Insecticidal characteristics and structural identification of the potential active compounds from Streptomyces sp. KR0006: Strain improvement through mutagenesis.

Pest control by biological means is an effective, eco-friendly, and promising method that typically involves compounds naturally derived from actinomycetes. Thus, the present study aimed to screen, characterize, and identify the structure of insecticidal compounds from Streptomyces sp. KR0006 and increase the activity through mutagenesis. In the examination of the insecticidal activity level of the isolates, Streptomyces sp. KR0006 metabolite showed significant activity against larvae and moths of Plutella xylostella. Taxonomic analyses of the 16S rRNA gene sequences revealed that the isolated KR0006 strain tended to be 99% consistent with Streptomyces cinereoruber strain NBRC 12756. Three active compounds isolated from the culture filtrate of KR0006 were purified by solvent partition, mid-pressure liquid chromatography (MPLC), Sephadex LH20 column chromatography, and high-performance liquid chromatography (HPLC). By performing 1H-NMR, 13C-NMR, and 2D-NMR experiments, and high-resolution electrospray ionization mass spectrometry analysis, the 316-HP2, 316-HP3, and 316-HP5 compounds were inferred as antimycin A3a (MW, 519.; C26H36N2O9), antimycin A8a (MW, 534; C27H38N2O9), and antimycin A1a (MW, 548; C28H40N2O9) respectively. Mutant U67 obtained from exposure to ultraviolet (UV) irradiation (254 nm, height 17 cm) for 70 seconds resulted in a 70% more larval mortality than that of the initial wild culture. The second mutation of the culture broth enhanced insecticidal activity by 80 and 100% compared with the first mutation and initial medium, respectively. Our study found that Streptomyces sp. KR0006 strain produces insecticidal active compounds and could be used for practical pest management.

Herbicidal characteristics and structural identification of a potential active compound produced by Streptomyces sp. KRA18-249.

The KRA18-249 strain, isolated from a natural recreational forest near Jeongseon, Gangwon-do, when applied to plants induced signs of wilting within 24 h, leading to plant death. The isolated actinomycete was identified as Streptomyces gardneri based on 16S rRNA gene homogeneity analysis. The culture filtrate was solvent fractionated to obtain the active substance, and the active compound 249-Y1 was isolated from the purified fractions via a herbicide activity test using Digitaria ciliaris. NMR and ESI-MS analyses revealed that the molecular formula of 249-Y1 is C20H16O6 [MW = 352.0947] and is an anthraquinone (rubiginone D2) produce by polyketide synthetase system. The active compound 249-Y1 showed strong (100%) herbicidal activity against several weeds at 500 µg mL-1 concentration. Twisting symptoms began to appear within 24 h of treatment and intensified over time. The KRA18-249 strain produced the herbicidal compound under specific culture conditions, that is, at 200 rpm, 35 °C, for eight days at an initial pH of 10. We also found that 249-Y1 inhibited chlorophyll, but was not a radical generator. Overall, the secondary metabolite 249-Y1, produced by KRA18-249, can be used as a new biological agent for weed control.

Herbicidal Characteristics and Structural Identification of the Potential Active Compounds from Streptomyces sp. KRA17-580.

Weeds are notorious plant species exhibiting a harmful impact on crops. Biological weed control is an efficient and environmentally friendly technique, usually constitutes naturally derived compounds, including bioherbicidal metabolites produced by Streptomyces sp. The isolation and structural identification of phytotoxic compounds from Streptomyces have recently been proposed as an effective way to the discovery of novel bioherbicides. In the screening of bioherbicidal agents, isolated Streptomyces strain KRA17-580 demonstrated significant phytotoxic activity against Digitaria ciliaris. Phylogenetic analysis of the 16S rRNA sequence indicated that isolated KRA17-580 is similar to Streptomyces olivochromogenes. The bacterial culture conditions were optimized for temperature, agitation, and initial pH. Streptomyces strain KRA17-580 showed intense phytotoxic activity and high cell mass at an initial pH of 5.5-7.0, more than 150 rpm, and 25-30 °C. The herbicidal compounds isolated from the culture filtrate of strain KRA17-580 were purified by solvent partition, C18, Sephadex LH20 column chromatography, and high-performance liquid chromatography. By 1D-NMR, 2D-NMR, and electrospray ionization mass spectrometry analysis, the 580-H1 and 580-H2 compounds were identified as a cinnoline-4-carboxamide (MW, 173.0490; C9H7N3O2) and cinnoline-4-carboxylic acid (MW, 174.0503; C9H6N2O2), respectively. Only these two herbicidal compounds showed strong phytotoxic activity against D. ciliaris in foliar applications. However, compound 580-H2 was more phytotoxic than 580-H1 and the toxicity was dose-dependent. The herbicidal metabolite KRA17-580 produced by Streptomyces sp. is a new bioherbicidal candidate that may provide a new lead molecule for more efficient phytotoxic compounds.

