Functional characterization and target discovery of glycoside hydrolases from the digestome of the lower termite Coptotermes gestroi

<p>Abstract</p> <p>Background</p> <p>Lignocellulosic materials have been moved towards the forefront of the biofuel industry as a sustainable resource. However, saccharification and the production of bioproducts derived from plant cell wall biomass are complex and lengthy processes. The understanding of termite gut biology and feeding strategies may improve the current state of biomass conversion technology and bioproduct production.</p> <p>Results</p> <p>The study herein shows comprehensive functional characterization of crude body extracts from <it>Coptotermes gestroi </it>along with global proteomic analysis of the termite's digestome, targeting the identification of glycoside hydrolases and accessory proteins responsible for plant biomass conversion. The crude protein extract from <it>C. gestroi </it>was enzymatically efficient over a broad pH range on a series of natural polysaccharides, formed by glucose-, xylose-, mannan- and/or arabinose-containing polymers, linked by various types of glycosidic bonds, as well as ramification types. Our proteomic approach successfully identified a large number of relevant polypeptides in the <it>C. gestroi </it>digestome. A total of 55 different proteins were identified and classified into 29 CAZy families. Based on the total number of peptides identified, the majority of components found in the <it>C. gestroi </it>digestome were cellulose-degrading enzymes. Xylanolytic enzymes, mannan- hydrolytic enzymes, pectinases and starch-degrading and debranching enzymes were also identified. Our strategy enabled validation of liquid chromatography with tandem mass spectrometry recognized proteins, by enzymatic functional assays and by following the degradation products of specific 8-amino-1,3,6-pyrenetrisulfonic acid labeled oligosaccharides through capillary zone electrophoresis.</p> <p>Conclusions</p> <p>Here we describe the first global study on the enzymatic repertoire involved in plant polysaccharide degradation by the lower termite <it>C. gestroi</it>. The biochemical characterization of whole body termite extracts evidenced their ability to cleave all types of glycosidic bonds present in plant polysaccharides. The comprehensive proteomic analysis, revealed a complete collection of hydrolytic enzymes including cellulases (GH1, GH3, GH5, GH7, GH9 and CBM 6), hemicellulases (GH2, GH10, GH11, GH16, GH43 and CBM 27) and pectinases (GH28 and GH29).</p

A Phage-Like Plasmid Carrying blaKPC-2 Gene in Carbapenem-Resistant Pseudomonas aeruginosa

Background: Lateral gene transfer plays a central role in the dissemination of carbapenem resistance in bacterial pathogens associated with nosocomial infections, mainly Enterobacteriaceae and Pseudomonas aeruginosa. Despite their clinical significance, there is little information regarding the mobile genetic elements and mechanism of acquisition and propagation of lateral genes in P. aeruginosa, and they remain largely unknown.Objectives: The present study characterized the genetic context of blaKPC-2 in carbapenem-resistant P. aeruginosa strain BH9.Methods:Pseudomonas aeruginosa BH9 sequencing was performed using the long-read PacBio SMRT platform and the Ion Proton System. De novo assembly was carried out using the SMRT pipeline and Canu, and gene prediction and annotation were performed using Prokka and RAST.Results:Pseudomonas aeruginosa BH9 exhibited a 7.1 Mb circular chromosome. However, the blaKPC-2 gene is located in an additional contig composed by a small plasmid pBH6 from P. aeruginosa strain BH6 and several phage-related genes. Further analysis revealed that the beginning and end of the contig contain identical sequences, supporting a circular plasmid structure. This structure spans 41,087 bp, exhibiting all the Mu-like phage landmarks. In addition, 5-bp direct repeats (GGATG) flanking the pBH6 ends were found, strongly indicating integration of the Mu-like phage into the pBH6 plasmid. Mu phages are commonly found in P. aeruginosa. However, for the first time showing a potential impact in shaping the vehicles of the dissemination of antimicrobial (e.g., plasmid pBH6) resistance genes in the Pseudomonas genus.Conclusion: pBH6 captured the Mu-like Phage BH9, creating a co-integrate pBH6::Phage BH9, and this phage-plasmid complex may represent novel case of a phage-like plasmid

Biorefining Potential of Wild-Grown Arundo donax, Cortaderia selloana and Phragmites australis and the Feasibility of White-Rot Fungi-Mediated Pretreatments

