Objective: The objective of this study was to observe pregnancy 
            outcomes in mice infected transvaginally with Chlamydia trachomatis

Bishop, Karen  D.

Blanco, Jorge D.

Graham, Jack M.

Oshiro, Bryan T.

Seraj, Ibrahim M.

English

PubMed

Bryan T. Oshiro

Jack M. Graham

Jorge D. Blanco

Ibrahim M. Seraj

Karen D. Bishop

Crossref

Infectious Diseases in Obstetrics and Gynecology 1:242-245 (1994)
(C) 1994 Wiley-Liss, Inc.
Pregnancy Outcome in Swiss-Webster Mice Infected
With Chlamydia trachomatis
Bryan T. Oshiro, Jack M. Graham, Jorge D. Blanco,
Ibrahim M. Seraj, and Karen D. Bishop
Department of Obstetrics, Gynecology, and Reproductive Sciences, University of Texas Health Science
Center, Houston, TX (B.T.O., J.M.G., J.D.B., K.D.B.), and Department of Obstetrics and
Gynecology, Loma Linda University, Loma Linda, CA (I.M.S.)
ABSTRACT
Objective: The objective of this study was to observe pregnancy outcomes in mice infected transvag-
inally with Chlamydia trachomatis.
Methods: Pregnant mice were inoculated transvaginally with either C. trachomatis (CT) or sterile
calf serum (CON) on pregnancy day 4. Pregnancy outcomes as well as genital tract histology and
culture were compared. Statistical analysis was performed using Fisher’s exact test and Student’s
t-test.
Results: Twenty-four of 26 CT mice had positive uterine cultures for C. trachomatis. Inflamma-
tion occurred in 9 (34.6%) (P 0.002, 95% confidence interval 1.7-3.5) and intrauterine fetal
demise occurred in 5 (19.2%) (P 0.05, 95% confidence interval 1.6-2.9) ofCT mice. No mice in
the CON group (0/24) had positive uterine cultures, developed inflammation, or experienced intrau-
terine fetal demise.
Conclusions: Lower genital tract chlamydial infection is associated with intrauterine fetal demise
in Swiss-Webster mice. (C) 1994 Wiley-Liss, Inc.
KEY WORDS
Chlamydia trachomatis, fetal demise, Swiss-Webster mice
hlamydia trachomatis is recognized as the most
common sexually transmitted organism, with
over 4 million cases reported annually. C. tra-
chomatis is responsible for causing inclusion con-
junctivitis and pneumonia in the neonate and is the
leading cause of preventable blindness in the
world. 2
Although C. trachomatis has clearly been shown
to cause mucopurulent cervicitis, 3 acute salpingi-
tis, 4 and neonatal disease, its effect on pregnancy
outcome remains controversial. Martin and col-
leagues5 reported that the incidence of" prematurity
and perinatal death was higher in women with a C.
trachomatis infection identified before 20 weeks. In
contrast, other authors6’7 have not found an associ-
ation between C. trachomatis infection and adverse
pregnancy outcome.
The Swiss-Webster mouse has been used exten-
sively to study the effects of C. trachomatis on the
female genital tract. 8-1 Rank et al. 11 have re-
ported infecting mice transvaginally with C. tra-
chomatis. However, to our knowledge, an animal
model has not been developed to study the effects of
C. trachomatis lower genital tract infection on preg-
nancy outcome.
To determine whether adverse pregnancy out-
come is associated with C. trachomatis genital tract
infection in an animal model, we studied the effect
of lower genital tract C. trachomatis infection in
pregnant Swiss-Webster mice.
Address correspondence/reprint requests to Dr. Bryan T. Oshiro, Department ofPerinatology, McKay-Dee Hospital Center,
3939 Harrison Boulevard, Ogden, UT 84409.
Received November 15, 1993
Basic Science Article Accepted March II, 1994
PREGNANCY OUTCOME IN MICE INFECTED WITH CT OSHIRO ET AL.
SUBJECTS AND METHODS
Mice
Seventy-six 6-8-week-old female Swiss-Webster
mice (Sasco, Inc., Omaha, NE)with unlimited
