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Transformation of the oomycete pathogen, Phytophthora infestans.

A stable transformation procedure has been developed for Phytophthora infestans, an oomycete fungus that causes the late blight diseases of potato and tomato. This is the first description of reliable methods for transformation in an oomycete pathogen. Drug-resistant transformants were obtained by using vectors that contained bacterial genes for resistance to hygromycin B or G418 fused to promoters and terminators from the Hsp70 and Ham34 genes of the oomycete, Bremia lactucae. Using polyethylene glycol and CaCl2, vector DNA was introduced into protoplasts as a complex with cationic liposomes or with carrier DNA only. Transformants were obtained at similar frequencies with each combination of promoter and selectable marker and were confirmed by DNA and RNA hybridization and phosphotransferase assays. Transformation occurred through the integration of single or tandemly repeated copies of the plasmids into genomic DNA, conferring mitotically stable drug-resistant phenotypes. The sizes of the marker gene mRNAs in each transformant and the results of transcript mapping studies were consistent with the function of the B. lactucae regulatory sequences in P. infestans. A hygromycin-resistant transformant was tested and found to maintain pathogenicity, indicating that the gene transfer procedure will be useful for the molecular analysis of genes relevant to disease.

Base Sequence↗

Quantification of melatonin in phototrophic organisms.

Melatonin, the chief secretory product of the vertebrate pineal gland is also known to occur in numerous photoautotrophic organisms. The indoleamine is suspected to act as a transducer of photoperiodic information and/or to participate in antioxidative protection. In higher plants and other photoautotrophic organisms, contradictory results for melatonin content for samples from the same species show that further improvement of methods for reliable quantification is required. In the present study, melatonin was quantified in tomatoes, ginger and the marine green macroalga, Ulva lactuca, after extraction with three different extraction methods based on ether, acetone or perchloric acid. Melatonin was determined by enzyme-linked immunosorbent assay (ELISA) in high-performance liquid chromatography (HPLC)-purified extracts. The same HPLC system used for purification of extracts was used for parallel quantifications after derivatization of melatonin under alkaline conditions in the presence of hydrogen peroxide (HPLC-PD). Both quantification methods gave similar results with a high correlation [f(x) = 0.99x + 3.01; R(2) = 0.99]. In ginger, the melatonin concentration was below 5 pg/g (fresh weight, f.w.), whereas in tomatoes about 1200 pg/g (f.w.) were found, and in the green alga, U. lactuca, approximately 12 pg/g (f.w.). Taking into account the recovery rates for synthetic melatonin added prior to extraction, no substantial differences were observed in melatonin quantification between different extraction methods. The demonstrated methods based on HPLC purification and subsequent quantification by ELISA and HPLC-PD allow highly sensitive melatonin determinations in diverse photoautotrophic organisms with a low risk of overestimations by false-positive results.

Chromatography, High Pressure Liquid↗

Competitive interactions in mixed-species biofilms containing the marine bacterium Pseudoalteromonas tunicata.

Pseudoalteromonas tunicata is a biofilm-forming marine bacterium that is often found in association with the surface of eukaryotic organisms. It produces a range of extracellular inhibitory compounds, including an antibacterial protein (AlpP) thought to be beneficial for P. tunicata during competition for space and nutrients on surfaces. As part of our studies on the interactions between P. tunicata and the epiphytic bacterial community on the marine plant Ulva lactuca, we investigated the hypothesis that P. tunicata is a superior competitor compared with other bacteria isolated from the plant. A number of U. lactuca bacterial isolates were (i) identified by 16S rRNA gene sequencing, (ii) characterized for the production of or sensitivity to extracellular antibacterial proteins, and (iii) labeled with a fluorescent color tag (either the red fluorescent protein DsRed or green fluorescent protein). We then grew single- and mixed-species bacterial biofilms containing P. tunicata in glass flow cell reactors. In pure culture, all the marine isolates formed biofilms containing microcolony structures within 72 h. However, in mixed-species biofilms, P. tunicata removed the competing strain unless its competitor was relatively insensitive to AlpP (Pseudoalteromonas gracilis) or produced strong inhibitory activity against P. tunicata (Roseobacter gallaeciensis). Moreover, biofilm studies conducted with an AlpP- mutant of P. tunicata indicated that the mutant was less competitive when it was introduced into preestablished biofilms, suggesting that AlpP has a role during competitive biofilm formation. When single-species biofilms were allowed to form microcolonies before the introduction of a competitor, these microcolonies coexisted with P. tunicata for extended periods of time before they were removed. Two marine bacteria (R. gallaeciensis and P. tunicata) were superior competitors in this study. Our data suggest that this dominance can be attributed to the ability of these organisms to rapidly form microcolonies and their ability to produce extracellular antibacterial compounds.

