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PubMed1990

Urogenital, maternal and neonatal isolates of Haemophilus influenzae: identification of unusually virulent serologically non-typable clone families and evidence for a new Haemophilus species.

A collection of 117 strains of Haemophilus influenzae, including 112 non-typable isolates recovered predominantly in the USA and France from genital, obstetric and neonatal sources, was characterized by the electrophoretic mobilities of 10 metabolic enzymes. Eighty-six distinctive multilocus chromosomal genotypes (electrophoretic types, ETs) were distinguished on the basis of allele profiles at the enzyme loci. Isolates of five allied biotype IV ETs were highly divergent from all other strains and hybridization of chromosomal DNA revealed that they undoubtedly represent a previously unrecognized species of Haemophilus. Isolates representing these ETs were recovered predominantly from obstetric infections and serious neonatal diseases and apparently possess specific tropism for the genital tract. Strains of these five ETs were present in samples from both the USA and France, but only in the USA did they cause bacteraemia and meningitis, an occurrence which probably reflects differences in patient management between the two countries. Although strains assigned to H. influenzae (sensu stricto) were strongly polymorphic in multilocus enzyme genotype, 69% of isolates recovered from patients with meningitis and/or septicaemia were assigned to only two clone families, a result suggesting that some serologically nontypable strains of H. influenzae originating from the genital tract are unusually virulent.

Alleles↗
PubMed1993

Ecology of Haemophilus influenzae and Haemophilus parainfluenzae in sputum and saliva and effects of antibiotics on their distribution in patients with lower respiratory tract infections.

Nine patients with lower respiratory tract infections were used to study in detail the effect of ampicillin or erythromycin on the colonization patterns of Haemophilus influenzae and Haemophilus parainfluenzae in sputum and saliva. H. influenzae was isolated from purulent sputum of eight patients before the start of treatment. Ampicillin was more effective than erythromycin at clearing H. influenzae from sputum and in decreasing purulence. By careful characterization of multiple strains, the changes in biotype distribution and antibiotic susceptibility patterns were shown. Five biotypes of H. influenzae were associated with chest infection, with type II predominating. Mixed biotype infections occurred in five patients. Most saliva contained multiple biotypes of H. parainfluenzae. Neither antibiotic selected resistant haemophili in saliva or sputum. After treatment with ampicillin, the mucoid sputum was colonized with ampicillin-susceptible H. parainfluenzae biotypes previously found in saliva. We postulate that as inflammation decreases at the bronchial mucosa, the ampicillin concentration drops, allowing ampicillin-susceptible oral H. parainfluenzae isolates to seed the residual mucoid sputum.

Ampicillin↗
PubMed1989

Typing of urogenital, maternal, and neonatal isolates of Haemophilus influenzae and Haemophilus parainfluenzae in correlation with clinical source of isolation and evidence for a genital specificity of H. influenzae biotype IV.

Over a period of 6 years, 114 strains of Haemophilus influenzae and Haemophilus parainfluenzae were isolated from genital, mother-infant, or neonatal infections. Their serotypes, biotypes, antibiotic resistance phenotypes, and outer membrane protein (OMP) electrophoretic patterns were characterized and correlated with the various clinical outcomes. Genital H. influenzae and H. parainfluenzae appeared to behave mostly as opportunistic pathogens; for instance, 62% of the cases of endometritis or pelvic inflammatory disease were related to the presence of an intrauterine device. However, as seen clearly in one case, the strains may be sexually transmitted. The analysis of OMP patterns proved to be a very convenient method to seek evidence for the sexual origin of the infection. H. influenzae was more often involved in complicated genital infections than was H. parainfluenzae. Nontypeable and biotype II H. influenzae strains were the more frequent isolates, except in pelvic inflammatory diseases, in which biotype I prevailed, and in mother-infant infections, in which one-fourth of the cases were due to biotype IV. Characterization of H. influenzae isolates did not support a general concept of specific genital strains. However, strains of biotype IV clearly stood out with two characteristics: (i) a peritrichous fimbriation and (ii) a very peculiar homogeneous OMP pattern comprising an OMP of molecular weight approximately 18,000 unique to this biotype. These characteristics were also found in H. influenzae biotype IV strains isolated from genital infections in the United States and used as controls. H. influenzae biotype IV strains may thus correspond to a group somewhat adapted to the genital tract.