access to food and water were placed 2 per cage with
a male of established fertility over a 24-h period.
The mice were then separated into individual cages.
Pregnancy was confirmed by the presence of a cop-
ulation plug and later by palpation of a pregnant
uterus. Twenty-six non-pregnant mice were re-
moved from the study. This experiment was ap-
proved by the Animal Welfare Committee of the
University ofTexas Health Science Center at Hous-
ton, and all practices were in accordance with this
committee’s guidelines.
Chlamydia
The mouse pneumonitis strain of C. trachomatis
ATCC BR-123 was reconstituted to a concentration
of approximately 106 inclusion-forming
units/ml in sterile calf serum as described by Blanco
et al. 10
Experimental Design
The female mice were separated into 2 groups: a
group for chlamydial (CT) inoculation and a con-
trol group (CON). The inoculation technique used
in this experiment has been reported by Rank et
al. 1 On pregnancy day 4, the CT group was inoc-
ulated transvaginally with chlamydia suspended in
50 I1 of sterile calf serum. The CON group was
similarly inoculated with 50 I1 of sterile calf se-
rum. On pregnancy day 7, lower genital tract cul-
tures were obtained on all mice using a dacron-
tipped aluminum swab which was placed in
chlamydia transport media and immediately frozen
at -70C. The mice were observed daily for signs
of clinical infection or preterm delivery. The mice
were sacrificed and a laparotomy was performed on
the day of delivery. If delivery did not occur by the
normal date of confinement (day 21), the observa-
tion period was extended to pregnancy day 24 at
which time all undelivered mice were sacrificed
and explored. The abdominal cavity and reproduc-
tive tract were grossly examined for evidence of
inflammation and the uterus was removed. The
distal most portion of the uterus was transected and
placed in chlamydia transport media and frozen at
-70C. The remainder of the uterus was placed in
10% formalin. The pups were counted and
weighed.
Culture
The lower genital tract culture swab specimens were
allowed to thaw at room temperature and vortexed
for 15 min. The uterine tissue was also thawed,
then morcelated and vortexed. The supernatants
were removed and placed on McCoy cells which
were previously washed with Mg2+, Ca2+, and
phosphate-buffered saline. The cells were then cen-
trifuged for h at 3,400g and incubated at 37C
for 48 h. A second-pass culture was performed on
all samples. The culture technique is standard and
has been described elsewhere. 2 The cells were
stained with a chlamydia-specific fluorescent anti-
body stain (Bartels Diagnostic Division, McGaw
Park, IL) and examined by indirect fluorescent
microscopy in a blinded fashion.
Histology
The formalin-fixed tissue was placed in paraffin
blocks, sliced, stained with hematoxylin and eosin,
and mounted on glass slides. The examiner was
blinded as to whether the specimens originated from
CT or CON animals. The tissue was classified as
inflamed if plasma cells or a polymorphonuclear
infiltrate were present on examination of the slides.
Statistical Analysis
Statistical analysis was performed by Fisher’s exact
test and Student’s t-test where appropriate.
P < 0.05 was considered statistically significant.
RESULTS
Twenty-six mice in the CT group and 24 mice in
the CON group became pregnant and were in-
cluded in the study. All 26 (100%) of the CT mice
had positive lower genital tract swab cultures for
chlamydia on day 7. Uterine tissue cultures were
positive in 24/26 (92.3%) of CT mice. Inflamma-
tion of the uterine tissues and intrauterine fetal
death were significantly increased in the CT mice
