Evaluation of serology and nasopharyngeal cultures for diagnosis of pertussis in a vaccine efficacy trial

Nasopharyngeal cultures and titer rises in paired sera were evaluated in a placebo-controlled pertussis vaccine efficacy trial. IgG ELISA for filamentous hemagglutinin (FHA) identified 30 (88%) of 34 placebo recipients and 33 (89%) of 37 vaccine recipients with culture-verified Bordetella pertussis infections, whereas IgG ELISA for pertussis toxin (PT) showed higher diagnostic sensitivity in the placebo group than in the vaccine groups. The CHO cell assay did not improve sensitivity. Children with Bordetella parapertussis infections had rises of titers of antibody to FHA of the same magnitude as children with B. pertussis infections. Sensitive serologic criteria, based on the intraassay variations, identified 105 additional culture-negative cases with significant titer rises in paired sera. IgG ELISA for FHA and PT and IgA ELISA for FHA were reliable assays, and bacterial isolation rates were lower in vaccine recipients than in placebo recipients with serologically defined pertussis.     

Correlation between pertussis toxin IgG antibodies in postvaccination sera and subsequent protection against pertussis

All acellular pertussis vaccines contain pertussis toxoid and induce protection against pertussis. This study investigated the relation between the postvaccination levels of pertussis toxin (PT) serum IgG and protection against pertussis. PT IgG was determined in sera obtained 21-77 days after the third vaccination from 813 children who received 3 doses of pertussis toxoid. The children were followed for 21-33 months after vaccination for the occurrence of pertussis. Of the children, 126 were exposed to pertussis in their households. The median PT IgG concentration was 79 U/mL in those who developed severe pertussis (>/=21 day of paroxysmal cough), 156 U/mL with mild pertussis (<21 days of paroxysmal cough), and 246 U/mL in those who did not develop pertussis (79 vs. 246, P<.0001). Corresponding values in the 687 children with no household exposure were 99, 124, and 155 U/mL, respectively (99 vs. 155, P<.0001). Thus, there is a highly significant correlation between the level of vaccine-induced serum PT IgG and protection against pertussis.     

Serum IgG, IgA, and IgM responses to pertussis toxin, filamentous hemagglutinin, and agglutinogens 2 and 3 after infection with Bordetella pertussis and immunization with whole-cell pertussis vaccine

Class antibody responses to pertussis toxin (PT), filamentous hemagglutinin (FHA), and agglutinogens 2 and 3 (Agg 2/3) were studied in 114 patients with pertussis, 49 family contacts of patients with pertussis, and 139 infants attending vaccination clinics. IgG responses to FHA and PT were similar in patients with pertussis and in infants immunized three times with whole-cell pertussis vaccine. IgA responses to FHA and PT were greater in patients with pertussis than in vaccines, and a serum log10 titer of IgA to PT greater than 1.9 was a useful discriminant of Bordetella pertussis infection. When compared with patients with pertussis, vaccinees had a greater IgG response to Agg 2/3 and an equivalent IgA response to Agg 2/3. Serologic responses to FHA, PT, and Agg 2/3 were common in family contacts but were smaller than the responses in patients with pertussis. Initial titers of IgG to FHA and IgA to FHA were significantly higher (P less than .005 and P less than .05, respectively) in family contacts aged 3-10 y than in patients with pertussis in an identical age group.     

Immune responses and antibody decay after immunization of adolescents and adults with an acellular pertussis vaccine: the APERT Study

As part of a prospective acellular pertussis (ACP) vaccine efficacy trial, 5 serum samples were obtained, over an 18-month period, from 101 ACP-vaccine recipients and 99 control subjects, to assess ACP antibody response and decay. Immunoglobulin (Ig) G and IgA antibodies to pertussis toxin (PT), filamentous hemagglutinin (FHA), pertactin (PRN), and fimbriae 2/3 (FIM) were measured by enzyme-linked immunosorbant assay, and titers of agglutinin were determined. Of the subjects, 16%-19% had preimmunization values of antibodies to PT that were above the assay's limit of quantitation (LOQ); in contrast, 36%-63% of the subjects had preimmunization values of antibodies to FHA, PRN, or FIM that were above the LOQ. Substantial increases in titers of IgG and IgA antibodies to the 3 ACP antigens (PT, FHA, and PRN) were observed. Over the 18-months, the percent decay in IgG and IgA antibodies ranged from 56% to 73% and from 57% to 70%, respectively; the IgG antibody response and decay suggests that geometric mean titers likely remain above the LOQ for 2-9 years and above the threshold of detection for 4-13 years. These findings support the use of ACP booster immunizations for adolescents and adults, to provide sustained levels of antibody.     