Impact of Organic Manure on Growth, Nutrient Content and Yield of Chilli Pepper under Various Temperature Environments.

Expected climatic changes likely elicit serious challenges for crop production. Therefore, it is indispensable to investigate the response of crop growth parameters and yield under temperature variability environments. The current experiment on chilli pepper growth was conducted in a field, rain-shelter plastic house, and plastic greenhouse, with accumulated temperatures of 2832 °C, 2967 °C, and 3105 °C in 2017; and 2944 °C, 3091 °C, and 3168 °C in 2018 growing seasons. Based on soil analysis, 132.7 kg ha-1 (1× of livestock manure compost as an optimum and 265.4 kg ha-1 (2×) as a double amount of organic matter were applied to each simulated temperature condition. The results showed that organic manure application favorably affects the growth attributes and nutrient uptake of chilli pepper with the highest values found in the plastic greenhouse, followed by the rain-shelter house, over the open field cultivation condition. The highest growth of chilli pepper was at the 2× rate of organic manure application, whereas the highest yield was found at the 1× rate of organic manure application. The application of organic manure at the 1× rate in the greenhouse increased root, shoot, and fruit dry weights of chilli pepper by 21.4%, 52.4%, and 79.7%, respectively, compared to the control values. These results indicate that the rational use of organic amendments might be the best solution for chilli pepper production under variable climate conditions.

Piperolein B and piperchabamide D isolated from black pepper (Piper nigrum L.) as larvicidal compounds against the diamondback moth (Plutella xylostella).

BACKGROUND: There is growing demand for the development of alternative pest control agents that are effective as well as non-toxic to human health and the environment. Plant protection products derived from plant extracts are an eco-friendly alternative to synthetic pesticides. The aim of this study was to identify larvicidal compounds isolated from a natural source against Plutella xylostella L. In a larvicidal activity assay, several solvent fractions from the methanol extract of Piper nigrum L. fruit showed larvicidal effects against P. xylostella. RESULTS: Screening results indicated that chloroform extract was the most effective against P. xylostella larvae. Two compounds with insecticidal activity in the chloroform fraction were identified as piperolein B and piperchabamide D by spectroscopic analyses, including mass spectrometry and NMR, and by comparison to published data. At applications of 0.1 mg mL-1 concentration, piperolein B and piperchabamide D, respectively, induced 96.7 ± 5.8% and 79.2 ± 16.6% mortality rates of P. xylostella larvae 4 days post-application. CONCLUSION: Our results demonstrate that piperolein B and piperchabamide D isolated from P. nigrum are the major constituents of the extract demonstrating insecticidal properties for the control of P. xylostella larvae. These plant-derived compounds should become useful alternatives to synthetic chemicals after studying their insecticidal mechanisms. © 2017 Society of Chemical Industry.

A comparative study of proteomic differences between pencil and storage roots of sweetpotato (Ipomoea batatas (L.) Lam.).

