Skip directly to site content Skip directly to page options Skip directly to A-Z link Skip directly to A-Z link Skip directly to A-Z link
Volume 24, Number 3—March 2018

Capsule Typing of Haemophilus influenzae by Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry1

Viktor MånssonComments to Author , Janet R. Gilsdorf, Gunnar Kahlmeter, Mogens Kilian, J. Simon Kroll, Kristian Riesbeck2, and Fredrik Resman2
Author affiliations: Lund University, Malmö, Sweden (V. Månsson, K. Riesbeck, F. Resman); University of Michigan, Ann Arbor, Michigan, USA (J.R. Gilsdorf); Växjö Hospital, Växjö, Sweden (G. Kahlmeter); Aarhus University, Aarhus, Denmark (M. Kilian); Imperial College London, London, UK (J.S. Kroll)

Main Article

Table 1

Validation of the original MALDI-TOF mass spectrometry capsule typing database (n = 31) by classification of the remaining 227 isolates in the evaluation set and 4 capsule transformants of Haemophilus influenzae*

Capsule type No. No. correct† No. inconclusive‡ No. incorrect§ Sensitivity, % Specificity, %
Hia, ST21-related 4 4 0 0 100 99.1
Hia, ST23-related 1 1 0 0 100 100
Hib, ST6-related 17 17 0 0 100 100
Hib, ST222-related 1 1 0 0 100 100
Hic 2 2 0 0 100 100
Hid 1 1 0 0 100 98.2
Hie 12 12 0 0 100 97.7
Hif 23 23¶ 0 0 100 99.0
All encapsulated isolates, a–f 61 61 0 0 100 92.2
Nontypeable 166 122 31 13# 73.5 100
Rb-negative capsule transformants 4 1** 0 3** NA NA

*Hia, H. influenzae type a; Hib, H. influenzae type b; Hic, H. influenzae type c; Hid, H. influenzae type d; Hie, H. influenzae type e; Hif, H. influenzae type f; M ALDI-TOF, matrix-assisted laser desorption/ionization time-of-flight; NA, not applicable; NTHi, nontypeable H. Influenzae; ST, sequence type.
>5/6 spectra classified to the same correct type.
<4/6 spectra classified to the same type.
§>5/6 spectra classified to the same incorrect type.
¶All classified to the established ST124-related Hif lineage.
#Two isolates classified as Hia (ST21-related), 4 as Hid, 5 as Hie, and, 2 as Hif (ST124-related).
**All classified as type d (i.e., the same type as the parental strain Rd) (30,31), resulting in correct classification of isolate Rb–/d+:02 and incorrect classification of isolates Rb+:02, Rb–/a+:02, and Rb–/c+:02.

