Fimbriation, hemagglutination and adherence properties of fresh clinical isolates of Branhamella catarrhalis.

This study investigated the fimbriation on 24 fresh clinical isolates of Branhamella catarrhalis by electron microscopy. All the strains were isolated from patients with respiratory infections. The Branhamella catarrhalis strains were classified into three groups according to the grade of fimbriation. Among these 24 strains the incidence of densely fimbriated, moderately fimbriated and sparsely fimbriated isolates were 12 (50%), 7 (29%) and 5 (21%), respectively. After five-times serial subculture on Brain Heart Infusion agar, the average number of fimbriae per bacteria was decreased from 174 to 114 in the densely fimbriated strain and from 48 to 10 in the moderately fimbriated strain. Moreover, 20% of the population became non-fimbriated in moderately fimbriated strain after the serial subculture. In strains with higher hemagglutination titer the number of fimbriae was significantly (P < 0.04) more than in strains with lower hemagglutination titer.     

A note on hydrolysis of tributyrin by Branhamella and Neisseria

Sixty-three strains of Branhamella and Neisseria were tested by two methods for their ability to hydrolyse glycerol tributyrate. After the conventional plate test, gas liquid chromatographical (GLC) analysis of the agar medium was carried out to detect the hydrolysis product, butyric acid, and other volatile fatty acids. All strains of Branhamella catarrhalis, Neisseria caviae, N. cuniculi and N. ovis but no other Neisseria spp. gave positive results with the conventional test. With GLC, however, most strains of Branhamella and Neisseria were shown to liberate butyric acid. In addition, some strains liberated acetic and isovaleric acids. Greater amounts of butyric acid were produced by clinical strains, in particular B. catarrhalis , compared with reference strains. It was concluded that the conventional plate test for tributyrin hydrolysis differentiates B. catarrhalis, N. caviae, N. cuniculi and N. ovis from other Neisseria.     

Expression of Fimbriae and Host Response in Branhamella catarrhalis Respiratory Infections

Sputum during the acute exacerbation of chronic respiratory diseases were observed under the electron microscope, to determine the in vivo expression of surface structures of Branhamella catarrhalis (B. catarrhalis), the polymorphonuclear neutrophil (PMN) response to B. catarrhalis infections, and the composition of sputum. It was found that during infection fimbriae are expressed in B. catarrhalis. However, there were sparsely to densely fimbriated bacteria in each sputum sample. The length of the fimbriae were from 50 to 76 nm. In the sparsely fimbriated B. catarrhalis, external to the cell wall, a thin, granular, electron-dense layer was observed. Due to the presence of fimbriae, this layer was not seen in densely fimbriated B. catarrhalis. Blebs were also found in B. catarrhalis. PMNs were found to phagocytose both B. catarrhalis and debris. Evidence was found that debris were formed mainly by the destruction of PMNs. Bacteria as well as debris were phagocytosed by PMNs.     

A note on hydrolysis of tributyrin by Branhamella and Neisseria

Sixty-three strains of Branhamella and Neisseria were tested by two methods for their ability to hydrolyse glycerol tributyrate. After the conventional plate test, gas liquid chromatographical (GLC) analysis of the agar medium was carried out to detect the hydrolysis product, butyric acid, and other volatile fatty acids. All strains of Branhamella catarrhalis, Neisseria caviae, N. cuniculi and N. ovis but no other Neisseria spp. gave positive results with the conventional test. With GLC, however, most strains of Branhamella and Neisseria were shown to liberate butyric acid. In addition, some strains liberated acetic and isovaleric acids. Greater amounts of butyric acid were produced by clinical strains, in particular B. catarrhalis, compared with reference strains. It was concluded that the conventional plate test for tributyrin hydrolysis differentiates B. catarrhalis, N. caviae, N. cuniculi and N. ovis from other Neisseria.     

