Use of Sanger protocols to identify variants of concern, key mutations and track evolution of SARS-CoV-2.

Vaccination and the emergence of SARS-CoV-2 variants mark the second year of the pandemic. Variants have amino acid mutations at the spike region, a viral protein central in the understanding of COVID-19 pathogenesis and vaccine response. Variants may dominate local epidemics, as Gamma (P.1) in Brazil, emerging in 2020 and prevailing until mid-2021. Different obstacles hinder a wider use of Next-Generation Sequencing for genomic surveillance. We describe Sanger based sequencing protocols: i) Semi-nested RT-PCR covering up to 3.684 kb (>96 %) spike gene; ii) One-Step RT-PCR for key Receptor Binding Domain (RBD) mutations (codons 417-501); iii) One-Step RT-PCR of partial N region to improve genomic capability. Protocols use leftovers of RNA extracted from nasopharyngeal swabs for quantitative RT-PCR diagnosis; with retro-transcribed DNA sequenced at ABI 3500 using dye termination chemistry. Analyses of sequences from 95 individuals (late 2020/early 2021) identified extensive amino acid variation, 57 % with at least one key mutation at the Receptor Binding Domain, with B.1.1.28 lineage most prevalent, followed by Gamma and Zeta variants, with no Delta variant observed. The relatively low cost and simplicity may provide an accessible tool to improve surveillance of SARS-CoV-2 evolution, monitor new variants and vaccinated breakthroughs.

Effects of substrates containing different concentrations of sewage sludge on physiological parameters and quality of Alibertia edulis (Rubiaceae) seedlings.

Sewage sludge (SS), a solid residue of effluent treatment, is rich in organic matter and nutrients, while also containing heavy metals and other potential contaminants. The feasibility of employing SS as a substrate component for seedling production depends on its composition and dose, as well as on the tolerance limit of individual plant species. To expand the knowledge base on the use of SS in the production of plants native to the Brazilian Cerrado biome, we evaluated the physiological responses and quality of Alibertia edulis seedlings grown under distinct SS concentrations. Chlorophyll a fluorescence (ChlF), stomatal conductance (gs), leaf temperature (Lt), biomass, growth, and seedling quality were investigated. At 25%, SS improved growth, biomass, and seedling quality, while substrates containing 50% or more SS affected gs, Lt, and ChlF. Seedling quality was strongly worsened by SS at 75% and 100%, as shown by lower biomass and impaired growth, including leaf symmetry loss and leaf deformities possibly related to contaminants, particularly heavy metals. Accordingly, we conclude that SS exhibited potential as a fertilizer at concentrations below 50%, but exerted a toxic effect on seedlings at higher concentrations.

Physiological mechanisms and phytoremediation potential of the macrophyte Salvinia biloba towards a commercial formulation and an analytical standard of glyphosate.

Glyphosate (Gly) is the most widely used herbicide in the world and has broad-spectrum and non-selective activity. Its indiscriminate use hence risks contamination of water bodies and can affect living organisms, especially sensitive or resistant non-target plants. Despite this, studies on physiological mechanisms and Gly remediation in Neotropical aquatic plants remain limited. This study aims to evaluate the physiological mechanisms of the aquatic macrophyte Salvinia biloba on exposure to different concentrations of a Gly commercial formulation (Gly-CF) and a Gly analytical standard (Gly-AS). Furthermore, using square-wave voltammetry (SWV), we determined whether the studied plant could remove Gly from water. Our data suggest that Gly-AS and Gly-CF induce similar physiological responses in S. biloba. However, Gly-CF was more phytotoxic. Depending on the concentration, the two forms of Gly affected the plants, decreasing the chlorophyll a and b contents and the photosystem II (PSII) photochemical activity. The data also revealed that Gly promoted oxidative stress and increased the shikimic acid concentration. At the same time, the plants removed Gly from water, with 100% removal for 1 mg L-1 Gly and above 60% removal for the other concentrations studied. Therefore, our results suggest that S. biloba may be a potential phytoremediation agent for low Gly concentrations, since 1 mg L-1 Gly was completely removed and exhibited low phytotoxicity. This study deepens our scientific understanding of the Gly impact on and the phytoremediation potential of S. biloba.

Purification of IgG against ribonucleoprotein by a homemade immunoaffinity chromatography column for rabies diagnosis.

Polyclonal or monoclonal antibodies against rabies virus ribonucleoprotein (RNP) conjugated to fluorescein isothiocyanate (FITC) have been employed for Rabies virus (RABV) antigen detection by the direct fluorescent antibody test (DFA). To date, these biomolecules have been purified by traditional methods such as precipitation by ammonium sulfate or ion exchange chromatography followed by ammonium sulfate precipitation, which allows only for partial detection of the protein of interest. In this study, we aimed to purify anti-RNP polyclonal horse IgG antibodies by cation-exchange chromatography in combination with a homemade immunoaffinity chromatography on RNP immobilized (RNP-IAC). Furthermore, to evaluate the accuracy of the prepared anti-RNP IgG fluorescent antibody in diagnostic purposes, DFA was applied for RABV antigen detection in suspected brain samples of different animal species. The combination of these two techniques made it possible to obtain antibodies with high selectivity and purity. Compared with the performance of the traditional method, anti-RNP IgG antibodies purified by RNP-IAC can be obtained from a smaller volume of hyperimmune serum and with greater avidity. Furthermore, the results obtained by DFA analyses revealed that the prepared anti-RNP IgG fluorescent antibody achieved 100% diagnostic specificity and sensitivity for RABV antigen detection. Thus, two-technique chromatographic, including RNP-IAC technology could be appropriate methods for the purification of polyclonal anti-RNP IgG for the use as a diagnostic reagent for rabies.

Teste piloto de checklist de cirurgia segura: relato de experiência / Test check pilot list of safe surgery: experience report

Objetivo: relatar a experiência de um teste piloto de utilização de checklist de cirurgia segura. Metodologia:trata-se de uma ação realizada por estudantes e docentes de enfermagem da Universidade Federal doRecôncavo da Bahia e a equipe do centro cirúrgico de um hospital escola. O desenvolvimento do teste pilotoenvolveu a comunicação setorial, a construção de um impresso, a integração do instrumento à prática e umareunião de equipe. Resultados: foram realizados quatro encontros com a participação da equipe, no total de35 funcionários. Foram aplicados 30 checklists em procedimentos anestésicos-cirúrgicos. Ao final, discutimosos resultados e consideramos que foi possível reconhecer os desafios de segurança no setor, as dificuldades epotencialidades da ação e do instrumento proposto. Conclusão: essa experiência agregou ensino-assistênciapesquisae proporcionou o desenvolvimento de estratégias para a segurança do paciente.

Objective: report the experience of a pilot test of the use of a checklist for safe surgery. Methodology: thedevelopment of the pilot test involved the communication sector, the construction of a form, the integrationof the instrument to practice and a team meeting. Results: were conducted four meetings with theparticipation of the team, for a total of 35 employees. Were applied 30 checklists in anesthetic proceduresand surgical. In the end, we discuss the results and we believe that it was possible to recognize the securitychallenges in the sector, the difficulties and possibilities of action and the proposed instrument. Conclusion:this experience has added teaching and assistance-research and has provided for the development ofstrategies for patient safety.