Corrigendum: New perspectives on an old grouping: the genomic and phenotypic variability of Oxalobacter formigenes and the implications for calcium oxalate stone prevention.

[This corrects the article DOI: 10.3389/fmicb.2022.1011102.].

New perspectives on an old grouping: The genomic and phenotypic variability of Oxalobacter formigenes and the implications for calcium oxalate stone prevention.

Oxalobacter formigenes is a unique bacterium with the ability to metabolize oxalate as a primary carbon source. Most kidney stones in humans are composed of calcium and oxalate. Therefore, supplementation with an oxalate-degrading bacterium may reduce stone burden in patients suffering from recurrent calcium oxalate-based urolithiasis. Strains of O. formigenes are divided into two groups: group I and group II. However, the differences between strains from each group remain unclear and elucidating these distinctions will provide a better understanding of their physiology and potential clinical applications. Here, genomes from multiple O. formigenes strains underwent whole genome sequencing followed by phylogenetic and functional analyses. Genetic differences suggest that the O. formigenes taxon should be divided into an additional three species: Oxalobacter aliiformigenes sp. nov, Oxalobacter paeniformigenes sp. nov, and Oxalobacter paraformigenes sp. nov. Despite the similarities in the oxalyl-CoA gene (oxc), which is essential for oxalate degradation, these strains have multiple unique genetic features that may be potential exploited for clinical use. Further investigation into the growth of these strains in a simulated fecal environment revealed that O. aliiformigenes strains are capable of thriving within the human gut microbiota. O. aliiformigenes may be a better therapeutic candidate than current group I strains (retaining the name O. formigenes), which have been previously tested and shown to be ineffective as an oral supplement to mitigate stone disease. By performing genomic analyses and identifying these novel characteristics, Oxalobacter strains better suited to mitigation of calcium oxalate-based urolithiasis may be identified in the future.

Phosphate-regulated expression of the SARS-CoV-2 receptor-binding domain in the diatom Phaeodactylum tricornutum for pandemic diagnostics.

The worldwide COVID-19 pandemic caused by the SARS-CoV-2 betacoronavirus has highlighted the need for a synthetic biology approach to create reliable and scalable sources of viral antigen for uses in diagnostics, therapeutics and basic biomedical research. Here, we adapt plasmid-based systems in the eukaryotic microalgae Phaeodactylum tricornutum to develop an inducible overexpression system for SARS-CoV-2 proteins. Limiting phosphate and iron in growth media induced expression of the receptor-binding domain (RBD) of the SARS-CoV-2 spike protein from the P. tricornutum HASP1 promoter in the wild-type strain and in a histidine auxotrophic strain that alleviates the requirement for antibiotic selection of expression plasmids. The RBD was purified from whole cell extracts (algae-RBD) with yield compromised by the finding that 90-95% of expressed RBD lacked the genetically encoded C-terminal 6X-histidine tag. Constructs that lacked the TEV protease site between the RBD and C-terminal 6X-histidine tag retained the tag, increasing yield. Purified algae-RBD was found to be N-linked glycosylated by treatment with endoglycosidases, was cross-reactive with anti-RBD polyclonal antibodies, and inhibited binding of recombinant RBD purified from mammalian cell lines to the human ACE2 receptor. We also show that the algae-RBD can be used in a lateral flow assay device to detect SARS-CoV-2 specific IgG antibodies from donor serum at sensitivity equivalent to assays performed with RBD made in mammalian cell lines. Our study shows that P. tricornutum is a scalable system with minimal biocontainment requirements for the inducible production of SARS-CoV-2 or other coronavirus antigens for pandemic diagnostics.

Superior Conjugative Plasmids Delivered by Bacteria to Diverse Fungi.

Fungi are nature's recyclers, allowing for ecological nutrient cycling and, in turn, the continuation of life on Earth. Some fungi inhabit the human microbiome where they can provide health benefits, while others are opportunistic pathogens that can cause disease. Yeasts, members of the fungal kingdom, have been domesticated by humans for the production of beer, bread, and, recently, medicine and chemicals. Still, the great untapped potential exists within the diverse fungal kingdom. However, many yeasts are intractable, preventing their use in biotechnology or in the development of novel treatments for pathogenic fungi. Therefore, as a first step for the domestication of new fungi, an efficient DNA delivery method needs to be developed. Here, we report the creation of superior conjugative plasmids and demonstrate their transfer via conjugation from bacteria to 7 diverse yeast species including the emerging pathogen Candida auris. To create our superior plasmids, derivatives of the 57 kb conjugative plasmid pTA-Mob 2.0 were built using designed gene deletions and insertions, as well as some unintentional mutations. Specifically, a cluster mutation in the promoter of the conjugative gene traJ had the most significant effect on improving conjugation to yeasts. In addition, we created Golden Gate assembly-compatible plasmid derivatives that allow for the generation of custom plasmids to enable the rapid insertion of designer genetic cassettes. Finally, we demonstrated that designer conjugative plasmids harboring engineered restriction endonucleases can be used as a novel antifungal agent, with important applications for the development of next-generation antifungal therapeutics.