Anti-Bacterial Agents↗

Structure elucidation and phytotoxicity of ecdysteroids from Chenopodium album.

The leaves of Chenopodium album were infused in H2O/MeOH. The extract treated with cold acetone gave heavy precipitation, which was removed by centrifugation. Solid material was fractionated into acidic and neutral fractions. The acidic material was subjected to different silica-gel column chromatographies, and then it was purified by reversed-phase HPLC to afford four known ecdysteroids and the new 3beta,14alpha-dihydroxy-5beta-pregn-7-ene-2,6,20-trione that were characterized by extensive spectroscopic investigation, especially by 1D- and 2D-NMR. Their effects on germination and growth of Lactuca sativa L. have been studied. The results are reported as percentage differences of germination, root elongation and shoot elongation, from the control at concentrations ranging from 10(-4) to 10(-7) M.

Chenopodium album↗

Quality of water types in Ukraine evaluated by WaterTox bioassays.

The quality of river, ground-, and tap water was analyzed using the basic set of WaterTox bioassays [Daphnia (Daphnia magna), Hydra (Hydra attenuata), and lettuce (Lactuca sativa)] as well as two additional bioassays, onion (Allium cepa) and microalga (Selenastrum gracile). Samples of these waters were also concentrated fivefold using a solid-phase procedure. The results of the Daphnia and Hydra bioassays showed that the winter and spring concentrated and nonconcentrated samples from the Dnieper and Desna rivers, the main water supply sources for Kiev, were nontoxic. In spring, after concentration, the two river samples brought about the same relative decrease in the lettuce root length (by 35%, p < 0.001), where the Desna River sample considerably reduced (by 79.1%, p < 0.001) the number of microalga cells. Samples of groundwater from countryside wells studied in autumn in several villages of the Kiev region were toxic mainly to Hydra (sublethal effects were found in 11%-78%) and lettuce (the root length decreased 15%-56%). Studies of tap water samples from two of the largest cities of Ukraine, Kiev and Kharkiv, were found to be nontoxic to both plants, lettuce and onion, but showed increased sublethal and lethal effects on both animals, Daphnia and Hydra, as well as a reduced number of microalgae. Different bioassays were sensitive to varying degrees to different water types. This reinforces the necessity of using sets of bioassays in toxicity evaluation. In general, all the tested water samples demonstrated some toxicity. These data suggest that drinking water quality in Ukraine needs improvement.

Animals↗

Overview of results from the WaterTox intercalibration and environmental testing phase II program: part 2, ecotoxicological evaluation of drinking water supplies.

Because of rapid population growth, industrial development, and intensified agricultural production increasing amounts of chemicals are being released into the environment, polluting receiving water bodies around the world. Given the potential health risk associated with the presence of toxicants in water sources used for drinking yet the scarcity of available data, there is a need to evaluate these waters and develop strategies to reduce and prevent their contamination. The present study examined the applicability of a battery of simple, inexpensive bioassays in environmental management and the relevance of the test results in establishing the toxicological quality of water sources and drinking water within the framework of the eight-country WaterTox Network, sponsored by the International Development Research Centre, Ottawa, Canada. Seventy-six samples were collected from surface and groundwater sources and seven samples from drinking water treatment plants. Each sample was tested with a core battery of bioassays (Daphnia magna, Hydra attenuata, and Lactuca sativa root inhibition tests) and a limited set of physical and chemical parameters. In addition, three labs included the Selenastrum capricornutum test. When no toxic effects were found with the battery, samples were concentrated 10x using a solid-phase extraction (SPE) procedure. Nonconcentrated natural water samples produced a toxic response in 24% of cases with all three core bioassays. When all bioassays are considered, the percentage of raw samples showing toxicity with at least one bioassay increased to 60%. Of seven treated drinkingwater samples, four showed toxicity with at least one bioassay, raising the possibility that treatment processes in these instances were unable to remove toxic contaminants. The Daphnia magna and Hydra attenuata tests indicated a high level of sensitivity overall. Although only three of the eight countries used S. capricornutum, it proved to be an efficient and reliable bioassay for toxicity assessment.

Animals↗

Interlaboratory study for the validation of an ecotoxicological procedure to monitor the quality of septic sludge received at a wastewater treatment plant.

