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Response of Microcystis aeruginosa BCCUSP 232 to barley ('Hordeum vulgare' L.) straw degradation extract and fractions

Gustavo Franciscatti Mecina, Anne Lígia Dokkedal, Luiz Leonardo Saldanha, Mathias Ahii Chia, Micheline Kézia Cordeiro-Araújo, Maria do Carmo Bittencourt-Oliveira, Regildo Márcio Gonçalves da Silva

The eutrophication of aquatic ecosystems is a serious environmental problemthat leads to increased frequency of cyanobacterial blooms and concentrations of cyanotoxins. These changes in aquatic chemistry can negatively affect animal and human health. Environment-friendly methods are needed to control bloom forming cyanobacteria. We investigated the effect of Hordeum vulgare L. (barley) straw degradation extract and its fractions on the growth, oxidative stress, antioxidant enzyme activities, and microcystins content of Microcystis aeruginosa (Kützing) Kützing BCCUSP232. Exposure to the extract significantly (p b 0.05) inhibited the growth of M. aeruginosa throughout the study, whereas only the highest concentration of fractions 1 and 2 significantly (p b 0.05) reduced the growth of the cyanobacterium on day 10 of the experiment. The production of reactive oxygen species (ROS), lipid peroxidation and antioxidant enzyme activities were significantly (p b 0.05) altered by the extract and fractions 1 and 2. Phytochemical profiling of the extract and its fractions revealed that the barley strawdegradation process yielded predominantly phenolic acids. These results demonstrate that barley straw extract and its fractions can efficiently interfere with the growth and development of M. aeruginosa under laboratory conditions.


Plant growth regulators on sweet sorghum: physiological and nutritional value analysis

Willian Rodrigues Macedo, Diego Kitahara Araújo, Valdinei Moreira Santos, Paulo Roberto de Camargo e Castro, Gisele Machado Fernandes

Sweet sorghum has gained attention in tropical and subtropical regions because of its potential as a bioenergy crop. The present research was carried out to evaluate the physiological, biochemical and nutritional characteristics of sweet sorghum submitted to six plant growth regulators (thiamethoxam, biostimulant mixture, gibberellic acid, chlormequat chloride, ethephon, and trinexapac-ethyl). The compounds were applied via foliar spraying in order to increase the productive potential of plants and reduce the carbohydrates sink strength by inflorescences. The experiment was conducted in pots and the following variables were evaluated: plant height, inflorescence dry matter, soluble solids content, shoot dry matter, crude protein, ashes, neutral detergent fiber and in vitro digestibility. It were observed the action of trinexapac-ethyl and ethephon to reduce the inflorescence dry matter, chlormequat chloride to increase the shoot dry matter, and trinexapac-ethyl to improve ashes content and reduce the neutral detergent fiber contents. These results indicate that chlormequat chloride and trinexapac-ethyl are effective in restricting the plant growth and increasing sorghum nutritional quality.


Loss of type-IV glandular trichomes is a heterochronic trait in tomatoand can be reverted by promoting juvenility

Eloisa Vendemiatti, Agustin Zsögön, Geraldo Felipe Ferreira e Silva,Frederico Almeida de Jesus, Lucas Cutri, Cassia Regina Fernandes Figueiredo,Francisco André Ossamu Tanaka, Fábio Tebaldi Silveira Nogueiraa,Lázaro Eustáquio Pereira Peres

Glandular trichomes are structures with widespread distribution and deep ecological significance. In the Solanum genus, type-IV glandular trichomes provide resistance to insect pests. The occurrence of thesestructures is, however, poorly described and controversial in cultivated tomato (Solanum lycopersicum).Optical and scanning electron microscopy were used to screen a series of well-known commercial tomatocultivars, revealing the presence of type-IV trichomes on embryonic (cotyledons) and juvenile leaves. Atomato line overexpressing the microRNA miR156, known to promote heterochronic development, andmutants affecting KNOX and CLAVATA3 genes possessed type-IV trichomes in adult leaves. A re-analysis ofthe Woolly (Wo) mutant, previously described as enhancing glandular trichome density, showed that thiseffect only occurs at the juvenile phase of vegetative development. Our results suggest the existence ofat least two levels of regulation of multicellular trichome formation in tomato: one enhancing differenttypes of trichomes, such as that controlled by the WOOLLY gene, and another dependent on develop-mental stage, which is fundamental for type-IV trichome formation. Their combined manipulation couldrepresent an avenue for biotechnological engineering of trichome development in plants.


