Bordetella pertussis Infection: Pathogenesis, Diagnosis, Management, and the Role of Protective Immunity

 Whooping cough is presently one of the ten most common causes of death from infectious disease worldwide. Despite a high vaccine uptake, resurgences of this disease have been observed in several countries. Virulence factors of Bordetella pertussis include agglutinogens, fimbriae, P.69/pertactin, pertussis toxin, filamentous haemagglutinin, adenylate cyclase, tracheal cytotoxin, dermonecrotic toxin, lipopolysaccharide, tracheal colonisation factor, serum resistance factor, and type III secretion. Virulence factor expression is regulated by the bvgAS locus, a two-component signal transduction system. The pathophysiologic sequence consists of attachment (fimbriae, P.69/pertactin, tracheal colonisation factor, pertussis toxin, filamentous haemagglutinin), evasion of host defence (adenylate cyclase, pertussis toxin, serum resistance factor), local effects (tracheal cytotoxin), and systemic effects (pertussis toxin). Bordetella pertussis is transmitted by respiratory droplets and causes disease only in humans. Various diagnostic methods are available, including culture, serological methods, and the polymerase chain reaction. Serotyping of isolates to detect agglutinogens 2 and 3 is useful because serotype 1,2 may be associated with higher mortality, and antibodies to these antigens (agglutinins) may be protective in both animals and humans. Immunisation using whole-cell vaccine is effective but is reactogenic. Acellular vaccines containing one to five components are being used increasingly in various countries. Protective immunity to pertussis correlates with high levels of antibody to each of pertactin, fimbriae, and pertussis toxin; however, doubt remains as to the relationship between agglutinogen 3 and fimbria 3, making results of trials investigating these virulence factors difficult to interpret.     

Pertussis Diagnosis in the 21st Century: Progress and Pitfalls, Part I

Bordetella pertussis is the causative agent of pertussis, also called whooping cough or the cough of 100 days. Infection can result in significant morbidity and mortality, particularly in young infants. Prior to the availability of effective vaccines, pertussis was a major cause of childhood disease. With the advent of such vaccines, the incidence of disease declined dramatically into the 1970s. However, pertussis is still present, with peaks occurring every 3 to 5 years, and the number of cases has been increasing in the United States since the 1980s. With recent reports of numerous outbreaks of pertussis, there is heightened interest in the control and diagnosis of the disease. Efforts to increase immunity through vaccination and also to improve the clinical and laboratory diagnosis of the disease are very important. Part I of this two-part article will review a recent outbreak of pertussis that occurred in California and discuss the biology of the genus Bordetella, followed by the clinical presentation of disease and recommendations for recent vaccination protocols and guidelines for diagnosis. Part II of this article will be published in the August 15 issue of this newsletter and will review laboratory methods available for diagnosis, along with their problems and pitfalls.     

Pertussis Diagnosis in the 21st Century: Progress and Pitfalls, Part II

Bordetella pertussis is the causative agent of pertussis, also called whooping cough or the cough of 100 days. Infection can result in significant morbidity and mortality, particularly in young infants. Prior to the availability of effective vaccines, pertussis was a major cause of childhood disease. With the advent of such vaccines, the incidence of disease declined dramatically into the 1970s. However, pertussis is still present, with peaks occurring every 3 to 5 years, and the number of cases has been increasing in the United States since the 1980s. With recent reports of numerous outbreaks of pertussis, there is heightened interest in the control and diagnosis of the disease. Efforts to increase immunity through vaccination and also to improve the clinical and laboratory diagnosis of the disease are very important. Part II of this two-part article will discuss methods for the diagnosis of pertussis infection with an emphasis on laboratory tests using PCR and serology. The advantages of newer testing methods include increased sensitivity and decreased time to result. There are potential problems, however, related to specificity and cross reactions that can complicate the interpretation of test results for pertussis diagnosis.     

Cryptococcus gattii: a Review of the Epidemiology, Clinical Presentation, Diagnosis, and Management of This Endemic Yeast in the Pacific Northwest

Cryptococcus gattii emerged on Vancouver Island, British Columbia, Canada, in 1999. Subsequent spread to the Vancouver lower mainland, Washington state, and Oregon has been documented. Unlike classic Cryptococcus neoformans, which causes disease in immunosuppressed individuals, C. gattii tends to cause disease in essentially immunocompetent individuals. This article reviews the epidemiology, clinical presentation, diagnosis, and treatment of infections caused by this now endemic yeast in the Pacific Northwest.     

