Genomic evolution and virulence association of Clostridioides difficile sequence type 37 (ribotype 017) in China¶

To investigate a possible Clostridioides difficile reservoir in the Czech Republic, we performed a study in 297 calves from 29 large-scale dairy farms. After enrichment, faecal samples were inoculated onto selective agar for C. difficile. From the 297 samples, 44 C. difficile isolates were cultured (prevalence of 14.8%, 10 farms). The Holstein breed and use of digestate were associated with C. difficile colonisation (p ˂ 0.05). C. difficile isolates belonged to the ribotype/sequence type: RT033/ST11 (n = 37), RT126/ST11 (n = 6) and RT046/ST35 (n = 1). A multiple-locus variable-number tandem-repeat analysis revealed four clonal complexes of RT033 isolates and one clonal complex of RT126 isolates. All isolates were sensitive to amoxicillin, metronidazole and vancomycin. Forty isolates were resistant to ciprofloxacin, twenty-one to clindamycin, seven to erythromycin, seven to tetracycline and six to moxifloxacin. Moxifloxacin resistant isolates revealed an amino-acid substitution Thr82Ile in the GyrA. In conclusion, the calves of Holstein breed from farms using digestate as a product of bio-gas plants are more likely to be colonised by clonally-related C. difficile of ST 11 represented by ribotypes 033 and 126. The identified resistance to moxifloxacin with a Thr82Ile substitution in the GyrA highlights the need for further monitoring by the “One health approach”.

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A global to local genomics analysis of Clostridioides difficile ST1/RT027 identifies cryptic transmission events in a northern Arizona healthcare network

Microbial Genomics ◽

10.1099/mgen.0.000271 ◽

2019 ◽

Vol 5 (7) ◽

Cited By ~ 2

Author(s):

Charles H. D. Williamson ◽

Nathan E. Stone ◽

Amalee E. Nunnally ◽

Heidie M. Hornstra ◽

David M. Wagner ◽

...

Keyword(s):

Type Species ◽

Sequence Type ◽

Infection Prevention And Control ◽

Polymorphism Analysis ◽

Whole Genome ◽

Content Type ◽

Northern Arizona ◽

Healthcare Network ◽

Link Type ◽

Clostridioides Difficile

Clostridioides difficile is a ubiquitous, diarrhoeagenic pathogen often associated with healthcare-acquired infections that can cause a range of symptoms from mild, self-limiting disease to toxic megacolon and death. Since the early 2000s, a large proportion of C. difficile cases have been attributed to the ribotype 027 (RT027) lineage, which is associated with sequence type 1 (ST1) in the C. difficile multilocus sequence typing scheme. The spread of ST1 has been attributed, in part, to resistance to fluoroquinolones used to treat unrelated infections, which creates conditions ideal for C. difficile colonization and proliferation. In this study, we analysed 27 isolates from a healthcare network in northern Arizona, USA, and 1352 publicly available ST1 genomes to place locally sampled isolates into a global context. Whole genome, single nucleotide polymorphism analysis demonstrated that at least six separate introductions of ST1 were observed in healthcare facilities in northern Arizona over an 18-month sampling period. A reconstruction of transmission networks identified potential nosocomial transmission of isolates, which were only identified via whole genome sequence analysis. Antibiotic resistance heterogeneity was observed among ST1 genomes, including variability in resistance profiles among locally sampled ST1 isolates. To investigate why ST1 genomes are so common globally and in northern Arizona, we compared all high-quality C. difficile genomes and identified that ST1 genomes have gained and lost a number of genomic regions compared to all other C. difficile genomes; analyses of other toxigenic C. difficile sequence types demonstrate that this loss may be anomalous and could be related to niche specialization. These results suggest that a combination of antimicrobial resistance and gain and loss of specific genes may explain the prominent association of this sequence type with C. difficile infection cases worldwide. The degree of genetic variability in ST1 suggests that classifying all ST1 genomes into a quinolone-resistant hypervirulent clone category may not be appropriate. Whole genome sequencing of clinical C. difficile isolates provides a high-resolution surveillance strategy for monitoring persistence and transmission of C. difficile and for assessing the performance of infection prevention and control strategies.

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Hypervirulent clade 2, ribotype 019/sequence type 67 Clostridioides difficile strain from Japan

Gut Pathogens ◽

10.1186/s13099-019-0336-3 ◽

2019 ◽

Vol 11 (1) ◽

Cited By ~ 2

Author(s):

Ryoichi Saito ◽

Yukino Usui ◽

Alafate Ayibieke ◽

Jun Nakajima ◽

Isaac Prah ◽

...

