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Natalia Valdiviezo,1 Cindy Alcarraz,1,2 Denisse Castro,3,4 Renzo Salas,3 Victor Begazo-Mollo,5 Marco Galvez-Villanueva,6 Luz Medina Aguirre,7 Elica Garcia-León,8 Isela Quispe-Santivañez,9 Carmen Cornejo-Raymundo,10 Eduardo Paz-Cornejo,11 Luis Sanchez-Vilela,12 Vanessa Bermudez-Alfaro,13 Juan Carlos Vargas-Nina,14 Carlos Pérez-Ramos,15 Andrea Meza-Hoces,16 Paolo R Valdez Barreto,17 Ruth Huaringa-Leiva,18 Johanny Muro-Cieza,19 Valeria Aguilar-Vásquez,20 Eduardo Yache Cuenca,21 Silvia Neciosup-Delgado,1 Nathaly Poma-Nieto,22 Sheyla Chavez-Gavino,23 Lenin Fernandez-Rosas,24 Jhajaira M Araujo,2 Eduardo Payet,25 Henry L Gomez1 1Departamento de Medicina Oncológica, Instituto Nacional de Enfermedades Neoplásicas, Lima, 15038, Peru; 2Escuela Profesional de Medicina Humana, Universidad Privada San Juan Bautista, Lima, Perú; 3Departamento de Oncología Médica, Hospital Nacional Edgardo Rebagliati Martins, Lima, 15072, Peru; 4Centro de Investigación de Medicina de Precisión, Universidad de San Martin de Porres, Lima, 15024, Peru; 5Departamento de Medicina Oncológica, Instituto Regional de Enfermedades Neoplásicas Sur, Arequipa, Perú; 6Departamento de Medicina Oncológica, Instituto Regional de Enfermedades Neoplásicas Norte, La Libertad, Perú; 7Departamento de Medicina Oncológica, Hospital Goyeneche, Arequipa, 04600, Perú; 8Departamento de Medicina Oncológica, Hospital Regional de Lambayeque, Lambayeque, 14001, Perú; 9Departamento de Medicina Oncológica, Hospital Santa Rosa, Lima, 15084, Peru; 10Departamento de Medicina Oncológica, Hospital Nacional Dos de Mayo, Lima, Peru; 11Departamento de Medicina Oncológica, Hospital Nacional Alberto Sabogal Sologuren, Lima, Peru; 12Departamento de Medicina Oncológica, Hospital de Apoyo II Santa Rosa, Piura, Peru; 13Departamento de Medicina Oncológica, Hospital Nacional Guillermo Almenara Irigoyen, Lima, 15033, Peru; 14Departamento de Medicina Oncológica, Hospital II EsSalud de Cajamarca, Cajamarca, Peru; 15Departamento de Medicina Oncológica, Hospital María Auxiliadora, Lima, Peru; 16Departamento de Medicina Oncológica, Hospital II E de Lamas, San Martin, Peru; 17Departamento de Medicina Oncológica, Hospital de Alta complejidad Virgen de la Puerta, La Libertad, Peru; 18Departamento de Medicina Oncológica, Hospital Militar Central, Lima, Peru; 19Departamento de Medicina Oncológica, Hospital Regional Docente de Cajamarca, Cajamarca, Peru; 20Departamento de Promoción de la Salud, Prevención y Control del Cáncer, Instituto Regional de Enfermedades Neoplásicas Norte, La Libertad, Perú; 21Departamento de Epidemiología, Instituto Regional de Enfermedades Neoplásicas Norte, La Libertad, Perú; 22Departamento de Medicina Oncológica, Hospital Nacional Daniel Alcides Carrión, Lima, Perú; 23Departamento de Medicina Oncológica, Hospital Regional Eleazar Guzmán Barrón, Ancash, Perú; 24Departamento de Cirugía Oncológica, Hospital Regional Eleazar Guzmán Barrón, Ancash, Perú; 25Jefatura Institucional, Instituto Nacional de Enfermedades Neoplásicas, Lima, 15038, PeruCorrespondence: Natalia Valdiviezo, Departamento de Medicina Oncológica, Instituto Nacional de Enfermedades Neoplásicas, Av. Angamos 2520, Surquillo, Lima, 15038, Peru, Tel +51 992825055, Email nivaldivieso@gmail.comPurpose: The aim of this study is to evaluate the oncological care during the first state of national emergency due to the COVID-19 pandemic in several public cancer hospitals in Peru.Materials and Methods: A multicentric cross-sectional descriptive study was conducted by interviewing adult cancer patients diagnosed and treated between January 2019 and February 2020 from 18 hospitals. This study was carried out in September 2020, the last month of the first state of national emergency. Demographic and clinical characteristics were evaluated, including COVID-19 status and cancer treatment features.Results: A total of 1472 patients were included; the median age was 55 years (range 19– 97). Most patients (85.8%, n = 1263) had solid neoplasia, 13.5% (n = 198) hematologic neoplasia, and 0.7% (n = 11) others. SARS-CoV-2 infection was confirmed in 8.6% (n = 126), 1.2% (n = 18) were probable, 1.6% (n = 24) suspected, and 88.6% (n = 1304) negative cases. Overall, 51.6% of patients (n = 759) had cancer treatment delays, 42.5% (n = 626) changed treatment delivery (endovenous to oral systemic therapy), and 12.6% (n = 185) of cases cancer therapy was discontinued. In total, 10.3% (n = 117) of patients whose disease was controlled or in remission, experienced progression of disease during the state of emergency. A total of 6.7% (n = 98) of patients died, of whom 73.5% (n = 72) died from disease progression; 18.4% (n = 18) from SARS-CoV-2 infection and 8.1% (n = 8) from undetermined causes. Patients with hematological malignancies [hazard ratio (HR): 5.11 (95% confidence interval (CI): 1.99– 13.07)] and no response to therapy before the onset of the pandemic [5.01 (1.44– 17.42)] had an increased risk of death among COVID-19 infected individuals, whereas advanced clinical stage [5.09 (2.37– 10.95)] and discontinuation of treatment [3.66 (1.97– 6.78)] were risk factors among non-COVID-19 patients.Conclusion: Our study suggests that the COVID-19 pandemic has an adverse impact on the outcomes of Peruvian cancer patients. In our cohort, cancer mortality was higher than COVID-19 disease mortality.Keywords: COVID-19 pandemic, cancer care, Peru

