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Highly Divergent Clostridium difficile Strains Isolated from the Environment

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ABSTRACT

Clostridium difficile is one of the most important human and animal pathogens. However, the bacterium is ubiquitous and can be isolated from various sources. Here we report the prevalence and characterization of C. difficile in less studied environmental samples, puddle water (n = 104) and soil (n = 79). C. difficile was detected in 14.4% of puddle water and in 36.7% of soil samples. Environmental strains displayed antimicrobial resistance patterns comparable to already published data of human and animal isolates. A total of 480 isolates were grouped into 34 different PCR ribotypes. More than half of these (52.9%; 18 of 34) were already described in humans or animals. However, 14 PCR ribotypes were new in our PCR ribotype library and all but one were non-toxigenic. The multilocus sequence analysis of these new PCR ribotypes revealed that non-toxigenic environmental isolates are phylogenetically distinct and belong to three highly divergent clades, two of which have not been described before. Our data suggest that environment is a potential reservoir of genetically diverse population of C. difficile.

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Phylogenetic relationship of C. difficile based on MLST sequences.The Maximum likelihood phylogenetic tree was constructed based on the alignment of concatenated DNA sequences of the seven housekeeping genes. Clades 1–5 and C-I were already described and clades C-II and C-III are new. In clade C-II a strain with ST200 (toxinotype XXXII) is present and is described in our recent publication [36].
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pone.0167101.g001: Phylogenetic relationship of C. difficile based on MLST sequences.The Maximum likelihood phylogenetic tree was constructed based on the alignment of concatenated DNA sequences of the seven housekeeping genes. Clades 1–5 and C-I were already described and clades C-II and C-III are new. In clade C-II a strain with ST200 (toxinotype XXXII) is present and is described in our recent publication [36].

Mentions: Representatives of PCR ribotypes which were newly identified in this study were further analyzed by MLST. Identity of these isolates was confirmed by the 16S rDNA analysis (S1 Table) and analysis of two additional genes rpoB (S2 Table, S2 Fig) and gyrB (S3 Table and S3 Fig). Fifteen strains, one toxigenic and 14 non-toxigenic (where 115-bp insertion could not be PCR amplified) belonging to 13 distinct ribotypes (for PCR ribotype SLO 204, three isolates were included) were sequenced and their MLST- sequence types were determined from the sequences. Twelve sequence types were identified, all of which were new (ST 335 to ST339 and ST341 to ST347, Table 3). Phylogenetic tree based on concatenated MLST sequences of environmental strains and representatives of all previously described clades [34] demonstrated two new, highly divergent lineages, here designated as C-II and C-III (following the designations introduced by Dingle et al. [34]). Clade C-II included two isolates and clade C-III contained 12 isolates represented by 10 different PCR ribotypes, all but one (toxinotype XXXII (A-B+), PCR ribotype SLO 240) were non-toxigenic. Only a single strain was found in recently described clade C-I (Fig 1).


Highly Divergent Clostridium difficile Strains Isolated from the Environment
Phylogenetic relationship of C. difficile based on MLST sequences.The Maximum likelihood phylogenetic tree was constructed based on the alignment of concatenated DNA sequences of the seven housekeeping genes. Clades 1–5 and C-I were already described and clades C-II and C-III are new. In clade C-II a strain with ST200 (toxinotype XXXII) is present and is described in our recent publication [36].
© Copyright Policy
Related In: Results  -  Collection

License
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getmorefigures.php?uid=PMC5120845&req=5

pone.0167101.g001: Phylogenetic relationship of C. difficile based on MLST sequences.The Maximum likelihood phylogenetic tree was constructed based on the alignment of concatenated DNA sequences of the seven housekeeping genes. Clades 1–5 and C-I were already described and clades C-II and C-III are new. In clade C-II a strain with ST200 (toxinotype XXXII) is present and is described in our recent publication [36].
Mentions: Representatives of PCR ribotypes which were newly identified in this study were further analyzed by MLST. Identity of these isolates was confirmed by the 16S rDNA analysis (S1 Table) and analysis of two additional genes rpoB (S2 Table, S2 Fig) and gyrB (S3 Table and S3 Fig). Fifteen strains, one toxigenic and 14 non-toxigenic (where 115-bp insertion could not be PCR amplified) belonging to 13 distinct ribotypes (for PCR ribotype SLO 204, three isolates were included) were sequenced and their MLST- sequence types were determined from the sequences. Twelve sequence types were identified, all of which were new (ST 335 to ST339 and ST341 to ST347, Table 3). Phylogenetic tree based on concatenated MLST sequences of environmental strains and representatives of all previously described clades [34] demonstrated two new, highly divergent lineages, here designated as C-II and C-III (following the designations introduced by Dingle et al. [34]). Clade C-II included two isolates and clade C-III contained 12 isolates represented by 10 different PCR ribotypes, all but one (toxinotype XXXII (A-B+), PCR ribotype SLO 240) were non-toxigenic. Only a single strain was found in recently described clade C-I (Fig 1).

View Article: PubMed Central - PubMed

ABSTRACT

Clostridium difficile is one of the most important human and animal pathogens. However, the bacterium is ubiquitous and can be isolated from various sources. Here we report the prevalence and characterization of C. difficile in less studied environmental samples, puddle water (n = 104) and soil (n = 79). C. difficile was detected in 14.4% of puddle water and in 36.7% of soil samples. Environmental strains displayed antimicrobial resistance patterns comparable to already published data of human and animal isolates. A total of 480 isolates were grouped into 34 different PCR ribotypes. More than half of these (52.9%; 18 of 34) were already described in humans or animals. However, 14 PCR ribotypes were new in our PCR ribotype library and all but one were non-toxigenic. The multilocus sequence analysis of these new PCR ribotypes revealed that non-toxigenic environmental isolates are phylogenetically distinct and belong to three highly divergent clades, two of which have not been described before. Our data suggest that environment is a potential reservoir of genetically diverse population of C. difficile.

No MeSH data available.


Related in: MedlinePlus