Female↗
PubMed1989

Haemophilus influenzae type b polysaccharide vaccine: an efficacy study. Haemophilus Vaccine Efficacy Study Group.

The Haemophilus influenzae type b polysaccharide vaccine was licensed for use in the United States in April 1985. Postlicensure case-control efficacy studies have yielded markedly different estimates of efficacy, leading to contradictory recommendations to practicing physicians. To obtain additional information about the efficacy of the vaccine, we studied cases of invasive Haemophilus influenzae type b disease ascertained through active surveillance in areas with a total population of 34 million. We enrolled children 24 to 59 months of age who did not attend day-care centers. (Data from our day-care study have been published elsewhere.) For each case child, as many as three 24- to 59-month-old control children were chosen from a roster of acquaintances supplied by the child's parent. Conditional logistic regression was used, and vaccine efficacy was estimated to be 62% (95% confidence interval = 0%, 85%), which did not change significantly after adjusting for age and parental smoking, variables that were significantly different for case and control children. Results of this study support our previous finding of a positive protective efficacy, albeit lower than the efficacy of 90% found in children 18 to 71 months of age in the Finnish prelicensure trial.

Bacterial Capsules↗
PubMed2001

Preservation of Haemophilus influenzae and Haemophilus parainfluenzae at -70 degrees C.

We have studied the viability of Haemophilus spp. preserved for 5 to 12 months at -70 degrees C. The following media were used: Laboratoire de Santé Publique du Québec (LSPQ) preservation medium, trypticase soy broth with 10 degrees C (vol/vol) glycerol and 40 degrees C (vol/vol) horse serum (TSBG), and Levinthal's broth (LB) medium. Three clinical isolates of both H. influenzae and H. parainfluenzae were used. After 5 months no differences in viability were observed between strains preserved in TSBG and strains preserved in LB, but a significant loss of viability was observed in strains preserved in LSPQ medium. No significant changes in antimicrobial susceptibility were observed after 5-month storage in any medium. After 12 months, TSBG appeared to be the most suitable cryopreservation medium for the six strains tested. We conclude that TSBG represents a good medium for the maintenance of Haemophilus spp. at -70 degrees C for up to 1 year.

Animals↗
PubMed1984

Antibiotic treatment of Haemophilus influenzae and Haemophilus parainfluenzae infections in patients with cystic fibrosis.

During the period 1977-81, 188 Cystic fibrosis patients have been treated with 1 254 anti-haemophilus treatments in the Danish Cystic Fibrosis Center. On an average, each patient received 1.7 courses of chemotherapy per year. Haemophilus infections were more prevalent in the youngest age group of patients. The average number of H. influenzae precipitins increased with age, and this pattern was consistent throughout the whole study. The standard treatment was orally administered pivampicillin or amoxicillin given for 14 days. 1 case of allergy or ampicillin-resistant strains, erythromycin was given, in most cases alone, in some cases in combination with rifampicin. The overall results showed, that H. influenzae was eradicated from sputum by a single course of chemotherapy in 73% of the courses and H. parainfluenzae in 100% of the courses. Six per cent of the patients suffered from chronic H. influenzae infection in spite of repeated treatments. No decrease of treatment efficiency was found even after many repeated chemotherapy courses. The prevalence of patients harbouring ampicillin-resistant H. influenzae increased in the beginning of the period (6%-20%), but a slight fall was subsequently observed.

Adolescent↗
PubMed1982

Differentiation of Haemophilus aegyptius and Haemophilus influenzae.