Prevalence of pertussis antibodies in maternal delivery, cord, and infant serum

BACKGROUND: Passively acquired maternal antibodies protect infants from many pathogens. With increasing reports of infant pertussis, we evaluated pertussis antibodies in maternal-infant paired sera from 1999-2000. METHODS: Antibodies to pertussis toxin (PT), filamentous hemagglutinin (FHA), and fimbrial proteins (FIM) were measured by validated IgG-specific enzyme-linked immunosorbant assay (ELISA) in 64 maternal-umbilical cord serum pairs and in 61 of 64 infant sera. Geometric mean concentrations (GMCs) of pertussis antibodies and cord : maternal GMC ratios were calculated. RESULTS: Mean maternal age and gestation were 29.7 years (range, 19-42) and 39.3 weeks (range, 35.6-40.9), and 81% of mothers were white. GMCs of maternal antibodies at delivery (ELISA units/mL) were 2.4 for PT, 6.9 for FHA, and 13 for FIM. Cord GMCs were 169%, 178%, and 157% of maternal delivery values for PT, FHA, and FIM, respectively, demonstrating active placental transfer (P<.001). Pertussis-specific IgG values for each antigen decayed to below the threshold of detection by age 2 months. CONCLUSIONS: Despite efficient placental transfer, low maternal pertussis antibody levels and their rapid decay in infant sera leave infants with little humoral protection against pertussis. These data support the rationale for maternal or neonatal immunization, with acellular pertussis vaccines, to prevent life-threatening pertussis in early infancy.     

A trial of acellular pertussis vaccine in hospital workers during the Cincinnati pertussis epidemic of 1993.

The safety and immunogenicity of acellular pertussis (AP) vaccine in outbreak control was determined in a randomized, double-blind, controlled trial. Participants received AP vaccine (n=102), which contained 25 microg of pertussis toxoid (PT) and 3 microg of filamentous hemagglutinin (FHA), or licensed meningococcal vaccine (MN; n=97). Local reactions (pain or tenderness, redness, swelling, and induration) and systemic reactions (fever, sleepiness or lethargy, and irritability) were similar among AP and MN vaccinees. One month after AP vaccination, the geometric mean level of IgG anti-PT was 33.1 microg/mL, with 2-fold increases in 85% of patients and 4-fold increases in 73% of patients; for IgG anti-FHA, the respective values were 34.7 microg/mL, 92%, and 63%. After 6 months of follow-up, no serological evidence of pertussis was seen among symptomatic or asymptomatic subjects. However, recent evidence of Bordetella pertussis infection before immunization was shown. Thus, AP vaccine was safe and immunogenic in adults.     

Serologic diagnosis of pertussis: evaluation of pertussis toxin and other antigens in enzyme-linked immunosorbent assay

IgM, IgA, and IgG antibodies to Bordetella pertussis were measured in paired sera from 34 patients who were culture-positive for pertussis by enzyme-linked immunosorbent assay (ELISA) with disrupted B. pertussis bacteria, purified pertussis toxin, or outer membrane proteins (OMP) as antigens. Paired sera from 50 patients with other respiratory infections were used as controls. The sensitivities of the assays from paired sera were 61%, 90%, and 90% and specificities were 98%, 92%, and 72%, respectively. Of the patients culture-positive for pertussis, 68% had positive levels of antibody to pertussis toxin antigen in their first serum samples, obtained at the same time as samples for culture. Infants had antibody responses to pertussis toxin antigen, in contrast to weak antibody responses measured by B. pertussis antigen. The results from this study indicate that ELISA, especially measuring pertussis toxin IgA, is a valuable additional tool for diagnosing pertussis and can be used as a complementary test with cultures.     

