Expression analysis of <i>OsSERK</i>, <i>OsLEC1</i> and <i>OsWOX4</i> genes in rice (<i>Oryza</i> sativa L.) callus during somatic embryo development

Somatic embryogenesis is an asexual reproduction process that occurs in many plant species, including rice. This process contains several totipotency markers such as Somatic Embryogenesis Receptor-like Kinase (SERK), Leafy Cotyledon1 (LEC1) and WUSCHEL-Related Homeobox4 (WOX4) and also a helpful model for embryo development and clones and transformations. Here, we report the gene expression during somatic embryo development correlates with regeneration frequency in 14 Javanica rice (pigmented and non-pigmented) using modifified N6 media supplemented with Kinetin (2.0 mg/L) and NAA (1.0 mg/L). Although there have been advances in understanding the genetic basis of somatic embryogenesis in other varieties, rice is still unexplored, especially during somatic embryo development. Moreover, for the formation of callus induction from immature embryos, 2,4-D (2.0 mg/L, 3.0 mg/L) was used. This study analysed the gene expression of OsSERK, OsWOX4 and OsLEC1 genes through RT-PCR analysis. Higher expression of the OsLEC1 gene indicates that their function may correlate in the in vitro with the high response of rice after transfer to regeneration media. This study found that rice varieties of pigmented rice (MS Pendek and Gogoniti II) and non-pigmented rice (Pandan Ungu) showed high regeneration frequency, showing higher OsLEC1 expression than other varieties because OsLEC1 promotes the maturation of somatic embryos in plant regeneration on day 14. However, the contrast with Genjah nganjuk may be effective because of other regulatory genes. RT-PCR analysis showed OsSERK had less expression level than OsLEC1 and OsWOX4 in the varieties, which correlate with the percentage of plant regeneration, but not for Gogoniti II. In conclusion, the higher percentage of plant regeneration correlates with the higher expression level of OsLEC1 at day 14 of media regeneration of rice.     

Phytohormonal and metabolism analysis of <i>Brassica rapa</i> L. ssp. <i>pekinensis</i> with different resistance during <i>Plasmodiophora brassicae</i> infection

Clubroot of Chinese cabbage (Brassica rapa L. ssp. pekinensis), caused by the obligate parasite Plasmodiophorabrassicae, accounts for serious yield losses. The aim of our study was to explore the phytohormone levels and metabolomechanges in the roots of resistant and susceptible B. rapa genotypes at a late stage of infection, i.e., 28 days post-infection.Both genotypes showed decreased auxin levels after P. brassicae infection except for indole-3-acetic acid. Overall, thesusceptible genotype had higher auxin and cytokinin levels after infection, with the exception of trans-zeatin and 3-indolebutyric acid as compared to the resistant genotype. Jasmonic acid levels declined after infection regardless of thegenotype. Resistance against clubroot was evident with the increased levels of salicylic acid in the resistant genotype.The susceptible genotype had a higher number of differentially accumulated metabolites (DAMs) (262) than theresistant genotype (238) after infection. Interestingly, 132 DAMs were commonly detected in both genotypes wheninfected with the pathogen, belonging to metabolite classes such as phenolic acids, amino acids, and derivatives,glucosinolates, organic acids, flavonoids, nucleotides and derivatives, and fatty acids. The differential metaboliteanalysis revealed that metabolites related to amino acid biosynthesis, fatty acid biosynthesis and elongation,glutathione metabolism, and glucosinolate metabolism were highly accumulated in the resistant genotype, suggestingtheir essential roles in resistance against P. brassicae infection.     

Response of <i>MaHMA2</i> gene expression and stress tolerance to zinc stress in mulberry (<i>Morus alba</i> L.)

