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Master's Dissertation
DOI
https://doi.org/10.11606/D.5.2021.tde-23092021-125854
Document
Author
Full name
Thays Almeida Franco de Barcellos
E-mail
Institute/School/College
Knowledge Area
Date of Defense
Published
São Paulo, 2021
Supervisor
Committee
Camargo, Carlos Henrique (President)
Girardello, Raquel
Perdigão Neto, Lauro Vieira
Silva, Rodrigo Cayô da
Title in Portuguese
Análise genômica e caracterização da suscetibilidade antimicrobiana em clones emergentes de Acinetobacter baumannii resistentes aos carbapenêmicos provenientes de diferentes instituições brasileiras
Keywords in Portuguese
Acinetobacter baumannii
Antimicrobianos
Complexo clonal
Epidemiologia molecular
Infecção hospitalar
Oxacilinases
Resistência
Abstract in Portuguese
Altamente prevalente nos hospitais, Acinetobacter baumannii tem gerado grande apreensão devido sua capacidade de causar Infecções Relacionadas à Assistência à Saúde em todo o mundo, impactando nas elevadas taxas de morbidade e mortalidade e à sua capacidade de causar surtos e disseminar clones resistentes. As altas taxas de resistência frente aos carbapenêmicos, assim como os aminoglicosídeos, tetraciclinas e quinolonas, juntamente com a falta de novas opções terapêuticas, tem levado à necessidade de recorrer novamente a fármacos antigos, como a polimixina B. O contexto epidemiológico molecular de A. baumannii é complexo, as cepas pertencentes aos Complexos Clonais CC1, CC15 e CC79, denominadas clones de alto risco, são as mais relatadas no Brasil, porém a ocorrência de outros clones alerta para a necessidade de constante monitoramento, como é o caso do CC25, que recorrentemente é encontrado em nosso país. O objetivo deste estudo foi realizar a análise genômica e caracterizar a susceptibilidade aos antimicrobianos em clones emergentes de 13 isolados de A. baumannii provenientes de diferentes hospitais brasileiros entre os anos de 2016 e 2017, porém com o intuito de selecionar isolados representantes de pulsotipos diferentes e aumentar o número de amostras a serem estudadas, foram acrescentados três isolados de 2013 e dois de 2018, resultando em 18 isolados neste estudo. Os isolados foram submetidos às técnicas de PCR multiplex para detecção das oxacilinases, teste de suscetibilidade aos antimicrobianos, tipagem molecular por eletroforese em campo pulsado (PFGE), tipagem por 3 loci modificado (m3LST), sequenciamento de genoma completo e perfil de análise populacional (PAP) para polimixina B. Detectou-se principalmente os genes blaOXA-64 em 13 isolados (72,2%), blaOXA-23 em 16 isolados (88,9%), blaOXA-72 em dois (11,1%) e blaOXA-253 em apenas um (5,6%). Houve dois isolados (11,1%) que foram carreadores de blaOXA-23 e blaOXA-72 simultaneamente. Todos os isolados, (n=17; 94,5%) foram classificados como XDR, com excessão de um (n=1; 5,5%) que foi considerado MDR. A minociclina foi o único antibiótico que se manteve 100% eficaz contra todas as cepas. Através da técnica de PFGE e m3LST conseguimos, presuntivamente, agrupar os isolados de acordo com suas diversidades genéticas e similaridades clonais. Através do sequenciamento de genoma completo, obtivemos quinze isolados identificados como pertencentes ao CC25 e três isolados pertencentes aos CC49, CC33 e CC162. Oito cepas (44.4%) apresentaram resistência frente à polimixina B, sendo seis delas pertencentes ao CC25. Estes resultados fornecem subsídios que ressaltam a necessidade de monitoramento e controle de clones emergentes circulando em nosso meio
Title in English
Genomic analysis and characterization of antimicrobial susceptibility in emerging clones of carbapenem-resistant Acinetobacter baumannii from different Brazilian institutions
Keywords in English
Acinetobacter baumannii
Antimicrobials
Clonal complex
Molecular epidemiology
Nosocomial infection
Oxacillinases
Resistance
Abstract in English
Highly prevalent in hospitals, Acinetobacter baumannii has raised great concern due to its ability to cause Healthcare-associated Infections worldwide, to cause epidemiological outbreaks and to spread resistant clones, impacting morbidity and mortality rates. The high rates of carbapenems resistance, as well as aminoglycosides, tetracyclines and quinolones, coupled with the lack of new therapeutic approaches, has led the need to revisit old drugs, such as polymyxin B. The epidemiological-molecular context of A. baumannii is complex. The strains belonging to the Clonal Complexes CC1, CC15 and CC79, known as high-risk clones, are the most frequent in Brazil, however the occurrence of other clones highlight the importance for constant monitoring, as is the case of CC25, which is recurrently reported in our country. The aim of this study was to perform a genomic analysis and characterize antimicrobial susceptibility in emerging clones from 13 A. baumannii isolates from different Brazilian hospitals in the years 2016 and 2017. In order to select isolates representing different pulsotypes and increase the number of samples, three isolates from 2013 and two from 2018 were also investigated, resulting in 18 isolates in this study. The isolates were submitted to multiplex PCR techniques for the detection of oxacillinases, antimicrobial susceptibility testing, molecular typing by pulsed field gel electrophoresis (PFGE), modified PCR based trilocus sequence-based typing (m3LST), complete genome sequencing and Population Analysis Profile (PAP) for polymyxin B. The blaOXA-64 genes were mainly detected in 13 isolates (72.2%), blaOXA-23 in 16 isolates (88.9%), blaOXA-72 in two (11.1%) and blaOXA-253 in only one (5.6%). There were two isolates (11.1%) that were carriers of blaOXA-23 and blaOXA-72 simultaneously. All isolates (n=17; 94.5%) were classified as XDR (extensively resistant drug), except for one (n=1; 5.5%) that was considered MDR (multi-drug resistant). Minocycline was the only antibiotic that remained 100% effective against all strains. By PFGE and m3LST, we presumably group the isolates according to their genetic diversity and clonal similarities. By complete genome sequencing we obtained fifteen isolates, which were identified as belonging to CC25 and three isolates belonging to CC49, CC33 and CC162. Eight strains (44.4%) showed resistance to polymyxin B, six of which belong to CC25. These results provide subsidies that highlight the importance of monitoring and control of circulating emerging clones in our environment
 
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Publishing Date
2021-09-23
 
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