Sulfur Induces Resistance against Canker Caused by Pseudomonas syringae pv. actinidae via Phenolic Components Increase and Morphological Structure Modification in the Kiwifruit Stems.

Bacterial canker caused by Pseudomonas syringae pv. actinidiae (Psa) has led to considerable losses in all major kiwifruit-growing areas. There are no commercial products in the market to effectively control this disease. Therefore, the defense resistance of host plants is a prospective option. In our previous study, sulfur could improve the resistance of kiwifruit to Psa infection. However, the mechanisms of inducing resistance remain largely unclear. In this study, disease severity and protection efficiency were tested after applying sulfur, with different concentrations in the field. The results indicated that sulfur could reduce the disease index by 30.26 and 31.6 and recorded high protection efficiency of 76.67% and 77.00% after one and two years, respectively, when the concentration of induction treatments was 2.0 kg/m3. Ultrastructural changes in kiwifruit stems after induction were demonstrated by scanning electron microscopy (SEM) and transmission electron microscopy (TEM), and the activities of phenylalanine ammonia-lyase (PAL), peroxidase (POD) and polyphenol oxidase (PPO), and the accumulation of lignin were determined by biochemical analyses. Our results showed that the morphological characteristics of trichomes and lenticels of kiwifruit stem were in the best defensive state respectively when the sulfur concentration was 3.0 kg/m3 and 1.5 kg/m3. Meanwhile, in the range of 0.5 to 2.0 kg/m3, the sulfur could promote the chloroplast and mitochondria of kiwifruit stems infected with Psa to gradually return to health status, increasing the thickness of the cell wall. In addition, sulfur increased the activities of PAL, POD and PPO, and promoted the accumulation of lignin in kiwifruit stems. Moreover, the sulfur protection efficiency was positively correlated with PPO activity (p < 0.05) and lignin content (p < 0.01), which revealed that the synergistic effect of protective enzyme activity and the phenolic metabolism pathway was the physiological effect of sulfur-induced kiwifruit resistance to Psa. This evidence highlights the importance of lignin content in kiwifruit stems as a defense mechanism in sulfur-induced resistance. These results suggest that sulfur enhances kiwifruit canker resistance via an increase in phenolic components and morphology structure modification in the kiwifruit stems. Therefore, this study could provide insights into sulfur to control kiwifruit canker caused by Psa.

Ampelomyces strains isolated from diverse powdery mildew hosts in Japan: Their phylogeny and mycoparasitic activity, including timing and quantifying mycoparasitism of Pseudoidium neolycopersici on tomato.

A total of 26 Ampelomyces strains were isolated from mycelia of six different powdery mildew species that naturally infected their host plants in Japan. These were characterized based on morphological characteristics and sequences of ribosomal DNA internal transcribed spacer (rDNA-ITS) regions and actin gene (ACT) fragments. Collected strains represented six different genotypes and were accommodated in three different clades of the genus Ampelomyces. Morphology of the strains agreed with that of other Ampelomyces strains, but none of the examined characters were associated with any groups identified in the genetic analysis. Five powdery mildew species were inoculated with eight selected Ampelomyces strains to study their mycoparasitic activity. In the inoculation experiments, all Ampelomyces strains successfully infected all tested powdery mildew species, and showed no significant differences in their mycoparasitic activity as determined by the number of Ampelomyces pycnidia developed in powdery mildew colonies. The mycoparasitic interaction between the eight selected Ampelomyces strains and the tomato powdery mildew fungus (Pseudoidium neolycopersici strain KTP-03) was studied experimentally in the laboratory using digital microscopic technologies. It was documented that the spores of the mycoparasites germinated on tomato leaves and their hyphae penetrated the hyphae of Ps. neolycopersici. Ampelomyces hyphae continued their growth internally, which initiated the atrophy of the powdery mildew conidiophores 5 days post inoculation (dpi); caused atrophy 6 dpi; and complete collapse of the parasitized conidiphores 7 dpi. Ampelomyces strains produced new intracellular pycnidia in Ps. neolycopersici conidiophores ca. 8-10 dpi, when Ps. neolycopersici hyphae were successfully destroyed by the mycoparasitic strain. Mature pycnidia released spores ca. 10-14 dpi, which became the sources of subsequent infections of the intact powdery mildew hyphae. Mature pycnidia contained each ca. 200 to 1,500 spores depending on the mycohost species and Ampelomyces strain. This is the first detailed analysis of Ampelomyces strains isolated in Japan, and the first timing and quantification of mycoparasitism of Ps. neolycopersici on tomato by phylogenetically diverse Ampelomyces strains using digital microscopic technologies. The developed model system is useful for future biocontrol and ecological studies on Ampelomyces mycoparasites.

