title: Exploratory Investigation ofBacteroides fragilisTranscriptional Response duringIn vitroExposure to Subinhibitory Concentration of Metronidazole.
abstract: Bacteroides fragilis, member from commensal gut microbiota, is an important pathogen associated to endogenous infections and metronidazole remains a valuable antibiotic for the treatment of these infections, although bacterial resistance is widely reported. Considering the need of a better understanding on the global mechanisms by whichB. fragilissurvive upon metronidazole exposure, we performed a RNA-seq transcriptomic approach with validation of gene expression results by qPCR. Bacteria strains were selected afterin vitrosubcultures with subinhibitory concentration (SIC) of the drug. From a wild typeB. fragilisATCC 43859 four derivative strains were selected: first and fourth subcultures under metronidazole exposure and first and fourth subcultures after drug removal. According to global gene expression analysis, 2,146 protein coding genes were identified, of which a total of 1,618 (77%) were assigned to a Gene Ontology term (GO), indicating that most known cellular functions were taken. Among these 2,146 protein coding genes, 377 were shared among all strains, suggesting that they are critical forB. fragilissurvival. In order to identify distinct expression patterns, we also performed a K-means clustering analysis set to 15 groups. This analysis allowed us to detect the major activated or repressed genes encoding for enzymes which act in several metabolic pathways involved in metronidazole response such as drug activation, defense mechanisms against superoxide ions, high expression level of multidrug eﬄux pumps, and DNA repair. The strains collected after metronidazole removal were functionally more similar to those cultured under drug pressure, reinforcing that drug-exposure lead to drastic persistent changes in theB. fragilisgene expression patterns. These results may help to elucidateB. fragilisresponse during metronidazole exposure, mainly at SIC, contributing with information about bacterial survival strategies under stress conditions in their environment.
authors.affiliation: Departamento de Parasitologia, Microbiologia e Imunologia, Universidade Federal de Juiz de Fora Juiz de Fora, Brazil.
authors.forename: Michele C R
authors.initials: MC
authors.lastname: de Freitas
authors.affiliation: Departamento de Parasitologia, Microbiologia e Imunologia, Universidade Federal de Juiz de Fora Juiz de Fora, Brazil.
authors.forename: Juliana A
authors.initials: JA
authors.lastname: Resende
authors.affiliation: Departamento de Parasitologia, Microbiologia e Imunologia, Universidade Federal de Juiz de Fora Juiz de Fora, Brazil.
authors.forename: Alessandra B
authors.initials: AB
authors.lastname: Ferreira-Machado
authors.affiliation: Laboratório de Bioinformática and Laboratório Nacional de Computação Científica Petrópolis, Brazil.
authors.forename: Guadalupe D R Q
authors.initials: GD
authors.lastname: Saji
authors.affiliation: Laboratório de Bioinformática and Laboratório Nacional de Computação Científica Petrópolis, Brazil.
authors.forename: Ana T R
authors.initials: AT
authors.lastname: de Vasconcelos
authors.affiliation: Departamento de Parasitologia, Microbiologia e Imunologia, Universidade Federal de Juiz de Fora Juiz de Fora, Brazil.
authors.forename: Vânia L
authors.initials: VL
authors.lastname: da Silva
authors.affiliation: Laboratório de Bioinformática and Laboratório Nacional de Computação Científica Petrópolis, Brazil.
authors.forename: Marisa F
authors.initials: MF
authors.lastname: Nicolás
authors.affiliation: Departamento de Parasitologia, Microbiologia e Imunologia, Universidade Federal de Juiz de Fora Juiz de Fora, Brazil.
authors.forename: Cláudio G
authors.initials: CG
authors.lastname: Diniz
keywords: Bacteroides fragilis; K-means clustering analysis; RNA-seq; metronidazole; subinhibitory concentration