Arundo donax, Cortaderia selloana and Phragmites australis are high-biomass-producing perennial Poalean species that grow abundantly and spontaneously in warm temperate regions, such as in Mediterranean-type climates, like those of Southern Europe, Western United States coastal areas, or in regions of South America, South Africa and Australia. Given their vigorous and spontaneous growth, biomass from the studied grasses often accumulates excessively in unmanaged agro-forestry areas. Nonetheless, this also creates the demand and opportunity for the valorisation of these biomass sources, particularly their cell wall polymers, for biorefining applications. By contrast, a related crop, Miscanthus × giganteus, is a perennial grass that has been extensively studied for lignocellulosic biomass production, as it can grow on low-input agricultural systems in colder climates. In this study Fourier transform mid-infrared spectroscopy (FTIR), high-performance anion-exchange chromatography (HPAEC) and lignin content determinations were used for a comparative compositional characterisation of A. donax, C. selloana and P. australis harvested from the wild, in relation to a trial field-grown M. × giganteus high-yielding genotype. A high-throughput saccharification assay showed relatively high sugar release values from the wild-grown grasses, even with a 0.1M NaOH mild alkali pretreatment. In addition to this alkaline pretreatment, biomass was treated with white-rot fungi (WRF), which preferentially degrade lignin more readily than holocellulose. Three fungal species were used: Ganoderma lucidum, Pleurotus ostreatus and Trametes versicolor. Our results showed that neutral sugar contents are not significantly altered, while some lignin is lost during the pretreatments. Furthermore, sugar release upon enzymatic saccharification was enhanced, and this was dependent on the plant biomass and fungal species used in the treatment. To maximise the potential for lignocellulose valorisation, the liquid fractions from the pretreatments were analysed by high performance liquid chromatography – photodiode array detection – electrospray ionisation tandem mass spectrometry (HPLC-PDA-ESI-MS(n)). This study is one of the first to report on the composition of WRF-treated grass biomass, while assessing the potential relevance of breakdown products released during the treatments, beyond more traditional sugar-for-energy applications. Ultimately, we expect that our data will help promote the valorisation of unused biomass resources, create economic value, while contributing to the implementation of sustainable biorefining systems

Pathophysiological Mechanisms In Gaseous Therapies For Severe Malaria.

Over 200 million people worldwide suffer from malaria every year, a disease that causes 584,000 deaths annually. In recent years, significant improvements have been achieved on the treatment of severe malaria, with intravenous artesunate proving superior to quinine. However, mortality remains high at 8% in children and 15% in adults in clinical trials, and even worse in the case of cerebral malaria (18% and 30%, respectively). Moreover, some individuals who do not succumb to severe malaria present long-term cognitive deficits. These observations indicate that strategies focused only on parasite killing fail to prevent neurological complications and deaths associated with severe malaria, possibly because clinical complications are associated in part with a cerebrovascular dysfunction. Consequently, different adjunctive therapies aimed at modulating malaria pathophysiological processes are currently being tested. However, none of these therapies has shown unequivocal evidence in improving patients' clinical status. Recently, key studies have shown that gaseous therapies based mainly on nitric oxide (NO), carbon monoxide (CO) and hyperbaric (pressurized) oxygen (HBO) alter vascular endothelium dysfunction and modulate host immune response to infection. Considering gaseous administration as a promising adjunctive treatment against severe malaria cases, we review here the pathophysiological mechanisms and the immunological aspects of such therapies.8

Expanding the knowledge on lignocellulolytic and redox enzymes of worker and soldier castes from the lower termite coptotermes gestroi