In the absence of the herbicide option, weed control is the major constraint to organic soybean production in southern Brazil and eastern Paraguay. A joint entrepreneurial initiative introduced innovative 6000-V electro-shock desiccation for pre-plant weed control in organic soybeans, under zero-tillage-conservation agriculture, complemented by already-proven inter-row cultivation and biological control methods for insects and diseases. To develop an efficient operating procedure, a field trial with the electro-shock treatment was carried out and showed 78.6% overall weed control 28 days after electro-shock treatment, and no significant difference between operating speeds of 3.42, 9.49 and 16.57 km [h.sup.-1]. Further research is required to improve equipment performance and prove the medium- and long-term economic and social benefits to policy-makers, in order to justify environmental services payments for this environmentally friendly system. Additional keywords: biological control, cover crop, electro-shock, ground-driven horizontal disc, weed control., Introduction To supply demand for organic soybean (Glycine max L. Merr.) ingredients in their organic food products, the Associafao Gebana Brasil Solidaria and Gebana Brasil Ltda (jointly abbreviated to Gebana), [...]

The leaves of Mikania (Asteraceae) species are used in folk medicine as antispasmodic, antiulcerogenic and antirheumatic agents. Phytochemical screening of the crude hydroalcoholic 70% extract (CHE) of Mikania laevigata Shultz [...]

This study aimed to compare the conventional soybean ( Glycine max L.) cultivation method with integrated systems in an Latossolo Vermelho Acriférrico típico and how these systems affect soil cover biomass production, initial nutrient concentration in plant residues, soil respiration and microclimate, as well as soybean growth, physiology and productivity. A comparative analysis of microclimate and soil respiration, plant physiology, and growth was conducted between a conventional soybean monoculture (soybean grown without plant residues on the soil from the previous crop) and soybean grown in soil containing maize residues. Additionally, experiments were conducted to evaluate the effect of monocultures and previous integration between maize, three cultivars of Panicum maximum (Zuri, Tamani, and Quênia guinea grass) and Pigeon pea ( Cajanus cajan cv. BRS Mandarim) on soil health, physiological aspects, and soybean production. Our results indicated that all cultivars of Panicum maximum can be used in integrated systems. The triple consortium resulted in greater production of ground cover biomass and a higher concentration of nitrogen, phosphorus, potassium and sulphur, which contributed to lower soil temperature and greater humidity, without a concomitant increase in soil respiration. Consequently, soybeans grown in the resulting integrated systems cover biomass showed a higher net photosynthesis rate and increased leaf chlorophyll index, resulting in taller plants, with higher above-ground biomass production and 21.0% and a 36.8% increase in grain yield when compared to soybean cultivated on maize biomass and on soil without cover residue, respectively. The data presented in this study demonstrated that integrated systems, with the presence of grasses and legumes, improve soil climatic conditions and nutrient availability, enhancing soybean physiology and productivity characteristics, thus contributing to the sustainability of agricultural production, even in the short term. Further long-term research is strongly recommended., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2025 da Silva, Habermann, Costa, Costa, Silva, Severiano, Vilela, Silva, da Silva, Marques, Rodrigues and Martinez.)

Background: A vaccination programme against severe acute respiratory syndrome coronavirus 2 was initiated in Portugal in December 2020. In this study, we report the findings of a prospective cohort study implemented with the objective of monitoring antibody production in response to COVID-19 vaccination. Methods: The humoral immune response to vaccination was followed up using blood samples collected from 191 healthcare workers. Participants were split into three groups: the Oxford-AstraZeneca (Vaxzevria) vaccine group ( n = 68), the Pfizer-BioNTech COVID-19 (Comirnaty) vaccine group ( n = 51), and the Post-COVID group ( n = 72). The kinetics of anti-spike antibody production were evaluated until 56 days on average after the third dose (booster). Results: We observed that antibody titres peaked approximately one month after full vaccination and declined steadily thereafter. We also found that mRNA vaccination induces higher titres of antibodies than viral vector vaccination, and both generate greater antibody responses than mild or moderate COVID-19. Additionally, whilst the booster for the Oxford-AstraZeneca and Pfizer-BioNTech groups led to antibody levels higher than those at any previous sample collection point, the booster for the Post-COVID group (persons with a history of COVID-19 prior to vaccination) led to antibody levels lower than those attained one month after the second dose. Interpretation: Our results indicate that there are different kinetics of antibody production between individuals who received the Pfizer-BioNtech mRNA vaccine and those who received the Oxford-AstraZeneca vector vaccine, or individuals who had COVID-19 before being vaccinated. Additionally, we observed that exposure to either natural infection or vaccination modulates the response to subsequent vaccination. This is particularly evident after administration of the third dose to the Post-COVID group, where our findings point to a hindrance in vaccine boosting, probably due to unwanted feedback by high titres of pre-existing antibodies.

Background: During the COVID-19 pandemic, increased hand hygiene practices using water, soap and hand disinfectants, became prevalent, particularly among frontline workers. This study investigates the impact of these practices on the skin's ability to retain the allergenic metals nickel, cobalt, and chromium. The study constitutes three parts: (I) creating an impaired skin barrier, (II) exposing treated and untreated skin to nickel alone, and (III) in co-exposure with cobalt and chromium., Methods: Using full-thickness skin from stillborn piglets, in vitro experiments were conducted to assess retention of metals in skin at conditions mimicking intense hand hygiene practices. Treatment of skin with varying concentrations of sodium lauryl sulphate (SLS), to impair its barrier integrity was assessed. This was followed by exposure of treated and untreated skin to the metals, that were dissolved in Milli-Q water, 0.5% SLS, and ethanol respectively., Results: Results showed that pre-treatment with 5% SLS impaired the skin barrier with regards to the measure of trans epidermal water loss (TEWL). Metal amounts retained in the skin were generally higher in treated than untreated skin. The highest amounts of metal retained in skin were observed for exposure to nickel in ethanol. Co-exposure to nickel, cobalt, and chromium in 0.5% SLS resulted in the highest amounts of total metal retention., Conclusions: The in vitro findings highlight the increased risk of metal retention in skin due to an impaired barrier. The SLS concentration used in the current study corresponds to those used in many hand hygiene products. Hence, occupational settings with frequent exposure to water, soap and disinfectants need to consider protective measures not only for the irritant exposures themselves but also simultaneous exposure to allergenic metals., (© 2024. The Author(s).)