Rapid diagnosis of pertussis in young infants: comparison of culture, PCR, and infant's and mother's serology.

The contribution of maternal pertussis serology comparing prepartum serum to serum collected during the infant's disease to the diagnosis of pertussis in infants was evaluated for 28 pairs of young infants with pertussis syndrome and their mothers and was compared to those of culture and PCR. Infants had a nasopharyngeal aspiration tested by PCR, and acute and convalescent sera were collected during their disease. Mothers had a first acute serum collected concomitantly with the infant's acute serum, and both acute sera were compared to a prepartum serum. Sera were analyzed by immunoblotting for the detection of anti-pertussis toxin (PT) antibodies. Serological evidence of pertussis in infants was assessed as either an increase in anti-PT antibody levels between the mother's prepartum and acute sera or the presence of antibodies in the infant's acute serum and their absence in both the mother's acute and prepartum sera. Culture and PCR sensitivity were 43 and 89%, respectively. Most infants (18 of 24) had no pertussis antibody detectable in their acute sera, confirming a delayed immune response at this age. A comparison of infant's and mother's serology, using prepartum serum, rapidly confirmed the diagnosis in 57% of the cases. Although less sensitive than PCR, this serological method should be used for a rapid diagnosis of pertussis in young infants when culture and PCR are either not available or negative.     

The diagnosis of pertussis: which method to choose?

Despite the introduction of routine vaccination against pertussis for more than a half century, leading to a drastic decline in the number of reported cases, pertussis continues to be an important respiratory disease afflicting unvaccinated infants and previously vaccinated children as well as adults in whom immunity has waned. The diagnosis of pertussis is challenging and accurate laboratory identification of Bordetella infections remains problematic. Common laboratory diagnostic methods used for pertussis diagnosis include culture, direct-fluorescent-antibody testing (DFA), serology and polymerase chain reaction (PCR). Culture of Bordetella pertussis is highly specific but fastidious and has limited sensitivity. DFA provides a much more rapid result, but has the disadvantage of poor sensitivity and specificity. Serology is not useful in infants. In older persons, it is hampered by the limitations of paired sera and it provides mainly a retrospective diagnosis. Such limitations of conventional diagnosis testing have led to the development of PCR assays. Notwithstanding its lack of standardization, PCR has been found to be more sensitive and more specific than other methods. In this report, we aimed to review current knowledge about the available diagnostic methods and tests that accurately diagnose pertussis.     

An Epidemic of Pertussis Among Elderly People in a Religious Institution in The Netherlands

An epidemic of pertussis is described among elderly people in a religious institution in the Netherlands in 1992. Subjects were evaluated for their vaccination status and for history and presence of respiratory symptoms. Specimens were collected for culture, polymerase chain reaction, and serological evaluation. None of the 75 residents and 19 of 24 nonresident personnel had been vaccinated against pertussis. The overall attack rate of clinical pertussis, defined as persistent cough lasting at least 2 weeks, was 49%. In five subjects with clinical pertussis, either culture or polymerase chain reaction or both were positive for Bordetella pertussis. A significant (at least 4-fold) change in specific antibody titre was observed in 85% (41/48) and 20% (10/49) of subjects with and without clinical pertussis, respectively (P<0.0001, chi-square 41.1). The attack rate of laboratory-confirmed pertussis was 42% (41/98). This rate was 5% (1/19), 20% (1/5), and 53% (39/74) in vaccinated personnel, nonvaccinated personnel, and nonvaccinated residents, respectively (not significant). Among residents aged between 55–74 years and 75–94 years, the attack rates were 47% (17/36) and 58% (22/38), respectively (relative risk=0.8; 95% confidence interval 0.5–1.3). Four of 75 residents (5%) died from intracranial bleeding, while they were symptomatic for pertussis. It is concluded that the attack rate of pertussis was high among nonvaccinated elderly and that pertussis tended to increase with age. There may be a considerable risk of mortality from pertussis in this population. Physicians should be alert to the diagnosis of pertussis in the elderly with nocturnal and prolonged periods of coughing.     

