Barryklavsen7718

nto seven phylogroups, which may differ in their reservoirs and epidemiology. Proper management of this public health hazard requires a better understanding of Kp ecology and routes of transmission to humans. So far, detection of these microorganisms in complex matrices such as food or the environment has been difficult due to a lack of accurate and sensitive methods. Here, we describe a novel method based on real-time PCR, which enables detection of all Kp phylogroups with high sensitivity and specificity. We used this method to detect Kp isolates from environmental samples, and show based on genomic sequencing that they differ in antimicrobial resistance and virulence gene content, from human clinical Kp isolates. The ZKIR PCR assay will enable rapid screening of multiple samples for Kp presence and will thereby facilitate tracking the dispersal patterns of these pathogenic strains across environmental, food, animal and human sources. Copyright © 2020 Barbier et al.In Saccharomyces cerevisiae, Y family DNA polymerase Rev1 is involved in the repair of DNA damage by translesion DNA synthesis (TLS). In the current study, to elucidate the role of Rev1 in oxidative stress-induced DNA damage in S. cerevisiae, REV1 was deleted and overexpressed; transcriptome analysis of these mutants along with the wild-type strain was performed to screen potential genes that could be associated with REV1 during response to DNA damage. Under 2 mM H2O2 treatment, the deletion of REV1 resulted in a 1.5- and 2.8-fold decrease in the survival rate and mutation frequency, respectively, whereas overexpression of REV1 increased the survival rate and mutation frequency by 1.1- and 2.9-fold, respectively, as compared to the survival rate and mutation frequency of the wild-type strain. Transcriptome and phenotypic analyses identified that Sml1 aggravated oxidative stress in the yeast cells by inhibiting the activity of Rev1. This inhibition was due to the physical interaction between the BRCT domain of Rev1 and the amino acid residues 36-70 of Sml1, the cell survival rate and mutation frequency increased by 1.8 and 3.1-fold, respectively, upon blocking this interaction. We also found that Sml1 inhibited Rev1 phosphorylation under oxidative stress, deletion of SML1 increased the phosphorylation of Rev1 by 46 %, whereas overexpression of SML1 reduced phosphorylation of Rev1. Overall, these findings demonstrate that Sml1 could be a novel regulator that mediates Rev1 dephosphorylation to inhibit its activity during oxidative stress.IMPORTANCE Rev1 was critical for cell growth in S. cerevisiae, and the deletion of REV1 caused a severe growth defect under exposure to oxidative stress with 2 mM H2O2 Furthermore, we found that Sml1 physically interacted with Rev1 and inhibited Rev1 phosphorylation, thereby inhibited Rev1 DNA antioxidant activity. These findings indicate that Sml1 could be a novel regulator for Rev1 in response to DNA damage by oxidative stress. Copyright © 2020 American Society for Microbiology.The siderophore synthetic system in Shewanella species is able to synthesize dozens of macrocyclic siderophores in vitro with synthetic precursors. In vivo, however, although three siderophores are produced naturally in S. algae B516 which carries a lysine decarboxylase (AvbA) specific for siderophore synthesis, only one siderophore can be detected from many other Shewanella species. read more In this study, we examined a siderophore-overproducing mutant of S. oneidensis, which lacks an AvbA counterpart, and found that it can also produce these three siderophores. We identified both SpeC and SpeF as promiscuous decarboxylases for both lysine and ornithine to synthesize siderophore precursors cadaverine and putrescine respectively. Intriguingly, putrescine is mainly synthesized from arginine through an arginine decarboxylation pathway in a constitutive manner, not liable to the concentrations of iron and siderophores. Our results provide further evidence that the substrate availability plays a determining role in siderolability. In addition to use the ADC pathway for putrescine synthesis, cells optimize the putrescine pool for siderophore production. Our work provides an insight into coordinated synthesis of multiple siderophores by harnessing promiscuous enzymes in bacteria and underscores the importance of substrate pools for biosynthesis of natural products. Copyright © 2020 American Society for Microbiology.Escherichia coli O157H7 and Salmonella enterica are leading causes of foodborne outbreaks linked to fresh produce. Both species can enter the "viable but non-culturable" (VBNC) state that precludes detection using conventional culture-based or molecular methods. In this study, we assessed propidium monoazide (PMA)-qPCR assays and novel methods combining PMA and loop-mediated isothermal amplification (LAMP) for the detection and quantification of VBNC E. coli O157H7 and S. enterica in fresh produce. The performance of PMA-LAMP assays targeting wzy gene of E. coli O157H7 and agfA gene of S. enterica and PMA-qPCR assays were compared in pure culture and spiked tomato, lettuce and spinach. No cross-reaction was observed in the specificity tests. The limit of detection (LOD) with PMA-LAMP was 9.0 CFU/reaction for E. coli O157H7 and 4.6 CFU/reaction for S. enterica in pure culture, and 5.13×103-4 CFU/g for VBNC E. coli O157H7 and 1.05×104-5 CFU/g for VBNC S. enterica in fresh produce, which was comparable to that oy to determine VBNC E. coli O157H7 and S. enterica in fresh produce, which potentially decreases the risks related to the consumption of fresh produce contaminated by enteric pathogens in this state. PMA-LAMP can be further applied in the field study to enhance our understanding of the fate of VBNC pathogens in the pre- and post-harvest stages of fresh produce. Copyright © 2020 American Society for Microbiology.Ergosterol plays an important role in maintaining cell membrane sterol homeostasis in fungi, and as such, is considered as an effective target in antifungal chemotherapy. In yeast, the enzyme acetyl-CoA acetyltransferase (ERG10) catalyzes the Claisen condensation of two acetyl-CoA molecules to acetoacetyl-CoA in the ergosterol biosynthesis pathway and is reported critical for cell viability. Using yeast ERG10 for alignment, two orthologues AfERG10A (AFUB_000550) and AfERG10B (AFUB_083570) were discovered in opportunistic fungal pathogen Aspergillus fumigatus Despite the essentiality of AfERG10B has been previously validated, the biological function of AfERG10A remains unclear. In this study, we have characterized recombinant AfERG10A as a functional acetyl-CoA acetyltransferase catalyzing both synthetic and degradative reactions. Unexpectedly, AfERG10A localizes to mitochondria in A. fumigatus as shown by C-terminal GFP-tag fusion. Both knockout and inducible promoter strategies demonstrate that Aferg10A is essential for the survival of A.