Keyword(s):

Base Pair ◽

Reference Strain ◽

Draft Genome ◽

Sequence Type ◽

Cytotoxicity Assay ◽

Nucleotide Identity ◽

Limited Data ◽

Nucleotide Substitutions ◽

Clostridioides Difficile ◽

First Time

Abstract Background Clostridioides difficile ribotype (RT) 019/sequence type (ST) 67 strains belong to a hypervirulent lineage closely related to RT027/ST1; however, limited data are available for hypervirulent clade 2 lineages in Japan. Herein, we report the draft genome of a C. difficile strain B18-123 belonging to clade 2, RT019/ST67 for the first time in Japan. Results The pathogenicity locus carried by B18-123 (19.6 kb) showed higher homology (97.29% nucleotide identity) with strain R20291 (RT027/ST1) than the reference strain 630 (RT012/ST54), and B18-123 harbored 8-nucleotide substitutions in tcdC. However, it did not contain an 18-base pair (bp) deletion or a single-bp deletion at position 117 in tcdC, which was identified in the previous strain R20291. A cytotoxicity assay revealed similar cytotoxicity levels between strains B18-123 and ATCC BAA-1870 (RT027/ST1). The B18-123 strain was found to be susceptible to metronidazole and vancomycin. Conclusion Our findings contribute to the further understanding of the characteristics of hypervirulent clade 2 including RT019/ST67 lineages.

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Evolutionary and Genomic Insights intoClostridioides difficileSequence Type 11: a Diverse Zoonotic and Antimicrobial-Resistant Lineage of Global One Health Importance

mBio ◽

10.1128/mbio.00446-19 ◽

2019 ◽

Vol 10 (2) ◽

Cited By ~ 22

Author(s):

Daniel R. Knight ◽

Brian Kullin ◽

Grace O. Androga ◽

Frederic Barbut ◽

Catherine Eckert ◽

...

Keyword(s):

Clostridium Difficile ◽

Antimicrobial Resistance ◽

Long Range ◽

One Health ◽

Core Genome ◽

Sequence Type ◽

Content Type ◽

Pan Genome ◽

Clostridioides Difficile ◽

Animal Populations

ABSTRACTClostridioides difficile(Clostridium difficile) sequence type 11 (ST11) is well established in production animal populations worldwide and contributes considerably to the global burden ofC. difficileinfection (CDI) in humans. Increasing evidence of shared ancestry and genetic overlap of PCR ribotype 078 (RT078), the most common ST11 sublineage, between human and animal populations suggests that CDI may be a zoonosis. We performed whole-genome sequencing (WGS) on a collection of 207 ST11 and closely related ST258 isolates of human and veterinary/environmental origin, comprising 16 RTs collected from Australia, Asia, Europe, and North America. Core genome single nucleotide variant (SNV) analysis identified multiple intraspecies and interspecies clonal groups (isolates separated by ≤2 core genome SNVs) in all the major RT sublineages: 078, 126, 127, 033, and 288. Clonal groups comprised isolates spread across different states, countries, and continents, indicative of reciprocal long-range dissemination and possible zoonotic/anthroponotic transmission. Antimicrobial resistance genotypes and phenotypes varied across host species, geographic regions, and RTs and included macrolide/lincosamide resistance (Tn6194[ermB]), tetracycline resistance (Tn6190[tetM] and Tn6164[tet44]), and fluoroquinolone resistance (gyrA/Bmutations), as well as numerous aminoglycoside resistance cassettes. The population was defined by a large “open” pan-genome (10,378 genes), a remarkably small core genome of 2,058 genes (only 19.8% of the gene pool), and an accessory genome containing a large and diverse collection of important prophages of theSiphoviridaeandMyoviridae. This study provides novel insights into strain relatedness and genetic variability ofC. difficileST11, a lineage of global One Health importance.IMPORTANCEHistorically,Clostridioides difficile(Clostridium difficile) has been associated with life-threatening diarrhea in hospitalized patients. Increasing rates ofC. difficileinfection (CDI) in the community suggest exposure toC. difficilereservoirs outside the hospital, including animals, the environment, or food.C. difficilesequence type 11 (ST11) is known to infect/colonize livestock worldwide and comprises multiple ribotypes, many of which cause disease in humans, suggesting CDI may be a zoonosis. Using high-resolution genomics, we investigated the evolution and zoonotic potential of ST11 and a new closely related ST258 lineage sourced from diverse origins. We found multiple intra- and interspecies clonal transmission events in all ribotype sublineages. Clones were spread across multiple continents, often without any health care association, indicative of zoonotic/anthroponotic long-range dissemination in the community. ST11 possesses a massive pan-genome and numerous clinically important antimicrobial resistance elements and prophages, which likely contribute to the success of this globally disseminated lineage of One Health importance.

