Unveiling the microbial dynamics in vermicomposting with coir pith as earthworm substrate.

This study explored the impact of incorporating coir pith, a byproduct of the coconut industry, into the vermicomposting substrate of Eudrilus eugeniae earthworms. The groups were compared based on their diets: cow manure only or cow manure mixed with varying amounts of coir pith. The aim was to assess the effects of coir pith on earthworm growth, mortality and the microbial community involved in vermicomposting. Earthworms fed with higher proportions of coir pith (70 % w/w) experienced reduced growth (0.81 g/worm) and increased mortality (24.67 %) after 5 weeks of vermicomposting. These effects were attributed to the high level of total phenolic content in the system. Coir pith required specific bacteria for digestion and detoxification, and excessive intake disrupted the earthworms' digestion, thus hindering nutrient absorption. The study also examined the microbial composition of the vermicast samples and identified variations based on the diet. Bacterial taxa involved in lignocellulose degradation, such as Bacteriodota, Azospirillum, Chitinophagaceae, Marinomonas and Pantoea, exhibited decreased abundances in treatments with coir pith. Conversely, the abundances of potentially harmful bacteria, such as Aeromonas, increased with higher coir pith inclusion levels. This pioneering investigation sheds light on the feasibility of coir pith use in vermicomposting and emphasises the importance of optimising earthworm diets to enhance microbial ecological functions and improve vermicompost quality.

Production of Neuraminidase Virus Like Particles by Stably Transformed Insect Cells: A Simple Process for NA-Based Influenza Vaccine Development.

Neuraminidase (NA) is a second major surface protein of the influenza virus and has recently been suggested as a supplemental antigen to the major immunodominant hemagglutinin (HA) antigen in the influenza vaccine. NA is less affected by antigenic drift compared to the HA, induces strong anti-neuraminidase immune responses, and provides broader protection against many influenza strains. However, the NA amount in currently licensed influenza virus vaccines is much lower than that of HA, and not standardized. A platform to produce NA antigen, in the form of virus-like particles (VLPs), was thus developed, to facilitate supplementation of NA antigen in the influenza vaccine formula. Stably transformed Sf9 insect cells had been engineered to express the influenza A virus (H5N1) NA gene under a baculovirus OpMNPV IE2 promoter. Recombinant NA protein was synthesized and assembled into VLPs, in the intact cellular environment provided by insect cells. Approximately 150 µg/ml of NA-VLPs was obtained in the culture medium. Purification of the NA-VLPs was achieved by a sucrose density gradient ultracentrifugation. The purified NA-VLPs effectively induced anti-NA antibodies with neuraminidase inhibition activities in mice. This work demonstrates a simple process to produce an immunocompetent NA-VLPs antigen, exclusively made of only neuraminidase, by insect cells.

Interplays of enzyme, substrate, and surfactant on hydrolysis of native lignocellulosic biomass.

Tracking enzyme, substrate, and surfactant interactions to reach maximum reducing sugar production during enzymatic hydrolysis of plant biomass may provide a better understanding of factors that limit the lignocellulosic material degradation in native rice straw. In this study, enzymes (Cellic Ctec2 cellulase and Cellic Htec2 xylanase) and Triton X-100 (surfactant) were used as biocatalysts for cellulose and xylan degradation and as a lignin blocking agent, respectively. The response surface model (R2 = 0.99 and R2-adj = 0.97) indicated that Cellic Ctec2 cellulase (p < 0.0001) had significant impacts on reducing sugar production, whereas Cellic Htec2 xylanase and Triton X-100 had insignificant impacts on sugar yield. Although FTIR analysis suggested binding of Triton X-100 to lignin surfaces, the morphological observation by SEM revealed similar surface features (i.e., smooth surfaces with some pores) of rice straw irrespective of Triton X-100. The reducing sugar yields from substrate hydrolysis with or without the surfactant were comparable, suggesting similar exposure of polysaccharides accessible to the enzymes. The model analysis and chemical and structural evidence suggest that there would be no positive effects on enzymatic hydrolysis by blocking lignins with Triton X-100 if high lignin coverage exists in the substrate due to the limited availability of hydrolyzable polysaccharides.

Recombinant neuraminidase pseudotyped baculovirus: a dual vector for delivery of Angiotensin II peptides and DNA vaccine.

