Influence of Antibiotic Pressure on Five Plasmid-based Bioluminescent Gram-negative Bacterial Strains.

PURPOSE: The present study aims to develop five Gram-negative bacteria expressing bacterial luciferase for use to evaluate the influence of different antibiotics on bacterial bioluminescence. PROCEDURES: The pBBR-lux plasmid was introduced into five Gram-negative bacteria; the bioluminescent signals and colony-forming unit (CFU)/ml of all the bioluminescent strains were monitored with six antibiotics at various concentrations. RESULTS: Dose-dependent bioluminescence signals can be used for rapid bacterial antibiotic susceptibility test (AST). All five bioluminescent bacterial strains have similar bioluminescence and CFU enhancement at sub-minimum inhibitory concentration (MIC) of six different antibiotics. CONCLUSION: The bioluminescent signals and CFU enhancement at sub-MIC antibiotic concentrations should be of value in the research of new antibiotic drugs and bioluminescent imaging.

Long noncoding RNA MRCCAT1 promotes metastasis of clear cell renal cell carcinoma via inhibiting NPR3 and activating p38-MAPK signaling.

BACKGROUND: Recent evidences showed that long noncoding RNAs (lncRNAs) are frequently dysregulated and play important roles in various cancers. Clear cell renal cell carcinoma (ccRCC) is one of the leading cause of cancer-related death, largely due to the metastasis of ccRCC. However, the clinical significances and roles of lncRNAs in metastatic ccRCC are still unknown. METHODS: lncRNA expression microarray analysis was performed to search the dysregulated lncRNA in metastatic ccRCC. quantitative real-time PCR was performed to measure the expression of lncRNAs in human ccRCC samples. Gain-of-function and loss-of-function experiments were performed to investigate the biological roles of lncRNAs on ccRCC cell proliferation, migration, invasion and in vivo metastasis. RNA pull-down, RNA immunoprecipitation, chromatin immunoprecipitation, and western blot were performed to explore the molecular mechanisms underlying the functions of lncRNAs. RESULTS: The microarray analysis identified a novel lncRNA termed metastatic renal cell carcinoma-associated transcript 1 (MRCCAT1), which is highly expressed in metastatic ccRCC tissues and associated with the metastatic properties of ccRCC. Multivariate Cox regression analysis revealed that MRCCAT1 is an independent prognostic factor for ccRCC patients. Overexpression of MRCCAT1 promotes ccRCC cells proliferation, migration, and invasion. Depletion of MRCCAT1 inhibites ccRCC cells proliferation, migration, and invasion in vitro, and ccRCC metastasis in vivo. Mechanistically, MRCCAT1 represses NPR3 transcription by recruiting PRC2 to NPR3 promoter, and subsequently activates p38-MAPK signaling pathway. CONCLUSIONS: MRCCAT1 is a critical lncRNA that promotes ccRCC metastasis via inhibiting NPR3 and activating p38-MAPK signaling. Our results imply that MRCCAT1 could serve as a prognostic biomarker and therapeutic target for ccRCC.

Bioluminescence Imaging of Colonization and Clearance Dynamics of Brucella Suis Vaccine Strain S2 in Mice and Guinea Pigs.

PURPOSE: The goal of this study was to develop a plasmid-based lux bio-reporter for use to obtain in vivo images of Brucella suis vaccine strain 2 (B.suis S2) infection with high resolution and good definition. PROCEDURES: The pBBR-lux (pBBR1MCS-2-lxCDABE) plasmid that carries the luxCDABE operon was introduced into B. suis S2 by electroporation yielding B. suis S2-lux. The spatial and temporal transit of B. suis S2 in mice and guinea pigs was monitored by bioluminescence imaging. RESULTS: The plasmid pBBR-lux is stable in vivo and does not appear to impact the virulence or growth of bacteria. This sensitive luciferase reporter could represent B. suis S2 survival in real time. B. suis S2 mainly colonized the lungs, liver, spleen, and uterus in mice and guinea pigs as demonstrated by bioluminescence imaging. CONCLUSION: The plasmid-based lux bioreporter strategy can be used to obtain high resolution in vivo images of B. suis S2 infection in mice and guinea pigs.

Development of Bioluminescent Cronobacter sakazakii ATCC 29544 in a Mouse Model.

Cronobacter sakazakii is an emerging pathogen that causes severe and life-threatening conditions including meningitis, bacteremia, and necrotizing enterocolitis. An animal model study for extrapolation of C. sakazakii infection can provide a better understanding of pathogenesis. However, methods for real-time monitoring of the course of C. sakazakii infection in living animals have been lacking. We developed a bioluminescent C. sakazakii strain (ATCC 29544) that can be used for real-time monitoring of C. sakazakii infection in BALB/c mice. C. sakazakii ATCC 29544 mainly colonized brain, liver, spleen, kidney, and gastrointestinal tract, as indicated by bioluminescence imaging. This work provides a novel approach for studying the progression of C. sakazakii infection and evaluating therapeutics in a living mouse model.