Invasive meningococcal disease: still challenging

1
Invasive meningococcal disease still challenging

• Pavla Krizova
• National Reference Laboratory
• for Meningococcal Infections,
• NIPH, Prague, Czech Republic
• pavla.krizova_at_szu.cz
• http//www.szu.cz/cema/hpcema.htm

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200 years history

• 1805 first reported outbreaks in Geneva
• 1806 reported outbreaks in New England
• 1840 first reported epidemics in Africa
• 1887 Weichselbaum identified diplococcus
  intracellularis meningitidis

Cartwright, 1995 Invasive meningococcal disease
is one of the most aggressive of the infectious
diseases of humans caused by bacteria.
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• Healthy carrier
• 10 in healthy population
• herd immunity

• Invasive meningococcal disease
• meningococcal meningitis
• Waterhause-Friderichsen syndrome
• meningococcaemia/sepsis
• localised meningococcal disease
• endocarditis
• pericarditis
• pneumonia
• appendicitis

Carriership is a normal host-bacterium
relationship of meningococcus.
? recovery sequelae complications death
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Meningococcal infection

• Characterisation of N. meningitidis
• Laboratory diagnosis of IMD
• Molecular epidemiology
• Virulence of N. meningitidis
• Epidemiology of IMD
• Risk factors for IMD
• Healthy carriers of N. meningitidis
• Herd immunity against N. meningitidis
• Treatment of IMD
• Measures recommended for contacts
• Vaccination

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Laboratory methods of meningococcal laboratory
in Prague

• 1970 N. m. identification, serogrouping, ATB
  susceptibility testing
• 1970 direct Latex-agglutination from clinical
  material
• 1976 serotyping (Ouchterlony, WCE)
• 1980 Ab measurement bactericidal assay, ELISA
• 1993 MLEE
• 1998 PFGE
• 1999 behavior of virulent/non-virulent
  meningococci on cell cultures
• 2000 PCR from clinical material
• 2000 MLST
• 2001 MLSTPCR
• 2002 RAPD
• 2004 porA, porB
• 2005 fetA

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Participation in International QAS

• Poolman J.T., Kriz P., Ashton F., Bibb W.,
  Dankert J., Demina A., Froholm L.O., Hassan-King.
  M., Jones D.M., Lind I., Prakash K., Xujing H.
  Serotypes and subtypes of Neisseria meningitidis
  Results of an international study comparing
  sensitivities and specifities of monoclonal
  antibodies. Clin. Diagn. Labor. Immunol. 1995,
  2(1) 69-72.
• Maslanka S.E., Gheesling L.L., Libutti D.E.,
  Donaldson K.B.J., Harakeh H.S., Dykes J.K., Arhin
  F.F., Devi S.J.N., Frasch C.E., Huang J.C.,
  Kriz-Kuzemenska P., Lemmon R.D., Lorange M.,
  Peeters C.A.M., Quataert S., Tai J.Y., Carlone
  G.M. Standardization and a multilaboratory
  comparison of Neisseria meningitidis serogroup A
  and C serum bactericidal assays. Clin. Diagn.
  Labor. Immunol. 1997, 4(2) 156-167.
• Vázquez J. A., Arreaza L., Block C., Ehrhard I.,
  Gray S. J., Heuberger S., Hoffmann S., Kriz P.,
  Nicolas P., Olcen P., Skoczynska A., Spanjaard
  L., Stefanelli P., Taha M.K., Tzanakaki G.
  Interlaboratory Comparison of Agar Dilution and
  E-test Methods for Determining the MICs of
  Antibiotics Used in Management of Neisseria
  meningitidis Infections. Antimicrob Agents and
  Chemother 2003, 47(11) 3430-3434.
•    Taha M.K., Alonso J.M., Cafferkey M., Caugant
  D.C., Clarje S.C., Diggle M.A., Fox A., Frosch
  M., Gray S.J., Guiver M., Heuberger S., Kalmusova
  J., Kesanopoulos K., Klem A.M., Kriz P., Marsh
  J., Molling P., Murphy K., Olcén P., Sanou O.,
  Tzanakaki G., Vogel U. Interlaboratory comparison
  of PCR-based identification and genogrouping of
  Neisseria meningitidis. J Clin Microbiol.
  200543(1)144-9.