Genetic and physiological characterization of three natural allelic variations affecting the organogenic capacity in tomato ('Solanum lycopersicum' cv. Micro-Tom)

Maísa de Siqueira Pinto, Chanaka Roshan Abeyratne, Vagner Augusto Benedito, Lázaro Eustáquio Pereira Peres

The study of allelic variations affecting organogenic capacity is not only relevant for manipulating plant traits but also to understand the fundamental mechanisms involved in plant development. Here, we report the characterization of three tomato (Solanum lycopersicum) loci (RG3C, RG7H and RG8F) whose alleles from its wild relative Solanum pennellii enhance in vitro shoot and root regeneration. S. pennellii alleles were introgressed into tomato cv. Micro-Tom (MT), creating near-isogenic lines. We evaluated the time taken for shoot induction and acquisition of competence by quantifying organogenesis after transferring explants, respectively, from the shoot-inducing medium (SIM) to the basal medium (BM) and from root-inducing medium (RIM) to the SIM. Concomitantly, we monitored the expression of key developmental genes. MT-Rg3C and MT-Rg7H started shoot induction, respectively, at 48 and 24 h earlier than MT and MT-Rg8F, while MT-Rg3C and MT-Rg8F acquired competence 24 h before MT. The impact of MT-Rg3C and MT-Rg8F in the acquisition of competence to assume different fates is consistent with their effect enhancing both shoot and root regeneration. MT-Rg7H seems to affect shoot induction specifically, which is in agreement with the enhanced expression of the shoot-related genes WUSCHEL and SHOOT MERISTEMLESS. Phenotypic characterization of greenhouse-grown plants showed that Rg3C has increased branching when compared to MT. Conversely, the normal branching observed in MT-Rg7H and MT-Rg8F indicates that adventitious in vitro shoot formation and ex vitro axillary bud formation/outgrowth are induced by different genetic pathways. These natural variations are thus useful for breeding highly regenerating varieties without undesirable effects on plant architecture.


Morpho-histological, histochemical, and molecular evidences related to cellular reprogramming during somatic embryogenesis of the model grass Brachypodium distachyon

Evelyn Jardim Oliveira, Andréa Dias Koehler, Diego Ismael Rocha, Lorena Melo Vieira, Marcos Vinícius Marques Pinheiro, Elyabe Monteiro de Matos, Ana Claudia Ferreira da Cruz, Thais Cristina Ribeiro da Silva, Francisco André Ossamu Tanaka, Fabio Tebaldi Silveira Nogueira, Wagner Campos Otoni
The wild grass species Brachypodium distachyon (L.) has been proposed as a new model for temperate grasses. Among the biotechnological tools already developed for the species, an efficient induction protocol of somatic embryogenesis (SE) using immature zygotic embryos has provided the basis for genetic transformation studies. However, a systematic work to better understanding the basic cellular and molecular mechanisms that underlie the SE process of this grass species is still missing. Here, we present new insights at the morpho-histological, histochemical, and molecular aspects of B. distachyon SE pathway. Somatic embryos arose from embryogenic callus formed by cells derived from the protodermal-dividing cells of the scutellum. These protodermal cells showed typical  meristematic features and high protein accumulation which were interpreted as the first observable steps towards the acquisition of a competent state. Starch content decreased along embryogenic callus differentiation supporting the idea that carbohydrate reserves are essential to morphogenetic processes. Interestingly, starch accumulation was also observed at late stages of SE process. Searches in databanks revealed three sequences available annotated as BdSERK, being two copies corresponding to SERK1 and one showing greater identity to SERK2. In silico analysis confirmed the presence of characteristic domains in a B. distachyon Somatic Embryogenesis Receptor Kinase genes candidates (BdSERKs), which suggests SERK functions are conserved in B. distachyon. In situ hybridization demonstrated the presence of transcripts of BdSERK1 in all development since globular until scutellar stages. The results reported in this study convey important information about the morphogenetic events in the embryogenic pathway which has been lacking in B. distachyon. This study also demonstrates that B. distachyon provides a useful model system for investigating the genetic regulation of SE in grass species. 

