Legumes tend to be nodulated by competitive rhizobia that do not maximize nitrogen (N
) fixation, resulting in suboptimal yields. Rhizobial nodulation competitiveness and effectiveness at N
fixation ...are independent traits, making their measurement extremely time-consuming with low experimental throughput. To transform the experimental assessment of rhizobial competitiveness and effectiveness, we have used synthetic biology to develop reporter plasmids that allow simultaneous high-throughput measurement of N
fixation in individual nodules using green fluorescent protein (GFP) and barcode strain identification (Plasmid ID) through next generation sequencing (NGS). In a proof-of-concept experiment using this technology in an agricultural soil, we simultaneously monitored 84 different
strains, identifying a supercompetitive and highly effective rhizobial symbiont for peas. We also observed a remarkable frequency of nodule coinfection by rhizobia, with mixed occupancy identified in ∼20% of nodules, containing up to six different strains. Critically, this process can be adapted to multiple
-legume symbioses, soil types, and environmental conditions to permit easy identification of optimal rhizobial inoculants for field testing to maximize agricultural yield.
We present a Bacterial Expression Vector Archive (BEVA) for the modular assembly of bacterial vectors compatible with both traditional and Golden Gate cloning, utilizing the Type IIS restriction ...enzyme Esp3I, and have demonstrated its use for Golden Gate cloning in
. Ideal for synthetic biology and other applications, this modular system allows a rapid, low-cost assembly of new vectors tailored to specific tasks. Using the principles outlined here, new modules (e.g., origin of replication for plasmids in other bacteria) can easily be designed for specific applications. It is hoped that this vector construction system will be expanded by the scientific community over time by creation of novel modules through an open source approach. To demonstrate the potential of the system, three example vectors were constructed and tested. The Golden Gate level 1 vector pOGG024 (pBBR1-based broad-host range and medium copy number) was used for gene expression in laboratory-cultured
. The Golden Gate level 1 vector pOGG026 (RK2-based broad-host range, lower copy number and stable in the absence of antibiotic selection) was used to demonstrate bacterial gene expression in nitrogen-fixing nodules on pea plant roots formed by R.
. Finally, the level 2 cloning vector pOGG216 (RK2-based broad-host range, medium copy number) was used to construct a dual reporter plasmid expressing green and red fluorescent proteins.
Legumes tend to be nodulated by competitive rhizobia that do not maximize nitrogen (N₂) fixation, resulting in suboptimal yields. Rhizobial nodulation competitiveness and effectiveness at N₂ fixation ...are independent traits, making their measurement extremely time-consuming with low experimental throughput. To transform the experimental assessment of rhizobial competitiveness and effectiveness, we have used synthetic biology to develop reporter plasmids that allow simultaneous high-throughput measurement of N₂ fixation in individual nodules using green fluorescent protein (GFP) and barcode strain identification (Plasmid ID) through next generation sequencing (NGS). In a proof-of-concept experiment using this technology in an agricultural soil, we simultaneously monitored 84 different Rhizobium leguminosarum strains, identifying a supercompetitive and highly effective rhizobial symbiont for peas. We also observed a remarkable frequency of nodule coinfection by rhizobia, with mixed occupancy identified in ~20% of nodules, containing up to six different strains. Critically, this process can be adapted to multiple Rhizobium-legume symbioses, soil types, and environmental conditions to permit easy identification of optimal rhizobial inoculants for field testing to maximize agricultural yield.
Biological nitrogen fixation by
Rhizobium
-legume symbioses represents an environmentally friendly and inexpensive alternative to the use of chemical nitrogen fertilizers in legume crops. Rhizobial ...inoculants, applied frequently as biofertilizers, play an important role in sustainable agriculture. However, inoculants often fail to compete for nodule occupancy against native rhizobia with inferior nitrogen-fixing abilities, resulting in low yields. Strains with excellent performance under controlled conditions are typically selected as inoculants, but the rates of nodule occupancy compared to native strains are rarely investigated. Lack of persistence in the field after agricultural cycles, usually due to the transfer of symbiotic genes from the inoculant strain to naturalized populations, also limits the suitability of commercial inoculants. When rhizobial inoculants are based on native strains with a high nitrogen fixation ability, they often have superior performance in the field due to their genetic adaptations to the local environment. Therefore, knowledge from laboratory studies assessing competition and understanding how diverse strains of rhizobia behave, together with assays done under field conditions, may allow us to exploit the effectiveness of native populations selected as elite strains and to breed specific host cultivar-rhizobial strain combinations. Here, we review current knowledge at the molecular level on competition for nodulation and the advances in molecular tools for assessing competitiveness. We then describe ongoing approaches for inoculant development based on native strains and emphasize future perspectives and applications using a multidisciplinary approach to ensure optimal performance of both symbiotic partners.
