BioTech

Acidogenic fermentation is a promising biotechnology for converting organic wastes into carboxylic acid (CA), which has significant commercial value and diverse applications in the food, chemical, pharmaceutical, and cosmetic industries. However, major challenges such as limited substrate hydrolysis and lower CA production hinder further development of this biotechnology towards full-scale implementation. This review provides a comprehensive overview of the current status of acidogenic fermentation, focusing on substrate composition, inoculum, and reactor design, along with potential strategies to overcome reactor-specific limitations and enhance CA production. It was found that the substrate composition, particularly its carbohydrate, protein, and lipid contents, strongly influences both CA production and yield. Specifically, carbohydrate-rich substrates yield higher CA production compared to protein- and lipid-rich substrates. These substrates have been investigated in different reactor configurations for CA production. Among them, the leachate bed reactor and anaerobic membrane bioreactor have demonstrated superior performance, achieving higher CA production with acetic and butyric acids as the dominant CA composition. These reactors are generally operated using three types of inocula: aerobic and anaerobic inoculum, enriched inoculum, and rumen microorganisms. Interestingly, rumen microorganisms are effective in degrading complex substrates, whereas enriched inoculum accelerates hydrolysis and acidogenesis processes within a shorter fermentation time. The findings presented herein will provide valuable information for addressing the challenges associated with acidogenic fermentation and lay the foundation for future research aimed at upscaling this biotechnology to a commercial scale.

Carotenoid-based pigmentation is crucial for the ornamental and commercial value of the cherry shrimp (Neocaridina denticulata sinensis). While several genes are known to influence carotenoid metabolism, the genetic basis for specific color strains remains largely unexplored. Here, we functionally characterized NinaB-like, a homolog of a carotenoid oxygenase, in cherry shrimp pigmentation. We employed qPCR to gain gene expression profiles, utilized RNAi technology to analysize the relation between its expression level and carotenoid accumulation, and performed GT-seq to identify genotypes of different color strains. Significant differential expression of NinaB-like was observed not only across distinct color strains but also during embryonic development of cherry shrimp (p < 0.05), peaking at the red strain and post-larval stage of cherry shrimp. RNA interference-mediated knockdown of NinaB-like resulted in a marked increase in red pigment deposition at the metanauplius and pre-zoea stages, confirming its role as a negative regulator of carotenoid accumulation. Importantly, we identified two tightly linked, non-synonymous SNPs (927C > A and 935A > C) within the NinaB-like coding region that exhibited a strong association with body color. Our study provides the first functional evidence that NinaB-like is a negative regulator of carotenoid degradation and a major genetic determinant for body color in cherry shrimp, providing new insights for genetic breeding and biological research.

Human embryonic kidney (HEK293) cells are a widespread choice for recombinant protein expression. To optimise yields, the hydrolysate Tryptone N1 (TN1) is commonly added post-transfection. TN1 is obtained by controlled enzymatic digestion of casein. As an animal by-product, TN1 faces stricter regulations during cross-country shipments than plant-based products. This raises the question of whether plant-derived peptides are a suitable alternative to TN1. Using polyethyleneimine (PEI) as a cationic polymer, we transfected HEK293-6E cells grown in suspension in serum-free medium and divided the transfectants into four groups (each in triplicate). Two plant-based hydrolysates each derived from pea and broad bean were compared with TN1 and a no-hydrolysate control group. We monitored the cultures for total cell numbers and viability at days 1, 4, and 5 post-transfection. Both plant-based hydrolysates and TN1 showed similar live cell percentages, in contrast to the no-hydrolysate control, which showed lower viability. Five days post-transfection, the expressed His-tagged protein, a tegumental antigen from the eukaryotic parasite Echinococcus granulosus, was retrieved from the serum-free culture supernatant, and the expressed recombinant protein was quantified. The linear ranges for the protein load on the stain-free blot and for the use of the fluorescent anti-His-Tag Alexa₄₈₈ antibody were determined. Using these parameters, stain-free Western blotting and total protein normalization were performed. The plant-derived pea and broad bean hydrolysates reproducibly resulted in similar expression levels as animal-derived TN1; all three hydrolysates were better than no hydrolysate. We conclude that plant-derived hydrolysates are a suitable, more sustainable replacement for TN1.