Evaluation of culture, immunofluorescence, and serology for the diagnosis of pertussis.

Nasopharyngeal culture, direct immunofluorescence, and serology of acute-phase and paired serum specimens were compared for the laboratory diagnosis of infections due to Bordetella pertussis in a community-based pediatric population with both high vaccine usage and high pertussis incidence. In 77 (37%) of 210 patients evaluated, one or more tests were positive for pertussis. A clinical illness compatible with pertussis was present in 52 (71%) of 73 pertussis test-positive and 42 (35%) of 119 test-negative patients (P less than 0.001). Nasopharyngeal culture was of low sensitivity (20 [26%] of 77 positive tests) but was most commonly confirmed by another positive pertussis test (85%). Direct immunofluorescence was both insensitive and nonspecific; only 6 (30%) of 20 cases positive by culture were positive by immunofluorescence, and only 4 (33%) of 12 of the culture-negative, immunofluorescence-positive cases could be confirmed by another positive pertussis test. Although serology by enzyme immunoassay proved to be the most sensitive of the laboratory tests (87%), this sensitivity could be achieved only by assaying both acute-phase and paired serum specimens and measuring immunoglobulin G (IgG), IgA, and IgM antibodies to two pertussis antigens (pertussis toxin and filamentous hemagglutinin). Loss of sensitivity occurred with any reduction in the number of these serologic assays performed. Optimal laboratory diagnosis of endemic pertussis in a pediatric population requires both nasopharyngeal culture and serology by enzyme immunoassay.     

Tdap5 Vaccine (Covaxis®)

Covaxis (also licensed as Triaxis or Adacel in individual countries) is a combined tetanus toxoid, reduced diphtheria toxoid, five component acellular pertussis (namely detoxified pertussis toxin, filamentous hemagglutinin, pertactin, and fimbriae types 2 and 3) vaccine for the prevention of diphtheria, tetanus, and pertussis. It is approved for use in Europe as a single intramuscular booster dose in children (aged ≥ 4 years), adolescents, and adults, and in the US it is approved for use in individuals aged 11-64 years. In large, randomized, controlled clinical trials conducted in the UK and North America, a single intramuscular booster dose of Covaxis induced robust immune responses for all of its component antigens when given to children (aged ≥ 4 years), adolescents, and adults. In addition, Covaxis vaccine was safe and generally well tolerated in terms of solicited and unsolicited local injection-site and systemic adverse events, most of which were of mild intensity and resolved without sequelae. Furthermore, the immunogenicity of each individual component and the reactogenicity of Covaxis vaccine in children, adolescents, and adults was generally similar to that of comparator vaccines. Despite being a vaccine-preventable disease and having >90% primary vaccination coverage worldwide, pertussis remains uncontrolled, particularly amongst adolescents and adults. Given the changing epidemiology of pertussis and the requirement to reduce infection in adolescents and adults (including healthcare workers) and thereby prevent transmission of the disease from these individuals to very young infants, the new 'cocoon strategy' recommended in current vaccination guidelines has become a key strategy in the management of morbidity and mortality associated with pertussis. This strategy focuses on the immunization of healthcare workers, and the parents and family members of infants who are too young to have undergone primary immunization, so as to prevent the transmission of pertussis to these young at-risk infants. The implementation of the 'cocoon strategy' may finally give countries the ability to control pertussis infections in these at-risk infants and ultimately provide the desired herd immunity against pertussis. In line with this strategy, a booster dose of Covaxis vaccine provides a valuable option to reduce pertussis morbidity and mortality, and to maintain seroprotection against diphtheria and tetanus in children (aged ≥ 4 years), adolescents, and adults.     