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Heme is crucial for medium-dependent metronidazole resistance in clinical isolates of C. difficile

10.1101/2020.11.18.388959 ◽

2020 ◽

Author(s):

Ilse M. Boekhoud ◽

Igor Sidorov ◽

Sam Nooij ◽

Céline Harmanus ◽

Ingrid M.J.G. Bos-Sanders ◽

...

Keyword(s):

Inhibitory Concentration ◽

Sequence Type ◽

Clinical Isolates ◽

Whole Genome Sequence ◽

Nucleotide Polymorphism ◽

First Line ◽

Single Nucleotide ◽

Clostridioides Difficile ◽

Metronidazole Resistance ◽

First Line Treatment

AbstractUntil recently, metronidazole was the first-line treatment for Clostridioides difficile infection and it is still commonly used. Though resistance has been reported due to the plasmid pCD-METRO, this does not explain all cases. Here, we investigate resistance to metronidazole in a collection of clinical isolates of C. difficile. We find that nearly all isolates demonstrate a heme-dependent increase in the minimal inhibitory concentration for metronidazole, which in some cases leads to isolates being qualified as resistant (MIC > 2 mg/L). Moreover, whole genome sequence analysis reveals a single nucleotide polymorphism in the heme responsive gene hsmA, which defines a metronidazole resistant lineage of PCR ribotype 010 / multilocus sequence type 15 isolates that also includes pCD-METRO containing strains. Together our data demonstrate that heme is crucial for medium-dependent metronidazole resistance in C. difficile.

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Comparative Whole Genome Sequence Analysis and Biological Features of Clostridioides difficile Sequence Type 2‡

Frontiers in Microbiology ◽

10.3389/fmicb.2021.651520 ◽

2021 ◽

Vol 12 ◽

Author(s):

Xingxing Xu ◽

Qiao Bian ◽

Yun Luo ◽

Xiaojun Song ◽

Shan Lin ◽

...

Keyword(s):

Phylogenetic Analysis ◽

Antibiotic Resistance ◽

Sequence Type ◽

Gene Clustering ◽

Whole Genome ◽

Toxin B ◽

Clostridioides Difficile ◽

Significant Difference ◽

Sporulation Capacity

Clostridioides difficile sequence type 2 (ST2) has been increasingly recognized as one of the major genotypes in China, while the genomic characteristics and biological phenotypes of Chinese ST2 strains remain to be determined. We used whole-genome sequencing and phylogenetic analysis to investigate the genomic features of 182 ST2 strains, isolated between 2011 and 2017. PCR ribotyping (RT) was performed, and antibiotic resistance, toxin concentration, and sporulation capacity were measured. The core genome Maximum-likelihood phylogenetic analysis showed that ST2 strains were distinctly segregated into two genetically diverse lineages [L1 (67.0% from Northern America) and L2], while L2 further divided into two sub-lineages, SL2a and SL2b (73.5% from China). The 36 virulence-related genes were widely distributed in ST2 genomes, but in which only 11 antibiotic resistance-associated genes were dispersedly found. Among the 25 SL2b sequenced isolates, RT014 (40.0%, n = 10) and RT020 (28.0%, n = 7) were two main genotypes with no significant difference on antibiotic resistance (χ2 = 0.024–2.667, P > 0.05). A non-synonymous amino acid substitution was found in tcdB (Y1975D) which was specific to SL2b. Although there was no significant difference in sporulation capacity between the two lineages, the average toxin B concentration (5.11 ± 3.20 ng/μL) in SL2b was significantly lower in comparison to those in L1 (10.49 ± 15.82 ng/μL) and SL2a (13.92 ± 2.39 ng/μL) (χ2 = 12.30, P < 0.05). This study described the genomic characteristics of C. difficile ST2, with many virulence loci and few antibiotic resistance elements. The Chinese ST2 strains with the mutation in codon 1975 of the tcdB gene clustering in SL2b circulating in China express low toxin B, which may be associated with mild or moderate C. difficile infection.

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Draft Genome Sequence of a Clostridioides difficile Sequence Type 97 Strain Belonging to Hypervirulent Clade 2

Microbiology Resource Announcements ◽

10.1128/mra.00245-20 ◽

2020 ◽

Vol 9 (14) ◽

Author(s):

Yukino Usui ◽

Yoko Nukui ◽

Ryuji Koike ◽

Shuji Tohda ◽

Ryoichi Saito

Keyword(s):

North America ◽

Genome Sequence ◽

Draft Genome ◽

Sequence Type ◽

Draft Genome Sequence ◽

Genome Sequences ◽

Content Type ◽

Clostridioides Difficile

Clostridioides difficile hypervirulent clade 2 lineages are the major lineages responsible for the outbreaks in North America and Europe. However, the genome sequences of Japanese isolates are scarcely available. Herein, we report the draft genome sequence of C. difficile strain TMD0138 sequence type 97 (ST97), belonging to hypervirulent clade 2, isolated in Japan.

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