Septic tank sludge is regularly hauled to the Montreal Urban Community (MUC) wastewater treatment plant. It is then discharged and mixed with the wastewater inflow before entering the primary chemical treatment process. An ecotoxicological procedure integrating chemical and toxicological analyses has been recently developed and applied to screen for the illicit discharge of toxic substances in septic sludge. The toxicity tests used were the Microtox, the bacterial-respiration, and the lettuce (Lactuca sativa) root elongation tests. In order to validate the applicability of the proposed procedure, a two-year interlaboratory study was carried out. In general, the results obtained by two independent laboratories (MUC and the Centre d'expertise en analyse environnementale du Quebec) were comparable and reproducible. Some differences were found using the Microtox test. Organic (e.g., phenol and formaldehyde) and inorganic (e.g., nickel and cyanide) spiked septic sludge were detected with good reliability and high efficiency. The relative efficiency to detect spiked substances was > 70% and confirms the results of previous studies. In addition, the respiration test was the most efficient toxicological tool to detect spiked substances, whereas the Microtox was the least efficient (< 15%). Efficiencies to detect spiked contaminants were also similar for both laboratories. These results support previous data presented earlier and contribute to the validation of the ecotoxicological procedure used by the MUC to screen toxicity in septic sludge.

Environmental Monitoring↗

Hysteresis and positive cooperativity of iceberg lettuce polyphenol oxidase.

A kinetic study of the diphenolase activity of latent polyphenol oxidase (PPO), purified from Iceberg lettuce (Lactuca sativa L), revealed a sigmoid relationship between the reaction rate and the substrate concentration with a high Hill coefficient (n(H) = 3.8). This positive cooperativity had not been previously described for any PPO. Furthermore, the enzyme showed a lag phase in the expression of this activity, suggesting a hysteretic nature of the enzyme. The kinetic behavior, the latency and the lag phase varied at different steps of the purification process. PPO showed hyperbolic or cooperative kinetics depending on the pH assay and the sodium dodecyl sulfate (SDS) concentration. Substrate-induced slow conformational change of the oligomeric enzyme is suggested. The conformational change would be toward a more active enzyme form with higher affinity for the substrate and favoured by acid pH and SDS.

Catechol Oxidase↗

Influence of growth conditions on the value of crisphead lettuce. 4. Quality changes during storage.

Crisphead lettuce (Lactuca sativa L. var capitata cvs. Marius and Saladin) were grown with a nitrogen supply from 50 to 200 kg N/hectare. Heads were stored for one or two weeks at 1 degree C in cold storage or ice bank cooling. Samples were taken for measurement of dry matter, sugars, vitamin C and nitrate. The content of dry matter, sugars (glucose, fructose) and vitamin C decreased with increasing level of nitrogen, and the content of nitrate increased. Except for nitrate the contents of the other quality attributes decreased at all nitrogen supply levels during storage. No differences were found between the storage systems, and beside fructose no significant differences were found between the two cultivars. The content of dry matter, vitamin C, and nitrate decreased from the outer to the inner head fraction, while the content of sugars increased. Trimming decreased the content of dry matter, vitamin C and nitrate and increased the content of sugars. To obtain heads from storage with a relatively high content of dry matter, sugars and vitamin C, and a relatively low content of nitrate the nitrogen supply must be as low as possible. Except for nitrate where no distinct results were found in this experiment it must also be recommended to store the heads as short time as possible. Possibly the cv. Saladin has some advantage quality attributes after storage compared with the cv. Marius.

Ascorbic Acid↗

Molecular and functional characterization of Slide, an Ac-like autonomous transposable element from tobacco.

A new transposable element of tobacco, Slide, was isolated from the tl mutant line, which shows somatic instability, after its transposition into a locus encoding nitrate reductase (NR). The Slide-124 element is 3733 bp long and its coding sequences show similarities with conserved domains of the transposases of Ac, Tam3 and hobo. Excision from the NR locus is detectable in somatic leaf tissues and Slide mobility is triggered by in vitro tissue culture. Slide excision events create footprints similar to those left by Ac and Tam3. Tobacco lines derived from the tl mutant line seem characterized by unmethylated copies of a few members of the highly repetitive Slide family. Slide mobility was monitored in transient expression assays. In wild-type tobacco protoplasts, the complete Slide element, as well as a defective copy, is able to excise. The complete Slide element, but not the defective version, is able to excise in protoplasts of the heterologous species lettuce (Lactuca sativa). These results show that Slide carries the functions required for its own mobility, and represents the first autonomous Ac-like element characterized in Solanaceae species.

