Presentation from the 2000
Emerging Infectious Diseases Conference in Atlanta, Georgia
Adaptation of Bordetella pertussis
to Vaccination: A Cause for Its Reemergence?
Frits R. Mooi, Inge H. M. van Loo, and Audrey King
National Institute for Public Health and the Environment (RIVM)
Bilthoven, The Netherlands
| In
The Netherlands, as in many other western
countries, pertussis vaccines have been used
extensively for more than 40 years. Therefore, it
is conceivable that vaccine-induced immunity has
affected the evolution of B. pertussis.
Consistent with this notion, pertussis has
reemerged in The Netherlands, despite high
vaccination coverage. Further, a notable change in
the population structure of B. pertussis
was observed in The Netherlands subsequent to the
introduction of vaccination in the 1950s. Finally,
we observed antigenic divergence between clinical
isolates and vaccine strains, in particular with
respect to the surface-associated proteins
pertactin and pertussis toxin. Adaptation may have
allowed B. pertussis to remain endemic
despite widespread vaccination and may have
contributed to the reemergence of pertussis in The
Netherlands. |
Bordetella pertussis and Bordetella parapertussis
are the etiologic agents of whooping cough or pertussis, a
respiratory disease that is most severe in infants and young
children. Compared to B. pertussis, B. parapertussis
is isolated less frequently from pertussis patients (1% to 5% of
pertussis patients in The Netherlands) and generally causes less
severe symptoms. Pertussis is highly contagious, and, in the
prevaccination era, nearly every child contracted this disease. The
clinical course of pertussis is characterized by paroxysms, or
bursts, of numerous, rapid coughs followed by a long inspiratory
effort, which may be accompanied by a characteristic high-pitched
whoop (hence the designation whooping cough). During such an attack,
patients may turn blue due to lack of oxygen. In serious cases, this
oxygen deprivation may lead to brain damage. The most common
complication of pertussis, however, is secondary pneumonia. Young
infants are at highest risk for pertussis-associated complications.
More than 50% of infants less than 6 months old who contract
pertussis require hospitalization. Treatment of pertussis is
primarily supportive, and adequate control of the disease depends on
effective immunization. Before vaccination was introduced in the
1950s, pertussis was a major cause of infant death throughout the
world (1). Widespread vaccination of young children
has been successful in controlling the disease (1).
Vaccination
The high rate of illness and death caused by pertussis stimulated
the early development of vaccines composed of whole, killed
bacteria. These whole-cell vaccines were introduced in many
countries in the 1950s and 1960s and have been highly successful in
reducing the incidence of pertussis (1). The desire
to avoid the side effects of whole-cell vaccines has stimulated the
development of less reactogenic, acellular, vaccines composed of
purified B. pertussis proteins (2).
Acellular vaccines are replacing whole-cell vaccines in many
countries.
Despite vaccination, pertussis is an endemic disease. Various
sero-epidemiologic studies have shown that the frequency of
infection may be as high as 1%-4% (de Melker and Schellekens, pers.
comm.) (3). Further up to 30% of people with a
persistent cough were found to have been infected with B.
pertussis (4). It is possible that vaccination
initially reduced the circulation of B. pertussis and that
adaptation allowed the B. pertussis population to restore
its high circulation rate. This assumption predicts a change in the
makeup of the B. pertussis population after the
introduction of vaccination, a phenomenon that has indeed been
observed in The Netherlands.
Changes in B. pertussis in a highly
vaccinated population
In some countries with highly vaccinated populations such as
Australia (5), Canada (6), and The
Netherlands (7) (Figure 1),
pertussis has reemerged. Such a phenomenon may have been caused by
changes in the accuracy of notifications, decreases in vaccine
coverage, or changes in vaccine quality. These possibilities have
been excluded for The Netherlands (7), and we have
proposed another possible cause: adaptation of B. pertussis
to the vaccine. To investigate this hypothesis, B. pertussis
strains collected in The Netherlands from 1949 to 1996 were
characterized by DNA fingerprinting and sequencing of genes coding
for surface proteins (8,9).
Initially, we studied changes in the B. pertussis
population by IS1002-based DNA fingerprinting (9,10). Strains collected from 1949 to 1996 were
stratified in periods of 5 to 8 years, and the frequency of
fingerprint types in each period was determined (Figure 2). Widespread vaccination was
introduced in The Netherlands in 1953, and we assumed that the
B. pertussis population was not significantly affected by
vaccination from 1949 to 1954 (defined as the prevaccination period
in Figure 2). Notable differences were found
between the populations from the prevaccination era and the
subsequent period, both in the type and frequency of fingerprint
types (e.g., the major fingerprint type found in strains collected
from 1965-1972 [Ft-29] was absent during the prevaccination period).
These qualitative observations were confirmed by the trend in
genotypic diversity (Figure 2). Genotypic
diversity decreased significantly after the introduction of
vaccination and subsequently increased to prevaccination levels. In
the 1980s, a second decrease in genotypic diversity occurred. Apart
from sampling artifacts, a drop in genotypic diversity may be caused
by a decrease in population size or clonal expansion. Indeed, we
found that the reduction in genotypic diversity in the 1960s and
1980s was associated with the expansion of antigenically distinct
strains.
