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The severe acute respiratory syndrome originated by the new coronavirus (SARS-CoV-2) that emerged in late 2019, known to be a highly transmissible and pathogenic disease, has caused the COVID-19 global pandemic outbreak. Thus, diagnostic devices that help epidemiological public safety measures to reduce undetected cases and isolation of infected patients, in addition to significantly help to control the population’s immune response to vaccine, are required. To address the negative issues of clinical research, we developed a Diagnostic on a Chip platform based on a disposable electrochemical biosensor containing laser-induced graphene and a protein (SARS-CoV-2 specific antigen) for the detection of SARS-CoV-2 antibodies. The biosensors were produced via direct laser writing using a CO2 infrared laser cutting machine on commercial polyimide sheets. The presence of specific antibodies reacting with the protein and the K3[Fe(CN)6] redox indicator produced characteristic and concentration-dependent electrochemical signals, with mean current values of 9.6757 and 8.1812 µA for reactive and non-reactive samples, respectively, proving the effectiveness of testing in clinical samples of serum from patients. Thus, the platform is being expanded to be measured in a portable microcontrolled potentiostat to be applied as a fast and reliable monitoring and mapping tool, aiming to assess the vaccinal immune response of the population.

Plants have been studied as bioreactors for over 30 years. Their advantages over other expression methods arouse interest in this heterologous protein expression system. Over the years, several methods of plant transformation were developed, and among them, the transient expression system - that occurs temporarily and without integration into the host genome. The use of viral vectors for this type of expression stands out, and the viral genome can be “full-length” or “deconstructed”. Some genome modifications in the “deconstructed” form help searching for efficient protein expression, and several types of viruses are studied to develop these vectors. This review reports different types of viral vectors, especially those particularly suitable for the development of effective vectors of fast and high yield recombinant protein production, based on Tobamoviruses, Potexviruses, Geminiviruses and Comoviruses.

Endoglucanases are carbohydrate-degrading enzymes widely used for bioethanol production as part of the enzymatic cocktail. However, family 5 subfamily 5 (GH5_5) endoglucanases are still poorly explored in depth. The Trichoderma reesei representative is the most studied enzyme, presenting catalytic activity in acidic media and mild temperature conditions. Though biochemically similar, its modular structure and synergy with other components vary greatly compared to other GH5_5 members and there is still a lack of specific studies regarding their interaction with other cellulases and application on novel and better mixtures. In this regard, the threedimensional structure elucidation is a highly valuable tool to both uncover basic catalytic mechanisms and implement engineering techniques, proved by the high success rate GH5_5 endoglucanases show. GH5_5 enzymes must be carefully evaluated to fully uncover their potential in biomass-degrading cocktails: the optimal industrial conditions, synergy with other cellulases, structural studies, and enzyme engineering approaches. We aimed to provide the current understanding of these main topics, collecting all available information about characterized GH5_5 endoglucanases function, structure, and bench experiments, in order to suggest future directions to a better application of these enzymes in the industry.

Background: In December 2019, China announced the first case of an infection caused by an, until then, unknown virus: SARS-CoV-2. Since then, researchers have been looking for viable alternatives for the treatment and/or cure of viral infection. Among the possible complementary solutions are lectins, proteins that are reversibly bound to different carbohydrates. The Spike protein, present on the viral surface, can interact with different cell receptors: ACE2, CD147, and DC-SIGNR. Since lectins have an affinity for different carbohydrates, the binding with the glycosylated cell receptors represents a possibility of preventing the virus from binding to the receptors of host cells. Objective: In this review, we discuss the main lectins that are possible candidates for use in the treatment of Covid-19, highlighting those that have already demonstrated antiviral activity in vivo and in vitro, including mannose-binding lectin, Griffithsin, BanLec, and others. We also aim to discuss the possible mechanism of action of lectins, which appears to occur through the mediation of viral fusion in host cells, by binding of lectins to glycosylated receptors found in human cells and/or binding of these proteins with the spike glycoprotein, present in virus surface. Moreover, we discuss the use of lectins in clinical practice. Conclusion: Even with the development of effective vaccines, new cases of viral infection with the same virus, or new outbreaks with different viruses can occur; so, the development of new treatments should not be discarded. Moreover, the discussions made in this work are relevant regarding the anti-viral properties of lectins.

Background: Lipid nanocarriers have been widely tested as drug delivery systems to treat diseases due to their bioavailability, controlled release, and low toxicity. For the pulmonary route, the Food and Drug Administration favors the use of substances generally recognized as safe, as well as biodegradable and biocompatible to minimize the possibility of toxicity. Tuberculosis (TB) remains a public health threat worldwide, mainly due to the long treatment duration and adverse effects. Therefore, new drug delivery systems for treating TB are needed. Objective: Physicochemical characterization of different lipid-based nanocarriers was used to optimize carrier properties. Optimized systems were incubated with Mycobacterium tuberculosis to assess whether lipid-based systems act as the energy source for the bacteria, which could be counterproductive to therapy. Method: Several excipients and surfactants were evaluated to prepare different types of nanocarriers using high-pressure homogenization. Results: A mixture of trimyristin with castor oil was chosen as the lipid matrix after differential scanning calorimetry analysis. A mixture of egg lecithin and PEG-660 stearate was selected as an optimal surfactant system, as this mixture formed the most stable formulations. Three types of lipid nanocarriers, solid lipid nanoparticles, nanostructured lipid carriers (NLC), and nanoemulsions, were prepared, with the NLC systems showing the most suitable properties for further evaluation. It may provide the advantages of increasing the entrapment efficiency, drug release, and the ability to be lyophilized, producing powder for pulmonary administration as an alternative to entrap poor water-soluble molecules. Conclusion: Furthermore, the NLC system can be considered for use as a platform for the treatment of TB through the pulmonary route.

