; non-
paired analysis for patient vs. control).
Results
All patients with RVVC had the primary idiopathic form
with no known causes. As shown in the Table 1, there
was a statistical significant difference (P < 0.001) in the
levels of Ca in RVVC women both in attacks (mean
value ¼ 2.21 mmol l)1) and in remissions (mean ¼
2.23 mmol l)1) comparing controls (2.38 mmol l)1).
No significant difference between the Ca levels during
attacks and remissions (paired t-test) were found. All
average levels were within reference intervals (2.17–
2.65 mmol l)1), nevertheless the Ca values of RVVC
patients were close to the lower limit of the standard
while the mean value of the control group corresponded
with the mid point of the standard values.

Table 1 The levels of serum zinc,

Value Significance magnesium, calcium, and iron in patients
with recurrent vulvovaginal candidosis
Metal Group (n) Mean SD A–R A–C R–C
during attack (regardless of the cycle)
Zinc (lmol l)1) Attack (44) 13.25 2.69 n.s. P £ 0.001 P £ 0.001 and remission in comparison with healthy
Remission (44) 13.16 2.28 controls (both subgroup between 2lst
Control (30) 17.52 5.62 and 23rd day of the cycle).
Magnesium (mmol l)1) Attack (44) 0.79 0.14 P £ 0.01 P £ 0.001 P £ 0.001
Remission (44) 0.82 0.14
Control (30) 0.87 0.17
Calcium (mmol l)1) Attack (44) 2.21 0.35 n.s. P £ 0.001 P £ 0.001
Remission (44) 2.23 0.36
Control (30) 2.38 0.47
Iron (lmol l)1) Attack (44) 17.53 8.74 n.s. n.s. n.s.
Remission (44) 17.22 7.78
Control (30) 13.68 5.14

A, attack; R, remission; C, Control; SD, standard deviation; n.s., statistically non-significant.

392
2005 Blackwell Publishing Ltd • Mycoses, 48, 391–395


Relative level (%)
Figure 1 The relative levels of serum zinc,
magnesium, calcium, and iron in patients
with recurrent vulvovaginal candidosis
during attack and remission in compar-
ison with healthy controls. (mean control
level ¼ 100%).
Similarly, there were significantly lower levels
of Mg (P < 0.001) in patients during attacks (mean ¼
0.79 mmol l)1) and remissions (0.82 mmol l)1) than in
controls (0.87 mmol l)1). The same was true if the Mg
levels were compared in attacks and in remissions (paired
t-test, Table 1). Despite the differences, all average levels
of Mg were within the normal range relevant for age/sex-
matched controls.
The levels of total Fe were insignificantly elevated in
both the attack group (mean value ¼ 17.53 lmol l)1)
and the remission group (17.22 lmol l)1) in compar-
ison with controls (13.68 lmol l)1).
The levels of zinc in plasma obtained both during
attack (13.25 lmol l)1) and remission (13.16 lmol l)1)
were significantly (P < 0.001) lower than values in
control group (17.52 lmol l)1). There was no signifi-
cant difference between the Zn levels in attacks and
remissions (paired t-test). All average levels fell into the
normal ranges (see Figure 1).
Discussion
While the levels of all metals were within the normal
range, the differences were evidenced from group to
group. The levels of serum Zn, Mg and Ca were in RVVC
patients significantly lowered, the levels of Fe showed in
this group a tendency towards an increased level
(insignificantly). We question whether these results
relate to the development of RVVC attack. The recurrent
form of vulvovaginal candidiasis appears to have many
factors, which participate in its etiopathogenesis inclu-
ding yeast virulence and patient resistance. The question
is which of the factors are responsible for the establish-