Background: During the COVID-19 pandemic, increased hand hygiene practices were implemented. Impaired skin health on the hands among healthcare workers has been reported previously. Knowledge of how worker in other occupations have been affected is scarce., Objectives: To investigate self-reported hand water-, and soap exposure and use of hand disinfectants, and hand eczema (HE) in frontline workers outside the hospital setting and in IT personnel during the COVID-19 pandemic., Methods: In this cross-sectional study, a questionnaire was sent out between 1 March and 30 April in 2021, to 6060 randomly selected individuals representing six occupational groups., Results: A significant increase in water exposure and hand disinfectant use was shown: Relative position (RP) 19; 95% confidence interval (CI) 0.17-0.21 and RP = 0.38: 95% CI 0.36-0.41, respectively. Newly debuted HE was reported by 7.4% of the population, more frequently among frontline workers (8.6%) compared to IT personnel (4.9%)., Conclusions: Water and soap exposure and use of hand disinfectants increased during COVID-19 pandemic, which may increase the risk of hand eczema. This highlights the importance of communication and implementation of preventive measures to protect the skin barrier also in occupations other than healthcare workers., (© 2024 The Authors. Contact Dermatitis published by John Wiley & Sons Ltd.)

Background: Considering the challenges posed by nitrogen (N) pollution and its impact on food security and sustainability, it is crucial to develop management techniques that optimize N fertilization in croplands. Our research intended to explore the potential benefits of co-inoculation with Azospirillum brasilense and Bacillus subtilis combined with N application rates on corn plants. The study focused on evaluating corn photosynthesis-related parameters, oxidative stress assay, and physiological nutrient use parameters. Focus was placed on the eventual improved capacity of plants to recover N from applied fertilizers (AFR) and enhance N use efficiency (NUE) during photosynthesis. The two-year field trial involved four seed inoculation treatments (control, A. brasilense, B. subtilis, and A. brasilense + B. subtilis) and five N application rates (0 to 240 kg N ha -1, applied as side-dress)., Results: Our results suggested that the combined effects of microbial consortia and adequate N-application rates played a crucial role in N-recovery; enhanced NUE; increased N accumulation, leaf chlorophyll index (LCI), and shoot and root growth; consequently improving corn grain yield. The integration of inoculation and adequate N rates upregulated CO 2 uptake and assimilation, transpiration, and water use efficiency, while downregulated oxidative stress., Conclusions: The results indicated that the optimum N application rate could be reduced from 240 to 175 kg N ha -1 while increasing corn yield by 5.2%. Furthermore, our findings suggest that replacing 240 by 175 kg N ha -1 of N fertilizer (-65 kg N ha -1 ) with microbial consortia would reduce CO 2 emission by 682.5 kg CO 2   -e ha -1 . Excessive N application, mainly with the presence of beneficial bacteria, can disrupt N-balance in the plant, alter soil and bacteria levels, and ultimately affect plant growth and yield. Hence, highlighting the importance of adequate N management to maximize the benefits of inoculation in agriculture and to counteract N loss from agricultural systems intensification., (© 2024. The Author(s).)

Objective: This study aimed to evaluate the rate of major adverse cardiac events (MACEs; the sum of death, myocardial infarction, and revascularization rates) according to interventional strategies guided by invasive physiological methods in both sexes in a Brazilian population during long-term follow-up for an average of 2 years., Methods: This retrospective single-center study included 151 consecutive patients (232 lesions) between January 2018 and January 2022. The participants were divided into two groups: the female group (FG), comprising 59 patients with 88 lesions, and the male group (MG), comprising 92 patients with 144 lesions., Results: The FG had a greater mean age (FG: 67.96 ± 13.12 vs. MG: 62.36 ± 12.01 years, p  = 0.009) and lower mean creatinine clearance (FG: 79.35 ± 38.63 vs. MG: 92.02 ± 38.62 mL/min, p  = 0.02) than did the MG. The percentage of lesions in the left main coronary artery was higher in the FG than in the MG (12.5% vs. 2.78%, p  = 0.006). The mean follow-up time was longer in the MG than in the FG (795.61 ± 350 vs. 619.19 ± 318 days, respectively; p  = 0.001). MACE occurred in 11.86% and 13.04% of patients in the FG and MG, respectively ( p  = 0.850). Secondary outcomes, such as death, reinfarction, and the need for new revascularization, showed no significant between-sex differences., Conclusions: Our study demonstrated the safety of invasive physiological methods to determine coronary revascularization in both male and female patients in a Brazilian population, as evidenced by the low rates of adverse cardiac events and death after a long-term follow-up., Competing Interests: The authors report no financial relationships or conflicts of interest regarding the content herein., (© Clarissa Campo Dall'Orto et al., 2024; Published by Mary Ann Liebert, Inc.)