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International collaboration of meningococcal
laboratory in Prague

• 1970 meningococcal laboratory in the NIPH created
• 1980 National Reference Laboratory for
  Meningococcal Infections
• 1987-1998 European Surveillance of bacterial
  meningitis
• since 1993 EMGM http//neisseria.org/nm/emgm/meet
  ings/
• 2005 8th EMGM in Dublin
• 1994 new serotype 22 Mab reference strain
  hybridoma to NIBSC
• 1999 MLST project with University of Oxford
• since 1999 EU-IBIS http//www.euibis.org/
• 2001-2005 EU project MenNet http//neisseria.org/n
  m/emgm/eumennet/

molecular methods 2005 80 meningo NRL in
Europe 2007 100
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Collection of N. meningitidis isolated in the
Czech Republic

• since 1970 strains isolated from
• invasive meningococcal disease 1620 strains
• contacts of IMD 610 strains
• healthy carriers 1430 strains
• respiratory infections 440 strains
• others 70 strains
• unknown 140 strains
• TOTAL 4310 strains covering more than 30 years

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Characterisation of meningococcal strains in the
collection of the National Reference Laboratory
for Meningococcal Infections in Prague 4310
strains covering more than 30 yearslaboratory
epidemiological data are computerized

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Laboratory diagnosis of invasive meningococcal
disease in the Czech Republic

• Classical diagnosis
• direct microscopy
• cultivation
• direct latex agglutination
• serology increased antibodies in reconvalescent
  serum
• PCR diagnosis
• MLST directly from clinical material
• Kriz P., Kalmusova J., Felsberg J. Multilocus
  sequence typing of
• Neisseria meningitidis directly from
  cerebrospinal fluid.
• Epidemiol. Infect. 2002, 128, 157-160.

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Laboratory confirmation of invasive meningococcal
disease Czech Republic, 1993 2006 (May)
MLST from PCR products in culture-negative IMD
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MLST

• 100 characterisation of strains
• the most valuable epidemiological marker
  sequence type ST
• internet portable and comparable data
• global epidemiology
• evolutionary analysis of bacterial populations
• detection of hypervirulent complexes

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Database statistics for Neisseria PubMLST
database http//pubmlst.org/neisseria/ - access
June 14, 2006
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• Populations of meningococci from patients versus
  carriers are different.
• Population study of meningococcal carriage
  strains showed that this population is highly
  diverse and most nucleotide sequence diversity
  resulted from the reassortment of alleles by
  horizontal genetic exchange.
• Jolley K.A., Kalmusova J., Feil E.J., Gupta S.,
  Musilek M., Kriz P., Maiden M.J. Carried
  Meningococci in the Czech Republic a Diverse
  Recombining Population.
• J. Clin. Microbiol., 2000, 38 4492-4498.

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• The genetic differentiation between disease and
  carriage isolates indicated that, whereas certain
  genotypes were overrepresented among disease
  isolates ("hyperinvasive" lineages),
  disease-associated and carried meningococci
  exhibited remarkably little differentiation at
  the level of individual nucleotide polymorphisms.
  
•  
• Jolley KA, Wilson DJ, Kriz P, McVean G, Maiden
  MC. The Influence of Mutation, Recombination,
  Population History, and Selection on Patterns of
  Genetic Diversity in Neisseria meningitidis. Mol
  Biol Evol. 2005, 22(3)562-569.
•  

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Difference of Czech meningococcal population was
probable before MLST availability

• High of NT/NST meningococci by WCE
• New serotype of N.m.B discovered by NRL in Prague
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• Reference strain hybridoma to NIBSC in 1996
• Kríová P., Musílek M., Danielová V., Holubová
  J. New serotype candidate of Neisseria
  meningitidis. Cent. Eur. J. Public. Health. 1996,
  4(3) 169-172.
• Tzanakaki G., Kriz P., Kremastinou J., Musílek
  M., Smart L.E., Blackwell C.C. Reactivity of the
  new monoclonal antibody 22 with meningococcal
  strains isolated from patients and carriers in
  Greece. FEMS Immunol. Med. Microbiol. 1997, 19
  1-5.
• Musilek M., Giorgini D., Hamadouche N., Kriz P.,
  Taha M.K. Genetic heterogeneity of strains of
  Neisseria meningitidis belonging to serotype 22
  isolated in the Czech Republic. J. Clin.
  Microbiol. 1998, 36(2) 563-565.
• Urwin R., Fox A.J., Musilek M., Kriz P., Maiden
  M.C.J. Heterogeneity of the PorB protein in
  serotype 22 Neisseria meningitidis. J. Clin.
  Microbiol. 1998, 36(12) 3680-3682.

• Eastern Germany
• Russian immigrants to Greece
• Poland

  Pizza M. et al., Identification of vaccine
candidates against serogroup B meningococcus by
whole-genome sequencing. Science.
2000287(5459)1816-20.
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Kríová, P., J. Kalmusová, M. Musílek. Invazivní
meningokokové onemocnení v Ceské republice v roce
2005. Zprávy CEM (SZÚ Praha) 200615(3-4),
45-51. htpp//www.szu.cz/cema/hpcema.htm
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• Higher and quicker adhesion to epithelial cells
  with reduced phagocytosis as revealed in strains
  isolated from invasive meningococcal disease
  compared to those from healthy carriers also
  confirmed diversity of these meningococcal
  populations.

