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Advani A.,Swedish Institute for Communicable Disease Control | Hallander H.O.,Swedish Institute for Communicable Disease Control | Dalby T.,Statens Serum Institute | Krogfelt K.A.,Statens Serum Institute | And 10 more authors.
Journal of Clinical Microbiology | Year: 2013

Between 1998 and 2009, Bordetella pertussis clinical isolates were collected during three periods, i.e., 1998 to 2001 (n=102), 2004 to 2005 (n=154), and 2007 to 2009 (n=140), from nine countries with distinct vaccination programs, i.e., Denmark, Finland, France, Germany, The Netherlands, Norway, Poland, Sweden, and the United Kingdom. Pulsed-field gel electrophoresis (PFGE) analysis was performed according to standardized recommendations for epidemiological typing of B. pertussis. There were 81 different PFGE profiles, five of which (BpSR3, BpSR5, BpSR10, BpSR11, and BpSR12) were observed in 61% of the 396 isolates and shown to be predominant in almost all countries. The major profile, BpSR11, showed a decreasing trend from 25% to 30% in 1998 to 2005 to 13% in 2007 to 2009, and there were increases in BpSR3 and BpSR10 from 0% and 8% to 21% and 22%, respectively. One difference between these profiles is that BpSR11 contains isolates harboring the fim3-2 allele and BpSR3 and BpSR10 contain isolates harboring the fim3-1 allele. The total proportion of the five predominant profiles increased from 44% in 1998 to 2001 to 63% in 2004 to 2005 to 70% in 2007 to 2009. In conclusion, common PFGE profiles were identified in B. pertussis populations circulating in European countries with different vaccination programs and different vaccine coverages. These prevalent isolates contain the novel pertussis toxin promoter ptxP3 allele. However, there is evidence for diversifying selection between ptxP3 strains characterized by distinct PFGE profiles. This work shows that, even within a relatively short time span of 10 years, successful isolates which spread through Europe and cause large shifts in B. pertussis populations may emerge. Copyright © 2013, American Society for Microbiology.


Guiso N.,Institute Pasteur Paris | Berbers G.,National Institute for Public Health and the Environment | Fry N.,Public Health England | He Q.,University of Turku | And 16 more authors.
European Journal of Clinical Microbiology and Infectious Diseases | Year: 2011

Bordetella pertussis-specific antibodies can be detected by enzyme-linked immunosorbent assays (ELISAs) or multiplex immunoassays. Assays use purified or mixed antigens, and only pertussis toxin (PT) is specific for B. pertussis. The interpretation of results can be based on dual-sample or single-sample serology using one or two cut-offs. The EU Pertstrain group recommends that: (i) ELISAs and multiplex immunoassays should use purified non-detoxified PT as an antigen, that they should have a broad linear range and that they should express results quantitatively in International Units per millilitre (IU/ml); (ii) a single or dual diagnostic cut-off for single-serum serology using IgG-anti-PT between 50 and 120 IU/ml should be used, and diagnostic serology cannot be validly interpreted for one year after vaccination with acellular pertussis (aP) vaccines; (iii) IgA-anti-PT should only be used with indeterminate IgG-anti-PT levels or when a second sample cannot be obtained. This group discourages using: (i) other antigens in routine diagnostics, as they are not specific; (ii) micro-agglutination, due to its lack of sensitivity; (iii) immunoblots for pertussis serodiagnosis, as results cannot be quantified; (iv) other methods, such as complement fixation or indirect immunofluorescence, due to their low sensitivity and/or specificity. © 2010 The Author(s).