To investigate consistency and accessibility of asthma and skin allergy hazard information in safety data sheets (SDSs) for cleaning agents on the Swedish market, we compiled a database of 504 SDSs and 351 therein declared ingredients. Labelling of products was compared to that of ingredients according to harmonised classification. For each ingredient, also notified classification and three additional sources on sensitising properties were compared. Product labelling most frequently indicated corrosion and irritation hazards. Only 3% of products were labelled as skin sensitisers and none as asthmagens. According to harmonised classification, 9% of products contained skin sensitisers, using other information sources increased the number to 46%. While 2% of products contained respiratory sensitisers according to harmonised classification, the number increased to 17% when using other information sources. Furthermore, sensitisers were declared across several sections of the SDSs, hampering easy access of such information. In conclusion, there are inconsistencies in hazard identification of cleaning agents and their ingredients. Hence, SDSs may not altogether fulfil its hazard information role. Improved criteria for identifying sensitisers and respiratory irritants are warranted. Additionally, we argue that all ingredients should be listed in section 3 regardless of concentration, to facilitate access of information about sensitising properties., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Early phylogenetic analysis of Pythium insidiosum, the etiologic agent of pythiosis in mammals, showed the presence of a complex comprising three monophyletic clusters. Two included isolates recovered from cases of pythiosis in the Americas (Cluster I) and Asia (Cluster II), whereas the third cluster included four diverged isolates three from humans in Thailand and the USA, and one isolate from a USA spectacled bear (Cluster III). Thereafter, several phylogenetic analyses confirmed the presence of at least three monophyletic clusters, with most isolates placed in clusters I and II. Recent phylogenetic analyses using isolates from environmental sources and from human cases in India, Spain, Thailand, and dogs in the USA, however, showed the presence of two monophyletic groups each holding two sub-clusters. These studies revealed that P. insidiosum possesses different phylogenetic patterns to that described by early investigators. In this study, phylogenetic, population genetic and protein MALDI-TOF analyses of the P. insidiosum isolates in our culture collection, as well as those available in the database, showed members in the proposed cluster III and IV are phylogenetically different from that in clusters I and II. Our analyses of the complex showed a novel group holding two sub-clusters the USA (Cluster III) and the other from different world regions (Cluster IV). The data showed the original P. insidiosum cluster III is a cryptic novel species, now identified as P. periculosum. The finding of a novel species within P. insidiosum complex has direct implications in the epidemiology, diagnosis, and management of pythiosis in mammalian hosts., Competing Interests: Declaration of competing interest No conflict of interest., (Copyright © 2022 British Mycological Society. Published by Elsevier Ltd. All rights reserved.)

Background: Fermented cocoa beans (Theobroma cacao L.) are a pivotal raw material for chocolate production. A cocktail yeast applied in the cocoa fermentation process can promote the formation of pleasant metabolites. Saccharomyces, Pichia and Hanseniaspora have been widely used in fermentation to improve the final product organoleptic profile, highlighting that fermentation is a critical point for chocolate flavour precursor production. This study aims to evaluate the impact of Pichia kluyveri and Saccharomyces cerevisiae strains as starter cultures on the fermentation for two cocoa hybrids, FA13 and CEPEC2002., Results: During fermentation processes, volatile organic compounds (VOCs) and protein profiles were assessed. Chocolates produced were also assessed regarding the presence of VOCs. Eighty VOCs were identified using gas chromatography coupled to mass spectrometry analysis. Mass spectrometry provided the protein profile evolution during fermentation and showed that the profiles changed with inoculation type (spontaneous versus inoculated fermentation). Chocolate obtained from FA13 inoculated with S. cerevisiae strain contained a greater amount of organics acids, being categorised as sourer than chocolate produced by spontaneous fermentation of FA13. CEPEC2002 inoculated with S. cerevisiae strain in co-culture with P. kluyveri strain generated less sour and sweeter chocolate than spontaneous fermentation only., Conclusions: Chocolates from inoculated assays with starter cultures were more accepted by evaluators, highlighting that P. kluyveri and S. cerevisiae influence the composition of VOCs. Besides, protein profiles also changed throughout fermentation. Further investigation should be conducted to clarify protein degradation dynamics during inoculated fermentations to define which of the microbial cultures positively affect the chocolate sensory characteristics. © 2021 Society of Chemical Industry., (© 2021 Society of Chemical Industry.)

The objective of this study was to evaluate the effect of heat stress on meta-taxonomic and metabolic profiles of prokaryotes in beef cattle rumen. Six pure-breed Nellore heifers with ruminal cannulas were used in the study. Six treatments were tested in a 6 × 6 Latin Square with six periods of 21days. The treatments were evaluated in a 2 × 2 + 2 factorial arrangement, consisting of 4 combinations: two temperatures conditions (thermoneutral, TN: 24 °C; and heat stress, HS: 34 °C) and two dietary energy concentration [low-energy (37% non-fibrous carbohydrates - NFC, 12 Mcal of metabolizable energy per kg of dry matter) or high-energy concentration (50.5% NFC, 18.49 Mcal of metabolizable energy per kg of dry matter)] plus two additional treatments with animals maintained in TN conditions but with your intake restricted (TN-RI) to the same of the heifers in HS with the two dietary energy concentration. The meta-genome was sequenced by MiSeq Sequencing System platform, and the DNA sequences were analysed using Geneious 10.2.3 software. The metabolic profile was evaluated by liquid and gas chromatography. Animals under HS presented lower (P = 0.04) prokaryote richness than animals under TN conditions. The genera Flavonifractor (1.4%), Treponema (0.6%) and Ruminococcus (0.9%) showed the lowest (P < 0.04) and Carnobacterium (7.7%) the highest (P = 0.02) relative abundance when the animals were submitted to HS, in relation to animals in TN. A total of 49 different metabolites were identified in the ruminal samples. The concentration of isobutyric acid (4.32 mM) was highest in bovine rumen under HS conditions. Heat stress influenced the microbiota and concentration of some organic acids in beef cattle rumen. There was a reduction in the richness of rumen in cattle under heat stress, but the diversity of prokaryotes was not affected., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

The Saccharomyces cerevisiae has been used for many years in the elaboration of food and beverage products, mainly associated with fermentation processes. The objective of this study was to characterize different indigenous S. cerevisiae strains and guide the notable strains for potential use in productions of fermented maize-based beverages. Initially, 81 strains isolated from different spontaneous food fermentations were evaluated. About 31% of strains showed phytase activity, an important characteristic for their application in cereals beverages production. All strains were able to grow in pH values 2.0, 3.0, and 5.0 and the presence of 5, 15, and 30% of glucose, but none could grow at 42 °C. Only 29.6% of the evaluated strains were able to efficiently grow in up to 1.0 mol L -1 of NaCl. The Rep-PCR and RAPD-PCR tools showed that the strains were differently grouped by the two techniques, and the grouping was not completely correlated with isolation source. A total of 65 volatile compounds were identified from the maize beverage produced. The profiles of volatile compounds produced by the strains were strain specific. S. cerevisiae strains isolated from the same source showed different chemical and genetic profiles, emphasize the importance to characterize the performance of each strain when searching for starter culture to develop or improve fermented beverages.