2005 Blackwell Publishing Ltd • Mycoses, 48, 391–395
Recurrent vulvovaginal candidiasis
Attack (n = 44)
Remission (n = 44)
200 Control (n = 30)
P ≤ 0.001 P ≤ 0.001
P ≤ 0.001
P ≤ 0.001 P ≤ 0.001
150
P ≤ 0.001
P ≤ 0.01
100
50
0
Zn Mg Ca Fe
Mean control level = 100%
ment of an individual episode and which of them act as
long-term predisposing conditions involved in the devel-
opment of RVVC. Several factors have been suggested to
be responsible for the recurrent attacks such as Ôhigh-
riskÕ behaviour,7, 8 transient weakening of host defences
including reduced vaginal immunity, hormonal imbal-
ance, abnormal vaginal bacterial flora and increased
number of yeast and/or their virulence.2, 9
There is a lack of information about a possible
association between the levels of some elements and
RVVC. For example there is no published data available
regarding hypomagnesaemia, hypocalcaemia and yeast
infections. In RVVC patients could be the lower level of
Ca hypothetically related to the insufficient level of Ca in
food. Considering the fact in cases of RVVC, Ôyoghurt
therapyÕ is recommended,10, 11 the finding of calcaemia
at the lower limit of a norm is an additional reason why
dairy products with probiotic microflora should be
recommended to such patients. Both promotion probi-
otic vaginal microflora and Ca supplementation can be
achieved with regular yoghurt consumption although
the beneficial effect of these factors remains to be
supported with randomized controlled studies.
There are few reports on the role of Fe in candidiasis.
We found insignificantly elevated total Fe in both
subgroups in RVVC patients in comparison with healthy
controls. The data from Davidson’s study12 are partly in
contrast to results of our study and these authors
concluded that Fe played a protective role in resistance
against RVVC.12 The results of other studies concerning
a role of Fe in RVVC showed that this element was
important not only for pathogenic yeast, but also for
normal function of host immunity. The virulence of
393
Jiri Spacek et al.
Candida albicans is dependent on the level of Fe, as the
absence of this metal has been shown to reduced the
virulence of this strain.13 It is assumed that a sufficient
amount of the Fe e.g. during hypersideraemia can
support the development of candidosis – as shown
experimentally in mice.14, 15 A relative excess of Fe
maybe responsible for the yeast invasion into the
epithelium.16 The importance of Fe for yeast results
from a special mechanism of Fe intake from host’s
erythrocytes in Candida cells.17 On the contrary, there is
a protective effect of Fe binding substances on the
antimicrobial immunity including the resistance to the
Candida.18
Zinc represents one of the crutial elements of the
immune system19, 20 and its supplementation is con-
sidered as the significant immunomodulating interven-
tion.21, 22 The Zn deficiency has been originally
described in relation to the impairment of functions of
the T-lymphocytes including (premature) thymus invo-
lution.20 Zn is an essential component of thymus
hormone thymulin, which supports the maturation of
the T-lymphocytes both in thymus and in extrathymic
places. The T-lymphocytes in the vaginal mucosa
represent a population with a significant portion of
cells matured in extrathymic places,23 so the manifes-
tation of Zn deficiency cannot be excluded in this
location. Hypozincaemia has been described in patients
with RVVC,6 although others did not confirm this
finding.5 In addition, Zn is also necessary for the correct
function of an innate immunity (phagocytosis, NK
cells).24
We found a significant drop in the serum Zn levels in
RVVC patients (Table 1). Approximately 75% of values
were just above the lower limit of normal (Table 1).
These data supports the hypothesis that Zn plays an
important role in anti-Candida resistance. Another fact
to be taken into consideration is that Zn deficiency is a
typical finding in acrodermatitis enteropatica, which is
accompanied by Candida infections.25 In view of the
suggestions, RVVC can be considered a disease deter-
mined by subtle immunological changes following from
a relative hypozincaemia. The yeast–host interaction
takes place on the epithelial surface and is influenced by
the local defence mechanisms and the indigenous
micro-organisms. In addition, interactions with steroid
hormones (gestagens), vitamins and trace elements can
be expected. It is suggested that Zn is involved in both
epithelial integrity and resistance to invasion. It has
been postulated that hypozincaemia could contribute to
the epithelial vulnerability to infections in the respirat-
ory tract,26 periodontal tissues27 or genitourinary
tract.28 Zn supplementation improves the anti-infective
394
resistance including the anti-Candida immunity in
patients with secondary immunodeficiency.19,20 Polizzi
et al.22 described a case of a woman with chronic
mucocutaneous candidiasis who considerably improved
in terms of frequency of vaginal and rectal Candida
episodes and increase of CD4 cells counts after
16 months of Zn suplementation. Furthermore, Zn is a
potential modulator of the yeast virulence, because it
inhibits the transition of the blastic (yeast) form into the
more virulent hyphal morphology in vitro.29 Although
this effect of micromolar Zn concentrations on morpho-
logical transition and cell cycle changes of yeast was
observed only in lower temperature (25 C) and acidic
pH (<5.5),29 it is not excluded that hypozincaemia can
be one of the predisposing factors for RVVC attack.
Thus, Zn supplementation to restore physiological Zn
level could be offered to RVVC patients.
Acknowledgment
The study was supported by the research project IGA
No.3694–3 of the Czech Ministry of Health.
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