The decline of Haemophilus influenzae type b disease in Australia

Between July 1993 and June 1996, there were 412 cases of invasive Haemophilus influenzae type b (Hib) disease reported to the Hib Case Surveillance Scheme, 71% in children under the age of five years. Meningitis was the most frequent illness reported, followed by epiglottitis, septicaemia and pneumonia. There were 18 deaths. Thirty-four cases were classified as vaccine failures. The number of vaccine failures increased over time and the total number of cases of Hib disease fell, consistent with an increase in Hib vaccine coverage. Based on an estimated vaccine coverage of 50% in April 1995, the vaccine efficacy for all vaccines in the period was estimated to be 89%. Invasive Hib is a serious illness of childhood which is being significantly reduced by the use of Hib vaccines, and has the potential to be eliminated from this country. Vaccination providers should aim to immunise all children against Hib disease on time and according to the National Health and Medical Research Council Standard Vaccination Schedule.     

Mathematical models of Haemophilus influenzae type b

Electron microscopic anterograde autoradiography has been used to analyze the morphology and postsynaptic relationships of area 17 cortical terminals in the lateral division of the lateral posterior nucleus (LPl) of the cat and medial division of the inferior pulvinar nucleus (IPm) of the owl monkey. Such terminals are thought to arise exclusively from layer 5 in the cat and primate (Lund et al. [1975] J. Comp. Neurol. 164:287-304; Abramson and Chalupa [1985] Neuroscience 15:81-95). All labeled terminals in both nuclei exhibited the morphology of ascending "lemniscal" afferents. That is, they contained round vesicles, were large, made asymmetrical synaptic and filamentous nonsynaptic contacts, and were classified as RLs. These cortical RLs also exhibited the postsynaptic relationships of lemniscal afferents. Thus, they were presynaptic to large dendrites within glial encapsulated glomeruli, where a majority was involved in complex synaptic arrangements called triads. They also were found adjacent to terminal profiles with pleomorphic vesicles but never adjacent to small terminals containing round vesicles. Our results suggest that the layer 5 projection from area 17 provides a functional "drive" for some LPl and IPm neurons. Information carried over this "re-entrant" pathway (Guillery [1995] J. Anat. 187:583-592) could be modified within the LPl and IPm by both cortical and subcortical pathways and subsequently conveyed to higher visual cortical areas, where it could be integrated with messages carried through the well-documented corticocortical pathways (Casagrande and Kaas [1994] Cerebral cortex New York: Plenum Press).     

Reactivity of antibodies against conserved regions of pilins of Haemophilus influenzae type b

To identify epitopes on pilins of Haemophilus influenzae type b (Hib) that may also be immunologically available on assembled pili, antisera were developed against eight synthetic peptides that represent conserved and hydrophilic regions of Hib pilin. Seven of the eight peptides were immunogenic. Binding of the anti-peptide antibodies to purified pili of Hib strain Eagan was weak. However, when the purified pili were denatured by heating, binding of the anti-peptide antibodies improved considerably, suggesting that the epitopes defined by the peptides were more available for anti-peptide antibody binding on the denatured pilins than on purified pili. On Western blot analysis, strain variation was seen in the binding of some of the anti-peptide antibodies, notably those directed against peptides in the N-terminal half of the pilin. Thus, when pilins are assembled into pili, the epitopes defined by the seven immunogenic peptides appear to be altered so that binding of the anti-peptide antibodies is greatly reduced.     

The epidemiology of Haemophilus influenzae invasive disease in Scotland prior to immunisation

Immunisation against Haemophilus influenzae b (Hib) was added to the UK childhood vaccination schedule on 1 October 1992. Based on reports of laboratory isolations from blood and/or CSF, the epidemiology of Haemophilus influenzae invasive disease in Scotland during the last full year before immunisation (1991) is reviewed. In children aged under five years the estimated incidence of infection (25.5 per 100,000) is higher than that previously reported from Scotland, but lower than estimates from Glasgow and other UK studies. However, the age-sex and seasonal distribution is consistent with previous surveys. As in England and Wales, there appears to be regional variation in incidence within Scotland, although this may simply reflect differences in the completeness of laboratory reporting. In addition to 113 laboratory reports of H. influenzae invasive infection, a retrospective search of hospital discharge data and death registrations identified a further 51 and two cases respectively, some of whom may be genuine. In spite of reservations about hospital discharge data, this raises the possibility that there may be an element of under-reporting by laboratories. With the advent of record linkage of hospital discharge data, it would be prudent to monitor the impact of the Hib vaccine programme using this data source in addition to laboratory reports and death registrations.     