Ergot Alkaloids and their Hallucinogenic Potential in Morning Glories.

Naturally occurring and semisynthetic ergot alkaloids play a role in health care or as recreational drugs in Western and indigenous Mexican societies. Evidence is summarized that ergot alkaloids present in Central American Convolvulaceae like Turbina corymbosa, Ipomoea violacea, and Ipomoea asarifolia are colonized by different species of a newly described clavicipitaceous fungal genus named Periglandula. The fungi are associated with peltate glandular trichomes on the adaxial leaf surface of its host plants. The Periglandula fungi are not yet culturable in vitro but were demonstrated to have the capacity to synthesize ergot alkaloids. The alkaloids do not remain in the fungal mycelium but are translocated via the glandular trichomes into their plant host. Both fungi and host benefit from a symbiotic lifestyle. In evolutionary terms the alkaloid biosynthetic gene cluster in the Periglandula/Ipomoea symbiosis is likely to have a conserved (basic) structure while biosynthetic ergot gene clusters within the genera Claviceps and Epichloe were under ecological selection for alkaloid diversification.

Computer Vision for High-Throughput Quantitative Phenotyping: A Case Study of Grapevine Downy Mildew Sporulation and Leaf Trichomes.

Quantitative phenotyping of downy mildew sporulation is frequently used in plant breeding and genetic studies, as well as in studies focused on pathogen biology such as chemical efficacy trials. In these scenarios, phenotyping a large number of genotypes or treatments can be advantageous but is often limited by time and cost. We present a novel computational pipeline dedicated to estimating the percent area of downy mildew sporulation from images of inoculated grapevine leaf discs in a manner that is time and cost efficient. The pipeline was tested on images from leaf disc assay experiments involving two F1 grapevine families, one that had glabrous leaves (Vitis rupestris B38 × 'Horizon' [RH]) and another that had leaf trichomes (Horizon × V. cinerea B9 [HC]). Correlations between computer vision and manual visual ratings reached 0.89 in the RH family and 0.43 in the HC family. Additionally, we were able to use the computer vision system prior to sporulation to measure the percent leaf trichome area. We estimate that an experienced rater scoring sporulation would spend at least 90% less time using the computer vision system compared with the manual visual method. This will allow more treatments to be phenotyped in order to better understand the genetic architecture of downy mildew resistance and of leaf trichome density. We anticipate that this computer vision system will find applications in other pathosystems or traits where responses can be imaged with sufficient contrast from the background.

O-Acyl Sugars Protect a Wild Tobacco from Both Native Fungal Pathogens and a Specialist Herbivore.

O-Acyl sugars (O-AS) are abundant trichome-specific metabolites that function as indirect defenses against herbivores of the wild tobacco Nicotiana attenuata; whether they also function as generalized direct defenses against herbivores and pathogens remains unknown. We characterized natural variation in O-AS among 26 accessions and examined their influence on two native fungal pathogens, Fusarium brachygibbosum U4 and Alternaria sp. U10, and the specialist herbivore Manduca sexta At least 15 different O-AS structures belonging to three classes were found in N. attenuata leaves. A 3-fold quantitative variation in total leaf O-AS was found among the natural accessions. Experiments with natural accessions and crosses between high- and low-O-AS accessions revealed that total O-AS levels were associated with resistance against herbivores and pathogens. Removing O-AS from the leaf surface increased M. sexta growth rate and plant fungal susceptibility. O-AS supplementation in artificial diets and germination medium reduced M. sexta growth and fungal spore germination, respectively. Finally, silencing the expression of a putative branched-chain α-ketoacid dehydrogenase E1 ß-subunit-encoding gene (NaBCKDE1B) in the trichomes reduced total leaf O-AS by 20% to 30% and increased susceptibility to Fusarium pathogens. We conclude that O-AS function as direct defenses to protect plants from attack by both native pathogenic fungi and a specialist herbivore and infer that their diversification is likely shaped by the functional interactions among these biotic stresses.

Glycan-binding F-box protein from Arabidopsis thaliana protects plants from Pseudomonas syringae infection.

