Clostridioides difficile in South American Camelids in Germany: First Insights into Molecular and Genetic Characteristics and Antimicrobial Resistance
Abstract
:1. Introduction
2. Results
2.1. Prevalence and Molecular Characteristics
2.2. Whole Genome Sequencing (WGS)
2.3. Antimicrobial Susceptibility Testing (AST)
3. Discussion
3.1. Clostridioides difficile and (South American) Camelids in Literature—A Short Review
3.2. Prevalence of Clostridioides difficile in German South American Camelids
3.3. Isolates of Different Toxinogenic Ribotypes Resistant to Antimicrobial Agents Were Detected
3.4. South American Camelids and Antimicrobial Agents
3.5. Further Investigation Is Needed
4. Materials and Methods
4.1. Sampling
4.2. Isolation of C. difficile
4.3. Molecular Characterisation
4.4. Whole Genome Sequencing
4.5. Antimicrobial Susceptibility Testing
5. Conclusions
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Isolate | Farm-ID | Federal State | Ribotype (RT) | WEBRIBO-ID | Sequence Type (ST) | Toxin A | Toxin B | Binary Toxin |
---|---|---|---|---|---|---|---|---|
tcdA | tcdB | cdtA/cdtB | ||||||
19S0105 | 008 | Thuringia | AI-75 | PR28224 | 8 | + | + | −/− |
19S0136 | 016 | Saxony | 078 | PR28225 | 11 | + | + | +/+ |
19S0160 | 018 | Saxony | 015 | PR26350 | 10 | + | + | −/− |
19S0161 | 018 | Saxony | 015 | PR28235 | 10 | + | + | −/− |
19S0260 | 039 | Saxony-Anhalt | 002/2 | PR28226 | 8 | + | + | −/− |
19S0262 | 040 | Saxony-Anhalt | 002/2 | PR28226 | 8 | + | + | −/− |
19S0264 | 041 | Saxony-Anhalt | 002/2 | PR26352 | 8 | + | + | −/− |
19S0266 | 042 | Saxony-Anhalt | 029 | PR28237 | 16 | + | + | −/− |
Isolate | 19S0105 | 19S0136 | 19S0160 | 19S0161 | 19S0260 | 19S0262 | 19S0264 | 19S0266 | CD630 |
---|---|---|---|---|---|---|---|---|---|
19S0105 | 0 | 91994 | 9286 | 9286 | 116 | 116 | 121 | 9306 | 9147 |
19S0136 | 91994 | 0 | 91623 | 91623 | 91999 | 91999 | 91996 | 91714 | 92383 |
19S0160 | 9286 | 91623 | 0 | 6 | 9310 | 9310 | 9307 | 3955 | 10527 |
19S0161 | 9286 | 91623 | 6 | 0 | 9310 | 9310 | 9307 | 3953 | 10531 |
19S0260 | 116 | 91999 | 9310 | 9310 | 0 | 0 | 5 | 9330 | 9173 |
19S0262 | 116 | 91999 | 9310 | 9310 | 0 | 0 | 5 | 9330 | 9173 |
19S0264 | 121 | 91996 | 9307 | 9307 | 5 | 5 | 0 | 9327 | 9168 |
19S0266 | 9306 | 91714 | 3955 | 3953 | 9330 | 9330 | 9327 | 0 | 10484 |
CD630 | 9147 | 92383 | 10527 | 10531 | 9173 | 9173 | 9168 | 10484 | 0 |
Isolate | Resistance Genes |
---|---|
19S0105 | blaCDD-1; vanG; vanR; vanS; vanT; vanZ1 |
19S0136 | aadE; blaCDD-1; tet(40); tet(M); vanZ1 |
19S0160 | blaCDD-1; vanG; vanR; vanS; vanT; vanZ1 |
19S0161 | blaCDD-1; vanG; vanR; vanS; vanT; vanZ1 |
19S0260 | blaCDD-1; vanG; vanR; vanS; vanT; vanZ1 |
19S0262 | blaCDD-1; vanG; vanR; vanS; vanT; vanZ1 |
19S0264 | blaCDD-1; vanG; vanR; vanS; vanT; vanZ1 |
19S0266 | blaCDD-1; vanG; vanR; vanS; vanT; vanZ1 |
Isolate | Protein | Amino Acid Substitutions |
---|---|---|
19S0136 | GyrA | Thr-82-Val * Lys-413-Asn |
GyrB | Gln-160-His Ser-366 #-Val Ser-416-Ala * | |
19S0160 | GyrB | Val-130-Ile |
19S0161 | GyrB | Val-130-Ile |
Isolate (RT) | Antimicrobials | MEZ | VAN | MOX | CIP | AMP | PEN | AMC | MEP | ERY | CLI | CHL | TET | LZD |
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
BP/ECOFF (mg/L) | >2 a | >2 a | ≤2/≥8 b | 32 c | ≤0.5/≥2 b | ≤0.5/≥2 b | ≤4/2/≥16/8 b | ≤4/≥16 b | 4 c | ≤2/≥8 b | ≤8/≥32 b | ≤4/≥16 b | - | |
19S0105 (RT AI-75) | 0.19 | 0.75 | 1.5 | >32 | 1.5 | 1.5 | 0.75 | 1.5 | 2 | 8 | 12 | 0.38 | 3 | |