Amino Acid Sequence↗

Nitric oxide stimulates seed germination and de-etiolation, and inhibits hypocotyl elongation, three light-inducible responses in plants.

Seed germination, greening of etiolated plants and inhibition of hypocotyl elongation are stimulated by light, which is sensed by various types of photoreceptor. Nitric oxide (NO) has proven to be a bioactive molecule, especially in mammalian cells and, most recently, in plants. Like some phytochrome-dependent processes, many NO-mediated ones are accomplished through increases in cGMP levels. Given these similarities, we proposed that NO could take part in light-mediated events in plants. Here we show that NO promotes seed germination and de-etiolation, and inhibits hypocotyl and internode elongation, processes mediated by light. Two NO donors, sodium nitroprusside (SNP) and S-nitroso-N-acetylpenicillamine induced germination of lettuce (Lactuca sativa L. cv. Grand Rapids) seeds in conditions in which this process is dependent on light (e.g. 26 degrees C). This was a dose-dependent response and was arrested by addition of an NO scavenger, carboxy-PTIO. In addition, nitrite and nitrate, two NO-decomposition products were ineffective in stimulating germination. Wheat seedlings sprayed with SNP and grown in darkness contained 30-40% more chlorophyll than control seedlings. Nitric-oxide-mediated partial greening was increased by light pulses, wounding and biotic stress. Arabidopsis thaliana (L.) Heynh. (ecotype Columbia) and lettuce seedlings grown in the dark had 20%-shorter hypocotyls in NO treatments than in control ones. On the other hand, internode lengths of potato plants growing under low light intensity and sprayed with 100 microM SNP were also 20% shorter than control ones. These results implicate NO as a stimulator molecule in plant photomorphogenesis, either dependent on or independent of plant photoreceptors.

Chlorophyll↗

Phytotoxicity of 2,4,6-trinitrotoluene (TNT) and octahydro-1,3,5,7-tetranitro-1,3,5,7-tetrazocine (HMX) in spiked artificial and natural forest soils.

Toxicity of 2,4,6-trinitrotoluene (TNT) and octahydro-1,3,5,7-tetranitro-1,3,5,7-tetrazocine (HMX) using two terrestrial plant species, lettuce (Lactuca sativa) and barley (Hordeum vugare), was assessed in artificial soil (silica) and forest soil. Lettuce emergence was significantly decreased after 5 days of exposure to TNT nominal spiked concentrations >/= 1,040 mg/kg dry soil in silica. Barley emergence was significantly reduced after 14 days of exposure at initial (t = 0) TNT concentrations >/= 55.9 +/- 4.5 mg/kg dry soil in silica and at >/= 291.9 +/- 42.8 mg/kg dry forest soil. Biomasses of shoot and roots of barley seeds were significantly reduced after 14 days of exposure at TNT initial exposure concentrations >/= 55.9 +/- 4.5 (LOEC) mg/kg dry soil in silica. Results were similar with the forest soil (LOEC = 91.4 +/- 7.9 mg TNT/kg dry soil) using the root growth parameter, but the shoot biomass was reduced only at concentrations >/= 291.9 +/- 42.8 mg TNT/kg dry soil. Plants were not affected by an HMX exposure up to 3,320 +/- 1,019 mg/kg dry soil using silica or 1,866 +/- 438 mg/kg dry soil using a forest soil. During the 14-day experiments, TNT was partially transformed in the spiked soil samples, as indicated by the presence of its amino metabolites (2-ADNT and 4-ADNT). Higher quantities of metabolites were detected in forest soils having higher initial TNT concentrations (</= 1,849.4 +/- 228.2 mg/kg) compared to silica (</= 239.3 +/- 88.0 mg TNT/kg). After 14 days, TNT concentrations in spiked silica and forest soil were reduced up to 80.5% at 55.9 +/- 4.5 mg/kg initial concentration and 94.4% at 91.4 +/- 7.9 mg/kg initial concentration, respectively. Data indicate that TNT is the probable phytotoxicant because it decreased plant emergence and growth in the presence and absence of the ADNT metabolites.

Azocines↗

Short-term bioassay responses to sludge products and leachate.

