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Objective: This cross sectional clinical trial aimed at
exploring the association between chlamydia trachomatis infections and
peri-menopausal bleeding presenting at Al-Azhar University Maternity Hospital
outpatient clinics.
Patients and methods: This was a
cross-sectional study involving 150 women divided into two equal groups of 75
women in each group. Women were
allocated from the out-patient clinic of Al-Azhar Maternity University Hospital
and they were counseled and arranged for Pipelle endometrial samples. Women of
group I suffered from peri-menopausal bleeding while group II were presented at
the hospital due to any cause other than vaginal bleeding. Pipelle endometrial biopsy was taken and sent
for detection of Chlamydia trachomatis
by real time PCR.
Results: In group-I, 44 (58.7%) samples were
positive for Chlamydial infection while the other 31 (41.3%) samples were
negative for Chlamydia. In group II 17 (22.7%) samples were PCR positive. There
was an apparent significant difference between the two groups regarding the
prevalence of C. trachomatis among symptomatic peri-menopausal women suffering
from uterine bleeding.
Conclusion: It is advisable to screen for Chlamydia
trachomatis in women with peri-menopausal bleeding.
Keywords: Chlamydia
endometritis; Peri-menopausal
bleeding.
INTRODUCTION
Worldwide, C. trachomatis is considered the most prevalent
sexually transmitted bacterial disease. C. trachomatis is responsible for a
diverse spectrum of genital infections as cervicitis, endometritis and
salpingitis. Pelvic inflammatory disease (PID) is a major problem affecting the
female genital system, C. trachomatis is the main organism causing it.
Infertility in women and men, ectopic pregnancy and chronic pelvic pain,
epididymitis and proctitis and arthritis in both men and women are spectra and
common sequelae of this infection [1].
Pregnancy complications and implantation failure after in vitro fertilization also are reported as common sequels of these bacteria [2–5]. Young adolescent females having, multiple sexual partners with unprotected intercourse, are at highest risk to contract C. trachomatis. In about 70% of the cases, the progression of infection from the lower into the upper genital tract occurs without symptoms (subclinical PID) [1]. Reproduction is often delayed until a woman reaches her twenties, thus, early detection of C. trachomatis infection to prevent permanent tissue damage to the upper genital tract is critical. Histopathological and microbiological assessments of the endometrium have been useful in identifying cases of subclinical PID. Quantitative changes in endometrial histopathology and microbiology were observed as the disease progressed from the lower to the upper genital tract in these women [6–8].
The diagnosis of endometritis based
on the number of neutrophils in the glandular epithelium and plasma cells in
the endometrial stroma, however, continues to be problematic [9]. This is
likely due to different levels of experience of the observers in identification
of plasma cells and neutrophils.
The presence of chlamydial antigens
in the endometrium has also been linked to abnormal uterine bleeding (AUB) in
women on birth control pills [10]. Because so many young women present with
this condition, AUB is the most common clinical diagnosis for patients of
reproductive age who undergo minor gynecologic procedures. The condition is
initiated by either an infectious process (endometritis) or by structural
alterations of micro vessels [11]. Whether AUB is a risk factor for subclinical
PID remains unknown. In a proof of the concept study AUB improved after
antibiotic treatment, lending further support to an infectious etiology of this
condition.12 In spite of the extensive research of the endometrium in
subclinical PID and the suggestion for an association between AUB, plasma cell
endometritis and C. trachomatis, there remains an absence of markers for
identification of C. trachomatis infection of the uterine lining.
This cross-sectional study
utilizing endometrial samples aimed at confirmation of the possible relation
between the clinical diagnosis of AUB and C. trachomatis infection of the
endometrium.
PATIENTS AND METHODS
This was a cross-sectional study
performed in Al-Azhar Maternity University Hospital involving 150 women; group
I: 75 cases with peri-menopausal bleeding and a control group II: 75
peri-menopausal women with normal menstruation attending gynecologic outpatient
clinic, for any reason other than bleeding. Pipelle endometrial biopsies were
collected and sent for detection of Chlamydia trachomatis by PCR. This study was carried out in the period from
January 2014 to December 2015. Explanation of the procedure and verbal consent
was taken from every patient.