In this review, our analyses of the metabolic routes of limonoids and quassinoids—with respect to their occurrence in the families Rutaceae, Meliaceae, and Simaroubaceae—suggest a disjunction between the Simaroubaceae (characterized by quassinoids) and the Rutaceae and Meliaceae (distinguished by limonoids). However, even with the latter two families, clear-cut chemical differences could be discerned. Meliaceae exploit different biogenetic pathways, which lead to more diverse limonoids than in Rutaceae. The complex limonoids and prenylated chromones isolated from Spathelia, Sohnreyia, Dictyoloma, Harrisonia, Cneorum, and Cedrelopsis provided firm support for including them into the Cneoroideae. Ptaeroxylon produces several types of chromones, justifying to include it near the allied taxa, also into Cneoroideae. However, considering the complex limonoids found in Flindersia and Hortia, their connections to the Rutoideae subfamily were thus rather difficult to envisage; they suggest a closer relationship with the Cneoroideae genera. Until recently, there was no chemical evidence to support the placement of Picramnia into Simaroubaceae or near the allied taxa Rutaceae and Meliaceae. However, our investigations revealed several nortriterpenes involving expansion of the A-ring to a 4,4-dimethyl-tetrahydro-2H-furo[1,10]pyran-3(6H)-one as in limonin-type limonoids, whose derivatives have been isolated from Rutaceae genera. In addition, liquid chromatography-tandem mass spectrometry-(MS/MS)-based molecular networking showed profiling of quassinoids from P. glazioviana. These data convincingly point to affinities between Picramnia species and Cneoroideae genera, suggesting Picramniaceae is sister to Rutaceae, Simaroubaceae, and Meliaceae.

Cochlospermum regium is well-known as "Algodãozinho do cerrado" in folk Brazilian medicine, and is used to fight infections, inflammation and skin disorders.To identify the phytochemical constituents and the effects of the ethanolic extract of C. regium leaves (EECR) on inflammation and pain, and the effects of C. regium gel (GEECR) on wound healing.Animals were treated with EECR (30-300 mg/kg) or GEECR (1.25 and 2.5%) and studies were conducted using carrageenan-induced pleurisy and paw edema tests, formalin-induced pain model, and excision wound model.In total, 25 compounds, including quercitrin, methyl gallate, and 1,2,3,4,6-pentagalloylhexose, with highest detectability were identified. The treatments reduced leukocyte migration, nitric oxide production, protein extravasation, edema, mechanical hyperalgesia, pain in both phases (neurogenic and inflammatory), cold hypersensitivity, and improved wound closure and tissue regeneration.The present findings established the anti-inflammatory, anti-nociceptive, and wound healing potential of the leaves of C. regium, confirming the potential therapeutic effect of this plant.

Bartonella henselae is a Gram-negative bacterium that causes cat scratch disease (CSD), as well as bacteremia, endocarditis, and other clinical presentations. CSD remains one of the most common infections caused by bacteria in the genus Bartonella, and it is transmitted to humans through a scratch or cat bite. Vaccination and more efficient diagnostic methods would represent a promising and sustainable alternative measure for CSD control in humans and animals. Here, we described the in silico analyses and design of three recombinant chimeric proteins (rC1, rC2, and rC3), for use in the control of CSD. The chimeras were constructed with epitopes identified from the sequences of the GroEL, 17 kDa, P26, BadA, Pap31, OMP 89, and OMP 43, previously described as the most important B. henselae antigens. The rC1, rC2, and rC3 were expressed and purified using a heterologous system based on Escherichia coli and reacted with antibodies present in the sera of humans naturally infected. The chimeric proteins were used to immunize mice using Freund adjuvant, and the humoral immune response was evaluated. Animals immunized with rC1 and rC3 showed a significant IgG antibodies response from the 28th day (P 0.05), and the animals immunized with the rC2 from the 35th day (P 0.05) remained until the 56th day of experimentation, with a titer of 1:3200 (P 0.05), 1:1600 (P 0.05) and 1:1600 (P 0.05) from rC1, rC2, and rC3, respectively. Significant production of IgA and IgG1 isotype was detected in animals immunized with rC1 and rC2 proteins. Additionally, analysis using 13 serum samples from naturally infected patients showed that the proteins are recognized by antibodies present in sera, reinforcing the possibility of using these chimeras for CSD control. KEY POINTS: • The recombinant chimeras were expressed in Escherichia coli with 37 kDa (rC1), 35 kDa (rC2), and 38 kDa (rC3). • Animals immunized with rC1, rC2, and rC3 showed significant antibody response. • The chimeras were recognized by the sera of naturally infected patients.

Bovine viral diarrhea virus (BVDV) and bovine alphaherpesvirus type 1 (BoHV-1) are responsible for major economic losses of livestock worldwide, making their eradication an important objective of veterinary research. Vaccines against these infectious agents are commercially available but have some limitations due to the specific features of these viral agents. The development of new antiviral drugs is therefore essential. Native banana lectin (BanLec) is a lectin isolated from banana fruit (Musa acuminata) and has a high affinity for mannose glycans found in several viral envelopes. The inhibitory properties of this lectin against several viruses has already been demonstrated. The aim of this work was therefore to test the antiviral and virucidal activities of BanLec against BVDV-1 and BoHV-1. Its antiviral activity was assessed by measuring the viral titer and viability of susceptible Madin-Darby Bovine Kidney cells (MDBK) treated with BanLec before and after viral infection. The virucidal properties of BanLec were determined by preincubation of the lectin with the viruses, followed by measurement of the viral load in exposed cells. Treatment with 25 μg/mL BanLec resulted in high levels of inhibition against BVDV-1 (99.98%) and BoHV-1 (99.68%) without affecting cell viability, demonstrating promising potential as an antiviral agent.

Aristolochia triangularis Cham. has been used in Brazilian traditional medicine for various therapeutic purposes, including as a leaf-based infusion for diabetes management.This study was designed to chemically characterize an infusion of in natura A. triangularis leaves and evaluate the in vivo anti-hyperglycemic properties of this infusion.Chemical composition was examined using liquid-liquid extraction procedure, chromatographic methods, NMR, and LC-MS/MS. The in vivo anti-hyperglycemic activity of the freeze-dried infusion of A. triangularis leaves (Inf-L-At) was assessed using oral glucose tolerance test (OGTT). Initially, normoglycemic male rats were pre-treated with orally administered Inf-L-At at doses of 62.5, 125, and 250 mg/kg for two consecutive days. On the day of the OGTT, fasting animals received a glucose load (4 g/kg) 30 min after treatment with Inf-L-At, and the blood glucose levels were verified at 15, 30, 60, and 180 min. Intestinal maltase, lactase, and sucrase activities and muscle and liver glycogen contents were also assessed after the OGTT.Inf-L-At extract led to glycemic reduction with no dose-response at 15, 30, and 60 min comparable to that of the antidiabetic drug glibenclamide and was accompanied by an increase in hepatic and muscle glycogen contents. Additionally, there was a significant statistically decrease in the in vitro activity of disaccharidases. Maltase and sucrase activities were inhibited at all doses, whereas lactase activity was inhibited only at 62.5 and 250 mg/kg. In total, 75 compounds were found in the infusion, including seven new ones, (7S*,8S*,7ꞌS*,8ꞌR*)-4,4ꞌ-dihydroxy-3,3ꞌ-dimethoxy-7,9ꞌ-epoxylignan-7ꞌ-ol; 4ꞌ-hydroxy-3ꞌ-methoxy-3,4-methylenedioxy-7,9ꞌ-epoxylignan-9,7ꞌ-diol; triangularisines A, B, and C; N-ethyl-N-methyl-affineine; and N-methyl pachyconfine, and one previously not described as a natural product, epi-secoisolariciresinol monomethyl ether.The results demonstrated the anti-hyperglycemic activity of the infusion from A. triangularis leaves and showed that it is a rich source of lignoids, alkaloids, and glycosylated flavonoids, which are known to exhibit antidiabetic effects and other biological properties that can be beneficial for patients with chronic hyperglycemia, thus certifying the popular use of this herbal drink.