Recycling of sewage sludge is needed in a sustainable society. Quality aspects of sludge include hygiene (pathogens), nutrients (N and P), and toxicants (metals and organics). Metals are of particular concern because they are not degradable, but their hazards are related to their bioavailability and chemical speciation. In this article, the effect on sludge quality of two treatment methods, incineration and pelletization, has been determined for digested sludge from two treatment plants. The combined effect of nutrients and toxicants in sludge and sludge product was determined for spring wheat, Triticum aestivum, and the toxicity of the leachate water was determined for water fleas, Daphnia magna, and seeds of lettuce, Lactuca sativa. Toxicity Identification Evaluation was used to determine whether metals were possible toxicants. The results indicated that incineration decreased toxicity, whereas leachability of metals and the fertilizing effect was unaffected. Pelletization seemed to increase toxicity and leachability of metals and also decreased the fertilizing effect of the sludge. Thus, the results suggest that pelletization of digested sewage sludge increased the toxicity of the sludge and thus decreased the quality of the sludge, whereas incineration apparently reduced toxicity for the two sludges tested.

Animals↗

Biosynthetic potential of the culturable foliar fungi associated with field-grown lettuce.

Fungal endophytes and epiphytes associated with plant leaves can play important ecological roles through the production of specialized metabolites encoded by biosynthetic gene clusters (BGCs). However, their functional capacity, especially in crops like lettuce (Lactuca sativa L.), remains poorly understood. We sequenced the genomes of nine fungal isolates, representing Fusarium sp., Fulvia sp., Alternaria alternata, and Alternaria postmessia, from leaves of lettuce grown under field conditions in Arizona, USA. We used antibiotics and secondary metabolite analysis shell (antiSMASH) and the database for automated carbohydrate-active enzyme annotation (dbCAN3), to predict BGCs and carbohydrate-active enzymes (CAZymes) for each strain, and then compared them to conspecific strains from other environments and substrates. Foliar lettuce-associated fungi featured 39-95 BGCs per genome, with substantial overlap between isolates occurring in association with lettuce leaves vs. from other substrates. Species identity was a significant determinant of BGC count, while host type, isolation source, and lifestyle were not. Several BGCs, including those for alternariol and 1,3,6,8-Tetrahydroxynaphthalene (T4HN), showed 100% similarity to characterized minimum information about a biosynthetic gene cluster (MIBiG) clusters based on antiSMASH predictions. Although analysis by biosynthetic gene similarity clustering and prospecting engine (BiG-SCAPE) identified gene cluster families (GCFs) across the dataset, these reference-matching clusters were not always grouped, reflecting methodological differences in how the tools assess similarity. Comparative CAZyme analysis in a focal species (Fulvia sp.) revealed higher gene counts in a foliar lettuce-derived isolate than in tomato (Solanum lycopersicum)-associated strains, challenging assumptions about host chemical complexity. These results highlight the importance of phylogenetic context in shaping fungal functional potential and suggest that selection on microbial traits in edible leafy crops may be more subtle and species-specific than previously assumed. KEY POINTS: &#x2022; Lettuce-associated fungi feature diverse biosynthetic potential &#x2022; Phylogeny predicts fungal BGC content more strongly than ecological lifestyle &#x2022; Findings support genome-informed microbiome strategies for leafy crops.

Lactuca↗

Inhibition of endogenous trypsin- and chymotrypsin-like activities in transgenic lettuce expressing heterogeneous proteinase inhibitor SaPIN2a.

SaPIN2a, a proteinase inhibitor II from American black nightshade (Solanum americanum Mill.) is highly expressed in the phloem and could be involved in regulating proteolysis in the sieve elements. To further investigate the physiological role of SaPIN2a, we have produced transgenic lettuce (Lactuca sativa L.) expressing SaPIN2a from the CaMV35S promoter by Agrobacterium-mediated transformation. Stable integration of the SaPIN2a cDNA and its inheritance in transgenic lines were confirmed by Southern blot analysis and segregation analysis of the R1 progeny. SaPIN2a mRNA was detected in both the R0 and R1 transformants on northern blot analysis but the SaPIN2a protein was not detected on western blot analysis using anti-peptide antibodies against SaPIN2a. Despite an absence of significant inhibitory activity against bovine trypsin and chymotrypsin in extracts of transgenic lettuce, the endogenous trypsin-like activity in each transgenic line was almost completely inhibited, and the endogenous chymotrypsin-like activity moderately inhibited. Our finding that heterogeneously expressed SaPIN2a in transgenic lettuce inhibits plant endogenous protease activity further indicates that SaPIN2a regulates proteolysis, and could be potentially exploited for the protection of foreign protein production in transgenic plants.

Base Sequence↗

Phytotoxic, clastogenic and bioaccumulation effects of the environmental endocrine disruptor bisphenol A in various crops grown hydroponically.