Riffelmann M.,Labor Medizin Krefeld MVZ GmbH | Hunfeld K.-P.,Institute for Laboratory Medicine Microbiology and Infection Control | Hunfeld K.-P.,Society for Promoting Quality Assurance in Medical Laboratories E.V. INSTAND E.V. | Muller I.,Institute for Laboratory Medicine Microbiology and Infection Control | And 4 more authors.
European Journal of Clinical Microbiology and Infectious Diseases | Year: 2013

The purpose of this investigation was to test the performance of pertussis serology in diagnostic laboratories. The World Health Organization (WHO) Reference Reagent (06/142) and a sample with a low level of antibodies were sent to 200 participants of an external quality assessment (EQA) programme in Germany. The results were reported qualitatively and quantitatively, and were converted into IU/ml when possible. A total of 183 participants reported results. IgG, IgA and IgM enzyme-linked immunosorbent assays (ELISAs) with mixed antigens were used by 111, 110 and 113 participants, respectively, and 69 and 44 participants used IgG and IgA ELISAs with purified pertussis toxin (PT), respectively. IgG, IgA and IgM immunoblots were employed by 62, 63 and 11 participants, respectively. Most tests could distinguish between the positive and negative samples, but quantitative results were reported partly in non-comparable units. Only 37 % of participants used ELISAs that gave results comparable to the expected values in IU/ml and that could be interpreted according to published recommendations. © 2012 Springer-Verlag Berlin Heidelberg.


Riffelmann M.,Institute For Hygiene Und Labormedizin | Thiel K.,Zentrum fur Kinder und Jugendmedizin | Schmetz J.,Institute For Hygiene Und Labormedizin | Wirsing Von Koenig C.H.,Institute For Hygiene Und Labormedizin
Journal of Clinical Microbiology | Year: 2010

Measuring antibodies to Bordetella pertussis antigens is mostly done by enzyme-linked immunosorbent assays (ELISAs). We compared the performance of ELISA kits that were commercially available in Germany. Eleven measured IgG antibodies, and nine measured IgA antibodies. An in-house ELISA with purified antigens served as a reference method. Samples included two WHO reference preparations, the former Food and Drug Administration (FDA)/Center for Biologics Evaluation and Research (CBER) reference preparations, serum samples from patients with clinically suspected pertussis, and serum samples from patients having received a combined tetanus, diphtheria, and pertussis (Tdap) vaccination. Kits using pertussis toxin (PT) as an antigen showed linearity compared to the WHO Reference preparation (r2 between 0.82 and 0.99), and these kits could quantify antibodies according to the reference preparation. ELISA kits using mixed antigens showed no linear correlation to the reference preparations. Patient results were compared to results of in-house ELISAs using a dual cutoff of either ≥100 IU/ml anti-PT IgG or ≥40 IU/ml anti-PT IgG together with ≥12 IU/ml anti-PT IgA. The sensitivities of kits measuring IgG antibodies ranged between 0.84 and 1.00. The specificities of kits using PT as an antigen were between 0.81 and 0.93. The specificities of kits using mixed antigens were between 0.51 and 0.59 and were thus not acceptable. The sensitivities of kits measuring IgA antibodies ranged between 0.53 and 0.73, and the specificities were between 0.67 and 0.94, indicating that IgA antibodies may be of limited diagnostic value. Our data suggest that ELISAs should use purified PT as an antigen and be standardized to the 1st International Reference preparation. Copyright © 2010, American Society for Microbiology. All Rights Reserved.


Kennerknecht N.,Labor Medizin Krefeld Medizinisches Versorgungszentrum | Kennerknecht N.,Institute For Hygiene Und Labormedizin | Riffelmann M.,Labor Medizin Krefeld Medizinisches Versorgungszentrum | Riffelmann M.,Institute For Hygiene Und Labormedizin | And 3 more authors.
European Journal of Clinical Microbiology and Infectious Diseases | Year: 2011

Bordetella pertussis infection is mostly diagnosed by serological tests, such as by enzyme-linked immunosorbent assays (ELISAs) or by immunoblots. We compared immunoblots from five different manufacturers. Immunoblots from Euroimmun, Mikrogen, Trinity Biotech, Viramed and Virotech were used. All kits except the kit from Trinity Biotech measured IgG and IgA antibodies separately. The kits were used according to the kit inserts. Various reference preparations from the World Health Organization (WHO), the National Institute for Biological Standards and Control (NIBSC) and the Center for Biologics Evaluation and Research/Food and Drug Administration (CBER/FDA) were analysed. Patient sera with high antibody titres in ELISA, sera from patients with compatible clinical symptoms and sera from vaccinees were compared. An algorithm for interpreting quantitative values for IgG and IgA anti-pertussis toxin (PT) from in-house ELISAs was used as a reference. The sensitivity and specificity of the assays was variable when comparing the qualitative results of immunoblots with expected values of reference preparations and ELISA interpretation of patient sera. The interpretation of semi-quantitative reading of the immunoblots did not compare well to the ELISA results. Adenylate cyclase toxin as an additional antigen in two immunoblots did not effectively distinguish between infection and vaccination. Due to the lack of quantification of antibody concentrations, IgG and IgA immunoblots are of limited value in the serological diagnosis of pertussis. © 2011 Springer-Verlag.


