Small RNA molecules are short, non-coding RNAs identified for their crucial role in post-transcriptional regulation. A well-studied example includes miRNAs (microRNAs) which have been identified in several model organisms including the freshwater flea and planktonic crustacean Daphnia. A model for epigenetic-based studies with an available genome database, the identification of miRNAs and their potential role in regulating Daphnia gene expression has only recently garnered interest. Computational-based work using Daphnia pulex, has indicated the existence of 45 miRNAs, 14 of which have been experimentally verified. To extend this study, we took a sequencing approach towards identifying miRNAs present in a small RNA library isolated from Daphnia magna. Using Perl codes designed for comparative genomic analysis, 815,699 reads were obtained from 4 million raw reads and run against a database file of known miRNA sequences. Using this approach, we have identified 205 putative mature miRNA sequences belonging to 188 distinct miRNA families. Data from this study provides critical information necessary to begin an investigation into a role for these transcripts in the epigenetic regulation of Daphnia magna.

Listeria monocytogenes is a ubiquitous opportunistic foodborne pathogen capable of survival in various adverse environmental conditions. Pathogenesis of L. monocytogenes is tightly controlled by a complex regulatory network of transcriptional regulators that are necessary for survival and adaptations to harsh environmental conditions both inside and outside host cells. Among these regulatory pathways are members of the DeoR-family transcriptional regulators that are known to play a regulatory role in sugar metabolism. In this study, we deciphered the role of FruR, a DeoR family protein, which is a fructose operon transcriptional repressor protein, in L. monocytogenes pathogenesis and growth. Following intravenous (IV) inoculation in mice, a mutant strain with deletion of fruR exhibited a significant reduction in bacterial burden in liver and spleen tissues compared to the parent strain. Further, the ΔfruR strain had a defect in cell-to-cell spread in L2 fibroblast monolayers. Constitutive activation of PrfA, a pleiotropic activator of L. monocytogenes virulence factors, did not restore virulence to the ΔfruR strain, suggesting that the attenuation was not a result of impaired PrfA activation. Transcriptome analysis revealed that FruR functions as a positive regulator for genes encoding enzymes involved in the pentose phosphate pathway (PPP) and as a repressor for genes encoding enzymes in the glycolysis pathway. These results suggested that FruR may function to facilitate NADPH regeneration, which is necessary for full protection from oxidative stress. Interestingly, deletion of fruR increased sensitivity of L. monocytogenes to H2O2, confirming a role for FruR in survival of L. monocytogenes during oxidative stress. Using anti-mouse neutrophil/monocyte monoclonal antibody RB6-8C5 (RB6) in an in vivo infection model, we found that FruR has a specific function in protecting L. monocytogenes from neutrophil/monocyte-mediated killing. Overall, this work clarifies the role of FruR in controlling L. monocytogenes carbon flow between glycolysis and PPP for NADPH homeostasis, which provides a new mechanism allowing metabolic adaptation of L. monocytogenes to oxidative stress.