The contribution of human activities to environmental reservoirs of antibiotic resistance is poorly understood. The purpose of this study was to determine if oxytetracycline (OTC) use in aquaculture facilities increased the detection frequency (i.e., prevalence) of tetracycline resistance (tetR) genes relative to facilities with no recent OTC treatment. We used polymerase chain reaction to screen water and sediment from four noncommercial fish farms in northwestern Wisconsin for the presence of ten tetR determinants: tet(A), tet(B), tet(D), tet(E), tet(G), tet(M), tet(O), tet(Q), tet(S), and tet(W). Water from farms with recent OTC use had significantly higher tetR detection frequencies than did water from farms without recent OTC use, with prevalence in raceways and rearing ponds of farms with recent OTC use exceeding by more than twofold that of farms not using OTC. Effluent from all farms, regardless of treatment regime, had higher tetR detection frequencies than their corresponding influent for all genes, but the specific combinations of tetR genes detected in a sample were not different from their corresponding influent. Although OTC use was associated with the increased occurrence and diversity of tetR genes in water samples, it was not found to relate to tetR gene occurrence in sediment samples. Sediment samples from facilities with no recent OTC use had significantly higher frequencies of tetR gene detection than did samples from facilities with recent OTC use. All of the tetR genes were detected in both the medicated and nonmedicated feed samples analyzed in this study. These findings suggest that both OTC treatment in aquaculture facilities and the farms themselves may be sources of tetR gene introduction to the environment. To our knowledge, this is the first study to use genotypic and cultivation-independent methods to examine tetR gene occurrence associated with OTC use in aquaculture.
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We thank Roderick Mackie and Satoshi Koike for kindly providing positive controls for tetR gene amplification. We also thank Trevor Ghylin, Ashley Shade, and Stuart Jones for helping collect the samples and for the discussions and Azivy Che Aziz, Aaron Jones, and Emma Ingebretsen for the help with processing the samples. This research was funded by a grant from Wisconsin SeaGrant (R/AQ-40) to K.D.M and J.A.P.
Electronic Supplementary Materials
Below is the link to the electronic supplementary material.
Summary of tetR genes detected in water samples throughout each facility. A “+” indicates that the gene was detected more than once in replicate PCR reactions across N samples, “+/−” indicates that the gene was detected only once across replicate PCR reactions in N samples, and “−” indicates that the gene was never detected in the sample (DOC 42 kb)
Summary of tetR genes detected in sediment samples throughout each facility. A “+” indicates that the gene was detected more than once in replicate PCR reactions across N samples, “+/−” indicates that the gene was detected only once across replicate PCR reactions in N samples, and “−” indicates that the gene was never detected in the sample (DOC 56 kb)
Summary of tetR gene detected in feed samples collected from each facility. “Before, During, and After” dates are in reference to OTC treatment at Facility 4. A “+” indicates the gene was detected more than once in the sample, “+/−” indicates that the gene was detected only once in the sample, and “−” indicates that the gene was never detected in the sample. The feed column indicates the difference in feed sizes (“1” being the smallest size). tet(O) was not run on individual feed samples, rather it was run on pooled samples and was detected in each case (DOC 57 kb)
Summary of influent water quality data (DOC 51 kb)
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Seyfried, E.E., Newton, R.J., Rubert, K.F. et al. Occurrence of Tetracycline Resistance Genes in Aquaculture Facilities with Varying Use of Oxytetracycline. Microb Ecol 59, 799–807 (2010). https://doi.org/10.1007/s00248-009-9624-7
- Sediment Sample
- Antibiotic Resistance Gene
- Detection Frequency
- Feed Sample
- Influent Water