Ryhára Dias Batista

Background/Objectives: Tuberculosis continues to be a significant global health concern, causing 1.3 million deaths in 2022, particularly affecting children under 5 years old. The Bacillus Calmette-Guérin (BCG) vaccine, developed in 1921, remains the primary defense against tuberculosis but requires modernized production methods. The recombinant BCG-pertussis strain shows potential in providing dual protection against tuberculosis and whooping cough, especially for vulnerable newborns, and enhanced efficacy against bladder cancer. Implementing submerged cultivation techniques for rBCG-pertussis production can offer increased productivity and standardization. Methods: This study explores a fed-batch cultivation strategy with pH-stat control to feed L-glutamic acid through the acid pump into 1 L bioreactor. Three pH values were evaluated for fed-batch and a simple batch without pH control was done for comparison. The viable cell concentration was compared before and after freeze-drying samples harvested during the cultures. Results: L-glutamic acid was identified as the preferred substrate for rBCG-pertussis. While the fed-batch strategy did not enhance the maximum specific growth rate compared to simple batch cultivation, it did improve the specific growth rate after day 4 in the pH 7.4-controlled fed-batch cultures, thereby reducing the cultivation time. Fed-batch cultures controlled at three pH levels exhibited lower optical density than the simple batch, although the viable cell counts were similar. Notably, samples harvested after day 8 from the simple batch cultures showed a reduction in CFU/mL after freeze-drying, whereas all fed-batch samples exhibited high recovery of viable cell counts post lyophilization. Conclusions: The additional glutamate supplied to the fed-batch cultures may have protected the cells during the lyophilization process.

Fructooligosaccharides are prebiotic sugars that are widely used in the production of functional foods, which can be produced enzymatically by the transfructosylation reaction of sucrose. This work aimed to optimize the production of an invertase with high transfructosylation activity from Aspergillus carbonarius PC-4 using pineapple crown as the inducer substrate and evaluate its biochemical properties. The culture medium was optimized using a Plackett–Burman experimental design and a central composite rotatable design, resulting in a maximum transfructosylation activity of 65.33 U/mL at 72 h of cultivation. The cultivation parameters were Yp/s = 1070.75 U/g and P_P = 2771.48 U/h, which showed an increase of 5.2-fold in the enzyme produced. The optimum temperature (50 °C) and pH (5.0) for the enzymatic activity were obtained by a CCR design. The enzyme showed a half-life of 60 min at 40 °C. In conclusion, the invertase produced from A. carbonarius PC-4 using agro-industrial waste (pineapple crown) and an inorganic nitrogen source (ammonium nitrate) exhibits high transfructosylation activity that can be used as a potential source for the production of fructooligosaccharides.

The aim of this work was to evaluate the use of different agro-industrial residues for the production of xylanase by Aspergillus sp. ART 100.1, as well as analyzing the biochemical properties of the enzyme. Agroindustrial residues malt bagasse, pineapple crown, açaí bagasse and soybean husk present in large quantities in the Tocantins region were used to evaluate the production of xylanase. Cultivation conditions for xylanase production were evaluated in submerged and solid-state cultivation. The highest production of xylanase in submerged cultivation was obtained using soybean husk residue (23.60 U/mL), while, for solid-state cultivation conditions, the highest production of xylanase was obtained with malt bagasse (110.00 U/g). The effect of additives to the culture medium was also evaluated, with the best result for the use of the xylose additive in the pineapple crown in solid-state cultivation. The enzyme produced in solid-state cultivation was characterized in terms of pH and temperature. The optimum activity pH was observed at 5.0 and for temperature at 55 °C. The xylanase was stable in a pH range between 4 and 5 and retained 50% of its activity at 45 °C after 110 minutes. The Aspergillus sp. ART 500.1 presents potential for the production of xylanase using agro-industrial residues, enabling the development of bioprocesses for the scaling of production.

Research background. Microbial β-fructofuranosidases are widely employed in food industry to produce inverted sugar or fructooligosaccharides. In this study, a newly isolated Aspergillus carbonarius PC-4 strain was used to optimize the β-fructofuranosidase production in a cost-effective process and the sucrose hydrolysis was evaluated to produce inverted sugars. Experimental approach. Optimization of nutritional components of culture medium was carried out using simplex lattice mixture design for 72 and 120 h at 28 °C. One-factor-at-a-time methodology was used to optimize the physicochemical parameters. Crude enzyme was used for sucrose hydrolysis at different concentrations. Results and conclusions. The optimized conditions of enzyme production were achieved from cultivations containing pineapple crown waste (1.3 %, m/V) and yeast extract (0.3 %, m/V) after 72 h with an enzyme activity of 9.4 U/mL, obtaining R²=91.85 %, R²adjusted=85.06 %, highest F-value (13.52) and low p-value (0.003). One-factor-at-a-time used for optimizing the physicochemical conditions showed optimum temperature (20 °C), pH (5.5), agitation (180 rpm) and time course (72 h) with a 3-fold increase of enzyme production. The invertase-induced sucrose hydrolysis showed the maximum yield (3.45 mmol of reducing sugars) using 10 % of initial sucrose concentration. Higher sucrose concentrations caused the inhibition of invertase activity, possibly due to the saturation of substrate or formation of sucrose aggregates, making it difficult for the enzyme to access sucrose molecules within the created clusters. Therefore, a cost-effective method was developed for the invertase production using agroindustrial waste and the produced enzyme can be used efficiently for inverted sugar production at high sucrose concentration. Novelty and scientific contribution. This study presents an efficient utilization of pineapple crown waste to produce invertase by a newly isolated Aspergillus carbonarius PC-4 strain. This enzyme exhibited a good potential for inverted sugar production at high initial sucrose concentration, which is interesting for industrial applications.

Although the productivity of common bean in Tocantins is economically favorable, it has been infected by various pathogens found in soil. Among the major diseases is the web blight and root rot caused by Rhizoctonia solani and collar rot caused by the fungus Sclerotium rolfsii . This study aimed to evaluate the fungitoxic activity of methanol extracts of eight plant species on the inhibition of mycelial growth of S. rolfsii and R. solani . The fungitoxic activities were carried out over the inhibition of mycelial growth by means in vitro assays. The extracts were applied in concentrations of 250, 500, 1000, 2500 e 5000 µg ml -1 in PDA culture medium. In bioassays, it was found the significant effect of plant, concentration and also their interaction on the antifungal activity of the extracts. However, some extracts showed no inhibition of mycelial growth of the pathogens studied. Among those who had higher inhibitions is the extract of Lantana trifolia , which inhibited the mycelial growth of S. rolfsii in all concentrations, being the same as 97% for the highest concentration. When the methanol extract of Piper amplum Kunth, inhibition of the highest concentration was 83% for S. rolfsii and 74% for R. solani . These results show the potential of methanolic extract of Lantana trifolia and Piper amplum Kunth in the control set of plant pathogens studied.