Navegando por Palavras-chave "Acinetobacter Spp"

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    Aplicação do sequenciamento completo do genoma ba caracterização de amostras de acinetobacter baumannii produtores de carbapenemase
    (Universidade Federal de São Paulo (UNIFESP), 2018-03-20) Boettger, Bruno Cruz [UNIFESP]; Pignatari, Antonio Carlos Campos [UNIFESP]; http://lattes.cnpq.br/9461346610553865; http://lattes.cnpq.br/9782967668875923
    Introdução: Infecções por Acinetobacter baumanni apresentam elevadas taxas de morbidade e mortalidade, particularmente quando os isolados são resistentes a antibióticos carbapenêmicos. Surtos de infecções relacionadas à assistência a saúde tem sido detectados com disseminação de clones intra e inter hospitais incluindo a determinação de genes codificadores de oxacilinases, o principal mecanismos de resistência a carbapenêmicos. Técnicas moleculares têm sido utilizadas para tipagem molecular, como Pulsed-Fiel Gel Electrophoresis (PFGE) e MLST por sequenciamento de housekeeping genes e caracterização de genes de resistência aos antimicrobianos por PCR convencional e real time. Recentemente, o sequenciamento de genoma completo tem sido utilizado como um método que permite a tipagem molecular e a caracterização de genes de resistência em diferentes gêneros e espécies bacterianas. Objetivos: Identificar genes codificadores de resistência a antimicrobianos e caracterizar perfis clonais de A. baumannii resistentes a carbapenêmicos por sequenciamento de genoma completo. Métodos: Foram estudadas seis amostras de A. baumannii resistentes a carbapenêmicos isoladas de pacientes internados em hospital geral da cidade de São Paulo no período de 20 de novembro a 03 de dezembro de 2016. O perfil de resistência a antimicrobianos foi realizado por microdiluição em caldo e a tipagem molecular por PFGE. As sequencias de DNA do genoma completo foram obtidas através da plataforma Illumina, anotação de bioinformática pelo software Prokka e análise pelo Center for Genomic Epidemiology. Resultados: Somente a polimixina B, minociclina e tigeciclina apresentaram atividade frente a todos os isolados de A. baumannii resistentes aos carbapenêmicos. Genes de resistência aos aminoglicosídeos, β-lactâmicos, fenicois, macrolídeos, quinolonas, sulfonamidas, tetraciclinas e trimetroprim foram encontrados no genoma dos isolados. Uma coprodução de OXA-23 e OXA-72 foram detectadas em um isolado de A. baumannii pertencente ao ST1/CC1. Dois isolados apresentando mesmo padrão de PFGE pertencente ao ST79/CC79 carreavam os genes blaOXA-23 e blaOXA-72 separadamente localizados no cromossomo e no plasmídeo, respectivamente. O gene qnrD-like foi detectado no isolado de A. baumannii coprodutor de OXA-23 e OXA-72, sendo o primeiro isolado de Acinetobacter spp. reportado a carrear este gene globalmente. Conclusão: O sequenciamento completo de genoma de A. baumanii revelou a presença de genes de resistência a diversos antimicrobianos, incluindo os codificadores de oxacilinases, e contribuiu para a tipagem molecular com finalidades epidemiológicas.