(Table 1). In the 5 pregnancies in which fetal death
occurred, all of the pups in the litters were dead.
Placental histology could not be determined in each
pregnancy affected by intrauterine fetal demise be-
cause of the various stages of resorption of the
pregnancy products, but extensive inflammation
INFECTIOUS DISEASES IN OBSTETRICS AND GYNECOLOGY 243
PREGNANCY OUTCOME IN MICE INFECTED WITH CT OSHIRO ET AL.
TABLE I. Adverse pregnancy outcome data in mice
infected with C. trachomatis
95%
CT CON Relative Confidence
(n- 26) (n 24) P risk interval
Inflammation 9 (35%) 0 (0%) 0.002 2.4 1.7-3.5
Fetal deaths 5 (19%) 0 (0%) 0.05 2.2 1.6-2.9
TABLE 2. Delivery data in mice with liveborn pupsa
CT CON
(n 21) (n= 24) Significance
No. live pup-mouse 9.0 -+ 1.7 8.9 -+ 2.2 NS
Pup weight (g) 1.59 0.24 1.59 -+ 0.15 NS
Pregnancy days 21.6 -+ I.I 21.2 -+ 1.6 NS
aValues are mean --+ SD. NS not significant.
was observed in the placentas that were examined.
In 4 of the pregnancies in which fetal death oc-
curred, the gestational sacs appeared to be intact.
Because of advanced resorption of pregnancy prod-
ucts in the 5th pregnancy, the status of the sacs
could not be determined. There were no instances
of preterm delivery in the CT group, and none of
the mice with intrauterine fetal death delivered their
pups by the end of the normal confinement period.
The fetal deaths occurred only in the group of mice
with uterine inflammation. None of the mice in the
CON group had positive cultures, evidence of an
inflammatory reaction in uterine tissues, or intrau-
terine fetal deaths. Table 2 shows the pregnancy
outcome data in mice with liveborn pups. No sig-
nificant difference was found in the mean duration
of pregnancy, the mean number of live pups born
per mouse, or the mean weight ofthe liveborn pups
between the CT and the CON groups if fetal death
did not occur.
DISCUSSION
The role of C. trachomatis in adverse pregnancy
outcomes is controversial. In humans, several stud-
ies have not shown an association between C. tra-
chomatis infection and adverse pregnancy out-
comes. 6’7’13-18 However, others have implicated
C. trachomatis in premature rupture of the mem-
branes, preterm birth, low birth weight, intraam-
niotic infection, and stillbirth, s’ 19-24
One of the first studies to suggest that C. tra-
chomatis was associated with adverse pregnancy out-
comes was performed by Martin and colleagues5 in
1982. In their study, 18 of 268 (6.7%) pregnant
women screened before 20 weeks gestation were
positive for C. trachomatis. Twenty-eight percent
of the chlamydia-positive patients delivered prema-
turely compared with 6% of the chlamydia-nega-
tive patients. Most significantly, the perinatal mor-
tality in the infected group was 33% compared with
0.4% in the uninfected group. Gravett et al. 19
found that chlamydial infection of the cervix was
associated with low birth weight, premature rup-
ture of the membranes, and preterm birth. Martius
et al.2 and The Johns Hopkins Study Group for
Cervicitis and Adverse Pregnancy Outcomes21 also
found that cervical infection with C. trachomatis
was significantly associated with preterm birth.
Chlamydial infections of the cervix may ascend
and infect the membranes and fetus. Thorp et al. 22
reported a case of fetal death attributed to an in-
traamniotic chlamydial infection across intact mem-
branes. Although chlamydia was never cultured
from the fetus, the formalin-fixed lung tissue was
highly positive for fluorescent antibody against
chlamydia. Other authors23’24 have also reported
in utero chlamydial infection across intact mem-
branes.
In this study, we found that lower genital tract
infection with chlamydia caused infection ofuterine
