Visible laser and UV-A radiation impact on a PNP degrading Moraxella strain and its rpoS mutant

The role of stationary phase sigma factor gene (rpoS) in the stress response of Moraxella strain when exposed to radiation was determined by comparing the stress responses of the wild-type (WT) and its rpoS knockout (KO) mutant. The rpoS was turned on by starving the WT cultures for 24 h in minimal salt medium. Under non-starved condition, both WT and KO planktonic Moraxella cells showed an increase in mortality with the increase in duration of irradiation. In the planktonic non-starved Moraxella, for the power intensity tested, UV radiation caused a substantially higher mortality rate than did by the visible laser light (the mortality rate observed for 15-min laser radiation was 53.4 +/- 10.5 and 48.7 +/- 8.9 for WT and KO, respectively, and 97.6 +/- 0 and 98.5 +/- 0 for 25 s of UV irradiation in WT and KO, respectively). However, the mortality rate decreased significantly in the starved WT when exposed to these two radiations. In comparison, rpoS protected the WT against the visible laser light more effectively than it did for the UV radiation. The WT and KO strains of Moraxella formed distinctly different types of biofilms on stainless steel coupons. The KO strain formed a denser biofilm than did the WT. Visible laser light removed biofilms from the surfaces more effectively than did the UV. This was true when comparing the mortality of bacteria in the biofilms as well. The inability of UV radiation to penetrate biofilms due to greater rates of surface absorption is considered to be the major reason for the weaker removal of biofilms in comparison to that of the visible laser light. This result suggests that high power visible laser light might be an effective tool for the removal of biofilms.     

Rapid typing of Moraxella catarrhalis subpopulations based on outer membrane proteins using mass spectrometry

Moraxella catarrhalis is a major mucosal pathogen of the human respiratory tract both in children and in adults. Two subpopulations of this organism have been described that differ in 16S rRNA gene sequence and virulence traits. Three 16S rRNA types have been defined. 2-DE followed by protein identification by MS revealed significant differences in the outer membrane protein (OMP) patterns of each M. catarrhalis 16S rRNA type. Approximately 130 features were detected on the 2-DE map of each M. catarrhalis 16S rRNA type. However, only 50 features were expressed by all strains. Furthermore, direct profiling of isolated OMP using MALDI-TOF MS resulted in a characteristic spectral fingerprint for each 16S rRNA type. Fingerprints remained identical when intact cells instead of isolated OMP were analyzed. This finding suggests that the source of desorbed ions is the outer membrane. Based on the fingerprint we were able to assign 18 well-characterized clinical M. catarrhalis isolates to the correct subpopulation. Therefore, MALDI-TOF of intact M. catarrhalis provides a rapid and robust tool for M. catarrhalis strain typing that could be applied in epidemiological studies.     

Unveiling electrotransformation of Moraxella catarrhalis as a process of natural transformation

The human respiratory tract pathogen Moraxella catarrhalis is a naturally competent microorganism. However, electrotransformation has long been used to introduce foreign DNA into this organism. This study demonstrated that electrotransformants obtained with linear or circular nonreplicating plasmid DNA originated exclusively from natural transformation processes taking place during the recovery phase after the application of current. Only replicating plasmid DNA could be introduced into M. catarrhalis by electrotransformation, in a type IV pilus-independent manner. Electrotransformation with homologous genomic DNA indicated that restriction of double-stranded DNA was independent of type III restriction-methylation systems. Nontransformability of M. catarrhalis by electrotransformation was observed using double- as well as single-stranded DNA. In addition, the study showed that natural competence is a very constant feature of M. catarrhalis.     

Moraxella catarrhalis does not grow on nutrient agar without sodium chloride supplementation

None of the 58 Moraxella catarrhalis strains grew on nutrient agar without sodium chloride supplementation, whereas 49 of 51 commensal Neisseria spp. strains tested did. Growth on nutrient agar without sodium chloride supplementation could be used for screening between M. catarrhalis and commensal Neisseria spp.     