Baculovirus is a promising vaccine deliver vector due to its biosafety profiles, gene transfer efficiency, ability to display small foreign antigens on its surface, strong adjuvant activities, etc. A dual vector for peptide antigens and a DNA vaccine delivery was constructed. In this vector, a tetrameric glycoprotein neuraminidase (NA) from influenza A virus (H5N1) serves as a baculovirus surface protein to improve baculovirus transduction efficiency and a partner for displaying the target peptide antigen. Nucleotides encoding target peptides could be fused to a full length NA gene, at the lower part of its head structure, integrated into Autographa californica multinucleopolyhedrovirus genome and expressed under the control of a White Spot Syndrome Virus IE-1 shuttle promoter. Angiotensin II (AngII) peptides, a potent vasoconstrictor that causes high blood pressure, was our target antigen. The recombinant NA-AngII pseudotyped baculovirus had the AngII peptides fused to the NA and displayed on its surface. In vitro studies revealed that this recombinant baculovirus successfully delivered AngII peptides, as DNA vaccine, into human HEK293A cells. A single subcutaneous injection of the recombinant NA-AngII pseudotyped baculovirus into moderately high blood pressure rats at 4 × 109 pfu/rat, stimulated anti-AngII antibody production and their systolic blood pressure (SBP) levels were found to have decreased. In addition, a single intranasal immunization at 8 × 108 pfu/rat, raised anti-AngII antibodies in a rat and its SBP was also reduced. The recombinant neuraminidase pseudotyped baculovirus is a potential vector for AngII peptide antigen and DNA vaccine for subcutaneous or intranasal immunization for treatment of hypertension.

Utilization of wastewater originated from naturally fermented virgin coconut oil manufacturing process for bioextract production: physico-chemical and microbial evolution.

Production of virgin coconut oil via natural fermentation has led to large amount of wastes being generated, i.e., coconut pulp and wastewater containing coconut crème. Objective of this study is to gain more insight into the feasibility of utilization of such wastes as raw materials together with several types of wastes such as fish waste and/or pineapple peel for bioextract production. Chemical, physico-chemical and biological changes including phytotoxicity of the fermented mixture were closely monitored. Physical observation suggested that fermentation of bioextract obtained with fish waste appeared to be complete within the first month of fermentation while bioextract obtained using pineapple waste seemed to be complete after 8 months post-fermentation. Fermentation broth is of blackish color with alcoholic as well as acidic odour with no gas bubble and/or yeast film present on top of the surface. During the whole fermentation interval, several attributes of both bioextracts, e.g., pH, chemical oxygen demand (COD) and organic acids, were statistically different. Further, the total bacteria and lactic acid bacteria present in pineapple bioextract were statistically higher than those of the fish bioextract (p<0.01). The highest germination indices of 123 and 106 were obtained at 21 and 14 days post-fermentation for fish and pineapple bioextracts, respectively. In addition, qualities of both bioextracts conformed well with those specified by the Thai standard for liquid biofertilizer after 1 month fermentation. Results further showed that wastewater derived from virgin coconut oil manufacturing process could effectively be employed together with other types of wastes such as fish waste and pineapple peel for bioextract production. However, for the best bioextract quality, fermentation should be carefully planned since over fermentation led to bioextract of low qualities.

Murine leukemia virus clearance by flocculation and microfiltration.

Clearance of murine leukemia virus from CHO cell suspensions by flocculation and microfiltration was investigated. Murine leukemia virus is a retrovirus that is recommended by the U.S. Food and Drug Administration for validating clearance of retrovirus-like particles. Due to biosafety considerations, an amphotropic murine leukemia virus vector (A-MLV) that is incapable of self-replication was used. Further, A-MLV is incapable of infecting CHO cells, thus ensuring that infection of the CHO cells in the feed did not result in a reduced virus titer in the permeate. The virus vector contains the gene for the enhanced green fluorescent protein (EGFP) to facilitate assaying for infectious virus particles. The virus particles are 80-130 nm in size. The feed streams were flocculated using a cationic polyelectrolyte. Microfiltration was conducted using 0.1 and 0.65 microm pore size hollow fiber membranes. The level of virus clearance in the permeate was determined. For the 0.1 microm pore size membranes a 1,000-fold reduction in the virus titer in the permeate was observed for feed streams consisting of A-MLV, A-MLV plus flocculant, A-MLV plus CHO cells, and A-MLV plus flocculant and CHO cells. While the flocculant had little effect on the level of virus clearance in the permeate for 0.1 microm pore size membranes, it did lead to higher permeate fluxes for the CHO cell feed streams. Virus clearance experiments conducted with 0.65 microm pore size membranes indicate little clearance of A-MLV from the permeate in the absence of flocculant. However, in the presence of flocculant the level of virus clearance in the permeate was similar to that observed for 0.1 microm pore size membranes. The results obtained here indicate that significant clearance of A-MLV is possible during tangential flow microfiltration. Addition of a flocculant is essential if the membrane pore size is greater than the diameter of the virus particles. Flocculation of the feed stream leads to an increase in the permeate flux.