Does antifouling paint select for antibiotic resistance? Carl-Fredrik Flach a,b, ⁎, Chandan Pal a,b , Carl Johan Svensson a,b , Erik Kristiansson b,c , Marcus Östman d , Johan Bengtsson-Palme a,b , Mats Tysklind d , D.G. Joakim Larsson a,b a Department of Infectious Diseases, Institute of Biomedicine, Sahlgrenska Academy, University of Gothenburg, Gothenburg, Sweden b Centre for Antibiotic Resistance Research (CARe), University of Gothenburg, Gothenburg, Sweden c Department of Mathematical Sciences, Chalmers University of Technology, Gothenburg, Sweden d Department of Chemistry, Umeå University, Umeå, Sweden HIGHLIGHTS • Antifouling paints often contain metals that may co-select for antibiotic resis- tance. • Marine microbial biofilms established on painted surfaces were studied. • The heavy-metal based paint co- selected for certain antibiotic resistant bacteria. • The paint did not enrich known mobile antibiotic resistance genes in the com- munities. • The paint selected for RND efflux sys- tems and genes involved in mobiliza- tion of DNA. GRAPHICAL ABSTRACT abstract article info Article history: Received 24 November 2016 Received in revised form 27 January 2017 Accepted 30 January 2017 Available online xxxx Editor: Kevin V. Thomas There is concern that heavy metals and biocides contribute to the development of antibiotic resistance via co- selection. Most antifouling paints contain high amounts of such substances, which risks turning painted ship hulls into highly mobile refuges and breeding grounds for antibiotic-resistant bacteria. The objectives of this study were to start investigate if heavy-metal based antifouling paints can pose a risk for co-selection of antibiotic-resistant bacteria and, if so, identify the underlying genetic basis. Plastic panels with one side painted with copper and zinc-containing antifouling paint were submerged in a Swedish marina and biofilms from both sides of the panels were harvested after 2.5–4 weeks. DNA was isolated from the biofilms and subjected to metagenomic sequencing. Biofilm bacteria were cultured on marine agar supplemented with tetracycline, genta- micin, copper sulfate or zinc sulfate. Biofilm communities from painted surfaces displayed lower taxonomic di- versity and enrichment of Gammaproteobacteria. Bacteria from these communities showed increased resistance to both heavy metals and tetracycline but not to gentamicin. Significantly higher abundance of metal and biocide resistance genes was observed, whereas mobile antibiotic resistance genes were not enriched in these communities. In contrast, we found an enrichment of chromosomal RND efflux system genes, including such with documented ability to confer decreased susceptibility to both antibiotics and biocides/heavy metals. This was paralleled by increased abundances of integron-associated integrase and ISCR transposase genes. The results show that the heavy metal-based antifouling paint exerts a strong selection pressure on marine bacterial communities and can co-select for certain antibiotic-resistant bacteria, likely by favoring species and strains car- rying genes that provide cross-resistance. Although this does not indicate an immediate risk for promotion of Keywords: Marine bacteria Metagenomics Antibiotic resistance Metal resistance RND efflux pump Integron Science of the Total Environment xxx (2017) xxx–xxx ⁎ Corresponding author at: University of Gothenburg, Department of Infectious Diseases, Guldhedsgatan 10A, SE-41346 Gothenburg, Sweden. E-mail address: carl-fredrik.flach@microbio.gu.se (C.-F. Flach). STOTEN-21926; No of Pages 8 http://dx.doi.org/10.1016/j.scitotenv.2017.01.213 0048-9697/© 2017 Elsevier B.V. All rights reserved. Contents lists available at ScienceDirect Science of the Total Environment journal homepage: www.elsevier.com/locate/scitotenv Please cite this article as: Flach, C.-F., et al., Does antifouling paint select for antibiotic resistance? Sci Total Environ (2017), http://dx.doi.org/ 10.1016/j.scitotenv.2017.01.213