Kalmusova J, Novotny J, Hulinska D, Musilek M
and Kriz P. Interactions of invasive and
non-invasive strains of Neisseria meningitidis
with monkey epithelial cells, mouse monocytes
and human macrophages. Microbiologica, 2000,
23185-200.
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Invasive meningococcal disease in the Czech
Republic, 1943-2005morbidity per 100 000
population
NRL for meningococcal infections
active surveilance
ET-15/37, ST-11
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IMD age specific morbidityCzech Republic, 1992
- 2005
N.m.B
N.m.C
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Invasive Meningococcal Disease in the Czech
Republic, 19922006 (May 15)
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IMD emergency situation in the Czech Republic in
1993

• increased IMD morbidity 1.3/100000
• high IMD case fatality rate 10 (20)
• age shift of IMD patients 15-19 y
• severe/unusual clinical symptoms of IMD sepsis,
  W-F
• antigenic shift of N. meningitidis C2aP1.2,5
• genetic shift of N. meningitidis ET-15/37,
  ST11-complex
• Kríová P., Musílek M. Changing epidemiology of
  meningococcal invasive disease in the Czech
  Republic caused by new clone Neisseria
  meningitidis C2aP1.2(P1.5), ET-15/37.
• Cent. Eur. J. Public Health. 1995, 3(4)
  189-194.
• Kriz P. Surveillance of invasive meningococcal
  disease in the Czech Republic.
• Eurosurveillance Monthly, 2004, 9 (11), 11-12.

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Distribution of serogroups of Neisseria
meningitidis isolated from invasive meningococcal
disease, Czech Republic, 1970-2005
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Risk factors

• socio-economic
• poor living conditions
• overcrowding
• household contacts
• close contacts
• siblings
• damage to respiratory musoca
• harmattan in Africa
• antecedent viral infections
• smoking
• immunological
• low levels of IgG/IgM bactericidal antibodies
• complement deficiency

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Case control prospective study for risk factors
in the Czech Republic in 1990th

• The risk of IMD is strongly associated with
  parental smoking
• Risk ratios adjusted for smoking
• 3.5 (1.4-8.7) for smoking of mother
• 3.2 (1.5-6.9) for smoking of father
• 8.23 (2.30-29.3) for smoking of both parents
• 2.7 (1.3-5.4) for every 20 cigarettes smoked at
  home on an average day
• The risk of IMD is inversely related to
• maternal education
• favourable socioeconomic conditions
• fathers education
• Kriz P., Bobak M., Kriz B. Parental smoking,
  socioeconomic factors, and risk of invasive
  meningococcal disease in children a population
  based case-control study.
• Arch. Dis. Child. 2000, 83 117-121.

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Healthy carriers of N. meningitidis

• percentage
• duration
• multiplicity
• exchange of genetic information between carried
  strains
• meningococcal lineages?
• carriage lineages
• IMD lineages, hypervirulent lineages

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IMD versus carrier populations

• Meningococcal isolates from invasive
  meningococcal disease and carriers in the Czech
  Republic, Greece and Norway were compared.
• Differences in the distribution of meningococcal
  genotypes between disease and carrier isolates
  and among countries were found.
• Yazdankhah, P. S., P. Kriz, G. Tzanakaki, J.
  Kremastinou, J. Kalmusova, M. Musilek, T.
  Alvestad, K. A. Jolley, D. J. Wilson, N. D.
  McCarthy, D. A. Caugant and M. C. J. Maiden.
  Distribution of serogroups and genotypes among
  diseaes-associated and carried isolates of
  Neisseria meningitidis from the Czech Republic,
  Greece and Norway. J. Clin. Microbiol. 2004, 42
  (11) 5146-5153.

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Prospective carriage study

• October 2002 March 2003 6 months
• 206 students sampled monthly
• 190 students were present in all 6 samplings
  92.2
• nasopharyngeal laryngeal swabs
• inoculation to selective medium
• questionnaires (ATB, smoking)
• each colony was investigated and stored
• in case of massive primoculture, the maximum of
  colonies investigated and stored from one carrier
  was 20 (47 in one case)
• RAPD (1242 isolates)
• MLST (35 isolates)
• serogroup, serotype subtype, ATB (35 isolates)

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Conclusions of recent carriage study

• Carriage of N. meningitidis has a long duration
  
• 19 carriers of 33 (57.6).
• Carried lineages are stable.
• Carriage of multiple lineages is rare
• 2 carriers of 33 (6.1).
• Carriage of one lineage do not protect against
  colonization by other lineage.
• Meningococcal population isolated from carriers
  showed a heterogeneity in RAPD patterns and
  phenotypes.
• Hypervirulent ST-complexes are rare among strains
  from carriers.