Binding of bacteria to HEp-2 cells infected with influenza A virus

Epidemiological studies indicate influenza virus infection increases susceptibility to bacterial respiratory pathogens and to meningococcal disease. Because density of colonisation is an important factor in the development of bacterial disease, the objectives of the study were to use flow cytometry methods for assessment of bacterial binding and detection of cell surface antigens to determine: (1) if HEp-2 cells infected with human influenza A virus bind greater numbers of bacteria than uninfected cells; (2) if influenza infection alters expression of cell surface antigens which act as receptors for bacterial binding; (3) if neuraminidase affects binding of bacteria to HEp-2 cells. There was significantly increased binding of all isolates tested regardless of surface antigen characteristics. There were no significant differences between virus-infected and -uninfected Hep-2 cells in binding of monoclonal antibodies to Lewisb, Lewisx or H type 2. There were significant increases in binding of monoclonal antibodies to CD14 (P < 0.05) and CD18 (P < 0.01). Treatment of cells with monoclonal antibodies significantly reduced binding of Neisseria meningitidis strain C:2b:P1.2, CD14 (P < 0.001) and CD18 (P < 0.001). No reduction in binding of a strain of Streptococcus pneumoniae (12F) was observed in these experiments. Neuraminidase treatment of HEp-2 cells increased binding of monoclonal antibodies to CD14 (P < 0.01) and CD18 (P < 0.01). In three experiments, the increase in binding of meningococcal strain C:2b:P1.2 to neuraminidase-treated cells was not significant, but binding of Staphylococcus aureus strain NCTC 10655 was significant (P < 0.05).     

Construction of a Combined NotI/SmaI Physical and Genetic Map of Moraxella (Branhamella) catarrhalis Strain ATCC25238

Using pulsed field gel electrophoresis (PFGE) and Southern hybridization techniques, a physical map of Moraxella catarrhalis strain ATCC25238 was constructed to provide basic genetic knowledge of this bacterium that has attracted attention in recent years as a human pathogen. Restriction endonuclease NotI cut the genome into 10 fragments and SmaI into 9, and the molecular size of the genome was estimated to be 1,940 kilobases. Location of the 12 genes participating in the biosynthesis of purine, pyrimidine and nine kinds of amino acids were determined on the circular physical map of the strain.     

Structure of the peptidoglycans of Moraxella glucidolytica and Moraxella lwoffi grown on hydrocarbons

The peptidoglycans of Moraxella glucidolytica and Moraxella lwoffi grown on aliphatic hydrocarbons were isolated. They contained muramic acid. glucosamine, alanine, d-glutamic acid and meso-diaminopimelic acid in a molar ratio of about 0.5:0.5:1.6:1.0:1.0 (M. glucidolytica) and 0.8:0.7:1.3:1.0:1.0 (M. lwoffi).The peptidoglycans were lysozyme-resistant. However, when treated with formamide, they could be partially degraded by lysozyme. The fragments were purified and their structure determined. In both strains, the peptide subunits consisted mainly of tripeptides (l-Ala-d-Glu-meso-DAP) and tetrapeptides (l-Ala-d-Glu-meso-DAP-d-Ala), most of them being directly cross-linked. It is concluded that in both strains the primary structures of the peptidoglycans are closely related.     

Identification of a Moraxella catarrhalis gene that confers adherence to various human epithelial cell lines in vitro

Moraxella catarrhalis is a pathogen of the human airways. We found that expression of the M. catarrhalis gene mcmA by Escherichia coli increases adherence to epithelial cells 100-fold. Furthermore, we discovered that disrupting mcmA decreases M. catarrhalis adherence to laryngeal and lung cells, which are relevant to pathogenesis by the bacterium.     

Attachment of Moraxella catarrhalis to pharyngeal epithelial cells is mediated by a glycosphingolipid receptor

Abstract Moraxella catarrhalis is one of the major pathogens of respiratory infections and has the ability to attach to the pharyngeal cells via fimbriae. We characterized the epithelial cell receptor to which fimbriate M. catarrhalis binds. Neuraminidase pretreatment of pharyngeal epithelial cells resulted in a significant decrease of M. catarrhalis attachment, suggesting interaction with the sialic acid component. The attachment was not decreased in M. catarrhalis pretreated with 2 and 1 mg/ml of fucose, N -acetyl-neuraminic acid, N -acetyl-glucosamine, N -acetyl-galactosamine, acetyl-salicylic acid and colominic acid. However, M. catarrhalis treated with gangliosides M1, M2, D1a, D1b and T1a at a concentration of 2.5 μg/ml had significantly decreased the attachment compared to the control. In contrast treatment with gangliosides M3 and asialoganglioside M1 did not decrease the attachment of M. catarrhalis and thereby provided evidence for specificity of the inhibition. Concentration dependent effects of ganglioside M2 on the attachment were also observed. Other fimbriate isolates of M. catarrhalis showed decrease in attachment after treatment with ganglioside M2. However there was no effect on attachment when a nonfimbriate isolate was treated with ganglioside M2. This study indicates that the receptor of fimbriate M. catarrhalis on pharyngeal epithelial cells resides in the sequences of ganglioside M2.