2026-10-14T08:17:59Z https://api.figshare.com/v2/oai

oai:figshare.com:article/34000910 2026-09-25T18:20:09Z category_7 category_8 category_13 category_21 category_24 category_272 category_39 category_734 category_64 category_132 category_134 portal_5 item_type_3 month_year_09_2026

Strains, plasmids, and oligonucleotides used. Jocelyn A. Hammond (25111760) Rachel L. Ehrlich (8293938) Ariel Gonzalez (22389474) Begoña Euba (563215) Kevin M. Raible (25111763) Jill M. Lawrence (25111766) Ari B. Gordon (25111769) Evangeline Williams (11024931) Mary C. Marino (25111772) Jason Limbo (25111775) Steven Lang (7532681) Bhaswati Sen (4599163) Sergey V. Balashov (25111778) Azad Ahmed (121246) Junkal Garmendia (217415) Joris Beld (136102) Garth D. Ehrlich (60143) Michael E. Pichichero (7880588) Ravinder Kaur (1871197) Joshua Chang Mell (7490840) Medicine Microbiology Genetics Biotechnology Evolutionary Biology Environmental Sciences not elsewhere classified Ecology Biological Sciences not elsewhere classified Cancer Infectious Diseases Virology sought new insights predicted functions consistent informed association tests candidate “ anti infected middle ears conferred alkaline sensitivity commonly afflicts children middle ear washes om ” operon div >< p clustered homologous genes bacterial disease burden haemophilus influenzae </ om disease pathogenesis middle ear prone children alkaline tolerance influenzae </ vitro </ spef </ prevalent om om pathogens om disease om arises om ), 12 om significant difference selective advantage polyamine metabolism ph homeostasis persistent colonization otitis media nitrogen metabolism laboratory strain genome sequenced gene absences fold increase distinct lineages clonal lineages associated bacterium across assemblies 9 healthy <div>Infection of the middle ear, or otitis media (OM), is a disease that commonly afflicts children, and Haemophilus influenzae is the most prevalent OM-associated bacterium. We sought new insights into mechanisms of pathogenesis by identifying bacterial genes associated with OM disease. We whole-genome sequenced >200 clinical isolates collected over time from 9 healthy and 12 OM-prone children. Genome analysis first annotated and clustered homologous genes from across assemblies, inferred the phylogeny, and identified closely related set of strains, or clonal lineages, finding both transient and persistent colonization by distinct lineages. Phylogeny-informed association tests then identified 7 strong associations of genes/operons with health or disease. Remarkably, 5 associations were with gene absences and OM, and all 5 had predicted functions consistent with roles in pH homeostasis and nitrogen metabolism, specifically in transport and modification of polyamines. Using a laboratory strain, knockouts of candidate “anti-OM” operon potE-speF were tested. In vitro growth assays varying pH and supplementation with polyamines showed that the presence of potE-speF conferred alkaline sensitivity to cells in the presence of ornithine. Measurements of human middle ear fluid pH found higher alkalinity in specimens where H. influenzae was collected over those with other OM pathogens, supporting the hypothesis that the alkaline middle ear microenvironment in OM arises from or selects for H. influenzae infection. Experiments using a mouse OM model found that absence of potE-speF caused a ~ 63-fold increase in bacterial disease burden in middle ear washes. By contrast, a mouse lung infection model showed no significant difference. We conclude that H. influenzae lacking potE-speF have a selective advantage in OM disease pathogenesis, because these strains thrive in the otherwise hostile alkaline environment that occurs in the infected middle ears of children.</div> 2026-09-25T17:56:18Z Dataset Dataset 10.1371/journal.ppat.1014649.s005 https://figshare.com/articles/dataset/_p_Strains_plasmids_and_oligonucleotides_used_p_/34000910 CC BY 4.0