{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,30]],"date-time":"2026-07-30T18:09:03Z","timestamp":1785434943601,"version":"3.56.0"},"reference-count":48,"publisher":"American Society for Microbiology","issue":"8","license":[{"start":{"date-parts":[[2025,8,5]],"date-time":"2025-08-05T00:00:00Z","timestamp":1754352000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"},{"start":{"date-parts":[[2025,8,5]],"date-time":"2025-08-05T00:00:00Z","timestamp":1754352000000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/journals.asm.org\/non-commercial-tdm-license"}],"funder":[{"DOI":"10.13039\/501100001807","name":"Funda\u00e7\u00e3o de Amparo \u00e0 Pesquisa do Estado de S\u00e3o Paulo","doi-asserted-by":"publisher","award":["2021\/06894-0, 2021\/10577-0"],"award-info":[{"award-number":["2021\/06894-0, 2021\/10577-0"]}],"id":[{"id":"10.13039\/501100001807","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100008353","name":"Funda\u00e7\u00e3o de Apoio ao Ensino, Pesquisa e Assist\u00eancia do Hospital das Cl\u00ednicas da Faculdade de Medicina de Ribeir\u00e3o Preto da Universidade de S\u00e3o Paulo","doi-asserted-by":"publisher","id":[{"id":"10.13039\/501100008353","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001871","name":"Funda\u00e7\u00e3o para a Ci\u00eancia e a Tecnologia","doi-asserted-by":"publisher","award":["doi.org\/10.54499\/UIDB\/04004\/2020, doi.org\/10.54499\/LA\/P\/0092\/2020"],"award-info":[{"award-number":["doi.org\/10.54499\/UIDB\/04004\/2020, doi.org\/10.54499\/LA\/P\/0092\/2020"]}],"id":[{"id":"10.13039\/501100001871","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["journals.asm.org"],"crossmark-restriction":true},"short-container-title":["Microbiol Spectr"],"published-print":{"date-parts":[[2025,8,5]]},"abstract":"ABSTRACT<\/jats:title>\n \n \n \n Bacterial resistance to \u03b2-lactam antibiotics mediated by \u03b2-lactamase enzymes is widespread worldwide.\n Chromobacterium violaceum<\/jats:italic>\n , an environmental Gram-negative bacterial pathogen, is intrinsically resistant to some \u03b2-lactam antibiotics. In this work, we found that mutations in an\n ampD<\/jats:italic>\n gene, encoding a peptidoglycan-recycling amidase, cause hyperproduction of two chromosomal \u03b2-lactamases (AmpC and CmcB), conferring high \u03b2-lactam resistance in\n C. violaceum<\/jats:italic>\n . Susceptibility tests using \u0394\n ampC<\/jats:italic>\n , \u0394\n cmcB<\/jats:italic>\n , and \u0394\n cmcB\u0394ampC<\/jats:italic>\n mutant strains revealed specific susceptibility profiles to penicillin, cephalosporin, and carbapenem \u03b2-lactams, suggesting that AmpC is a broad-spectrum \u03b2-lactamase (penicillinase and cephalosporinase), while CmcB is a narrow-spectrum metallo-carbapenemase. \u03b2-galactosidase assays indicate that the expression of\n ampC<\/jats:italic>\n and\n cmcB<\/jats:italic>\n increased in response to \u03b2-lactams. We isolated\n C. violaceum<\/jats:italic>\n spontaneous mutants resistant to the antibiotic ceftazidime and found that most mutants were also resistant to several other \u03b2-lactams and overexpressed\n ampC<\/jats:italic>\n and\n cmcB<\/jats:italic>\n . DNA sequencing of the three paralog genes encoding the\n C. violaceum<\/jats:italic>\n AmpD amidases revealed mutations of different types in AmpD1 (CV_0566) in most of the spontaneous mutants, but no mutation was found in AmpD2 or AmpD3. Analysis of single and combined null amidase mutants revealed overexpression of both \u03b2-lactamases and increased resistance to \u03b2-lactams only in mutants with deleted\n ampD1<\/jats:italic>\n . When introduced into\n ampD1<\/jats:italic>\n null or spontaneous mutants, the\n ampD1<\/jats:italic>\n gene rescued the antibiotic-related phenotypes. The AmpD1 amidase from\n C. violaceum<\/jats:italic>\n has a unique architecture with an N-terminal acetyltransferase domain. Our work offers new insights into the mechanisms of \u03b2-lactamase-mediated antibiotic resistance and opens perspectives to improve the treatment of\n C. violaceum<\/jats:italic>\n infections.