A novel siphovirus, vB_PagS_MED16 (MED16) was isolated in Lithuania using Pantoea agglomerans strain BSL for the phage propagation. The double-stranded DNA genome of MED16 (46,103 bp) contains 73 predicted open reading frames (ORFs) encoding proteins, but no tRNA. Our comparative sequence analysis revealed that 26 of these ORFs code for unique proteins that have no reliable identity when compared to database entries. Based on phylogenetic analysis, MED16 represents a new genus with siphovirus morphology. In total, 35 MED16 ORFs were given a putative functional annotation, including those coding for the proteins responsible for virion morphogenesis, phage–host interactions, and DNA metabolism. In addition, a gene encoding a preQ₀ DNA deoxyribosyltransferase (DpdA) is present in the genome of MED16 and the LC–MS/MS analysis indicates 2′-deoxy-7-amido-7-deazaguanosine (dADG)-modified phage DNA, which, to our knowledge, has never been experimentally validated in genomes of Pantoea phages. Thus, the data presented in this study provide new information on Pantoea-infecting viruses and offer novel insights into the diversity of DNA modifications in bacteriophages.

Objective: To identify potential serum biomarkers in women with peritoneal endometriosis (PE) by first looking at its source in the peritoneal fluid (PF). Design: Case-control pilot studies, comprising independent discovery and validation sets. Setting: KK Women’s and Children’s Hospital, Singapore. Patient(s): Women with laparoscopically confirmed PE and absence of endometriosis (control). Intervention(s): None. Main Outcome Measure(s): In the discovery set, we used untargeted liquid chromatography-mass spectrometry (LC-MS/MS) metabolomics, multivariable and univariable analyses to generate global metabolomic profiles of PF for endometriosis and to identify potential metabolites that could distinguish PE (n=10) from controls (n=31). Using targeted metabolomics, we validated the identified metabolites in PF and sera of cases (n=16 PE) and controls (n=19). We performed the area under the receiver-operating characteristics curve (AUC) analysis to evaluate the diagnostic performance of PE metabolites. Result(s): In the discovery set, PF phosphatidylcholine (34:3) and phenylalanyl-isoleucine were significantly increased in PE than controls groups, with AUC 0.77 (95% confidence interval 0.61-0.92; p =0.018) and AUC 0.98 (0.95-1.02; p <0.001), respectively. In the validation set, phenylalanyl-isoleucine retained discriminatory performance to distinguish PE from controls in both PF (AUC 0.77; 0.61-0.92; p =0.006) and serum samples (AUC 0.81; 0.64-0.99; p =0.004). Conclusion(s): Our preliminary results propose phenylalanyl-isoleucine as a potential biomarker of PE, which may be used as a minimally-invasive diagnostic biomarker of PE.

SUMMARY: Shigellosis causes most diarrheal deaths worldwide, particularly affecting children. Shigella invades and replicates in the epithelium of the large intestine, eliciting inflammation and tissue destruction. To understand how Shigella rewires macrophages prior to epithelium invasion, we performed genome-wide and focused secondary CRISPR knockout and CRISPR interference (CRISPRi) screens in Shigella flexneri -infected human monocytic THP-1 cells. Knockdown of the Toll-like receptor 1/2 signaling pathway significantly reduced pro-inflammatory cytokine and chemokine production, enhanced host cell survival, and controlled intracellular pathogen growth. Knockdown of the enzymatic component of the mitochondrial pyruvate dehydrogenase complex enhanced THP-1 cell survival. Small molecule inhibitors blocking key components of these pathways had similar effects; these were validated with human monocyte-derived macrophages, which closely mimic the in vivo physiological state of cells post-infection. High-throughput CRISPR screens can elucidate how S. flexneri triggers inflammation and redirects host pyruvate catabolism for energy acquisition before killing macrophages, pointing to new shigellosis therapies.