Characterization of polymorphic microsatellite loci in the tropical ant‐plant Leonardoxa africana (Fabaceae: Caesalpinioideae)

Ten microsatellite loci were isolated from the African ant‐plant Leonardoxa africana (Fabaceae: Caesalpinioideae). They differentiate the two most divergent subspecies of this polytypic complex, L. a. africana and L. a. gracilicaulis, showing promise for the study of gene flow. In each of these two subspecies, high levels of within‐population variation were observed, with a number of alleles ranging from one to 10 in ssp. africana and from one to 7 in ssp. gracilicaulis, and heterozygosity from 0 to 0.933 in ssp. africana and to 0.867 in ssp. gracilicaulis. All loci amplified successfully in the two other subspecies of L. africana.     

Exploration of Phytoconstituents from Mussaenda roxburghii and Studies of Their Antibiofilm Effect

In the context of ethno botanical importance with no phytochemical investigations, Mussaenda roxburghii have been investigated to explore it's phytoconstituents and studies of their antibiofilm activity. Four compounds have been isolated from the aerial parts of this plant and were characterized as 2α,3β,19α,23‐tetrahydroxyurs‐12‐en‐28‐oic acid ( 1 ), β‐sitosterol glucoside ( 4 ), lupeol palmitate ( 5 ), and myoinositol ( 6 ). All these compounds were tested for antibacterial and antibiofilm activity against Pseudomonas aeruginosa. Compound 1 exhibited three times more antibiofilm activity with minimum inhibitory concentration (MIC) at 0.74 mm compared to that of streptomycin. Molecular docking studies exhibited a very high binding affinity of 1 with P. aeruginosa quorum sensing proteins and motility associated proteins viz. LasR and PilB, PilY1, PilT, respectively. Compound 1 was also found to be non‐cytotoxic against sheep RBC and murine peritoneal macrophages at selected sub‐MIC doses.     

Impact of Delftia tsuruhatensis and Achromobacter xylosoxidans on Escherichia coli dual-species biofilms treated with antibiotic agents

Recently it was demonstrated that for urinary tract infections species with a lower or unproven pathogenic potential, such as Delftia tsuruhatensis and Achromobacter xylosoxidans, might interact with conventional pathogenic agents such as Escherichia coli. Here, single- and dual-species biofilms of these microorganisms were characterized in terms of microbial composition over time, the average fitness of E. coli, the spatial organization and the biofilm antimicrobial profile. The results revealed a positive impact of these species on the fitness of E. coli and a greater tolerance to the antibiotic agents. In dual-species biofilms exposed to antibiotics, E. coli was able to dominate the microbial consortia in spite of being the most sensitive strain. This is the first study demonstrating the protective effect of less common species over E. coli under adverse conditions imposed by the use of antibiotic agents.     

Repellent,Insecticidal and Antimicrobial Activities of Leaf Essential Oils from Three Eucalyptus Species

Developing effective and eco‐friendly antimicrobials and pesticides has become a highly important issue. The repellent, insecticidal and antimicrobial activity of essential oils (EOs) isolated by hydrodistillation from dried leaves of the three Eucalyptus species (E. cloeziana, E. umbellata and E. benthamii) were investigated. During GC/MS analysis, α‐pinene (47.36 %), 1,8‐cineol (38.53 %) and α‐pinene (35.31 %) were identified as major components of E. cloeziana, E. umbellata and E. benthamii, respectively. The EOs from E. cloeziana exhibited the longest effective protection time (465 min, at 50.0 % w/w) for humans among the EOs studied. The effective protection time was 30 min and 300 min at concentrations of 12.5 % (w/w) and 25.0 % (w/w), respectively. Fumigating insecticidal activity of EOs from three Eucalyptus species was tested by airtight fumigation in conical flask, which indicated that essential oils had a highly and rapidly insecticidal activity on Culex pipiens quinquefasciatus. The antimicrobial activity of EOs was evaluated by using disc diffusion and agar dilution methods. There was no significant difference in the antibacterial activity of EOs from E. cloeziana and E. umbellate and they had the same MICs (20 mL/L) on Staphylococcus aureus, Salmonella typhi, Bacillus subtilis and Escherichia coli. E. benthamii had the worst microbial inhibitory effect among the three Eucalyptus essential oils and the MIC value for the test species is 40 mL/L except for Rhodotorula Harrison (10 mL/L).     

