We report a high-throughput platform for delivering large cargo into 100,000 cells in 1 min. An array of micro-cavitation bubbles explode in response to laser pulsing, forming pores in adjacent cell membranes, and immediately thereafter, pressurized flows drive slow diffusing cargo through these pores into cells. The platform delivers large cargo including bacteria, enzymes, antibodies, and nanoparticles into diverse cell types with high efficiency and cell viability. We used this platform to explore the intracellular lifestyle of Francisella novicida and discovered that the iglC gene is unexpectedly required for intracellular replication even after phagosome escape into the cell cytosol.

The serum-free medium from Japanese encephalitis virus (JEV) infected Baby Hamster Kidney-21 (BHK-21) cell cultures was analyzed by liquid chromatography tandem mass spectrometry (LC-MS) to identify host proteins that were secreted upon viral infection. Five proteins were identified, including the molecular chaperones Hsp90, GRP78, and Hsp70. The functional role of GRP78 in the JEV life cycle was then investigated. Co-migration of GRP78 with JEV particles in sucrose density gradients was observed and co-localization of viral E protein with GRP78 was detected by immunofluorescence analysis in vivo. Knockdown of GRP78 expression by siRNA did not effect viral RNA replication, but did impair mature viral production. Mature viruses that do not co-fractionate with GPR78 displayed a significant decrease in viral infectivity. Our results support the hypothesis that JEV co-opts host cell GPR78 for use in viral maturation and in subsequent cellular infections.

We report a novel microfluidic integrated optoelectronic tweezers (OET) platform for single-cell sample preparation and analysis. Integration of OET and microfluidics is achieved by embedding single-wall carbon nanotube (SWNT) electrodes into multilayer PDMS structures. This integrated platform allows users to selectively pick up individual cells from a population with light beams based on their optical signatures such as size, shape, and fluorescence, and transport them into isolated chambers using light induced dielectrophoretic forces. Isolated cells can be encapsulated into nanoliter liquid plugs and transported out of the platform for downstream molecule analysis using standard commercial instruments.

Bacterial adherence to epithelial cells is a key virulence trait of pathogenic bacteria. The type 1 fimbriae and the P-fimbriae of uropathogenic Escherichia coli (UPEC) have both been described to be important for the establishment of urinary tract infections (UTI). To explore the interactions between the host and bacterium responsible for the different environments of UPEC invasion, we examined the effect of pH and osmolarity on UPEC strain J96 fimbrial expression, and subsequent J96-induced interleukin-8 (IL-8) expression in different uroepithelial cells. The J96 strain grown in high pH with low osmolarity condition was favorable for the expression of type 1 fimbriae; whereas J96 grown in low pH with high osmolarity condition was beneficial for P fimbriae expression. Type 1 fimbriated J96 specifically invaded bladder 5637 epithelial cells and induced IL-8 expression. On the contrary, P fimbriated J96 invaded renal 786-O epithelial cells and induced IL-8 expression effectively. Type 1 fimbriated J96-induced IL-8 induction involved the p38, as well as ERK, JNK pathways, which leads to AP-1-mediated gene expression. P fimbriated J96-induced augmentation of IL-8 expression mainly involved p38-mediated AP-1 and NF-κB transcriptional activation. These results indicate that different expression of fimbriae in J96 trigger differential IL-8 gene regulation pathways in different uroepithelial cells.

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