Main Article

  1. Pittman  M. Variation and type specificity in the bacterial species Haemophilus influenzae. J Exp Med. 1931;53:47192. DOIPubMedGoogle Scholar
  2. Peltola  H. Worldwide Haemophilus influenzae type b disease at the beginning of the 21st century: global analysis of the disease burden 25 years after the use of the polysaccharide vaccine and a decade after the advent of conjugates. Clin Microbiol Rev. 2000;13:30217. DOIPubMedGoogle Scholar
  3. Langereis  JD, de Jonge  MI. Invasive disease caused by nontypeable Haemophilus influenzae. Emerg Infect Dis. 2015;21:17118. DOIPubMedGoogle Scholar
  4. Resman  F, Ristovski  M, Ahl  J, Forsgren  A, Gilsdorf  JR, Jasir  A, et al. Invasive disease caused by Haemophilus influenzae in Sweden 1997-2009; evidence of increasing incidence and clinical burden of non-type b strains. Clin Microbiol Infect. 2011;17:163845. DOIPubMedGoogle Scholar
  5. Ladhani  S, Slack  MP, Heath  PT, von Gottberg  A, Chandra  M, Ramsay  ME; European Union Invasive Bacterial Infection Surveillance participants. Invasive Haemophilus influenzae Disease, Europe, 1996-2006. Emerg Infect Dis. 2010;16:45563. DOIPubMedGoogle Scholar
  6. Whittaker  R, Economopoulou  A, Dias  JG, Bancroft  E, Ramliden  M, Celentano  LP; European Centre for Disease Prevention and Control Country Experts for Invasive Haemophilus influenzae Disease. Epidemiology of Invasive Haemophilus influenzae Disease, Europe, 2007-2014. Emerg Infect Dis. 2017;23:396404. DOIPubMedGoogle Scholar
  7. Bender  JM, Cox  CM, Mottice  S, She  RC, Korgenski  K, Daly  JA, et al. Invasive Haemophilus influenzae disease in Utah children: an 11-year population-based study in the era of conjugate vaccine. Clin Infect Dis. 2010;50:e416. DOIPubMedGoogle Scholar
  8. Bruce  MG, Zulz  T, DeByle  C, Singleton  R, Hurlburt  D, Bruden  D, et al. Haemophilus influenzae serotype a invasive disease, Alaska, USA, 1983-2011. Emerg Infect Dis. 2013;19:9327. DOIPubMedGoogle Scholar
  9. Zanella  RC, Bokermann  S, Andrade  AL, Flannery  B, Brandileone  MC. Changes in serotype distribution of Haemophilus influenzae meningitis isolates identified through laboratory-based surveillance following routine childhood vaccination against H. influenzae type b in Brazil. Vaccine. 2011;29:893742. DOIPubMedGoogle Scholar
  10. Rotondo  JL, Sherrard  L, Helferty  M, Tsang  R, Desai  S. The epidemiology of invasive disease due to Haemophilus influenzae serotype a in the Canadian North from 2000 to 2010. Int J Circumpolar Health. 2013;72:72. DOIPubMedGoogle Scholar
  11. Bajanca-Lavado  MP, Simões  AS, Betencourt  CR, Sá-Leão  R; Portuguese Group for Study of Haemophilus influenzae invasive infection. Characteristics of Haemophilus influenzae invasive isolates from Portugal following routine childhood vaccination against H. influenzae serotype b (2002-2010). Eur J Clin Microbiol Infect Dis. 2014;33:60310. DOIPubMedGoogle Scholar
  12. Ladhani  SN, Collins  S, Vickers  A, Litt  DJ, Crawford  C, Ramsay  ME, et al. Invasive Haemophilus influenzae serotype e and f disease, England and Wales. Emerg Infect Dis. 2012;18:72532. DOIPubMedGoogle Scholar
  13. Ladhani  S, Heath  PT, Slack  MP, McIntyre  PB, Diez-Domingo  J, Campos  J, et al.; Participants of the European Union Invasive Bacterial Infections Surveillance Network. Haemophilus influenzae serotype b conjugate vaccine failure in twelve countries with established national childhood immunization programmes. Clin Microbiol Infect. 2010;16:94854. DOIPubMedGoogle Scholar
  14. Ladhani  SN. Two decades of experience with the Haemophilus influenzae serotype b conjugate vaccine in the United Kingdom. Clin Ther. 2012;34:38599. DOIPubMedGoogle Scholar