Volatiles produced by soil‐borne endophytic bacteria increase plant pathogen resistance and affect tritrophic interactions

Volatile organic compounds (VOCs) released by soil microorganisms influence plant growth and pathogen resistance. Yet, very little is known about their influence on herbivores and higher trophic levels. We studied the origin and role of a major bacterial VOC, 2,3‐butanediol (2,3‐BD), on plant growth, pathogen and herbivore resistance, and the attraction of natural enemies in maize. One of the major contributors to 2,3‐BD in the headspace of soil‐grown maize seedlings was identified as Enterobacter aerogenes, an endophytic bacterium that colonizes the plants. The production of 2,3‐BD by E. aerogenes rendered maize plants more resistant against the Northern corn leaf blight fungus Setosphaeria turcica. On the contrary, E. aerogenes‐inoculated plants were less resistant against the caterpillar Spodoptera littoralis. The effect of 2,3‐BD on the attraction of the parasitoid Cotesia marginiventris was more variable: 2,3‐BD application to the headspace of the plants had no effect on the parasitoids, but application to the soil increased parasitoid attraction. Furthermore, inoculation of seeds with E. aerogenes decreased plant attractiveness, whereas inoculation of soil with a total extract of soil microbes increased parasitoid attraction, suggesting that the effect of 2,3‐BD on the parasitoid is indirect and depends on the composition of the microbial community.     

Antagonistic Activity of Neocosmospora vasinfecta var. africana (von Arx) Cannon & Hawksworth against Soil-borne Fungi

An isolate of Neocosmospora vasinfecta var. africana (F-2) was assayed for its antagonistic activity against 15 soil-borne fungi; 14 of them were important plant pathogens. The fungus exhibited a strong antibiotic effect against most of the fungi under test. While Pythium debaryanum, Macrophomina phaseolina, Phytophthora capsici, Sclerotinia sclerotiorum, Cochliobolus sativus and Alternaria alternata showed a very high degree of sensitivity as evidenced by the respectivezones of inhibition caused by the antagonist, Phoma betae, Rhizoctonia solani and Verticillium dahliae proved to be the most resistant. A mutual antibiosis existed between N. vasinfecta var. africana and five of the test fungi. The culture filtrate of the antagonist, diluted ten times with PDA, suppressed the mycelial growth of P. debaryanum and P. capsici completely. Whereas in this test R. solani proved to be the least sensitive, the remaining test fungi showed some differences in their susceptibility, but in any case their growth was inhibited significantly as compared to the controls.     

Influence of plant polyphenols and medicinal plant extracts on antibiotic susceptibility of Escherichia coli

Aims: To investigate the influence of polyphenols and plant extracts on the susceptibility of Escherichia coli to antibiotics. Methods and Results: Susceptibility of E. coli to antibiotics in the presence of extracts and polyphenols was estimated by the determination of the minimum inhibitory concentrations (MICs). To study gene expression, we used strains of E. coli carrying fusions between promoters of genes katG, sodA, iucC and structural β‐galactosidase gene. Treatment with polyphenols and some plant extracts significantly decreased the antibacterial effects of antibiotics, to a larger extent, ciprofloxacin. The most remarkable protective effect was observed for the extracts of Chamerion (Epilobium) angustifolium, Filipendula vulgaris, Tanacetum vulgare and Serratula coronata. These extracts increased the MICs of ciprofloxacin by four and more times. In case of kanamycin, extracts of Artemisia austriaca and Artemisia pontica increased MICs by four and eight times, respectively. Polyphenol quercetin also caused protective effect against ciprofloxacin, increasing the MIC by four times. A positive correlation was found between protective effects of polyphenols and extracts and their antioxidant activity. Conclusion: Medicinal plant extracts and polyphenols may protect cells of E. coli against antibiotic toxicity. Significance and Impact of the Study: The results of this study may be used to enhance the efficiency of antibacterial therapies.     