Genotypic and phenotypic relatedness of 80 strains of Branhamella catarrhalis of worldwide origin

Abstract 80 clinical Branhamella catarrhalis strains of worldwide origin were examined for genotypic relatedness and phenotypic characteristics. Using a quantitative bacterial dot method for DNA-DNA hybridization the strains were found to form a homogeneous group with ΔT_m-values ranging from 0.0–2.3°C. In Minibact-N, an identification kit for oxidase positive, Gram-negative diplococci using eight phenotypic characteristics, all isolates were correctly identified and also demonstrated complete homogeneity except for β-lactamase production. Type strains representing the genera Branhamella, Moraxella and Neisseria were also examined for comparison. B. catarrhalis strain NCTC 4103-known to be atypical-had a ΔT_m-value of 5.7°C and produced γ-glutamylaminopeptidase, in contrast to all other B. catarrhalis strains. In GN MicroPlate^TM, a kit which tests utilizable carbon sources, B. catarrhalis strains were found to be able to utilize up to 16 to 95 carbon sources.     

A note on susceptibility of Branhamella catarrhalis to heavy metals

The susceptibility of 56 strains of Branhamella catarrhalis and ten Neisseria spp. to arsenate, silver, nickel, mercury, lead, cadmium, chromium, manganese, iron, cobalt and molybdenum was tested with an agar dilution technique. All but two strains of B. catarrhalis were resistant to multiple metal ions. There were not sufficient differences in susceptibility, however, to allow the development of a typing scheme based on resistograms. Heavy metal resistance in Branhamella was unrelated to beta-lactamase production. Neisseria spp. were more susceptible to metal ions than B. catarrhalis and this may form the basis of a simple diagnostic test.     

Ultrastructural study on the adherence of Branhamella catarrhalis to oropharyngeal epithelial cell.

In the present study, it was observed that Branhamella catarrhalis adhere to the microplicae of the oropharyngeal epithelial cells. Both long and short microplicae patterns are present on the surface of oropharyngeal epithelial cells and the adherence ability of fimbriated Branhamella catarrhalis also varies according to the microplicae pattern. It was found that Branhamella catarrhalis attached more to one surface of the epithelial cell than to the other, suggesting that the presence of receptors are more on one surface than on the other. Branhamella catarrhalis did not attach to the mucus layer but directly to the epithelial cell surface. Ruthenium red staining specimen showed that Branhamella catarrhalis attached to a granular ruthenium red positive layer on the microplicae and also to a ruthenium red positive component, external to the unit membrane of the epithelial cell membrane.     

A note on susceptibility of Branhamella catarrhalis to heavy metals

The susceptibility of 56 strains of Branhamella catarrhalis and ten Neisseria spp. to arsenate, silver, nickel, mercury, lead, cadmium, chromium, manganese, iron, cobalt and molybdenum was tested with an agar dilution technique. All but two strains of B. catarrhalis were resistant to multiple metal ions. There were not sufficient differences in susceptibility, however, to allow the development of a typing scheme based on resistograms. Heavy metal resistance in Branhamella was unrelated to beta-lactamase production. Neisseria spp. were more susceptible to metal ions than B. catarrhalis and this may form the basis of a simple diagnostic test.     

Haemagglutinins and Fimbriae in Different Serotypes and Biotypes of Yersinia enterocolitica

When cultured in static broth at 20°C, 46 of 115 strains of Yersinia enterocolitica , diverse in biotype and serotype, produced a broad-spectrum mannose-resistant (MR) adhesin that agglutinated the erythrocytes of all of 10 animal species examined. The production of haemagglutinin (HA) was associated with the presence of fimbriae of S nm diameter. Culture of HA+ strains at 37° resulted in the disappearance of haemagglutinating ability and loss of fimbrial production. Strains of Y. enterocolitica with K1 antigen produced an MR adhesin that agglutinated only fowl erythrocytes and was associated with fimbriae of 4–4.5 nm diameter. None of 14 strains of Y. pseudotuberculosis was haemagglutinating.     

Fimbriae and haemagglutinating properties in Edwardsiella and Levinea.

Using the haemagglutinating procedure, the presence of fimbriae was revealed in all 34 Edwardsiella strains tested and in 3 Levinea strains out of 17 examined. The great majority of fimbriate Edwardsiella strains caused haemagglutination of MR type; only two strains of Edwardsiella and all fimbriate strains of Levinea brought about haemagglutination of MS type. All the strains exhibited a similar spectrum of haemagglutinating activity when they were tested against red cells of various animal species. Under the electron microscope MR fimbriae of Edwardsiella were found to be thicker than MS fimbriae (6.5 nm and 6.0 nm respectively). MS fimbriae of Levinea had 6.5 nm in diameter.     

