l-Asparaginase Type II from Fusarium proliferatum: Heterologous Expression and In Silico Analysis.

The search for new drug-producing microorganisms is one of the most promising situations in current world scientific scenarios. The use of molecular biology as well as the cloning of protein and compound genes is already well established as the gold standard method of increasing productivity. Aiming at this increase in productivity, this work aims at the cloning, purification and in silico analysis of l-asparaginase from Fusarium proliferatum in Komagataella phaffii (Pichia pastoris) protein expression systems. The l-asparaginase gene (NCBI OQ439985) has been cloned into Pichia pastoris strains. Enzyme production was analyzed via the quantification of aspartic B-hydroxamate, followed by purification on a DEAE FF ion exchange column. The in silico analysis was proposed based on the combined use of various technological tools. The enzymatic activity found intracellularly was 2.84 IU/g. A purification factor of 1.18 was observed. The in silico analysis revealed the position of five important amino acid residues for enzymatic activity, and likewise, it was possible to predict a monomeric structure with a C-score of 1.59. The production of the enzyme l-asparaginase from F. proliferatum in P. pastoris was demonstrated in this work, being of great importance for the analysis of new methodologies in search of the production of important drugs in therapy.

Patient and Healthcare Professional Insights of Home- and Remote-Based Clinical Assessment: A Qualitative Study from Spain and Brazil to Determine Implications for Clinical Trials and Current Practice.

INTRODUCTION: The SARS-CoV-2 virus pandemic has accelerated the growing trend towards using home- and remote-based medical testing (H/RMT). The aim of this study was to gather insights and explore the opinions of patients and healthcare professionals (HCPs) in Spain and Brazil regarding H/RMT and the impact of decentralised clinical trials. METHODS: This qualitative study consisted of in-depth open question interviews of HCPs and patients/caregivers followed by a workshop that aimed to determine the advantages and barriers to H/RMT in general, and in the context of clinical trials. RESULTS: There were 47 participants in the interviews (37 patients, 2 caregivers, 8 HCPs) and 32 in the validation workshops (13 patients, 7 caregivers, 12 HCPs). The main advantages for the use of H/RMT in current practice were the comfort and convenience, the ability to improve the relationship between HCPs and patients and personalise patient care, and the increased patient awareness towards their disease. Barriers to H/RMT included accessibility, digitalisation, and the training requirements for both HCPs and patients. Furthermore, according to the Brazilian participants, there is a general distrust in the logistical management of H/RMT. Patients indicated that the convenience of H/RMT did not influence their decision to participate in a clinical trial, with the main reason for participating in a clinical trial being to improve health; however, H/RMT in clinical research does aid adherence to the long-term follow-up associated with trials and provides access to patients living far from the clinical sites. CONCLUSION: Insights from patients and HCPs suggest that the advantages of H/RMT may outweigh the barriers, and that social, cultural and geographical factors and the HCP-patient relationship are critical aspects to be considered. Moreover, the convenience of H/RMT does not appear to be a driver for participating in a clinical trial but can facilitate patient diversity and study adherence.

Preferências e expectativas de pacientes de ensaio clínico sobre exames em domicílio e testes remotos / Preferences and expectations of patients in clinical trial about laboratory exams at home and remote tests / Preferencias y expectativas de los pacientes en ensayo clínico sobre exámenes domiciliarios y pruebas remotas

Objetivo: Conhecer a aceitação de pacientes de ensaio clínico quanto a coleta de exames domiciliares, testes remotos e teleconsultas, compreendendo as suas preferências e expectativas. Método: Estudo qualitativo, exploratório, descritivo, com entrevistas em profundidade envolvendo profissionais de saúde (especialistas) e pacientes, com e sem experiência em ensaios clínicos, com posterior validação dos constructos por meio de workshop participativo. Resultados: Para os pacientes de ensaio clínico, a aceitação mostrou-se relativa. Diversos fatores influenciam suas preferências e expectativas, sendo as principais: a) O grau de confiabilidade nos processos e a acurácia dos resultados de exames domiciliares. b) Os testes remotos, apesar de bem aceitos, dependem da capacidade de entendimento no correto manuseio da tecnologia requerida. c) A teleconsulta, já bem utilizada e aceita, não substitui totalmente o encontro presencial com o médico em fases críticas. A preferência é pelo uso equilibrado do "presencial x remoto". Conclusão: A aceitação é influenciada por diversas variáveis. Devem ser definidas estratégias adequadas para cada caso, para garantir resultados seguros e confiáveis, com equilíbrio entre o presencial e remoto, através da participação ativa dos pacientes nas tomadas de decisão.