19S0136 (RT 078) | 0.125 | 0.75 | >32 | >32 | 0.75 | 1.5 | 0.38 | 0.75 | >256 | 2 | 6 | 16 | 3 | |
19S0160 (RT 015) | 0.38 | 0.75 | 1.5 | >32 | 1 | 1 | 1 | 1.5 | 2 | 12 | 6 | 0.38 | 6 | |
19S0161 (RT 015) | 0.125 | 0.75 | 1 | >32 | 1.5 | 1 | 0.75 | 1.5 | 2 | 12 | 6 | 0.38 | 4 | |
19S0260 (RT 002/2) | 0.38 | 1 | 1.5 | >32 | 1.5 | 1.5 | 0.5 | 1.5 | 2 | 8 | 8 | 0.19 | 3 | |
19S0262 (RT 002/2) | 0.38 | 0.75 | 1.5 | >32 | 1.5 | 1.5 | 0.75 | 1.5 | 2 | 8 | 8 | 0.25 | 3 | |
19S0264 (RT 002/2) | 0.38 | 0.75 | 1.5 | >32 | 1 | 1 | 0.5 | 1 | 2 | 12 | 8 | 0.094 | 3 | |
19S0266 (RT 029) | 0.38 | 1 | 1 | >32 | 3 | 4 | 0.75 | 1.5 | 2 | 6 | 8 | 0.5 | 8 |
PCR-MIX and -Programme | Test Components | Working Concentration | cdd3 | tcdA—Non-Repetitive Region | tcdA—Repetitive Region | tcdB | cdtA | cdtB |
---|---|---|---|---|---|---|---|---|
PCR-Mix | DNA | As obtained by DNA isolation | 2 µL | 2 µL | 2 µL | 2 µL | 2 µL | 2 µL |
DreamTaq Buffer (Thermo Fisher Scientific, Waltham, MA USA) | 10 x | 2.5 µL | 2.5 µL | 2.5 µL | 2.5 µL | 2.5 µL | 2.5 µL | |
dNTP-Mix (Carl Roth, Karlsruhe, Germany) | 10 mM | 0.5 µL | 1 µL | 0.5 µL | 1 µL | 0.5 µL | 0.5 µL | |
Primer 1 a | 10 pmol/µL | 0.5 µL | 1 µL | 1 µL | 1 µL | 1 µL | 1 µL | |
Primer 2 a | 10 pmol/µL | 0.5 µL | 1 µL | 1 µL | 1 µL | 1 µL | 1 µL | |
DreamTaq DNA Polymerase (Thermo Fisher Scientific, Waltham, MA USA) | 5 U/µL | 0.1 µL | 0.2 µL | 0.1 µL | 0.2 µL | 0.1 µL | 0.1 µL | |
MgCl2 (Qiagen, Hilden, Germany) | 25 mM | 1 µL | 1 µL | 1 µL | 1 µL | 1 µL | 1 µL | |
H2O | 17.9 µL | 16.3 µL | 16.9 µL | 16.3 µL | 16.9 µL | 16.9 µL | ||
PCR-Programme | Cycles | 35 | 35 | 35 | 35 | 35 | 35 | |
Initial Denaturation | 2 min, 95 °C | 2 min, 95 °C | 2 min, 95 °C | 2 min, 95 °C | 2 min, 95 °C | 2 min, 95 °C | ||
Denaturation | 30 s, 95 °C | 30 s, 95 °C | 30 s, 95 °C | 30 s, 95 °C | 30 s, 95 °C | 30 s, 95 °C | ||
Annealing | 45 s, 50 °C | 30 s, 55 °C | 30 s, 62 °C | 30 s, 55 °C | 45 s, 52 °C | 45 s, 52 °C | ||
Elongation | 1 min, 72 °C | 30 s, 72 °C | 1,5 min 72 °C | 30 s, 72 °C | 40 s, 72 °C | 40 s, 72 °C | ||
Finale Extension | 8 min, 72 °C | 5 min, 72 °C | 8 min, 72 °C | 5 min, 72 °C | 8 min, 72 °C | 8 min, 72 °C |
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Dost, I.; Abdel-Glil, M.; Schmoock, G.; Menge, C.; Berens, C.; González-Santamarina, B.; Wiegand, E.; Neubauer, H.; Schwarz, S.; Seyboldt, C. Clostridioides difficile in South American Camelids in Germany: First Insights into Molecular and Genetic Characteristics and Antimicrobial Resistance. Antibiotics 2023, 12, 86. https://doi.org/10.3390/antibiotics12010086
Dost I, Abdel-Glil M, Schmoock G, Menge C, Berens C, González-Santamarina B, Wiegand E, Neubauer H, Schwarz S, Seyboldt C. Clostridioides difficile in South American Camelids in Germany: First Insights into Molecular and Genetic Characteristics and Antimicrobial Resistance. Antibiotics. 2023; 12(1):86. https://doi.org/10.3390/antibiotics12010086
Chicago/Turabian StyleDost, Ines, Mostafa Abdel-Glil, Gernot Schmoock, Christian Menge, Christian Berens, Belén González-Santamarina, Elisabeth Wiegand, Heinrich Neubauer, Stefan Schwarz, and Christian Seyboldt. 2023. "Clostridioides difficile in South American Camelids in Germany: First Insights into Molecular and Genetic Characteristics and Antimicrobial Resistance" Antibiotics 12, no. 1: 86. https://doi.org/10.3390/antibiotics12010086