Conjugate vaccines and the carriage of Haemophilus influenzae type b

Pharyngeal carriage of Haemophilus influenzae type b (Hib) is important in the transmission of Hib organisms, the pathogenesis of Hib disease, and the development of immunity to the bacterium. The remarkable success of current vaccination programs against Hib has been due in part to the effect of conjugate Hib vaccines in decreasing carriage of Hib. This review explores evidence for this effect, and discusses the possible mechanisms of the mucosal influence of Hib conjugate vaccines.     

Molecular size characterization of Haemophilus influenzae type b polysaccharide-protein conjugate vaccines

Current vaccines against Haemophilus influenzae type b (Hib) consist of capsular polysaccharide, polyribosylribitol phosphate (PRP), chemically conjugated to a carrier protein. The stability of the conjugate is essential for vaccine efficacy, as the target population for this vaccine includes infants, who do not mount an immune response to free polysaccharide vaccines. A method has been developed for determining structural stability and batch-to-batch consistency of Hib vaccines by the application of fast protein liquid chromatography (FPLC). This FPLC method is fast, reproducible, and can be used to evaluate single human doses of Hib vaccines. We have shown that the FPLC elution profiles provide a suitable indicator of vaccine stability under normal and degradative conditions. The method may also be applicable to other conjugate vaccines such as meningococcal and pneumococcal protein-polysaccharide conjugates.     

Invasive infection with Haemophilus influenzae type b in spite of complete vaccination

Five patients, 4 boys and 1 girl aged 13-41 months, developed invasive Haemophilus influenzae type b (Hib) disease (2 epiglottitis, 3 meningitis) despite full (or at least 3 times) vaccination. At admission as well during convalescence, 3 out of 5 had IgG anti Hib antibody levels < or = 5 U/ml. Serum immunoglobulin levels, including IgG subclasses, as well as complement were normal in all cases. In 2 of the 3, booster vaccinations with Hib conjugate vaccine elicited adequate antibody titres. Since the incorporation of the conjugated Hib polysaccharide tetanus toxoid vaccine (HibTT) in the National Vaccination Programme in the Netherlands, the number of invasive infections caused by Hib has dropped significantly. Causes of Hib conjugate vaccine failures are mostly unknown. In about one-third of the cases serum immunoglobulin levels are deficient, most often IgG2 or IgM. Susceptibility to Hib infection is in part also genetically determined. In the follow-up of Hib vaccine failures, anti Hib antibody titres should be determined. Booster vaccinations may be necessary.     

A competitive enzyme-linked immunosorbent assay for measuring the levels of serum antibody to Haemophilus influenzae type b