tissues across intact membranes and furthermore
was significantly associated with intrauterine de-
mise in Swiss-Webster mice. We also found that,
in contradistinction to humans, intrauterine or in-
traamniotic chlamydial infection in mice did not
result in premature rupture of the membranes or
preterm delivery. Rather, the affected pregnancies
were retained past their normal term of gestation.
With the exception of the study by Martin and
colleagues, s human studies have not found an in-
crease in stillbirth associated with chlamydial infec-
tion in pregnancy. Many other studies looking at
the effect of C. trachomati in pregnant women
have had small sample sizes and did not specifically
look at stillbirths as an outcome variable. We have
shown that C. trachomati is capable of causing
intrauterine fetal death in Swiss-Webster mice.
These results may not correlate to adverse preg-
nancy outcomes in humans, but it would be inter-
esting to determine if C. trachomatis is a significant
244 INFECTIOU& DISEASES IN OBSTETRICS AND GYNECOLOGY
PREGNANCY OUTCOME IN MICE INFECTED WITH CT OSHIRO ET AL.
cause of stillbirth in human pregnancy as we have
demonstrated in the Swiss-Webster mice.
REFERENCES
1. Centers for Disease Control: Chlamydia trachomatis infec-
tions: Policy guidelines for prevention and control.
MMWR 34(Suppl 3):53s-74s, 1985.
2. Schachter J, Hanna L, Hill EC, Massad S, Sheppard
CW, Conte JE, Meyer KF: Are chlamydial infections the
most prevalent venereal disease? JAMA 231:1252-1255,
1975.
3. Brunham RC, Paavonen J, Stevens CE, et al.: Mucopu-
rulent cervicitis: The ignored counterpart in women of
urethritis in men. N Engl J Med 311:1-6, 1984.
4. Sweet RL: Chlamydial salpingitis and infertility. Fertil
Steril 38:530-533, 1982.
5. Martin DH, Koutsky K, Eschenbach DA, et al.: Prema-
turity and perinatal mortality in pregnancies complicated
by maternal Chlamydia trachomat# infections. JAMA 247:
1585-1588, 1982.
6. Harrison HR, Alexander ER, Weinstein L, et al.: Cer-
vical Chlamydia trachomatis and mycoplasmal infections
in pregnancy. Epidemiology and outcomes. JAMA 250:
1721-1727, 1983.
7. Sweet RL, Landers DV, Walker C, Schachter J: Chlamy-
dia trachomatis infection and pregnancy outcome. Am J
Obstet Gynecol 156:824-833, 1987.
8. Swenson CE, Donegan E, Schachter J: Chlamydia tra-
chomatis-induced salpingitis in mice. J Infect Dis 148:
1101-1107, 1983.
9. Patton DL, Landers DV, Schachler J: Experimental
Chlamydia trachomatis salpingitis in mice: Initial studies
on characterization of the leukocyte response to chlamy-
dial infection. J Infect Dis 159:1105-1110, 1989.
10. Blanco JD, Patterson RM, Ramzy I, Turner T: Clin-
damycin and ibuprofen effects on chlamydial salpingitis
in mice. Sex Trans Dis 16:192-194, 1989.
11. Rank RG, Ramsey KH, Hough AJ: Antibody-mediated
modulation ofarthritis induced by chlamydia. AmJ Pathol
132:372-381, 1988.
12. Ripa KT, Mardh P-A: Cultivation of Chlamydia trachom-
atis in cycloheximide-treated McCoy cells. J Clin Micro-
biol 6:328-331, 1977.
13. Heggie A, Lumicao GG, Stuart LA, Gyves MT:
Chlamydia trachomatis infection in mothers and infants: A
prospective study. Am J Dis Child 135:507-511, 1981.
14. Berman SM, Harrison HR, Boyce WT, Haffner WJ,
Lewis M, Arthur JB: Low birth weight, prematurity,
and post-partum endometritis. Association with prenatal
cervical Mycoplasma hominis and Chlamydia trachomatis
infection. JAMA 257:1189-1194, 1987.
15. Ross SM, Windson IM, Robins-Browne RM, Ballard
RC, Adhikari M, Fenn DB: Microbiological studies dur-
ing the perinatal period: An attempt to correlate selected
bacterial and viral infection with intrauterine deaths and