To understand the acceptance of clinical trial patients regarding in-house exams, remote tests and teleconsultations, revealing their preferences and expectations Method: Qualitative, exploratory-descriptive study, with in-depth interviews involving health professionals (specialists) and patients, with and without experience in clinical trials, with subsequent validation of the constructs through a participatory workshop. Results: For clinical trial patients, acceptance was relative. Several factors influence their preferences and expectations, the main ones being: a) The degree of reliability in the processes and the accuracy of the results of home exams. b) Remote tests, although well accepted, depend on the ability of the patient to understand the correct handling of the required technology. c) Teleconsultation, already well used and accepted, does not completely replace the face-to-face meeting with the doctor in critical phases. The preference is for the balanced use of "in person versus remote". Conclusion: Acceptance is influenced by several variables. Adequate strategies must be defined for each case, to ensure safe and reliable results, with a balance between face-to-face and remote, through the active participation of patients in decision-making

Objetivo: Conocer la aceptación de los pacientes de ensayos clínicos en cuanto a la recogida de exámenes domiciliarios, pruebas a distancia y teleconsultas, entendiendo sus preferencias y expectativas. Método: Estudio cualitativo, exploratorio-descriptivo, con entrevistas en profundidad involucrando a profesionales de la salud (especialistas) y pacientes, con y sin experiencia en ensayos clínicos, con posterior validación de los constructos a través de un taller participativo. Resultados: Para pacientes de ensayos clínicos, la aceptación fue relativa. Varios factores influyen en sus preferencias y expectativas, siendo los principales: a) El grado de confiabilidad en los procesos y la precisión de los resultados de los exámenes domiciliarios. b) Las pruebas a distancia, aunque bien aceptadas, dependen de la capacidad de comprender el manejo correcto de la tecnología requerida. c) La teleconsulta, ya muy utilizada y aceptada, no sustituye por completo al encuentro presencial con el médico en fases críticas. La preferencia es por el uso equilibrado de "en persona versus remoto". Conclusión: La aceptación está influenciada por varias variables. Se deben definir estrategias adecuadas para cada caso, para garantizar resultados seguros y confiables, con equilibrio entre presencial y remoto, a través de la participación activa de los pacientes en la toma de decisiones

Underlying factors influencing street food vendors' implementation of food safety behaviours

This study aimed to determine the underlying factors influencing street food vendors' implementation of food safety behaviours related to food handling. A questionnaire based on the Theory of Planned Behaviour was applied to 303 street food vendors in Southern Brazil. Open-ended questions identified that consequences related to consumers were important factors influencing street food vendors' food safety behaviour. Also, water supply and lack of water were identified, respectively, as possible facilitators and barriers to carry out food safety behaviours such as handwashing and cleaning the shelf used to prepare food. Optimistic bias was identified and the majority of street food vendors gave positive results related to perceived behavioural control, subjective norm, attitude and behavioural intention. Improvements are needed in government regulation, infrastructure and food safety training for this sector, with measures that reflect and are adapted to the street food environment.

Este estudio tuvo como objetivo determinar los factores subyacentes que influyen en la implementación del comportamiento de seguridad de los alimentos en la manipulación de alimentos por vendedores ambulantes de alimentos. Se aplicó un cuestionario basado en la Teoría del Comportamiento Planificado a 303 vendedores ambulantes de alimentos en el sur de Brasil. Las preguntas abiertas identificaron que las consecuencias relacionadas con los consumidores eran factores importantes que influían en el comportamiento de seguridad de los alimentos por parte de los vendedores ambulantes de alimentos. También se identificaron el suministro de agua y la falta de agua, respectivamente, como posibles facilitadores y barreras para realizar comportamientos de seguridad de los alimentos como lavarse las manos y limpiar la mesa de trabajo que utilizan para preparar los alimentos. Se identificó sesgo optimista y la mayoría de los vendedores ambulantes de alimentos dieron resultados positivos relacionados con el control conductual percibido, la norma subjetiva, la actitud y la intención de comportamiento. Se necesitan mejoras en la regulación gubernamental, la infraestructura y la capacitación en seguridad de los alimentos para el sector, con medidas que reflejen y se adapten al entorno de la comida callejera.

Accelerated apoptosis, oxidative stress, and cholinergic inflammation in blood of metalworkers.

