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Genetic tools for studying Capnocytophaga canimorsus.

ABSTRACT: Capnocytophaga canimorsus, a commensal bacterium from canine oral flora, has been isolated throughout the world from severe human infections caused by dog bites. Due to the low level of evolutionary relationship to Proteobacteria, genetic methods suitable for the genus Capnocytophaga needed to be established. Here, we show that Tn4351, derived from Bacteroides fragilis, could be introduced by conjugation into C. canimorsus and conferred resistance to erythromycin. By mapping and sequencing a naturally occurring plasmid isolated from a clinical isolate of C. canimorsus, we identified a repA gene that allowed us to construct Escherichia coli-Capnocytophaga shuttle vectors. Most commonly used antibiotic markers were not functional in C. canimorsus, but cefoxitin (cfxA), tetracycline (tetQ), and erythromycin (ermF) resistances could be used as markers for plasmid maintenance in C. canimorsus and even in some other Capnocytophaga spp. Shuttle vectors were introduced into C. canimorsus either by conjugation using the origin of transfer (oriT) of RP4 or by electrotransformation. Taking advantage of the promoter of ermF, an expression vector was constructed. Finally, a method that allows site-directed mutagenesis is described. All these genetic tools pave the way, not only for molecular studies of the pathogenesis of C. canimorsus, but also for studies of other oral Capnocytophaga species.

SUBMITTER: Mally M

PROVIDER: S-EPMC2570293 | biostudies-literature | 2008 Oct

REPOSITORIES: biostudies-literature

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Genetic tools for studying Capnocytophaga canimorsus.

Mally Manuela M Cornelis Guy R GR

Applied and environmental microbiology 20080822 20

Capnocytophaga canimorsus, a commensal bacterium from canine oral flora, has been isolated throughout the world from severe human infections caused by dog bites. Due to the low level of evolutionary relationship to Proteobacteria, genetic methods suitable for the genus Capnocytophaga needed to be established. Here, we show that Tn4351, derived from Bacteroides fragilis, could be introduced by conjugation into C. canimorsus and conferred resistance to erythromycin. By mapping and sequencing a nat ...[more]

PMID: 18723653

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Project description:Capnocytophaga canimorsus and Capnocytophaga cynodegmi are commensal bacteria in the oral cavities of dogs. Both are zoonotic pathogens that could infect humans via dog bites. C. canimorsus may cause life-threatening infections in humans, whereas C. cynodegmi infections tend to be milder and more localized. Capsular serovars A-C of C. canimorsus seem to be virulence-associated. Some of the C. canimorsus serovars described to date can also be detected in other Capnocytophaga species, including C. cynodegmi. The objective of this pilot study was to investigate the emergence of C. canimorsus and C. cynodegmi after birth in oral cavities of puppies and to evaluate the impact of the dam's Capnocytophaga spp. carrier status on the emergence. Ten litters, altogether 59 puppies, were included in the study. The puppies and their dams were sampled at five time points over seven weeks after whelping. Oral swab samples taken were investigated for the presence of C. canimorsus and C. cynodegmi by species-specific polymerase chain reaction (PCR), the specificity of which was verified by sequencing a selection of the PCR products. Samples that were positive in Capnocytophaga PCR reactions were also capsular-typed by PCR to gain more knowledge about the Capnocytophaga spp. present in the samples. Altogether 10.2% and 11.9% of puppies, or 20.0% and 30.0% of litters tested PCR-positive for C. canimorsus and C. cynodegmi, respectively. Capnocytophaga PCR-positive puppy samples were always positive for only C. cynodegmi or C. canimorsus, not both. Most Capnocytophaga PCR-positive puppies became positive at the age of 5 to 7 weeks. Only a minority (5/16) of the C. cynodegmi PCR-positive dog samples were positive in capsular typing PCR, whereas all C. canimorsus PCR-positive dog samples were negative in capsular typing PCR. For all Capnocytophaga PCR-positive puppies, their dam was positive for the same Capnocytophaga species. These results suggest that puppies become colonized by C. cynodegmi or C. canimorsus from their dams at the time of deciduous teeth eruption.

Dataset Information

Genetic tools for studying Capnocytophaga canimorsus.

Publications

Genetic tools for studying Capnocytophaga canimorsus.

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