This study aimed at clarifying the relationship of Haemophilus aegyptius and Haemophilus influenzae isolated from acute conjunctivitis in Egypt. Twenty-nine freshly isolated strains selected from a large clinical material were examined for morphological and growth characteristics, biochemical properties and susceptibility to selected antibiotics. H. aegyptius strains were clearly differentiated from strains of H. influenzae by their inability to grow on tryptic soy agar containing X + V factors, by their susceptibility to trooleandomycin, by a distinct bacillary morphology, and, in part, by not fermenting xylose. The results confirm that H. aegyptius is distinct from H. influenzae and provides reproducible means of differentiating the two species.

Anti-Bacterial Agents↗
PubMed1986

Comparative in vitro activity of imipenem against Haemophilus influenzae and Haemophilus parainfluenzae.

A microdilution broth method was used to test 77 clinical isolates of Haemophilus influenzae and Haemophilus parainfluenzae, including beta-lactamase-positive and -negative strains, for susceptibility to ampicillin, chloramphenicol, and imipenem. Except for ampicillin against beta-lactamase-producing strains (MIC for 90% of strains [MIC90], greater than or equal to 128 micrograms/ml), all three antimicrobial agents had comparable in vitro activity (MIC90, less than or equal to 1 microgram/ml) against these bacterial strains.

Ampicillin↗
PubMed2001

PCR-based detection, restriction endonuclease analysis, and transcription of tonB in Haemophilus influenzae and Haemophilus parainfluenzae isolates obtained from children undergoing tonsillectomy and adenoidectomy.

We developed and evaluated a PCR-based-restriction endonuclease analysis method to detect and analyze the tonB gene of Haemophilus influenzae and Haemophilus parainfluenzae from pediatric patients undergoing tonsillectomy and adenoidectomy. Multiple sites from the same patient, including the surface of adenoids and tonsils, as well as the core of tonsils, were cultured on chocolate agar and identified using standard procedures and the API NH Kit. A total of 55 H. influenzae isolates were recovered from different sites of 20 patients, and 32 H. parainfluenzae isolates were recovered from various sites of 12 patients. DNA was extracted from American Type Culture Collection strains and test isolates by the PureGene kit. Two primers, G1 (21-mer) and G2 (23-mer), were designed by us to amplify by PCR the tonB gene that consists of an 813-bp fragment. A nested PCR using primers T1 (23-mer) and T2 (24-mer) that flank an internal sequence to the gene of the order of 257 bp and restriction endonuclease digestion using XhoI and BglII were done to detect whether heterogeneity within the gene exists between the two species. Reverse transcription-PCR (RT-PCR) was finally done to detect transcription of the gene in both species. Our data have shown that the tonB gene was detected in both species. It is known to encode a virulent protein, TonB, in H. influenzae; however, demonstration of its presence in H. parainfluenzae is novel. Nested-PCR and restriction endonuclease analysis have shown that the tonB gene is apparently structurally the same in both species, with possible differences that may exist in certain H. parainfluenzae isolates. RT-PCR done on selected numbers of H. influenzae and H. parainfluenzae have shown that the tonB gene was transcribed in both species. This shows that the TonB protein, if expressed, may play a different role in the virulence in H. parainfluenzae since it is not needed for heme or heme complexes uptake as with H. influenzae.

Adenoidectomy↗
PubMed1992

The hbpA gene of Haemophilus influenzae type b encodes a heme-binding lipoprotein conserved among heme-dependent Haemophilus species.

A membrane-associated lipoprotein of Haemophilus influenzae type b has previously been shown to bind heme in vitro and to promote binding of this compound by Escherichia coli recombinants expressing this protein. The H. influenzae type b heme-binding protein A (HbpA) was found to be highly conserved with respect to both antigenicity and apparent molecular weight among heme-requiring Haemophilus species pathogenic for humans. To further the characterization of the structure and function of HbpA, the complete nucleotide sequence of its gene, hbpA, was determined. Analysis of the nucleotide sequence revealed a single large open reading frame of 1,638 bp encoding a protein of 546 amino acid residues, with a molecular weight of 60,695. The sequence of the amino-terminal end of this protein contained a potential site for lipid acylation and for cleavage by signal peptidase II, consistent with earlier biochemical evidence which indicated that HbpA is a lipoprotein. A search of GenBank for proteins with amino acid sequence similarity to HbpA revealed that the periplasmic dipeptide transport protein of E. coli, DppA, has 53% sequence identity to HbpA.