HMA2 (heavy metal ATPase 2) plays a crucial role in extracellular and intracellular Zn²⁺ transport across biomembranes, maintaining ion homeostasis, and playing an important role in the normal physiological metabolism, growth, and development of plants. In our study, a novel HMA2 gene, named MaHMA2, was isolated and cloned from white mulberry (Morus alba L.). The gene sequence obtained was 1,342 bp long, with an open reading frame of 1,194 bp, encoding a protein of 397 amino acids, with a predicted molecular mass of 42.852 kD and an isoelectric point of 7.53. This protein belonged to the PIB-type ATPase transport protein family. We analyzed the expression of the MaHMA2 gene by quantitative real-time PCR. The results showed that the level of MaHMA2 gene expression decreased to a Zn concentration of 800 mg/kg. Malondialdehyde and proline levels increased and responded to increasing Zn when the MaHMA2 gene was silenced, whereas the activities of peroxidase and superoxide dismutase tended to increase in response to increasing Zn²⁺ ion stress concentrations but were lower in the gene-silenced plants. These findings suggested that the MaHMA2 gene played an active role in the tolerance response of mulberry to Zn stress.     

Structure of the stigma and style in sunflower (<i>Helianthus annuus</i> L.)

This is the first report of the ultrastructure of the stigma and style during and after anthesis in Helianthus annuus L. using light and transmission electron microscopy. The stigma is bifid with unicellular papillae. There is no secretion of lipids, carbohydrates or proteins at anthesis. The style is semisolid in the upper portion, closer to the stigma, and becomes solid below. Ultrastructural changes on cells of the stigma and the style are described. The transmitting tissue of the ovule is first evident 40 minutes after pollination and persists during the first stages of embryogenesis. Only one pollen tube per micropyle was observed growing through this tissue.     

Application of two forms of silicon and their impact on the postharvest and the content of bioactive compounds in cucumber (<i>Cucumis sativus</i> L.) fruits

The metabolic activity of the fruits continues even after harvest, which results in the loss of bioactive compounds, a decrease in the quality of the fruits, softening and browning, among other negative effects. The use of certain elements such as silicon can improve postharvest quality, since it is involved in the metabolic, physiological and structural activity of plants, moreover can increase the quality of the fruits. In addition, nanotechnology has had a positive impact on crop yield, nutritional value, fruit quality and can improve antioxidant activity. For these reasons, the use of beneficial elements such as silicon in the form of nanoparticles can be a viable option to improve the characteristics of the fruits. In the present study was evaluated the application of potassium silicate (125, 250 and 500 mg L⁻¹) and SiO₂ nanoparticles (125, 250 and 500 mg L⁻¹) during the development of the crop. The results showed that the application of silicon (potassium silicate and silicon nanoparticles) increased the content of total soluble solids (up to 15.6% higher than control), titratable acidity (up to 38.8% higher than control), vitamin C (up to 78.2% higher than control), phenols (up to 22% higher than control), flavonoids (up to 64.6% higher than control), and antioxidant activity in lipophilic compounds (up to 56.2% higher than control). This study suggests that the use of silicon can be a good option to increase the content of bioactive compounds in cucumber fruits when they are applied during the development of the crop.     

Antiproliferative effect of extracts of <i>Sida rhombifolia</i> L. (Malvaceae) on the <i>Allium cepa</i> cell cycle

Field collected roots of four populations of Sida rhombifolia were used for preparing aqueous decoctions at two concentrations: 4g/L; and 16g/L. Afterwards, we used three groups of six onion (Allium cepa) bulbs for testing each population. Slides were made with all bulbs through the smashing technique. Cells in all phases of the cell cycle of A. cepa were analyzed. The mitotic index (% of cells in mitosis) was calculated, and the statistical analysis through the χ2 test was carried out at 5% probability. The results showed that the aqueous extracts of S. rhombifolia have antiproliferative activity at high concentrations. Practically no chromosomal aberrations were induced by treatments.     

Screening and evaluation of chilli (<i>Capsicum annuum</i> L.) genotypes for waterlogging tolerance at seedling stage

