The Fis nucleoid-associated protein controls the expression of a large and diverse regulon of genes in Gram-negative bacteria. Fis production is normally maximal in bacteria during the early exponential phase of batch culture growth, becoming almost undetectable by the onset of stationary phase. We tested the effect on the Fis regulatory network in Salmonella of moving the complete fis gene from its usual location near the origin of chromosomal replication to the position normally occupied by the dps gene in the right macrodomain of the chromosome, and vice versa, creating the gene exchange (GX) strain. In a parallel experiment, we tested the effect of rewiring the Fis regulatory network by placing the fis open reading frame under the control of the stationary-phase-activated dps promoter at the dps genetic location within the right macrodomain, and vice versa, creating the open reading frame exchange (OX) strain. Chromatin immunoprecipitation sequencing (ChIP-seq) was used to measure global Fis protein binding levels and to determine gene expression patterns. Strain GX showed few changes compared with the wild type, although we did detect increased Fis binding at Ter, accompanied by reduced binding at Ori. Strain OX displayed a more pronounced version of this distorted Fis protein-binding pattern together with numerous alterations in the expression of genes in the Fis regulon. OX, but not GX, had a reduced ability to infect cultured mammalian cells. These findings illustrate the inherent robustness of the Fis regulatory network with respect to the effects of rewiring based on gene repositioning alone and emphasize the importance of fis expression signals in phenotypic determination. IMPORTANCE We assessed the impact on Salmonella physiology of reciprocally translocating the genes encoding the Fis and Dps nucleoid-associated proteins (NAPs) and of inverting their growth-phase production patterns such that Fis was produced in stationary phase (like Dps) and Dps was produced in exponential phase (like Fis). Changes to peak binding of Fis were detected by ChIP-seq on the chromosome, as were widespread impacts on the transcriptome, especially when Fis production mimicked Dps production. Virulence gene expression and the expression of a virulence phenotype were altered. Overall, these radical changes to NAP gene expression were well tolerated, revealing the robust and well-buffered nature of global gene regulation networks in the bacterium.

The importance of infection control procedures in hospital radiology departments has become increasingly apparent in recent months as the impact of COVID-19 has spread across the world. Existing disinfectant procedures that rely on the manual application of chemical-based disinfectants are time consuming, resource intensive and prone to high degrees of human error. Alternative non-touch disinfection methods, such as Ultraviolet Germicidal Irradiation (UVGI), have the potential to overcome many of the limitations of existing approaches while significantly improving workflow and equipment utilization. The aim of this research was to investigate the germicidal effectiveness and the practical feasibility of using a robotic UVGI device for disinfecting surfaces in a radiology setting. We present the design of a robotic UVGI platform that can be deployed alongside human workers and can operate autonomously within cramped rooms, thereby addressing two important requirements necessary for integrating the technology within radiology settings. In one hospital, we conducted experiments in a CT and X-ray room. In a second hospital, we investigated the germicidal performance of the robot when deployed to disinfect a CT room in <15 minutes, a period which is estimated to be 2–4 times faster than current practice for disinfecting rooms after infectious (or potentially infectious) patients. Findings from both test sites show that UVGI successfully inactivated all of measurable microbial load on 22 out of 24 surfaces. On the remaining two surfaces, UVGI reduced the microbial load by 84 and 95%, respectively. The study also exposes some of the challenges of manually disinfecting radiology suites, revealing high concentrations of microbial load in hard-to-reach places. Our findings provide compelling evidence that UVGI can effectively inactivate microbes on commonly touched surfaces in radiology suites, even if they were only exposed to relatively short bursts of irradiation. Despite the short irradiation period, we demonstrated the ability to inactivate microbes with more complex cell structures and requiring higher UV inactivation energies than SARS-CoV-2, thus indicating high likelihood of effectiveness against coronavirus.