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    Caracterização de plasmídeos de isolados clínicos de acinetobacter spp. produtores de carbapenemases
    (Universidade Federal de São Paulo (UNIFESP), 2017-12-05) Silva, Adriana Pereira de Matos Marques [UNIFESP]; Gales, Ana Cristina [UNIFESP]; Silva, Rodrigo Cayô da [UNIFESP]; http://lattes.cnpq.br/5699739668358897; http://lattes.cnpq.br/8402272715765172; http://lattes.cnpq.br/1117846722738557; Universidade Federal de São Paulo (UNIFESP)
    ABSTRACT Acinetobacter spp. is a nosocomial pathogen frequently isolated in ICU patients. High resistance rates have been detected among Acinetobacter spp.. Carbapenem Hydrolyzing Class D β-lactamases (CHDL) production is the most frequent and the most important resistance mechanism. This species is able to acquire plasmids and mobile genetic elements (MGEs) that mobilize resistance genes. This study aimed to describe plasmids of carbapenemase producing Acinetobacter spp. clinical isolates. Six carbapenem resistant Acinetobacter spp. strains were selected as OXA-58, OXA-143, OXA-231 and OXA-253 producing. Strain identification was performed on MALDI-TOF MS and confirmed by rpoB sequencing. Carbapenemase detection was confirmed by PCR and susceptibility testing was performed by broth microdilution. MLST was performed to evaluate the ancestrality. The pool of plasmids was extracted using Mini Prep Extraction Kit® and quantified in Qubit® 3.0. Libraries were constructed using Illumina® TruSeq Nano DNA LT Library Preparation Kit – SetA generating ~550 bp and sequencing was performed in Illumina® MiSeq 2x300 bp in paired-end mode. Sequences were assembled using softwares Newbler 3.0 and Ray 2.3.1. Automatic annotation was performed using SABIA/LNCC platform and manual validation was performed using NCBI Blast, UniProt, ISFinder e ResFinder 2.1 softwares. The illustration of the circularized plasmids and in silico analysis of replicase gene group (AbGR) was obtained of Snap Gene® program. Promoter sequences were suggested by BPROM tool. Five of six strains were identified as A. baumanni and one was identified as A. seifertii. High MIC rates were observed to the β-lactams tested. Only polymyxin B and minocyclin presented good activity to the tested strains, except A. seifertii that showed resistance to polymyxin B (MIC: 4μg/mL). OXA-231 producing strain was included on ST107, OXA-253 on ST25 and OXA-58 producing A. seifertii and A. baumannii were included on ST551 and ST15, respectively. A total of 23 plasmids were obtained from the six analyzed strains. Of strain A-58.015 plasmids pAb58015_a (118.738 bp/173 ORFs; blaOXA-143 carrying), pAb58015_b (55.356 bp/62 ORFs), pAb58015_c (6.520 bp/9 ORFs) and pAb58015_d (2.368 bp/3 ORFs) were obtained; on strain 81048, three identic plasmids pAb58015_b, pAb58015_c and pAb58015_d were observed and also plasmids pAb81048_a (113.646 bp/177 ORFs), pAb81048_b (87.915 bp/101 ORFs), pAb81048_c (13.910 bp/21 ORFs), pAb81048_d (9.304 bp/15 ORFs), pAb81048_e (7.763 bp/6 ORFs; blaOXA-143 carrying), pAb81048_f (5.300 bp/6 ORFs) and pAb81048_g (2.845 bp/5 ORFs) were obtained; on strain A-41.652, plasmids pAb41652_a (101.946 bp/137 ORFs), pAb41652_b (11.844 bp/12 ORFs) and pAb41652_c (3.955 bp/3 ORFs; blaOXA-231 carrying) were observed; on strain 86, plasmids pAb86_a (234.383 bp/256 ORFs) and pAb86_b (13.453 bp/14 ORFs; blaOXA-253 carrying) were sequenced; on strain 1069, plasmids pAs1069_a (24.672 bp/44 ORFs; blaOXA-58 carrying) and pAs1069_b (13.129 bp/14 ORFs); and from strain A-45.063, plasmids pAb45063_a (183.767 bp/209 ORFs) and pAb45063_b (19.808 bp/24 ORFs; blaOXA-58 carrying) were verified. Plasmids were classified in six different AbGR groups (GR2, GR3, GR4, GR6, GR8 and GR19). Resistance genes of distinct antimicrobial classes were found in approximately half of the sequenced plasmids. Nine plasmids had two or three virulence genes. A new genetic environment in OXA-143 and variants OXA-231 and OXA-253 was described in our isolates. The presence of an ISAba825 upstream blaOXA-58 in A. seifertii was the main difference on the genetic environment of blaOXA-58 inserted in plasmid pAb45063_b in A. baumannii. The number of MGEs found in plasmids evaluated in this study suggests a high mobilization capacity of those structures and consequently horizontal transfer of resistance.