Zeddeman A.,National Health Research Institute | van Gent M.,National Health Research Institute | Heuvelman C.J.,National Health Research Institute | van der Heide H.G.,National Health Research Institute | And 14 more authors.
Eurosurveillance | Year: 2014

Pathogen adaptation has been proposed to contribute to the resurgence of pertussis. A striking recent example is the emergence of isolates deficient in the vaccine component pertactin (Prn). This study explores the emergence of such Prn-deficient isolates in six European countries. During 2007 to 2009, 0/83 isolates from the Netherlands, 0/18 from the United Kingdom, 0/17 Finland, 0/23 Denmark, 4/99 Sweden and 5/20 from Norway of the isolates collected were Prn-deficient. In the Netherlands and Sweden, respectively 4/146 and 1/8 were observed in a later period (2010–12). The Prn-deficient isolates were genetically diverse and different mutations were found to inactivate the prn gene. These are indications that Prn-deficiency is subject to positive selective pressure. We hypothesise that the switch from whole cell to acellular pertussis vaccines has affected the balance between ‘costs and benefits’ of Prn production by Bordetella pertussis to the extent that isolates that do not produce Prn are able to expand. The absence of Prn-deficient isolates in some countries may point to ways to prevent or delay the spread of Prn-deficient strains. In order to substantiate this hypothesis, trends in the European B. pertussis population should be monitored continuously. © 2007-2013. All rights reserved.


Riffelmann M.,Labor Medizin Krefeld Medizinisches Versorgungszentrum GmbH | Riffelmann M.,Institute For Hygiene Und Labormedizin | Mohr J.,Zentrum fur Kinder und Jugendmedizin | Hellenbrand W.,Robert Koch Institute | Wirsing Von Koenig C.H.,Labor Medizin Krefeld Medizinisches Versorgungszentrum GmbH
European Journal of Clinical Microbiology and Infectious Diseases | Year: 2014

We evaluated whether the results of diagnostic polymerase chain reaction (PCR) testing combined with time since last vaccine dose could be used to monitor the effectiveness of acellular pertussis vaccines. In 258 consecutive nasopharyngeal swabs from children and adolescents with typical pertussis symptoms, 80 were positive and 178 were negative in PCR for Bordetella pertussis DNA (IS 481). Time since last vaccine dose was available for 152 patients, of which 120 were fully immunised. Among the fully vaccinated patients, the median age of 41 PCR-positive patients was 8.4 years (range 0.9-12.3) and that of 79 PCR-negative cases was 3.3 years (range 0.4-14.1) (p<0.01). The median time since last pertussis vaccine dose was 6.05 years [95 % confidence interval (CI): 0.5-10.9] in PCR-positive cases and 2.22 years (95 % CI: 0.04-9.23) in PCR-negative cases (p<0.001). The use of diagnostic PCR results from pertussis cases together with time since last vaccine dose permits estimates of the duration of protection after vaccination with acellular pertussis vaccines that are in keeping with more complex studies. © 2013 Springer-Verlag.


Guiso N.,Institute Pasteur Paris | Wirsing von Konig C.-H.,Institute For Hygiene Und Labormedizin | Forsyth K.,Flinders University | Tan T.,Northwestern University | Plotkin S.A.,University of Pennsylvania
Vaccine | Year: 2011

