## 4 Discussion
While the detailed taxonomic knowledge required for microscopy is not needed for HTS, the methodology for such molecular sequencing currently appears too lengthy to fulfil this time requirement. As such HTS is probably best suited to a broader risk assessment role, with prospects for further refinement. The following are available online at [[http://www.mdpi.com/2076-3417/9/12/2441/s1](http://www.mdpi.com/2076-3417/9/12/2441/s1)]([http://www.mdpi.com/2076-3417/9/12/2441/s1](http://www.mdpi.com/2076-3417/9/12/2441/s1)), Figure S1: Alpha Diversity at Phylum Level. [Sample Number (1-55)]. Figure S2: Alpha Diversity Categorized by Water Temperature at Phylum Level. Figure S3: Alpha Diversity categorized by Lake at Phylum Level. Figure S4: Alpha Diversity at Class Level. Figure S5: Alpha Diversity Categorized by Water Temperature at Class Level. Figure S6: Diversity Categorized by Lake at Class Level. Table S1: Microscopic Phytoplankton Counts in Ballast Water from Indiana Harbor, Indiana. Table S2: Microscopic Phytoplankton Counts in Ballast Water from Burns Harbor, Indiana. Table S3: Microscopic Phytoplankton Counts in Ballast Water from Monroe, Michigan. Table S4: Microscopic Phytoplankton Counts in Ballast Water from St. Clair, Michigan. Table S5: Microscopic Phytoplankton Counts in Ballast Water from Detroit, Michigan. Table S6: Microscopic Phytoplankton Counts from Ashtabula, Ohio. Table S7: Microscopic Phytoplankton Counts in Ballast Water from Essexville Michigan, Two Harbors Minnesota and Superior Wisconsin. The specific roles of the authors of this paper were as follows: Conceptualization, D.A.W., C.L.M. and A.P. ; D.A.W. secured funding and logistical support from ASC; D.A.W. and [PERSON] were responsible for Project Administration; Liaison with ASC and Sample Processing were carried out by C.L.M. ; Light microscopy was performed by [PERSON]. (ERS); [PERSON] and [PERSON] were responsible for nucleic acid sequencing and statistical treatment and presentation of molecular data; [PERSON] prepared the first draft of the manuscript, which was then reviewed and edited by [PERSON], [PERSON] and [PERSON]. **Funding:** This research was funded by a private grant from the American Steamship Company (ASC) to Environmental Research Services (ERS) who sub-contracted the University of Maryland Center for Environmental Science Chesapeake Biological Laboratory (CBL) and the Institute for Environmental Technology (IET) to perform molecular analyses. ASC played no part in sample processing and analysis or in the presentation or interpretation of results. **Acknowledgments:** The authors are grateful for the extraordinary efforts to develop innovative approaches to ballast water treatment and testing by the American Steamship Company (ASC) as shown by their support of this work and previous studies. This project could not have been carried out without the initiative and support of [PERSON] and [PERSON] of ASC and the efforts of [PERSON] and crews of the M/Vs _American Spirit_, _American Integrity and Burns Harbor_. Thanks are also due to [PERSON] of Proper Design, Cardiff U.K. for data formatting, and [PERSON], NetsCo., for logistical support. This is contribution No. UMCS 5631 from the University of Maryland Center for Environmental Science and contribution No. MET 19-010 from the University of Maryland Institute for Marine Environmental Technology. **Conflicts of Interest:** The authors declare no conflicts of interest. ASC had no role in the design of this study or data interpretation. ASC crews were responsible for sample collection as part of their normal ship operations and according to strict procedures laid down by [PERSON] (University of Maryland, CES, who supplied the sampling equipment and supervised sample shipment and preparation). Publication and dissemination of results was entirely the decision of the authors.