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| Journal of the New Zealand Medical Association,
20-February-2004, Vol 117 No 1189
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Serological evidence of
Toxoplasma gondii infection among
pregnant women in Auckland
Arthur Morris and Margaret
Croxson
Primary Toxoplasma
gondii infection in pregnancy is usually asymptomatic, nevertheless
infection may be transmitted to the fetus and cause severe
damage.1 Infections occurring in the first
trimester have a 10% chance of transmission to the fetus and have the worst
prognosis because of the risk of extensive central nervous system
involvement.2 Infection occurring in later
pregnancy is transmitted to the fetus more frequently but does not cause such
extensive damage. Some countries, such as France, Switzerland, Austria, Germany,
Norway and Italy, screen pregnant women for T.
gondii infection during pregnancy. Other countries including New Zealand
do not screen.
Screening for toxoplasmosis is made difficult by the unusual
persistence of the IgM antibody response.3, 4
With most other infections, the predominate early response is IgM antibody,
which then dwindles to undetectable levels within a few weeks. After primary
T. gondii infection, however, IgM
antibody frequently remains positive for many months or even years. A screening
programme aimed at pregnant women in their first trimester would ideally detect
only very recent infection, since pre-conception infection is not a risk to the
subsequent pregnancy. An antenatal screening programme would therefore need to
include tests for IgM antibody. As a consequence of the persistent IgM response
a number of women would be detected with
Toxoplasma IgM in whom primary
infection pre-dated conception. In these women the fetus is not at risk of
congenital toxoplasmosis,5, 6 but the problem
is how to distinguish them from women with true first-trimester
infection.
In order to provide data on the immune status of pregnant
women, 500 serum samples submitted for routine antenatal screening were tested
for antibody to T. gondii.
MethodsSerum
selection One hundred consecutive serum samples from each of five age
groups (<20, 21–25, 26–30, 31–35, >36 years) were tested
anonymously for antibodies to T.
gondii. Those that were IgG positive were tested for IgM antibody. The
samples had been submitted for the first routine set of antenatal blood
tests.
Test methods
Serum samples were tested in an enzyme immunoassay (EIA) on the AxSym system
using the IMXII kit (Abbott Laboratories). Sera with positive IgM results were
sent to the Immunology Department of Auckland Hospital for further testing. This
comprised an in-house immunofluorescent assay (IFA) using reconstituted
T. gondii trophozoites (BioMerieux)
fixed onto pre-marked wells on clean slides. A 1:64 dilution of the serum
sample, using rabbit anti-human IgG antiserum as the diluent, was added to the
well. Slides were then incubated for 30 minutes at 37˚C and rinsed.
Specific, fluorescent labelled anti-IgM conjugate (Fluoline-M, BioMerieux) was
then added and the slides incubated for further 30 minutes at 37˚C.
Subsequently, slides were washed, mounted and read using UV illumination.
Fluorescence was recorded as negative, weakly positive, or positive. Sera were
read without knowledge of the EIA result along with a random selection of
negative sera. Sera IgM positive by both EIA and IFA methods were deemed IgM
positive.
Estimation of
screening results The last calendar year for which full information is
available is 2000, when 14 530 mothers gave birth at National Women’s and
Middlemore Hospitals (personal communication, Maternity Services, 2003). To
estimate the results of routine screening for toxoplasmosis the number of women
giving birth in the respective age groups above was multiplied by the proportion
with positive IgM results in our sample.
Statistics To
assess the significance of trend the Cochran-Armitage Trend Test was used.
Statistical analysis was performed using SAS release 8.0 Software (Cary, North
Carolina).
The study was approved by the Ethics Committee North
Health (reference number 97/118).
ResultsOne hundred and sixty three (33%)
women had IgG antibody to Toxoplasma
(Table 1). Sixteen samples that were IgM positive by EIA were sent for IFA
testing and 12 (2.4%) were confirmed as being IgM positive. There was a trend
for the prevalence of IgG antibodies to increase with age (Table 1, p = 0.13).
In 2000, for the five age groups listed in Table 1, the estimated numbers of
women with positive IgM giving birth were 49/1227, 141/2814, 0/4049, 84/4207,
and 22/2233 respectively. We estimate, therefore, that 296 of 14 530 women
(2.03%, 95% CI: 1.8–2.2%) would have had a positive IgM for
T. gondii if routine screening had been
performed. Thus, up to 2.2% of women could have initial antenatal serology
consistent with recent infection.
Table 1. Toxoplasma
gondii serology results of 500 pregnant Auckland women
*100 serum samples tested in each age group;
†p= 0.13 for trend of increasing
seroprevalence with age
DiscussionAntenatal screening for infectious
diseases presupposes there can be a better outcome for the baby. There are clear
guidelines as to what constitutes a worthwhile screening
programme,7 and understanding the disease
incidence is an essential starting point. Other requirements are the
availability of simple and safe tests that clearly differentiate between
infected and non-infected individuals and, most importantly, the availability of
effective interventions.
Antenatal screening in New Zealand currently includes
rubella, hepatitis B and syphilis. In each of these infections there are
acceptable interventions based on the known risk of adverse outcome to the fetus
and the known efficacy of medical therapy. There are strong arguments for adding
HIV testing to this programme because, with proper management, the risk of fetal
infection can be drastically reduced. Unfortunately, the situation with
toxoplasmosis is much less clear. First, the incidence of congenital
toxoplasmosis in New Zealand is unknown, so the risks/benefits of a screening
programme cannot be assessed. Second, there are no good data on the benefits of
treatment during pregnancy.8–10 Third,
current serological tests may not distinguish between recently acquired and
remote infection. 11 Consequently, there is the
very real possibility that more harm might result from positive screening tests
than potential benefit from detection of pregnancies actually at risk.
