Characterizing Antibiotic and Heavy Metal Resistance Genes from Bacteria in Commercial Ship Ballast Water Discharged into the Duluth-Superior Harbor

Abstract

Ballast water discharge is a powerful vector for introducing invasive species into aquatic ecosystems and microorganisms numerically dominate the discharge. Invasive bacteria may not only alter the diversity of native bacterial communities but also transfer genetic resistance to antibiotics and heavy metals into these communities. Antibiotic and heavy metal resistance was characterized for bacteria found in ballast water collected from commercial ships actively discharging ballast water into the Duluth-Superior Harbor during 2011 and 2012. Six fosmid libraries containing metagenomic DNA were constructed from ballast water and Duluth-Superior Harbor water. These libraries were screened for antibiotic resistance to benzylpenicillin, cefotaxime, and levofloxacin and heavy metal resistance to cadmium, zinc and mercury to determine resistance by bacteria in each water sample. There were differences between the proportions of microbial fosmids showing resistance to the three antibiotics from different ballast waters than originated from within the Great Lakes. The order of increasing proportion of resistance to benzylpenicillin was: Burns Harbor, IN=Hamilton, Ont.<Duluth, MN=Cleveland, OH=Detroit, MI. A similar pattern of resistance relative to the sources of the ballast water was seen for the other two antibiotics as well. The order of increasing proportion of resistance to cefotaxime was: Burns Harbor, IN<Hamilton, Ont.=Duluth, MN<<Cleveland, OH=Detroit, MI. For the antibiotic levofloxacin, the order from less resistant to most resistant was: Burns Harbor, IN<<Detroit, MI=Duluth, MN=Hamilton, Ont.<Cleveland, OH. These patterns of resistance to the three antibiotics appeared to be related to the population density of the urban areas adjacent to the Great Lakes harbors that were the sources of the ship ballast water. Typically, ballast waters from Great Lakes cities with a population density less than 1,300 people per square mile had a smaller proportion of microbial fosmid clones resistant to benzylpenicillin, cefotaxime, and levofloxacin than microbial fosmids created from ballast water originating from harbors in larger metropolitan areas like Cleveland, OH and Detroit, MI. The percentage of fosmids demonstrating resistance to heavy metals was less obvious between the three heavy metals compared to the three antibiotics. Ballast water received from ports in larger, more urbanized cities may include more bacteria with antibiotic resistance genes and cause greater concern for the spread of antibiotic resistance among native bacterial populations in the Duluth-Superior Harbor than ballast water received from harbors in smaller metropolitan areas.