Essential oils as soil biofumigants for the control of the root‐knot nematode Meloidogyne incognita on tomato

The effectiveness of soil fumigation with 50, 100 and 200 µL kg^?1 soil of essential oils (EOs) from the plant species Eucalyptus citriodora, Eucalyptus globulus, Mentha piperita, Pelargonium asperum and Ruta graveolens was assessed against the root‐knot nematode Meloidogyne incognita on potted tomato. Plant growth parameters and number of galls, nematode eggs and juveniles on tomato roots were evaluated after two months of maintenance of the treated plants at 25°C in greenhouse. EOs of E. globulus and P. asperum significantly reduced nematode multiplication and gall formation on tomato roots at all the tested rates, whereas the EOs of E. citriodora, M. piperita and R. graveolens were more suppressive at levels greater than 50 µL kg^?1 soil. Biofumigation with EOs of E. globulus and P. asperum resulted also in the largest increase of tomato plant top and root biomass. The five samples of EOs had a different chemical composition as determined by GC and GC‐MS. Structure–activity relationship based on the main constituents of the tested EOs and their nematicidal effect on M. incognita is discussed.     

Antisense gene inhibition by phosphorothioate antisense oligonucleotide in Arabidopsis pollen tubes

Sexual reproduction is an essential biological event for proliferation of plants. The pollen tube (PT) that contained male gametes elongates and penetrates into the pistils for successful fertilization. However, the molecular mechanisms of plant fertilization remain largely unknown. Here, we report a transient inhibition of gene function using phosphorothioate antisense oligodeoxynucleotides (AS‐ODNs) without cytofectin, which is a simple way to study gene function in Arabidopsis thaliana PTs. The PTs treated with AS‐ODNs against both ANX1 and ANX2 showed short, knotted, and ruptured morphology in vitro/semi‐in vitro, whereas normal PT growth was shown in its sense control in vitro/semi‐in vitro. PT growth was impaired in a manner dependent on the dose of AS‐ODNs against both ANX1 and ANX2 above 10 μm . The treatment with AS‐ODNs against ROP1 and CalS5 resulted in waving PTs and in short PTs with a few callose plugs, respectively. The expression levels of the target genes in PTs treated with their AS‐ODNs were lower than or similar to those in the sense control, indicating that the inhibition was directly or indirectly related to the expression of each mRNA. The AS‐ODN against fluorescent protein (sGFP) led to reduced sGFP expression, suggesting that the AS‐ODN suppressed protein expression. This method will enable the identification of reproductively important genes in Arabidopsis PTs.     

Biocontrol Ability of <Emphasis Type="Italic">Lysobacter antibioticus</Emphasis> HS124 Against <Emphasis Type="Italic">Phytophthora</Emphasis> Blight Is Mediated by the Production of 4-Hydroxyphenylacetic Acid and Several Lytic Enzymes

Several rhizobacteria play a vital role in plant protection, plant growth promotion and the improvement of soil health. In this study, we have isolated a strain of Lysobacter antibioticus HS124 from rhizosphere and demonstrate its antifungal activity against various pathogens including Phytophthora capsici, a destructive pathogen of pepper plants. L. antibioticus HS124 produced lytic enzymes such as chitinase, β-1,3-glucanase, lipase, protease, and an antibiotic compound. This antibiotic compound was purified by diaion HP-20, silica gel, sephadex LH-20 column chromatography and high performance liquid chromatography. The purified compound was identified as 4-hydroxyphenylacetic acid by gas chromatography-electron ionization (GC-EI) and gas chromatography-chemical ionization (GC-CI) mass spectrometry. This antibiotic exhibited destructive activity toward P. capsici hyphae. In vivo experiments utilizing green house grown pepper plants demonstrated the protective effect of L. antibioticus HS124 against P. capsici. The growth of pepper plants treated with L. antibioticus culture was enhanced, resulting in greater protection from fungal disease. Optimum growth and protection was found when cultures were grown in presence of Fe(III). Additionally, the activities of pathogenesis-related proteins such as chitinase and β-1,3-glucanase decreased in roots, but increased in leaves with time after treatment compared to controls. Our results demonstrate L. antibioticus HS124 as a promising candidate for biocontrol of P. capsici in pepper plants.     