Mass vaccination of children with pertussis toxoid--decreased incidence in both vaccinated and nonvaccinated persons.

During 1979-1995, there was no vaccination against pertussis in Sweden. With the aim of studying the epidemiology and transmission of pertussis, mass vaccination with pertussis toxoid of children born during the 1990s was instituted in the G?teborg area (population, 778,597) in 1995. Infants were offered 3 doses of pertussis toxoid combined with diphtheria and tetanus toxoids. Children aged > or =1 year were offered 3 doses of pertussis toxoid alone. From June 1995 through February 1999, 167,810 doses of pertussis toxoid were given to 61,219 children born during the 1990s (56% received 3 doses). The number of Bordetella pertussis isolates per year declined from 1214 (1993-1995) to 64 (January 1997 through June 1999; P<.0001), and hospitalizations due to pertussis declined from 62 to 5 (P<.0001). Significant decreases in B. pertussis isolates and hospitalizations occurred in all age groups, including adults and nonvaccinated infants. Thus, mass vaccination of children with pertussis toxoid decreases spread of B. pertussis in the population.     

Human serum antibody responses to Bordetella pertussis infection and pertussis vaccination

We used an immunoblotting technique to compare the serum antibody responses to pertussis toxin (PT), filamentous hemagglutinin (FHA), a 69-kilodalton (kDa) adenylate cyclase-associated protein (69 KD protein), and Bordetella pertussis outer membrane proteins (OMPs) following either B. pertussis infection or immunization with whole-cell pertussis vaccine. Infection and vaccination induced nearly equally intense antibody responses to PT and to FHA, but vaccination induced stronger antibody responses to the 69 KD protein and to many OMPs. The importance of serum antibody responses to the 69 KD protein and to B. pertussis OMPs other than PT and FHA in conferring immunity to pertussis after vaccination is unknown. Serum antibody responses to PT following either infection or vaccination were almost exclusively to the 28-kDa enzymatic subunit (S1) and only rarely and weakly to the lesser molecular weight binding subunits (S2-S5).     

Immune responses to Bordetella pertussis infection and vaccination

To assess antibody and cellular immune responses, 156 healthy children were immunized at approximately 18 months of age with acellular diphtheria-tetanus-pertussis vaccine. Changes in antibody responses to filamentous hemagglutinin (FHA) and to pertussis toxin (PT) were similar in pattern, and antibody titers reached values equal to those from patients with convalescent-stage pertussis. The FHA-induced DNA synthesis in peripheral blood mononuclear cells was maximum at 4 weeks after the primary series, and these levels were equal to those of patients with pertussis. High amounts of PT-induced DNA synthesis were observed in both immunized and nonimmunized children; thus, PT seemed to act mainly as a nonspecific mitogen. Almost the same responses to several mitogens that activate different subsets of lymphocytes were observed in young infants compared with older children. Furthermore, young infants who had Bordetella pertussis infection responded by FHA stimulation almost as well as older children.     

Induction of protective immunity for Bordetella pertussis by nasal inoculation of pertussis vaccine in mice

Although nasal vaccination has emerged as an interesting alternative to intramuscular or oral vaccination, knowledge is scarce about the immune responses after such immunization. In the present study, we inoculated purified Pertussis Toxin (PT) and Filamentous Haemagglutinin (FHA) with or without adjuvant (kayexalate), or Diphtheria acellular Pertussis Tetanus (DaPT) combined vaccine to mice intranasally three times every four weeks to investigate the references of the immunoresponses between nasal and intramuscular vaccination. The levels of pertussis specific serum IgG antibodies (Abs) and secretory IgA Abs in the nasal wash were measured by ELISA, and cytotoxic T cell activities were examined by proliferative response, and compared with the result from intramuscular inoculation. We also studied the efficacy of adjuvant in the nasal vaccination. The intramuscular inoculation of pertussis vaccine induced serum IgG antibodies and cellular immunity against PT and FHA, but did not induce local IgA antibodies. On the other hand, the nasal inoculation induced both serum and local antibody responses. Moreover, it also induced significant cellular immunity to pertussis antigen. In nasal vaccination, the inoculation with adjuvant was superior to inoculation without adjuvant for the induction of both humoral and cellular immunity.     