Waterlogging is an illustrious abiotic stress and the constrictions it enforces on plant roots have negative effectson growth and development. This study was undertaken to investigate waterlogging stress tolerant potential in chilli(Capsicum annum L.) genotypes through evaluating morphological, physiological, biochemical and anatomicalparameters. Thirty-five days old seedlings of 10 chilli genotypes were exposed to waterlogging stress maintainingwater height 3–5 cm over the soil surface artificially for three days. This duration (36–38 DAE) was termed aswaterlogging period, and subsequent withdrawal of waterlogging condition (39–45 DAE) was regarded as a recoveryphase. Based on their survival performance, two tolerant genotypes viz., SRC-517 and BARI morich-2 and twosusceptible genotypes viz., AHM-206 and RI-1(6) were selected for studying stress tolerance mechanism. Underwaterlogging, however, both genotypes (tolerant and susceptible) exhibited reduced root shoot length, dry weightratio, petiole weight and leaf area, and noticeable reduction regarding these parameters was observed in susceptiblegenotypes. Moreover, tolerant genotypes displayed a higher recovery than susceptible genotypes after removal ofwaterlogging stress. Lower reduction of leaf area and photosynthetic pigments as well as higher reduction of relativewater content (RWC) were noticed in susceptible genotypes. Higher accumulation of proline and total antioxidantcapacity (TAC) during waterlogging condition in tolerant genotypes suggested lower oxidative damage. Although bothgenotypes lost total soluble sugar (TSS) relative to control at waterlogging stress, better performance was recorded intolerant genotypes. During the period after the removal of extra water, a similar genotypic response in terms of TSSgain was seen. Undoubtedly, under flooding conditions, the development of aerenchyma cells in tolerant genotypes isa means of tolerance mechanism for long-term survival. Thus, the morpho-physiological and biochemical changeshelp to understand the tolerance mechanism in chilli under waterlogging stress.     

Induction of adaptive response <i>in utero </i>by ionizing radiation: A radiation quality dependent phenomenon

Investigation on possible induction of adaptive response (AR) by high-liner energy transfer (LET) particleradiation for protection against low-LET photon radiation-induced detrimental effects has not yet been performed inutero. This study verified if an AR could be induced by high-LET particle radiation from accelerated heavy ionsagainst low-LET X-ray radiation-induced detrimental effects on fetal mice. Total body irradiation of pregnantC57BL/6J mice were performed by delivering a priming dose ranging from 10 mGy to 320 mGy of particle radiationon gestation day 11 followed one day later by a challenge dose at 3500 mGy from X-ray radiation. The monoenergeticbeams of carbon, silicon and iron with the LET values of about 15, 55, and 200 KeV/μm, respectively, were examined.Significant suppression by the priming radiation of the detrimental effects (fetal death, malformation, or low bodyweight) was used as the endpoints for judgment of a successful AR induction on gestation day 18. Existence of ARwas not observed. On the other hand, the priming dose of high-LET particle radiation, in some cases, even increasedthe detrimental effects induced by the challenge dose from low-LET X-ray radiation. Although existence of ARinduced by high-LET radiation in cultured mammalian cells in vitro and in certain tissues of laboratory mice in vivowas demonstrated, the present study did not suggest that low dose of high-LET particle radiation could induce an ARin fetal mice inutero under the setup of our experimental system.     

Micropropagation of <i>Prosopis chilensis</i> (Mol.) Stuntz from young and mature plants

Prosopis chilensis (Mol.) Stuntz (Algarrobo de Chile) is an important native tree species that can be grown in arid and semiarid regions for wood and forage production and environmental protection. Developing a simple and reliable in vitro protocol for cloning it would enable to improve it genetically. Explants of P.chilensis were taken from 4 months-old plants grown in the greenhouse or from adult trees grown in a natural environment. Nodal segments 1 – 2 cm long containing an axillary bud were selected from elongating shoots. These cuttings were aseptically cultured on two agar-solid basal media, MS or BTMm, and treated with 0.05 mg L^-1 BA and 3 mg L^-1 of either IAA, IBA or NAA. Sucrose (3% w/v) was used as carbon source. The percentage of sprouted cuttings and whole plant regeneration as well as its shoot and root length were recorded. Number, length and dry weight of shoots and roots were also measured. Rooting was successful with cuttings taken from young or adult plants, but explants from young plants showed a better response. Culturing in BTMm resulted in significantly greater shoot and root biomass than culturing in MS. Moreover, this response was higher in young explants when IBA was used as growth regulator. This paper reports a simple and effective method to micropropagate P. chilensis from young and adult plants.     

