title: Identification of genes contributing to the intracellular replication of Brucella abortus within HeLa and RAW 264.7 cells.
abstract: Brucella abortus, the causative agent of brucellosis, can survive and replicate within host cells. Understanding bacterial virulence factors and bacteria-host cell interactions is critical for controlling brucellosis, yet very little is known about the virulence strategies and signaling pathways activated in phagocytes during infection to ensure their growth and survival. B. abortus was mutagenized by mini-Tn5Km2 transposon mutagenesis to identify virulence genes related to the internalization and intracellular replication of the bacteria. Of the total 2300 mutants used to infect HeLa cells, 23 mutants defective for intercellular growth and the mutated genes were identified. Sequence analysis of DNA flanking the transposon showed various insertion sites in bacterial genes that might be associated with virulence, including genes associated with lipoproteins, amino acid metabolism, translation, transcription, carbohydrate transport, coenzyme transport, inorganic ion transport, energy metabolism, membrane transport, and cell wall/membrane biogenesis. Moreover, mutants were classified into class I, class II and class III as higher, similar, and lower internalization, respectively, into HeLa cells. Furthermore, defective mutants for intracellular growth in HeLa cells were found to be defective in RAW 264.7 cells. Taken together, we suggest that the identified virulence associated genes might contribute to the intracellular growth and survival of B. abortus in phagocytes.
authors.affiliation: College of Veterinary Medicine, Gyeongsang National University, Jinju 660-701, Republic of Korea.
authors.forename: Dong Hyeok
authors.initials: DH
authors.lastname: Kim
authors.forename: Jeong Ju
authors.initials: JJ
authors.lastname: Lim
authors.forename: Jin Ju
authors.initials: JJ
authors.lastname: Lee
authors.forename: Dae Geun
authors.initials: DG
authors.lastname: Kim
authors.forename: Hu Jang
authors.initials: HJ
authors.lastname: Lee
authors.forename: Wongi
authors.initials: W
authors.lastname: Min
authors.forename: Kwang Dong
authors.initials: KD
authors.lastname: Kim
authors.forename: Hong Hee
authors.initials: HH
authors.lastname: Chang
authors.forename: Man Hee
authors.initials: MH
authors.lastname: Rhee
authors.forename: Masahisa
authors.initials: M
authors.lastname: Watarai
authors.forename: Suk
authors.initials: S
authors.lastname: Kim
mesh_terms: D000818:Animals; D002003:Brucella abortus / Q000166:cytology / Q000235:genetics / Q000472:pathogenicity / Q000502:physiology*; D002006:Brucellosis / Q000382:microbiology*; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D004251:DNA Transposable Elements / Q000235:genetics; D005260:Female; D005798:Genes, Bacterial / Q000235:genetics; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D016296:Mutagenesis; D009154:Mutation; D010586:Phagocytes / Q000382:microbiology; D014774:Virulence / Q000235:genetics
chemical_list: D004251:DNA Transposable Elements