On the verge of colistin resistance. (c2019)

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dc.contributor.author Alousi, Sahar
dc.date.accessioned 2019-11-27T07:35:51Z
dc.date.available 2019-11-27T07:35:51Z
dc.date.copyright 2019 en_US
dc.date.issued 2019-11-27
dc.date.submitted 2019-05-13
dc.identifier.uri http://hdl.handle.net/10725/11582
dc.description.abstract Colistin has often become the last option to treat infections by carbapenemase producing Klebsiella pneumoniae (CPKP). The PhoPQ two-component system (TCS), insertional inactivation of mgrB gene, and crrAB TCS have recently gained attention as mediators of colistin resistance and lipid A remodeling. Through whole-genome sequencing and lipidomic approaches, genetic alterations and lipid A modifications responsible for colistin resistance can be detected with high precision. Three colistin resistant, seven heteroresistant and one susceptible isolate were studied to explore the underlying mechanisms of colistin resistance. Colistin broth microdilution antimicrobial susceptibility testing was performed, coupled with Etest phenotypic testing. The nucleotide sequence of genes encoding for two-component regulatory systems such as phoP, phoQ, pmrA, pmrB, crrA, and crrB were screened in silico. Plasmid encoded mcr-1 gene was screened through in silico PCR. The mgrB gene was screened through PCR and was manually sequenced through sanger sequencing. Core genome single nucleotide polymorphisms (cg-SNP) were analyzed to determine the phylogenetic evolution among the isolates. Lipid A extraction procedure was optimized for Lipopolysaccharide hydrolysis from both Escherichia coli and K. pneumoniae using acetic acid extraction procedure. For the purpose of lipid A detection and profiling, MALDI-TOF MS was operated in the negative reflectron mode and served as a golden standard for lipid A analysis. None of the plasmid encoded colistin resistance genes were detected. PCR amplification of mgrB revealed insertional inactivation Δ𝑚𝑔𝑟𝐵 in three of the studied isolates (designated as KP5, KP6, and KP16) showing MICs ≥ 64 mg/L. ISKpn14 was associated with KP5 and KP6, while IS903 was detected in KP16. Wild-type mgrB gene in the remaining seven isolates might suggest the involvement of other mechanisms underlying their heteroresistance to colistin. All 11 isolates were negative for the crrA and crrB genes. Lipid A major mass ion was observed at (m/z 1840) in all KP isolates. KP5 had an altered lipid A moiety corresponding to 4-amino-4-deoxy-L-arabinose (L-Ara4N) modification (1892 m/z). This is the first study detecting lipid A modifications in colistin resistant K. penumoniae in Lebanon and the first reporting colistin heteroresistant subpopulations. en_US
dc.language.iso en en_US
dc.subject Lebanese American University -- Dissertations en_US
dc.subject Dissertations, Academic en_US
dc.subject Klebsiella pneumoniae en_US
dc.subject Drug resistance in microorganisms en_US
dc.subject Polymyxin en_US
dc.title On the verge of colistin resistance. (c2019) en_US
dc.type Thesis en_US
dc.title.subtitle Genetic determinants mediating colistin resistance in klebsiella pneumoniae en_US
dc.term.submitted Spring en_US
dc.author.degree MS in Molecular Biology en_US
dc.author.school SAS en_US
dc.author.idnumber 201005632 en_US
dc.author.commembers Wex, Brigitte
dc.author.commembers Khalil, Christian
dc.author.department Natural Sciences en_US
dc.description.embargo 24M en_US
dc.description.physdesc 1 hard copy: xiv, 81 leaves: col. ill., charts; 30 cm. available at RNL. en_US
dc.author.advisor Tokajian, Sima
dc.author.advisor Khnayzer, Rony
dc.keywords Klebsiella pneumoniae en_US
dc.keywords Colistin en_US
dc.keywords Heteroresistance en_US
dc.keywords mgrB en_US
dc.keywords lipid A en_US
dc.keywords lipidomic en_US
dc.keywords MALDI-TOF MS en_US
dc.description.bibliographiccitations Bibliography: (leaves 66-81). en_US
dc.identifier.doi https://doi.org/10.26756/th.2019.148 en_US
dc.author.email sahar.alousi@lau.edu en_US
dc.identifier.tou http://libraries.lau.edu.lb/research/laur/terms-of-use/thesis.php en_US
dc.publisher.institution Lebanese American University en_US
dc.author.affiliation Lebanese American University en_US

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