<i>Trypanosoma rangeli:</i> growth in mammalian cells <i>in vitro</i> and action of a repositioned drug (17-AAG) and a natural extract (<i>Artemisia sp</i>. essential oil)

Trypanosoma rangeli and T. cruzi are both parasitic unicellular species that infect humans. Unlike T. cruzi,the causative agent of Chagas disease, T. rangeli is an infective and non-pathogenic parasite for humans, but pathogenicfor vectors from the Rhodnius genus. Because both species can coexist in different hosts and overlap their infectivecycles but very little is known about the infection of T. rangeli in mammalian cells, we decided to characterize both thedevelopment of this parasite in cell culture and the effect of therapeutic agents with potential trypanocidal action onit. We found that T. rangeli exhibits a cycle of infection in Vero cells similar to that for T. cruzi and that the repurposeddrug, 17-AAG, and the natural extract Artemisia sp. essential oil produce a toxic effect on epimastigotes showinga trypanocidal action from the fifth day of culture. Both treatments also affected the infection of trypomastigotesand reduced the capacity of replication of amastigotes of T. rangeli. Since T. cruzi / T. rangeli coinfection cases havebeen reported, the finding of drugs with potential activity against both species could be significant in the future.Furthermore, studies of susceptibility of both species to drugs could also help to know the different mechanisms ofpathogenicity in humans displayed by T. cruzi that are absent in T. rangeli     

Pollen viability of <i>Polygala paniculata</i> L. (Polygalaceae) using different staining methods

Polygala paniculata L. is a medicinal plant that grows in the Brazilian Atlantic coast, known as ‘barba-de-São-João’, ‘barba-de-bode’, ‘vassourinha branca’, and ‘mimosa’. In this study, pollen viability was estimated by three different staining methods: 2% acetic orcein, 2% acetic carmine, and Alexander’s stain. The young inflorescences of twenty accessions were collected and fixed in a solution of ethanol: acetic acid (3:1) for 24 hours, then stored in ethanol 70% under refrigeration. Six slides per plant, two for each stain, were prepared by squashing, and 300 pollen grains per slide were analyzed. Pollen viability was high (>70%) for most accessions of P. paniculata using the Alexander’s stain, which proved the most adequate method to estimate pollen viability.     

Brief Note : Stimulation of jasmonic acid production in <i>Zea Mays</i> L. infected by the maize rough dwarf virus - Río Cuarto. Reversion of symptoms by salicylic acid

In the present paper we study the possible biological relevance of endogenous jasmonic acid (JA) and exogenous salicylic acid (SA) in a plant-microbial system maize-virus. The virus disease "Mal de Río Cuarto" is caused by the maize rough dwarf virus - Río Cuarto. The characteristic symptoms are the appearance of galls or "enations" in leaves, shortening of the stem internodes, poor radical system and general stunting. Changes in JA and protein pattern in maize control and infected plants of a virus-tolerant cultivar were investigated. Healthy and infected-leaf discs were collected for JA measurement at different post-infection times (20, 40, 60 and 68 days). JA was also measured in roots on day 60 after infection. For SDS-PAGE protein analysis, leaf discs were also harvested on day 60 after infection. Infected leaves showed higher levels of JA than healthy leaves, and the rise in endogenous JA coincided with the enation formation. The soluble protein amount did not show differences between infected and healthy leaves; moreover, no difference in the expression of soluble protein was revealed by SDS-PAGE. Our results show that the octadecanoid pathway was stimulated in leaves and roots of the tolerant maize cultivar when infected by this virus. This finding, together with fewer plants with the disease symptoms, suggest that higher foliar and roots JA content may be related to disease tolerance. SA exogenous treatment caused the reversion of the dwarfism symptom.     

Gene expression of 49 kDa apyrase,cytoskeletal proteins,ATPase, ADPase and amino acid contents of <i>Pisum sativum</i> (L.) cells germinated in E<i>uryops arabicus</i> (Steud. ex Jaub. & Spach) water extract

