The regulation by two transcriptional activators of flagellar expression (FlhD and FlhC) and the chemotaxis methyl-accepting protein Aer was studied with glass slide DNA microarrays. An flhD::Kan insertion and an aer deletion were independently introduced into two Escherichia coli K-12 strains, and the effects upon gene regulation were investigated. Altogether, the flhD::Kan insertion altered the expression of 29 operons of known function. Among them was Aer, which in turn regulated a subset of these operons, namely, the ones involved in anaerobic respiration and the Entner-Doudoroff pathway. In addition, FlhD/FlhC repressed enzymes involved in aerobic respiration and regulated many other metabolic enzymes and transporters in an Aerindependent manner. Expression of 12 genes of uncharacterized function was also affected. FlhD increased gltBD, gcvTHP, and ompT expression. The regulation of half of these genes was subsequently confirmed with reporter gene fusions, enzyme assays, and real-time PCR. Growth phenotypes of flhD and flhC mutants were determined with Phenotype MicroArrays and correlated with gene expression.The Escherichia coli flagellar regulon consists of 14 operons expressed in a transcriptional hierarchy (for a review see reference 35). The flagellar master operon flhDC consists of flhD and flhC (4). The active complex is a heterotetramer (C 2 D 2 ) that binds to the upstream sequences of class II promoters (24). FlhD/FlhC also has nonflagellar targets. Previous work with Panorama E. coli gene arrays (Sigma GenoSys, The Woodlands, Tex.) and lacZ fusions determined that FlhD/FlhC regulates the transporter for galactose (mglBAC), the rodshape determination proteins (mreBCD), the tricarboxylic acid (TCA) cycle enzyme malate dehydrogenase (mdh), and five enzymes involved in anaerobic respiration (41).Although FlhD alone does not bind to class II flagellar promoters, FlhD without the involvement of FlhC is involved in the regulation of the cell division rate (36). Mutants with mutations in flhD, but not those with mutations in flhC, continued to divide rapidly as they entered stationary phase. This was mediated by cadA (39), the second gene in the cadBA operon, encoding lysine decarboxylase (28,29,58).In this study, glass slide genomic microarrays were used to study transcriptional regulation by FlhD and FlhC. Altogether, 29 operons that have a characterized function and 12 genes of unknown function were affected by FlhD or FlhD/FlhC. While the regulation of anaerobic respiration involved the oxygen sensor Aer, aerobic regulation appeared to be unaffected by this protein. MATERIALS AND METHODSBacterial strains and plasmids. Strains and plasmids are presented in Table 1. MC1000 was derived by crossing the araD139 ⌬(araA-leu)7697 allele of D7091 Hfr into M182 (M. Casadaban). MC1000 flhD::Kan contains a kanamycin resistance gene inserted into flhD which exhibits a polar effect upon the expression of flhC (26). YK410 was used as a second parental strain (23). YK4131 (flhD A32D) and YK4136 (flhC N70T) contain p...

SARS CoV-2 and its associated disease COVID-19 has devastated the world during 2020. Masks and social distancing could be efficient if done by large proportions of the population, but pandemic fatigue has decreased their efficacy. Economic shut downs come with large price tags and cannot be a long term solution either. The announcements by three vaccine manufacturers in November that their vaccines are 90% or more effective has given hope to at least those in the population who plan to get vaccinated as soon as a scientifically and medically sound vaccine becomes available. This review summarizes the underlying design strategies and current status of development of the nine vaccines that were in phase III trial on 8 November 2020. Contracts between vaccine manufacturing companies and governments aim at distributing the vaccine to a large part of the world population. Questions remain how the temperature sensitive mRNA vaccines will be transported and/or stored and how vaccination will be prioritized within each country. Additionally, current contracts do not cover all countries, with a serious gap in Africa and South America. The second part of this review will detail current distribution plans and remaining challenges with vaccine accessibility and acceptance.

Aims:  The aim of this study was to develop an assay system that can quantify the amount of biomass in biofilms formed by different isogenic mutants of an Escherichia coli K‐12 strain. Methods and Results:  The reported assay, which is based on the BacTiter‐Glo™ assay from Promega, uses bioluminescence to detect the intracellular concentration of ATP, which correlates with viable bacterial cell numbers. The quantitative data obtained with this ATP assay were compared to those obtained with the conventional crystal violet assay. As a qualitative control, scanning electron microscopy was performed. Conclusions:  The ATP assay, the crystal violet assay and scanning electron microscopy yielded similar results for six of the eight strains tested. For the remaining two strains, the images from the scanning electron microscopy confirmed the results from the ATP assay. Significance and Impact of the Study:  The ATP assay, in combination with other quantitative and qualitative assays, will allow us to perform genetic studies on the regulatory network that underlies the early steps in E. coli biofilm formation.

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Bacterial biofilms are communities of bacteria that form on surfaces and are extremely difficult to remove by conventional physical or chemical techniques, antibiotics or the human immune system. Despite advanced technologies, biofilm still contributes to 60 to 80% of human bacterial infections (NIH and CDC) and cause problems in many natural, environmental, bioindustrial or food processing settings. The discovery of novel substances that inhibit biofilm without increasing the virulence of the bacteria opens doors for countless applications where a reduction of biofilm is desired.