Inclusion criteria:
1.
Peri-menopausal females; age 40 – 50 years.
2.
Complain of dysfunctional uterine bleeding in group I
3.
No gross uterine lesions were detected by vaginal US.
Exclusion criteria:
1.
Patients who are immediately post-partum or post
abortion or known cases of sexually transmitted diseases
2. Patients
with any uterine abnormality detected by transvaginal sonar or hysteroscopy.
3. Patients
with suspicion of pregnancy or malignancy.
All patients were subjected to:
1) Full history taking and physical
examination.
2) Counseling and verbal consent
was taken from every patient.
3) Pipelle endometrial biopsies
were taken and sent to confirm Chlamydia trachomatis endometritis by
real time PCR. The endometrial biopsy was placed into a transport medium
composed of 2-sucrose phosphate buffer (PH 7.0) supplemented with 5% fetal
bovine serum, 50 ug of streptomycin / ml, 100 ug of vancomycin per ml and 12.5
ug of amphotericin B (Fungizone) per ml (Phosphate Buffer Saline).
Detection of the C. trachomatis
DNA in the collected specimens
A- Extraction of DNA: This
was performed using the QI Amp DNA mini kit (QIAGEN GmbH, Hilden, Germany Cat.
No.51304) as described by the manufacturer.
B- Real-time PCR assay:
Each run included the testing of
the positive control, Tris-EDTA buffer in four reactions (no-template
controls), and diethyl-pyrocarbonate-treated water (QIAGEN Germany) in
duplicate reactions (negative reagent controls). The no-template controls and
negative reagent controls were used to detect any nonspecific fluorescent
signal or carry-over contamination. Run acceptability required obtaining the
expected results from each control. Samples were considered positive if the
amplification plots (i.e., change in normalized reporter signal versus PCR cycle
number) showed definite exponential increase in fluorescent signal. If the
fluorescent signal did not increase within 45 cycles, the sample was considered
negative.
Statistical methodology:
Retrieved data were recorded on an investigative report form. The data were analyzed with SPSS® for
Windows®, version 15.0 (SPSS, Inc, USA). Description of quantitative
(numerical) variables was performed in form of mean, standard deviation (SD)
and range. Description of qualitative (categorical) data was performed in the
form of numbers and percent. Analysis of numerical variables was performed by
using student’s unpaired t-test (for two groups) or ANOVA (for more than two
groups). Analysis of categorical data was performed by using Fischer’s exact
test and Chi-squared test. Significance level was set at 0.05.
RESULTS
This cross-sectional study involved 150 women
consented to participate in this study; group I (test group): 75 cases with
peri-menopausal bleeding and group II (control group): 75 peri-menopausal women
with normal menstruation recruited from outpatient clinic, and complaining from
any reason other than bleeding. Pipelle endometrial biopsies were collected and
sent for detection of C. trachomatis by real time PCR. Both groups were comparable in terms of age,
body mass index, gravidity, parity, duration of marriage, frequency of coitus
per week, mode of delivery (vaginal or cesarean), level of education,
occupation (house wife or employed/business woman) and previous use of IUCD or
hormonal methods as shown in Table 1.
In group I, 44 (58.7%) specimens
were positive for Chlamydia and the other 31 (41.3%) specimens were negative
for Chlamydia, while in group II only 17 (22.7%) specimens were PCR positive
for chlamydia. There was a significant difference between the two groups about
the prevalence of C. trachomatis among symptomatic peri-menopausal women
complaining of abnormal uterine bleeding (Table
2).
There was no statistically
significant correlation (P value > 0.05) between the presence of chlamydial
endometritis in peri-menopausal women and the age of participants, parity, form
of bleeding, history of abortion, type of infertility and duration of marriage
(Table 3-8).
DISCUSSION
AUB and C. trachomatis infection of
the endometrium have a strong link as shown by the results of the current
study. When plasma cells were absent, AUB alone lost its diagnostic value,
while plasma cells alone without peri-menopausal bleeding remained predictive.
This is possibly due to many factors linked to the diagnosis of bleeding.