The present research reports of quick and marked changes induced by plant extract of Euryops arabicus in thegene expression of 49-kDa apyrases, cytoskeletal proteins, ATPases, ADPase and amount of amino acid of pea (Pisumsativum L. var. Alaska). Pellets of cytoskeletals proteins (27000 xg) were probed with anti-apyrase antibody, biotinylatedanti-rat, actin and alpha and beta-tubulin for Western blotting. ATPase and ADPase activities were determined basedon the hydrolytic efficacy of adenine triphosphate and adenine diphosphate. By 72 hours, the abundance of apyrases,cytoskeletal proteins and amount of amino acid in pellets of 27000 xg of germinated pea seeds in E. arabicus extractswere sharply increased than those sown in distilled water. All the samples exhibited that the stems had more amountfrom apyrases, cytoskeletal proteins, amino acids and ATPase and ADPase activities than primary leaves and primaryroots that were germinated either on E. arabicus water extract or in distilled water. Based on the enzyme’s capability tohydrolyse nucleotide triphosphate and nucleotide diphosphate as well as the direct association between expression of49-kDa apyrase and cytoskeletal proteins, E. arabicus water extract had an important effect on plant germinations.     

Ultrastructure of the ovariole sheath in <i>Diatraea saccharalis</i> (Lepidoptera: Pyralidae)

The ultrastructure of the ovariole sheath along the Diatraea saccharalis ovariole was studied by scanning and transmission electron microscopy. Each ovariole is surrounded by an epithelial sheath, a tunica propria and scattered lumen cells. These three components of the ovariole sheath show different ultrastructural features along the ovariole, in the germarium or in the vitellarium; these differences are more evident in the epithelial sheath cells. The epithelial sheath is composed by two layers of cells, the external one running longitudinally and the internal one running circularly in the ovariole. These cells, in vitellarium, present cytoplasmic bundles of myofilaments that are arranged parallel to the long axis of the cells; these myofilaments are apparently related to the contraction movements of the follicles within the ovariole. The acellular tunica propria, composed of finely filamentous material, is attached to the adjacent follicle cells by adhesive dense plates. Between the epithelial sheath and the tunica propria there is a population of lumen cells, with morphological features of secretory activity     

Genome-wide identification,characterization, and expression analysis of aluminum-activated malate transporter genes (ALMTs) in <i>Gossypium hirsutum</i> L.

Aluminum-activated malate transporters (ALMT) are widely involved in plant growth and metabolic processes, including adaptation to acid soils, guard cell regulation, anion homeostasis, and seed development. Although ALMT genes have been identified in Arabidopsis, wheat, barley, and Lotus japonicus, little is known about its presence in Gossypium hirsutum L. In this study, ALMT gene recognition in diploid and tetraploid cotton were done using bioinformatics analysis that examined correlation between homology and evolution. Differentially regulated ALMT genetic profile in G. hirsutum was examined, using RNA sequencing and qRT-PCR, during six fiber developmental time-points, namely 5 d, 7 d, 10 d, 15 d, 20 d, and 25 d. We detected 36 ALMT genes in G. hirsutum, which were subsequently annotated and divided into seven sub-categories. Among these ALMT genes, 34 had uneven distribution across 14/26 chromosomes. Conserved domains and gene structure analysis indicated that ALMT genes were highly conserved and composed of exons and introns. The GhALMT gene expression profile at different DPA (days post anthesis) in different varieties of G. hirsutum is indicative of a crucial role of ALMT genes in fiber development in G. hirsutum. This study provides basis for advancements in the cloning and functional enhancements of ALMT genes in enhancing fiber development and augmenting high quality crop production.     

Micropropagation of <i>Ilex dumosa</i> (Aquifoliaceae) from nodal segments in a tissue culture system

Micropropagation of Ilex dumosa var. dumosa R. (“yerba señorita”) from nodal segments containing one axillary bud was investigated. Shoot regeneration from explants of six-year-old plants was readily achieved in ¹/₄ strength Murashige and Skoog medium (¹/₄ MS) plus 30 gr·L^-1 sucrose and supplemented with 4.4 µM BA. Further multiplication and elongation of the regenerated shoots were obtained by subculture in a fresh medium of similar composition with 1.5 gr·L^-1 sucrose. Rooting induction from shoots were achieved in two steps: 1) 7 days in ¹/₄ MS (30 gr·L^-1 sucrose, 0.25 % Phytagel^®) with 7.3 µM IBA and 2) 21 days in the same medium without IBA and 20 µM of cadaverine added. Regenerated plants were successfully transferred to soil. This micropropagation schedule can be implemented in breeding programs of Ilex dumosa.