Caio Roberto Soares Bragança

Artisanal cheeses bearing the ARTE seal are authorized for nationwide commercialization and are produced in accordance with quality standards established by specific regulations. In contrast, cheeses without this certification have their sale restricted to municipal or state-level commerce, subject to registration with official inspection services. This study evaluated the microbiological quality of Minas artisanal cheeses produced in the Serra da Canastra region, Minas Gerais, by comparing products from farms with and without the ARTE seal. A total of 69 samples were analyzed, including raw milk, pasteurized milk, curd, cheese, and swabs from cheese molds, collected from 16 farms, eight certified with the ARTE seal and eight uncertified. Microbiological quantification techniques, culture methods, and polymerase chain reaction (PCR) assays were employed for pathogen identification, alongside phenotypic antimicrobial susceptibility testing and genetic similarity analysis of Staphylococcus aureus isolates via pulsed-field gel electrophoresis (PFGE). Neither Salmonella spp. nor Listeria monocytogenes were detected. Elevated counts of Enterobacteriaceae and coagulase-positive staphylococci were more frequently observed in ARTE-certified farms; however, Staphylococcus aureus was isolated exclusively from uncertified properties. These findings suggest that the ARTE seal may contribute to more effective sanitary practices and enhanced microbiological safety of artisanal cheeses.

The biotechnology industry plays a crucial role in the advancement of agronomy, contributing increasingly to each stage of its development. Microbial bioprospecting is a highly diverse field with significant agroindustrial potential, as nature provides a vast range of microorganisms suitable for such applications — including lipase-producing strains. As part of an initiative to establish a fungal collection at the Universidade do Estado de Minas Gerais (UEMG) – Passos Unit, 22 fungal isolates were obtained from various natural sources, 17 of which demonstrated lipase production. Fungal isolates with the highest lipolytic activity were derived from wet soil in calf paddocks, oral residues from calves post-nursing, and treated sewage sludge. This study introduces a standardized method using LED light for detecting lipolytic activity — a technique that to the best of our knowledge has not previously been applied to microorganisms. LED light presents a safer alternative to UV light, which can compromise sample integrity and pose health risks to users. This novel approach highlights the diversity of lipase-producing fungi from distinct environmental sources. The study aims to evaluate and qualitatively assess fungal enzymatic activity through this innovative methodology, facilitating the selection of strains with high enzymatic potential for agroindustrial applications.

Lectins are glycan-binding proteins involved in diverse biological processes and have gained attention for their potential applications in biotechnology and immunomodulation. BOL (Brassica oleracea lectin) is a unique ~34 kDa lectin isolated from Brassica oleracea var. botrytis, composed exclusively of TRAF-like domains, where TRAF stands for tumor necrosis factor receptor–associated factor. To overcome the limitations of plant-based extraction, we aimed to produce recombinant BOL in Escherichia coli. Various strains and expression vectors were tested under distinct induction conditions to optimize solubility and yield. While expression using pET28a was unsuccessful, GST-tagged BOL was efficiently expressed in E. coli BL21-R3-pRARE2(DE3) and purified using affinity chromatography. Functional assays demonstrated that the recombinant protein retained lectin activity, as evidenced by hemagglutination of goat erythrocytes. Protein identity was confirmed by MALDI-TOF/TOF mass spectrometry, with tryptic peptides matching the BOL lectin sequence in the National Center for Biotechnology Information (NCBI) database. Our findings highlight the importance of codon optimization, temperature modulation, and fusion tag selection for the successful expression of eukaryotic lectins in E. coli. This work provides a platform for future functional studies of BOL and supports its potential application in plant immunity and biomedical research.

The BOL lectin, a 34 kDa protein with a hemagglutination titer of 64 hemagglutination units (HU), was extracted from cauliflower (Brassica oleracea spp. botrytis L.), purified by affinity and ion exchange chromatography, and confirmed, in this study, by Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis (SDS-PAGE). The antibiofilm activity of BOL was evaluated at two concentrations (0.1 mg/mL and 1.0 mg/mL) against bacterial strains of importance to human health (Bacillus cereus ATCC 10876, Escherichia coli ATCC 25922, Staphylococcus aureus ATCC 29213, and Streptococcus agalactiae ATCC 12403). In addition to a biofilm formation assay, a pre-formed biofilm assay was conducted, with biofilm structure analyzed by Scanning Electron Microscopy (SEM). The antimicrobial potential of BOL was also investigated using the Minimum Inhibitory Concentration (MIC) assay in 96-well microplates. Among the tested bacterial strains, BOL exhibited activity against S. aureus at 1.0 mg/mL, interfering with both biofilm formation and disrupting pre-formed biofilms, which may be explained by a possible interaction between BOL and the components present in the biofilm matrix. However, no antibiofilm activity was observed against E. coli, B. cereus, or S. agalactiae, possibly due to differences in the composition of their biofilm matrices. Furthermore, BOL showed no detectable bactericidal or bacteriostatic activity in the antimicrobial assays. In conclusion, BOL lectin, at the tested concentrations, does not exhibit direct antimicrobial activity but effectively disrupts the extracellular matrix in S. aureus ATCC 29213.