Plants engineer the rhizosphere to their advantage by secreting various nutrients and secondary metabolites. Coupling transcriptomic and metabolomic analyses of the pea (Pisum sativum) rhizosphere, a ...suite of bioreporters has been developed in Rhizobium leguminosarum bv viciae strain 3841, and these detect metabolites secreted by roots in space and time. Fourteen bacterial lux fusion bioreporters, specific for sugars, polyols, amino acids, organic acids, or flavonoids, have been validated in vitro and in vivo. Using different bacterial mutants (nodC and nifH), the process of colonization and symbiosis has been analyzed, revealing compounds important in the different steps of the rhizobium-legume association. Dicarboxylates and sucrose are the main carbon sources within the nodules; in ineffective (nifH) nodules, particularly low levels of sucrose were observed, suggesting that plant sanctions affect carbon supply to nodules. In contrast, high myo-inositol levels were observed prior to nodule formation and also in nifH senescent nodules. Amino acid biosensors showed different patterns: a g-aminobutyrate biosensor was active only inside nodules, whereas the phenylalanine bioreporter showed a high signal also in the rhizosphere. The bioreporters were further validated in vetch (Vicia hirsuta), producing similar results. In addition, vetch exhibited a local increase of nod gene-inducing flavonoids at sites where nodules developed subsequently. These bioreporters will be particularly helpful in understanding the dynamics of root exudation and the role of different molecules secreted into the rhizosphere.
Exotic species pose a threat to most ecosystems because of their potential to establish negative interactions with native biota. However, exotic species can also offer resources to native species, ...especially within highly modified environments such as urban ecosystems. We studied 17 exotic-native pairs of species with the potential to compete with one another, or in which one of the species could offer resources to the other, in an urban ecological reserve located within Mexico City. We used two-species occupancy models to analyze the potential association between the presence of the exotic species and the spatial distribution of the native species, as well as to assess if these species tend to avoid each other (negative spatial interaction) or to co-occur more often than expected under the hypothesis of independent occurrences (positive spatial interaction). Our results revealed few cases in which the exotic species influenced occupancy of the native species, and these spatial interactions were mainly positive, indicated by the fact that the occupancy of the native species was usually higher when the exotic species was also present. Seven of the eight observed non-independent patterns of co-occurrence were evident during the dry months of the year, when resources become scarce for most species. Our results also demonstrate that the observed patterns of species co-occurrence depend on the distance to the nearest urban structure and the amount of herb, shrub, and tree cover, indicating that these habitat features influence whether native species avoid or co-occur with exotic species. Our study represents an important contribution to the understanding of temporal dynamics in the co-occurrence between exotic and native species within urban ecological reserves.
'Speed breeding' (SB) shortens the breeding cycle and accelerates crop research through rapid generation advancement. SB can be carried out in numerous ways, one of which involves extending the ...duration of plants' daily exposure to light, combined with early seed harvest, to cycle quickly from seed to seed, thereby reducing the generation times for some long-day (LD) or day-neutral crops. In this protocol, we present glasshouse and growth chamber-based SB approaches with supporting data from experimentation with several crops. We describe the conditions that promote the rapid growth of bread wheat, durum wheat, barley, oat, various Brassica species, chickpea, pea, grass pea, quinoa and Brachypodium distachyon. Points of flexibility within the protocols are highlighted, including how plant density can be increased to efficiently scale up plant numbers for single-seed descent (SSD). In addition, instructions are provided on how to perform SB on a small scale in a benchtop growth cabinet, enabling optimization of parameters at a low cost.
Non-nutritive sweeteners (NNS) are widely consumed by humans due to their apparent innocuity, especially sucralose. However, several studies link sucralose consumption to weight gain and metabolic ...derangements, although data are still contradictory.
To determine the effect of acute and chronic consumption of sucralose on insulin and glucose profiles in young healthy adults.