preterm labor. S Afr Med J 66:598-603, 1984.
16. FitzSimmons J, Callahan C, Shanahan B, Jungkind D:
Chlamydia infection in pregnancy. J Reprod Med 31:19-
22, 1986.
17. Hardy PH, Nell EE, Spence MR, et al.: Prevalence of
six. sexually transmitted disease agents among pregnant
inner-city adolescents and pregnancy outcome. Lancet
2:333-337, 1984.
18. Thompson S, Lopez B, Wong K-H, et al.: A prospective
study of chlamydia and mycoplasma infections during
pregnancy: Relation to pregnancy outcome and maternal
morbidity. In Mardh P-A, et al. (eds): Chlamydia Infec-
tions. Amsterdam: Elsevier Biomedial Press, pp 15-18,
1982.
19. Gravett, MG, Nelson HP, DeRouen T, Critchlow C,
Eschenbach DA, Holmes KK: Independent associations
of bacterial vaginosis and Chlamydia trachomatis infection
with adverse pregnancy outcome. JAMA 256:1899-1903,
1986.
20. Martius J, Krohn M, Hillier S, et al.: Relationships of
vaginal Lactobacillus species, cervical Chlamydia trachom-
atis, and bacterial vaginosis to preterm birth. Obstet Gy-
necol 71:89-95, 1988.
21. The Johns Hopkins Study Group for Cervicitis and Ad-
verse Pregnancy Outcomes: Association of Chlamydia tra-
chomatis and Mycoplasma hominis with intrauterine growth
retardation and preterm delivery. Am J Epidemiol 129:
1247-1251, 1989.
22. ThorpJM Jr, Katz VL, Fowler LJ, KurtzmanJT, Bowes
WA Jr: Fetal death from chlamydial infection across in-
tact amniotic membranes. Am J Obstet Gynecol 161:
1245-1246, 1989.
23. Givner LB, Rennels MB, Woodward CL, Huang S-W:
Chlamydia trachomatis infection in infant delivered by
cesarean section. Pediatrics 68:420-421, 1981.
24. La Scolea LJ, Paroski JS, Burzynski L, Faden HS:
Chlamydia trachomatis infection in infant, delivered by
cesarean section. Clin Pediatr 23:118-120, 1984.
INFECTIOUS DISEASES IN OBSTETRICS AND GYNECOLOGY 245
Submit your manuscripts at
http://www.hindawi.com
Stem Cells
International
Hindawi Publishing Corporation
http://www.hindawi.com Volume 2014
Hindawi Publishing Corporation
http://www.hindawi.com Volume 2014
MEDIATORS
INFLAMMATION
of
Hindawi Publishing Corporation
http://www.hindawi.com Volume 2014
Behavioural 
Neurology
Endocrinology
International Journal of
Hindawi Publishing Corporation
http://www.hindawi.com Volume 2014
Hindawi Publishing Corporation
http://www.hindawi.com Volume 2014
Disease Markers
Hindawi Publishing Corporation
http://www.hindawi.com Volume 2014
BioMed 
Research International
Oncology
Journal of
Hindawi Publishing Corporation
http://www.hindawi.com Volume 2014
Hindawi Publishing Corporation
http://www.hindawi.com Volume 2014
Oxidative Medicine and 
Cellular Longevity
Hindawi Publishing Corporation
http://www.hindawi.com Volume 2014
PPAR Research
The Scientific 
World Journal
Hindawi Publishing Corporation 
http://www.hindawi.com Volume 2014
Immunology Research
Hindawi Publishing Corporation
http://www.hindawi.com Volume 2014
Journal of
Obesity
Journal of
Hindawi Publishing Corporation
http://www.hindawi.com Volume 2014
Hindawi Publishing Corporation
http://www.hindawi.com Volume 2014
 Computational and  
Mathematical Methods 
in Medicine
Ophthalmology
Journal of
Hindawi Publishing Corporation
http://www.hindawi.com Volume 2014
Diabetes Research
Journal of
Hindawi Publishing Corporation
http://www.hindawi.com Volume 2014
Hindawi Publishing Corporation
http://www.hindawi.com Volume 2014
Research and Treatment
AIDS
Hindawi Publishing Corporation
http://www.hindawi.com Volume 2014
Gastroenterology 
Research and Practice
Hindawi Publishing Corporation
http://www.hindawi.com Volume 2014
Parkinson’s 
Disease
Evidence-Based 
Complementary and 
Alternative Medicine
Volume 2014
Hindawi Publishing Corporation
http://www.hindawi.com