  15. Juarez  MD, Rancaño  C, Neyro  S, Biscayart  C, Katz  N, Pasinovich  M, et al. What’s happening with Haemophilus influenzae type B invasive disease in Latin America region? Argentina’s experience. In: Abstracts of IDWeek 2016, New Orleans, October 26–30. Abstract no. 768 [cited 2017 Dec 18].
  16. World Health Organization. Global and regional immunization profile, 2016 global summary [cited 2017 Nov 9].
  17. Musser  JM, Kroll  JS, Moxon  ER, Selander  RK. Clonal population structure of encapsulated Haemophilus influenzae. Infect Immun. 1988;56:183745.PubMedGoogle Scholar
  18. Meats  E, Feil  EJ, Stringer  S, Cody  AJ, Goldstein  R, Kroll  JS, et al. Characterization of encapsulated and noncapsulated Haemophilus influenzae and determination of phylogenetic relationships by multilocus sequence typing. J Clin Microbiol. 2003;41:162336. DOIPubMedGoogle Scholar
  19. Erwin  AL, Sandstedt  SA, Bonthuis  PJ, Geelhood  JL, Nelson  KL, Unrath  WC, et al. Analysis of genetic relatedness of Haemophilus influenzae isolates by multilocus sequence typing. J Bacteriol. 2008;190:147383. DOIPubMedGoogle Scholar
  20. University of Oxford. Haemophilus influenzae MLST website [cited 2017 Nov 9].
  21. Myers  AL, Jackson  MA, Zhang  L, Swanson  DS, Gilsdorf  JR. Haemophilus influenzae type b invasive disease in Amish children, Missouri, USA, 2014. Emerg Infect Dis. 2017;23:1124. DOIPubMedGoogle Scholar
  22. LaClaire  LL, Tondella  ML, Beall  DS, Noble  CA, Raghunathan  PL, Rosenstein  NE, et al.; Active Bacterial Core Surveillance Team Members. Identification of Haemophilus influenzae serotypes by standard slide agglutination serotyping and PCR-based capsule typing. J Clin Microbiol. 2003;41:3936. DOIPubMedGoogle Scholar
  23. Satola  SW, Collins  JT, Napier  R, Farley  MM. Capsule gene analysis of invasive Haemophilus influenzae: accuracy of serotyping and prevalence of IS1016 among nontypeable isolates. J Clin Microbiol. 2007;45:32308. DOIPubMedGoogle Scholar
  24. Falla  TJ, Crook  DW, Brophy  LN, Maskell  D, Kroll  JS, Moxon  ER. PCR for capsular typing of Haemophilus influenzae. J Clin Microbiol. 1994;32:23826.PubMedGoogle Scholar
  25. Davis  GS, Sandstedt  SA, Patel  M, Marrs  CF, Gilsdorf  JR. Use of bexB to detect the capsule locus in Haemophilus influenzae. J Clin Microbiol. 2011;49:2594601. DOIPubMedGoogle Scholar
  26. Lâm  TT, Elias  J, Frosch  M, Vogel  U, Claus  H. New diagnostic PCR for Haemophilus influenzae serotype e based on the cap locus of strain ATCC 8142. Int J Med Microbiol. 2011;301:1769. DOIPubMedGoogle Scholar
  27. van Ketel  RJ, de Wever  B, van Alphen  L. Detection of Haemophilus influenzae in cerebrospinal fluids by polymerase chain reaction DNA amplification. J Med Microbiol. 1990;33:2716. DOIPubMedGoogle Scholar
  28. Clark  AE, Kaleta  EJ, Arora  A, Wolk  DM. Matrix-assisted laser desorption ionization-time of flight mass spectrometry: a fundamental shift in the routine practice of clinical microbiology. Clin Microbiol Rev. 2013;26:547603. DOIPubMedGoogle Scholar
  29. Månsson  V, Resman  F, Kostrzewa  M, Nilson  B, Riesbeck  K. Identification of Haemophilus influenzae type b isolates by use of matrix-assisted laser desorption ionization−time of flight mass spectrometry. J Clin Microbiol. 2015;53:221524. DOIPubMedGoogle Scholar
  30. Zwahlen  A, Kroll  JS, Rubin  LG, Moxon  ER. The molecular basis of pathogenicity in Haemophilus influenzae: comparative virulence of genetically-related capsular transformants and correlation with changes at the capsulation locus cap. Microb Pathog. 1989;7:22535. DOIPubMedGoogle Scholar
