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Providing high school students access to research experiences improves learning, can have positive and long-lasting impacts on their attitudes toward science, and gives them the opportunity to make impactful scientific contributions. Through EvolvingSTEM, students evolve populations of\n                      <jats:italic toggle=\"yes\">Pseudomonas fluorescens<\/jats:italic>\n                      in a bead model that includes daily cycles of bacterial dispersal, attachment, and biofilm growth, and observe heritable changes in colony morphology. We sequenced the genomes of 69 evolved clones with unique colony morphologies and identified parallel mutations in genes known to regulate or affect biofilm growth (\n                      <jats:italic toggle=\"yes\">wsp<\/jats:italic>\n                      ,\n                      <jats:italic toggle=\"yes\">yfiBNR<\/jats:italic>\n                      ,\n                      <jats:italic toggle=\"yes\">morA,<\/jats:italic>\n                      and\n                      <jats:italic toggle=\"yes\">fuzY<\/jats:italic>\n                      ). We also uncovered novel adaptations: loss-of-function mutations in phosphodiesterase PFLU0185 that did not alter colony morphology and mutations affecting periplasmic disulfide bond formation producing small colonies. PFLU0185 mutants rapidly and consistently reached high frequencies, and phenotyping revealed roles in cyclic di-GMP regulation, biofilm formation, and motility, prompting us to name this gene\n                      <jats:italic toggle=\"yes\">bmo<\/jats:italic>\n                      (\n                      <jats:underline>b<\/jats:underline>\n                      iofilm and\n                      <jats:underline>mo<\/jats:underline>\n                      tility regulator). Competition experiments and microscopy demonstrated that\n                      <jats:italic toggle=\"yes\">bmo<\/jats:italic>\n                      mutants are ecological generalists that compete with their ancestor and specialist mutants. Consequently, phenotypic diversity is maintained, with smooth (ancestral and\n                      <jats:italic toggle=\"yes\">bmo<\/jats:italic>\n                      ) colonies consistently outnumbering wrinkly and fuzzy variants. This study advances our understanding of biofilm genetic architecture while demonstrating that student-led research can uncover mechanisms of microbial adaptation relevant to\n                      <jats:italic toggle=\"yes\">Pseudomonas<\/jats:italic>\n                      infection biology.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>IMPORTANCE<\/jats:title>\n                    <jats:p>\n                      Bacterial biofilms dominate microbial life; however, their evolutionary genetics remain incompletely understood. Extensive replication of biofilm selection experiments by secondary school students can provide valuable insights into mechanisms of adaptation. Notably, mutations in a conserved phosphodiesterase, PFLU0185\/\n                      <jats:italic toggle=\"yes\">bmo<\/jats:italic>\n                      , dominate evolved populations without changing colony morphology. Phenotypic assays reveal that\n                      <jats:italic toggle=\"yes\">bmo<\/jats:italic>\n                      plays a generalist role and competes with its ancestor and other diverse, less frequent biofilm-specialist mutants, resulting in populations that have differentiated to fill multiple niches within the biofilm life cycle. This work also demonstrates the power of distributed research networks for addressing complex biological questions while promoting scientific literacy to the public.\n                    <\/jats:p>\n                  <\/jats:sec>","DOI":"10.1128\/aem.02499-25","type":"journal-article","created":{"date-parts":[[2026,3,30]],"date-time":"2026-03-30T13:00:21Z","timestamp":1774875621000},"update-policy":"https:\/\/doi.org\/10.1128\/asmj-crossmark-policy-page","source":"Crossref","is-referenced-by-count":0,"title":["Genetic diversification of\n                    <i>Pseudomonas fluorescens<\/i>\n                    maintained by multi-niche selection within biofilms"],"prefix":"10.1128","volume":"92","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-4090-9927","authenticated-orcid":false,"given":"Abigail M.","family":"Matela","sequence":"first","affiliation":[{"name":"University of Pittsburgh School of Medicine","place":["Pittsburgh, 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