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Microbially produced 3-acyltetramic acids display a diverse range of biological activities. The pyreudiones are new members of this class that were isolated from bacteria of the genus Pseudomonas . Here, we performed a structure-activity relationship study and determined their mode of action. An efficient biomimetic synthesis was developed to synthesize pyreudione A. Pyreudiones and synthetic analogs thereof were tested for their amoebicidal, antibacterial, antiproliferative, and cytotoxic activities. The length of the alkyl side chain and the nature of the amino acid residues within the tetramic acid moiety strongly affected activity, in particular against mycobacteria. The mode of action was shown to correlate with the ability of pyreudiones to act as protonophores. Removal of the acidic proton by methylation of pyreudione A resulted in a loss of bioactivity.

Re-engineering biosynthetic assembly lines, including nonribosomal peptide synthetases (NRPS) and related megasynthase enzymes, is a powerful route to new antibiotics and other bioactive natural products that are too complex for chemical synthesis. However, engineering megasynthases is very challenging using current methods. Here, we describe how CRISPR-Cas9 gene editing can be exploited to rapidly engineer one of the most complex megasynthase assembly lines in nature, the 2.0 MDa NRPS enzymes that deliver the lipopeptide antibiotic enduracidin. Gene editing was used to exchange subdomains within the NRPS, altering substrate selectivity, leading to ten new lipopeptide variants in good yields. In contrast, attempts to engineer the same NRPS using a conventional homologous recombination-mediated gene knockout and complementation approach resulted in only traces of new enduracidin variants. In addition to exchanging subdomains within the enduracidin NRPS, subdomains from a range of NRPS enzymes of diverse bacterial origins were also successfully utilized., (© 2021. The Author(s).)

Purpose: To investigate the pre-soaking of hamstring grafts in topical vancomycin, in addition to IV prophylaxis, during anterior cruciate ligament (ACL) reconstruction to reduce the incidence of post-operative infection, and to describe an evidence-based diagnostic and treatment algorithm to facilitate early diagnosis and appropriate management of possible knee sepsis post-operatively after ACL reconstruction., Methods: This study is a controlled observational series comprising of 1585 individuals who underwent ACL reconstruction over a 13-year period. All surgeries were performed by a single surgeon at the same hospital. Group 1 consisted of 285 patients who received pre-operative IV antibiotics without topical graft pre-soaking. Group 2 consisted of 1300 individuals who received IV antibiotics and graft pre-soaking in a vancomycin solution of 5 mg/mL., Results: In group 1, a total of four patients suffered a post-operative joint infection (1.4 %). Three out of the four were culture positive for Staphylococcus species. The fourth was culture negative but was managed as an acute infection. Group 2 suffered no post-operative infections (0 %). Statistical analysis of the vancomycin pre-soak with IV antibiotics group, compared with IV antibiotics-alone group, revealed a significantly reduced post-operative infection rate using a Fisher's exact test (P = 0.0011) and Chi-square test with Yates' correction (P = 0.0003)., Conclusions: Pre-soaking of hamstrings grafts with topical vancomycin reduced the rate of post-operative infection when compared to IV antibiotics alone. This technique should be utilised by surgeons to reduce the overall incidence of knee sepsis post-ACL reconstruction., Level of Evidence: III.