This was a randomized, parallel, double-blind, placebo-controlled trial conducted in healthy young adults from 18 to 35 years old, without insulin resistance. A hundred thirty seven participants were randomized into three groups: a) volunteers receiving 48 mg sucralose, b) volunteers receiving 96 mg sucralose, and c) controls receiving water as placebo. All participants underwent a 3-h oral glucose tolerance test (OGTT) preceded by consuming sucralose or placebo 15 min before glucose load, at two time points: week zero (Wk0) and week ten (Wk10). Serum insulin and glucose were measured every 15 min during both OGTTs.
Compared to Wk0, consumption of sucralose for 10 weeks provoked 1) increased insulin concentrations at 0 min (7.5 ± 3.4 vs 8.8 ± 4.1 μIU/mL; p = 0.01), 30 min (91.3 ± 56.2 vs 110.1 ± 49.4 μIU/mL; p = 0.05), 105 min (47.7 ± 24.4 vs 64.3 ± 48.2 μIU/mL; p = 0.04) and 120 min (44.8 ± 22.1 vs 63.1 ± 47.8 μIU/mL; p = 0.01) in the 48 mg sucralose group; 2) increased blood glucose at - 15 min (87.9 ± 4.6 vs 91.4 ± 5.4 mg/dL; p = 0.003), 0 min (88.7 ± 4 vs 91.3 ± 6 mg/dL; p = 0.04) and 120 min (95.2 ± 23.7 vs 106.9 ± 19.5 mg/dL; p = 0.009) in the 48 mg sucralose group; 3) increased area under the curve (AUC) of insulin in both 48 and 96 mg sucralose groups (9262 vs 11,398; p = 0.02 and 6962 vs 8394; p = 0.12, respectively); and 4) reduced Matsuda index in the 48 mg sucralose group (6.04 ± 3.19 vs 4.86 ± 2.13; p = 0.01).
These data show that chronic consumption of sucralose can affect insulin and glucose responses in non-insulin resistant healthy young adults with normal body mass index (between 18.5 and 24.9 kg/m
), however, the effects are not consistent with dose; further research is required.
NCT03703141.
Background: A diet containing non-caloric sweeteners (NCS) could reduce calorie intake; conversely, some animal studies suggest that NCS consumption may increase functional gastrointestinal disorder ...symptoms (FGDs). This study aimed to compare the effect of consuming a diet containing NCS (c-NCS) versus a non-caloric sweetener-free diet (NCS-f) on FGDs. Methods: We conducted a randomized, controlled, parallel-group study using two different diets for five weeks: the c-NCS diet contained 50−100 mg/day NCS, whereas the NCS-f diet had less than 10 mg/day NCS. At the beginning of the study (PreTx) and at the end (PostTx), we assessed FGDs, dietary intake, and NCS consumption. Results: The percentage of participants with diarrhea (PreTx = 19% vs. PstTx = 56%; p = 0.02), post-prandial discomfort (PreTx = 9% vs. PstTx = 39%; p = 0.02), constipation (PreTx = 30% vs. PostTx = 56%; p < 0.01), and burning (PreTx = 13% vs. PostTx = 33%; p < 0.01) increased in the c-NCS diet group. Conversely, abdominal pain (PreTx = 15% vs. PostTx = 3%; p = 0.04), post-prandial discomfort (PreTx = 26% vs. PostTx = 6%; p = 0.02), burning (PreTx = 15% vs. PostTx = 0%; p = 0.02), early satiety (PreTx = 18% vs. PostTx = 3%; p < 0.01), and epigastric pain (PreTx = 38% vs. PostTx = 3%; p < 0.01) decreased in the NCS-f diet group. Conclusion: A c-NCS diet is associated with increased FGDs, including diarrhea, post-prandial discomfort, constipation, and burning or retrosternal pain. The NCS-f diet also decreased FGDs, as well as abdominal pain, post-prandial discomfort, burning or retrosternal pain, early satiety, and epigastric pain.