Metalworkers are exposed to numerous chemicals in their workplace environment, such as solvents, heavy metals, and metalworking fluids, that have a negative impact on their health. Furthermore, there is an increase in the prevalence of chronic diseases among metalworkers; however, the molecular mechanisms involved in this increased predisposition to chronic diseases are unclear. Considering that occupational exposure represents a potential risk for metalworkers, the aim of this study was to measure biomarkers of oxidative stress, inflammation, and cytotoxicity in the peripheral blood of metalworkers from Southern Brazil. The study included 40 metalworkers and 20 individuals who did not perform activities with any recognized exposure to chemical substances, such as those working in administration, commerce, and education, as controls. Cellular and molecular biomarkers as leukocyte viability, intracellular production of reactive species, mitochondrial mass and membrane potential and plasma lipid peroxidation, sulfhydryl groups, total antioxidant capacity, and butyrylcholinesterase activity were evaluated in the blood of metalworkers and controls. Metalworkers were found to have higher rates of apoptosis, increased production of reactive species, and increased mitochondrial potential and mass in leukocytes associated with decreased antioxidant defenses and increased activity of the butyrylcholinesterase enzyme in their plasma. It can be concluded that cytotoxicity, oxidative stress, and inflammation are involved in the multiplicity of health outcomes related to chemical exposure in the metalworking industry.

Evaluation of Ogataea (Hansenula) polymorpha for Hyaluronic Acid Production.

Hyaluronic acid (HA) is a biopolymer formed by UDP-glucuronic acid and UDP-N-acetyl-glucosamine disaccharide units linked by ß-1,4 and ß-1,3 glycosidic bonds. It is widely employed in medical and cosmetic procedures. HA is synthesized by hyaluronan synthase (HAS), which catalyzes the precursors' ligation in the cytosol, elongates the polymer chain, and exports it to the extracellular space. Here, we engineer Ogataea (Hansenula) polymorpha for HA production by inserting the genes encoding UDP-glucose 6-dehydrogenase, for UDP-glucuronic acid production, and HAS. Two microbial HAS, from Streptococcus zooepidemicus (hasAs) and Pasteurella multocida (hasAp), were evaluated separately. Additionally, we assessed a genetic switch using integrases in O. polymorpha to uncouple HA production from growth. Four strains were constructed containing both has genes under the control of different promoters. In the strain containing the genetic switch, HA production was verified by a capsule-like layer around the cells by scanning electron microscopy in the first 24 h of cultivation. For the other strains, the HA was quantified only after 48 h and in an optimized medium, indicating that HA production in O. polymorpha is limited by cultivation conditions. Nevertheless, these results provide a proof-of-principle that O. polymorpha is a suitable host for HA production.

Use of Phanerochaete Chrysosporium in a Semibatch Reactor for Removal of Indigo Carmine from Synthetic Textile Wastewater

Abstract The textile industry demonstrates a polluting potential from the planting of cotton to the release of wastewater. The presence of dyes in water bodies decreases the passage of sun rays and directly affects the photosynthetic organisms and the ecosystem. Fungi have potential in the treatment of wastewater containing dyes with complex organic structures due to enzymes that they produce. This study evaluated the use of Phanerochaete chrisosporium in the treatment of synthetic effluent from textile industry containing indigo carmine (20 mg/L). The fungus was immobilized in a semibatch reactor. Glucose was the cosubstrate employed in the experiment and it was used in the system at 1g/L at the beginning of the process and 0.5 g /L after 24 hours of reaction. Average dye removal was 84±10% and chemical oxygen demand removal was 79±14%. For nitrogen compounds, the removal efficiencies were 87±11%, 81±11% and 91±9% for ammonia, nitrite and nitrate, respectively. The pH of the medium remained in the acidic range (2.57 to 5.00) throughout the process, with the lowest values recorded in the effluent of each cycle, justified by the release of organic acids from fungi metabolism. There was contamination of the medium by bacteria (710,000 CFU/mL), but the colonies count showed a predominance of fungi (1,365,000 CFU/mL). With the use of the semibatch system after reading of glucose it was observed that the efficiency of dye removal evolved from 72±17% to 84±10%, producing a final effluent with 3.35±1.99 mg/L of indigo, which proves that treatment configuration analyzed is satisfactory for dye removal.

Physicochemical characterization and microbiological analysis of Oligosarcus robustus raw fish meat and development a fish hamburger / Caracterización físico-química y análisis microbiológico de la carne cruda de pescado Oligosarcus robustus y desarrollo de una hamburguesa de pescado

ABSTRACT This study aimed to evaluate the microbiological, physicochemical and fatty acid profile of the of "Tambica" (Oligosarcus robustus) raw fish meat and develop and characterize a fish burger. The fish burger was subjected to a sensory analysis. The raw fish meat showed a high content of moisture (82.3%) and minerals (3.1%), a low content of fat (1.5%) and a considerable protein content (13.1%). The raw fish showed a high count of positive coagulase Staphylococcus. Tambica lipid fraction was composed of 41.9% unsaturated and 58.2% saturated fatty acids. Palmitic acid and oleic acid were the major fatty acids in the raw meat fish. The fish burger was well accepted by sensory analysis.