Made available in DSpace on 2022-04-29T08:30:07Z (GMT). No. of bitstreams: 0 Previous issue date: 2021-09-01 In this work, three new silver(I) complexes, [Ag(NO2-NN)(tu)]NO3 (1), [Ag(NON-NN)(tu)]NO3 (2), and [Ag(CN-NN)(tu)]NO3 (3) and (R-N'N”= extended phenanthroline ligand with aromatic diamines replaced with R = NO2; NON (furazan); CN (nitrile); tu = thiourea) were synthesized and fully characterized by the techniques of UV–Vis spectroscopy; Infrared spectroscopy; 1H and 13C{1H} NMR; molar conductivity; elemental analysis (CHN); high-resolution electrospray ionization mass spectrometry (HRMS-ESI); and X-ray diffraction (compound 3). The compounds showed themselves to be stable to light, air, and the DMSO solution. The DNA interaction assays indicated that compounds 2 and 3 interacted with the DNA, by interaction via the minor groove. Compounds 2 and 3 showed outstanding cytotoxicity towards the human cell lines A2780cis (cisplatin-resistant ovarian carcinoma), the DU-145 (prostate cancer) cells, and MRC-5 (human lung fibroblasts). The IC50 (concentration of a compound that is required for 50% inhibition of the cell viability, in vitro) values substantially overcome that of cisplatin. This data has suggested that such a molecular design is promising to develop silver(I) complexes, with high anticancer activities. Departamento de Química Univ. Federal de São Carlos UNESP – Univ. Estadual Paulista Instituto de Química Departamento de Química Analítica Físico-Química e Inorgânica Instituto de Física de São Carlos Universidade de São Paulo Avenida Trabalhador São-Carlense 400 UNESP – Univ. Estadual Paulista Instituto de Química Departamento de Química Analítica Físico-Química e Inorgânica

The Xanthomonas genus, comprises more than 30 species of gram-negative bacteria, most of which are pathogens of plants with high economic value, such as rice, common bean, and maize. Transcription activator-like effectors (TALEs), which act by regulating the host gene expression, are some of the major virulence factors of these bacteria. We present a novel tool to identify TALE genes in the genome of Xanthomonas strains and their respective targets. The analysis of the results obtained by TargeTALE in a proof-of-concept validation demonstrate that, at optimum setting, approximately 93% of the predicted target genes with available expression data were confirmed as upregulated during the infection, indicating that the tool might be useful for researchers in the field.

In this study, we describe the characterization of a new lectin, BfL-II, purified from the seeds of Bauhinia forficata, which is distinct, at sequence-level, from the previously reported lectin from the same specie (BfL). In addition, the gene for this lectin was cloned and expressed in Escherichia coli and its antiproliferative activity was evaluated against human breast and colorectal cancer cells (MCF-7 and HT-29, respectively). The treatment with 100 μg/μL of either native or recombinant BfL (nBfL or rBfL) significantly reduced the proliferation of both cancer cell lines (p

The genus Xanthomonas comprises Gram-negative bacteria, many of which are phytopathogens. Xanthomonas fuscans subsp. fuscans is one of the most devastating pathogens affecting the bean plant, resulting in the common bacterial blight of bean (CBB). The disease is mainly foliar and affects a wide variety of bean species, thus acting as the yield-limiting factor for the bean crop. Here, we report the whole-genome sequencing of a new strain of X. fuscans subsp. fuscans, named Xff49, isolated from the infected and symptomatic beans from Capão do Leão, Southern Brazil. The genetic analysis demonstrated the presence of single-nucleotide variants (SNVs) in this strain, potentially affecting the mobilome, cell mobility, and inorganic ion metabolism. In addition, the analysis resulted in the identification of a new plasmid similar to the pAX22 derived from Achromobacter denitrificans, which was named plX, along with plA and plC, previously reported in other strains of X. fuscans subsp. fuscans. Xff49 represents the first Brazilian genome of X. fuscans subsp. fuscans and might provide useful information applicable to the studies of phylogenetics, evolution, and pathogenomics, thereby allowing a better understanding of the genomic features present in the Brazilian strains. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1007/s42770-019-00050-0) contains supplementary material, which is available to authorized users.

The prolactin hormone is involved in several biological functions, although its main role resides on reproduction. As it interferes on fertility changes, studies focused on human health have established a linkage of this hormone to fertility losses. Regarding animal research, there is still a lack of information about the structure of prolactin. In case of horse breeding, prolactin has a particular influence; once there is an individualization of these animals and equines are known for presenting several reproductive disorders. As there is no molecular structure available for the prolactin hormone and receptor, we performed several bioinformatics analyses through prediction and refinement softwares, as well as manual modifications. Aiming to elucidate the first computational structure of both molecules and analyse structural and functional aspects related to these proteins, here we provide the first known equine model for prolactin and prolactin receptor, which obtained high global quality scores in diverse software's for quality assessment. QMEAN overall score obtained for ePrl was (- 4.09) and QMEANbrane for ePrlr was (- 8.45), which proves the structures' reliability. This study will implement another tool in equine genomics in order to give light to interactions of these molecules, structural and functional alterations and therefore help diagnosing fertility problems, contributing in the selection of a high genetic herd.

The lectin of Bauhinia forficata (nBfL) is a protein able to bind reversibly to N-acetylgalactosamine, performing several functions and one of them is the antiproliferative activity in tumor cells, but its effects have not yet been evaluated in female gametes. The objective of the present study was to determine the additional effect of B. forficata recombinants lectins in the medium of maturation in vitro of bovine oocytes in expression of genes related to oxidative stress pathways. To get the proteins, the gene for this recombinant lectin (rBfL) and its truncated isoform (rtBfL) were cloned and expressed in Escherichia coli (E.coli). The oocytes obtained through follicular puncture were incubated in IVM medium for 24 h containing concentrations of 10 μg/mL, 50 μg/mL and 100 μg/mL of nBfL, rBfL and rtBfL, and a no treated group as a control. In the groups treated with the concentration of 100 μg / mL, the gene expression of genes involved in oxidative stress SOD2, CAT, GPX-1, GSR, NOS2 and apoptosis BAX, CASP3 were evaluated. The rtBfL increased the expression of the SOD2, GSR and NOS2 genes and all the tested lectins increased the expression of the CASP3 gene compared to the control group. These findings indicate that the tested concentrations of the B. forficata recombinants lectins probably influence the expression of oxidative stress genes and increase the expression of the apoptotic gene CASP3 during in vitro maturation of bovine oocytes.