Antigen-specific immunotherapy (ASI) has been proposed as an alternative treatment strategy for type 1 diabetes (T1D). ASI aims to induce a regulatory, rather than stimulatory, immune response in order to reduce, or prevent, autoimmune mediated β-cell destruction, thus preserving endogenous insulin production. The abundance of immunocompetent antigen presenting cells (APCs) within the skin makes this organ an attractive target for immunotherapies. Microneedles (MNs) have been proposed as a suitable drug delivery system to facilitate intradermal delivery of autoantigens in a minimally invasive manner. However, studies to date have employed single peptide autoantigens, which would restrict ASI to patients expressing specific Human Leukocyte Antigen (HLA) molecules, thus stratifying the patient population. This study aims to develop, for the first time, an intradermal MN delivery system to target proinsulin, a large multi-epitope protein capable of inducing tolerance in a heterogeneous (in terms of HLA status) population of T1D patients, to the immunocompetent cells of the skin. An optimized three component coating formulation containing proinsulin, a diluent and a surfactant, facilitated uniform and reproducible coating of >30 μg of the active pharmaceutical ingredient on a stainless steel MN array consisting of thirty 500 μm projections. When applied to a murine model these proinsulin-coated MNs efficiently punctured the skin and after a limited insertion time (150 s) a significant proportion of the therapeutic payload (86%) was reproducibly delivered into the local tissue. Localized delivery of proinsulin in non-obese diabetic (NOD) mice using the coated MN system stimulated significantly greater proliferation of adoptively transferred antigen-specific CD8 + T cells in the skin draining lymph nodes compared to a conventional intradermal injection. This provides evidence of targeted delivery of the multi-epitope proinsulin antigen to skin-resident APCs, in vivo, in a form that enables antigen presentation to antigen-specific T cells in the local lymph nodes. The development of an innovative coated MN system for highly targeted and reproducible delivery of proinsulin to local immune cells warrants further evaluation to determine translation to a tolerogenic clinical outcome., (Copyright © 2020 Elsevier B.V. All rights reserved.)

We generalize the linear discrete dimensional scaling approach for the repulsive Hubbard model to obtain a nonlinear scaling relation that yields accurate approximations to the ground-state energy in both two and three dimensions, as judged by comparison to auxiliary-field quantum Monte Carlo (QMC) data. Predictions are made for the per-site ground-state energies in two and three dimensions for n (filling factor) and U (Coulomb interaction) values for which QMC data are currently unavailable.

Burkholderia harbors versatile Gram-negative species and is β-Proteobacteria. Recently, it was proposed to split the genus in two main branches: one of animal and plant pathogens and another, Paraburkholderia, harboring environmental and plant-beneficial species. Currently, Paraburkholderia comprises more than 70 species with ability to occupy very diverse environmental niches. Herein, we sequenced and analyzed the genome of Paraburkholderia kururiensis type strain KP23 T , and compared to P. kururiensis M130, isolated in Brazil, and P. kururiensis susbp. thiooxydans, from Korea. This study focused on the gene content of the three genomes with special emphasis on their potential of plant-association, biocontrol, and bioremediation. The comparative analyses revealed several genes related to plant benefits, including biosynthesis of IAA, ACC deaminase, multiple efflux pumps, dioxygenases, and degradation of aromatic compounds. Importantly, a range of genes for protein secretion systems (type III, IV, V, and VI) were characterized, potentially involved in P. kururiensis well documented ability to establish endophytic association with plants. These findings shed light onto bacteria-plant interaction mechanisms at molecular level, adding novel information that supports their potential application in bioremediation, biofertilization, and biocontrol of plant pathogens. P. kururiensis emerges as a promising model to investigate adaptation mechanisms in different ecological niches., (© 2019 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.)

In Brazil, the visceral leishmaniasis (VL) is caused by Leishmania infantum, while the tegumentary leishmaniasis (TL) etiological agents are mainly Leishmania braziliensis and Leishmania amazonensis. The canine visceral leishmaniasis (CVL) diagnosis is an important step of the VL control program in Brazil, which involves the elimination of infected dogs, the main urban VL reservoirs. The current serology-based diagnostic tests have shown cross-reactivity between these three species, whereas molecular diagnosis allows high sensitivity and specie identification. In the present study, 349 dogs of the metropolitan region of Belo Horizonte (Minas Gerais state) were screened by conjunctival swab and the samples analyzed by ITS-1 nested PCR. Thirty dogs (8.5%) tested positive. The RFLP of amplicons using HaeIII demonstrated that 17/30 samples presented a banding pattern compatible with L. infantum, 4/30 matched with L. amazonenis, 1/30 with L. braziliensis and 8/30 showed a mixed infection pattern. The samples that were distinct of L. infantum or presented a mixed pattern were submitted to RFPL with HaeIII and RsaI enzymes that confirmed the mixed pattern. Such patterns were also confirmed by Sanger Sequencing. The results pointed eight dogs with mixed infections and the establishment of TL causing species in the Belo Horizonte dog population. These findings highlight the need for more comprehensive epidemiological studies, since the TL transmission profile might be changing. This study also shows the potential of the ITS1-nPCR associated with RFLP for the proper Leishmania diagnosis and typing in the dog population., (Copyright © 2019 Elsevier B.V. All rights reserved.)

The use of lactic bacteria in the development of functional foods has increased in recent years. In addition to their probiotic characteristics, they can ferment a variety of substrates, such as cereals, roots, and tubers. Phytase producer lactic acid bacteria strains and their behavior during the fermentation process of yam-based food were studied. Leuconostoc lactis CCMA 0415, Lactobacillus plantarum CCMA 0744, and Lactobacillus fermentum CCMA 0745 were selected due to phytase production, pH reduction, and growth during 24 h of fermentation. Oxalate activity was not detected in all assays, suggesting its concentration was reduced due to the bleaching process. Among the selected strains, L. lactis CCMA 0415 appeared to be a promising strain in yam-based fermentations because it maintained a cell viability above 8 log CFU/mL and did not reduce diosgenin concentrations (around 8.0 μg/mL) after fermentation for 24 h, thereby, generating a potentially functional yam food. Furthermore, this strain promoted the decrease of pH value from 6.1 to 3.8 and produced 8.1 g/L lactic acid, at 6 h of fermentation. The L. lactis CCMA 0415 was reported as a starter culture in fermented products based on cereals, roots, and tubers.