Multitarget PCR for diagnosis of pertussis and its clinical implications

PCR has greatly facilitated pertussis diagnosis due to the speed, sensitivity, and specificity of this assay compared to other detection methods. Various single-target PCR assays are currently utilized, but none is universally considered to be the "gold standard." Our aim was to assess the use of multitarget versus single-target PCR for the diagnosis of pertussis in clinical samples. PCR assays targeting insertion sequence IS481 (IS), pertussis toxin ptxA promoter region (PT), and outer membrane porin (PO), or recA (RA) were evaluated in respiratory specimens collected from 4,442 patients with suspected pertussis. The diagnosis of pertussis was confirmed in 309 (6.96%) patients by the 3-target IS-PT-PO/RA PCR versus 247 (5.56%) by the conventional single-target IS (P=0.007). Compared to single-target IS, the three-target combination increased the proportion of positive specimens by 1.25-fold, and two-target combinations increased the proportion of positive specimens by 1.10- to 1.24-fold. In addition, nine cases of B. parapertussis infection were also confirmed by using the discriminative features of this multitarget PCR. Of the 89 culture-proven pertussis cases, 17 (19.1%) and 5 of the 16 patients (31.3%) admitted to intensive care unit would have been missed had only the single-target IS PCR been applied. Patients with mild disease (P=0.004) and shorter hospitalization (P=0.006) were less likely to have positive cultures. This consensus generating real-time PCR approach permits a sensitive detection, as well as an accurate species identification of the causative Bordetella pathogens for the timely management of patients.     

Comparison of polymerase chain reaction, culture, and western immunoblot serology for diagnosis of Bordetella pertussis infection.

Polymerase chain reaction (PCR) amplification of the pertussis toxin promoter region was used to detect Bordetella pertussis infection in nasopharyngeal aspirates collected from 24 infants and children infected with pertussis and 13 adult contacts during an epidemiological study. The sensitivity of this PCR assay was approximately one bacterium, and the assay was specific for B. pertussis in tests with other Bordetella species and other respiratory pathogens. The pertussis case definition required a cough with a duration of more than 21 days for infants and children and laboratory confirmation by serology as the primary detection method for infants, children, and adults. The sensitivity of PCR and culture on Bordet-Gengou agar medium was assessed with regard to the case definitions. In the group of infants and children (index cases), the sensitivities of the culture and the PCR were 54.1% (13 of 24) and 95.8% (23 of 24), respectively. In the adult group (household contacts), the sensitivities of the two methods were 15.4% (2 of 13) and 61.5% (8 of 13), respectively. PCR combined with pertussis-specific serology appears to be a useful tool for diagnosis of pertussis especially in epidemiological studies.     

Congenital Athymia: Genetic Etiologies,Clinical Manifestations,Diagnosis, and Treatment

Congenital athymia is an ultra-rare disease characterized by the absence of a functioning thymus. It is associated with several genetic and syndromic disorders including FOXN1 deficiency, 22q11.2 deletion, CHARGE Syndrome (Coloboma, Heart defects, Atresia of the nasal choanae, Retardation of growth and development, Genitourinary anomalies, and Ear anomalies), and Complete DiGeorge Syndrome. Congenital athymia can result from defects in genes that impact thymic organ development such as FOXN1 and PAX1 or from genes that are involved in development of the entire midline region, such as TBX1 within the 22q11.2 region, CHD7, and FOXI3. Patients with congenital athymia have profound immunodeficiency, increased susceptibility to infections, and frequently, autologous graft-versus-host disease (GVHD). Athymic patients often present with absent T cells but normal numbers of B cells and Natural Killer cells (T^?B⁺NK⁺), similar to a phenotype of severe combined immunodeficiency (SCID); these patients may require additional steps to confirm the diagnosis if no known genetic cause of athymia is identified. However, distinguishing athymia from SCID is crucial, as treatments differ for these conditions. Cultured thymus tissue is being investigated as a treatment for congenital athymia. Here, we review what is known about the epidemiology, underlying etiologies, clinical manifestations, and treatments for congenital athymia.

     

Pertussis vaccine--an analysis of benefits, risks and costs.

Using decision analysis, we estimated the benefits, risks and costs of routine childhood immunization against pertussis. Without an immunization program, we predict that there would be a 71-fold increase in cases and an almost fourfold increase in deaths (2.0 to 7.6) per cohort of one million children. With a vaccination program, we predict 0.1 case of encephalitis associated with pertussis and five cases of post-vaccination encephalitis; without a program, there would be only 2.3 cases of encephalitis associated with pertussis. Community vaccination would reduce by 61 per cent the costs related to pertussis. Our analysis supports continuation of vaccination in routine childhood immunization programs, but suggests the need for more reliable data on complications from the vaccine, further study of the epidemiology of pertussis and development of a less toxic vaccine.     

