{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,5]],"date-time":"2026-10-05T17:42:12Z","timestamp":1791222132385,"version":"4.1.0"},"reference-count":59,"publisher":"American Society for Microbiology","issue":"10","license":[{"start":{"date-parts":[[2010,5,15]],"date-time":"2010-05-15T00:00:00Z","timestamp":1273881600000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/journals.asm.org\/non-commercial-tdm-license"}],"content-domain":{"domain":["journals.asm.org"],"crossmark-restriction":true},"short-container-title":["Appl Environ Microbiol"],"published-print":{"date-parts":[[2010,5,15]]},"abstract":"<jats:title>ABSTRACT<\/jats:title>\n          <jats:p>\n            <jats:italic>Burkholderia cenocepacia<\/jats:italic>\n            is a multidrug-resistant opportunistic pathogen that infects the airways of patients with cystic fibrosis (CF) and can survive intracellularly in macrophages and epithelial cells. The gentamicin protection assay, which relies on the poor ability of gentamicin or other aminoglycosides to permeate eukaryotic cell membranes, is traditionally employed to quantify intracellular bacteria. However, the high resistance of these bacteria to aminoglycosides hampers the use of the gentamicin protection assay to investigate intracellular infection by\n            <jats:italic>B. cenocepacia<\/jats:italic>\n            . Here, we report the construction of gentamicin-sensitive strains of\n            <jats:italic>B. cenocepacia<\/jats:italic>\n            carrying a deletion of the BCAL1674, BCAL1675, and BCAL1676 genes that form an operon encoding an AmrAB-OprA-like efflux pump. We show that bacteria carrying this deletion are hypersensitive to gentamicin and also delay phagolysosomal fusion upon infection of RAW 264.7 murine macrophages, as previously demonstrated for the parental strain. We also demonstrate for the first time that low concentrations of gentamicin can be used to effectively kill extracellular bacteria and reliably quantify the intracellular infection by\n            <jats:italic>B. cenocepacia<\/jats:italic>\n            , which can replicate in RAW 264.7 macrophages.\n          <\/jats:p>","DOI":"10.1128\/aem.03024-09","type":"journal-article","created":{"date-parts":[[2010,3,27]],"date-time":"2010-03-27T02:54:25Z","timestamp":1269658465000},"page":"3170-3176","update-policy":"https:\/\/doi.org\/10.1128\/asmj-crossmark-policy-page","source":"Crossref","is-referenced-by-count":91,"title":["Construction of Aminoglycoside-Sensitive\n            <i>Burkholderia cenocepacia<\/i>\n            Strains for Use in Studies of Intracellular Bacteria with the Gentamicin Protection Assay"],"prefix":"10.1128","volume":"76","author":[{"given":"Mohamad A.","family":"Hamad","sequence":"first","affiliation":[{"name":"Infectious Diseases Research Group, Siebens-Drake Research Institute, Department of Microbiology and Immunology, University of Western Ontario, London, Ontario, Canada N6A 5C1"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Alexander M.","family":"Skeldon","sequence":"additional","affiliation":[{"name":"Infectious Diseases Research Group, Siebens-Drake Research Institute, Department of Microbiology and Immunology, University of Western Ontario, London, Ontario, Canada N6A 5C1"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Miguel A.","family":"Valvano","sequence":"additional","affiliation":[{"name":"Infectious Diseases Research Group, Siebens-Drake Research Institute, Department of Microbiology and Immunology, University of Western Ontario, London, Ontario, Canada N6A 5C1"},{"name":"Department of Medicine, University of Western Ontario, London, Ontario, Canada N6A 5C1"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"235","reference":[{"key":"e_1_3_2_2_2","doi-asserted-by":"publisher","DOI":"10.1164\/ajrccm.161.4.9907147"},{"key":"e_1_3_2_3_2","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.87.14.5509"},{"key":"e_1_3_2_4_2","doi-asserted-by":"publisher","DOI":"10.1046\/j.1462-5822.1999.00024.x"},{"key":"e_1_3_2_5_2","doi-asserted-by":"publisher","DOI":"10.1093\/embo-reports\/kve062"},{"key":"e_1_3_2_6_2","doi-asserted-by":"publisher","DOI":"10.1128\/IAI.01257-08"},{"key":"e_1_3_2_7_2","doi-asserted-by":"publisher","DOI":"10.1128\/iai.64.10.4054-4059.1996"},{"key":"e_1_3_2_8_2","doi-asserted-by":"publisher","DOI":"10.1186\/1471-2180-9-200"},{"key":"e_1_3_2_9_2","doi-asserted-by":"publisher","DOI":"10.1128\/IAI.00263-08"},{"key":"e_1_3_2_10_2","doi-asserted-by":"publisher","DOI":"10.1128\/IAI.00985-09"},{"key":"e_1_3_2_11_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.jcf.2006.05.007"},{"key":"e_1_3_2_12_2","doi-asserted-by":"publisher","DOI":"10.1128\/JB.01258-06"},{"key":"e_1_3_2_13_2","doi-asserted-by":"publisher","DOI":"10.1128\/AEM.02430-07"},{"key":"e_1_3_2_14_2","doi-asserted-by":"publisher","DOI":"10.1016\/S0035-9203(08)70031-2"},{"key":"e_1_3_2_15_2","doi-asserted-by":"publisher","DOI":"10.1046\/j.1462-2920.2003.00471.x"},{"key":"e_1_3_2_16_2","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.69.8.2110"},{"key":"e_1_3_2_17_2","doi-asserted-by":"publisher","DOI":"10.1007\/BF03256660"},{"key":"e_1_3_2_18_2","first-page":"2885","volume":"135","year":"1989","unstructured":"Craig, F. 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