Chocolate production suffered a vast impact with the emergence of the "witches' broom" disease in cocoa plants. To recover cocoa production, many disease-resistant hybrid plants have been developed. However, some different cocoa hybrids produce cocoa beans that generate chocolate with variable quality. Fermentation of cocoa beans is a microbiological process that can be applied for the production of chocolate flavor precursors, leading to overcoming the problem of variable chocolate quality. The aim of this work was to use a cocktail of microorganisms as a starter culture on the fermentation of the ripe cocoa pods from PH15 cocoa hybrid, and evaluate its influence on the microbial communities present on the fermentative process on the compounds involved during the fermentation, and to perform the chocolate sensorial characterization. According to the results obtained, different volatile compounds were identified in fermented beans and in the chocolate produced. Bitterness was the dominant taste found in non-inoculated chocolate, while chocolate made with inoculated beans showed bitter, sweet, and cocoa tastes. 2,3-Butanediol and 2,3-dimethylpyrazine were considered as volatile compounds making the difference on the flavor of both chocolates. Saccharomyces cerevisiae UFLA CCMA 0200, Lactobacillus plantarum CCMA 0238, and Acetobacter pasteurianus CCMA 0241 are proposed as starter cultures for cocoa fermentation.

Recent innovations in designing purification processes for biopharmaceutical production have enabled initial screening (optimization) of chromatographic conditions for binding to be performed in miniaturized batch format. The present report demonstrates the possibility of using this format to screen for selectivity and illustrates the need for careful adjustment of protocols when highly abundant, tightly-binding impurities are present in the sample. This batch format approach was used to choose a chromatography medium (resin) from a selection of available resins for the purification of recombinant insulin expressed in E. coli and to screen binding and elution conditions. Subsequent optimization was performed in small packed columns using a Design of Experiments (DoE) approach with statistical modeling before scaling up to a small pilot scale experiment. In this study insulin was effectively purified from the more tightly-binding C-peptide, and a reduction in insulin variants was also noted using the optimized conditions., (Copyright © 2014 Elsevier B.V. All rights reserved.)

To date, there is no vaccine available against human leishmaniasis. Although some vaccination protocols can induce immunity in murine models, they fail to induce protection in humans. The reasons for that remain unclear. The aim of the present study was to characterize the changes in the pattern of the immune response during subcutaneous vaccination with Leishvacin® in mice. We also investigated whether IFN-γ and nitric oxide synthase are indispensable for the protection elicited by the vaccine. C57BL/6 WT vaccinated mice showed smaller lesions and fewer numbers of parasites in footpads until 8 weeks post-infection. Up to this time, they produced higher levels of IFN-γ, IL-2, IL-4, IL-17A and IL-10 and higher specific antibody response than control non-vaccinated mice. Moreover, we showed that IFN-γ, most likely by induction of iNOS expression, is essential for immunity. However, after 12 weeks of infection, we observed loss of difference in lesion size and parasite burden between the groups. Loss of resistance was associated with the disappearance of differences in cytokine patterns between vaccinated and control mice, but not of antibody response, which remained different until a later time of infection. The reversal of resistance to L. amazonensis could not be explained by upregulation of regulatory cytokines. Our data point to a subversion of the host immune response by L. amazonensis even when a protective response was previously induced., (Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.)

Endophthalmitis caused by fungi is commonly diagnosed around the world in apparently healthy and immunocompromised individuals. An accurate clinical diagnosis for endophthalmitis confirmed by laboratory techniques is essential for early treatment with antifungal drugs, such as amphotericin B, imidazoles, and other antifungals. Here, we review endophthalmitis caused by fungi according to its classification into endogenous fungal endophthalmitis (EFE) and exogenous fungal endophthalmitis (EXFE). EFE is caused by endogenously acquired fungi, whereas the traumatic implantation of opportunistic fungal pathogens is the main feature of EXFE. We highlight the most important etiologies causing endophthalmitis and the steps required for a rapid diagnosis and management.

This study presents results regarding the successful cloning of the bacterial xylose isomerase gene (xylA) of Burkholderia cenocepacia and its functional expression in Saccharomyces cerevisiae. The recombinant yeast showed to be competent to efficiently produce ethanol from both glucose and xylose, which are the main sugars in lignocellulosic hydrolysates. The heterologous expression of the gene xylA enabled a laboratorial yeast strain to ferment xylose anaerobically, improving ethanol production from a fermentation medium containing a glucose-xylose blend similar to that found in sugar cane bagasse hydrolysates. The insertion of xylA caused a 5-fold increase in xylose consumption, and over a 1.5-fold increase in ethanol production and yield, in comparison to that showed by the WT strain, in 24h fermentations, where it was not detected accumulation of xylitol. These findings are encouraging for further studies concerning the expression of B. cenocepacia xylA in an industrial yeast strain., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

A 33 year-old HIV-positive Brazilian female patient was diagnosed with a cutaneous inflammatory reaction on her left forearm. The lesion spread rapidly affecting most of her forearm. The clinical diagnosis of tinea corporis (ringworm) was confirmed by wet mount preparations on 20% KOH and by the isolation of Trichophyton rubrum on pure cultures. Treatment with Fluconazole for a period of four weeks successfully cured the infection.

Proinsulin C-peptide has been recently described as an endogenous peptide hormone, responsible for important physiological functions others than its role in proinsulin processing. Accumulating evidences that C-peptide exerts beneficial effects in the treatment of long term complications of patients with type 1 diabetes mellitus indicate that this molecule may be administered together with insulin in future therapies. Despite its clear pharmacological interest, the secondary and three-dimensional (3D) structures of human C-peptide are still points of controversy. In the present work we report molecular dynamics (MD) simulations of human, rat I and rat II C-peptides. A common experimental strategy applied to all peptides consisted of homology building followed by multinanosecond MD simulations in vacuum and water. Circular dichroism (CD) experiments of each peptide in the absence and presence of 2,2,2-trifluoroethanol (TFE) were performed to support validation of the theoretical models. A multiple sequence alignment of 23 known mammalian C-peptides was constructed to identify significant conserved sites that would be important for the maintenance of secondary and tertiary structures. The analysis of the molecular dynamics trajectories for the human, rat I and rat II molecules have shown quite different general behavior, being the human C-peptide more flexible than the two others. Human and rat C-peptides exhibit very stable turn-like structures at the middle and C-terminal regions, which have been described as potential active sites of C-peptides. Human C-peptide also presented a short alpha-helix throughout the MD, which was not found in the rat molecules. CD data is in very good agreement with the MD results and both methods were able to identify a greater structural stability and potential in rat C-peptides when compared to the human C-peptide. The simulation results are discussed and validated in the light of multiple sequence alignment, recent experimental data from the literature and our own CD experiments.

