Antibiotic-Resistant Bacteria across a Wastewater Treatment Plant
Abstract
:1. Introduction
2. Material and Methods
2.1. Sampling and Processing
2.2. Antibiotic Susceptibility Assays
2.3. Phylogenetic Determination with 16S rRNA Sequencing
2.4. Detection of Antibiotic Resistance Genes and Class 1 Integrons
3. Results and Discussion
3.1. Isolation Results and Identification
3.2. Antibiotic Susceptibility Testing
3.3. Presence of Mobile Genetic Elements
4. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Target | Primer Sequence | Amplicon Expected Size (bp) | Annealing Temperature | Program | Reference |
---|---|---|---|---|---|
16S rRNA | 27F—AGA GTT TGA TCM TGG CTC AG | ~1400 | 56 °C | 95 °C—5 min (1×) 95 °C—1 min, 56 °C—1 min, 72 °C—1.5 min, (30×) 72 °C—10 min (1×) | [22] |
1492R—TAC GGY TAC CTT GTT ACG ACT T | |||||
Class 1 integron | Int 5′CS—GGC ATC CAA GCA GCA AG | Variable | 55 °C | 94 °C—5 min (1×) 94 °C—30 s, 55 °C—30 s, 72 °C—6 min, (35×) 72 °C—16 min (1×) | [28] |
Int 3′CS—AAG CAG ACT TGA CCT GA | |||||
Tet(M) | tetMF—GTG GAC AAA GGT ACA ACG AG | 406 | 55 °C | 95 °C—3 min (1×) 95 °C—30 s, 55 °C—30 s, 72 °C—30 s, (35×) 72 °C—10 min (1×) | [29] |
tetMR—CGG TAA AGT TCG TCA CAC AC | |||||
blaSHV | SHV-F—GGG TTA TTC TTA TTT GTC GC | 930 | 56 °C | 94 °C—10 min (1×) 94 °C—30 s, 56 °C—30 s, 72 °C—30 s, (35×) 72 °C—10 min (1×) | [30,31] |
SHV-R—TTA GCG TTG CCA GTG CTC | |||||
blaTEM | TEM-F—ATG AGT ATT CAA CAT TTC CG | 847 | 58 °C | 94 °C—3 min (1×) 94 °C—30 s, 58 °C—30 s, 72 °C—30 s, (35×) 72 °C—10 min (1×) | [32] |
TEM-R—CTG ACA GTT ACC AAT GCT TA | |||||
blaCTX-M | CTX-M-F′—TTT GCG ATG TGC AGT ACC AGT AA | 590 | 51 °C | 94 °C—10 min (1×) 94 °C—30 s, 51 °C—30 s, 72 °C—30 s, (35×) 72 °C—10 min (1×) | [33] |
CTX-M-R′—CGA TAT CGT TGG TGG TGC CAT A |
Phenotype (n.) | |||
---|---|---|---|
Antibiotic | Resistant | Intermediate | Susceptible |
Ampicillin | 9 | 2 | 25 |
Cefotaxime | 0 | 3 | 11 |
Imipenem | 3 | 0 | 19 |
Tetracycline | 13 | 0 | 27 |
Vancomycin | 0 | 6 | 19 |
Gentamicin | 2 | 9 | 6 |
Ciprofloxacin | 5 | 15 | 27 |
Isolate ID | Resistance Phenotype | Intermediate Phenotype | Affiliation | Antibiotic Resistance Genes | Integron Content |
---|---|---|---|---|---|
E73 | AMP; TE; CIP | CTX; CN | Citrobacter sp. | blaTEM * | ANT(3″ )-IIa * |
E79 | AMP | CN; CIP | Kluyvera sp. | blaCTX-M * | |
E82 | TE | VA | Enterococcus sp. | tet(M) * | ANT(3″ )-IIa * |
E84 | TE | Enterococcus sp. | tet(M) | ||
E112 | Brevudimonas sp. | tet(M) * | |||
E113 | AMP; CN; CIP | Shigella sp. | blaTEM * | ||
E126 | Stenotrophomonas sp. | tet(M) * | |||
E127 | IMP | CIP | Pseudomonas sp. | ND | dfrA25 * |
E142 | AMP; TE; CIP | CN | Klebsiella sp. | blaSHV * | |
E145 | CN | Pseudomonas sp. | ND | aadA6 * | |
E148 | AMP; TE | CIP | Raoultella sp. | blaSHV * | dfrA25 * |
E154 | TE | Enterococcus sp. | tet(M) * | ||
E159 | TE; CIP | Enterococcus sp. | blaSHV | dfrA25 * | |
E160 | TE | CIP | Enterococcus sp. | tet(M) * | |
E161 | TE | Enterococcus sp. | tet(M) | ||
E164 | TE | CIP | Enterococcus sp. | tet(M) * | |
E170 | TE | Enterococcus sp. | tet(M) * | ||
E171 | TE | VA | Enterococcus sp. | tet(M) | |
E179 | TE | VA | Enterococcus sp. | tet(M) * | |
E183 | Enterococcus sp. | tet(M) * | |||
E188 | Enterococcus sp. | tet(M) | |||
E195 | Enterococcus sp. | tet(M) * | |||
E208 | TE | Enterococcus sp. | tet(M) * | ||
E209 | Enterococcus sp. | blaSHV * |
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Godinho, O.; Lage, O.M.; Quinteira, S. Antibiotic-Resistant Bacteria across a Wastewater Treatment Plant. Appl. Microbiol. 2024, 4, 364-375. https://doi.org/10.3390/applmicrobiol4010025
Godinho O, Lage OM, Quinteira S. Antibiotic-Resistant Bacteria across a Wastewater Treatment Plant. Applied Microbiology. 2024; 4(1):364-375. https://doi.org/10.3390/applmicrobiol4010025
Chicago/Turabian StyleGodinho, Ofélia, Olga Maria Lage, and Sandra Quinteira. 2024. "Antibiotic-Resistant Bacteria across a Wastewater Treatment Plant" Applied Microbiology 4, no. 1: 364-375. https://doi.org/10.3390/applmicrobiol4010025