Damgaardlambertsen9594

Polyketide-terpenoid hybrid compounds are one of the largest families of meroterpenoids, with great potential for drug development for resistant pathogens. Genome sequence analysis of secondary metabolite gene clusters of a phytopathogenic fungus, Bipolaris sorokiniana 11134, revealed a type I polyketide gene cluster, consisting of highly reducing polyketide synthase, non-reducing polyketide synthase, and adjacent prenyltransferase. MS- and UV-guided isolations led to the isolation of ten meroterpenoids, including two new compounds 19-dehydroxyl-3-epi-arthripenoid A (1) and 12-keto-cochlioquinone A (2). The structures of 1-10 were elucidated by the analysis of NMR and high-resolution electrospray ionization mass spectroscopy data. Compounds 5-8 and 10 showed moderate activity against common Staphylococcus aureus and methicillin-resistant S. aureus, with minimum inhibitory concentration (MIC) values of 12.5-100 μg/mL. Compound 5 also exhibited activity against four clinical resistant S. aureus strains and synergistic antifungal activity against Candida albicans with MIC values of 12.5-25 μg/mL. The biosynthetic gene cluster of the isolated compounds and their putative biosynthetic pathway are also proposed. KEY POINTS • Ten meroterpenoids were identified from B. sorokiniana, including two new compounds. • Cochlioquinone B (5) showed activity against MRSA and synergistic activity against C. albicans. • The biosynthetic gene cluster and biosynthetic pathway of meroterpenoids are proposed. • Genome mining provided a new direction to uncover the diversity of meroterpenoids.The ability to rapidly detect viable pathogens in food is important for public health and food safety reasons. Culture-based detection methods, the traditional means of demonstrating microbial viability, tend to be laborious, time consuming and slow to provide results. Several culture-independent methods to detect viable pathogens have been reported in recent years, including both nucleic acid-based (PCR combined with use of cell viability dyes or reverse-transcriptase PCR to detect messenger RNA) and phage-based (plaque assay or phage amplification and lysis plus PCR/qPCR, immunoassay or enzymatic assay to detect host DNA, progeny phages or intracellular components) methods. Some of these newer methods, particularly phage-based methods, show promise in terms of speed, sensitivity of detection and cost compared with culture for food testing. This review provides an overview of these new approaches and their food testing applications, and discusses their current limitations and future prospects in relation to detection of viable pathogens in food. KEY POINTS • Cultural methods may be 'gold standard' for assessing viability of pathogens, but they are too slow. • Nucleic acid-based methods offer speed of detection but not consistently proof of cell viability. • Phage-based methods appear to offer best alternative to culture for detecting viable pathogens.Dicarboxylic acids are widely used in fine chemical and food industries as well as the monomer for polymerisation of high molecular material. Given the problems of environmental contamination and sustainable development faced by traditional production of dicarboxylic acids based on petrol, new approaches such as bio-based production of dicarboxylic acids drew more attentions. The yeast, Saccharomyces cerevisiae, was regarded as an ideal organism for bio-based production of dicarboxylic acids with high tolerance to acidic and hyperosmotic environments, robust growth using a broad range of substrates, great convenience for genetic manipulation, stable inheritance via sub-cultivation, and food compatibility. In this review, the production of major dicarboxylates via S. cerevisiae was concluded and the challenges and opportunities facing were discussed.Key Points• Summary of current production of major dicarboxylic acids by Saccharomyces cerevisiae.• Discussion of influence factors on four-carbon dicarboxylic acids production by Saccharomyces cerevisiae.• Outlook of potential production of five- and six-carbon dicarboxylic acids by Saccharomyces cerevisiae.This study evaluated the inactivation effect of ultrasonic treatment combined with acidic electrolyzed water (AEW) on Bacillus cereus spores. AEW treatment reduced the spores by 1.05-1.37 log CFU/mL while the sporicidal effect of ultrasound was minor. More strikingly, simultaneous ultrasonic and AEW treatments for 30 min led to 2.29 log CFU/mL reduction and thus, considered a synergistic effect. Flow cytometry combined with SYTO/PI staining analysis revealed that ultrasound hydrolyzed the cortex while the AEW partially damaged the integrity of the inner membrane. Scanning and transmission electron microscopies were used to characterize the ultrastructural changes. The detachment of the exosporium induced by ultrasound was the most apparent difference compared with the control group, and the electron density of spores appeared to be heterogeneous after treatment with AEW. These results indicated that combining ultrasound with AEW is a promising decontamination technology with potential uses in the food industry and environmental remediation.The methylotrophic bacterium Methylorubrum extorquens AM1 holds a great potential of a microbial cell factory in producing high value chemicals with methanol as the sole carbon and energy source. However, many gene functions remain unknown, hampering further rewiring of metabolic networks. Clustered regularly interspaced short palindromic repeat interference (CRISPRi) has been demonstrated to be a robust tool for gene knockdown in diverse organisms. In this study, we developed an efficient CRISPRi system through optimizing the promoter strength of Streptococcus pyogenes-derived deactivated cas9 (dcas9). When the dcas9 and sgRNA were respectively