Are vaccination programs and isolate polymorphism linked to pertussis re-emergence?

Whooping cough remains an endemic disease, and the re-emergence of pertussis in older children and adolescents has been reported in several countries, despite high vaccine coverage. Polymorphism of Bordetella pertussis has been observed over time, and some characteristics of pertussis isolates have gradually diverged from the vaccine strains. The present review summarizes the current knowledge on B. pertussis variability in countries with different vaccination programs and discusses its potential impact on the recently observed increased incidence of whooping cough. No direct association between B. pertussis isolate variability and vaccination programs has been observed to date, except for shifts from fimbriae Fim2 to Fim3. More likely explanations for the re-emergence of pertussis include the change in the epidemiology and transmission patterns of pertussis in highly vaccinated populations, and a shift of disease from young children to adolescents and adults due to waning protective immunity.     

Characterization of serological responses to pertussis

We have compared the use of five nonvaccine antigens to the use of conventional vaccine antigens, pertussis toxin (PT), and filamentous hemagglutinin (FHA) for the serological diagnosis of pertussis by enzyme-linked immunosorbent assay (ELISA). The nonvaccine antigens included the catalytic region of adenylate cyclase toxin (CatACT), the C-terminal region of FHA (C-FHA), lipooligosaccharide (LOS), the peptidoglycan-associated lipoprotein (PAL), and the BrkA protein. The serological responses of individuals with culture-confirmed pertussis were compared to those of adults with no recent history of a coughing disease. An immunoglobulin G (IgG) ELISA for PT was the most sensitive (92.2%) test for the serodiagnosis of pertussis. Of the nonvaccine antigens, ELISA for IgG responses to CatACT (sensitivity, 62.8%), C-FHA (sensitivity, 39.2%), and LOS IgA (sensitivity, 29.4%) were less sensitive but could also distinguish culture-positive individuals from control individuals. The use of a combination of multiple ELISA targets improved the sensitivity of the assay for serological diagnosis. Elevated IgG and IgA antibody titers persisted for more than a year in the individuals with culture-confirmed pertussis.     

Variation in the Bordetella pertussis Virulence Factors Pertussis Toxin and Pertactin in Vaccine Strains and Clinical Isolates in Finland

There is evidence that pertussis is reemerging in vaccinated populations. We have proposed, and provided evidence for, one explanation for this phenomenon in The Netherlands: antigenic divergence between vaccine strains and circulating strains. Finland has a pertussis vaccination history very similar to that of The Netherlands, and yet there is no evidence for an increase in the incidence of pertussis to the extent that it was observed in The Netherlands. A comparison of the Bordetella pertussis strains circulating in the two countries may shed light on the differences in pertussis epidemiology. Here we investigated whether temporal changes had occurred in pertussis toxin and pertactin types produced by the Finnish B. pertussis population. We show that strains isolated before 1964 produced the same pertussis toxin and pertactin variants as the vaccine strains. However, these vaccine types were replaced in later years, and in the 1990s most strains were distinct from the vaccine strains with respect to the two proteins. These trends are similar to those found in the Dutch B. pertussis population. An interesting difference between the contemporary Finnish and Dutch B. pertussis populations was found in the frequencies of pertactin variants, possibly explaining the distinct epidemiology of pertussis in the two countries.     

Antibody response to pertussis toxin in patients with clinical pertussis measured by enzyme-linked immunosorbent assay

Serum antibody response to pertussis toxin was measured by enzyme-linked immunosorbent assay in 172 patients with clinical symptoms typical of whooping cough. The diagnosis was verified by culture in 100 patients. Serum antibodies were either not detectable or present only at low levels in sera obtained in the early stage of disease. Significant changes in serum levels of IgG, IgM and/or IgA were demonstrated in 143 patients (83 %). The lack of comparable increases in most of the other patients may be due to inappropriate timing of serum collection. Thus, detection of antibodies against pertussis toxin in paired serum samples can be used for serological diagnosis of pertussis. However, the presence of IgM and/or IgA in a single serum sample does not confirm a diagnosis of pertussis, since such antibodies were found in healthy adults as well as in patients two years after the disease. High levels of these antibodies are, however, suggestive of on-going or recent disease.     