Relationships among Broad-Spectrum and Narrow-Spectrum Mannose-Resistant Fimbrial Hemagglutinins in Different Yersinia Species

When 117 strains of Yersinia were grown in serial broth culture at 22 or 30 C, 44 (38%) formed a broad-spectrum, mannose-resistant hemagglutinin (MR/Y-HA) associated with thick (8 nm) channelled (MR/Y) fimbriae; eight other strains (7%) formed a narrow-spectrum, mannose-resistant hemagglutinin (MR/K-like HA) associated with thin (4 nm) non-channelled (type 3-like) fimbriae. The distribution of these two fimbrial hemagglutinins in different Yersinia species is discussed and their properties are compared with those of other reported mannose-resistant hemagglutinins. The thick fimbriae of four Yersinia species were antigenically similar as judged by immunoelectronmicroscopy. Likewise, the thin fimbriae of three Yersinia species were antigenically similar, though different from the thick fimbriae.     

Possible presence of a capsule in Branhamella catarrhalis.

Clinical isolates of Branhamella catarrhalis from patients with respiratory infections were used in this study. Electron microscopic observation after treating Branhamella catarrhalis with immune serum and ruthenium red revealed the capsule. In the phagocytosis test, most organisms were not ingested by human polymorphonuclear neutrophils in the presence of normal rabbit serum (NRS), while organisms were primarily cell associated and apparently ingested in the presence of immunized rabbit serum (IRS). The capsule may be one of the virulence factors in this bacteria. This study demonstrates the possible presence of a capsule in Branhamella catarrhalis.     

Moraxella (Branhamella) catarrhalis: Restriction enzyme analysis typing with HinfI, HaeIII and PstI

Abstract Restriction enzyme analysis typing with Hin fI, Hae III and Pst I was performed on Moraxella (Branhamella) catarrhalis strains consecutively collected from children suspected of respiratory tract infection and the type strain. Use of Hin fI gave the most distinct patterns. Great polymorphism was seen among strains.     

Isolation and characterization of conditional adherent and non-type 1 fimbriated Salmonella typhimurium mutants

Mutations in the genes encoding the type 1 fimbriae of Salmonella typhimurium were isolated by selecting for the deletion of Tn10 inserted adjacent to the chromosomal fim+ genes and screening for the loss of mannose-sensitive haemagglutination (HA) activity. S. typhimurium strains with Tn10 insertions in ahp were hypersensitive to peroxides, and tetracycline-sensitive derivatives of ahp::Tn10 mutants displayed two fim mutant phenotypes. The predominant class of fim mutants did not synthesize type 1 fimbriae. A second type of fim mutant synthesized type 1 fimbriae and exhibited a conditional lipoic acid requirement for HA. A fim-lip conditional mutant synthesized type 1 fimbriae when grown in Mueller-Hinton broth but the haemagglutinating activity of the fimbriae was dependent upon the addition of lipoic acid to the growth medium. Independently isolated lip mutations did not demonstrate a similar pleiotropic effect on HA. Western blots of fimbriae extracted from a fim-lip conditional mutant that was grown under permissive and restrictive conditions indicated the presence of 33 and 36.6 kDa proteins in HA+ fimbriae that were absent in HA- fimbriae. The HA+ phenotype of both conditional and non-fimbriated mutants was restored by transformation with cloned genes encoding S. typhimurium type 1 fimbriae.     

Immunological relationships of some non-pathogenic Neisseria.

A collection of nine stock strains of non-pathogenic Neisseria species and one strain of Branhamella catarrhalis has been examined serologically by a simple immunodiffusion technique utilising cultures growing on solid medium.     

A new mannose-resistant haemagglutinin in Klebsiella

Strains of Klebsiella of the species (or 'patho-bio-sero-geogroups') Kl. atlantae, Kl. edwardsii and Kl. rhinoscleromatis produced neither haemagglutinins (HAs) nor fimbriae; strains of Kl. ozaenae were HA- but some produced type-6 fimbriae; and strains of Kl. pneumoniae (sensu stricto) and Kl. aerogenes that produced the mannose-sensitive HA (MS-HA) formed type-1 fimbriae. Most strains of Kl. aerogenes produced, in addition, one or both of the mannose-resistant HAs, MR/K-HA or MR/P-HA. The former, associated with type-3 fimbriae, was produced by 95%, and the latter by 57%, of the Kl. aerogenes strains. Some of the properties of the MR/P-HA, apparently a non-fimbrial HA not previously recognised in Klebsiella, are described.     