Dengue is a mosquito-borne viral disease caused by the dengue virus (DENV), which affects millions worldwide. The envelope (E) protein, particularly domain III (EDIII), plays a critical role in viral entry and is a major target for diagnostic and vaccine development. In this study, we aimed to express and purify EDIII from DENV-3 using Komagataella phaffii as an expression system. The recombinant gene was codonoptimized, cloned into the pPICZαA vector, and integrated into the K. phaffii genome. Protein expression was induced with methanol and purified using Ni²⁺-affinity chromatography, yielding 209–309 mg/L. SDS-PAGE and Western blot confirmed protein expression. The recombinant antigen was evaluated in an indirect ELISA with 92 serum samples, demonstrating an AUC of 0.94 (IgM) and 0.828 (IgG), with 92% specificity for IgM. These results suggest that K. phaffii is a suitable host for recombinant dengue antigen production, offering a scalable and cost-effective strategy for diagnostic applications.

Efficient co-fermentation of glucose and xylose remains a critical hurdle in second-generation bioethanol production. In this study, we evaluated two non-Saccharomyces yeasts—Wickerhamomyces anomalus UEMG-LF-Y2 and Diutina rugosa UEMG-LF-Y4—under mixed-sugar conditions. D. rugosa exhibited superior xylose metabolism and ethanol productivity, achieving a maximum volumetric productivity (Q_P) of 0.55 g/L·h in a medium containing 20 g/L glucose and 40 g/L xylose. Its highest ethanol yield (Y_P/S) reached 0.45 g EtOH/g sugar, comparable to results from engineered Saccharomyces cerevisiae strains. By contrast, W. anomalus displayed lower ethanol yields (0.24–0.34 g/g) and greater sensitivity to catabolite repression induced by 2-deoxyglucose (2-DG). Xylose consumption by D. rugosa exceeded 80% in high-xylose media, while W. anomalus left residual xylose under all tested conditions. A strong inverse correlation (r < −0.98) between ethanol accumulation and xylose uptake was observed, especially for W. anomalus, indicating ethanol-induced inhibition as a key challenge. These findings highlight the potential of D. rugosa as a robust non-Saccharomyces platform for lignocellulosic bioethanol processes, whereas W. anomalus may benefit from further metabolic or process optimizations. Future research should address ethanol tolerance, inhibitory byproducts, and large-scale feasibility to fully exploit these strains for second-generation bioethanol production.

Pre- and post-milking teat antisepsis is one of the most effective methods for reducing the incidence of mastitis. Plant extracts have proven effective in reducing microorganisms, providing a natural alternative for antisepsis, along with additional benefits for teat health. This study aimed to evaluate the antimicrobial and antiseptic effects of two newly formulated products, used as pre-dipping and post-dipping agents, respectively, based on plant extracts from papain (Carica papaya), aloe vera (Aloe barbadensis), andiroba (Carapa guianensis), copaiba (Copaifera officinalis), tea tree (Melaleuca alternifolia), and barbatimão (Stryphnodendron barbatiman). Minimum inhibitory concentration tests were performed in vitro, along with microbiological analyses of different bacterial groups and in vivo compliance assessments to evaluate the antimicrobial activity of the formulations, as well as the molecular identification of Staphylococcus aureus. The results demonstrated that the products were effective in vitro against the main pathogens that cause mastitis, including Staphylococcus aureus, Streptococcus agalactiae, and Escherichia coli. In the in vivo assays, similar performance was observed between the tested products and the control products, indicating that the plant-extract-based formulations, applied as pre-dipping and post-dipping agents, effectively reduced the microbial load on the teats. The effects were equivalent to those of hydrogen-peroxide- and iodine-based products, being able to reduce the total microbial load by more than 80%.

With the increase in pollution and improper waste disposal, aquatic ecosystems are experiencing escalating degradation leading to various detrimental effects, including eutrophication and adverse impacts on the health of the population reliant on these water resources. Consequently, microalgae have demonstrated efficacy in nutrient removal, minimal environmental disruption, and superior cost-effectiveness in comparison to traditional treatment methods. Thus, this study aimed to investigate wastewater treatment in an aerobic batch system, using two strains of non-axenic mixotrophic chlorophytes, Chlorella sp. and Desmodesmus sp., across distinct light regimes: continuous light exposure for 24 h, a photoperiod of 12 h light and 12 h darkness, and complete absence of light for 24 h. The Desmodesmus sp. strain exhibited superior efficiency in the proposed biological treatment, yielding more favorable nutrient removal results across all conditions, except for total nitrogen removal under the 24-h continuous light condition in which Chlorella sp. removed 0.199 ± 0.02% by biomass. In other parameters, Desmodesmus sp., remediated by biomass 0.408 ± 0.013% of inorganic phosphorus in 24 h light, 0.372 ± 0.011% of COD and 0.416 ± 0.004% of carbohydrate in 24 h dark. While Chlorella sp. removed 0.221 ± 0.01% of inorganic phosphorus in 24 h light, 0.164 ± 0.02% of COD in 24 h light and 0.214 ± 0.002% of carbohydrates in 24 h dark. Nevertheless, both strains displayed potential as viable alternatives for wastewater biological treatment, indicating that nutrient removal is achievable across all tested light conditions, albeit with variations in efficiency depending on the specific nutrient type.