Herbicidal activity of volatile oils from Eucalyptus citriodora against Parthenium hysterophorus

The herbicidal effect of volatile oils from leaves of Eucalyptus citriodora against the noxious weed Parthenium hysterophorus was tested. In a laboratory bioassay, seed germination and seedling length, chlorophyll content and respiratory activity of Parthenium decreased with increased concentration of eucalypt oils from 0.2 to 5.0 nL mL^‐1. Germination was completely inhibited at 5.0 nL mL^‐1 eucalyptus oils. Further, for 4‐week‐old plants of Parthenium sprayed with different concentrations of volatile oils, visible damage increased and chlorophyll content and respiratory activity decreased with increased concentration from 0 to 100 μL mL^‐1, the week after spraying. At concentrations up to 50 μL mL^‐1, plants showed some recovery over time but plants sprayed with 75 and 100 μL mL^‐1 died 2 weeks after treatment. Plants sprayed with 50 μL mL^‐1 and higher concentrations of eucalypt oils were desiccated and wilted in appearance. At concentrations of 5–75 μL mL^‐1, eucalypt oils caused a rapid electrolyte leakage from the Parthenium plants thereby indicating an effect on membrane integrity. It is concluded that volatile oils from E. citriodora possess weed‐suppressing ability and could be used as a potential bioherbicide for future weed management programmes.     

Evaluation of the antimicrobial potential of two flavonoids isolated from limnophila plants

The antimicrobial potential of two bioflavonoids, i.e., 5,7‐dihydroxy‐4′,6,8‐trimethoxyflavone ( 1 ) and 5,6‐dihydroxy‐4′,7,8‐trimethoxyflavone ( 2 ), isolated from Limnophila heterophylla Benth . and L. indica (Linn .) Druce (Scrophulariaceae), respectively, were evaluated against the microbial strains Bacillus subtilis, Staphylococcus aureus, Escherichia coli, Salmonella typhimurium, Alternaria solani, and Candida albicans. Compounds 1 and 2 exhibited moderate but broad antimicrobial activities against both Gram‐positive and Gram‐negative bacteria and also against the fungal pathogens. Moreover, the mechanism of action of 1 and 2 on the cellular functions or structures of some of the microorganisms was studied. Compound 1 showed a bactericidal effect against E. coli and S. aureus (MICs of 200 and 250 μg/ml, resp.), while compound 2 was found to effectively kill B. subtilis by cell lysis. The growth of A. solani and C. albicans was inhibited by compounds 1 and 2 , respectively. The effects of the flavonoids on the cellular structures and the carbohydrate metabolic pathways were studied by scanning electron microscopy (SEM) of the treated cells and by assessing the specific activity of key enzymes of the pathways, respectively. At sublethal doses, they enhanced the activity of gluconeogenic fructose bisphosphatase, but decreased the activity of phosphofructokinase and isocitrate dehydrogenase, the key enzymes of the Embden Meyerhof Parnas pathway and the tricarboxylic acid cycle, respectively.     

IDL6‐HAE/HSL2 impacts pectin degradation and resistance to Pseudomonas syringae pv tomato DC3000 in Arabidopsis leaves

Plant cell walls undergo dynamic structural and chemical changes during plant development and growth. Floral organ abscission and lateral root emergence are both accompanied by cell‐wall remodeling, which involves the INFLORESCENCE DEFICIENT IN ABSCISSION (IDA)‐derived peptide and its receptors, HAESA (HAE) and HAESA‐LIKE2 (HSL2). Plant cell walls also act as barriers against pathogenic invaders. Thus, the cell‐wall remodeling during plant development could have an influence on plant resistance to phytopathogens. Here, we identified IDA‐like 6 (IDL6), a gene that is prominently expressed in Arabidopsis leaves. IDL6 expression in Arabidopsis leaves is significantly upregulated when the plant is suffering from attacks of the bacterial Pseudomonas syringae pv. tomato (Pst) DC3000. IDL6 overexpression and knockdown lines respectively decrease and increase the Arabidopsis resistance to Pst DC3000, indicating that the gene promotes the Arabidopsis susceptibility to Pst DC3000. Moreover, IDL6 promotes the expression of a polygalacturonase (PG) gene, ADPG2, and increases PG activity in Arabidopsis leaves, which in turn reduces leaf pectin content and leaf robustness. ADPG2 overexpression restrains Arabidopsis resistance to Pst DC3000, whereas ADPG2 loss‐of‐function mutants increase the resistance to the bacterium. Pst DC3000 infection elevates the ADPG2 expression partially through HAE and HSL2. Taken together, our results suggest that IDL6‐HAE/HSL2 facilitates the ingress of Pst DC3000 by promoting pectin degradation in Arabidopsis leaves, and Pst DC3000 might enhance its infection by manipulating the IDL6‐HAE/HSL2‐ADPG2 signaling pathway.     

