Project description:Salmonella enterica is an important foodborne pathogen. Here, we present the construction and characterization of a high-density transposon sequencing library of the Salmonella Typhimurium ATCC 14028 strain. Essential, advantageous, and disadvantageous genes for growth in rich culture medium were identified on the chromosome and the pSLT plasmid.
Project description:The goal of this study was to determine how the method of pooling (combining mutants) for bacteral Tn libraries affected mutant distribution. Previously we generated a pooled version of a~6,800 Tn mutant library in Enterococcus faecalis OG1RF by plating and scraping individual mutants. Here, we combined liquid cultures grown in deep well plates. DNA from the new pooled library was extracted, and TnSeq was used to compare mutant distribution between the old (plate scraping) and new (liquid pooling) Tn library formats. This is important because it will guide best practices for handling large collections of bacterial mutants.
Project description:Salmonella enterica is one of the most important foodborne pathogens that infect a variety of animals and birds. In humans, S. Typhimurium causes gastroenteritis, leading to vomiting, diarrhea, fever, and abdominal cramps. We mainly get infected with Salmonella by ingesting comminated poultry products. Therefore, developing an oral live attenuated vaccine for the poultry industry is our best bet against Salmonella infection. In this article, we investigated the potential of the next generation of Salmonella vaccines. We generated a library of potentially attenuated S. Typhimurium mutants and compared fitness to that of a commercial vaccine. We also investigated the invasion and survival potential of these mutants in chicken macrophages. Our data indicate that although these mutants had no significant growth defects, they were much sensitive to macrophage attack. Analyzing the transcriptome data from infected primary chicken macrophages, we concluded that these mutants elicit a robust immune response by activating several immunoregulatory pathways. Our data also indicates that by combining phoPQ deletion with an already existing cya-crp deletion in MeganVac1, a much stronger immune response can be generated.
Project description:We developed a surface-displayed library of 117 human antimicrobial peptides (AMPs) and over 3000 human AMP variants in a laboratory E. coli strain expressing PhoP and PhoQ from Salmonella Typhimurium. We used sort-seq (fluorescence-activated cell sorting followed by next-generation sequencing) to characterize PhoPQ activation by these surface-displayed AMPs.