RESUMEN Este estudio tuvo como objetivo evaluar el perfil microbiológico, fisicoquímico y de ácidos grasos de la carne de pescado cruda "Tambica" (Oligosarcus robustus); y desarrollar y caracterizar una hamburguesa de pescado. La hamburguesa de pescado fue sometida a análisis sensorial. La carne de pescado cruda mostró un alto contenido de humedad (82,3%) y minerales (3,1%), bajo contenido de grasa (1,5%) y un contenido de proteína considerable (13,1%). El pescado crudo mostró un recuento alto de Staphylococcus coagulasa positivo. La fracción lipídica de Tambica estaba compuesta por un 41,9% de ácidos grasos insaturados y 58,2% de ácidos grasos saturados. El ácido palmítico y el ácido oleico fueron los principales ácidos grasos en el pescado de carne cruda. La hamburguesa de pescado fue bien aceptada por el análisis sensorial.

Exploring the Brazilian diversity of Aspergillus sp. strains for lovastatin and itaconic acid production.

Filamentous fungi are well known for producing secondary metabolites applied in various industrial segments. Among these, lovastatin and itaconic acid, produced by Aspergillus terreus, have applications in the pharmaceutical and chemical industries. Lovastatin is primarily used for the control of hypercholesterolemia, while itaconic acid is a building block for the production of synthetic fibers, coating adhesives, among others. In this study, for the first time, 35 strains of Aspergillus sp. from four Brazilian culture collections were evaluated for lovastatin and itaconic acid production and compared to a reference strain, ATCC 20542. From an initial screening, the strains ATCC 20542, URM 224, URM1876, URM 5061, URM 5254, URM 5256, URM 5650, and URM 5961 were selected for genomic comparison. Among tested strains, the locus corresponding to the lovastatin genomic cluster was assembled, showing that all genes essential for lovastatin biosynthesis were present in producing URM 5961 and URM 5650 strains, with 100% and 98.5% similarity to ATCC 20542, respectively. However, in the no producing URM 1876, URM 224, URM 5254, URM 5061, and URM 5256 strains, this cluster was either fragmented or missing. Among the 35 strains evaluated for itaconic acid production in this study, only three strains had titers above 0.5 g/L, 16 strains had production below 0.5 g/L, and the remaining 18 strains had no production, with the highest production of itaconic acid observed in the URM 5254 strain with 2.2 g/L. The essential genes for itaconic acid production, mttA, cadA msfA were also mapped, where all three genes linked to itaconic acid production were found in a single contig in the assembly of each strain. In contrast to lovastatin loci, there is no correlation between the level of itaconic acid production and genetic polymorphisms in the genes associated with its biosynthesis.

The role of Optimistic Bias in safe food handling behaviours in the food service sector.

This study evaluates the influences on food handlers' intention to conduct safe handling behaviours in food service settings in Brazil and the United Kingdom, focusing on Optimistic bias (OB). Although OB has been identified in food handlers in other studies, this is the first study evaluating the role of OB in determining behavioural intention. Three hundred participants, from the United Kingdom (n = 150) and Brazil (n = 150), completed a Theory of Planned Behaviour - based questionnaire, including additional questions about OB. For Brazilian food handlers, Attitude had a significant influence on Behavioural Intention followed by Subjective Norm, while Perceived Behavioural Control and OB did not significantly influence Behavioural Intention. For the United Kingdom, Perceived Behavioural Control had the largest influence, followed by Attitude, and then OB. Results suggest that cultural differences may influence predictors of behavioural intention, and that OB can contribute to predicting engagement in safe handling behaviours. Results contribute to a better understanding of the influences on food safety related behaviours.

Emprego de Aspergillus niger AN 400 em reatores de bancada para remover pesticida de matriz aquosa / Employment of Aspergillus niger AN 400 in batch reactors to remove pesticide aqueous matrix

