Synthesis, biological evaluation and computational studies of pyrazole derivatives as Mycobacterium tuberculosis CYP121A1 inhibitors.

A series of imidazole and triazole diarylpyrazole derivatives were prepared using an efficient 5-step synthetic scheme and evaluated for binding affinity with Mycobacterium tuberculosis (Mtb) CYP121A1 and antimycobacterial activity against Mtb H37Rv. Antimycobacterial susceptibility was measured using the spot-culture growth inhibition assay (SPOTi): the imidazoles displayed minimum inhibitory concentration (MIC90) in the range of 3.95-12.03 µg mL-1 (10.07-33.19 µM) with 11f the most active, while the triazoles displayed MIC90 in the range of 4.35-25.63 µg mL-1 (11.88-70.53 µM) with 12b the most active. Assessment of binding affinity using UV-vis spectroscopy showed that for the imidazole series, the propyloxy (11f) and isopropyloxy (11h) derivatives of the 4-chloroaryl pyrazoles displayed Mtb CYP121A1 type II binding affinity with K d 11.73 and 17.72 µM respectively compared with the natural substrate cYY (K d 12.28 µM), while in the triazole series, only the methoxy substitution with the 4-chloroaryl pyrazole (12b) showed good type II Mtb CYP121A1 binding affinity (K d 5.13 µM). Protein-detected 1D 19F-NMR spectroscopy as an orthogonal strategy was used to evaluate ligand binding independent of perturbations at the haem. For imidazole and triazole compounds, perturbations were more intense than cYY indicating tighter binding and confirming that ligand coordination occurs in the substrate-binding pocket despite very modest changes in UV-vis absorbance, consistent with computational studies and the demonstrated potential anti-tuberculosis properties of these compounds.

Engineering a suite of E. coli strains for enhanced expression of bacterial polysaccharides and glycoconjugate vaccines.

BACKGROUND: Glycoengineering, in the biotechnology workhorse bacterium, Escherichia coli, is a rapidly evolving field, particularly for the production of glycoconjugate vaccine candidates (bioconjugation). Efficient production of glycoconjugates requires the coordinated expression within the bacterial cell of three components: a carrier protein, a glycan antigen and a coupling enzyme, in a timely fashion. Thus, the choice of a suitable E. coli host cell is of paramount importance. Microbial chassis engineering has long been used to improve yields of chemicals and biopolymers, but its application to vaccine production is sparse. RESULTS: In this study we have engineered a family of 11 E. coli strains by the removal and/or addition of components rationally selected for enhanced expression of Streptococcus pneumoniae capsular polysaccharides with the scope of increasing yield of pneumococcal conjugate vaccines. Importantly, all strains express a detoxified version of endotoxin, a concerning contaminant of therapeutics produced in bacterial cells. The genomic background of each strain was altered using CRISPR in an iterative fashion to generate strains without antibiotic markers or scar sequences. CONCLUSIONS: Amongst the 11 modified strains generated in this study, E. coli Falcon, Peregrine and Sparrowhawk all showed increased production of S. pneumoniae serotype 4 capsule. Eagle (a strain without enterobacterial common antigen, containing a GalNAc epimerase and PglB expressed from the chromosome) and Sparrowhawk (a strain without enterobacterial common antigen, O-antigen ligase and chain length determinant, containing a GalNAc epimerase and chain length regulators from Streptococcus pneumoniae) respectively produced an AcrA-SP4 conjugate with 4 × and 14 × more glycan than that produced in the base strain, W3110. Beyond their application to the production of pneumococcal vaccine candidates, the bank of 11 new strains will be an invaluable resource for the glycoengineering community.

Antimicrobial resistance and COVID-19: Intersections and implications.

Before the coronavirus 2019 (COVID-19) pandemic began, antimicrobial resistance (AMR) was among the top priorities for global public health. Already a complex challenge, AMR now needs to be addressed in a changing healthcare landscape. Here, we analyse how changes due to COVID-19 in terms of antimicrobial usage, infection prevention, and health systems affect the emergence, transmission, and burden of AMR. Increased hand hygiene, decreased international travel, and decreased elective hospital procedures may reduce AMR pathogen selection and spread in the short term. However, the opposite effects may be seen if antibiotics are more widely used as standard healthcare pathways break down. Over 6 months into the COVID-19 pandemic, the dynamics of AMR remain uncertain. We call for the AMR community to keep a global perspective while designing finely tuned surveillance and research to continue to improve our preparedness and response to these intersecting public health challenges.

The importance of the glycosylation of antimicrobial peptides: natural and synthetic approaches.

Glycosylation is one of the most prevalent post-translational modifications of a protein, with a defining impact on its structure and function. Many of the proteins involved in the innate or adaptive immune response, including cytokines, chemokines, and antimicrobial peptides (AMPs), are glycosylated, contributing to their myriad activities. The current availability of synthetic coupling and glycoengineering technology makes it possible to customise the most beneficial glycan modifications for improved AMP stability, microbicidal potency, pathogen specificity, tissue or cell targeting, and immunomodulation.

Intracellular replication of the well-armed pathogen Burkholderia pseudomallei.

The Burkholderia genus contains a group of soil-dwelling Gram-negative organisms that are prevalent in warm and humid climates. Two species in particular are able to cause disease in animals, B. mallei primarily infects Equus spp. and B. pseudomallei (BPS), that is able to cause potentially life-threatening disease in humans. BPS is naturally resistant to many antibiotics and there is no vaccine available. Although not a specialised human pathogen, BPS possesses a large genome and many virulence traits that allow it to adapt and survive very successfully in the human host. Key to this survival is the ability of BPS to replicate intracellularly. In this review we highlight recent advances in our understanding of the intracellular survival of BPS, including how it overcomes host immune defenses and other challenges to establish its niche and then spread the infection. Knowledge of these mechanisms increases our capacity for therapeutic interventions against a well-armed foe.