Fibrous roots of sweetpotato (Ipomoea batatas (L.) Lam.) usually develop into both pencil and storage roots. To understand protein function in root development, a proteomic analysis was conducted on the pencil and storage roots of the light orange-fleshed sweetpotato cultivar, Yulmi. Two-dimensional gel electrophoresis showed that expression of 30 protein spots differed between pencil and storage roots: 15 proteins were up-regulated or expressed in pencil roots and 15 in storage roots. Differentially expressed proteins spots were investigated using matrix-assisted laser desorption/ionization time of flight mass spectrometry, and 10 proteins from pencil roots were identified as binding protein isoform A, catechol oxidase, peroxidases, ascorbate peroxidase, endochitinase, flavanone 3-hydroxylase and unknown proteins. Of the proteins up-regulated in, or restricted to, storage roots, 13 proteins were identified as protein disulfide isomerase, anionic peroxidase, putative ripening protein, sporamin B, sporamin A and sporamin A precursor. An analysis of enzyme activity revealed that catechol oxidase and peroxidase as the first and last enzymes of the lignin biosynthesis pathway, and ascorbate peroxidase had higher activities in pencil than in storage roots. The total concentration of phenolic compounds was also far higher in pencil than in storage roots, and lignin accumulated only in pencil roots. These results provide important insight into sweetpotato proteomics, and imply that lignin biosynthesis and stress-related proteins are up-regulated or uniquely expressed in pencil roots. The results indicate that the reduction of carbon flow toward phenylpropanoid biosynthesis and its delivery to carbohydrate metabolism is a major event in storage root formation.

Comparative proteomic study between tuberous roots of light orange- and purple-fleshed sweetpotato cultivars.

This study compares the differences in proteomes expressed in tuberous roots of a light orange-fleshed sweetpotato (Ipomoea batatas (L.) Lam. cultivar Yulmi) and a purple-fleshed sweetpotato cultivar (Shinjami). More than 370 protein spots were reproducibly detected by two-dimensional gel electrophoresis, in which 35 spots were up-regulated (Yulmi vs. Shinjami) or uniquely expressed (only Yulmi or Shinjami) in either of the two cultivars. Of these 35 protein spots, 23 were expressed in Yulmi and 12 were expressed in Shinjami. These protein spots were analyzed by matrix-assisted laser desorption/ionization-time of flight mass spectrometry and electrospray ionization tandem mass spectrometry. Fifteen proteins in Yulmi and eight proteins in Shinjami were identified from the up-regulated (Yulmi vs. Shinjami) or uniquely expressed (only Yulmi or Shinjami) proteins, respectively. In Yulmi, α-amylase and isomerase precursor-like protein were uniquely expressed or up-regulated and activities of α-amylase, monodehydroascorbate reductase, and dehydroascorbate reductase were higher than in Shinjami. In Shinjami, peroxidase precursor and aldo-keto reductase were uniquely expressed or up-regulated and peroxidase and aldo-keto reductase activities were higher than in Yulmi. PSG-RGH7 uniquely expressed only in Shinjami and the cultivar was evaluated more resistant than Yulmi against the root-knot nematode, Meloidogyne incognita (Kofold and White, 1919) Chitwood 1949 on the basis of shoot and root growth. Egg mass formation was 14.9-fold less in Shinjami than in Yulmi. These results provide important clues that can provide a foundation for sweetpotato proteomics and lead to the characterization of the physiological function of differentially expressed proteins.

Gene flow from herbicide-tolerant GM rice and the heterosis of GM rice-weed F2 progeny.

Gene flow from genetically modified (GM) crops to non-GM cultivars or weedy relatives may lead to the development of more aggressive weeds. We quantified the amount of gene flow from herbicide-tolerant GM rice (Protox GM, derived from the cultivar Dongjin) to three cultivars (Dongjin, Aranghyangchal and Hwaseong) and a weedy rice line. Gene flow frequency generally decreased with increasing distance from the pollen donor. At the shortest distance (0.5 m), we observed a maximum frequency (0.039%) of gene flow. We found that the cultivar Dongjin received the greatest amount of gene flow, with the second being weedy rice. Heterosis of F2 inbred progeny was also examined between Protox GM and weedy rice. We compared growth and reproduction between F2 progeny (homozygous or hemizygous for the Protox gene) and parental rice lines (GM and weedy rice). Here, transgene-homozygous F2 progeny was significantly taller and produced more seeds than the transgene-hemizygous F2 progeny and parental lines. Although the gene flow frequency was generally low, our results suggest that F2 progeny between GM and weedy relatives may exhibit heterosis.