Protozoa of the genus Leishmania are intracellular parasites of macrophages and may cause diverse clinical forms of leishmaniasis, including cutaneous, diffuse cutaneous, mucocutaneous and visceral leishmaniasis. Infection with L. major in mice indicates that a protective immune response is achieved when Th1 cells are developed. Thus, adoptive or vaccine-induced protection against leishmaniasis is largely dependent on cell-mediated immunity and IFN-gamma production. Induction of a Th1 response is dependent on the presence of IL-12 whilst lymphocytes are activated. This study was aimed at evaluating the role of IL-12 during infection with L. amazonensis and after vaccination with Leishvacin (killed Leishmania amazonensis promastigotes), since the role of this cytokine in vaccine-induced immunity with this preparation in experimental models or in humans is not yet elucidated. Hence, C57BL/6 interleukin-12-deficient mice (IL-12p40(-/-)) and wild-type controls (wt) were infected with L. amazonensis and the course of infection, parasite burden and cytokine production were compared. IL-12p40(-/-) mice were more susceptible to L. amazonensis than wt: lesions and parasite burden were larger in IL-12p40(-/-) when compared to wt. Interestingly, IL-4 was not produced in the absence of IL-12 in response to infection with L. amazonensis. To evaluate the role of IL-12 in the vaccine-induced immunity against L. amazonensis infection, IL-12p40(-/-) wt mice were vaccinated in the base of the tail and subsequently challenged with L. amazonensis in the footpads. Surprisingly, vaccinated IL-12p40(-/-) mice developed smaller lesions and had fewer parasites in footpads than non-vaccinated controls. Lymph node and spleen cells from vaccinated IL-12p40(-/-) mice did not produce high levels of IFN-gamma in response do in vitro stimulus with antigen. Hence, partial protection against infection with L. amazonensis could be obtained in the absence of functional IL-12 and a typical Th1 response.

Inabilities to process particulate material and to allow the use of high flow rates are limitations of conventional chromatography. Membranes have been suggested as matrix for affinity separation due to advantages such as allowing high flow rates and low-pressure drops. This work evaluated the feasibility of using an iminodiacetic acid linked poly(ethylenevinyl alcohol) membrane in the immobilized metal ion affinity chromatography (IMAC) purification of a human proinsulin(His)(6) of an industrial insulin production process. The screening of metal ions showed Ni(2+) as metal with higher selectivity and capacity among the Cu(2+), Ni(2+), Zn(2+) and Co(2+). The membrane showed to be equivalent to conventional chelating beads in terms of selectivity and had a lower capacity (3.68 mg/g versus 12.26 mg/g). The dynamic adsorption capacity for human proinsulin(His)(6) was unaffected by the mode of operation (dead-end and cross-flow filtration).

The C-peptide of proinsulin is important for the biosynthesis of insulin, but has been considered for a long time to be biologically inert. Recent studies in diabetic patients have stimulated a new debate about its possible regulatory role, suggesting that it is a hormonally active peptide. We describe structural studies of the C-peptide using 2D NMR spectroscopy. In aqueous solution, the NOE patterns and chemical shifts indicate that the ensemble is a nonrandom structure and contains substructures with defined local conformations. These are more clearly visible in 50% H2O/50% 2,2,2-trifluoroethanol. The N-terminal region (residues 2-5) forms a type I beta-turn, whereas the C-terminal region (residues 27-31) presents the most well-defined structure of the whole molecule including a type III'beta-turn. The C-terminal pentapeptide (EGSLQ) has been suggested to be responsible for chiral interactions with an as yet uncharacterized, probably a G-protein-coupled, receptor. The three central regions of the molecule (residues 9-12, 15-18 and 22-25) show tendencies to form beta-bends. We propose that the structure described here for the C-terminal pentapeptide is consistent with the previously postulated CA knuckle, believed to represent the active site of the C-peptide of human proinsulin.

Strategies to differentiate progenitor cells into beta cells in vitro have been considered as an alternative to increase beta cell availability prior to transplantation. It has recently been suggested that nestin-positive cells could be multipotential stem cells capable of expressing endocrine markers upon specific stimulation; however, this issue still remains controversial. Here, we characterized short- and long-term islet cell cultures derived from three different human islet preparations, with respect to expression of nestin and islet cell markers, using confocal microscopy and semi-quantitative RT-PCR. The number of nestin-positive cells was found to be strikingly high in long-term cultures. In addition, a large proportion (49.7%) of these nestin-positive cells, present in long-term culture, are shown to be proliferative, as judged by BrdU incorporation. The proportion of insulin-positive cells was found to be high in short-term (up to 28 days) cultures and declined thereafter, when cells were maintained in the presence of 10% serum, concomitantly with the decrease in insulin and PDX-1 expression. Interestingly, insulin and nestin co-expression was observed as a rare event in a small proportion of cells present in freshly isolated human islets as well as in purified islet cells cultured in vitro for long periods of time. In addition, upon long-term subculturing of nestin-positive cells in 10% serum, we observed reappearance of insulin expression at the mRNA level; when these cultures were shifted to 1% serum for a month, expression of insulin, glucagon and somatostatin was also detected, indicating that manipulating the culture conditions can be used to modulate the nestin-positive cell's fate. Attempts to induce cell differentiation by plating nestin-positive cells onto Matrigel revealed that these cells tend to aggregate to form islet-like clusters, but this is not sufficient to increase insulin expression upon short-term culture. Our data corroborate previous findings indicating that, at least in vitro, nestin-positive cells may undergo the early stages of differentiation to an islet cell phenotype and that long-term cultures of nestin-positive human islet cells may be considered as a potential source of precursor cells to generate fully differentiated/ functional beta cells.