Histochemical staining with Syndecan-1 increases the plasma cells thus
assisting in the diagnosis of endometritis. Syndecan-1 staining increases the
identification rate of plasma cells between 10-50%, and is very useful to the
less experienced pathologist allowing for detection of many plasma cells that
are lacking the characters (clock-face chromatin, eccentrically placed nucleus
and peri-nuclear halo) [12-14].
In the current study we found that
Pipelle endometrial biopsies were collected and sent for detection of C.
trachomatis by real time PCR. Both
groups were comparable in terms of age, body mass index, gravidity, parity,
duration of marriage, frequency of coitus per week, mode of delivery (vaginal
or cesarean), level of education, occupation (house wife or employed/business
woman) and previous use of IUCD or hormonal methods.
In group I, 44 (58.7%) specimens
were positive for Chlamydia and the other 31 (41.3%) specimens were negative
for Chlamydia, while in group II only 17 (22.7%) specimens were PCR positive
for chlamydia. There was a significant difference between the two groups about
the prevalence of C. trachomatis among symptomatic peri-menopausal women
complaining of abnormal uterine bleeding.
There was no statistically
significant correlation (P value > 0.05) between the presence of chlamydial
endometritis in peri-menopausal women and the age of participants, parity, form
of bleeding, history of abortion, type of infertility and duration of marriage.
In this group of patients, the
presence of plasma cells correlated well with a positive C. trachomatis result
but only at a higher number (>5 PCE) than expected. The presence of ‘any
number of plasma cells’ was a much weaker predictor.
The number of C. trachomatis
positive cases in this group of patients is quite high (48%). Selection bias
may play a role. However, similarly high rates (38%) had been supported by
Kiviat et al. in a group of women with suspected subclinical PID. Using
species-specific monoclonal antibodies, these investigators also found C.
trachomatis in 18% of culture negative specimens [15]. As it was stated before,
close to one-third of our patients had at least previous evidence of a previous
infection. Immunocytochemistry methods employing monoclonal antibodies on
tissue samples appear to be superior to culture and/or to amplification assays.
Antigen detection by monoclonal antibody in women who are culture or PCR
negative likely represents a subclinical persistent infection that may have
been undetected or inadequately treated [16,17]. Recent data confirmed that
presumed recurrent infections by the same serovar of Chlamydia are indeed due
to persistence as evidenced by outer membrane protein (Omp1) genotyping [18].
We believe that this study has
identified a group of women with persistent C. trachomatis infection. These are
relatively older women dealing with the late consequences of an earlier, missed
or incompletely treated chlamydial infection.
The strong correlation between
endometrial macrophage count and C. trachomatis infection is a novel finding.
Macrophages have been described as the missing link in the pathogenesis of C.
trachomatis induced (previously ‘reactive’) arthritis and are considered the
vehicle by which C. trachomatis arrives to the synovial tissue from its point
of entry [19,20]. Chlamydia trachomatis infected macrophages can significantly
induce the apoptosis of autologous T-cell lymphocytes, the most important
defense line in C. trachomatis infection, allowing the bacterium to subvert the
immune system and develop the persistent state [20].
The major limitations of this
retrospective study are the lack of cervical/vaginal microbiology specimens and
serum samples to confirm chlamydial infection. It is feasible that many of our
study patients had had a negative lower genital tract C. trachomatis test.
Based on the data we suggest that the effect of
C. trachomatis on women of reproductive age is overwhelmingly underestimated.
An asymptomatic, preconceptional, persistent infection may lead to pregnancy
complications in reproductive aged women. AUB in women of reproductive age
should raise the suspicion of a persistent C. trachomatis infection.
Endometrial specimens from these women should be examined for the presence of
PCEs. Our data showed that Syndecan-1 could enhance PCE diagnosis in a clinical
setting and that an elevated macrophage count might indicate the presence of C.
trachomatis in the endometrium in a research setting. Large, prospective
studies are needed to confirm a cause-effect relationship between C.
trachomatis and AUB.
- Tavakoli
M, Craig AM, Malek M (2002) An economic analysis of screening and
treatment of patients with suspected Chlamydia. Health Care Manag Sci; 5:33–39.
