A phcA− marker-free mutant of Ralstonia solanacearum as potential biocontrol agent of tomato bacterial wilt

A phcA⁻ mutant of Ralstonia solanacearum strain ZJ3721 was created in a marker-free method. Expression of virulence-associated genes such as xpsR, egl, tek and epsE was significantly suppressed in the phcA⁻ mutant. The ability of the mutant to control tomato bacterial wilt was evaluated by potting experiment. Results showed that application of mutant with wild type (WT) at the same time only delayed the development of wilt for about one day and the population of WT in tomato rhizosphere soil was nearly 70-fold higher than that of the mutant, resulting in a 90% disease incidence at last, as high as that of control. If the phcA⁻ mutant was applied three days earlier than WT pathogen, tomato wilt disease incidence was only 6%, 80% lower than that of control and population of WT was about 0.5-fold as much as that of mutant. Under hydroponic conditions, phcA⁻ mutant significantly triggered the expressions of genes in salicylic acid pathway but inhibited the expressions of genes in jasmonic acid (JA) and ethylene (ET) pathways. The expressions of PR-1a and GluA genes (salicylic acid pathway) in phcA⁻ mutant were 66-fold and 7.5-fold higher than in WT pathogen after three days of inoculation.     

A review on the hybrids of hydroxycinnamic acid as multi-target-directed ligands against Alzheimer’s disease

Alzheimer’s disease (AD), a complex chronic progressive central nervous system degenerative disease and a public health problem of the world, often characters cognitive dysfunction accompaning aggression and depression, and may lead to death. More attentions should be paid on it because there is no modified strategy against AD till now. AD is featured with the loss of cholinergic neurons, the amyloid-beta peptide (Aβ) plaques and the neurofibrillary tangles and several hypotheses were established to explain the pathogenesis of AD. Hydroxycinnamic acids, including caffeic acid (CA) and ferulic acid (FA) are widely distributed in natural plants and fruits. CA and FA exert various pharmacological activities, including anti-inflammatory, antioxidant, neuroprotection, anti-amyloid aggregation and so on. All these pharmacological activities are associated with the treatment of AD. Here we summarized the pharmacological activities of CA and FA, and their hybrids as multi-target-directed ligands (MTDLs) against AD. The future application of CA and FA was also discussed, hoping to provide beneficial information for the development of CA- and FA-based MTDLs against AD.     

Synthesis of long-chain fatty acid derivatives as a novel anti-Alzheimer’s agent

In order to develop new drugs for Alzheimer’s disease, we prepared 17 fatty acid derivatives with different chain lengths and different numbers and positions of double bonds by using Wittig reaction and stereospecific hydrogenation of triple bonds as key reactions. Among them, (4Z,15Z)-octadecadienoic acid (10) and (23Z,34Z)-heptatriacontadienoic acid (16) showed the most potent neurite outgrowth activities on Aβ(25–35)-treated rat cortical neurons, which activities were comparable to that of a positive control, NGF. Both fatty acids 10 and 16 possess two (Z)-double bonds at the n-3 and n-14 positions, which might be important for the neurite outgrowth activity.     

Backbone resonance assignment of Alt a 1, a unique β-barrel protein and the major allergen of Alternaria alternata

Alt a 1 is the major allergen of the fungus Alternaria alternata and can be found in the cell wall of its spores. It is a cysteine linked homodimeric protein with a unique β-barrel fold as recently revealed by X-ray crystallography. Despite the elucidation of its structure, its biological function remains unknown. For Alternaria-sensitized patients, contact leads to respiratory allergy and in severe cases to asthma-related death. Here we report the sequence-specific Alt a 1 backbone ¹H, ¹⁵N and ¹³C chemical shift assignment.     

Silica-supported fluoroboric acid (HBF4–SiO2) catalyzed highly productive synthesis of thiomorpholides as activators of l-asparaginase as well as the antioxidant agent

An efficient solvent-free procedure for the synthesis of thiomorpholides in the presence of a catalytic amount of solid-supported fluoroboric acid (HBF₄–SiO₂) is described. The advantages of this method are high yields, short reaction times, ease of product isolation, low cost, and the catalyst can be recycled for a number of times without significant loss of activity. Three thiomorpholides possessing electron-donating group (4c, 4g, and 4h) were exhibiting excellent stimulatory activities against Erwinia carotovora l-asparaginase. The most potent activator, compound 4h displayed the following kinetic parameters, K_m = 75 μM and V_max = 1000 μmol mg^?1 min^?1 and K_A = 0.985 μM. Furthermore, these compounds (4g, 4h, 4c, 4f, 4a, and 4d) have also shown promising 2,2′-diphenyl-1-picrylhydrazyl (DPPH) reducing antioxidant activity (21–36%) at 1 mM concentration as compared to standard butylated hydroxyl anisole (72% at 1 mM).     