Our study set out to answer two questions: what percentage
of pregnant women in Auckland have antibody to
T.gondii at the time of their first
antenatal visit; and how many of these women have serological evidence of recent
primary infection.
Testing for toxoplasmosis involves testing for IgG and IgM
antibodies. With most infections, IgM antibody disappears some weeks after the
primary infection so detection of IgM implies recent infection. With
toxoplasmosis, the IgM antibody may persist for months or even
years.12 Attempts to increase the specificity
for recent infection include testing for IgA antibody and testing for avidity of
the IgG antibody.13–15 Avidity testing is
based on the maturation of the antibody response over time, with low-avidity
antibody being replaced by high-avidity antibody. The detection of high-avidity
IgG antibody may be useful by allowing us to place infection before conception.
This depends on the assay in question and the timing of the
test.13,14 There are, however, only few data on
the clinical outcome of children (n = 13) born to women who had high-avidity
antibody in their first trimester.16 Although
none of these children had evidence of T.
gondii infection at follow up, all the mothers had received spiramycin
treatment during their pregnancy. It is unclear, therefore, whether these 13
children did not become infected because the maternal infection truly pre-dated
conception, as suggested by the high-avidity IgG, or spiramycin protected the
fetus, or whether the result was observed by chance since the probability of
congenital infection following first-trimester infection is only about 10%.
Clearly, more data are required on the reliability of IgG avidity
testing.
Second-round tests involve looking directly for the
Toxoplasma parasite, which in practice
devolves to polymerase chain reaction (PCR) for
T. gondii
DNA.17 Searching for the parasite has its own
problems, first and foremost being the rapidity with which
T. gondii is cleared from the
circulation in the immunocompetent host. A study looked for
Toxoplasma DNA in the peripheral white
blood cells of immunocompetent individuals with a known exposure date.
T. gondii DNA could be recovered from
approximately 33% of the patients over the 10 weeks following primary infection,
with 53% of these positive results within five weeks of
infection.18 Since most maternal infections are
subclinical, the opportunities to test within that five-week period are rare.
Even if acute maternal infection can be established unambiguously, this does not
necessarily imply fetal infection.19 Attempts
have been made to monitor fetal status by ultrasound imaging, but this has low
sensitivity.20 Currently, amniocentesis and
testing for T. gondii by PCR is
considered the most reliable diagnostic approach, with high positive and
negative predictive values.21–26
Amniocentesis does, however, pose a risk to the pregnancy and requires
considerable medical resources.
Our data show that 2% (range 1.8–2.2%) of women tested
in the first trimester will have an antibody profile consistent with recent
primary T. gondii infection. If one
accepts that transmission to the fetus in the first trimester will occur in 10%
of these infections, that implies some 2/1000 cases of first-trimester
congenital toxoplasmosis in Auckland. Considering that first-trimester infection
is the most severe and would be clinically apparent, this is clearly a gross
overestimate, since this level of clinical disease is not occurring. Therefore,
as a result of screening, up to 2.2% of pregnant women will have the anxiety of
a potentially damaged baby and may undergo amniocentesis and/or termination of
the pregnancy. The overwhelming majority of these procedures will be
unnecessary.
Medical intervention is likewise of uncertain
benefit.27 Traditionally, spiramycin has been
advocated for treatment in early pregnancy, being apparently risk free to the
fetus and having putative parasitostatic
properties.28 It is hoped that the
administration of this antibiotic will prevent
Toxoplasma from replicating in the
placenta and thus avert fetal infection. There have been no controlled trials of
the effectiveness of spiramycin in preventing congenital toxoplasmosis; instead
all efficacy data are based on comparison with historical untreated series.
Treatment in later pregnancy generally includes either
pyrimethamine/sulphadiazine plus supplemental folinic acid or co-trimoxazole in
a bid to achieve active treatment of the infected fetus and, particularly, to
treat infection of the central nervous
system.29 Pyrimethamine/sulphadiazine treatment
is not without side-effects and may be teratogenic as a consequence of severe
bone marrow suppression unless adequate folinic acid is supplied.
It is apparent that the risk of
T. gondii infection currently does not
fulfil the requirements that would justify its inclusion in an antenatal
screening programme in New Zealand. Data on the incidence of congenital
infection is sparse. We have presented data on a small series of pregnant women
in Auckland, but this is an urban community and figures from rural districts may
be significantly different.30 The current
serological tests that might be offered in a screening programme lack the
necessary discrimination between recent and remote infection. The consequences
of this are the extensive and invasive investigations needed to assess the
possibility of true first-trimester infection. Nor is there a proven medical
intervention available, with scant data available on the efficacy of the
currently recommended treatment protocols.
In view of these major barriers to screening, we suggest at
this time that education for women on how to avoid
T. gondii infection should be the
preferred approach. Testing is best reserved for symptomatic patients, in whom
the prior probability of infection favourably weights the positive predictive
value of a positive IgM result.
Author information:
Arthur J Morris, Clinical Microbiologist, Clinical Microbiology Laboratory,
Diagnostic Medlab, Auckland; Margaret C Croxson, Clinical Virologist, Virology
and Immunology Laboratory, LabPlus, Auckland
Acknowledgements:
This research was funded by the Health Research Council of New Zealand, grant
number 98/330/PHR/PL/HRL.
Correspondence: Dr
Arthur Morris, Diagnostic Medlab, PO Box 14 743, Auckland. Fax: (09) 571 4091;
email: amorris@dml.co.nz
References:
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