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Two variants among Haemophilus influenzae serotype b strains with distinct bcs4, hcsA and hcsB genes display differences in expression of the polysaccharide capsule.

Schouls L, van der Heide H, Witteveen S, Zomer B, van der Ende A, Burger M, Schot C - BMC Microbiol. (2008)

Bottom Line: The DNA sequences of the complete capsular gene clusters of 9 Dutch Hib strains were assessed and two variants, designated type I and type II were found.NMR analysis of type I and type II capsule polysaccharides did not reveal structural differences.However, this phenomenon does not explain the increase in the number of Hib vaccine failures in the Netherlands.

View Article: PubMed Central - HTML - PubMed

Affiliation: Laboratory for Infectious Diseases and Perinatal screening, National Institute for Public Health and the Environment, Antonie van Leeuwenhoeklaan 9, 3721 MA Bilthoven, The Netherlands. LM.Schouls@rivm.nl

ABSTRACT

Background: Despite nearly complete vaccine coverage, a small number of fully vaccinated children in the Netherlands have experienced invasive disease caused by Haemophilus influenzae serotype b (Hib). This increase started in 2002, nine years after the introduction of nationwide vaccination in the Netherlands. The capsular polysaccharide of Hib is used as a conjugate vaccine to protect against Hib disease. To evaluate the possible rise of escape variants, explaining the increased number of vaccine failures we analyzed the composition of the capsular genes and the expressed polysaccharide of Dutch Hib strains collected before and after the introduction of Hib vaccination.

Results: The DNA sequences of the complete capsular gene clusters of 9 Dutch Hib strains were assessed and two variants, designated type I and type II were found. The two variants displayed considerable sequence divergence in the hcsA and hcsB genes, involved in transport of capsular polysaccharide to the cell surface. Application of hcsA type specific PCRs on 670 Hib strains collected from Dutch patients with invasive Hib disease showed that 5% of the strains collected before 1996 were type II. No endogenous type II Hib strains were isolated after 1995 and all type II strains were isolated from 0-4 year old, non-vaccinated children only. Analysis of a worldwide collection of Hib strains from the pre-vaccination era revealed considerable geographic differences in the distribution of the type I and type II strains with up to 73% of type II strains in the USA. NMR analysis of type I and type II capsule polysaccharides did not reveal structural differences. However, type I strains were shown to produce twice as much surface bound capsular polysaccharide.

Conclusion: Type II strains were only isolated during the pre-vaccination era from young, non-vaccinated individuals and displayed a lower expression of capsular polysaccharide than type I strains. The higher polysaccharide expression may have provided a selective advantage for type I strains resulting in the rapid elimination of type II from the Dutch Hib population after introduction of nationwide Hib vaccination. However, this phenomenon does not explain the increase in the number of Hib vaccine failures in the Netherlands.

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Minimum spanning trees of 667 strains on which MLVA was performed and of 247 Hib strains analyzed by MLST. Each circle represents a different type. The size of the circle indicates the number of strains with this particular type. The orange area in the circles indicates the portion of type II strains. The numbers in the circles denote the MLST sequence type or MLVA type. Only the numbers of the predominant types and the types carrying capsular genotype II are displayed.
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Figure 2: Minimum spanning trees of 667 strains on which MLVA was performed and of 247 Hib strains analyzed by MLST. Each circle represents a different type. The size of the circle indicates the number of strains with this particular type. The orange area in the circles indicates the portion of type II strains. The numbers in the circles denote the MLST sequence type or MLVA type. Only the numbers of the predominant types and the types carrying capsular genotype II are displayed.

Mentions: Assessing 667 of the Dutch Hib strains by MLVA [9], revealed that the type II strains were not randomly distributed among the 47 MLVA types. Of the 22 type II strains, 13 (59%) had MLVA type 40 which is the predominant type among Dutch strains, one strain belonged to MLVA type 19 and the remaining 8 strains formed a separate branch of MLVA types not found among type I strains (Fig. 2). Of the collection of Dutch strains available, 247 Hib strains were analyzed by MLST [10,9] and none of the 19 type II strains among this collection were of sequence type 6 (ST-6), which was by far the predominant sequence type (75%) among the 247 Hib strains (Fig 2). This suggested an inverse relationship between capsule locus type II and ST-6. To assess such a possible inverse relationship we amplified and subsequently sequenced the MLST fucK locus for all 89 type II Hib strains from our collection, including strains isolated in other countries. All strains with a fucK allele other than allele number 5 were classified as not having ST-6. Only a few type II Hib strains yielded fucK allele 5 and for these strains the MLST pgi locus was amplified and subsequently sequenced. None of these strains carried pgi allele 7. As pgi allele 7 and fucK allele 5 are part of the ST-6 allelic profile this showed that none of the type II isolates had ST-6, demonstrating an absolute inverse relationship between capsule locus type II and ST-6 genotype for the 89 type II Hib strains included in this study.