The effect of genetic complexity on the time-course of ribonucleic acid–deoxyribonucleic acid hybridization

1. The rate of RNA-DNA hybridization was studied under conditions of RNA excess, with RNA synthesized in vitro. The initial rate of the reaction was proportional to the initial RNA concentration. Throughout the observed course of the reaction there was a linear relationship between the reciprocal of the amount of RNA hybridized/mug. of DNA and the reciprocal of time. The slope of the reciprocal plot was inversely proportional to the initial RNA concentration. 2. A comparison was made of the hybridization of DNA from Escherichia coli and from bacteriophages T4 and lambda with homologous RNA. The initial rate of hybridization was inversely proportional to the genetic complexity of the hybridizing system. The slope of the reciprocal-time plot was directly proportional to genetic complexity. These results are interpreted to indicate that the rate of hybridization reflects the mean concentration of the various unique RNA species in a preparation.     

Hrp?mutant bacteria as biocontrol agents: Toward a sustainable approach in the fight against plant pathogenic bacteria

Sustainable agriculture necessitates development of environmentally safe methods to protect plants against pathogens. Among these methods, application of biocontrol agents has been efficiently used to minimize disease development. Here we review current understanding of mechanisms involved in biocontrol of the main Gram-phytopathogenic bacteria-induced diseases by plant inoculation with strains mutated in hrp (hypersensitive response and pathogenicity) genes. These mutants are able to penetrate plant tissues and to stimulate basal resistance of plants. Novel protection mechanisms involving the phytohormone abscisic acid appear to play key roles in the biocontrol of wilt disease induced by Ralstonia solanacearum in Arabidopsis thaliana. Fully understanding these mechanisms and extending the studies to other pathosystems are still required to evaluate their importance in disease protection.     

Expression of a Clostridium perfringens genome-encoded putative N-acetylmuramoyl–l-alanine amidase as a potential antimicrobial to control the bacterium

Clostridium perfringens is a gram-positive, spore-forming anaerobic bacterium that plays a substantial role in non-foodborne human, animal, and avian diseases as well as human foodborne disease. Previously discovered C. perfringens bacteriophage lytic enzyme amino acid sequences were utilized to identify putative prophage lysins or autolysins by BLAST analyses encoded by the genomes of C. perfringens isolates. A predicted N-acetylmuramoyl–l-alanine amidase or MurNAc–LAA (also known as peptidoglycan aminohydrolase, NAMLA amidase, NAMLAA, amidase 3, and peptidoglycan amidase; EC 3.5.1.28) was identified that would hydrolyze the amide bond between N-acetylmuramoyl and l-amino acids in certain cell wall glycopeptides. The gene encoding this protein was subsequently cloned from genomic DNA of a C. perfringens isolate by polymerase chain reaction, and the gene product (PlyCpAmi) was expressed to determine if it could be utilized as an antimicrobial to control the bacterium. By spot assay, lytic zones were observed for the purified amidase and the E. coli expression host cellular lysate containing the amidase gene. Turbidity reduction and plate counts of C. perfringens cultures were significantly reduced by the expressed protein and observed morphologies for cells treated with the amidase appeared vacuolated, non-intact, and injured compared to the untreated cells. Among a variety of C. perfringens strains, there was little gene sequence heterogeneity that varied from 1 to 21 nucleotide differences. The results further demonstrate that it is possible to discover lytic proteins encoded in the genomes of bacteria that could be utilized to control bacterial pathogens.     

Insight on the interaction of polychlorobiphenyl with nucleic acid–base

The interaction between one polychlorobiphenyl (3,3′,4,4′,-tetrachlorobiphenyl, coded PCB77) and the four DNA nucleic acid–base is studied by means of quantum mechanics calculations in stacked conformations. It is shown that even if the intermolecular dispersion energy is the largest component of the total interaction energy, some other contributions play a non negligible role. In particular the electrostatic dipole-dipole interaction and the charge transfer from the nucleobase to the PCB are responsible for the relative orientation of the monomers in the complexes. In addition, the charge transfer tends to flatten the PCB, which could therefore intercalate more easily between DNA base pairs. From these seminal results, we predict that PCB could intercalate completely between two base pairs, preferably between Guanine:Cytosine pairs.