Amino Acid Sequence↗
PubMed1995

Monoclonal antibodies against Haemophilus lipopolysaccharides: clone DP8 specific for Haemophilus ducreyi and clone DH24 binding to lacto-N-neotetraose.

Mouse monoclonal antibodies (MAbs) DP8 [immunoglobulin G1(kappa)] and DH24 [immunoglobulin M(kappa)], which are specific for Haemophilus ducreyi lipopolysaccharide (LPS), were generated by fusing mouse myeloma NS0 cells with spleen cells of BALB/c mice immunized with a total membrane preparation of H. ducreyi. MAb DP8 reacted in whole-cell enzyme immunoassay (EIA) and colony dot immunoblotting with all 50 strains of H. ducreyi but not with any other bacteria tested, which suggests an exposed and species-specific epitope on the H. ducreyi cell surface. This conclusion was supported by the finding that DP8 bound to all six H. ducreyi LPSs tested but not to any of the Haemophilus influenzae or enterobacterial LPSs or synthetic glycoconjugates. The MAb DH24 bound to 43 of 50 strains of H. ducreyi and to few strains of H. influenzae, Neisseria gonorrhoeae, and Neisseria meningitidis, as evaluated by whole-cell EIA and colony dot immunoblotting. The MAb DH24 reacted with five of the six H. ducreyi LPSs tested and with the lacto-N-neotetraose (Gal beta 1-->4GlcNAc beta 1-->3Gal beta 1-->4Glc) series of synthetic glycoconjugates, as determined by EIA. By using polysaccharides obtained after both mild acidic hydrolysis and strong alkali treatment and dephosphorylated samples as inhibitors of the MAbs binding to H. ducreyi LPS antigens, it could be shown that phosphate groups were essential for the binding of DP8 to LPS but that they did not affect antigenic recognition by DH24. None of the MAbs bound to isolated lipid A, but aggregation caused by the fatty acids of lipid A was essential for epitope recognition.

Animals↗
PubMed1985

Comparison of transformation mechanisms of Haemophilus parainfluenzae and Haemophilus influenzae.

Transformation pathways in two closely related bacterial species, Haemophilus parainfluenzae and Haemophilus influenzae, were studied. Both organisms rapidly take up transforming DNA within minutes into specialized membranous structures on the cell surface (transformasomes). DNA within transformasomes is in a protected state, inaccessible to external DNase or internal restriction and modification enzymes. However, the subsequent processing of donor DNA differs in these two organisms. In H. influenzae, linear DNA immediately undergoes degradation from one end at a constant rate, leaving a lower-molecular-weight intermediate in the transformasome. The end undergoing degradation is searching for homologous regions of the chromosome. Once pairing is initiated, the remaining lower-molecular-weight DNA exits from the transformasome, and a single strand undergoes efficient integration. In contrast, in H. parainfluenzae little degradation of donor DNA is observed, with the majority remaining intact within the transformasomes after 1 h. Thus, whereas only 10% of donor DNA molecules leave the protected state after 1 h, portions of each molecule appear to become quantitatively integrated.

Cell Membrane↗
PubMed1975

New satellitism test for isolation and identification of Haemophilus influenzae and Haemophilus parainfluenzae in sputum.

A new satellitism test designed to facilitate the isolation and identification of Haemophilus influenzae and Haemophilus parainfluenzae is described. In the basal medium, nicotinamide adenine dinucleotide is incorporated at a concentration of 0.2 mug per ml, an amount adequate for H. influenzae but not for H. parainfluenzae. Two disks are placed on the surface of the medium, one disk being impregnated with 60 mug of hemin and the other with 15 mug of nicotinamide adenine dinucleotide. Under these conditions, H. influenzae strains grow around the hemin disk only and the majority of H. parainfluenzae grow around the nicotinamide adenine dinucleotide disk. This procedure gives results which are more clear cut than other established methods, especially in sputum culture.