Anomalocalyx uleanus (Pax &amp, K. Hoffm.) Ducke (Euphorbiaceae) is a singular species in the genus and is restricted and exclusive to the Brazilian Amazon. A phytochemical study of A. uleanus leaves was performed, yielding the isolation of five major compounds: catechin/epicatechin, afzelin, quercetin 3-O-α-L-rhamnopyranoside, and astilbin. The phytochemical compositions of the methanolic extracts of leaves, roots, bark, and stem bark were determined using a dereplication approach. Forty-six compounds were annotated from the liquid chromatography-mass spectrometry (LC-MS/MS) data, while four lipids were identified using gas chromatography-mass spectrometry (GC-MS). In total, fifty compounds were detected, and they belonged to the primary metabolism and several classes of natural products such as flavonoids, flavonoids O-glycosides, flavonoids C-glycosides, biflavonoids, procyanidin, triterpene, triterpenes esterified with phenylpropanoids, phenylpropanoid derivatives, flavonolignans, coumarins, quinic acid derivatives, and benzoic acid derivatives. This is the first report on the phytochemical data of the genus Anomalocalyx, and the results of this study will contribute to the chemosystematic knowledge of the Euphorbiaceae family and justify the need for investigation of the pharmacological potential of the species A. uleanus.

Made available in DSpace on 2021-06-25T10:26:54Z (GMT). No. of bitstreams: 0 Previous issue date: 2021-01-01 This work has as the main focus, to analyze the behavior of physic-chemical variations from the fungus Xylaria sp., through the OSMAC (One Strain, Many Compounds) approach as an efficient way of obtaining new compounds. To perform such inductions and to explore the variability of the metabolic network of this microorganism, a factorial design was designed to induce variability (or enhancement) of metabolites. In view of chemometric insights, the planned inductions were imposed on the microorganism and variations in the chemical profile were observed in the crude extracts. Through mass spectrometry (HR-ESI–MS) and nuclear magnetic resonance-based profiles, combined with multivariate analysis through Principal Component Analysis (PCA), it was observed a marked variability of signals, confirming the efficacy in the metabolic alteration, defining the culture medium and agitation as the most important variables in the metabolic variability of the fungus. However, the extract mass is more significant for the agitation variable, and there is no relationship between the mass of crude extract and the amount of molecular signals of the complex matrices studied. Instituto Federal de Educação Ciência e Tecnologia de Rondônia IFRO, Campus Ji-Paraná, Street Rio Amazonas, 151 Instituto Federal de Educação Ciência e Tecnologia de Rondônia IFRO University of Manchester UoM Universidade Federal de São Carlos UFSCar Universidade Estadual Paulista “Júlio de Mesquita Filho” UNESP Universidade Estadual Paulista “Júlio de Mesquita Filho” UNESP

Made available in DSpace on 2020-12-12T02:14:00Z (GMT). No. of bitstreams: 0 Previous issue date: 2020-10-15 Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Fundação de Amparo à Pesquisa do Estado do Rio Grande do Sul Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) MicroRNAs have been hypothesized to be involved in the regulation of male fertility potential. The primary aim of our study was to demonstrate the effects of transfection with dendrimer nanostructure on the parameters of bovine sperm quality and to investigate whether the microRNA profile could be disturbed after cationic dendrimer-mediated exogenous DNA transfection of bovine spermatozoa. The binding of exogenous DNA was significantly increased when dendrimer-based transfection was implemented. However, cationic dendrimer transfection induced detrimental changes in the kinetics and sperm quality parameters, such as membrane integrity, acrosome reaction, and mitochondrial membrane potential, when compared to the control group. Sperm microRNA sequencing revealed 218 known and 106 novel microRNAs in the sperm samples, among which nine were dysregulated after transfection (one was upregulated and eight were downregulated), in comparison to the non-transfected sperm. All the dysregulated microRNAs were related to sperm quality and embryonic development. These results suggest that the transfection process using the dendrimer nanostructure has an impact on the quality and microRNA profile of bovine sperm. Laboratório de Genômica Estrutural Programa de Pós-Graduação em Biotecnologia Centro de Desenvolvimento Tecnológico Universidade Federal de Pelotas Laboratório de Reprodução Animal Programa de Pós-Graduação em Veterinária Universidade Federal de Pelotas Laboratório de Bioinformática e Proteômica Programa de Pós-Graduação em Biotecnologia Centro de Desenvolvimento Tecnológico Universidade Federal de Pelotas Laboratório de Biotecnologia do Câncer Programa de Pós-Graduação em Biotecnologia Centro de Desenvolvimento Tecnológico Universidade Federal de Pelotas Laboratório Genômica e Evolução Molecular Departamento de Genética Instituto de Biociências de Botucatu Universidade Estadual Paulista (UNESP) Unidad de Biología Molecular Institut Pasteur Montevideo Faculdade de Nutrição Universidade Federal de Pelotas Department of Biopharmaceutical Sciences Programs in Human Genetics and Biological Sciences University of California Laboratório Genômica e Evolução Molecular Departamento de Genética Instituto de Biociências de Botucatu Universidade Estadual Paulista (UNESP) Fundação de Amparo à Pesquisa do Estado do Rio Grande do Sul: 17/2551-0000953-3 CAPES: AUXPE 2537/2018 FAPESP: FAPERGS-FAPESP #19/2551-0002283-2

The occurrence of multidrug-resistant Serratia marcescens strains represents a serious public health threat. The purpose here is to report three cases of carbapenem-resistant S. marcescens infections with unfavorable clinical outcomes and provide a molecular description of the antibiotic resistance determinants at a genomic level. We performed bacterial identification by VITEK 2 and MALDI-TOF. The minimal inhibitory concentrations of antimicrobials were determined according to the Clinical and Laboratory Standards Institute guidelines, except for tigecycline, for which they were determined using Etest strips. Preliminary screening for the presence of carbapenemases was performed by ertapenem hydrolysis using MALDI-TOF MS. Whole-genome sequencing was provided to identify genes responsible for virulence and antimicrobial resistance. Here we report three challenging cases of S. marcescens that were resistant to the most commonly used antibiotics. Otherwise, we performed a genome description, which includes several genes involved in the resistance and virulence. These cases illustrate serious infection due to multidrug-resistant organisms and the complexity of treatment. Our results highlight the need to evaluate isolates regularly during long-term hospital stay to achieve optimal quality of clinical care and thus improve patient outcomes. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s42770-021-00477-4.