Introduction: The COVID-19 pandemic posed unprecedented challenges to the nursing staff, who were at the forefront of patient care and their families. Exposed to the virus due to strenuous work ...schedules and other factors, their health was at risk. However, they took proactive self-care actions to counteract these factors, promote self-health, and prevent the onset of diseases caused by stressors from the pandemic. These actions, identified in the literature published between 2020-2023, have significantly impacted their individual and collective health. Objective: To identify the self-care strategies implemented by nursing professionals during the COVID-19 pandemic and to analyze them in the light of Giddens’ theoretical proposal. Materials and methods: An integrative literature review under a descriptive scope was conducted between June and September 2023 in the following databases: Pubmed, Scielo, Science Direct, Lilacs, and Google Scholar; also, PRISMA recommendations were taken into account for the organization of the search. Results: 40 articles, published between 2020 and 2023 and representing for different methodological designs, were included in the review. The self-care implemented by nurses during the pandemic was assessed in the light of Giddens’ theoretical proposal under two types of actions: intentional, which account for physical, mental, and emotional health care activities, and deliberate, which focus on care at a social level. Conclusions: The identified self-care actions were oriented toward the promotion of individual and collective health of workers, the reduction of employee turnover, the recognition of stress, and the strengthening of resilience, which can also be implemented nowadays.
Introdução: durante a emergência da covid-19, os enfermeiros estiveram presentes no atendimento aos pacientes e seus familiares, sendo constantemente expostos ao vírus devido a jornadas de trabalho extenuantes e outros fatores que levaram à fragilização de sua saúde. Para neutralizar esses fatores, foram identificadas na literatura publicada entre 2020 e 2023 diferentes ações de autocuidado realizadas por profissionais de enfermagem para promover sua saúde e prevenir o aparecimento de doenças causadas por estressores derivados da pandemia. Objetivo: identificar as estratégias de autocuidado implementadas pelos profissionais de enfermagem durante a pandemia da covid-19 e analisá-las à luz da proposta teórica de Giddens. Materiais e método: foi realizada uma revisão integrativa da literatura com escopo descritivo, entre junho e setembro de 2023, nas seguintes bases de dados: PubMed, SciELO, Science Direct, Lilacs e Google Scholar; além disso, foram levadas em conta as recomendações PRISMA para a organização da busca. Resultados: 40 artigos foram incluídos na revisão, publicados entre 2020 e 2023, com diferentes desenhos metodológicos. O autocuidado realizado pelos enfermeiros durante a pandemia foi organizado à luz da proposta teórica de Giddens, por meio de dois tipos de ações: intencionais, que contêm atividades para o cuidado da saúde física, mental e emocional; e deliberadas, voltadas para o cuidado em nível social. Conclusões: as ações de autocuidado identificadas foram orientadas para a promoção da saúde individual e coletiva dos trabalhadores, para a redução da deserção do trabalho, para o reconhecimento do estresse e para o fortalecimento da resiliência, que também podem ser implementadas atualmente.
Introducción: durante la emergencia por covid-19 el personal de enfermería estuvo presente en el cuidado de los pacientes y sus familias, exponiéndose constantemente al virus debido a los extenuantes horarios de trabajo y a otros factores que llevaron al debilitamiento de su salud. Para contrarrestar dichos factores, se han identificado, en la literatura publicada entre el periodo 2020-2023, diferentes acciones de autocuidado realizadas por los profesionales de enfermería con el fin de promover su salud y prevenir la aparición de enfermedades causadas por estresores propios derivados de la pandemia. Objetivo: identificar las estrategias de autocuidado implementadas por los profesionales de enfermería durante la pandemia covid-19 y analizar las mismas a la luz de la propuesta teórica de Giddens. Materiales y Método: se realizó una revisión integrativa de la literatura con alcance descriptivo entre junio y septiembre de 2023 en las siguientes bases de datos: Pubmed, Scielo, Science Direct, Lilacs y Google Académico; asimismo, se tuvieron en cuenta las recomendaciones PRISMA para la organización de la búsqueda. Resultados: se incluyeron 40 artículos en la revisión, publicados entre 2020 y 2023, con diferentes diseños metodológicos. El autocuidado realizado por los enfermeros durante la pandemia se organizó a la luz de la propuesta teórica de Giddens, mediante dos tipos de acciones: intencionadas, las cuales contienen actividades para el cuidado de la salud física, mental y emocional; y deliberadas, enfocadas al cuidado a nivel social. Conclusión: las acciones de autocuidado identificadas se orientaron a la promoción de la salud individual y colectiva de los trabajadores, la disminución de la deserción laboral, el reconocimiento del estrés y el fortalecimiento de la resiliencia, las cuales también pueden implementarse en la actualidad.
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