Actinomycetes continue to be important sources for the discovery of secondary metabolites for applications in human medicine, animal health, and crop protection. With the maturation of actinomycete genome mining as a robust approach to identify new and novel cryptic secondary metabolite gene clusters, it is critical to continue developing methods to activate and enhance secondary metabolite biosynthesis for discovery, development, and large-scale manufacturing. This review covers recent reports on promising new approaches and further validations or technical improvements of existing approaches to strain improvement applicable to a wide range of Streptomyces species and other actinomycetes.

Genetic manipulation of secondary metabolite biosynthesis for improved production in Streptomyces and other actinomycetes

Engineering microbial hosts for production of bacterial natural products

Genetic platforms for heterologous expression of microbial natural products.

Direct Pathway Cloning (DiPaC) to unlock natural product biosynthetic potential.

The seamless ligation cloning extract (SLiCE) method is a novel seamless DNA cloning tool that utilizes homologous recombination activities in Escherichia coli cell lysates to assemble DNA fragments into a vector. Several laboratory E. coli strains can be used as a source for the SLiCE extract; therefore, the SLiCE-method is highly cost-effective.The SLiCE has sufficient cloning ability to support conventional DNA cloning, and can simultaneously incorporate two unpurified DNA fragments into vector. Recently, many seamless DNA cloning kits have become commercially available; these are generally very convenient, but expensive. In this study, we evaluated the cloning efficiencies between a simple and highly cost-effective SLiCE-method and a commercial kit under various molar ratios of insert DNA fragments to vector DNA. This assessment identified that the SLiCE from a laboratory E. coli strain yielded 30−85% of the colony formation rate of a commercially available seamless DNA cloning kit. The cloning efficiencies of both methods were highly effective, exhibiting over 80% success rate under all conditions examined. These results suggest that SLiCE from a laboratory E. coli strain can efficiently function as an effective alternative to commercially available seamless DNA cloning kits.

A simple and ultra-low cost homemade seamless ligation cloning extract (SLiCE) as an alternative to a commercially available seamless DNA cloning kit

A synbiotic multiparticulate microcapsule for enhancing inulin intestinal release and Bifidobacterium gastro-intestinal survivability.

Production of prebiotic chitooligosaccharide and its nano/microencapsulation for the production of functional yoghurt

Background: Integrating vectors based on the int/attP loci of temperate phages are convenient and used widely, particularly for cloning genes in Streptomyces spp. Results: We have constructed and tested a novel integrating vector based on g27, encoding integrase, and attP site from the phage, SV1. This plasmid, pBF3 integrates efficiently in S. coelicolor and S. lividans but surprisingly fails to generate stable integrants in S. venezuelae, the natural host for phage SV1. Conclusion: pBF3 promises to be a useful addition to the range of integrating vectors currently available for Streptomyces molecular genetics.

/pdf/a-novel-streptomyces-spp-integration-vector-derived-from-the-wmkab1u1d4.pdf

A novel Streptomyces spp. integration vector derived from the S. venezuelae phage, SV1

The potential application for the multiparticulate synbiotic microcapsule in yoghurt production was presented in the current study. Initially, three different formulae were fabricated to investigate the mechanism of action. This was followed by characterization the size of both of the nanoparticles and the final multiparticulate microcapsule. The yoghurt was then formed and the formulae were added and subjected to 4 weeks storage period. The viability of Bifidobacterium mixed cultures was monitored weekly. The results showed the viability of Bifidobacterium cultures was enhanced when double coated microcapsule was applied compared to the free one. The enhancement was further boosted when inulin was incorporated in either free form or in the form of nanoparticles. Scanning electron microscope image showed the final multiparticulate microcapsule has size of 700 μm while the laser diffraction particle size analyzer revealed the nanoparticles size is 118.1±4 nm. It was concluded that both of the double coat layer covering the multiparticulate microcapsule and the entrapped inulin, significantly enhanced the survivability of the encapsulated Bifidobacterium cultures in yoghurt during the shelf period.

The application of multi-particulate microcapsule containing probiotic bacteria and inulin nanoparticles in enhancing the probiotic survivability in yoghurt

Candida is the most common fungal class, causing both superficial and invasive diseases in humans. Although Candida albicans is the most common cause of fungal infections in humans, C. auris is a new emergent serious pathogen causing complications similar to those of C. albicans. Both C. albicans and C. auris are associated with high mortality rates, mainly because of their multidrug-resistance patterns against most available antifungal drugs. Although several compounds were designed against C. albicans, very few or none were tested on C. auris. Therefore, it is urgent to develop novel effective antifungal drugs that can accommodate not only C. albicans, but also other Candida spp., particularly newly emergent one, including C. auris. Inspired by the significant broad-spectrum antifungal activities of the essential oil cuminaldehyde and the reported wide incorporation of azoles in the antifungal drugs, a series of compounds (UoST1-11) was designed and developed. The new compounds were designed to overcome the toxicity of the aldehyde group of cuminaldehyde and benefit from the antifungal selectivity of azoles. The new developed UoST compounds showed significant anti-Candida activities against both Candida species. The best candidate compound, UoST5, was further formulated into polymeric nanoparticles (NPs). The new formula, UoST5-NPs, showed similar activities to the nanoparticles-free drug, while providing only 25% release after 24 h, maintainng prolonged activity up to 48 h and affording no toxicity. In conclusion, new azole formulations with significantly enhanced activities against C. albicans and C. auris, while maintaining prolonged action and no toxicities at lower concentrations, were developed.

https://www.mdpi.com/1420-3049/25/6/1463/pdf

Bahgat Fayed

Papers

A synbiotic multiparticulate microcapsule for enhancing inulin intestinal release and Bifidobacterium gastro-intestinal survivability.

Production of prebiotic chitooligosaccharide and its nano/microencapsulation for the production of functional yoghurt

A novel Streptomyces spp. integration vector derived from the S. venezuelae phage, SV1

The application of multi-particulate microcapsule containing probiotic bacteria and inulin nanoparticles in enhancing the probiotic survivability in yoghurt

Essential Oil-Based Design and Development of Novel Anti-Candida Azoles Formulation.