Prevalence,diagnosis, and disease course of pertussis in adults with acute cough: a prospective,observational study in primary care

Background

Most cases of adult pertussis probably remain undiagnosed.

Aim

To explore the prevalence, diagnosis, and disease course of acute pertussis infection in adult patients presenting with acute cough.

Design and setting

Prospective observational study between 2007 and 2010 in primary care in 12 European countries.

Method

Adults presenting with acute cough (duration of ≤28 days) were included. Bordetella pertussis infection was determined by polymerase chain reaction (from nasopharyngeal flocked swabs and sputa) and by measurement of immunoglobulin G antibodies to pertussis toxin (PT) in venous blood at day 28. An antibody titre to PT of ≥125 IU/ml or PCR positive result in a respiratory sample defined recent infection. Patients completed a symptom diary for 28 days.

Results

Serum and/or respiratory samples were obtained in 3074 patients. Three per cent (93/3074) had recent B. pertussis infection. Prior cough duration >2 weeks discriminated to some extent between those with and without pertussis (adjusted odds ratio 1.89, 95% confidence interval = 1.17 to 3.07; P = 0.010). Median cough duration after presentation was 17 and 12 days in patients with and without pertussis, respectively (P = 0.008). Patients with pertussis had longer duration of phlegm production (P = 0.010), shortness of breath (P = 0.037), disturbed sleep (P = 0.013) and interference with normal activities or work (P = 0.033) after presentation.

Conclusion

Pertussis infection plays a limited role among adults presenting with acute cough in primary care, but GPs should acknowledge the possibility of pertussis in uncomplicated lower respiratory tract infection. As in children, pertussis also causes prolonged symptoms in adults. However, pertussis is difficult to discern from other acute cough syndromes in adults at first presentation.     

Resurgence of pertussis in Taiwan during 2009–2015 and its impact on infants

Background/purposePertussis incidence markedly decreased due to universal vaccination, but outbreaks had been noted worldwide in recent decade. This study was conducted to know the epidemiology of pertussis and its impact on infants in Taiwan.MethodsEpidemiologic parameters for confirmed pertussis cases were collected from the Taiwan Centers for Disease Control. The incidence of each age group over years was calculated using population data. We also did retrospective reviews of laboratory-confirmed pertussis cases in NTUH to analyze clinical characteristics and disease severity.ResultsA total of 668 confirmed pertussis cases were obtained from the Taiwan CDC open database between 2003 and 2017. There was higher incidence during the period 2009–2015, with a mean incidence of 0.27 cases per 100,000 population, about 2-fold increase compared with mean incidence of 0.12 cases per 100,000 population during the period 2003–2008. Infants accounted for the highest proportion of all cases (49.8%), with mean incidence of 16.1 cases per 100,000 people per year during 2009–2015, and a trend of increase was found from 2003 to 2015. In NTUH, a total of 17 laboratory-confirmed pertussis cases were diagnosed during 2012–2016, and 14 cases were young infants. Among them, 9 infants had been admitted to intensive care unit and 2 infant needed invasive ventilator support.ConclusionThere was a resurgence of pertussis during 2009–2015 and it had significant impact on infants. Young infants with pertussis may be severe and need intensive care, so preventive strategy may be advocated for them.     

Use of monoclonal fluorescent antibodies for the detection of B. pertussis and B. parapertussis

While it has been said that PCR is likely to replace DFA as the test of choice, accessibility to and cost of the technique may limit the number of laboratories that are able to implement the process. Still, the advantage of DFA staining relative to other diagnostic methods, including PCR, is the provision of a rapid result and lower overall cost. The laboratory diagnosis of pertussis remains problematic, and polyclonal DFA reagents for its detection have brought into question the utility of DFA. However, the integration of a commercially available monoclonal DFA reagent, in combination with the development and standardization of existing procedures, should provide clinicians with an improved method for the diagnosis and epidemiology of pertussis.