1. The protein family of the neurotrophins, consisting of nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF) and Neurotrophin-3, -4/5, and -6 (NT-3; NT-4/5; NT-6) is well known to enhance the survival and to stabilize the phenotype of different populations of neurons in the central and the peripheral nervous system. These effects are mediated via binding to specific tyrosine kinase receptors (Trks) and to the low-affinity p75 neurotrophin receptor. 2. The neurotrophins NGF, BDNF, and NT-3 and the BDNF and NT-3 selective receptors (TrkB, TrkC) are expressed at high levels in neurons of the basal forebrain, the hippocampus, and the neocortex of the mammalian brain. The expression and secretion of NGF and BDNF in these brain areas is regulated by (physiological levels of) neuronal activity. 3. Exogenous application of the neurotrophins to hippocampal and neocortical neurons can enhance excitatory glutamatergic synaptic transmission via activation of Trk receptors. In addition, long-term potentiation (a potential cellular correlate for learning and memory formation in mammals) in the rodent hippocampus depends on endogenous supply of neurons with BDNF. 4. Judged by the analysis of electrophysiological data, the BDNF- and NT-3-induced enhancement of glutamatergic synapses is mediated by increasing the efficacy of glutamate release from the presynaptic neuron. However, neurotrophin-dependent postsynaptic enhancement of NMDA (but not AMPA) receptor responsiveness has also been shown. 5. On the molecular level, neither the pre- nor the postsynaptic modulation of glutamatergic synapses by neurotrophins is well understood. However, neurotrophins were shown to acutely affect intraneuronal Ca2+ levels and to influence molecular components of the transmitter release machinery, which could underly the presynaptic modifications, whereas BDNF-induced phosphorylation of NMDA-type glutamate receptors could account for the postsynaptic effects. 6. Taken together, these results suggest that the activity-dependent release of neurotrophins at frequently used synapses could modulate the synaptic efficacy at these junctions. Thus, neurotrophins might operate as locally released feedback modulators of synaptic transmission, and this could be a cellular correlate for certain aspects of information processing in the mammalian brain.     

A 20-kilodalton N-terminal fragment of the D15 protein contains a protective epitope(s) against Haemophilus influenzae type a and type b

A conserved 80-kDa minor outer membrane protein, D15, of Haemophilus influenzae has been shown to be a protective antigen in laboratory animals against H. influenzae type a (Hia) or type b (Hib) infection. To localize the protective B-cell epitope(s) within the D15 protein and to further explore the possibility of using synthetic peptides as vaccine antigens, a 20-kDa N-terminal fragment of D15 protein (truncated D15 [tD15]) was expressed as a fusion protein with glutathione S-transferase in Escherichia coli. The tD15 moiety was cleaved from glutathione S-transferase by using thrombin and purified to homogeneity. The purified soluble tD15 appeared to contain immunodominant protective epitope(s) against Hia and Hib, since rabbit antisera directed against tD15 were capable of protecting infant rats from Hia or Hib bacteremia. The ease of purification of soluble tD15, therefore, makes it a better candidate antigen than the full-length recombinant D15 which is produced as inclusion bodies in E. coli. Furthermore, both the purified tD15 fragment and a mixture of tD15-derived peptides spanning amino acid residues 93 to 209 of the mature D15 protein were capable of inhibiting the protection against Hib conferred on infant rats by rabbit anti-tD15 antiserum, indicating that the protective epitopes of D15 may not be conformational. However, the administration of pooled rabbit immune sera raised against the same panel of peptides failed to protect infant rats from Hib infection.     

Interlaboratory study evaluating quantitation of antibodies to Haemophilus influenzae type b polysaccharide by enzyme-linked immunosorbent assay

An interlaboratory study was conducted to determine whether an enzyme-linked immunosorbent assay (ELISA) with an antigen preparation composed of various-sized fragments of Haemophilus influenzae type b polysaccharide conjugated to human serum albumin could be standardized across laboratories and whether the ELISA-derived results from different laboratories are equivalent to those obtained by the standard radioactive antigen binding assay (RABA) for quantitation of anti-H, influenzae type b polysaccharide antibodies. Twenty coded human serum samples were quantitated by ELISA in 11 laboratories and by RABA in 5 laboratories. The mean RABA-derived values served as the basis for all comparisons. While the overall correspondence of antibody values between the two methods was good, significant differences were found among some of the 11 ELISA data sets and among the mean RABA values. Seven laboratories generated higher ELISA antibody values for low-titered sera. Four laboratories generated antibody concentrations that were not statistically different between the two assay methods. The results therefore indicate that the ELISA can tolerate substantial variations in protocol, such as the use of different plates and different antibody reagents, without affecting the quantitation of serum antibodies. However, attention should be focused on low-titered sera, as some assay conditions may yield spurious results. This ELISA is a serologic assay which can serve as an alternative to the RABA for quantitation of antibodies to H. influenzae type h polysaccharide.     

