ARCHIVED - Invasive Haemophilus influenzae disease in Manitoba in the post-vaccination era suggests a changing epidemiology

 

Canada Communicable Disease Report

1 June 2006

Volume 32
Number 11

Introduction

Surveillance for invasive Haemophilus influenzae (Hi) disease in Canada dates back to 1979(1), before the introduction of H. influenzae serotype b (Hib) vaccine. Prior to the introduction of Hib vaccines, most cases of invasive Hi disease were caused by Hib(2). Since the introduction of the polyribosylribitol phosphate conjugate vaccine in Canada in 1992, the incidence of Hib has decreased dramatically, reaching an all-time low in 2000 of only four cases noted by a network of 12 pediatric centres across Canada monitoring for vaccine preventable diseases(3,4). However, surveillance for invasive Hi disease in Canada captures only cases due to Hib, and there is limited information on the prevalence or incidence of invasive Hi disease due to non-b type Hi. Currently, it is not known whether Hib vaccination alters the epidemiology of invasive Hi disease by inducing capsule replacement. Capsule replacement in Hi disease has been reported from at least two countries (Brazil and Portugal) after extensive use of the Hib vaccine(5,6). An editorial comment recently raised concerns that non-b type Hi isolates are becoming a more common cause of invasive Hi disease(7). This raises the question of what should be the appropriate public health response to invasive non-b type Hi disease.

In order to address some of these concerns, we studied invasive Hi strains (defined by isolation from normally sterile body sites such as blood and cerebrospinal fluid [CSF]) isolated at the Health Sciences Centre (HSC) and Children's Hospital in Winnipeg, Manitoba, from 2000 to 2004 and characterized them according to their serotypes using serotype-specific antisera as well as molecular techniques to detect the presence of serotype-specific capsular polysaccharide synthesis genes.

Material and methods

Bacterial isolates

Isolates from patients with invasive Hi diseases were obtained from the Clinical Microbiology Laboratory at the HSC in Winnipeg. The strains were grown on chocolate agar plates and stored at
-800C in brain-heart infusion broth containing 20% glycerol. The identities of all isolates were re-confirmed by standard biochemical tests(8).

Serotyping by antiserum and PCR analysis of capsular polysaccharide synthesis genes

Serotyping was done by slide agglutination assay using antisera from two commercial sources (Difco, Oakville, Ont.; Denka Seiken, Tokyo). Polymerase chain reaction (PCR) amplification of serotype-specific and capsule transport bexA genes was carried out using the primers described by Falla et al.(9).

Results

The HSC and Children's Hospital constitute the largest tertiary health care centre in the province of Manitoba, serving residents of Manitoba, northwestern Ontario, and Nunavut. According to the HSC 2004-2005 annual report(10), approximately 19,748 adults and 5,452 children were admitted to the HSC and Children's Hospital, respectively.

Fifty-three cases of invasive Hi disease were identified by the Clinical Microbiology Laboratory at the HSC. We were able to retrieve the corresponding Hi isolates from 52 of these 53 cases; one isolate did not grow from the original HSC frozen stock. The serotype distribution of these 52 cases by year is presented in Table 1. The serotype identity of the 52 isolates was confirmed by PCR detection of their serotype-specific capsular polysaccharide synthesis genes. The capsular transport bexA gene was also detectable in all serotypeable strains. Serotype a was the most frequently identified serotype (26 cases), followed by non-serotypeable (NST) strains (20 cases); there were only three serotype b isolates and one isolate each of serotypes c, d, and f. There was no trend observed in the frequency of serotype a or NST isolates from 2000 to 2004. Although we did not have the vaccination history of the patients, it was of interest that the three cases of Hib infection were found in infants aged 5, 6, and 9 months. In Canada, the routine immunization schedule for the Hib vaccine is a primary dose given at 2 months of age, followed by boosters at 4, 6, and 18 months of age. Therefore, none of the three patients with Hib disease would have received all four doses of the vaccine.

Table 1. Serotype distribution of 52 Haemophilus influenzae isolates from patients with invasive disease

Serotype

No. of isolates by year of isolation

Total no. of isolates

2000

2001

2002

2003

2004

a

3

7

6

6

4

26

b

0

2

0

0

1

3

c

0

0

0

0

1

1

d

1

0

0

0

0

1

f

0

0

0

1

0

1

NST

4

7

4

1

4

20

Total

8

16

10

8

10

52


The overall age distribution of the patients with invasive Hi disease is presented in Figure 1. Thirty-two cases occurred in patients of ≤ 2 years, and nine cases occurred in patients aged ≥ 50 years. Serotype a was more likely to be isolated from children (of ≤ 2 years) than NST Hi (chi-square test, p < 0.01; odds ratio 5.0; 95% confidence interval 1.2-22.4). Overall, 41 Hi isolates were from blood culture and nine from CSF; three isolates were recovered from both blood and CSF cultures.

