Amplicon-Based High-Throughput Sequencing Method Capable of Species-Level Identification of Coagulase-Negative Staphylococci in Diverse Communities
Abstract
:1. Introduction
2. Materials and Methods
2.1. Bacterial Strains
2.2. Genomic DNA Extraction
2.2.1. Pure Cultures
2.2.2. Pooled Cultures
2.3. Primer Design
2.4. PCR Assays and Sequencing
2.5. Amplicon Sequence Data Analysis
2.6. Data Availability
3. Results and Discussion
3.1. Enumeration and Pooling of Pure Cultures
3.2. Sequencing and In Silico Analysis
3.3. Amplicon-Based HTS Method
4. Conclusions
Supplementary Materials
Author Contributions
Funding
Conflicts of Interest
References
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Species | Strain | Origin |
---|---|---|
Staphylococcus auricularis | IMDO-S3 | Teat apex skin |
G162 | Teat apex skin | |
Staphylococcus arlettae | IMDO-S2 | Teat apex skin |
G238 | Teat apex skin | |
Staphylococcus carnosus | IMDO-S14 | Fermented meat |
IMDO-S15 | Meat starter culture | |
Staphylococcus chromogenes | IMDO-S17 | Teat apex skin |
IMDO-S18 | Teat apex skin | |
Staphylococcus cohnii | IMDO-S19 | Milk |
IMDO-S21 | Teat apex skin | |
Staphylococcus epidermidis | IMDO-S29 | Teat apex skin |
IMDO-S30 | Fermented meat | |
Staphylococcus equorum | IMDO-S35 | Fermented meat |
IMDO-S36 | Fermented meat | |
Staphylococcus fleuretti | IMDO-S47 | Milk |
2.05 | Milk | |
Staphylococcus hominis | IMDO-S53 | Unknown |
E326 | Unknown | |
Staphylococcus haemolyticus | IMDO-S50 | Teat apex skin |
IMDO-S51 | Fermented meat | |
Staphylococcus lugdunensis | NA 14.37.1.5 | Fermented meat |
IMDO-S92 | Fermented meat | |
Staphylococcus pasteuri | IMDO-S54 | Fermented meat |
IMDO-S55 | Fermented meat | |
Staphylococcus saprophyticus | IMDO-S58 | Milk |
IMDO-S59 | Fermented meat | |
Staphylococcus sciuri | IMDO-S71 | Teat apex skin |
IMDO-S72 | Fermented meat | |
Staphylococcus simulans | IMDO-S66 | Unknown |
IMDO-S95 | Fermented meat | |
Staphylococcus succinus | IMDO-S67 | Fermented meat |
IMDO-S68 | Fermented meat | |
Staphylococcus warneri | IMDO-S73 | Milk |
IMDO-S74 | Milk | |
Staphylococcus xylosus | IMDO-S76 | Meat starter culture |
IMDO-S81 | Fermented meat |
Primer | Primer Sequence | Amplicon Size (bp) |
---|---|---|
Tuf108 | 5′- WCACGTTGACCAYGGTAAAAC -3′ | 301 |
Tuf408 | 5′-YTCACGMGTTTGWGGCATTGG -3′ | |
Tuf387 | 5′- YCCAATGCCWCAAACKCGTGA -3′ | 379 |
Tuf765 | 5′- RAYTTGHCCACGTTCAACAC -3′ | |
Tuf216 | 5′- WGAAGAAAMAGARCGTGGTA -3′ | 307 |
Tuf522 | 5′- RTCACGWACTTCCATTTCWACT -3′ | |
Tuf32 | 5′- AGAATAGGAGAGATTTAATAATGGC -3′ | 869 |
Tuf900 | 5′- VCCACGTAATAAHGCACC -3′ |
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Van Reckem, E.; De Vuyst, L.; Leroy, F.; Weckx, S. Amplicon-Based High-Throughput Sequencing Method Capable of Species-Level Identification of Coagulase-Negative Staphylococci in Diverse Communities. Microorganisms 2020, 8, 897. https://doi.org/10.3390/microorganisms8060897
Van Reckem E, De Vuyst L, Leroy F, Weckx S. Amplicon-Based High-Throughput Sequencing Method Capable of Species-Level Identification of Coagulase-Negative Staphylococci in Diverse Communities. Microorganisms. 2020; 8(6):897. https://doi.org/10.3390/microorganisms8060897
Chicago/Turabian StyleVan Reckem, Emiel, Luc De Vuyst, Frédéric Leroy, and Stefan Weckx. 2020. "Amplicon-Based High-Throughput Sequencing Method Capable of Species-Level Identification of Coagulase-Negative Staphylococci in Diverse Communities" Microorganisms 8, no. 6: 897. https://doi.org/10.3390/microorganisms8060897
APA StyleVan Reckem, E., De Vuyst, L., Leroy, F., & Weckx, S. (2020). Amplicon-Based High-Throughput Sequencing Method Capable of Species-Level Identification of Coagulase-Negative Staphylococci in Diverse Communities. Microorganisms, 8(6), 897. https://doi.org/10.3390/microorganisms8060897