Phytochemical Diversity,Classification and Antibacterial Activity of Some Medicinal Plant Species from Tabuk (Saudi Arabia)

The main objectives of this study were to investigate the intra-specific and inter-specific phytochemical diversity and classification of nine important medicinal plant species from Tabuk region (KSA), namely (Pulicaria undulata L., Pulicaria incisa Lam., Artemisia herba-alba Asso., Artemisia monosperma Delile, Artemisia judaica L. and Achillea fragrantissima Forssk. from Asteraceae family, Ducrosia flabellifolia Boiss. from Apiaceae family, Thymus vulgaris L. and Lavandula coronopifolia Poir. from Lamiaceae family); to evaluate the antibacterial potentials of the plant extracts, and to inspect the possible associations between phytochemical diversity and contents of different phytochemical classes with the antibacterial activities of plant extracts. GC/MS technique was used to identify phytochemicals in the plant extracts. The standard disk diffusion technique was used to conduct the antibiotic susceptibility against four pathogenic bacterial species (two Gram positive: Staphylococcus aureus and Bacillus subtilis and two Gram negative species: Pseudomonas aeruginosa and Escherichia coli. A total of 160 different phytochemicals belonging to 30 compound classes were separated and identified. A. fragrantissima had the highest phytochemical diversity and P. incisa had the lowest one. Phytochemical beta diversity was 6.2362. Ethanol outperformed other extraction solvents in terms of antibacterial activity, while Pulicaria undulata and T. vulgaris ranked highest among plants in this regard. Gram positive bacterial species were more sensitive to plant extracts compared to Gram negative species. Phytochemical diversity and antibacterial activity of plant extracts against E. coli and P. aeruginosa were positively correlative, terpenoid and benzene & substituted derivative contents exhibited significant (p<0.05) positive correlations with the antibacterial activity against E. coli, terpenoid contents also showed positive correlation with activity against P. aeruginosa; benzene & derivative showed positive correlation with activity against the rest of bacterial species,     

Effects of diet,habitat, and phylogeny on the fecal microbiome of wild African savanna (Loxodonta africana) and forest elephants (L. cyclotis)

The gut microbiome, or the community of microorganisms inhabiting the digestive tract, is often unique to its symbiont and, in many animal taxa, is highly influenced by host phylogeny and diet. In this study, we characterized the gut microbiome of the African savanna elephant (Loxodonta africana) and the African forest elephant (Loxodonta cyclotis), sister taxa separated by 2.6–5.6 million years of independent evolution. We examined the effect of host phylogeny on microbiome composition. Additionally, we examined the influence of habitat types (forest versus savanna) and diet types (crop‐raiding versus noncrop‐raiding) on the microbiome within L. africana. We found 58 bacterial orders, representing 16 phyla, across all African elephant samples. The most common phyla were Firmicutes, Proteobacteria, and Bacteroidetes. The microbiome of L. africana was dominated by Firmicutes, similar to other hindgut fermenters, while the microbiome of L. cyclotis was dominated by Proteobacteria, similar to more frugivorous species. Alpha diversity did not differ across species, habitat type, or diet, but beta diversity indicated that microbial communities differed significantly among species, diet types, and habitat types. Based on predicted KEGG metabolic pathways, we also found significant differences between species, but not habitat or diet, in amino acid metabolism, energy metabolism, and metabolism of terpenoids and polyketides. Understanding the digestive capabilities of these elephant species could aid in their captive management and ultimately their conservation.     

