Biology subjects

Sebastian, V.

Publications and source records attributed to Sebastian, V..

4 recordsLinked to original sources

bioRxiv2026

Integrated Continuous Biomanufacturing of Recombinant Adeno-Associated Virus

Recombinant adeno-associated virus (rAAV) manufacturing remains constrained by low yields, costly purification, and process complexity, limiting cost-effective access to gene therapy. Here, we demonstrate an integrated continuous biomanufacturing platform that transfers proven antibody-manufacturing technology to rAAV production by coupling perfusion to semi-continuous twin-column affinity capture (CaptureSMB). This architecture enables continuous harvest of extracellular rAAV without cell lysis, depth filtration, or endonuclease treatment, sustaining stable operation over five days. Relative to conventional batch processing, perfusion improved capsid or vector genome yield depending on the production process, while affinity capture alone reduced total DNA to batch-comparable levels, with no measurable loss in impurity clearance or potency. By consolidating multiple manual unit operations into one continuous workflow, the platform may reduce manual intervention and process footprint. Its modular design may generalize to other rAAV serotypes, inducible producer cell lines, and viral vector classes, offering a potential route toward more efficient gene therapy manufacturing.

bioengineering↗

bioRxiv2023

Enhancing Recovery from Gut Microbiome Dysbiosis and Alleviating DSS-Induced Colitis in Mice with a Consortium of Rare Short-Chain Fatty Acid-Producing Bacteria

The human gut microbiota is a complex community comprising hundreds of species, with a few present in high abundance and the vast majority in low abundance. The biological functions and effects of these low-abundant species on their hosts are not yet fully understood. In this study, we assembled a bacterial consortium (SC-4) consisting of B. paravirosa, C. comes, M. indica, and A. butyriciproducens, which are low-abundant, short-chain fatty acid (SCFA)-producing bacteria isolated from healthy human gut, and tested its effect on host health using germfree and human microbiota-associated colitis mouse models. The selection also favored these four bacteria being reduced in abundance in either Ulcerative Colitis (UC) or Crohns disease (CD) metagenome samples. Our findings demonstrate that SC-4 can colonize germ-free (GF) mice, increasing mucin thickness by activating MUC-1 and MUC-2 genes, thereby protecting GF mice from Dextran Sodium Sulfate (DSS)-induced colitis. Moreover, SC-4 aided in the recovery of human microbiota-associated mice from DSS-induced colitis, and intriguingly, its administration enhanced the alpha diversity of the gut microbiome, shifting the community composition closer to control levels. The results showed enhanced phenotypes across all measures when the mice were supplemented with Inulin as a dietary fiber source alongside SC-4 administration. We also showed a functional redundancy existing in the gut microbiome, resulting in the low abundant SCFA producers acting as a form of insurance, which in turn accelerates recovery from the dysbiotic state upon the administration of SC-4. SC-4 colonization also upregulated iNOS gene expression, further supporting its ability to produce an increasing number of goblet cells. Collectively, our results provide evidence that low-abundant SCFA-producing species in the gut may offer a novel therapeutic approach to IBD.

microbiology↗

bioRxiv2020

The extent of protein hydration dictates the preference for heterogeneous or homogeneous nucleation generating either parallel or antiparallel β-sheet α-synuclein aggregates

-Synuclein amyloid self-assembly is the hallmark of a number of neurodegenerative disorders, including Parkinsons disease, although there is still very limited understanding about the factors and mechanisms that trigger this process. Primary nucleation has been observed to be initiated in vitro at hydrophobic/hydrophilic interfaces by heterogeneous nucleation generating parallel {beta}-sheet aggregates, although no such interfaces have yet been identified in vivo. In this work, we have discovered that -synuclein can self-assemble into amyloid aggregates by homogeneous nucleation, without the need of an active surface, and with a preference for an antiparallel {beta}-sheet arrangement. This particular structure has been previously proposed to be distinctive of stable toxic oligomers and we here demonstrate that it indeed represents the most stable structure of the preferred amyloid pathway triggered by homogeneous nucleation under limited hydration conditions, including those encountered inside -synuclein droplets generated by liquid-liquid phase separation. In addition, our results highlight the key role that water plays not only in modulating the transition free energy of amyloid nucleation, and thus governing the initiation of the process, but also in dictating the type of preferred primary nucleation and the type of amyloid polymorph generated depending on the extent of protein hydration. These findings are particularly relevant in the context of in vivo -synuclein aggregation where the protein can encounter a variety of hydration conditions in different cellular microenvironments, including the vicinity of lipid membranes or the interior of membraneless compartments, which could lead to the formation of remarkably different amyloid polymorphs by either heterogeneous or homogeneous nucleation.

biophysics↗

bioRxiv2020

A simple RNA preparation method for SARS-CoV-2 detection by RT-qPCR

The technique RT-qPCR for viral RNA detection is the current worldwide strategy used for early detection of the novel coronavirus SARS-CoV-2. RNA extraction is a key pre-analytical step in RT-qPCR, often achieved using commercial kits. However, the magnitude of the COVID-19 pandemic is causing disruptions to the global supply chains used by many diagnostic laboratories to procure the commercial kits required for RNA extraction. Shortage in these essential reagents is even more acute in developing countries with no means to produce kits locally. We sought to find an alternative procedure to replace commercial kits using common reagents found in molecular biology laboratories. Here we report a method for RNA extraction that takes about 40 min to complete ten samples, and is not more laborious than current commercial RNA extraction kits. We demonstrate that this method can be used to process nasopharyngeal swab samples and yields RT-qPCR results comparable to those obtained with commercial kits. Most importantly, this procedure can be easily implemented in any molecular diagnostic laboratory. Frequent testing is crucial for individual patient management as well as for public health decision making in this pandemic. Implementation of this method could maintain crucial testing going despite commercial kit shortages.

molecular biology↗