Figure 1. Age distution of 52 cases of invasive Haemophilus influenzae disease

Figure 1. Age distution of 52 cases of invasive Haemophilus influenzae disease

Discussion

Although several studies have reported invasive disease due to Hia, most of them were based on reports of only a few cases(6,11-13). This is one of the few studies to date in which a large number of invasive Hia isolates were examined. Twenty-six (50%) of the 52 cases of invasive Hi infection in this study were caused by Hia. In contrast, only three cases (6%) were caused by Hib. Another 6% of the 52 cases were caused by serotypes other than Hia and Hib. Before the introduction of Hib vaccines, Hib was responsible for the majority of serious infections, such as meningitis, epiglottitis, pneumoniae, and septicemia in early childhood, suggesting that it has a more virulent nature than other serotypes.With the declining incidence of Hib, it appears from this study, as well as from others, that Hia may be the second most clinically virulent type among the six serotypes of Hi. Since capsular structure is related to virulence and serotyping of Hi, it is important to note that the Hia capsule structure is more similar to that of Hib than to those of other Hi serotypes. Both the Hia and Hib capsules contain the five-carbon sugar ribitol. In Hia, the ribitol is linked to glucose to form the polymer of glucose-ribitol phosphate, whereas in Hib the capsule is made up of a polymer of ribose-ribitol phosphate(14,15).

As found in other studies, most cases of Hia infection occurred in children(6,11-13). Twenty-one (81%) of our 26 Hia cases were found in children < 4 years, and 14 of those were < 12 months. In this study, there were also five cases in adults: one was 27 years of age, two were 37 years of age, and two were 50 and 60 years of age.

This study also detected a large number of cases due to NST strains (20 of the 52 cases or 38.5%). The virulence properties of NST Hi have been disputed(16); however, our findings indicate a clear link between NST Hi and invasive disease. Unlike the cases of Hia infection, there were more cases of NST infection among adolescents or adults (63%, or 12 out of 19 cases; age information was not available for 1 NST case) than infants or children (37%, or seven out of 19, cases were aged < 13 months).

The finding of NST Hi causing invasive disease in the adult population may be related to the fact that NST Hi is an important respiratory pathogen in patients with chronic obstructive pulmonary disease (COPD)(17). Hi strains isolated from patients with an acute exacerbation of COPD have been reported to induce more inflammation than colonizers, which may explain their increased pathogenicity in these patients(18). However, it is not possible from our laboratory-based study to know the proportion of cases of NST Hi infection with pulmonary involvement. Therefore, our second phase of characterizing the changing epidemiology of invasive Hi diseases would involve chart reviews to determine host factors that may play a role in invasive NST Hi diseases.

Although our data are based on findings from only one province, the observations have far-reaching implications regarding the changing epidemiology of invasive Hi disease in Canada. Therefore, we are proposing to enhance the national surveillance of invasive Hi disease by suggesting that the current national case definition of invasive Hi disease be modified to include cases with positive isolation of any of the six serotypes of Hi as well as NST Hi from normally sterile body sites. Such an initiative would potentially have an impact upon vaccine development for invasive Hi infection. Furthermore, we advocate that serotypes determined by bacterial agglutination should be confirmed by PCR in order to minimize potential error and to ensure that the serotype information for all cases of invasive Hi infection is accurate(19).

As a follow-up to this study, we are conducting a chart review of the 52 patients with invasive Hi disease in order to describe disease severity and outcomes, and to determine whether host risk factors can be identified in association with invasive Hia or invasive NST Hi disease. We are also planning further analyses of the NST Hi isolates in order to determine whether there are common virulence traits within this group of bacteria.

Acknowledgements

We thank Dr. A. Kabani and his staff at the Clinical Microbiology Laboratory of the University of Manitoba HSC for the provision of and permission to use the H. influenzae strains described in this study. R.S.W. Tsang thanks John Robbins of the National Institute of Health for the discussion on capsular structures of H. influenzae.

The materials presented in this report are taken from a previous publication by the authors in an article that appeared in the Journal of Clinical Microbiology(20).We thank the Journals Department of the American Society for Microbiology for permission to reproduce the work.

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Source : RSW Tsang, MMedSc, PhD, Laboratory for Vaccine Preventable Bacterial Diseases, National Microbiology Laboratory (NML), Public Health Agency of Canada (PHAC); S Mubareka, MD, Department of Medical Microbiology and Infectious Diseases, University of Manitoba (UM); ML Sill, BSc, NML, PHAC; J Wylie, PhD, Cadham Provincial Public Health Laboratory, Manitoba Health, Winnipeg, Man.; S Skinner, MD, Department of Medical Microbiology, UM; and DKS Law, BA, BSc, NML, Public Health Agency of Canada, Winnipeg, Manitoba.

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