RESUMO Esta pesquisa propõe utilizar o fungo Aspergillus niger AN 400 no tratamento de matriz aquosa contendo metil paration por meio de reatores em batelada, sendo dividida em duas etapas. Na etapa I, foram montados reatores controle (RCs) contendo apenas metil paration em matriz aquosa; reatores contendo metil paration e suspensão de esporos de fungo (RFs); e reatores contendo metil paration, suspensão de esporos de fungo e etanol (RFEs). A cada tempo de reação (TR), um RC, um RF e um RFE eram colocados fora de operação. Os TRs estudados foram distribuídos em 30 dias. Na etapa II, operou-se um reator com biomassa imobilizada (RBI) alimentado por metil paration e etanol, efetuando-se cinco ciclos cada um com duração de uma semana. Como resultados, na Etapa I, verificou-se que o pH variou de 5 a 8, atingindo remoções finais de 61% (RF) e 89% (RFE) de demanda química de oxigênio (DQO) bruta; 67% (RF) e 52% (RFE) de nitrogênio amoniacal; 34% (RF) e 45% (RFE) de nitrato; 2% (RF) e 15% (RFE) de fósforo total. Na Etapa II, o pH variou de 3 a 8 e as médias de remoção final foram: 94% de DQO filtrada, 88% de DQO bruta, 19% de nitrogênio amoniacal, 96% de nitrato, 12% de fósforo total e metil paration de 95%, com biomassa imobilizada e 73% (95%) com biomassa dispersa. Os resultados mostraram que a utilização do fungo Aspergillus niger AN 400 pode ser uma alternativa no tratamento de água residuária contendo metil paration.

ABSTRACT This research proposes the use of the fungus Aspergillus niger AN 400 in treating aqueous matrix containing methyl parathion in batch reactors. The research was divided into two steps. In Step I, were mounted control reactors (RC) containing only methyl parathion in aqueous matrix; reactors with fungus (RF) containing methyl parathion and suspension of fungus spores; and reactors with fungus and ethanol (RFE) containing methyl parathion, the suspension of fungus spores and ethanol. Each reaction time (TR), one RC, one RF and one RFE were put out of operation. The reaction times were studied in 30 days. In Step II, operated a reactor with immobilized biomass (RBI) with methyl parathion and ethanol, making up 5 cycles each lasting one week. As a result, in Step I, it was found that the pH varied from 5 to 8. The final removal of 61% (RF) and 89% (RFE) of COD; 67% (RF) and 52% (RFE) of ammonia nitrogen; 34% (RF) and 45% (RFE) nitrate; 2% (RF) and 15% (RFE) of total phosphorus. In Step II, the pH ranged from 3 to 8 and the average final removal were 94% of filtered COD, 88% gross COD, 19% of ammonia nitrogen, 96% nitrate, 12% of total phosphorus and 95% of methyl parathion with immobilized biomass and 73% with disperse biomass. The results showed that the use of the fungus Aspergillus niger AN 400 can be an alternative for treating wastewater containing methyl parathion.

Altered Developmental Expression of the Astrocyte-Secreted Factors Hevin and SPARC in the Fragile X Mouse Model.

Astrocyte dysfunction has been indicated in many neurodevelopmental disorders, including Fragile X Syndrome (FXS). FXS is caused by a deficiency in fragile X mental retardation protein (FMRP). FMRP regulates the translation of numerous mRNAs and its loss disturbs the composition of proteins important for dendritic spine and synapse development. Here, we investigated whether the astrocyte-derived factors hevin and SPARC, known to regulate excitatory synapse development, have altered expression in FXS. Specifically, we analyzed the expression of these factors in wild-type (WT) mice and in fragile X mental retardation 1 (Fmr1) knock-out (KO) mice that lack FMRP expression. Samples were collected from the developing cortex and hippocampus (regions of dendritic spine abnormalities in FXS) of Fmr1 KO and WT pups. Hevin and SPARC showed altered expression patterns in Fmr1 KO mice compared to WT, in a brain-region specific manner. In cortical tissue, we found a transient increase in the level of hevin in postnatal day (P)14 Fmr1 KO mice, compared to WT. Additionally, there were modest decreases in Fmr1 KO cortical levels of SPARC at P7 and P14. In the hippocampus, hevin expression was much lower in P7 Fmr1 KO mice than in WT. At P14, hippocampal hevin levels were similar between genotypes, and by P21 Fmr1 KO hevin expression surpassed WT levels. These findings imply aberrant astrocyte signaling in FXS and suggest that the altered expression of hevin and SPARC contributes to abnormal synaptic development in FXS.

Remoção de BTEX por fungos em reator aeróbio de escoamento contínuo / BTEX removal by fungi in continuous flow aerobic reactor