A new mannose-resistant haemagglutinin in Klebsiella

Strains of Klebsiella of the species (or 'patho-bio-sero-geogroups') Kl. atlantae, Kl. edwardsii and Kl. rhinoscleromatis produced neither haemagglutinins (HAs) nor fimbriae; strains of Kl. ozaenae were HA^- but some produced type-6 fimbriae; and strains of Kl. pneumoniae ( sensu stricto ) and Kl. aerogenes that produced the mannose-sensitive HA (MS-HA) formed type-1 fimbriae. Most strains of Kl. aerogenes produced, in addition, one or both of the mannose-resistant HAs, MR/K-HA or MR/P-HA. The former, associated with type-3 fimbriae, was produced by 95%, and the latter by 57%, of the Kl. aerogenes strains. Some of the properties of the MR/P-HA, apparently a non-fimbrial HA not previously recognised in Klebsiella , are described.     

Lipids of Branhamella catarrhalis and Neisseria gonorrhoeae.

Three strains of Branhamella catarrhalis and three strains of Neisseria gonorrhoeae were analyzed with regard to their phospholipid and neutral lipid composition. B. catarrhalis (ATCC 23246) contained 5.12 +/- 0.34% lipid, determined gravimetrically, compared to 8.56 +/- 0.15% and 9.73 +/- 0.06% for two strains of N. gonorrhoeae. Cardiolipin, phosphatidylglycerol, and phosphatidyl-ethanolamine were identified in extracts of both species. In addition, B. catarrhalis contained small amounts of phosphatidylcholine, and N. gonorrhoeae contained small amounts of lyso-phosphatidylethanolamine, which accumulated with autolysis accompanying late cell culture growth. The kinetics of change of relative amounts of phospholipids in both species were measured and found to differ substantially. Neutral lipid accounted for 30.4% of the total lipid of B. catarrhalis (ATCC 23246) and 7.6% of the total lipid of N. gonorrhoeae NYH 002. Hydrocarbons, triglycerides, free fatty acids, coenzyme Q, diglycerides, and free hydroxy fatty acids were identified in the neutral lipid fraction of both species. The three strains of N. gonorrhoeae, sensitive, intermediate, and resistant to penicillin, exhibited no significant difference in the composition or metabolism of phospholipid.     

Complement resistance is a virulence factor of Branhamella (Moraxella) catarrhalis

Abstract The purpose of this study was to investigate complement resistance in Branhamella (Moraxella) catarrhalis isolated from healthy schoolchildren or sputum-producing adult patients. Two techniques were used: a serum bactericidal assay as the gold standard and an easier ‘culture and spot’ test. Children (age 4–13; n = 303) and patients ( n = 1047) showed high colonization/infection rates with B. catarrhalis (31% and 19%, respectively). Complement resistance or intermediate sensitivity occurred frequently in patient isolates (62% and 27%, respectively) and less often in children (33% and 8.5%, respectively; P ⪡ 0.0001). In young children (age 4–5 years), the proportion of complement-resistant strains was around 50%. Complement resistance in B. catarrhalis is associated with illness and may hence be considered a virulence factor.     

Studies on the cellular and free lipopolysaccharides from Branhamella catarrhalis.

Cellular and free lipopolysaccharides obtained from Neisseria catarrhalis and Branhamella catarrhalis were found to be essentially identical. Both cellular and free lipopolysaccharides contained core-oligosaccharides of the following composition: D-glucose (4 mol), D-galactose (1 mol), 2-amino-2-deoxy-D-glucose (1 mol), and 3-deoxy-D-manno-octulosonic acid. Aldoheptose and phosphate components were below levels of detection. Several physical methods indicated that all core-oligosaccharide preparations were identical. Lipid A preparations from cellular and free lipopolysaccharides of both organisms were qualitatively and quantitatively similar; they were composed of decanoic acid, dodecanoic acid, 3-hydroxy dodecanoic acid, 2-amino-2-deoxy-D-glucose, phosphate, and ethanolamine. The results tend to justify the transfer of Neisseria catarrhalis to the genus Branhamella.