Calcium-dependent protein kinases (CDPKs) are encoded by a large gene family and play important roles against biotic and abiotic stresses and in plant growth and development. To date, little is known about the CDPK genes in strawberry (Fragaria x ananassa). In this study, analysis of Fragaria x ananassa CDPK gene family was performed, including gene structures, phylogeny, interactome and expression profiles. Nine new CDPK genes in Fragaria x ananassa were identified based on RNA-seq data. These identified strawberry FaCDPK genes were classified into four main groups, based on the phylogenetic analysis and structural features. FaCDPK genes were differentially expressed during fruit development and ripening, as well as in response to abiotic stress (salt and drought), and hormone (abscisic acid) treatment. In addition, the interaction network analysis pointed out proteins involved in the ABA-dependent response to plant stress via Ca2+ signaling, especially RBOHs. To our knowledge, this is the first report on CDPK families in Fragaria x ananassa, and it will provide valuable information for development of biofortified fruits and stress tolerant plants.

Plants may contain beneficial or potentially dangerous substances to humans. This study aimed to prepare and evaluate a new drug delivery system based on a glass-ionomer-Brazilian pepper extract composite, to check for its activity against pathogenic microorganisms of the oral cavity, along with its in vitro biocompatibility. The ethanolic Brazilian pepper extract (BPE), the glass-ionomer cement (GIC) and the composite GIC-BPE were characterized by scanning electron microscopy, attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR), and thermal analysis. The BPE compounds were identified by UPLC–QTOF–MS/MS. The release profile of flavonoids and the mechanical properties of the GIC-BPE composite were assessed. The flavonoids were released through a linear mechanism governing the diffusion for the first 48 h, as evidenced by the Mt/M∞ relatively to $$\sqrt t$$ t , at a diffusion coefficient of 1.406 × 10–6 cm2 s−1. The ATR-FTIR analysis indicated that a chemical bond between the GIC and BPE components may have occurred, but the compressive strength of GIC-BPE does not differ significantly from that of this glass-ionomer. The GIC-BPE sample revealed an ample bacterial activity at non-cytotoxic concentrations for the human fibroblast MRC-5 cells. These results suggest that the prepared composite may represent an alternative agent for endodontic treatment.

Lectins are proteins of non-immune origin present throughout all kingdoms of life. They are capable of binding to specific carbohydrates reversibly, thus performing several biological roles. Plant lectins are the most studied ones, with hundreds of isolated and characterized hemagglutinins. Most of the known lectins have been isolated from plants belonging to family Leguminosae, which includes genus Bauhinia. This genus comprises over 300 species located in the tropical zones of the planet, where these are utilized in folk medicine because of their numerous medicinal effects, such as anti-inflammatory and antidiabetic actions. Despite being studied for over fifty years, the literature regarding Bauhinia hemagglutinins is scarce, describing just ten proteins isolated from seven different species. Structurally as well as biophysically, there is great similarity among all the known Bauhinia lectins, which may classify them as chemotaxonomic markers; however, the carbohydrate-binding sites and further specificities are unique for each of these proteins. The activities identified for these lectins include growth inhibition in cancer cell lines, cell marking, anti-inflammatory and insecticidal effects, which are just a few among their various other activities of high economic importance. Besides their versatility, four recombinant Bauhinia lectins have already been successfully expressed in heterologous microbial systems, further suggesting that these proteins could serve as promising biotechnological products in future.

Investigations focused on the interactions of nanoparticles with lectins are relevant since it is well accepted that such proteins can be recognized by carbohydrates as parts of cell membranes. This can ultimately enhance the cellular uptake of the produced assemblies. In this framework, the physical interactions of phosphatidylcholine (PC) liposomes and the Bauhinia variegate lectin (BVL) are reported here. BVL-liposome interactions were characterized by a variety of techniques to understand the influence of BVL in the structural features, thermodynamic and spectroscopic properties of the hybrid material. The produced system is composed of 56% w/w lectin, and the scattering techniques show the presence of stable vesicular structures with a mean diameter DH ∼ 100 nm. The FTIR and NMR results showed a strong lectin effect on the PC choline region, restricting the rotational motion of the lipid group. The BVL-liposome interaction promoted hardening of the protein as evidenced by circular dichroism spectroscopy. The photophysics results suggest higher rigidity of the system in the presence of BVL. The BVL may be present in the inner or outer polar surface of the liposomes. The system was shown to be relatively stable and therefore potentially useful for carbohydrate recognition of nanoparticles.

Electrochemical sensors and biosensors are useful techniques for fast, inexpensive, sensitive, and easy detection of innumerous specimen. In face of COVID-19 pandemic, it became evident the necessity of a rapid and accurate diagnostic test, so the impedimetric immunosensor approach can be a good alternative to replace the conventional tests due to the specific antibody-antigen binding interaction and the fast response in comparison to traditional methods. In this work, a modified electrode with electrosynthesized PEDOT and gold nanoparticles followed by the immobilization of truncated nucleoprotein (N aa160–406aa) was used for a fast and reliable detection of antibodies against COVID-19 in human serum sample. The method consists in analyzing the charge-transfer resistance (RCT) variation before and after the modified electrode comes into contact with the positive and negative serum sample for COVID-19, using [Fe(CN)6]3-/4− as a probe. The results show a linear and selective response for serum samples diluted in a range of 2.5 × 103 to 20 × 103. Also, the electrode material was fully characterized by Raman spectroscopy, transmission electron microscopy and scanning electron microscopy coupled with EDS, indicating that the gold nanoparticles were well distributed around the polymer matrix and the presence of the biological sample was confirmed by EDS analysis. EIS measurements allowed to differentiate the negative and positive samples by the difference in the RCT magnitude, proving that the material developed here has potential properties to be applied in impedimetric immunosensors for the detection of SARS-CoV-2 antibodies in about 30 min., Graphical abstract Image, graphical abstract

Bacillus cereus is a gram positive bacterium with sporulation capacity. Here, we report the complete genome sequence of two native B. cereus strains (#25 and #29) isolated from intestinal tract of the crab Ucides sp. from Pacoti River in the State of Ceará, Brazil. The findings of this study might increase the molecular information for Bacillus strains. The data can be used in comparative analyses, origin and distribution, as well support for genetic engineering. Keywords: Gram positive bacteria, Genome sequence, Bacillus

