Microarray-based identification of differentially expressed genes in intracellular Brucella abortus within RAW264.7 cells.
Brucella spp. is a species of facultative intracellular Gram-negative bacteria that induces abortion and causes sterility in domesticated mammals and chronic undulant fever in humans. Important determinants of Brucella's virulence and potential for chronic infection include the ability to circumvent the host cell's internal surveillance system and the capability to proliferate within dedicated and non-dedicated phagocytes. Hence, identifying genes necessary for intracellular survival may hold the key to understanding Brucella infection. In the present study, microarray analysis reveals that 7.82% (244/3334) of all Brucella abortus genes were up-regulated and 5.4% (180/3334) were down-regulated in RAW264.7 cells, compared to free-living cells in TSB. qRT-PCR verification further confirmed a >5-fold up-regulation for fourteen genes. Functional analysis classified araC, ddp, and eryD as to partake in information storage and processing, alp, flgF and virB9 to be involved in cellular processes, hpcd and aldh to play a role in metabolism, mfs and nikC to be involved in both cellular processes and metabolism, and four hypothetical genes (bruAb1_1814, bruAb1_0475, bruAb1_1926, and bruAb1_0292) had unknown functions. Furthermore, we constructed a B. abortus 2308 mutant Δddp where the ddp gene is deleted in order to evaluate the role of ddp in intracellular survival. Infection assay indicated significantly higher adherence and invasion abilities of the Δddp mutant, however it does not survive well in RAW264.7 cells. Brucella may survive in hostile intracellular environment by modulating gene expression.
Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Science, Shanghai, PR China.
Mingxing
M
Tian
Jing
J
Qu
Xiangan
X
Han
Min
M
Zhang
Chan
C
Ding
Jiabo
J
Ding
Guanghua
G
Chen
Shengqing
S
Yu
D000818:Animals; D002003:Brucella abortus / Q000235:genetics*; D002460:Cell Line; D016000:Cluster Analysis; D015964:Gene Expression Regulation, Bacterial; D042541:Intracellular Space / Q000382:microbiology; D008264:Macrophages / Q000166:cytology / Q000382:microbiology*; D051379:Mice; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D059467:Transcriptome