\n <\/jats:p>\n \n IMPORTANCE<\/jats:title>\n \n Resistance to \u03b2-lactam antibiotics reduces the options for treating bacterial infections, posing a threat to public health. In this work, we demonstrated that the intrinsic resistance to \u03b2-lactam antibiotics in the environmental pathogen\n Chromobacterium violaceum<\/jats:italic>\n is mediated by two chromosomally encoded \u03b2-lactamases, AmpC and CmcB, and revealed the mechanism that contributes to their simultaneous expression. Our data indicate that mutations in the peptidoglycan recycling amidase\n ampD1<\/jats:italic>\n , but not in its paralogs\n ampD2<\/jats:italic>\n and\n ampD3<\/jats:italic>\n , lead to stable overexpression of both \u03b2-lactamases and increased resistance to \u03b2-lactam antibiotics. Remarkably, AmpD1 possesses a unique N-terminal acetyltransferase domain, suggesting a distinct functional mechanism for this enzyme. Our work offers an explanation for the limited effectiveness of many \u03b2-lactams in treating\n C. violaceum<\/jats:italic>\n infections. Understanding the mechanism of antimicrobial resistance is crucial for developing effective treatments and mitigating the spread of \u03b2-lactam-resistant bacteria.\n <\/jats:p>\n <\/jats:sec>\n <\/jats:sec>","DOI":"10.1128\/spectrum.00916-25","type":"journal-article","created":{"date-parts":[[2025,6,12]],"date-time":"2025-06-12T08:27:18Z","timestamp":1749716838000},"update-policy":"https:\/\/doi.org\/10.1128\/asmj-crossmark-policy-page","source":"Crossref","is-referenced-by-count":3,"title":["Mutations in\n ampD<\/i>\n cause hyperproduction of AmpC and CmcB \u03b2-lactamases and high resistance to \u03b2-lactam antibiotics in\n Chromobacterium violaceum<\/i>"],"prefix":"10.1128","volume":"13","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-6696-4177","authenticated-orcid":true,"given":"Lu\u00eds Gustavo","family":"Laranjeiro","sequence":"first","affiliation":[{"id":[{"id":"https:\/\/ror.org\/036rp1748","id-type":"ROR","asserted-by":"publisher"}],"name":"Departamento de Biologia Celular e Molecular e Bioagentes Patog\u00eanicos, Faculdade de Medicina de Ribeir\u00e3o Preto, Universidade de S\u00e3o Paulo","place":["Ribeir\u00e3o Preto, Brazil"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Carlos Eduardo M.","family":"Neme","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/036rp1748","id-type":"ROR","asserted-by":"publisher"}],"name":"Departamento de Biologia Celular e Molecular e Bioagentes Patog\u00eanicos, Faculdade de Medicina de Ribeir\u00e3o Preto, Universidade de S\u00e3o Paulo","place":["Ribeir\u00e3o Preto, Brazil"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Maristela","family":"Previato-Mello","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/036rp1748","id-type":"ROR","asserted-by":"publisher"}],"name":"Departamento de Biologia Celular e Molecular e Bioagentes Patog\u00eanicos, Faculdade de Medicina de Ribeir\u00e3o Preto, Universidade de S\u00e3o 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