The rice cultivar (Oryza sativa L.) BRS AG, developed by Embrapa Clima Temperado, is the first cultivar designed for purposes other than human consumption. It may be used in ethanol production and animal feed. Different abiotic stresses negatively affect plant growth. Soil salinity is responsible for a serious reduction in productivity. Therefore, the objective of this study was to evaluate the gene expression and the activity of antioxidant enzymes (SOD, CAT, APX and GR) and identify their functions in controlling ROS levels in rice plants, cultivar BRS AG, after a saline stress period. The plants were grown in vitro with two NaCl concentrations (0 and 136 mM), collected at 10, 15 and 20 days of cultivation. The results indicated that the activity of the enzymes evaluated promotes protection against oxidative stress. Although, there was an increase of reactive oxygen species, there was no increase in MDA levels. Regarding genes encoding isoforms of antioxidant enzymes, it was observed that OsSOD3-CU/Zn, OsSOD2-Cu/Zn, OsSOD-Cu/Zn, OsSOD4-Cu/Zn, OsSODCc1-Cu/Zn, OsSOD-Fe, OsAPX1, OsCATB and OsGR2 were the most responsive. The increase in the transcription of all genes among evaluated isoforms, except for OsAPX6, which remained stable, contributed to the increase or the maintenance of enzyme activity. Thus, it is possible to infer that the cv. BRS AG has defense mechanisms against salt stress.

Campylobacter jejuni is the most common bacterial cause of foodborne diarrheal disease worldwide and is among the antimicrobial resistant "priority pathogens" that pose greatest threat to public health. The genomes of two C. jejuni isolated from poultry meat sold on the retail market in Southern Brazil phenotypically characterized as multidrug-resistant (CJ100) and susceptible (CJ104) were sequenced and analyzed by bioinformatic tools. The isolates CJ100 and CJ104 showed distinct multilocus sequence types (MLST). Comparative genomic analysis revealed a large number of single nucleotide polymorphisms, rearrangements, and inversions in both genomes, in addition to virulence factors, genomic islands, prophage sequences, and insertion sequences. A circular 103-kilobase megaplasmid carrying virulence factors was identified in the genome of CJ100, in addition to resistance mechanisms to aminoglycosides, beta-lactams, macrolides, quinolones, and tetracyclines. The molecular characterization of distinct phenotypes of foodborne C. jejuni and the discovery of a novel virulence megaplasmid provide useful data for pan-genome and large-scale studies to monitor the virulent C. jejuni in poultry meat is warranted.

Alzheimer’s disease (AD) is a multifactorial pathology characterized by amyloid deposits, neurofibrillary formation, oxidative stress and cholinergic system dysfunction. In this sense, here we report the rational design of a multi-target directed ligand (MTDL) for AD based on virtual screening and bioinformatic analyses, exploring the molecular targets β-secretase (BACE-1), glycogen synthase kinase-3β (GSK-3β) and acetylcholinesterase (AChE). After this screening, the compound with higher molecular docking affinity was selected, the 1-(7-chloroquinolin-4-yl)-N-(4-methoxybenzyl)-5-methyl-1H-1,2,3-triazole-4 carboxamide(QTC-4-MeOBnE). To further our studies, the protective effect of QTC-4-MeOBnE (0.1 and 1 mg/kg for 20 days) on STZ-induced sporadic AD mice was determined. QTC-4-MeOBnE pretreatment attenuated cognitive and memory deficit induced by STZ in an object recognition test, Y-maze, social recognition test and step-down passive avoidance. The mechanisms underlying this action might be attributed to the reduction of lipid peroxidation and reactive species formation in the prefrontal cortex and hippocampus of mice submitted to STZ. In addition, QTC-4-MeOBnE pretreatment abolished the up-regulation of AChE activity and the overexpression of GSK 3β and genes involved in amyloid cascade such as BACE-1, protein precursor amyloid, у-secretase, induced by STZ. Moreover, toxicological parameters were not modified by QTC-4-MeOBnE chronic treatment. This evidence suggests that QTC-4-MeOBnE exerts its therapeutic effect through multiple pathways involved in AD.

Beans (Phaseolus vulgaris L.) contain nutritional and functional compounds, and form a staple diet for approximately one-third of the population worldwide. Due to the seasonal production of beans, the grains need to be stored for the off-season, and, in general, quality losses are observed. Among the main property changes, changes in color and cooking properties are easily perceived. Therefore, the storage conditions that prevent (low temperature and vacuum) and that favor (room temperature and absence of vacuum) the occurrence of these changes was studied to better understand the possible causative modifications and alterations in the proteomic profile of beans. The best storage condition involved the combination of low temperatures with vacuum; however, with one year of storage at this condition, a loss of quality with respect to coloration, cooking property and protein profile was observed.

Canine leptospirosis is often caused by Leptospira interrogans serovar Canicola. Infected dogs may become asymptomatic carriers of the pathogen, which leads to many public health concerns. In this work, we present the complete genome sequencing and in silico analysis from a virulent Brazilian strain of L. interrogans serovar Canicola, previously isolated from a stray dog in Sao Paulo City. Comparative genomic analysis with a reference genome allowed identification of 1031 INDELs and several arrangement variations. Out of 35,361 SNPs identified, 6780 were missense mutations and 16,114 were synonymous mutations. The Gene Ontology terms more affected by mutations were described. Interestingly, phylogenetic analyses indicated a genetic relatedness of the isolate with serovar Linhai strain 56,609. In addition, we found several virulence-related genes and main outer membrane proteins associated with pathogenesis. This genomic information about canine isolates may help to elucidate the molecular diversity and mechanisms of Leptospira spp. pathogenicity.

The introduction of next-generation sequencing (NGS) had a significant effect on the availability of genomic information, leading to an increase in the number of sequenced genomes from a large spectrum of organisms. Unfortunately, due to the limitations implied by the short-read sequencing platforms, most of these newly sequenced genomes remained as “drafts”, incomplete representations of the whole genetic content. The previous genome sequencing studies indicated that finishing a genome sequenced by NGS, even bacteria, may require additional sequencing to fill the gaps, making the entire process very expensive. As such, several in silico approaches have been developed to optimize the genome assemblies and facilitate the finishing process. The present review aims to explore some free (open source, in many cases) tools that are available to facilitate genome finishing.