Pregnancy Outcome in Swiss-Webster Mice Infected With
                    Chlamydia trachomatis

Karen  D. Bishop

Directory of Open Access Journals

Infectious Diseases in Obstetrics and Gynecology

Pregnancy Outcome in Swiss-Webster Mice Infected With Chlamydia trachomatis

A prospective study of chlamydia and mycoplasma infections during pregnancy: Relation to pregnancy outcome and maternal morbidity.

Bowes WA Jr: Fetal death from chlamydial infection across intact amniotic membranes.

Cervical Chlamydia trachomatis and mycoplasmal infections in pregnancy. Epidemiology and outcomes.

Chlamydia trachomatis infection and pregnancy outcome.

Chlamydia trachomatis-induced salpingitis in mice.

Chlamydial salpingitis and infertility.

Clindamycin and ibuprofen effects on chlamydial salpingitis in mice. Sex Trans Dis 16:192-194,

D: Chlamydia infection in pregnancy.

DB: Microbiological studies during the perinatal period: An attempt to correlate selected bacterial and viral infection with intrauterine deaths and preterm labor.

Disease Control: Chlamydia trachomatis infections: Policy guidelines for prevention and control.

Experimental Chlamydia trachomatis salpingitis in mice: Initial studies on characterization of the leukocyte response to chlamydial infection.

Gyves MT: Chlamydia trachomatis infection in mothers and infants: A prospective study.

Hough AJ: Antibody-mediated modulation ofarthritis induced by chlamydia.

HS: Chlamydia trachomatis infection in infant, delivered by cesarean section. Clin Pediatr 23:118-120,

Huang S-W: Chlamydia trachomatis infection in infant delivered by cesarean section. Pediatrics 68:420-421,

KF: Are chlamydial infections the most prevalent venereal disease?JAMA

KK: Independent associations of bacterial vaginosis and Chlamydia trachomatis infection with adverse pregnancy outcome.JAMA

Low birth weight, prematurity, and post-partum endometritis. Association with prenatal cervical Mycoplasma hominis and Chlamydia trachomatis infection.

Mardh P-A: Cultivation of Chlamydia trachomatis in cycloheximide-treated McCoy cells.

Mucopurulent cervicitis: The ignored counterpart in women of urethritis in men.

Prematurity and perinatal mortality in pregnancies complicated by maternal Chlamydia trachomat# infections.

Prevalence of six. sexually transmitted disease agents among pregnant inner-city adolescents and pregnancy outcome.

Relationships of vaginal Lactobacillus species, cervical Chlamydia trachomatis, and bacterial vaginosis to preterm birth. Obstet Gynecol 71:89-95,

The Johns Hopkins Study Group for Cervicitis and Adverse Pregnancy Outcomes: Association of Chlamydia trachomatis and Mycoplasma hominis with intrauterine growth retardation and preterm delivery.

http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=2366144

Pregnancy Outcome in Swiss-Webster Mice Infected With Chlamydia trachomatis

Abstract

Similar works

Full text

Available Versions

Crossref

Directory of Open Access Journals