RESUMO Foi estudada a remoção de compostos BTEX (benzeno, tolueno, etilbenzeno e xilenos) em reator de escoamento contínuo e mistura perfeita, com inóculo de Aspergillus niger AN 400, operado sob o tempo de detenção hidráulica (TDH) de 12 horas para avaliar a eficiência do sistema na biorremediação de água poluída com gasolina na presença (Etapa I) e na ausência (Etapa II) de glicose (0,5 g.L-1). A água poluída com gasolina foi preparada na proporção de 10:1000 mL (gasolina:água). Na primeira etapa foi ainda estudada a influência da adição de nutrientes no afluente sobre a eficiência do processo. A adição de nutrientes foi benéfica ao processo e, mesmo com a retirada da glicose, a eficiência foi mantida, devido à metabolização do etanol - presente na gasolina brasileira em 25% - pelos fungos, sendo ambos fontes de carbono de assimilação mais fácil para obtenção de energia; o etanol e a glicose, foram usados como cossubstratos na degradação dos BTEX. Assim, na Etapa 2, quando somente havia o etanol no meio, chegou-se a percentuais médios de 88% para benzeno, 90% para tolueno, 90% para etilbenzeno, e de 91% para meta, para e orto-xileno (m, p e o-xileno), com formação de subprodutos fenólicos. A população fúngica predominou no meio ao longo de toda a operação do reator.

ABSTRACT The removal of benzene, toluene, ethylbenzene and xylene (BTEX) compounds by Aspergillus niger AN 400 was studied in a continuous flow reactor which was operated at hydraulic retention time (HRT) of 12 hours evaluate the efficiency of the system in the bioremediation of water polluted with gasoline in the presence (Step 1) and absence (Step 2) of 0.5 g.L-1 glucose. In the first step it was also studied the influence of the addition of nutrients in the influent on the efficiency of the process. The addition of nutrients was beneficial to the process and, with the removal of glucose, the efficiency was maintained due to the use of ethanol - which is a Brazilian gasoline constituent (25%) - and was used as co-substrate. Both ethanol and glucose are more easily assimilated carbon and energy sources for fungi and were used as co-substrate in the degradation of BTEX. Thus, in Step 2, when only ethanol was in the biomedia, averages percentage of 88% for benzene, 90% in toluene, 90% ethylbenzene and 91% for meta, para and ortho-xylene (m, p and o-xylene) were reached, with the formation of phenolic by-products in between. The fungal population in the middle prevailed throughout the operation of the reactor.

Recombinant expression of Thermobifida fusca E7 LPMO in Pichia pastoris and Escherichia coli and their functional characterization.

The discovery of lytic polysaccharides monooxygenases copper dependent (LPMOs) revolutionized the classical concept that the cleavage of cellulose is a hydrolytic process in recent years. These enzymes carry out oxidative cleavage of cellulose (and other polysaccharides), acting synergistically with cellulases and other hydrolases. In fact, LPMOs have the potential for increasing the efficiency of the lignocellulosic biomass conversion in biofuels and high value chemicals. Among a small number of microbial LPMOs that have been characterized, some LPMOs were expressed and characterized biochemically from the bacteria Thermobifida fusca, using the host Escherichia coli. In this work, the E7 LPMO protein of T. fusca was expressed both in E. coli (native DNA sequence) and Pichia pastoris (codon-optimized DNA sequence), for further analysis of oxidative cleavage, with PASC (phosphoric acid swollen cellulose) and Avicel PH-101 substrates, using mass spectrometry analysis. Mass spectra results of Avicel PH-101 and PASC cleavages by purified E7 LPMO expressed in E. coli and in P. pastoris allowed the visualization of compounds corresponding to oxidized and non-oxidized oligosaccharides. Further optimization of reactions will be performed, since it was found only one degree of polymerization (DP 7). This work demonstrated that it is possible to produce the E7 LPMO from T. fusca in the host P. pastoris, and the recombinant protein was shown to be active on cellulose. The approach used in the present work could be an alternative to produce this bacterial LPMO for cellulose cleavage.

Complete genome sequence of tobacco mosqueado virus.

We describe the genomic characteristics of a new potyvirus isolated from tobacco plants showing mottling ("mosqueado" in Portuguese) in southern Brazil. The complete genomic sequence consists of 9896 nucleotides, without the poly(A) tail, and shares the highest pairwise nucleotide sequence identities of 68.5 % with pepper yellow mosaic virus and 68.2 % with Brugmansia mosaic virus isolate D437. These identity values are below the level of 76.0 % used as a criterion for species demarcation in the genus Potyvirus based on the complete genome sequence. The viral genomic organization and sequence comparison thus suggest that this virus, tentatively named "tobacco mosqueado virus" (TMosqV), represents a new potyvirus species.

Production of Brazilian human norovirus VLPs and comparison of purification methods.

Noroviruses (NVs) are responsible for most cases of human nonbacterial gastroenteritis worldwide. Some parameters for the purification of NV virus-like particles (VLPs) such as ease of production and yield were studied for future development of vaccines and diagnostic tools. In this study, VLPs were produced by the expression of the VP1 and VP2 gene cassette of the Brazilian NV isolate, and two purification methods were compared: cesium chloride (CsCl) gradient centrifugation and ion-exchange chromatography (IEC). IEC produced more and purer VLPs of NV compared to CsCl gradient centrifugation.