The effects of the endocrine disruptor bisphenol A (BPA) at concentrations of 10 and 50 mg l(-1) were evaluated on the germination and morphology, micronuclei (MN) content in root tip cells and BPA bioaccumulation of hydroponic seedlings of broad bean (Vicia faba L.), tomato (Lycopersicon esculentum Mill.), durum wheat (Triticum durum Desf.) and lettuce (Lactuca sativa L.) after 6 and 21 days of growth. In general, BPA at any dose used did not inhibit germination and early growth (6 days) of seedlings of the species examined, with the exception of primary root length of tomato which decreased at the higher BPA dose. In contrast, an evident phytotoxicity was induced by BPA in all species after 21 days of growth with evident morphological anomalies and significant reductions of the lengths and fresh and dry weights of shoots and roots of seedlings. With respect to the nutrient medium without seedlings, BPA concentration decreased markedly during the growth period in the presence of broad bean and tomato seedlings, and limitedly in the presence of durum wheat and, especially, lettuce. Further, the presence of BPA measured in roots and shoots of broad bean and tomato after 21-day growth indicated that bioaccumulation of BPA had occurred. The number of MN in broad bean and durum wheat root tip cells increased markedly by treatment with BPA at both concentrations, thus suggesting a potential clastogenic activity of BPA in these species.

Benzhydryl Compounds↗

How does glutamine synthetase activity determine plant tolerance to ammonium?

The wide range of plant responses to ammonium nutrition can be used to study the way ammonium interferes with plant metabolism and to assess some characteristics related with ammonium tolerance by plants. In this work we investigated the hypothesis of plant tolerance to ammonium being related with the plants' capacity to maintain high levels of inorganic nitrogen assimilation in the roots. Plants of several species (Spinacia oleracea L., Lycopersicon esculentum L., Lactuca sativa L., Pisum sativum L. and Lupinus albus L.) were grown in the presence of distinct concentrations (0.5, 1.5, 3 and 6 mM) of nitrate and ammonium. The relative contributions of the activity of the key enzymes glutamine synthetase (GS; under light and dark conditions) and glutamate dehydrogenase (GDH) were determined. The main plant organs of nitrogen assimilation (root or shoot) to plant tolerance to ammonium were assessed. The results show that only plants that are able to maintain high levels of GS activity in the dark (either in leaves or in roots) and high root GDH activities accumulate equal amounts of biomass independently of the nitrogen source available to the root medium and thus are ammonium tolerant. Plant species with high GS activities in the dark coincide with those displaying a high capacity for nitrogen metabolism in the roots. Therefore, the main location of nitrogen metabolism (shoots or roots) and the levels of GS activity in the dark are an important strategy for plant ammonium tolerance. The relative contribution of each of these parameters to species tolerance to ammonium is assessed. The efficient sequestration of ammonium in roots, presumably in the vacuoles, is considered as an additional mechanism contributing to plant tolerance to ammonium nutrition.

Carbohydrate Metabolism↗

Phytotoxic effects and chemical analysis of leaf extracts from three Phytolaccaceae species in South Korea.

We analyzed phenolic compounds and other elements in leaf extracts and compared morphology of three species of the Phytolaccaceae family found in South Korea. To test allelochemical effects of the three Phytolacca species, we also examined seed germination and dry weight of seedlings of Lactuca indica and Sonchus oleraceus treated with leaf extracts. The concentrations of total phenolic compounds were exotic Phytolacca esculenta (3.9 mg/l), native Phytolacca insularis (4.4 mg/l), and exotic Phytolacca americana (10.2 mg/l). There was no significant difference in concentrations between P. esculenta and P. insularis, but the concentration of total phenolics in P. americana was two times higher than either P. esculenta or P. insularis. Analysis of aqueous extracts by HPLC showed seven phenolic compounds (gallic acid, protocatechuic acid, chlorogenic acid, caffeic acid, m-hydroxybenzoic acid, p-coumaric acid, and cinnamic acid). Total phenolics in P. americana were eight to 16 times higher than either P. esculenta or P. insularis, respectively. P. americana inhibited seed germination and dry weight of the two assay species. The phytotoxic effects of the two Phytolacca species were different, despite the fact that P. esculenta and P. insularis had similar levels of total phenolic compounds. We also found that P. americana had invaded Ullung Island, which suggested that P. americana had excellent adaptability to the environment. The three species of Phytolaccaceae in South Korea can be distinguished by their different allelopathic potentials and morphologies.

Biological Assay↗