Two variants among Haemophilus influenzae serotype b strains with distinct bcs4, hcsA and hcsB genes display differences in expression of the polysaccharide capsule.

Schouls L, van der Heide H, Witteveen S, Zomer B, van der Ende A, Burger M, Schot C - BMC Microbiol. (2008)

Minimum spanning trees of 667 strains on which MLVA was performed and of 247 Hib strains analyzed by MLST. Each circle represents a different type. The size of the circle indicates the number of strains with this particular type. The orange area in the circles indicates the portion of type II strains. The numbers in the circles denote the MLST sequence type or MLVA type. Only the numbers of the predominant types and the types carrying capsular genotype II are displayed.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2267795&req=5

Figure 2: Minimum spanning trees of 667 strains on which MLVA was performed and of 247 Hib strains analyzed by MLST. Each circle represents a different type. The size of the circle indicates the number of strains with this particular type. The orange area in the circles indicates the portion of type II strains. The numbers in the circles denote the MLST sequence type or MLVA type. Only the numbers of the predominant types and the types carrying capsular genotype II are displayed.
Mentions: Assessing 667 of the Dutch Hib strains by MLVA [9], revealed that the type II strains were not randomly distributed among the 47 MLVA types. Of the 22 type II strains, 13 (59%) had MLVA type 40 which is the predominant type among Dutch strains, one strain belonged to MLVA type 19 and the remaining 8 strains formed a separate branch of MLVA types not found among type I strains (Fig. 2). Of the collection of Dutch strains available, 247 Hib strains were analyzed by MLST [10,9] and none of the 19 type II strains among this collection were of sequence type 6 (ST-6), which was by far the predominant sequence type (75%) among the 247 Hib strains (Fig 2). This suggested an inverse relationship between capsule locus type II and ST-6. To assess such a possible inverse relationship we amplified and subsequently sequenced the MLST fucK locus for all 89 type II Hib strains from our collection, including strains isolated in other countries. All strains with a fucK allele other than allele number 5 were classified as not having ST-6. Only a few type II Hib strains yielded fucK allele 5 and for these strains the MLST pgi locus was amplified and subsequently sequenced. None of these strains carried pgi allele 7. As pgi allele 7 and fucK allele 5 are part of the ST-6 allelic profile this showed that none of the type II isolates had ST-6, demonstrating an absolute inverse relationship between capsule locus type II and ST-6 genotype for the 89 type II Hib strains included in this study.

Bottom Line: The DNA sequences of the complete capsular gene clusters of 9 Dutch Hib strains were assessed and two variants, designated type I and type II were found.NMR analysis of type I and type II capsule polysaccharides did not reveal structural differences.However, this phenomenon does not explain the increase in the number of Hib vaccine failures in the Netherlands.

View Article: PubMed Central - HTML - PubMed

Affiliation: Laboratory for Infectious Diseases and Perinatal screening, National Institute for Public Health and the Environment, Antonie van Leeuwenhoeklaan 9, 3721 MA Bilthoven, The Netherlands. LM.Schouls@rivm.nl

ABSTRACT

Background: Despite nearly complete vaccine coverage, a small number of fully vaccinated children in the Netherlands have experienced invasive disease caused by Haemophilus influenzae serotype b (Hib). This increase started in 2002, nine years after the introduction of nationwide vaccination in the Netherlands. The capsular polysaccharide of Hib is used as a conjugate vaccine to protect against Hib disease. To evaluate the possible rise of escape variants, explaining the increased number of vaccine failures we analyzed the composition of the capsular genes and the expressed polysaccharide of Dutch Hib strains collected before and after the introduction of Hib vaccination.

Results: The DNA sequences of the complete capsular gene clusters of 9 Dutch Hib strains were assessed and two variants, designated type I and type II were found. The two variants displayed considerable sequence divergence in the hcsA and hcsB genes, involved in transport of capsular polysaccharide to the cell surface. Application of hcsA type specific PCRs on 670 Hib strains collected from Dutch patients with invasive Hib disease showed that 5% of the strains collected before 1996 were type II. No endogenous type II Hib strains were isolated after 1995 and all type II strains were isolated from 0-4 year old, non-vaccinated children only. Analysis of a worldwide collection of Hib strains from the pre-vaccination era revealed considerable geographic differences in the distribution of the type I and type II strains with up to 73% of type II strains in the USA. NMR analysis of type I and type II capsule polysaccharides did not reveal structural differences. However, type I strains were shown to produce twice as much surface bound capsular polysaccharide.

Conclusion: Type II strains were only isolated during the pre-vaccination era from young, non-vaccinated individuals and displayed a lower expression of capsular polysaccharide than type I strains. The higher polysaccharide expression may have provided a selective advantage for type I strains resulting in the rapid elimination of type II from the Dutch Hib population after introduction of nationwide Hib vaccination. However, this phenomenon does not explain the increase in the number of Hib vaccine failures in the Netherlands.

Show MeSH
Related in: MedlinePlus