Production of Brazilian human norovirus VLPs and comparison of purification methods

Abstract Noroviruses (NVs) are responsible for most cases of human nonbacterial gastroenteritis worldwide. Some parameters for the purification of NV virus-like particles (VLPs) such as ease of production and yield were studied for future development of vaccines and diagnostic tools. In this study, VLPs were produced by the expression of the VP1 and VP2 gene cassette of the Brazilian NV isolate, and two purification methods were compared: cesium chloride (CsCl) gradient centrifugation and ion-exchange chromatography (IEC). IEC produced more and purer VLPs of NV compared to CsCl gradient centrifugation.

Influência do excesso de nitrogênio amoniacal no tratamento de efluente têxtil em reator de bateladas sequenciais com Aspergillus niger AN 400 / Influence of excess nitrogen in the treatment of the textile effluent in sequential batch reactors with Aspergillus nigerAN 400

RESUMO Água residuária têxtil foi tratada com Aspergillus niger AN 400 em reator de bateladas sequenciais, em ciclos de 48 h para avaliar o efeito de fonte adicional de nitrogênio sobre a eficiência do sistema alimentado com efluente diluído (10% v/v) e glicose (3 g.L-1). O afluente recebeu fonte adicional de nitrogênio (etapa I), acarretando em excesso de nitrogênio amoniacal (192 mg.L-1) no meio, resultando em remoção de matéria orgânica de 85%, porém em apenas 55% de remoção de corante. Ao se diminuir a concentração média de amônia para 98 mg.L-1 (etapa II), a remoção de corante (83%) passou a ser superior à de matéria orgânica, de 69%, mostrando o efeito negativo de concentrações elevadas de amônia sobre o sistema, favorecendo ainda o crescimento de bactérias, o que provavelmente inibiu a produção de enzimas fúngicas e contribuiu para a perda de eficiência na remoção de corante e formação de subprodutos.

ABSTRACT Textile wastewater was treated by Aspergillus niger AN 400 in sequential batch reactor, which was operated in cycles of 48 h in order to evaluate the effect of an additional source of nitrogen on the system's efficiency, which was fed with effluent diluted (10% v/v) and glucose (3 g.L-1). The influent received addition of an external source of nitrogen (phase I), in which there was excess ammonia (192 mg.L-1) in the medium, resulting in organic matter removal of 85%, but in only 55% removal of dye. In the phase II, when the average concentration of ammonia in the medium was lower (98 mg.L-1), removal of dye (83%) became higher than that of organic matter (69%), showing the negative effect of high concentrations of ammonia over the system and favoring the growth of bacteria, which probably inhibit the production of fungal enzymes and contributed to the loss of efficiency in the removal of dye and the formation of byproducts.

ASSESSMENT OF GOOD PRACTICES IN HOSPITAL FOOD SERVICE BY COMPARING EVALUATION TOOLS.

INTRODUCTION: since food service in hospitals complements medical treatment, it should be produced in proper hygienic and sanitary conditions. It is a well-known fact that food-transmitted illnesses affect with greater severity hospitalized and immunosuppressed patients. AIMS: good practices in hospital food service are evaluated by comparing assessment instruments. METHODS: good practices were evaluated by a verification list following Resolution of Collegiate Directory n. 216 of the Brazilian Agency for Sanitary Vigilance. Interpretation of listed items followed parameters of RCD 216 and the Brazilian Association of Collective Meals Enterprises (BACME). Fisher's exact test was applied to detect whether there were statistically significant differences. Analysis of data grouping was undertaken with Unweighted Pair-group using Arithmetic Averages, coupled to a correlation study between dissimilarity matrixes to verify disagreement between the two methods. RESULTS AND DISCUSSION: Good Practice was classified with mean total rates above 75% by the two methods. There were statistically significant differences between services and food evaluated by BACME instrument. Hospital Food Services have proved to show conditions of acceptable good practices. CONCLUSION: the comparison of interpretation tools based on RCD n. 216 and BACME provided similar results for the two classifications.