Neoseiulus idaeus (Acari: Phytoseiidae) as a potential biocontrol agent of the two-spotted spider mite, Tetranychus urticae (Acari: Tetranychidae) in papaya: performance on different prey stage – host plant combinations

Development, reproduction, and life table parameters of the phytoseiid mite Neoseiulus idaeus Denmark & Muma preying on various stages of Tetranychus urticae Koch fed on papaya or snap bean plant were studied at 25 ± 2°C, 77 ± 2% RH and 12L:12D photoperiod. Most characteristics of the predator, including the duration of the immature stages, preoviposition and oviposition periods, and female longevity did not differ significantly among the different food combinations. During the adult stage, N. idaeus females survived up to 14 days. The net reproductive rate, the mean generation time, and the intrinsic rate of natural increase of N. idaeus were very similar on all T. urticae stage – host plant combinations, ranging from 5.15 to 6.61 females, 10.15 to 12.62 days, and 0.150 to 0.168, respectively. These results indicate that T. urticae reared either on papaya or snap bean is an adequate prey for the development and reproduction of N. idaeus. Moreover, the current findings demonstrate that the host plant of T. urticae did not affect the predator. The implications of these results for the success of N. idaeus as a biological control agent of T. urticae in papaya orchards are discussed.     

Use of koji prepared with a high citric acid producing mutant of Aspergillus usamii as a raw material for Saké brewing

There is a possibility of developing a new kind of saké in which the refreshing sour taste of citric acid is introduced. In this study, we bred a new mutant of Aspergillus usamii mut. shiro-usamii that produced much citric acid. The koji prepared with the mutant contained about 20 mg of citric acid per gram of dry koji, twice that of the koji of the parental strain. The activities of a-amylase, glucoamylase, and acidic protease in the koji prepared with the mutant were 82%, 94%, and 95%, respectively, those of the parental strain. Using this koji with the mutant, saké was produced. The levels of citric acid and isoamyl acetate were 5.1 and 1.4 times, respectively, those of saké prepared with koji of A. oryzae. Sensory tests indicated that saké made with koji with the mutant was refreshingly sour, with a good aroma.     

Effect of livestock grazing in the partitions of a semiarid plant–plant spatial signed network

In recent times, network theory has become a useful tool to study the structure of the interactions in ecological communities. However, typically, these approaches focus on a particular kind of interaction while neglecting other possible interactions present in the ecosystem. Here, we present an ecological network for plant communities that consider simultaneously positive and negative interactions, which were derived from the spatial association and segregation between plant species. We employed this network to study the structure and the association strategies in a semiarid plant community of Cabo de Gata-Níjar Natural Park, SE Spain, and how they changed in 4 sites that differed in stocking rate. Association strategies were obtained from the partitions of the network, built based on a relaxed structural balance criterion. We found that grazing simplified the structure of the plant community. With increasing stocking rate species with no significant associations became dominant and the number of partitions decreased in the plant community. Independently of stocking rate, many species presented an associative strategy in the plant community because they benefit from the association to certain ‘nurse’ plants. These ‘nurses’ together with species that developed a segregating strategy, intervened in most of the interactions in the community. Ecological networks that combine links with different signs provide a new insight to analyze the structure of natural communities and identify the species which play a central role in them.     

Entropy as a factor in the binding of γ-aminobutyric acid and nipecotic acid to the γ-aminobutyric acid transport system

Nipecotic acid is one of the most potent competitive inhibitors and alternative substrates for the high-affinity -aminobutyric acid transport system in neurons, but the structural basis of this potency is unclear. Because -aminobutyrate is a highly flexible molecule in solution, it would be expected to lose rotational entropy upon binding to the transport system, a change which does not favor binding. Nipecotic acid, in contrast, is a much less flexible molecule, and one would expect the loss of conformational entropy upon binding to be smaller thus favoring the binding of nipecotic acid over -aminobutyric acid. To investigate this possibility, the thermodynamic parameters, G°, H°, and S°, were determined for the binding of -aminobutyrate and nipecotic acid to the high affinity GABA transport system in synaptosomes. In keeping with expectations, the apparent entropy change for nipecotic acid binding (112±13 J·K^–1) was more favorable than the apparent entropy change for -aminobutyric acid binding (61.3±6.6 J·K^–1). The results suggest that restricted conformation per se is an important contributory factor to the affinity of nipecotic acid for the high-affinity transport system for -aminobutyric acid.This work was conducted when both authors were at the Department of Chemistry, University of Maryland, College Park.Special issue dedicated to Dr. Elling Kvamme.     

