Thursday 16 August | PS19 – Horizontal gene transfer and ecology of antibiotic resistance 334A - Functional metagenomics for the investigation of ancient microbial antibiotic resistance in permafrost Cristina Alcaraz*, Rachel Mackelprang California State University Northridge, United States Permafrost, or permanently frozen soil, is found in polar regions and underlies ~20% of the Earth’s surface. Despite subzero temperatures, permafrost hosts a diversity of microbial life. Permafrost serves as a substrate in which to study survival strategies in extreme cryoenvironments. To understand microbial community survival, we focused on a particularly important aspect of community function--antibiotic resistance. Antibiotic resistance is ancient and occurs naturally among soil dwelling microbes. Antibiotic resistance genes are involved in community signaling, environmental sensing and play a role in competition and defense interactions. To identify antibiotic resistance genes in bacterial communities that have never been exposed to modern synthetic antibiotics and to determine their importance as a survival strategy, we employed functional metagenomics. Using this approach, we extracted DNA directly from permafrost frozen for 19,000 to 33,000 years before present and cloned it into a plasmid vector. A metagenomics library was thus constructed and expressed in an E. coli surrogate host. We screened for antibiotic resistance, pooled resistant colonies, and sequenced the antibiotic resistance genes using high-throughput next-generation sequencing. Results obtained from an analogous permafrost substrate demonstrated a robust metagenomic library size of 90.9 GB with 1 out of 15,000 clones expressing antibiotic resistance. Sequence data confirmed the transfer and identification of antibiotic resistance genes from both gram positive and gram negative microorganisms as well as novel functional resistance genes. These results expand our knowledge of functional resistance genes and reveal specific modes of action diverse microbes must use to survive in extreme environments. 335A - Variation in genotype and phenotype of resistance to antibiotics involved in collateral sensitivity interactions in Escherichia coli Richard Allen*, Katia Pfrunder-Cardozo, Alex Hall ETH Zürich, Switzerland As antibiotic resistance spreads there is a lot of interest in using rational treatment strategies to slow the evolution of resistance. One strategy is to use combinations of drugs where mutations causing resistance to one drug sensitise the bacterium to one or more other drugs. Thus far work on this pleiotropic effect, termed collateral sensitivity or negative cross resistance, has focused on finding which pairs of drugs have collateral sensitivity interactions. However, little is known about whether the emergence and expression of known collateral sensitivity varies with environment. To investigate this, we isolated and sequenced independent mutants of a lab strain of Escherichia coli with resistance to the minimal inhibitory concentrations of five antibiotics (gentamicin, streptomycin, chloramphenicol, cefuroxime and trimethoprim) in four different lab environments. We then tested mutants, in the