The longevity of the immune response to filamentous hemagglutinin and pertussis toxin in patients with pertussis in a semiclosed community.

Titers of antibodies to the filamentous hemagglutinin (FHA) and pertussis toxin (PT) of Bordetella pertussis were studied in patients from a semiclosed community after an outbreak of the disease in 1985 and in the general population. In convalescent-phase serum obtained after B. pertussis infection in patients in the semiclosed community, high anti-FHA IgG and anti-PT IgG titers were observed; these titers tended to disappear over the next 5 years. On the other hand, among the medical staff, high anti-FHA and anti-PT IgG titers persisted during the same 5-year period. It seems likely that anti-FHA and anti-PT IgG levels reflect the frequency of exposure to the bacteria and that B. pertussis is widespread in the Japanese population. Although the anti-FHA IgG antibody response decreased in patients from the semiclosed community, the cellular immune response was maintained.     

Toxin-neutralizing antibodies in patients with pertussis, as determined by an assay using Chinese hamster ovary cells

Antibodies to pertussis toxin were measured by a neutralization test (Chinese hamster ovary cell assay) in paired serum samples from 122 patients with whooping cough. The results were compared with previously published titers of IgG, IgM, and IgA measured by ELISA. Significant (fourfold or more) increases in neutralizing antibodies developed in 69 patients. Sixty-eight of them also had significant increases in at least one antibody class as measured by ELISA. Seventeen patients had significant increases in at least one antibody class without increases in neutralizing antibodies. All 163 serum samples with neutralizing antibodies also had detectable IgG, whereas 14 samples with IgG and 18 with IgM were negative by neutralization test. Thus, toxin-neutralizing serum antibodies develop in most patients with pertussis. Our ELISA, however, was more sensitive in detecting low levels of antibodies and in demonstrating significant increases between acute- and convalescent-phase serum samples.     

Incidence of pertussis in persons < or =15 years of age in Valencia, Spain: seroprevalence of antibodies to pertussis toxin (PT) in children, adolescents and adults

PURPOSE: To determine the incidence of pertussis in persons < or =15 years in age in Valencia, Spain. To assess the prevalence of IgG antibodies to pertussis toxin (PT) in children, adolescents and adults. METHODS: Prospective study conducted at paediatric primary care centres. All persons < or =15 years in age presented with persistent cough were enrolled. Parents completed a brief questionnaire and immunization history was obtained from paediatrician records. A blood sample was obtained, for determination of IgG antibodies to Bordetella pertussis toxin (PT) by an ELISA method. A study confirmed-case was the presence of two conditions: (1) cough illness of > or =14 days duration; and (2) ELISA absorbance value of IgG to PT > or =2. Two subjects per clinical-case (same centre and range of age) and parents were asked to participate in the prevalence study. RESULTS: Sixty-one children < or =15 years in age presented with symptoms leading to a clinical diagnosis of pertussis were detected. Serological evidence of recent pertussis was found in five of these patients (incidence of 46.0/100,000 persons < or =15 years in age). Prevalence of antibodies to B. pertussis (> or =0.3) in children < or =15 years in age and adults was 39 and 33%, respectively. Only a minority of children, adolescents and adults had absorbance values indicative of immunity (> or =1). CONCLUSIONS: These incidence and seroprevalence results show that despite high immunization rates in infancy, B. pertussis is circulating in Spain.     

