TABLE 1.
Genome features, MICs by agar dilution, AMR determinants, and predicted intact prophages of the isolatesa
| Strain | Ribotype | MLST11 | Clade | MIC (μg/mL) determined by agar dilution (sensitivity criteria, source) |
AMR determinants (CARD database) |
Predicted intact prophages (PHAST) | |||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| VAN (S ≤ 2, R > 2; EUCAST ECOFFs) | MTZ (S ≤ 8, I = 16, R ≥ 32; CLSI) | FDX (S < 1, I > 1; literature) | RFX (S ≤ 0.0.004, I = 0.004 to 16, R ≥ 16; literature) | ARO term | AMR gene family | Resistance mechanism | % identity of matching region | % length of reference sequence | |||||
| TGH35 | RT027 | ST1 | 2 | 4 | 0.5 | ≤0.5 | ≤0.125 | APH(3′)-Ia | APH(3′) | Antibiotic inactivation | 100.0 | 30.63 | PHAGE_ Clostr_ phiMMP01_ NC_028883 |
| cdeA | Multidrug and toxic compound extrusion (MATE) transporter | Antibiotic efflux | 99.09 | 100.00 | |||||||||
| APH(3′)-Ia | APH(3′) | Antibiotic inactivation | 96.77 | 67.90 | |||||||||
| vanRG | Glycopeptide resistance gene cluster, vanR | Antibiotic target alteration | 77.45 | 99.15 | |||||||||
| tet(W/N/W) | Tetracycline-resistant ribosomal protection protein | Antibiotic target protection | 69.28 | 100.00 | |||||||||
| vanXYG | Glycopeptide resistance gene cluster, vanXY | Antibiotic target alteration | 58.82 | 105.51 | |||||||||
| TGH64 | RT027 | ST1 | 2 | 4 | 0.25 | ≤0.5 | ≥128 | dfrF | Trimethoprim resistant dihydrofolate reductase dfr | Antibiotic target replacement | 100.0 | 100.00 | PHAGE_ Clostr_ phiMMP01_ NC_028883 |
| cdeA | MATE transporter | Antibiotic efflux | 99.32 | 100.00 | |||||||||
| vanRG | Glycopeptide resistance gene cluster, vanR | Antibiotic target alteration | 77.45 | 99.15 | |||||||||
| vanXYG | Glycopeptide resistance gene cluster, vanXY | Antibiotic target alteration | 58.82 | 105.51 | |||||||||
| tet(W/N/W) | Tetracycline-resistant ribosomal protection protein | Antibiotic target protection | 69.28 | 100.00 | |||||||||
| ermB | Erm 23S ribosomal RNA methyltransferase | Antibiotic target alteration | 98.78 | 98.79 | |||||||||
| TGH120 | RT106 | ST42 | 1 | 2 | 0.5 | ≤0.5 | ≤0.125 | APH(3′)-Ia | APH(3′) | Antibiotic inactivation | 100.0 | 29.89 | - |
| cdeA | MATE transporter | Antibiotic efflux | 99.32 | 100.00 | |||||||||
| vanRG | Glycopeptide resistance gene cluster, vanR | Antibiotic target alteration | 77.87 | 99.15 | |||||||||
| vanXYG | Glycopeptide resistance gene cluster, vanXY | Antibiotic target alteration | 58.82 | 105.51 | |||||||||
| TGH33 | RT017 | ST37 | 4 | 2 | 0.5 | ≤0.5 | ≤0.125 | cdeA | MATE transporter | Antibiotic efflux | 98.41 | 100.00 | PHAGE_ Clostr_ phiCD506_ NC_028838 PHAGE_ Clostr_ phiMMP01_ NC_028883 |
| APH(3′)-Ia | APH(3′) | Antibiotic inactivation | 99.1 | 40.96 | |||||||||
| APH(3′)-Ia | APH(3′) | Antibiotic inactivation | 100.0 | 28.7 | |||||||||
| TGH51 | RT017 | ST37 | 4 | 1 | 0.5 | ≤0.5 | ≤0.125 | cdeA | MATE transporter | Antibiotic efflux | 98.41 | 100.00 | PHAGE_ Clostr_ phiCD506_ NC_028838 PHAGE_ Clostr_ phiCDHM19_ NC_028996 PHAGE_ Clostr_ phiCDHM19_ NC_028996 PHAGE_ Clostr_ phiMMP01_ NC_028883 |
| APH(3′)-Ia | APH(3′) | Antibiotic inactivation | 97.79 | 99.26 | |||||||||
The MICs of vancomycin via the agar dilution method for both control strains RT027/R20291 and RT012/CD630 were 2 μg/mL. The MICs of vancomycin were further determined via BHI broth microdilution: TGH35 (2 μg/mL), TGH64 (4 μg/mL), TGH120 (1 μg/mL), TGH33 (1 μg/mL), TGH51 (1 μg/mL), and R20291 (1 μg/mL). The MICs of vancomycin were also tested via Brucella blood agar dilution and E-tests at LUMC: TGH35 (1 μg/mL), TGH64 (2 μg/mL), TGH120 (0.25 μg/mL), TGH33 (0.5 μg/mL), R20291 (0.5 μg/mL). An EMBL Clustal Omega-based multiple sequence alignment was performed to explore the presence of the mutations that were previously reported to be associated with a reduced susceptibility to vancomycin in in vitro C. difficile strains. Pro108Leu in MurG was detected in all of the isolates in the present study, and G733T in rpoC was detected in none of them. MIC, minimum inhibitory concentration; AMR, antimicrobial resistance; ECOFFs, epidemiological cutoff values; VAN, vancomycin; MTZ, metronidazole; FDX, fidaxomicin; RFX, rifaximin; MLST, multi locus sequence typing; CARD, comprehensive antibiotic resistance database; ARO, Antibiotic resistance ontology. Resistance Gene Identifier criteria used by the CARD database: Strict. Detection criteria: Protein homolog model.