Enhancing sorgoleone levels in grain sorghum root exudates.

Sorgoleone, found in the root exudates of sorghum [(Sorghum bicolor (L.) Moench], has been a subject of continued research. Sorgoleone production in grain sorghum roots was investigated under different growth conditions. Methanol was the most effective solvent for extracting sorgoleone from grain sorghum roots. Sorgoleone production is high in young developing plants. The maximum concentration (microg mg(-1) root dry weight) was produced in 5-d-old seedlings; beyond this age, production declined. However, considering both root weight and sorgoleone content per seedling, 10-d-old seedlings had the highest total amounts (microg). Compared with the control, sorgoleone content increased 6.1, 8.6, and 14.2 times when sorghum seeds were treated with auxins, Hoagland solution, and a combination of auxins and Hoagland solution, respectively. Among the innate immunity response elicitors, cellulose (an elicitor of plant origin) stimulated higher sorgoleone production than the others, and it produced 6.2 times more sorgoleone than the control. Combined treatment of sorghum seeds with half strength Hoagland solution and 5 microg ml(-1) of IBA significantly increased both root growth and sorgoleone content in sorghum seedlings.

Investigating the mechanisms of glyphosate resistance in Lolium multiflorum.

Evolved resistance to the herbicide glyphosate has been reported in eleven weed species, including Lolium multiflorum. Two glyphosate-resistant L. multiflorum populations were collected, one from Chile (SF) and one from Oregon, USA (OR), and the mechanisms conferring glyphosate resistance were studied. Based on a Petri dish dose-response bioassay, the OR and the SF populations were two and fivefold more resistant to glyphosate when compared to the susceptible (S) population, respectively; however, based on a whole-plant dose-response bioassay, both OR and SF populations were fivefold more resistant to glyphosate than the S population, implying that different resistance mechanisms might be involved. The S population accumulated two and three times more shikimic acid in leaf tissue 96 h after glyphosate application than the resistant OR and SF populations, respectively. There were no differences between the S and the glyphosate-resistant OR and SF populations in 14C-glyphosate leaf uptake; however, the patterns of 14C-glyphosate translocation were significantly different. In the OR population, a greater percentage of 14C-glyphosate absorbed by the plant moved distal to the treated section and accumulated in the tip of the treated leaf. In contrast, in the S and in the SF populations, a greater percentage of 14C-glyphosate moved to non-treated leaves and the stem. cDNA sequence analysis of the EPSP synthase gene indicated that the glyphosate-resistant SF population has a proline 106 to serine amino acid substitution. Here, we report that glyphosate resistance in L. multiflorum is conferred by two different mechanisms, limited translocation (nontarget site-based) and mutation of the EPSP synthase gene (target site-based).

psbA mutation (Asn266 to Thr) in Senecio vulgaris L. confers resistance to several PS II-inhibiting herbicides.

DNA sequence analysis of the psbA gene encoding the D1 protein of photosystem II (PS II), the target site of PS II-inhibiting herbicides, identified a point mutation (Asn266 to Thr) in a bromoxynil-resistant Senecio vulgaris L. population collected from peppermint fields in Oregon. Although this mutation has been previously reported in Synechocystis, this is the first report of this particular point mutation in a higher plant exhibiting resistance to PS II-inhibiting herbicides. The resistant population displayed high-level resistance to bromoxynil and terbacil (R/S ratio 10.1 and 9.3, respectively) and low-level resistance to metribuzin and hexazinone (R/S ratio 4.2 and 2.6, respectively) when compared with the susceptible population. However, the population was not resistant to the triazine herbicides atrazine and simazine or to the urea herbicide diuron. A chlorophyll fluorescence assay confirmed the resistance levels and patterns of cross-resistance of the whole-plant studies. The resistant S. vulgaris plants produced fewer seeds. Differences in cross-resistance patterns to PS II-inhibiting herbicides and the difference in fitness cost could be exploited in a weed management program.