15 16 17 18 Running Tittle: SEM images of E. coli strains harboring the pks genomic island 19 Abstract 49Colibactin is a natural product made by numerous strains of E. coli that harbor the pks 50 genomic island. The deletion of one of the genes within the pks island, the peptidase 51 clbP, has been found to disrupt the maturation of colibactin, thus promoting the 52 accumulation in the periplasmic space of numerous biosynthesis intermediates, some of 53 which have been characterized chemically. To date, no one has reported the effect of 54 such an accumulation of intermediates on the cellular morphology of the producing E. 55 coli bacterium. In this report, we describe the scanning electron microscopy (SEM) 56 images of numerous clinical isolates of E. coli harboring the pks island, collected from 57Puerto Rico hospitals. We have observed that the wild type isolates that harbor the pks 58 island display lesions on the bacterial envelope surface. These lesions are absent in 59 isolates lacking the pks island. To determine whether this phenotype is associated with 60 colibactin production, we deleted the clbP gene from the extraintestinal pathogenic E. 61 coli strain IHE3034, thus disrupting its ability to make colibactin. The wild-type IHE3034 62 displayed a spherical shape with no envelope lesions, and was practically 63 indistinguishable from the ΔclbP deletion mutant. To our knowledge, this work provides 64 the first SEM images of a pks deletion mutant. 65 66 67 68 69 70 71 87 88 89 90 91 92 93 94
All gram‐negative bacteria secrete outer membrane vesicles (OMV) that have several functions, among them the transportation of bacterial virulence and survival factors. It has been found by other groups that certain OMV promote aberrant growth in mammalian cells upon contact. OMV provide an insoluble secretion pathway for gram‐negative bacteria and are responsible for the release of outer membrane and periplasm content out of the cell. The main objective of this investigation is to compare the composition and biological activity of OMV isolated from diverse clinical strains of E. coli. We will assess morphological effects of OMV from selected clinical isolates of E. coli on HeLa cells. Furthermore, we will also measure the lipid composition of OMV for the different strains and mutants by GC/MS. As preliminary results, we observed a significant difference in the lipid composition of the IHE3034 (pks+ strain) OMV and the IHE3034‐ΔclbP OMV. The clbP gene is presumed to be involved in the biosynthesis of colibactin. We observed that the deletion of clbP peptidase causes a decrease in the percentage of 12:0 fatty acids and an increase of 16:0 fatty acids in OMV, but not in the cellular membrane. These results begin to establish a correlation between the presence of colibactin genes and events at the bacterial cell surface.Support or Funding InformationNSF grant CHE0953254 NIH grant R25GM061838 (MBRS‐RISE program) to Ramón Gómez‐Moreno
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