Introducción: por ser un adyuvante al tratamiento médico, la alimentación hospitalaria debe ser producida en adecuadas condiciones higiénicas y sanitarias, considerando qué enfermedades transmitidas por los alimentos afectan con mayor severidad a pacientes hospitalizados e inmunodeprimidos. Objetivos: evaluar la adopción de buenas prácticas en los servicios de alimentación hospitalarios, comparando instrumentos de evaluación. Métodos: la evaluación de las buenas prácticas fue realizada utilizando una lista de verificación conforme la Resolución de Directora Colegiada ­ RDC n.º 216 de la Agencia Nacional de Vigilancia Sanitaria. Para la interpretación de los ítems listados fueron utilizados los parámetros de la RDC n.º 216 y de la Asociación Brasileña de Empresas de Comidas Colectivas (ABERC). Se aplicó el test exacto de Ficher para analizar si existían diferencias estadísticamente significativas y también fue realizado un análisis de agrupamiento de los datos per el método Unweighted Pair-group using Arithmetic Averages y un estudio de la correlación entre las matrices de disimilitud, objetivando verificar la concordancia entre los dos métodos de diagnóstico. Resultados y discusión: las buenas prácticas fueron clasificadas con valores medios totales superiores al 75% de adecuación, en los dos instrumentos utilizados. Se observaron diferencias estadísticamente significativas entre los servicios y la alimentación evaluados por el instrumento de la ABERC. Los servicios de alimentación hospitalarios, de modo general, presentaron condiciones de buenas prácticas aceptables. Conclusión: la comparación de los instrumentos de interpretación basados en la RDC n.º 216 y ABERC mostró resultados semejantes en ambas clasificaciones.

MICROBIOLOGICAL ASSESSMENT OF LETTUCE SALADS AND ANTIMICROBIAL RESISTANCE OF STAPHYLOCOCCUS SPP.

INTRODUCTION: self-service restaurants in which food is served ready to be consumed are liable to have some products contaminated by pathogenic microorganisms causing food-transmitted diseases. AIM: evaluates the microbiological quality of lettuce salads in restaurants in Pelotas RS Brazil by counts of thermo-tolerant coliforms, E. coli, Staphylococcus spp. and detection of Salmonella spp. Antimicrobial resistance of Staphylococcus spp. isolates are also assessed. METHODS: thirty-six samples of lettuce salads were collected from nine restaurants and thermotolerant coliforms, Escherichia coli and Staphylococcus spp. were quantified, coupled to a research on Salmonella spp., following methodology by the Bacteriological Analytical Manual. Staphylococcus spp. isolates underwent antimicrobial resistance test by the disc-diffusion method. RESULTS AND DISCUSSION: results showed that 61.1% of the salad samples contained more thermotolerant coliforms than allowed by Brazilian legislation and E. coli was confirmed in 5.6% of the samples. Positive and negative coagulase Staphylococcus occurred respectively in 5.6% and 77.8% of isolates, but no sample had Salmonella spp. Further, 56.7% of the thirty isolates of Staphylococcus spp. tested were resistant to penicillin; 46.7% to oxacillin; 26.7% to erythromycin and 23.3% were multi- resistant. CONCLUSION: inadequate quality of the salad was due to pathogenic microorganisms, while Staphylococcus spp. isolates had a high percentage of antimicrobial resistance.

Introducción: la procura por estabelecimientos que ofrecen alimentos prontos para consumo ha aumentado, sin embrago, los alimentos disponibles en estos locales pueden estar contaminados con microorganismos patogénicos, pudiendo causar enfermedades transmitidas por alimentos. Objetivos: evaluar la calidad microbiológica de las ensaladas de lechuga en los restaurantes de Pelotas RS Brasil, a través de los recuentos de coliformes termotolerantes, Escherichia coli, Staphylococcus spp. y la detección de Salmonella spp. Resistencia a los antimicrobianos de Staphylococcus spp. también se evalúan. Métodos: fueron colectadas 36 muestras de ensaladas de lechuga en nueve restaurantes y realizada la cuantificación de coliformes termotolerantes, Escherichia coli y Staphylococcus spp. e investigación de Salmonella spp., siguiendo la metodología del Bacteriological Analytical Manual. Los aislados de Staphylococcus spp. fueron sometidos al examen de resistencia a antimicrobianos por el método de difusión con discos. Resultados y discusión: de las 36 muestras de ensalada de lechuga, 61,1% presentaron cuantificación de coliformes termotolerantes superiores a lo permitido por la legislación brasileña, y hubo confirmación de E. coli en 5,6% de las muestras. La cuantificación de Staphylococcus coagulasa positiva representó 5,6% de los aislados y Staphylococcus coagulasa negativa representó 77,8%. Todas las muestras presentaron ausencia de Salmonella spp. De los 30 aislados de Staphylococcus spp. examinados, 56,7% fueron resistentes a penicilina, 46,7% a oxacilina, 26,7% a eritromicina y 23,3% fueron multirresistentes. Conclusión: la calidad microbiológica de las ensaladas de lechuga se mostró inadecuada debido a la presencia de microorganismos patogénicos, y los aislados de Staphylococcus spp. presentaron elevado porcentaje de resistencia antimicrobiana.