  31. Alexander  HE, Leidy  G. Determination of inherited traits of H. influenzae by desoxyribonucleic acid fractions isolated from type-specific cells. J Exp Med. 1951;93:34559. DOIPubMedGoogle Scholar
  32. Darriba  D, Taboada  GL, Doallo  R, Posada  D. jModelTest 2: more models, new heuristics and parallel computing. Nat Methods. 2012;9:772. DOIPubMedGoogle Scholar
  33. Guindon  S, Gascuel  O, Rannala  B. A simple, fast, and accurate algorithm to estimate large phylogenies by maximum likelihood. Syst Biol. 2003;52:696704. DOIPubMedGoogle Scholar
  34. López-Fernández  H, Santos  HM, Capelo  JL, Fdez-Riverola  F, Glez-Peña  D, Reboiro-Jato  M. Mass-Up: an all-in-one open software application for MALDI-TOF mass spectrometry knowledge discovery. BMC Bioinformatics. 2015;16:318. DOIPubMedGoogle Scholar
  35. Kehrmann  J, Wessel  S, Murali  R, Hampel  A, Bange  FC, Buer  J, et al. Principal component analysis of MALDI TOF MS mass spectra separates M. abscessus (sensu stricto) from M. massiliense isolates. BMC Microbiol. 2016;16:24. DOIPubMedGoogle Scholar
  36. Rizzardi  K, Åkerlund  T. High molecular weight typing with MALDI-TOF MS: a novel method for rapid typing of Clostridium difficile. PLoS One. 2015;10:e0122457. DOIPubMedGoogle Scholar
  37. Camoez  M, Sierra  JM, Dominguez  MA, Ferrer-Navarro  M, Vila  J, Roca  I. Automated categorization of methicillin-resistant Staphylococcus aureus clinical isolates into different clonal complexes by MALDI-TOF mass spectrometry. Clin Microbiol Infect. 2016;22:161.e17. DOIPubMedGoogle Scholar
  38. Josten  M, Reif  M, Szekat  C, Al-Sabti  N, Roemer  T, Sparbier  K, et al. Analysis of the matrix-assisted laser desorption ionization-time of flight mass spectrum of Staphylococcus aureus identifies mutations that allow differentiation of the main clonal lineages. J Clin Microbiol. 2013;51:180917. DOIPubMedGoogle Scholar
  39. Ojima-Kato  T, Yamamoto  N, Suzuki  M, Fukunaga  T, Tamura  H. Discrimination of Escherichia coli O157, O26 and O111 from other serovars by MALDI-TOF MS based on the S10-GERMS method. PLoS One. 2014;9:e113458. DOIPubMedGoogle Scholar
  40. Fothergill  LD, Chandler  CA, Spencer  M. Observations on the dissociation of meningitic strains of H. influenzae. J Immunol. 1936;31:40115.
  41. Hoiseth  SK, Connelly  CJ, Moxon  ER. Genetics of spontaneous, high-frequency loss of b capsule expression in Haemophilus influenzae. Infect Immun. 1985;49:38995.PubMedGoogle Scholar
  42. Tsang  RS, Li  YA, Mullen  A, Baikie  M, Whyte  K, Shuel  M, et al. Laboratory characterization of invasive Haemophilus influenzae isolates from Nunavut, Canada, 2000-2012. Int J Circumpolar Health. 2016;75:29798. DOIPubMedGoogle Scholar
  43. Sauget  M, Valot  B, Bertrand  X, Hocquet  D. Can MALDI-TOF mass spectrometry reasonably type bacteria? Trends Microbiol. 2017;25:44755. DOIPubMedGoogle Scholar

Main Article

1Preliminary results from this study were presented at the IDWeek 2016 Conference, October 26−30, 2016, New Orleans, Louisiana, USA.

2These senior authors contributed equally to this article.

Page created: February 15, 2018
Page updated: February 15, 2018
Page reviewed: February 15, 2018
The conclusions, findings, and opinions expressed by authors contributing to this journal do not necessarily reflect the official position of the U.S. Department of Health and Human Services, the Public Health Service, the Centers for Disease Control and Prevention, or the authors' affiliated institutions. Use of trade names is for identification only and does not imply endorsement by any of the groups named above.