Listeria monocytogenes Wall Teichoic Acid Glycosylation Promotes Surface Anchoring of Virulence Factors, Resistance to Antimicrobial Peptides, and Decreased Susceptibility to Antibiotics
Abstract
:1. Introduction
2. Results
2.1. WTA-glycosylation Promotes Efficient Surface Association of Lm Virulence Factors
2.2. WTA-glycosylation Promotes Lm Resistance to AMPs
2.3. WTA-glycosylation Promotes Lm Decreased Susceptibility to Some Antibiotics
3. Discussion
4. Materials and Methods
4.1. Bacterial Strains and Growth Conditions
4.2. Construction of Deletion Mutant Strains
Bacterial Strains and Plasmid | Lab Code | Relevant Characteristics | Source |
---|---|---|---|
L. monocytogenes | |||
EGD-e | DC 4 | Wild-type; Sv 1/2a | [41] |
EGD-e ∆rmlT | DC492 | EGD-e rmlT (lmo1080) deletion mutant | [17] |
EGD-e ∆lmo1079 | DC 858 | EGD-e lmo1079 deletion mutant | This study |
EGD-e ∆lmo1079∆rmlT | DC 899 | EGD-e lmo1079-lmo1080 deletion mutant | This study |
WSLC 1042 WT | DC 825 | Wild-type; Sv 4b | ATCC®23074 |
WSLC 1042 ∆gttA | DC 826 | WSLC 1042 gttA deletion mutant | [23] |
WSLC 1042 ∆gltB | DC 827 | WSLC 1042 gltB deletion mutant | [23] |
WSLC 1042 ∆gttA∆gltB | DC 828 | WSLC 1042 gttAgltB deletion mutant | [23] |
E. coli | |||
DH5α | Competent cells | Life Technologies | |
Plasmid | |||
pMAD | DC 48 | Ampr and Eryr | [39] |
Primers | Sequence (5’ → 3’) * | Restriction Enzymes | |
---|---|---|---|
1 | lmo1079 UP Fw | AGTCGGATCCGGAGCATCTTCTACATTAGGC | BamHI |
2 | lmo1079 UP Rv | AGTCGTCGACCCATTAACTTTCTCCCTCC | SalI |
3 | lmo1079 DW Fw | AGTCGTCGACTAAATGAGGGAAAACGTTAGG | SalI |
4 | lmo1079 DW Rv | AGTCCCATGGCACCGTGAATGAACGCC | NcoI |
5 | lmo1080 DW Fw | CGGGTCGACTAAGAATGGAGAGAAAAGAATGAAAGG | SalI |
6 | lmo1080 DW Rv | CGGCCATGGGGAATGCTTTTTCATTATAGC | NcoI |
Internal Primers | |||
7 | lmo1079 Fw | GCAAATTGGAATGGGAGGCG | |
8 | lmo1079 Rv | GGATGCCTTGTTGCCGAAAC | |
9 | lmo1080 Fw | TATTGCCACACGCTTTACCG | |
10 | lmo1080 Rv | CTTCCACGATTGAACGAACG | |
11 | lmo1492 Fw | GACGGATCCCGCAACTTCGCAAAATGGG | |
12 | lmo1492 Rv | AGCGTCGACGTCGCCATACCATCTGTTTG | |
pMAD Primers | |||
13 | pMAD Fw | TGATGGTCGTCATCTACCTGCC | |
14 | pMAD Rv | CCTACGTAGGATCGATCCGACC |
4.3. Growth Analysis in Vitro
4.4. Extracts of Lm Proteins
4.5. SDS-PAGE and Western Blot Analysis of Protein Extracts
4.6. Antimicrobial Peptides Susceptibility
4.7. Antibiotic Susceptibility
4.8. Statistical Analysis
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
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Antibiotic | Strip Concentration (μg/mL) |
---|---|
Gentamicin Ampicillin Benzylpenicillin | 0.016-256 0.016-256 0.002-32 |
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Meireles, D.; Pombinho, R.; Carvalho, F.; Sousa, S.; Cabanes, D. Listeria monocytogenes Wall Teichoic Acid Glycosylation Promotes Surface Anchoring of Virulence Factors, Resistance to Antimicrobial Peptides, and Decreased Susceptibility to Antibiotics. Pathogens 2020, 9, 290. https://doi.org/10.3390/pathogens9040290
Meireles D, Pombinho R, Carvalho F, Sousa S, Cabanes D. Listeria monocytogenes Wall Teichoic Acid Glycosylation Promotes Surface Anchoring of Virulence Factors, Resistance to Antimicrobial Peptides, and Decreased Susceptibility to Antibiotics. Pathogens. 2020; 9(4):290. https://doi.org/10.3390/pathogens9040290
Chicago/Turabian StyleMeireles, Diana, Rita Pombinho, Filipe Carvalho, Sandra Sousa, and Didier Cabanes. 2020. "Listeria monocytogenes Wall Teichoic Acid Glycosylation Promotes Surface Anchoring of Virulence Factors, Resistance to Antimicrobial Peptides, and Decreased Susceptibility to Antibiotics" Pathogens 9, no. 4: 290. https://doi.org/10.3390/pathogens9040290