controlled by medium PR/tetO and strong PmxaF-g promoters, dynamic repression efficacy of cell growth through disturbing a central metabolism gene glyA was achieved from 41.9 to 96.6% dependent on the sgRNA targeting sites. Furthermore, the optimized CRISPRi system was shown to effectively decrease the abundance of exogenous fluorescent protein gene mCherry over 50% ae pathway.Leishmaniosis is caused by the protozoa of the genus Leishmania with a wide spectrum of clinical and epidemiological manifestations which are characterized into four clinical groups cutaneous, mucocutaneous, diffuse cutaneous, and visceral. American visceral leishmaniosis (AVL) or visceral leishmaniosis (VL) has been known as the most severe form of the disease. However, despite the growing number of people exposed to the infection risk and the great effort done by the scientific community worldwide to significantly increase the knowledge about these diseases, there is no vaccine capable of preventing VL in humans. In this short review, we present some of the plasmids used for the expression of recombinant protein by Escherichia coli strains used mainly for the second generation of vaccines for leishmaniosis. It can be emphasized that currently, these vectors and hosts play an important role in developing vaccine strategies against the disease. Indeed, use of the E. coli BL21 (DE) strain is remarkable mainly due to its characteristics for being a stable protein producer as well as the use of histidine tags for antigen purification. KEY POINTS • Plasmid vectors and E. coli will continue being important for studies about leishmaniosis. • Protein purification exploiting histidine tags is a key technique.Quorum sensing (QS) and signal molecules used for interspecies communication are well defined in mesophiles, but there is still a plethora of microorganisms in which existence and mechanisms of QS need to be explored, thermophiles being among them. In silico analysis has revealed the presence of autoinducer-2 (AI-2) class of QS signaling molecules in thermophiles, synthesized by LuxS (AI-2 synthase), though the functions of this system are not known. In this study, LuxS of Meiothermus ruber was used for understanding the mechanism and functions of AI-2 based QS among thermophilic bacteria. The luxS gene of M. ruber was expressed in luxS- deletion mutant of Escherichia coli. Complementation of luxS resulted in significant AI-2 activity, enhanced biofilm formation, and antibiotic susceptibility. Transcriptome analysis showed significant differential expression of 204 genes between the luxS-complemented and luxS- deletion mutant of E. coli. Majority of the genes regulated by luxS belonged to efflux pumps. This elucidation may contribute towards finding novel alternatives against incessant antibiotic resistance in bacteria.Key Points • Expression of luxS in luxS - E. coli resulted in increase in biofilm index. • Reduction in the MIC of antibiotics was observed after complementation of luxS. • Downregulation of efflux pump genes was observed after complementation of luxS. • Transcriptome analysis showed that 204 genes were differentially regulated significantly.Here, we used codon usage technology to generate two codon-modified human papillomavirus (HPV)16 E7 genes and, together with wild-type E7, to construct three HPV16 E7 gene plasmids Wt-E7, HB1-E7, and HB2-E7. The three HPV 16 E7 plasmids were used to investigate how HPV16 E7 protein was expressed in different cells and how this oncoprotein deregulated cellular and molecular events in human keratinocytes to induce carcinogenesis. We discovered that codon usage of HPV16 E7 gene played a key role in determining expression of E7 oncoprotein in all tested cells. HPV16 E7 inhibited significantly expression of pRb to impair keratinocyte differentiation and disrupted development of skin epidermis in mice. HPV16 E7 increased substantially the number of G0/G1 cells associated with upregulation of cyclin D2 and downregulation of cyclin B1 in keratinocytes. HPV16 E7 not only inhibited expression of involucrin and α-spectrin but also disrupted the organization of involucrin filaments and spectrin cytoskeleton. Furthermore, HPV16 E7 inhibited expression of β-adducin, destroyed its cytoskeletal structure and induced phosphorylation of β-adducin(Ser662) in keratinocytes. Importantly, HPV16 E7 induced carcinogenesis in mice associated with expression of phosphorylated β-adducin(Ser662) and its nucleus-translocation. In conclusion, we provided evidence that HPV16 E7 oncoprotein inhibited keratinocyte differentiation in vitro and in vivo leading to carcinogenesis through cell cycle arrest and disruption of pRb/involucrin/spectrin/adducin cascade.Ethyl carbamate (EC) is a potential carcinogen to humans that is mainly produced through the spontaneous reaction between urea and ethanol during Chinese rice wine brewing. We metabolically engineered a strain by over-expressing the DUR3 gene in a previously modified strain using an improved CRISPR/Cas9 system to further decrease the EC level. Homologous recombination of the DUR3 over-expression cassette was performed at the HO locus by individual transformation of the constructed plasmid CRISPR-DUR3-gBlock-HO, generating the engineered strain N85DUR1,2/DUR3-c. Consequently, the DUR3 expression level was significantly enhanced in the modified strain, resulting in increased utilization of urea. The brewing test showed that N85DUR1,2/DUR3-c reduced urea and EC concentrations by 92.0% and 58.5%, respectively, compared with those of the original N85 strain. Moreover, the engineered strain showed good genetic stability in reducing urea content during the repeated brewing experiments. this website Importantly, the genetic manipulation had a negligible effect on the growth and fermentation characteristics of the yeast strain.