Antimicrobial peptides offer potential as novel therapeutics to combat food spoilage and poisoning caused by pathogenic and nonpathogenic bacteria. Our previous studies identified the peptide human beta-defensin 3 (HBD3) as a potent antimicrobial agent against a wide range of beer-spoiling bacteria. Thus, HBD3 is an excellent candidate for development as an additive to prevent food and beverage spoilage. To expand the repertoire of peptides with antimicrobial activity against bacteria associated with food spoilage and/or food poisoning, we carried out an in silico discovery pipeline to identify peptides with structure and activity similar to those of HBD3, focusing on peptides of plant origin. Using a standardized assay, we compared the antimicrobial activities of nine defensin-like plant peptides to the activity of HBD3. Only two of the peptides, fabatin-2 and Cp-thionin-2, displayed antimicrobial activity; however, the peptides differed from HBD3 in being sensitive to salt and were thermostable. We also compared the activities of several ultrashort peptides to that of HBD3. One of the peptides, the synthetic tetrapeptide O3TR, displayed biphasic antimicrobial activity but had a narrower host range than HBD3. Finally, to determine if the peptides might act in concert to improve antimicrobial activity, we compared the activities of the peptides in pairwise combinations. The plant defensin-like peptides fabatin-2 and Cp-thionin-2 displayed a synergistic effect with HBD3, while O3TR was antagonistic. Thus, some plant defensin-like peptides are effective antimicrobials and may act in concert with HBD3 to control bacteria associated with food spoilage and food poisoning. IMPORTANCEFood spoilage and food poisoning caused by bacteria can have major health and economic implications for human society. With the rise in resistance to conventional antibiotics, there is a need to identify new antimicrobials to combat these outbreaks in our food supply. Here we screened plant peptide databases to identify peptides that share structural similarity with the human defensin peptide HBD3, which has known antimicrobial activity against food-spoiling bacteria. We show that two of the plant peptides display antimicrobial activity against bacteria associated with food spoilage. When combined with HBD3, the peptides are highly effective. We also analyzed the activity of an easily made ultrashort synthetic peptide, O3TR. We show that this small peptide also displays antimicrobial activity against food-spoiling bacteria but is not as effective as HBD3 or the plant peptides. The plant peptides identified are good candidates for development as natural additives to prevent food spoilage.A ntimicrobial peptides (AMPs) are short oligopeptides ranging from 4 to 100 amino acids that display antimicrobial activity against a broad range of pathogens, including Gram-positive and -negative bacteria, fungi, viruses, protists, parasites, and even insects (1). AMPs form part of the innate immune response, acting as a first line of defense and...

The Integration Host Factor (IHF) is a heterodimeric nucleoid-associated protein that plays roles in bacterial nucleoid architecture and genome-wide gene regulation. The ihfA and ihfB genes encode the subunits and are located 350 kbp apart, in the Right replichore of the Salmonella chromosome. IHF is composed of one IhfA and one IhfB subunit. Despite this 1 : 1 stoichiometry, MS revealed that IhfB is produced in 2-fold excess over IhfA. We re-engineered Salmonella to exchange reciprocally the protein-coding regions of ihfA and ihfB, such that each relocated protein-encoding region was driven by the expression signals of the other’s gene. MS showed that in this 'rewired' strain, IhfA is produced in excess over IhfB, correlating with enhanced stability of the hybrid ihfB–ihfA mRNA that was expressed from the ihfB promoter. Nevertheless, the rewired strain grew at a similar rate to the wild-type and was similar in competitive fitness. However, compared to the wild-type, it was less motile, had growth-phase-specific reductions in SPI-1 and SPI-2 gene expression, and was engulfed at a higher rate by RAW macrophage. Our data show that while exchanging the physical locations of its ihf genes and the rewiring of their regulatory circuitry are well tolerated in Salmonella , genes involved in the production of type 3 secretion systems exhibit dysregulation accompanied by altered phenotypes.

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