Perspective: a five-country analysis of the impact of four different Haemophilus influenzae type b conjugates and vaccination strategies in Scandinavia

Prior to vaccinations against invasive Haemophilus influenzae type b (Hib) diseases in Scandinavia, first initiated in Finland in 1986, the incidence of cases in those five countries was 49/100,000/year in 0- to 4-year-olds and 3.5/100,000 overall. During the following decade, Hib conjugates administered to young children had approximately 95% effectiveness, regardless of which conjugate was used, whether two or three primary doses were administered, and at what age in early infancy the first vaccination was given. The herd immunity effect has extended protection to older age groups. A similar effectiveness of different conjugates in five countries despite considerable diversity in approach suggests that the same impact would occur in other regions with comparable epidemiology. The Scandinavian experience supports the view that three primary vaccine doses are not imperative, thus suggesting that reducing doses of costly Hib vaccines would be one way to facilitate their usage in regions with limited resources.     

Low genetic diversity of Haemophilus influenzae type b compared to nonencapsulated H. influenzae in a population in which H. influenzae is highly endemic

Immunization with Haemophilus influenzae type b (Hib) conjugate polysaccharide vaccines has dramatically reduced Hib disease worldwide. As in other populations, nasopharyngeal carriage of Hib declined markedly in Aboriginal infants following vaccination, although carriage has not been entirely eliminated. In this study, we describe the genetic characteristics and the carriage dynamics of longitudinal isolates of Hib, characterized by using several typing methods. In addition, carriage rates of nonencapsulated H. influenzae (NCHi) are high, and concurrent colonization with Hib and NCHi is common; we also observed NCHi isolates which were genetically similar to Hib. There is a continuing need to promote Hib immunization and monitor H. influenzae carriage in populations in which the organism is highly endemic, not least because of the possibility of genetic exchange between Hib and NCHi strains in such populations.     

Haemophilus influenzae type b osteoarthritis. A report of 7 cases and a review of the literature

OBJECTIVE: The epidemiological and clinical characteristics, treatment and evolution of osteoarthritis by Haemophilus influenzae type b (HIB) are reviewed since there has been little published on this subject in our country. PATIENTS AND METHODS: The clinical histories of the 7 children with osteoarthritis due to Hib infection are reviewed. The diagnostic criteria included classical signs and symptoms of septic arthritis, radiological alterations compatible with joint infection and isolation of microorganisms in joint effusion and/or in the blood. RESULTS: During a 24-year period (1973-1996), 248 cases of invasive infection by Hib were documented. Seven cases (2.82%) had osteoarticular infections. The ages were between 5 and 7 years and there were more males than females (71.4% vs. 28.6%). Four children/58%) had previous upper respiratory infections (URI). The microorganism was isolated in the joint effusion in 5 children and in the blood sample of the other 2. C-reactive protein was high and radiology showed alterations in 100% of the cases. Surgical treatment with articular drainage was necessary in 5 children/71.4%). In 6 cases (85.7%) initial medical treatment was i.v. beta-lactam antibiotics for 2-3 weeks followed by oral antibiotic treatment for a minimum of 6 weeks. Three children (42.8%) had sequelae. CONCLUSIONS: Early diagnosis of bacterial osteoarthritis by Hib is difficult. Final therapeutical success depends on an early clinical diagnosis and aggressive multidisciplinary treatment. Drainage of the hip joint is mandatory for successful outcome. Currently, arthritis by Hib can be avoided and its sequelae prevented by vaccination.     

A routine high-performance size-exclusion chromatography to determine molecular size distribution of Haemophilus influenzae type b conjugate vaccines

High-performance size exclusion chromatography has been used to determine the molecular size distribution of Haemophilus influenzae type b (Hib) conjugate vaccines. Both high molecular weight preparations of native Hib capsular polysaccharide coupled to tetanus toxoid and low molecular weight vaccines with Hib oligosaccharides linked to the CRM197 nontoxic mutant diphtheria protein were analysed. Columns with different fractionation ranges were used for the two kinds of vaccines. This method showed to be rapid, accurate and reproducible for different lots of Hib vaccine of different composition produced by various manufacturers. It could replace more time-consuming chromatographic methods enabling control authorities to employ a single methodological approach for different Hib vaccines.