Evaluating factors that predict the structure of a commensalistic epiphyte–phorophyte network

A central issue in ecology is the understanding of the establishment of biotic interactions. We studied the factors that affect the assembly of the commensalistic interactions between vascular epiphytes and their host plants. We used an analytical approach that considers all individuals and species of epiphytic bromeliads and woody hosts and non-hosts at study plots. We built models of interaction probabilities among species to assess if host traits and abundance and spatial overlap of species predict the quantitative epiphyte–host network. Species abundance, species spatial overlap and host size largely predicted pairwise interactions and several network metrics. Wood density and bark texture of hosts also contributed to explain network structure. Epiphytes were more common on large hosts, on abundant woody species, with denser wood and/or rougher bark. The network had a low level of specialization, although several interactions were more frequent than expected by the models. We did not detect a phylogenetic signal on the network structure. The effect of host size on the establishment of epiphytes indicates that mature forests are necessary to preserve diverse bromeliad communities.     

Synthesis and biodistribution of a novel (⁹⁹m)TcN complex of norfloxacin dithiocarbamate as a potential agent for bacterial infection imaging

Achieving a (??m)Tc-labeled fluoroquinolone derivative as a single photon emission computed tomography (SPECT) tracer is considered to be of great interest. The norfloxacin dithiocarbamate (NFXDTC) was synthesized and radiolabeled with a [(??m)TcN]2(+) intermediate to form the (??m)TcN-NFXDTC complex in high yield. The radiochemical purity of (??m)TcN-NFXDTC was over 90%, as measured by thin layer chromatography (TLC) and high performance liquid chromatography (HPLC), without any notable decomposition at room temperature over a period of 6 h. The partition coefficient and electrophoresis results indicated that (??m)TcN-NFXDTC was lipophilic and neutral. The bacterial binding assay studies showed tht (??m)TcN-NFXDTC had a good binding affinity. Biodistribution results in bacterial infected mice showed that (??m)TcN-NFXDTC had a higher uptake at the sites of infection and better abscess/blood and abscess/muscle ratios than those of (??m)Tc-ciprofloxacin and (??m)TcN-CPFXDTC (CPFXDTC = ciprofloxacin dithiocarbamate). The biodistribution results of (??m)TcN-NFXDTC in bacterially infected mice and in mice with turpentine-induced abscesses indicated that (??m)TcN-NFXDTC was suited to be a bacteria-specific infection imaging agent. Single photon emission computed tomography (SPECT) image studies showed there was a visible accumulation in infection sites, suggesting that it would be a promising candidate for bacterial infection imaging.     

Bioactivity and structure–activity relationship of cinnamic acid derivatives and its heteroaromatic ring analogues as potential high-efficient acaricides against Psoroptes cuniculi

A series of cinnamic acid derivatives and its heteroaromatic ring analogues were synthesized and evaluated for acaricidal activity in vitro against Psoroptes cuniculi, a mange mite. Among them, eight compounds showed the higher activity with median lethal concentrations (LC₅₀) of 0.36–1.07 mM (60.4–192.1 µg/mL) and great potential for the development of novel acaricidal agent. Compound 40 showed both the lowest LC₅₀ value of 0.36 mM (60.4 µg/mL) and the smallest median lethal time (LT₅₀) of 2.6 h at 4.5 mM, comparable with ivermectin [LC₅₀ = 0.28 mM (247.4 µg/mL), LT₅₀ = 8.9 h], an acaricidal drug standard. SAR analysis showed that the carbonyl group is crucial for the activity. The type and chain length of the alkoxy in the ester moiety and the steric hindrance near the ester group significantly influence the activity. The esters were more active than the corresponding thiol esters, amides, ketones or acids. Replacement of the phenyl group of cinnamic esters with α-pyridyl or α-furanyl significantly increase the activity. Thus, a series of cinnamic esters and its heteroaromatic ring analogues with excellent acaricidal activity emerged.     

Gold nanoparticles functionalized by gadolinium–DTPA conjugate of cysteine as a multimodal bioimaging agent

The synthesis and characterization of gold nanoparticles coated with Gd-chelate (Au@GdL), where L is a conjugate of DTPA and cysteine, is described. These particles are obtained by the replacement of citrate from the gold nanoparticle surfaces with gadolinium chelate (GdL). The average size of Au@GdL is 14 nm with a loading of GdL reaching up to 2.9 × 10³ per particles, and they demonstrate very high R1 relaxivity (～10⁵ mM^?1 s^?1) as well as X-ray attenuation. The R1 relaxivity per [Gd] is 17.9 mM^?1 s^?1. The present system also exhibits macrophage-specific property, as demonstrated by histological and TEM images as well as CT and MR, rendering itself as a new class of T1 multimodal CT/MR contrast agent.