Goat whey is normally discarded in the milk processing industry. However, several studies have addressed its biological properties and possible use in human or animal diet. The present study aimed to analysis the protein profile of goat whey to evaluate its possible oxidant, antioxidant, antibacterial, antitumour, and cytotoxic activities in vitro against human erythrocytes. Goat whey was skimmed, and crude protein extract (CPE) was obtained. Next, protein fractions (F) were obtained using ammonium sulphate precipitation method. The proteins were characterized by SDS-PAGE, two-dimensional electrophoresis and soluble protein measurements. No significant differences were observed in protein profile of CPE, F 30-60% and F 60-90%. The highest protein content was found in F 60-90% (0.41mgP/mL). All samples, except F 0-30% showed bacteriostatic activity against different bacterial strains. Only CPE at a concentration of 1000μg/mL was haemolytic against human erythrocytes. Oxidant activity against erythrocytes was not observed. Antioxidant activity was observed only for CPE. Cytotoxicity against C6 rat glioma cell line that was performed with CPE revealed tumour cell death>70% at concentrations of 0.05 and 0.1μg/mL. These results demonstrate at first time that CPE may be used as an antioxidant, bacteriostatic and cytotoxic compound against tumour cells.

Leptospira spp. are diderm (two membranes) bacteria that infect mammals causing leptospirosis, a public health problem with global implications. Thousands of people die every year due to leptospirosis, especially in developing countries with tropical climates. Prophylaxis is difficult due to multiple factors, including the large number of asymptomatic hosts that transmit the bacteria, poor sanitation, increasing numbers of slum dwellers, and the lack of an effective vaccine. Several leptospiral recombinant antigens were evaluated as a replacement for the inactivated (bacterin) vaccine; however, success has been limited. A prospective vaccine candidate is likely to be a surface-related protein that can stimulate the host immune response to clear leptospires from blood and organs. In this study, a comprehensive bioinformatics approach based on reverse and structural vaccinology was applied toward the discovery of novel leptospiral vaccine candidates. The Leptospira interrogans serovar Copenhageni strain L1-130 genome was mined in silico for the enhanced identification of conserved β-barrel (βb) transmembrane proteins and outer membrane (OM) lipoproteins. Orthologs of the prospective vaccine candidates were screened in the genomes of 20 additional Leptospira spp. Three-dimensional structural models, with a high degree of confidence, were created for each of the surface-exposed proteins. Major histocompatibility complex II (MHC-II) epitopes were identified, and their locations were mapped on the structural models. A total of 18 βb transmembrane proteins and 8 OM lipoproteins were identified. These proteins were conserved among the pathogenic Leptospira spp. and were predicted to have epitopes for several variants of MHC-II receptors. A structural and functional analysis of the sequence of these surface proteins demonstrated that most βb transmembrane proteins seem to be TonB-dependent receptors associated with transportation. Other proteins identified included, e.g., TolC efflux pump proteins, a BamA-like OM component of the βb transmembrane protein assembly machinery, and the LptD-like LPS assembly protein. The structural mapping of the immunodominant epitopes identified the location of conserved, surface-exposed, immunogenic regions for each vaccine candidate. The proteins identified in this study are currently being evaluated for experimental evidence for their involvement in virulence, disease pathogenesis, and physiology, in addition to vaccine development.

BACKGROUND Leptospirosis is the most widespread zoonotic disease. It is caused by infection with pathogenic Leptospira species, of which over 300 serovars have been described. The accurate identification of the causative Leptospira spp. is required to ascertain the pathogenic status of the local isolates. OBJECTIVES This study aimed to obtain the complete genome sequence of a virulent Leptospira interrogans strain isolated from southern Brazil and to describe its genetic features. METHODS The whole genome was sequenced by next-generation sequencing (Ion Torrent). The genome was assembled, scaffolded, annotated, and manually reviewed. Mutations were identified based on a variant calling analysis using the genome of L. interrogans strain Fiocruz L1-130 as a reference. FINDINGS The entire genome had an average GC content of 35%. The variant calling analysis identified 119 single nucleotide polymorphisms (SNPs), from which 30 led to a missense mutation. The structural analyses identified potential evidence of genomic inversions, translocations, and deletions in both the chromosomes. MAIN CONCLUSIONS The genome properties provide comprehensive information about the local isolates of Leptospira spp., and thereby, could facilitate the identification of new targets for the development of diagnostic kits and vaccines.

There is lack of suitable explants for efficient shoot proliferation in recalcitrant melon cultivars restricts genetic manipulation for crop improvement. In this study, we aimed to stimulate the regeneration capability of cells of melon (Cucumis melo L.) explants in order to increase shoot proliferation efficiency during regeneration. The media of seeds germination containing different concentrations of benzylaminopurine (BAP: 0.5, 1.0, and 1.5 mg L-1) can stimulate the shoot proliferation potential of cotyledonary nodes was described. The highest number of shoots per explant was obtained when the seeds and cotyledonary nodes were cultured on Murashige and Skoog media containing 1.0 and 0.5 mg L-1 BAP, respectively, and the regeneration process occurred through organogenesis. Key words: Cucumis melo L., organogenesis, cytokinin, explants.

A new galactose-binding lectin, termed BUL, has been purified from seeds of Bauhinia ungulata L. (Caesalpinoideae) by precipitation with solid ammonium sulfate followed by agarose–lactose affinity chromatography. B. ungulata lectin strongly agglutinated rabbit erythrocytes, both native and treated with proteolytic enzymes, and was inhibited by d -galactose and d -galactose-derived sugars, especially N-acetyl- d -galactosamine. BUL was shown to be a stable glycoprotein, maintaining its hemagglutinating activity after incubation at wide ranges of temperature and pH, but not after incubation with EDTA. By SDS-PAGE analysis, purified BUL showed an electrophoretic profile consisting of a single band with apparent molecular mass of 30 kDa. BUL showed intrinsic fluorescence typical of folded globular proteins, and CD spectra of lectin in the native state showed a predominance of β-sheet secondary structure. The N-terminal amino acid sequence of 19 residues showed a high sequential similarity to other galactose-specific lectins from the Bauhinia genus. In addition, BUL showed antifungal activity against phytopathogenic species and showed in vitro antiproliferative activity against the HT-29 cell line of human colon adenocarcinoma in a dose-dependent manner.

Neospora caninum is considerd a major cause of abortion in cattle worldwide. The antigenic domain of NcSRS2 in N. caninum is an important surface antigen present in the membrane of this parasite. In the present study, the Pichia pastoris expression system proved to be a useful tool for the production of recombinant protein. The truncated NcSRS2 gene (by removal of the N-terminal hydrophobic sequence), was cloned in the vector pPICZalphaB, and integrated on the genome of the methylotrophic yeast P. pastoris. Subsequently, the NcSRS2 protein was expressed, purified, and characterized using naturally infected cattle sera and Mab 6xhistag. The recombinant protein NcSRS2 was present in the supernatant of the culture, where later it was concentrated and purified using ammonium sulfate (∼100 mg/ml). An indirect immunoenzymatic assay (ELISA) was performed using cattle sera from endemic N. caninum area.

