Intracellular viability of toxigenic<i>Corynebacterium diphtheriae</i>strains in HEp-2 cells

Hirata, Raphael; Napoleão, Fátima; Monteiro-Leal, Luiz Henrique; Andrade, Arnaldo F.B.; Nagao, Prescilla Emy; Formiga, Luiz Carlos Duarte; Fonseca, Leila de Souza; Mattos‐Guaraldi, Ana Luíza

doi:10.1111/j.1574-6968.2002.tb11379.x

Cited by 35 publications

(12 citation statements)

References 19 publications

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“…Adhesion of different C. diphtheriae strains to lung, pharyngeal, and laryngeal epithelial cells and to erythrocytes has been reported (Colombo et al 2001;Bertuccini et al 2004;Hirata et al 2002Hirata et al , 2004. Although it is known that both cell-wall-linked surface proteins and fibrous protein polymers, for example fimbriae and pili, are of major importance in the adhesion process for many types of Gram-positive bacteria, many aspects of the underlying mechanisms remain unclear (Rogers et al 2011).…”

Section: Introductionmentioning

confidence: 97%

Contour and persistence length of Corynebacterium diphtheriae pili by atomic force microscopy

et al. 2012

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Many bacteria are characterized by nanoscale ultrastructures, for example S-layers, flagella, fimbriae, or pili. The last two are especially important for attachment to different abiotic and biotic surfaces and for host-pathogen interactions. In this study, we investigated the geometric and elastic properties of pili of different Corynebacterium diphtheriae strains by atomic force microscopy (AFM). We performed quantitative contour-length analysis of bacterial pili and found that the visible contour length of the pili can be described by a log-normal distribution. Our data revealed significant strain-specific variations in the mean visible contour length of the pili, ranging from 260 to 1,590 nm. To estimate their full contour length, which is not directly accessible from the AFM images, we developed a simple correction model. Using this model, we determined the mean full contour length as 510-2,060 nm. To obtain the persistence length we used two different methods of analysis, one based on the end-to-end distance of the pili and one based on the bending angles of short segments. In comparison, the bending angle analysis proved to be more precise and resulted in persistence lengths in the narrow range of 220-280 nm, with no significant strain-specific variations. This is small compared with some other bacterial polymers, for example type IV pili, F-pili, or flagella.

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Section: Introductionmentioning

confidence: 97%

Contour and persistence length of Corynebacterium diphtheriae pili by atomic force microscopy

et al. 2012

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“…Using a combination of gentamicin protection assays and thin-section electron microscopy, Hirata and co-workers [8] showed that toxigenic C. diphtheriae were not only able to adhere to laryngeal HEp-2 cells, but also enter these cells and survive after internalization. Similar observations were made for non-toxigenic strains [9] showing that also pharyngeal Detroit 562 cells can be invaded by C. diphtheriae .…”

Section: Introductionmentioning

confidence: 99%

Corynebacterium diphtheriae invasion-associated protein (DIP1281) is involved in cell surface organization, adhesion and internalization in epithelial cells

et al. 2010

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BackgroundCorynebacterium diphtheriae, the causative agent of diphtheria, is well-investigated in respect to toxin production, while little is known about C. diphtheriae factors crucial for colonization of the host. In this study, we investigated the function of surface-associated protein DIP1281, previously annotated as hypothetical invasion-associated protein.ResultsMicroscopic inspection of DIP1281 mutant strains revealed an increased size of the single cells in combination with an altered less club-like shape and formation of chains of cells rather than the typical V-like division forms or palisades of growing C. diphtheriae cells. Cell viability was not impaired. Immuno-fluorescence microscopy, SDS-PAGE and 2-D PAGE of surface proteins revealed clear differences of wild-type and mutant protein patterns, which were verified by atomic force microscopy. DIP1281 mutant cells were not only altered in shape and surface structure but completely lack the ability to adhere to host cells and consequently invade these.ConclusionsOur data indicate that DIP1281 is predominantly involved in the organization of the outer surface protein layer rather than in the separation of the peptidoglycan cell wall of dividing bacteria. The adhesion- and invasion-negative phenotype of corresponding mutant strains is an effect of rearrangements of the outer surface.

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“…Using a combination of gentamicin protection assays and thin-section electron microscopy, Hirata and co-workers [9] showed that toxigenic C. diphtheriae are not only able to adhere to laryngeal HEp-2 cells, but also enter these cells and survive after internalization. Similar observations were made for non-toxigenic strains [10] showing that also pharyngeal Detroit 562 cells can be invaded by C. diphtheriae and that viable intracellular bacteria can be detected up to 48 h after infection.…”

Section: Introductionmentioning

confidence: 99%

Strain-specific differences in pili formation and the interaction of Corynebacterium diphtheriae with host cells

et al. 2010

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BackgroundCorynebacterium diphtheriae, the causative agent of diphtheria, is well-investigated in respect to toxin production, while little is known about C. diphtheriae factors crucial for colonization of the host. In this study, we investigated strain-specific differences in adhesion, invasion and intracellular survival and analyzed formation of pili in different isolates.ResultsAdhesion of different C. diphtheriae strains to epithelial cells and invasion of these cells are not strictly coupled processes. Using ultrastructure analyses by atomic force microscopy, significant differences in macromolecular surface structures were found between the investigated C. diphtheriae strains in respect to number and length of pili. Interestingly, adhesion and pili formation are not coupled processes and also no correlation between invasion and pili formation was found. Using RNA hybridization and Western blotting experiments, strain-specific pili expression patterns were observed. None of the studied C. diphtheriae strains had a dramatic detrimental effect on host cell viability as indicated by measurements of transepithelial resistance of Detroit 562 cell monolayers and fluorescence microscopy, leading to the assumption that C. diphtheriae strains might use epithelial cells as an environmental niche supplying protection against antibodies and macrophages.ConclusionsThe results obtained suggest that it is necessary to investigate various isolates on a molecular level to understand and to predict the colonization process of different C. diphtheriae strains.

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Intracellular viability of toxigenicCorynebacterium diphtheriaestrains in HEp-2 cells

Cited by 35 publications

References 19 publications

Contour and persistence length of Corynebacterium diphtheriae pili by atomic force microscopy

Contour and persistence length of Corynebacterium diphtheriae pili by atomic force microscopy

Corynebacterium diphtheriae invasion-associated protein (DIP1281) is involved in cell surface organization, adhesion and internalization in epithelial cells

Strain-specific differences in pili formation and the interaction of Corynebacterium diphtheriae with host cells

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