In a second study of changes in the B. pertussis
population, we investigated whether antigenic shifts had occurred in
surface proteins (8). Very little polymorphism was
observed in most proteins studied. However, two virulence factors,
pertussis toxin and pertactin, were polymorphic. Interestingly,
antibodies against these proteins correlate with protection against
disease, which suggests they have an important role in inducing host
immunity (12,13). Essentially, all DNA
polymorphisms observed were nonconservative, indicating Darwinian
selection. Three pertactin and three pertussis toxin variants were
found in the Dutch B. pertussis population (Figure 3). Polymorphism in pertussis toxin was
restricted to the S1 subunit (PtxS1), which carries the toxic
activity. Variation in PtxS1 was observed in two regions.
Significantly, one of the polymorphic residues has been implicated
in binding to the T-cell receptor (14).
Polymorphism in pertactin was confined to a region comprised of
tandem repeats located proximally to the RGD motif involved in
adherence to host tissues (15). Regions with
tandem repeats are known to undergo rapid variation due to
slipped-strand mispairing during replication (16).
Pertactin and PtxS1 variants, identical to those included in the
Dutch whole-cell vaccine, were found in 100% of the strains from the
1950s, when the whole-cell vaccine was introduced in The Netherlands
(Figure 4). However, nonvaccine types of
pertactin and PtxS1 gradually replaced the vaccine types in later
years and were found in 90% of strains collected from 1990 to 1996.
These results suggest that vaccination has caused strains that are
antigenically distinct from vaccine strains to be selected. The drop
in genotypic diversity observed in the 1960s and the 1980s coincided
with the emergence of nonvaccine-type pertussis toxin and pertactin
variants, respectively, suggesting that the drop was caused by
clonal expansion. Antigenic divergence between vaccine strains and
clinical isolates was also observed in other countries with a long
history of pertussis vaccination, such as Finland and the United
States (17,18), and also in Italy, where vaccine
coverage has varied considerably.
Discussion
Is polymorphism in pertactin and pertussis toxin driven by host
immunity, or is it the result of random fixation due to genetic
drift? The latter possibility is highly unlikely since essentially
all DNA mutations we detected in the pertactin and pertussis toxin
genes were nonconservative. In contrast, random genetic drift is
characterized by a high degree of conservative mutations in protein
coding regions (20). Further, the polymorphic
regions interact directly with the immune system. The polymorphic
region of pertactin induces a protective immune response
(unpublished data). One of the polymorphic residues in PtxS1 has
been implicated in binding to the T-cell receptor (14).
Finally, the fact that the same temporal trends in allele
frequencies are observed in geographically distinct regions such as
Finland, the United States, and The Netherlands argues against
random genetic drift. It is possible that the polymorphic loci we
have identified are linked to other, as yet unknown, polymorphic
loci that increase fitness of strains in vaccinated populations
(hitchhiking).
Strains carrying nonvaccine-type pertactin or pertussis toxin
variants were not found in the prevaccination era. Although the
number of strains analyzed from this period was limited, these data
suggest that the nonvaccine-type variants are not able to displace
the vaccine-type strains in unvaccinated populations (i.e., they
have a lower fitness level?, or reproductive rate, in unvaccinated
communities). Alternatively, the nonvaccine-type strains may have
evolved relatively recently. Consistent with the first hypothesis,
we have observed that nonvaccine-type strains are less fit in naive
mice than vaccine-type strains. In immune mice the difference in
fitness between the two types of strains was much less pronounced
(unpublished data). Thus vaccination has acted to shift the
competitive balance between strains.
An important question to address is whether adaptation of the
B. pertussis population has affected vaccine efficacy, i.e.,
contributed to the reemergence of B. pertussis. Animal
experiments have indicated that variation in pertactin affects
vaccine efficacy (unpublished data). Further, we found vaccine-type
pertactin variants less frequently among vaccinated persons than
among unvaccinated persons, which would be expected if the vaccine
protects differentially against strains with distinct pertactin
types (8). However, the extent to which
polymorphism affects vaccine efficacy is probably dependent on the
vaccine used. It is conceivable that the increase in fitness
associated with nonvaccine types of pertactin and pertussis toxin in
vaccinated populations is substantial enough to drive expansion of
strains carrying these protein variants but that the effect is too
small to result in a measurable drop in vaccine efficacy. Further
studies are required to assess the effect of the observed
adaptations on the efficacy of pertussis vaccines. In this period,
when whole-cell vaccines are being replaced by acellular vaccines in
many countries, continued strain surveillance is of paramount
importance.
Frits R. Mooi is at the National Institute of Public Health and
the Environment, The Netherlands, and the Eijkman-Winkler Laboratory
of the University of Utrecht. His current interests are molecular
epidemiology and evolution of Bordetella spp.
This research was supported by the PraeventieFonds, grant numbers
25-2545 and 28-2852, the Ministry of Health, Welfare and Culture,
and the RIVM.
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