Next-generation sequencing has significantly reduced the cost of genome-sequencing projects, resulting in an expressive increase in the availability of genomic data in public databases. The cheaper and easier is to sequence new genomes, the more accurate the annotation steps have to be to avoid both the loss of information and the accumulation of erroneous features that may affect the accuracy of further analysis. In the case of bacteria genomes, a range of web annotation software has been developed; however, many applications have yet to incorporate the steps required to improve their result, including the removal of false-positive/spurious and a more complete identification of non-coding features. We present Genix, a new web-based bacterial genome annotation pipeline. A comparison of the results generated by Genix for four reference genomes against those generated by other annotation tools indicated that our pipeline is able to provide results that are closer to the reference genome annotation, with a smaller amount of false-positive proteins and missing functional annotated proteins. Additionally, the metrics obtained by Genix were slightly better than those obtained by Prokka, a state-of-art standalone annotation system. Our results indicate that Genix is a useful tool that is able to provide a more refined result, and may be a user-friendly way to obtain high-quality results.

Different rice (Oryza sativa L.) genotypes were subjected to high salinity and low temperature (150 mM NaCl and 13°C, respectively) for 0, 6, 24, 48, or 72 h. We evaluated the simultaneous expression of the genes OsCATA, OsCATB, and OsCATC, correlated gene expression with enzyme activity, and verified the regulation of these genes through identification of cis-elements in the promoter region. The hydrogen peroxide content increased in a tolerant genotype and decreased in a sensitive genotype under both stress conditions. Lipid peroxidation increased in the tolerant genotype when exposed to cold, and in the sensitive genotype when exposed to high salinity. Catalase activity significantly increased in both genotypes when subjected to 13°C. In the tolerant genotype, OsCATA and OsCATB were the most responsive to high salinity and cold, while in the sensitive genotype, OsCATA and OsCATC responded positively to saline stress, as did OsCATA and OsCATB to low temperature. Cis-element analysis identified different regulatory sequences in the catalase promoter region of each genotype. The sensitive genotype maintained a better balance between hydrogen oxyacid levels, catalase activity, and lipid peroxidation under low temperature than the resistant genotype. OsCATA and OsCATB were the most responsive in the salt-tolerant genotype to cold, OsCATA and OsCATC were the most responsive to saline stress, and OsCATA and OsCATB were the most responsive to chilling stress in the sensitive genotype. There were positive correlations between catalase activity and OsCATB expression in the tolerant genotype under saline stress and in the sensitive genotype under cold stress.

SUMMARYEffective control of gastrointestinal parasites is necessary in sheep production. The development of anthelmintics resistance is causing the available chemically based anthelmintics to become less effective. Biological control strategies present an alternative to this problem. In the current study, we tested the larvicidal effects ofBacillus thuringiensisvar.israelensisCry11Aa toxin againstHaemonchus contortuslarvae. Bacterial suspensions [2 × 108colony-forming units (CFU) g−1of the feces] ofB. thuringiensisvar.israelensisand recombinantEscherichia coliexpressing Cry11Aa toxin were added to naturallyH. contortusegg-contaminated feces. The larvae were quantified, and significant reductions of 62 and 81% (P< 0·001) were, respectively observed, compared with the control group. A 30 mL bacterial suspension (1 × 108CFU mL−1) ofB. thuringiensisvar.israelensisand recombinantE. coliexpressing Cry11Aa toxin were then orally administered to lambs naturally infected withH. contortus. Twelve hours after administration, feces were collected and submitted to coprocultures. Significant larvae reductions (P< 0·001) of 79 and 90% were observed respectively compared with the control group. The results suggest that the Cry11Aa toxin ofB. thuringiensisvar.israelensisis a promising new class of biological anthelmintics for treating sheep againstH. contortus.

Lectins are proteins able to interact specifically and reversibly with carbohydrates. They are present in all living beings, particularly in legume seeds, which have many biological functions. The aim of this study was to isolate, characterize and verify antioxidant, anti-hemolytic, antitumor and gastroprotective activities in a lectin present in seeds of Phaseolus lunatus L. var. cascavel (PLUN). The isolation of lectin was performed by size exclusion chromatography on Sephadex G-100, which was isolated from a protein capable of agglutinating only human erythrocytes type A, being this the only inhibited haemagglutination n -acetyl- d -galactosamine. Its weight was estimated by PAGE is 128 kDa. The lectin is thermostable up to 80 °C and is active between pH 2–11. As 8 M urea was able to denature the lectin. PLUN is a glycoprotein consisting of 2% carbohydrate and has antioxidant action with ascorbic acid equivalent antioxidant capacity (μMAA/g) of 418.20, 326 and 82.9 for total antioxidant activity, ABTS radical capture and capture of DPPH radical, respectively. The lectin has antitumor activity against melanoma derived cells at doses of 100 and 50 mg/ml, reducing up to 83% tumor cells, and gastroprotective action, reducing up to 63% damaged area of ​​the stomach induced by ethanol.

Lectins are non-immune proteins that reversibly bind to carbohydrates in a specific manner. Bauhinia variegata lectin I (BVL-I) is a Gal/GalNAc-specific, single-chain lectin isolated from Bauhinia variegata seeds that has been implicated in the inhibition of bacterial adhesion and the healing of damaged skin. Since the source of the native protein (nBVL) is limited, this study aimed to produce recombinant BVL-I in Pichia pastoris (rBVL-Ip). The coding sequence for BVL-I containing preferential codons for P. pastoris was cloned into the pPICZαB plasmid. A single expressing clone was selected and fermented, resulting in the secretion and glycosylation of the protein. Fed-batch fermentation in 7L-scale was performed, and the recombinant lectin was purified from culture supernatant, resulting in a yield of 1.5mg/L culture. Further, rBVL-Ip was compared to nBVL and its recombinant version expressed in Escherichia coli BL21 (DE3) (rBVL-Ie). Although it was expressed as a monomer, rBVL-Ip retained its biological activity since it was able to impair the initial adhesion of Streptococcus mutans and S. sanguinis in an in vitro model of biofilm formation and bacterial adhesion. In summary, rBVL-Ip produced in Pichia pastoris represents a viable alternative to large-scale production, encouraging further biological application studies with this lectin.