Termites are considered one of the most efficient decomposers of lignocelluloses on Earth due to their ability to produce, along with its microbial symbionts, a repertoire of carbohydrate-active enzymes (CAZymes). Recently, a set of Pro-oxidant, Antioxidant, and Detoxification enzymes (PAD) were also correlated with the metabolism of carbohydrates and lignin in termites. The lower termite Coptotermes gestroi is considered the main urban pest in Brazil, causing damage to wood constructions. Recently, analysis of the enzymatic repertoire of C. gestroi unveiled the presence of different CAZymes. Because the gene profile of CAZy/PAD enzymes endogenously synthesized by C. gestroi and also by their symbiotic protists remains unclear, the aim of this study was to explore the eukaryotic repertoire of these enzymes in worker and soldier castes of C. gestroi. Our findings showed that worker and soldier castes present similar repertoires of CAZy/PAD enzymes, and also confirmed that endo-glucanases (GH9) and beta-glucosidases (GH1) were the most important glycoside hydrolase families related to lignocellulose degradation in both castes. Classical cellulases such as exo-glucanases (GH7) and endo-glucanases (GH5 and GH45), as well as classical xylanases (GH10 and GH11), were found in both castes only taxonomically related to protists, highlighting the importance of symbiosis in C. gestroi. Moreover, our analysis revealed the presence of Auxiliary Activity enzyme families (AAs), which could be related to lignin modifications in termite digestomes. In conclusion, this report expanded the knowledge on genes and proteins related to CAZy/PAD enzymes from worker and soldier castes of lower termites, revealing new potential enzyme candidates for second-generation biofuel processes7CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICO - CNPQFUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULO - FAPESP140796/2013-4; 310186/2014-5; 442333/2014-511/20977-3; 15/06971-3; 12/19040-0; 14/10351-8; 06/59086-8; 14/20576- 7; 13/03061-0; 10/11469-1; 08/58037-9; 14/50371-8; 08/50114-

Sucesso adaptativo e percepções das perturbações hidrológicas por populações ribeirinhas em rios do semiárido brasileiro

Brazilian semi-arid streams are characterized by extreme hydrological events, with short periods of flooding and long periods of no surface water flow. Human populations living in the surroundings of these systems must be adapted to such hydrological disturbances. This paper evaluates the perceptions of riverine human populations in the Brazilian semi-arid, in relation to the hydrological disturbances, adaptations of such populations to withstand drought while maintaining social structure, and their utilization of the watershed during times of flooding. Data presented was collected from three riverine human populations that live near intermittent streams in the semi-arid region of Brazil. These populations rely mostly on the intermittent water supply from these systems and on the production from small-scaled agriculture and livestock. The surveyed populations maintain the stability of their social structures and organizations by producing an identification process through representations of hydrological disturbances (floods and drought). The resistance and resilience to these events basically involve strategies to avoid social ruptures, through the practice of subsistence agriculture, and the search for opportunities (e.g. adequate sites) to build weirs, wells and “cacimbas” (water holes). A simple yet well-structured scheme to utilize the watershed was found to be important during wet periods. The history of human adaptation to the Brazilian semi-arid depends on the configurations that both ecosystem and the social system take at each hydrological cycle. These aspects should be taken into account by decision-makers, and should be included in development-oriented policies adopted for the region. Any attempt to manage the semi-arid ecosystems should include the utilization of hydrological disturbances as part of the human element and its dimensions. Key words: riverine populations, survival strategies, flood, drought, stability, intermittent stream, semi-arid.Os rios do semiárido brasileiro são caracterizados por extremos hidrológicos, com curtos períodos de inundação e longos períodos de ausência de água. Populações humanas vivendo próximas a esses sistemas devem estar adaptadas a tais perturbações hidrológicas. Este trabalho avalia a percepção de populações ribeirinhas no semiárido brasileiro, considerando as perturbações hidrológicas, suas adaptações para resistir à seca enquanto mantêm a estrutura social e o uso da água durante os períodos de inundação. Este estudo foi realizado em três populações que vivem próximas a rios intermitentes na região semiárida do Brasil. Essas populações dependem principalmente da água de rios intermitentes para a agricultura em pequena escala e para a pecuária. As populações estudadas mantêm a estabilidade de suas estruturas e organizações sociais por meio das representações das perturbações hidrológicas (inundação e seca). A resistência e a resiliência a esses eventos envolvem basicamente estratégias para evitar rupturas sociais através da prática de agricultura de subsistência e pela procura de oportunidades (locais adequados) para construir açudes, poços e cacimbas. Um esquema simples e bem estruturado de utilização de água é também importante durante períodos de inundação. O histórico da adaptação humana no semiárido brasileiro resulta da conformação do ecossistema e do sistema social a cada ciclo hidrológico. Estes aspectos devem ser levados em conta por tomadores de decisões e devem ser considerados nas políticas que orientam o desenvolvimento dessa região. As ações de manejo nos ecossistemas do semiárido devem incluir a utilização das perturbações hidrológicas como parte do elemento humano e suas dimensões. Palavras-chave: populações ribeirinhas, estratégias de sobrevivência, inundação, seca, estabilidade, rio intermitente, semiárido