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Bactericidal antibodies against N. meningitidis
A, B and C (percentage of sera with protective
level) Czech Republic, 2001
Herd immunity in the Czech Republic
A 1027 - A4, 21P1.10 B 16B6 - B2aP1.2,P1.5 C
7606 - C2aP1.2,P1.5
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Bactericidal antibodies against N. meningitidis
C2aP1.2,5 (percentage of sera with protective
level) Czech Republic, 1989, 1996, 2001
Herd immunity in the Czech Republic
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Herd immunity in the Czech Republic - conclusion

• Hypervirulent complex ST-11 enhanced herd
  immunity against this meningococcal complex among
  the healthy Czech population.
• However, naturally occurring antimeningococcal
  antibodies are not able to stop the spread of the
  hypervirulent complex in the population.
• Kriz P., Kriz B., Svandova E., Musilek M.
  Antimeningococcal herd immunity in the Czech
  Republic - influence of an emerging clone,
  Neisseria meningitidis ET-15/37.
• Epidemiol. Infect. 1999, 123193-200.
• Kríová P. Immunological survey in the Czech
  Republic in 2001 invasive meningococcal
  disease. Cent. Eur. J. Public Health. 2003, 11,
  Supplement S62-S67.

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Treatment of IMD

• Preadmission ATB
• case fatality rate reduced
  twice
• Ronovský L., Kríová P. , truncová V., Dostál
  V., Plíek S., Kasal E., Burget I., Chalupa P.,
  Dlouhý P. Administration of antibiotics before
  admission in patients with meningococcal disease.
  Cent. Eur. J. Public Health. 2003, 11 (1),
  14-18.
• L. Ronovský et al., Standard of effective
  preadmission care for suspected IMD. Urgentní
  medicína, 3/2003 18-20 (in the Czech language).
• E. Kasal et al., guidelines for the treatment of
  invasive meningococcal disease. Praktický lékar,
  2005, 85, 4, 196-199 (in the Czech language).

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Case fatality rate of IMDCzech Republic, 1993
2005
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Prevention of IMD in the Czech Republic

• intensive medical surveillance of contacts (1
  week)
• limitation of physical effort of contacts (1
  week)
• limitation of exposure in contacts to infection
  (1 week)
• protective chemotherapy (PNC)
• immediately
• to risk contacts only
• vaccination of contacts (after one week)
• No secondary cases of IMD in the Czech Republic.
• Measures in a focus of IMD
• Bulletin, Ministry of Health of the Czech
  Republic, 1993

Actualisation of the Surveillance programmes in
2006
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Vaccination in Europe

• Conjugated vaccine against N. m. C is highly
  effective.
• It is desirable to replace the polysaccharide AC
  vaccine with conjugated Men C vaccine, with the
  exception of travellers.
• The usefulness of conjugate tetravalent vaccine
  ACYW135 for Europe is discussed.
• Effective vaccine against Men B is desirable.
• Universal vaccine against all meningococci is
  desirable.

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MenC vaccine in Europe

• excellent efficacy of MenC conjugate vaccine in
  Europe
• probably no capsule switching from MenC to MenB
  after MenC vaccination

• more than 90 reduction of morbidity from N.m.C
  IMD
• lower carriage of N.m.C in healthy population

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Progress during last 30 years

• Characterisation of N. meningitidis MLST
• factors of virulence
• genome defined
• Laboratory diagnosis of IMD PCR, PCR
  MLST
• Molecular epidemiology hypervirulent complexes
• Risk factors of IMD some of them defined
• Treatment of IMD preadmission ATB
• Prophylaxis of IMD measures for contacts
• Prevention of IMD MenC conjugate vaccine
• vaccination strategy

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Challenge in the 21st century

• Treatment of IMD reduction of CFR
• Prevention of IMD reduction of secondary
  cases reduction of incidence

universal meningococcal vaccine vaccination
strategy
Healthy carrier or patient ???
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Acknowledgement

• EMGM
• EU-IBIS
• MenNet
• NRL for Meningococcal Infections, NIPH, Prague
• Dr. Jitka Kalmusova Dr. Martin Musilek
• Alena Halouzkova
• Vlasta Pavlikova
• Renata Pospisilova
• Monika Heroldova

• Center of Epidemiology and Microbiology, NIPH,
  Prague
• Microbiological Institute of Academy of Science
• collaborators on Czech research projects on
  surveillance of IMD

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Happy stay in Prague
Thanks for your attention