Genome editing as a tool to achieve the crop ideotype and de novo domestication of wild relatives: Case study in tomato

Agustin Zsögön; Tomas Cermak; Dan Voytas; Lázaro Eustáquio Pereira Peres
The ideotype is a theoretical model of an archetypal cultivated plant. Recent progress in genome editing is aiding the pursuit of this ideal in crop breeding. Breeding is relatively straightforward when the traits in question are monogenic in nature and show Mendelian inheritance. Conversely, traits with a diffuse, polygenic basis such as abiotic stress resistance are more difficult to harness. In recent years, many genes have been identified that are important for plant domestication and act by increasing yield, grain or fruit size or altering plant architecture. Here, we propose that (a) key monogenic traits whose physiology has been unveiled can be molecularly tailored to achieve the ideotype; and (b) wild relatives of crops harboring polygenic stress resistance genes or other traits of interest could be de novo domesticated by manipulating monogenic yield-related traits through state-of-the-art gene editing techniques. An overview of the genomic and physiological challenges in the world’s main staple crops is provided. We focus on tomato and its wild Solanum (section Lycopersicon) relatives as a suitable model for molecular design in the pursuit of the ideotype for elite cultivars and to test de novo domestication of wild relatives.

Are plant growth retardants a strategy to decrease lodging and increase yield of sunflower?

Marcia Eugenia Amaral Carvalho, Paulo Roberto de Camargo e Castro, Marcos Vinicius de Castro Ferraz Junior, Ana Carolina Cabrera Machado Mendes

One of the major disadvantages of sunflower cultivation is the increased plant height, making it prone to the lodging. The use of plant growth retardants can be an alternative strategy to reduce plant height; however, these compounds may affect productivity. The aim of this study was to evaluate the effects of plant growth retardants on sunflower development and yield. Four treatments were studied: 1- control; 2- gibberellic acid (GA) 10 mg L−1; 3- trinexapac-ethyl (TE) 5 mL L−1, and 4- maleic hidrazide (MH) 8 mL L−1. TE and MH decreased plant height (16.9 and 35.9%, respectively); however, only TE positively influenced capitulim diameter and dry mass (46.7 and 311%, when compared to control) at 60 days after planting (DAP). At 81 DAP, dry mass of capitulum did not differ among control and TE-treated plants. On the other hand, MH impaired diameter and dry mass of capitulum (92.9 and 74.7%, respectively). It can be concluded that the application of TE is a potential strategy to decrease lodging probability without affecting sunflower yield. Furthermore, although MH negatively affected sunflower development, its use on the crop cannot be excluded since other doses, frequencies and moment of application can be studied.


RNA interference as a gene silencing tool to control "Tuta absoluta" in tomato ("Solanum lycopersicum")

Roberto A. Camargo; Guilherme O. Barbosa; Isabella Presotto Possignolo; Lazaro Eustáquio Pereira Peres; Eric Lam; Joni E. Lima; Antonio Figueira; Henrique Marques-Souza
RNA interference (RNAi), a gene-silencing mechanism that involves providing doublestranded RNA molecules that match a specific target gene sequence, is now widely used in functional genetic studies. The potential application of RNAi-mediated control of agricultural insect pests has rapidly become evident. The production of transgenic plants expressing dsRNA molecules that target essential insect genes could provide a means of specific gene silencing in larvae that feed on these plants, resulting in larval phenotypes that range from loss of appetite to death. In this report, we show that the tomato leafminer (Tuta absoluta), a major threat to commercial tomato production, can be targeted by RNAi.Weselected two target genes (Vacuolar ATPase-A and Arginine kinase) based on the RNAi response reported for these genes in other pest species. In view of the lack of an artificial diet for T. absoluta, we used two approaches to deliver dsRNA into tomato leaflets. The first approach was based on the uptake of dsRNA by leaflets and the second was based on "in planta-induced transient gene silencing" (PITGS), a well-established method for silencing plant genes, used here for the first time to deliver in planta-transcribed dsRNA to target insect genes. Tuta absoluta larvae that fed on leaves containing dsRNA of the target genes showed and 60% reduction in target gene transcript accumulation, an increase in larval mortality and less leaf damage. We then generated transgenic `Micro-Tom' tomato plants that expressed hairpin sequences for both genes and observed a reduction in foliar damage by T. absoluta in these plants. Our results demonstrate the feasibility of RNAi as an alternative method for controlling this critical tomato pest.