The risk of pertussis and diphtheria infections among pediatric healthcare workers in Japan

For infection control in pediatric hospitals, we investigated the risk of pertussis and diphtheria infections among pediatric healthcare workers. Forty-nine Japanese pediatric healthcare workers in 12 general hospitals were screened for antibodies of pertussis toxin (PT), filamentous hemagglutinin (FHA), and diphtheria toxin (DT). The seropositive rates of anti-PT IgG (protective level, > 10 U/mL), anti-FHA IgG (> 10 U/ mL), and anti-DT (> 0.11 U/mL) were 50, 82, and 59%, respectively. During this survey period (Oct. 2003-Feb. 2004), 16 (33%) of the healthcare workers were in contact with pertussis-infant (s). However, all culture and PCR tests for Bordetella pertussis were negative. One of the 16 exposed healthcare workers, a male pediatrician, had serological evidence of a pertussis infection, but no disease symptomatic of pertussis. Our observations indicate that i) 50 and 41% of Japanese pediatric healthcare workers were seronegative for pertussis (anti-PT IgG) and diphtheria antibodies, respectively, and ii) although the healthcare workers had a high rate of contact with pertussis-infant (s), the infection rate was low. For pertussis and diphtheria infection control in pediatric hospitals, it is important for healthcare workers to be aware of their own protection levels against these diseases.     

Cell-mediated immunity and antibody responses to Bordetella pertussis antigens in children with a history of pertussis infection and in recipients of an acellular pertussis vaccine

Cell-mediated immunity (CMI) and antibody responses to Bordetella pertussis antigens were assessed 4-6 years after primary infant immunization with diphtheria-tetanus tricomponent acellular pertussis (DTaP) or diphtheria-tetanus (DT) vaccine in a country with high endemicity of B. pertussis infection. CMI to the B. pertussis antigens (especially to the pertussis toxin [PT]) was more frequent and/or intense in DTaP than in DT recipients. No lymphoproliferation differences were found between those with and without a history of pertussis although the DT recipients produced very little interferon-gamma after antigen (particularly PT and filamentous hemagglutinin [FHA]) stimulation. In contrast, seropositivity to PT, but not to pertactin or FHA, was more frequent in DT recipients with history of pertussis than in all other subjects. Thus, years after disease or vaccination, CMI response to PT or circulating PT antibodies appears to be the main distinctive feature of pertussis-protected DTaP recipients or pertussis-affected DT recipients.     

Serum antibodies to the components of diphtheria-tetanus-pertussis vaccine in Polish children related to vaccination status

Summary In Poland vaccination against diphtheria, tetanus and pertussis (DTP) is recommended from 2–3 months of age. Three doses at approximately 6-week intervals are given. A booster dose of DTP is given at 19–24 months and boosters of DT at 6 and 14 years. In this study serum samples were obtained from 166 Polish children aged 2 weeks to 14 years. Vaccination status was verified from the children's Health Books. Antibodies were determined against pertussis toxin, filamentous hemagglutinin (FHA), pertactin, tetanus toxoid and diphtheria toxin. Antibodies of maternal origin against all five antigens were detected in almost all sera from infants not yet vaccinated. Antibody levels increased with the number of vaccinations given. Children who had recently received the fourth vaccination had the highest antibody levels. Antibody levels decreased with time after the fourth vaccination for all antibodies except FHA. It was concluded that the Polish whole cell pertussis vaccine stimulates antibodies against pertussis toxin, FHA and pertactin, but that antibodies against FHA probably also are stimulated by cross-reacting antigens. Diphtheria toxin and tetanus toxoid antibodies were above protective levels in all vaccinated children, but the long-term decreases justify the booster dose at 14 years. Twenty-five of 166 children (15%) had a vaccination status which deviated from recommendations demonstrating a need to increase the vaccination rate.

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Serumantikörper gegen die Komponenten der Diphtherie-Tetanus-Pertussis-Vakzine bei polnischen Kindern in Beziehung zum Impfstatus