Predicting the risk of non-target damage to a close relative of a target weed using sequential no-choice tests,paired-choice tests and olfactory discrimination experiments

We investigated host-plant utilisation by the candidate biocontrol agent Paradibolia coerulea (Coleoptera: Chrysomelidae) on the target plant Spathodea campanulata Beauv. (Bignoniaceae) and a closely related non-target plant, Kigelia africana (Lam.) Benth. (Bignoniaceae). Paired-choice and sequential no-choice experiments were performed and coupled with olfactory discrimination experiments to test the insects’ responses to volatiles from both plant species as well as to cues from conspecific beetles. Although K. africana was utilised by P. coerulea, S. campanulata was preferred for both adult feeding and oviposition. Interestingly, whereas females were attracted to olfactory cues emitted by S. campanulata, males demonstrated no such olfactory discrimination. Females were also attracted to cues deposited by males, and males were deterred by cues from other males, but neither sex responded to female olfactory cues. Very few eggs were recorded on K. africana and none of the larvae that hatched on K. africana survived the first instar. Both S. campanulata and K. africana are suitable for adult feeding, but persistent utilisation of K. africana in the field is unlikely because larval development is only possible on S. campanulata and because the adult females are strongly attracted to volatiles emitted by the target plant. Nevertheless, if P. coerulea is released as a biocontrol agent, spill-over adult feeding could potentially occur on K. africana growing sympatrically with S. campanulata. Because P. coerulea cannot complete its development on K. africana, non-target damage will only occur where the target plant is present, with an intensity dependent on densities of adult beetles locally.     

Elevated atmospheric CO2 increases microbial growth rates in soil: results of three CO2 enrichment experiments

Increasing the belowground translocation of assimilated carbon by plants grown under elevated CO₂ can cause a shift in the structure and activity of the microbial community responsible for the turnover of organic matter in soil. We investigated the long‐term effect of elevated CO₂ in the atmosphere on microbial biomass and specific growth rates in root‐free and rhizosphere soil. The experiments were conducted under two free air carbon dioxide enrichment (FACE) systems: in Hohenheim and Braunschweig, as well as in the intensively managed forest mesocosm of the Biosphere 2 Laboratory (B2L) in Oracle, AZ. Specific microbial growth rates (μ) were determined using the substrate‐induced respiration response after glucose and/or yeast extract addition to the soil. For B2L and both FACE systems, up to 58% higher μ were observed under elevated vs. ambient CO₂, depending on site, plant species and N fertilization. The μ‐values increased linearly with atmospheric CO₂ concentration at all three sites. The effect of elevated CO₂ on rhizosphere microorganisms was plant dependent and increased for: Brassica napus=Triticum aestivum<Beta vulgaris<Populus deltoides. N deficiency affected microbial growth rates directly (N limitation) and indirectly (changing the quantity of fine roots). So, 50% decrease in N fertilization caused the overall increase or decrease of microbial growth rates depending on plant species. The μ‐value increase was lower for microorganisms growing on yeast extract then for those growing on glucose, i.e. the effect of elevated CO₂ was smoothed on rich vs. simple substrate. So, the r/K strategies ratio can be better revealed by studying growth on simple (glucose) than on rich substrate mixtures (yeast extract). Our results clearly showed that the functional characteristics of the soil microbial community (i.e. specific growth rates) rather than total microbial biomass amount are sensitive to increased atmospheric CO₂. We conclude that the more abundant available organics released by roots at elevated CO₂ altered the ecological strategy of the soil microbial community specifically a shift to a higher contribution of fast‐growing r‐selected species was observed. These changes in functional structure of the soil microbial community may counterbalance higher C input into the soil under elevated atmospheric CO₂ concentration.     

Target Range of Zwittermicin A, an Aminopolyol Antibiotic from Bacillus cereus

Zwittermicin A is a novel antibiotic produced by Bacillus cereus UW85, which suppresses certain plant diseases in the laboratory and in the field. We developed a rapid method for large-scale purification of zwittermicin A and then studied the in vitro activity of zwittermicin A against bacteria, fungi, and protists. Zwittermicin A was highly active against the Oomycetes and their relatives, the algal protists, and had moderate activity against diverse Gram-negative bacteria and certain Gram-positive bacteria as well as against a wide range of plant pathogenic fungi. Zwittermicin A was more active against bacteria and fungi at pH 7–8 than at pH 5–6. When zwittermicin A was combined with kanosamine, another antibiotic produced by B. cereus, the two acted synergistically against Escherichia coli and additively against Phytophthora medicaginis, an Oomycete. The results indicate that there are diverse potential applications of this new class of antibiotic. Received: 1 December 1997 / Accepted: 9 January 1998