Pertussis remains endemic worldwide and is an important public health problem, even in countries with sustained high vaccination coverage. Resurgence of pertussis in the post-vaccination era has been reported in many areas of the world. The Global Pertussis Initiative (GPI) was established in 2001 to evaluate the ongoing problem of pertussis worldwide and to recommend appropriate pertussis control strategies. In addition to primary vaccinations, the GPI currently recommends a pertussis booster vaccination to pre-school children, adolescents and those adults at risk of transmitting. Bordetella pertussis infection to infants. At a meeting in Paris, France, in January 2010, GPI members discussed pertussis surveillance and testing then prepared recommendations on the implementation and utilisation of these activities. Issues and projects discussed included: national surveillance systems and their suitability for other countries; seroprevalence studies; ideal surveillance methodologies; ongoing efforts in obtaining biological samples; standardisation of sample treatment; culture; real-time polymerase chain reaction (PCR); and likely future advances such as antibody detection in saliva. Previous regional meetings of the GPI have confirmed that many countries have limited laboratory facilities for the detection of pertussis. The GPI hopes that the future introduction of increased laboratory capabilities and greater harmonisation of clinical definitions and detection methods will lead to enhanced surveillance and a better estimate of the burden of pertussis infection worldwide. This article provides a current guide on the appropriate use of laboratory diagnostics and optimal surveillance methodologies to assist countries in the control of pertussis disease. © 2010.


Sobotzki C.,Ludwig Maximilians University of Munich | Riffelmann M.,Labor Medizin Krefeld MVZ GmbH | Riffelmann M.,Institute For Hygiene Und Labormedizin | Kennerknecht N.,Institute For Hygiene Und Labormedizin | And 5 more authors.
Epidemiology and Infection | Year: 2016

Laboratory tests in adult outpatients with longer lasting coughs to identify a potential causal pathogen are rarely performed, and there is no gold standard for these diagnostic tests. While the diagnostic validity of serological tests for pertussis is well established their potential contribution for diagnosing adenovirus and influenza virus A and B infections is unclear. A sentinel study into the population-based incidence of longer lasting coughs in adults was done in Rostock (former East Germany) and Krefeld (former West Germany). A total of 971 outpatients who consulted general practitioners or internists were included. Inclusion criteria were coughing for 1 week and no chronic respiratory diseases. We evaluated the performance of polymerase chain reaction (PCR) as well as IgG and IgA serology, applying a latent class model for diagnosing infections with adenovirus, B. pertussis, and influenza virus A and B. The adult outpatients first sought medical attention when they had been coughing for a median of 3 weeks. In this situation, direct detection of infectious agents by PCR had a low sensitivity. Modelling showed that additional serological tests equally improved sensitivity and specificity for diagnosis for adenovirus, B. pertussis and influenza virus A and B infections. The combination of serology and PCR may improve the overall performance of diagnostic tests for B. pertussis and also for adenovirus, and influenza virus A and B infections. Copyright © Cambridge University Press 2015.


PubMed | Ludwig Maximilians University of Munich, Labor Medizin Krefeld MVZ GmbH, Institute For Hygiene Und Labormedizin, Landesamt fur Gesundheit und Soziales des Landes Mecklenburg Vorpommern and Trinity College Dublin
Type: Journal Article | Journal: Epidemiology and infection | Year: 2016

Laboratory tests in adult outpatients with longer lasting coughs to identify a potential causal pathogen are rarely performed, and there is no gold standard for these diagnostic tests. While the diagnostic validity of serological tests for pertussis is well established their potential contribution for diagnosing adenovirus and influenza virus A and B infections is unclear. A sentinel study into the population-based incidence of longer lasting coughs in adults was done in Rostock (former East Germany) and Krefeld (former West Germany). A total of 971 outpatients who consulted general practitioners or internists were included. Inclusion criteria were coughing for 1 week and no chronic respiratory diseases. We evaluated the performance of polymerase chain reaction (PCR) as well as IgG and IgA serology, applying a latent class model for diagnosing infections with adenovirus, B. pertussis, and influenza virus A and B. The adult outpatients first sought medical attention when they had been coughing for a median of 3 weeks. In this situation, direct detection of infectious agents by PCR had a low sensitivity. Modelling showed that additional serological tests equally improved sensitivity and specificity for diagnosis for adenovirus, B. pertussis and influenza virus A and B infections. The combination of serology and PCR may improve the overall performance of diagnostic tests for B. pertussis and also for adenovirus, and influenza virus A and B infections.

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