The peculiarities of molecular structures and the influence of reaction conditions on the folding efficiency of fusion proteins-biotechnological precursors of human insulin, expressed in Escherichia coli as inclusion bodies have been investigated. The fusion proteins contained proinsulin sequence with various leader peptides connected by an Arg residue to the insulin B-chain. The kind and the size of leader peptide do not have essential influence on folding efficiency. However, the efficiency of protein folding depends on the location of the (His)6 site, which is used for metal-chelating affinity chromatography. In our study the protein folding depends on the reaction medium composition (including additives), the presence of accompanied cell components, pH, temperature, concentrations of protein, and redox agents. A negative influence of nucleic acid and heavy metal ions on folding has been found. S-sulfonated fusion protein has proinsulin-like secondary structure (by CD-spectroscopy data) that is the key point for 95% efficient folding proceeding. Folded fusion proteins are transformed into insulin by enzymatic cleavage.

In the 70's, pancreatic islet transplantation arose as an attractive alternative to restore normoglycemia; however, the scarcity of donors and difficulties with allotransplants, even under immunosuppressive treatment, greatly hampered the use of this alternative. Several materials and devices have been developed to circumvent the problem of islet rejection by the recipient, but, so far, none has proved to be totally effective. A major barrier to transpose is the highly organized islet architecture and its physical and chemical setting in the pancreatic parenchyma. In order to tackle this problem, we assembled a multidisciplinary team that has been working towards setting up the Human Pancreatic Islets Unit at the Chemistry Institute of the University of São Paulo, to collect and process pancreas from human donors, upon consent, in order to produce purified, viable and functional islets to be used in transplants. Collaboration with the private enterprise has allowed access to the latest developed biomaterials for islet encapsulation and immunoisolation. Reasoning that the natural islet microenvironment should be mimicked for optimum viability and function, we set out to isolate extracellular matrix components from human pancreas, not only for analytical purposes, but also to be used as supplementary components of encapsulating materials. A protocol was designed to routinely culture different pancreatic tissues (islets, parenchyma and ducts) in the presence of several pancreatic extracellular matrix components and peptide growth factors to enrich the beta cell population in vitro before transplantation into patients. In addition to representing a therapeutic promise, this initiative is an example of productive partnership between the medical and scientific sectors of the university and private enterprises.

Various methods have been investigated for the isolation and purification of fusion proteins of precursors of human insulin in the form of S-sulfonates, from the biomass of transformed Escherichia coli cells. Fusion proteins were prepared with different sizes and structures of the leader peptide and the poly-His position (inserted for purification by metal chelate affinity chromatography). The fusion proteins contained an IgG-binding B domain of protein A from Staphylococcus aureus at the N-terminus and an Arg residue between the leader peptide of the molecule and the proinsulin sequence, for trypsin cleavage of the leader peptide. Six residues of Cys in proinsulin allow the chemical modification of the protein as a (Cys-S-SO(-)(3))(6) derivative (S-sulfonate), which increases its polyelectrolytic properties and improves the efficiency of its isolation. Various methods of oxidative sulfitolysis were compared with catalysis by sodium tetrathionate or cystine and Cu2+ or Ni2+ ions. An optimum scheme for the isolation and purification of S-sulfonated fusion proteins was developed by the combination of metal-chelating affinity and ion-exchange chromatography. Highly purified (95%) S-sulfonated fusion protein was recovered which was 85% of the fusion protein contained in the biomass of E. coli cells. Folding of fusion protein S-sulfonate occurred with high yield (up to 90-95%). We found that the fusion protein-S-sulfonate has proinsulin-like secondary structure. This structure causes highly efficient fusion protein folding., (Copyright 2001 Academic Press.)

Protection from cutaneous leishmaniasis, a chronic ulcerating skin lesion affecting millions, has been achieved historically using live virulent preparations of the parasite. Killed or recombinant Ags that could be safer as vaccines generally require an adjuvant for induction of a strong Th1 response in murine models. Murine rIL-12 as an adjuvant with soluble Leishmania Ag has been shown to protect susceptible mice. We used 48 rhesus macaques to assess the safety, immunogenicity, and efficacy of a vaccine combining heat-killed Leishmania amazonensis with human rIL-12 (rhIL-12) and alum (aluminum hydroxide gel) as adjuvants. The single s.c. vaccination was found to be safe and immunogenic, although a small transient s.c. nodule developed at the site. Groups receiving rhIL-12 had an augmented in vitro Ag-specific IFN-gamma response after vaccination, as well as increased production of IgG. No increase in IL-4 or IL-10 was found in cell culture supernatants from either control or experimental groups. Delayed hypersensitivity reactions were not predictive of protection. Intradermal forehead challenge infection with 107 metacyclic L. amazonensis promastigotes at 4 wk demonstrated protective immunity in all 12 monkeys receiving 2 microgram rhIL-12 with alum and Ag. Partial efficacy was seen with lower doses of rhIL-12 and in groups lacking either adjuvant. Thus, a single dose vaccine with killed Ag using rhIL-12 and alum as adjuvants was safe and fully effective in this primate model of cutaneous leishmaniasis. This study extends the murine data to primates, and provides a basis for further human trials.

Availability of a safe, immunogenic, and affordable vaccine would represent the best strategy for control of cutaneous leishmaniasis (CL). Stability in field conditions is a essential property for any candidate vaccine. The stability and immunogenicity of three different preparations (thimerosal-preserved, autoclaved, and lyophilized) of a killed Leishmania amazonensis vaccine were assessed using fresh products and after 12 months of storage at 4 degrees C. Autoclaving was associated with a time-dependent decrease in the immunogenicity of the vaccine, as measured by the leishmanin skin test and production of interferon-gamma. These findings are of importance in the decision of which preparation of candidate killed CL vaccines should move to phase III trials.

A process for purifying bovine pancreatic glucagon as a by-product of insulin production is described. The glucagon-containing supernatant from the alkaline crystallization of insulin was precipitated using ammonium sulfate and isoelectric precipitation. The isoelectric precipitate containing glucagon was then purified by ion-exchange chromatography on Q-Sepharose FF, gel filtration on Sephadex G-25 and ion-exchange chromatography on S-Sepharose FF. A pilot scale test was performed with a recovery of 87.6% and a purification factor of 8.78 for the first chromatographic step, a recovery of 75.1% and a purification factor of 3.90 for the second, and a recovery of 76.2% and a purification factor of 2.36 for the last one. The overall yield was 50%, a purification factor of 80.8 was obtained and the fraction containing active glucagon (suitable for pharmaceutical preparations) was 84% pure as analyzed by HPLC.