- Sulak PJ 2003 Sexually transmitted
diseases. Pediatr and Adolescent Gynecol 21:399–414.
- Romero R,
Espinoza J, Mazor M (2004) Can endometrial infection/inflammation explain
implantation failure, spontaneous abortion and preterm birth after in
vitro fertilization? Fertil Steril 82:799–804.
- Toth M,
Witkin SS, Ledger W, Thaler H (1987) The role of infection in the etiology
of preterm birth. Obstet Gynecol 71:723–726.
- Toth M,
Chaundray A, Witkin S (1993) Pregnancy outcome following pelvic infection.
Infect Dis Obstet Gynecol 1:12–15.
- Wiesenfeld
HC, Hillier SL, Krohn MA, Amortegui AJ, Heine RP (2002) Lower genital
tract infection and endometritis: insight into subclinical pelvic
inflammatory disease. Obstet Gynecol 100:456–463.
- Kiviat
NB, Wolner-Hanssen P, Eschenbach DA, Wasserheit JN, Paavonen JA, et al.
(1990) Endometrial histopathology in patients with culture-proved upper
genital tract infection and laparoscopically diagnosed salpingitis. Am J
Surg Pathol 14:167–175.
- Eckert
LO, Hawes SE, Wolner-Hanssen PK, Kiviat NB, Wasserheit JN, et al. (2002)
Endometritis: the clinico-pathologic syndrome. AM J Obstet Gynecol
186:690–695.
- Achilles
SL, Amortegui AJ, Wiesenfeld HC (2005) Endometrial plasma cells: Do they
indicate subclinical pelvic inflammatory disease? Sexually Transmitted
Diseases 32:185–188.
- Krettek
JE, Arkin SI, Chaisilwattana P, Monif Gr (1993) Chlamydia trachomatis in
patients who used oral contraceptives and had intermenstrual spotting.
Obstet Gynecol 81:728–731.
- Ferenczy
A (2003) Pathophysiology of endometrial bleeding. Maturitas 45:12–14.
- Eckert
LO, Watts DH, Thwin SS, Hillier SL, Kiviat NB, et al. (2004) Eschenbach
DA: The antimicrobial treatment of subacute endometritis: a proof of the
concept study. A Obstet Gynecol 190:305–313.
- Hsu SM,
Raine SI, Ranger H (1981) A comparative study of the
peroxidase–antiperoxidase method and an avidin–biotin complex method for
studying polypeptide hormones with radioimmunoassay antibodies. Am J Clin
Pathol 75:734–738.
- Bayer-Garner
IB, Nickell JA, Korourian S (2004) Routine Syndecan-1 immunohistochemistry
aids in the diagnosis of chronic endometritis. Arch Pathol Lab Med
128:1000–1003.
- Kiviat
NB, Wolner-Hanssen P, Peterson M, Wasserheit J, Stamm WE, et al. (1986)
Localization of Chlamydia trachomatis infection by direct
immunofluorescence and culture in pelvic inflammatory disease. Am J Obstet
Gynecol 154:865–873.
- Toth M,
Patton DL, Campbell LA, Carretta EI, Mouradian J, et al. (2000) Detection
of chlamydial antigenic material in ovarian, prostatic, ectopic pregnancy
and semen samples of culture-negative subjects. Am J Reprod Immunol
43:218–222.
- Patton
DL, Askienazy-Elbhar M, Henry-Suchet J (1994) Detection of Chlamydia
trachomatis in fallopian tube tissue in women with post infectious tubal
infertility. Am J Obstet Gynecol 171:95–101.
- Dean D,
Suchland RJ, Stamm W (2000) Evidence for long-term cervical persistence of
Chlamydia trachomatis by omp1 genotyping. J Infect Dis 182:909–916.
- Zeidler
H, Kuipers J, Kohler L (2004) Chlamydia-induced arthritis. Curr Opin
Rheumatol 6:380–392.
- Jendro
MC, Deutsch T, Korber B, Kohler L, Krausse-Opatz JGB et al. (2000)
Infection of human monocyte-derived macrophages with Chlamydia trachomatis
induces apoptosis of T cells: a potential mechanism for persistent
infection. Infect Immun 68:6704–6711.
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