Does anatoxin-a influence the physiology of 'Microcystis aeruginosa' and 'Acutodesmus acuminatus' under different light and nitrogen conditions?

Mathias Ahii Chia; Micheline Kézia Cordeiro-Araújo; Adriana Sturion Lorenzi; Maria do Carmo Bittencourt-Oliveira

Due to changing global climatic conditions, a lot of attention has been given to cyanobacteria and their bioactive secondary metabolites. These conditions are expected to increase the frequency of cyanobacterial blooms, and consequently, the concentrations of cyanotoxins in aquatic ecosystems. Unfortunately, there are very few studies that address the effects of cyanotoxins on the physiology of phytoplankton species under different environmental conditions. In the present study, we investigated the effect of the cyanotoxin anatoxin-a (ATX-A) on Microcystis aeruginosa (cyanobacteria) and Acutodesmus acuminatus (chlorophyta) under varying light and nitrogen conditions. Low light (LL) and nitrogen limitation (LN) resulted in significant cell density reduction of the two species, while the effect of ATX-A on M. aeruginosa was not significant. However, under normal (NN) and high nitrogen (HN) concentrations, exposure to ATX-A resulted in significantly (p < 0.05) lower cell density of A. acuminatus. Pigment content of M. aeruginosa significantly (p < 0.05) declined in the presence of ATX-A, regardless of the light condition. Under each light condition, exposure to ATX-A caused a reduction in total microcystin (MC) content of M. aeruginosa. The detected MC levels varied as a function of nitrogen and ATX-A concentrations. The production of reactive oxygen species (H2O2) and antioxidant enzyme activities of both species were significantly altered by ATX-A under different light and nitrogen conditions. Our results revealed that under different light and nitrogen conditions, the response of M. aeruginosa and A. acuminatus to ATX-A was variable, which demonstrated the need for different endpoints of environmental factors during ecotoxicological investigations.


Root growth restraint can be an acclimatory response to low pH and is associated with reduced cell mortality: a possible role of class III peroxidases and NADPH oxidases

J. P. Gracas; R. Ruiz-Romero; L. D. Figueiredo; L. Mattiello; Lázaro Eustáquio Pereira Peres; Victor Augusto Vitorello

Low pH (<5.0) can significantly decrease root growth but whether this is a direct effect of H+ or an active plant response is examined here. Tomato (Solanum lycopersicum cv Micro-Tom) roots were exposed directly or gradually to low pH through step-wise changes in pH over periods ranging from 4 to 24 h. Roots exposed gradually to pH 4.5 grew even less than those exposed directly, indicating a plant-coordinated response. Direct exposure to pH 4.0 suppressed root growth and caused high cell mortality, in contrast to roots exposed gradually, in which growth remained inhibited but cell viability was maintained. Total class III peroxidase activity increased significantly in all low pH treatments, but was not correlated with the observed differential responses. Use of the enzyme inhibitors salicylhydroxamic acid (SHAM) or diphenyleneiodonium chloride (DPI) suggest that peroxidase and, to a lesser extent, NADPH oxidase were required to prevent or reduce injury in all low pH treatments. However, a role for other enzymes, such as the alternative oxidase is also possible. The results with SHAM, but not DPI, were confirmed in tobacco BY-2 cells. Our results indicate that root growth inhibition from low pH can be part of an active plant response, and suggest that peroxidases may have a critical early role in reducing loss of cell viability and in the observed root growth constraint.


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