FTIR Screening to Elucidate Compositional Differences in Maize Recombinant Inbred Lines with ContrastingSaccharification Efficiency Yields

With a high potential to generate biomass, maize stover arises as an outstanding feedstock for biofuel production. Maize stover presents the added advantage of being a multiple exploitation of the crop as a source of food, feed, and energy. In this study, contrasting groups of recombinant inbred lines (RILs) from a maize multiparent advanced generation intercross (MAGIC) population that showed variability for saccharification efficiency were screened by FTIR-ATR spectroscopy to explore compositional differences between high and low saccharification yielders. High and low saccharification efficiency groups differed in cell wall compositional features: high saccharification RILs displayed higher proportions of S subunits, aromatic compounds, and hemicellulose as opposed to low saccharification efficiency groups in which FTIR predicted higher proportions of lignin, more precisely lignin being richer in subunits G, and greater proportions of crystalline cellulose and acetyl methyl esters. The application of FTIR-ATR spectroscopy in this material allowed us to obtain a rapid and broad vision of cell wall compositional features in contrasting groups of saccharification efficiency. These results helped us to deepen our knowledge into the relationship between cell wall composition and biorefining potential; they also allowed us to establish new targets for future research regarding lignocellulosic bioconversion

Ações de Terapia Ocupacional no território da cultura: a experiência de cooperação entre o Museu de Arte Contemporânea da USP (MAC USP) e o Laboratório de Estudos e Pesquisas Arte e Corpo em Terapia Ocupacional

This article's aim is to report a collaborative experience, started in 1996, between the Laboratory of Studies and Research in Art, Body and Occupation Therapy and the Museum of Contemporary Art of USP (MAC USP), by focusing on some actions and their unfolding for the construction of Occupational Therapy practices in the cultural field. For 10 years, joint actions in teaching, researching and extending services to the community were developed. The experience made it possible to follow up groups of elderly people and people in vulnerability, in proposals that articulate doing art, appreciate and contextualize artistic practices in art history. It made possible, also, deepen studies on contemporary art and the comprehension of occupational therapists' role to build up the access of vulnerable populations to the artistic-cultural field.Este trabalho se propõe a relatar a experiência de colaboração entre o Laboratório de Estudos e Pesquisas Arte e Corpo em Terapia Ocupacional e o Museu de Arte Contemporânea da USP (MAC USP), que teve início em 1996, focalizando algumas ações significativas e seus desdobramentos para a construção de práticas de Terapia Ocupacional no campo cultural. Durante mais de 10 anos foram desenvolvidas ações conjuntas nas esferas do ensino, da pesquisa e da extensão de serviços à comunidade. A experiência possibilitou o acompanhamento de grupos de idosos e pessoas em situação de vulnerabilidade, em propostas que compreendem o fazer artístico, a apreciação e a contextualização de práticas estéticas na história da arte; propiciou, também, o aprofundamento de estudos sobre arte contemporânea e a compreensão do papel do terapeuta ocupacional na construção de acesso para populações vulneráveis ao campo artístico-cultural

Setting performance indicators for coastal marine protected areas: An expert-based methodology

Marine Protected Areas (MPAs) require effective indicators to assess their performance, in compliance with the goals of relevant national and international commitments. Achieving and prioritizing shortlists of multidisciplinary indicators demands a significant effort from specialists to depict the multiple conservation and socioeconomic interests, and the large complexity of natural systems. The present paper describes a structured expert-based methodology (process and outputs) to co-define a list of multidisciplinary MPA performance indicators. This work was promoted by the management authority of coastal MPAs in mainland Portugal to gather a consensual and feasible list of indicators that would guide the design of a future national monitoring program. Hence, Portuguese coastal MPAs served as a case study to develop such a process between 2019 and 2020. In the end, participants (1) agreed on a shortlist of prioritized indicators (i.e., environmental, governance, and socioeconomic indicators) and (2) defined minimum monitoring frequencies for the indicators in this list, compatible with the potential replicability of the associated survey methods. The present approach recommends that management plans incorporate monitoring procedures and survey methods, with a validated list of indicators and associated monitoring periodicity, agreed among researchers, MPA managers and governance experts. The proposed methodology, and the lessons learned from it, can support future processes aiming to define and prioritize MPA performance indicatorsFundação para a Ciência e Tecnologia - FCT, European Maritime and Fisheries Fund (EMFF)info:eu-repo/semantics/publishedVersio