Bacteriological Techniques↗
PubMed2004

Oropharyngeal colonization by Haemophilus influenzae in healthy children from Taubaté (São Paulo), prior to the Haemophilus influenzae type B vaccination program in Brazil.

UNLABELLED: Haemophilus influenzae is one of the most important bacterial agents of otitis and sinusitis. H. influenzae type b (Hib) is one of the main causes of meningitis, pneumonia, and septicemia in nonvaccinated children under 6 years of age. The aims of this study were to determine the prevalence of H. influenzae and Hib oropharyngeal colonization prior to the onset of the Hib vaccination program in Brazil in previously healthy children and to assess the susceptibility profile of this microorganism to a selected group of antimicrobials that are used to treat acute respiratory infections. METHOD: Cultures of Haemophilus influenzae were made from oropharynx swabs from 987 children under 6 years of age who were enrolled in 29 day-care centers in Taubaté (a city of São Paulo state, Brazil) between July and December 1998. RESULTS: The prevalence of H. influenzae carriers was 17.4%, and only 5.5% of the strains were beta-lactamase producers. The prevalence of Hib carriers was high, 7.3% on average (range, 0.0 - 33.3%). CONCLUSIONS: The low prevalence of colonization by penicillin-resistant strains indicates that it is not necessary to substitute ampicilin or amoxicilin to effectively treat otitis and sinusitis caused by H. influenzae in Taubaté.

Anti-Bacterial Agents↗
PubMed1997

The tympanic membrane and middle ear mucosa during non-typeable Haemophilus influenzae and Haemophilus influenzae type b acute otitis media: a study in the rat.

Non-typeable Haemophilus influenzae (NTHi) and encapsulated Haemophilus influenzae type b (Hib) were inoculated into the middle ears of Sprague-Dawley rats. Tympanic membrane (TM) status was assessed otomicroscopically and specimens from various middle ear areas were prepared for light microscopy at various times during the acute phase and up to 6 months after inoculation. Irrespective of bacteria strain, acute otitis media (AOM) was present in all ears 4 days after inoculation. The Hib-infected ears showed initially a severe course of AOM, but all were otomicroscopically resolved by day 12, at which time a few NTHi-inoculated ears still exhibited middle ear effusion. The TMs infected with Hib had normalized without scar formation, whereas NTHi induced a persistent thickening of the TMs in half of all cases. The middle ear mucosa of NTHi-infected ears initially showed vigorous activity among the goblet cells, but the mucosa normalized after the acute phase. Hib, by contrast, induced prominent changes in the middle ear mucosa. Initially, no goblet cell granules or ciliated cells could be observed in the mucosa. Later on, the epithelium contained large, active goblet cells. Glands appeared beneath the mucosa which persisted as streaks of epithelial cells throughout the study period. The findings show that NTHi and Hib both induce AOM but with differing clinical courses, and affect different targets in the middle ear.

Acute Disease↗
PubMed1999

Oklahoma notes decline in Haemophilus influenzae: invasive Haemophilus influenzae disease among children aged < 5 years--Oklahoma, 1990-1997.

Haemophilus influenzae (Hi) causes many clinical illnesses such as meningitis, bacteremia, epiglottitis, pneumonia, otitis media, sinusitis and tracheobronchitis. Before the introduction of the Haemophilus influenzae type b (Hib) conjugate vaccine in 1988, Hib caused more than 95 percent of invasive Hi disease in developed countries. Conjugate vaccines were licensed for use in children > or = 15 months of age in 1989 and for use in children > or = 2 months of age in 1990. During 1987-1995, the incidence of invasive Hi disease among children < 5 years of age decreased 96 percent in the United States. This report summarizes the trend in invasive disease caused by Hi among Oklahoma children < 5 years of age. The data represents cases reported to the OSDH as part of the infectious disease surveillance system. Invasive Hi disease has been reportable by law in Oklahoma since 1983.

Child, Preschool↗