Zusammenfassung In Polen wird die Impfung gegen Diphtherie, Tetanus und Pertussis (DTP) vom 2.–3. Lebensmonat an empfohlen. Es werden drei Dosen in Abständen von etwa sechs Wochen appliziert. Die Boosterung erfolgt im Alter von 19–24 Monaten mit DPT und im Alter von 6 und 14 Jahren mit DT. In der vorliegenden Studie wurden Serumproben von 166 polnischen Kindern im Alter von zwei Wochen bis 14 Jahren untersucht. Der Impfstatus wurde den Gesundheitsbüchern der Kinder entnommen. Die Antikörper gegen Pertussis Toxin, filamentöses Hämagglutinin (FHA) und Pertactin sowie gegen Tetanus Toxoid und Diphtherie-Toxin wurden bestimmt. Mütterliche Antikörper gegen alle fünf Antigene fanden sich bei allen Seren der noch nicht geimpften Säuglinge. Die Antikörperspiegel stiegen mit der Zahl der verabreichten Impfdosen an. Die höchsten Antikörpertiter fanden sich bei Kindern, die vor kurzer Zeit die vierte Impfdosis erhalten hatten. Mit Ausnahme von FHA nahmen die Titer aller Antikörper nach der vierten Impfung mit der Zeit ab. Es wird angenommen, daß die polnische Ganzzell-Vakzine gegen Pertussis die Antikörperbildung gegen Pertussis-Toxin, FHA und Pertactin stimuliert, daß jedoch die Antikörperbildung gegen FHA wahrscheinlich auch durch kreuzreagierende Antigene angeregt wird. Die Antikörper gegen Diphtherie-Toxin und Tetanus-Toxoid waren bei allen Kindern oberhalb der protektiven Spiegel, aber der Abfall über lange Zeit rechtfertigt die Boosterung mit 14 Jahren. 25 der 166 Kinder (15%) hatten einen Impfstatus, der von den Empfehlungen abwich, was zeigt, daß es nötig ist, die Impfrate zu erhöhen.

     

Diphtheria, tetanus, and pertussis immunizations in adults

Although only few cases of diphtheria are reported annually in the United States, substantial numbers of adults have declining levels of protective serum antitoxin. A recent outbreak of diphtheria in Sweden emphasizes the importance of reimmunizing adults in diphtheria toxoid, included as part of routine use of tetanus/diphtheria toxoid. Tetanus is a completely preventable disease. Yet, most of the 60 to 100 annual cases of tetanus occur in older adults with no prior history of immunization. Tetanus immunization should be a routine part of medical care of all adults. Research currently is aimed at improving the efficacy and safety of pertussis vaccine. Future vaccines may be useful in protecting health care workers and other adults who have extensive contact with children against pertussis.     

Population-based incidence of pertussis among adolescents and adults, Minnesota, 1995-1996

To estimate the incidence of pertussis, a prospective study was done among members of a managed care organization in Minneapolis/St. Paul, Minnesota. Of 212 patients 10-49 years old enrolled from January 1995 through December 1996, 8 were found to be culture positive, 10 were found to be positive by polymerase chain reaction assay, 13 had a > or =2-fold increase in IgG or IgA to pertussis toxin (PT), and 18 had IgG to PT in a single serum specimen > or =3 SD above the mean of an age-matched control group. At least 1 positive laboratory test result for pertussis infection was found in 27 (13%) patients, among whom the duration of cough illness was a median of 42 days (range, 27-66 days). On the basis of any positive laboratory result, the estimated annual incidence of pertussis was 507 cases per 100,000 person-years (95% confidence interval, 307-706 cases). Bordetella pertussis infection may be a more common cause of cough illness among adolescents and adults than was recognized previously.     

Frequency of pertussis in children with prolongued cough

To determine the frequency of pertussis in children < or = 16 y who had prolonged cough (> or = 14 d), a prospective study was conducted at an outpatient clinic of a paediatric hospital. Nasopharyngeal swabs were taken for culture and nucleic acid testing by polymerase chain reaction (PCR) for Bordetella pertussis. Immunoglobulin A and immunoglobulin G antibodies against pertussis toxin (PT) were tested by ELISA in paired serum samples. A total of 148 patients were recruited during 1 y. Pertussis was detected in 25 (16.9%) patients with at least 1 of the tests. PCR was positive in 12 patients, and 9 cases was diagnosed serologically. Both PCR and serology were positive in 4 children. Duration of cough was longer in the patients with pertussis (median 33 vs 20, p = 0.03). Seropositivity of pertussis toxin was higher in pertussis negative patients during enrollment (24% vs 65%, p = 0.005). From the results of this study, B. pertussis seems to be common in our population despite high immunization rates with whole cell vaccine. Although the duration of cough is defined as longer than 21 d in some studies for pertussis case definition criteria, it was shorter than this in 3 of our cases.