BACKGROUND: A small group of F-box proteins consisting of a conserved F-box domain linked to a domain homologous to the glycan-binding protein has been identified within the genome of Arabidopsis thaliana. Previously, the so-called F-box-Nictaba protein, encoded by the gene At2g02360, was shown to be a functional lectin which binds N-acetyllactosamine structures. Here, we present a detailed qRT-PCR expression analysis of F-box-Nictaba in Arabidopsis plants upon different stresses and hormone treatments. RESULTS: Expression of the F-box-Nictaba gene was enhanced after plant treatment with salicylic acid and after plant infection with the virulent Pseudomonas syringae pv. tomato strain DC3000 (Pst DC3000). ß-glucuronidase histochemical staining of transgenic Arabidopsis plants displayed preferential activity of the At2g02360 promoter in trichomes present on young rosette leaves. qRT-PCR analyses confirmed high expression of F-box-Nictaba in leaf trichomes. A. thaliana plants overexpressing the gene showed less disease symptoms after Pst DC3000 infection with reduced bacterial colonization compared to infected wild type and F-box-Nictaba knock-out plants. CONCLUSIONS: Our data show that the Arabidopsis F-box-Nictaba gene is a stress-inducible gene responsive to SA, bacterial infection and heat stress, and is involved in salicylic acid related plant defense responses. This knowledge enriched our understanding of the physiological importance of F-box-Nictaba, and can be used to create plants with better performance in changing environmental conditions.

Maize leaf trichomes represent an entry point of infection for Fusarium species.

Fifteen day old maize seedlings were inoculated with Fusarium graminearum, Fusarium proliferatum, and Fusarium verticillioides. More than 90 % F. proliferatum and F. verticillioides conidia and 50 % of F. graminearum formed one germ tube whereas the other 50 % of F. graminearum conidia formed two to three germ tubes. The germ tubes of F. graminearum conidia were longer than those of F. proliferatum and F. verticillioides. The three species of Fusarium infected bi-cellular trichomes by adhering and growing along the trichomes or by attaching to the cap cell of the trichomes 48 h after inoculation. Hyphae penetrated into the trichomes at the base, the side or at the top of the cap cells. The hyphae colonized the cap cells and then spread to base cells. Prickle trichomes were infected 72 h after inoculation. The hyphae either wrapped around prickle trichomes or formed a mass of hyphae around the top of prickle trichomes or formed appressorium. Macro trichomes were infected by F. graminearum 7 d after inoculation. Following penetration, the fungus spread to adjacent epidermal cells and to the subcuticle. This investigation provides the first assessment of F. graminearum, F. proliferatum, and F. verticillioides infection via trichomes of maize leaves.

Microbe-based technology ameliorates glandular trichomes, secondary metabolites and antioxidants in Pelargonium graveolens L'Hér.

BACKGROUND: Despite the vast exploration of microbes for plant health, there is a lack of knowledge about the synergistic effects of specific microorganisms in sustainable agriculture, especially in medicinal plants such as Pelargonium graveolens L'Hér. The aim of this study was to evaluate how synergistic microbes Trichoderma harzianum ThU, Glomus intraradices and Bacillus subtilis CIM affected crop productivity, secondary metabolites and glandular trichome number in P. graveolens. RESULTS: The results demonstrated a significant (P < 0.05) increase in plant growth, secondary metabolites, total chlorophyll, carotenoids, carbohydrates, total phenolics, total flavonoids, free radical-scavenging activity and total antioxidant capacity of P. graveolens treated with synergistic bioinoculants as compared with the control. Most interestingly, an increase in essential oil by 32% in the treatment with all three microbes was observed. Furthermore, the principal aroma compounds citronellol and geraniol also increased in the same treatment. A positive and direct correlation was observed between essential oil content and number of glandular trichomes in all treatments. CONCLUSION: The present study highlights an explicit amalgamation of prospective microbes showing potential for synergism that act as biostimulants in enhancing plant production and improving the antioxidant and aroma profile of P. graveolens. © 2016 Society of Chemical Industry.

The key role of peltate glandular trichomes in symbiota comprising clavicipitaceous fungi of the genus periglandula and their host plants.

Clavicipitaceous fungi producing ergot alkaloids were recently discovered to be epibiotically associated with peltate glandular trichomes of Ipomoea asarifolia and Turbina corymbosa, dicotyledonous plants of the family Convolvulaceae. Mediators of the close association between fungi and trichomes may be sesquiterpenes, main components in the volatile oil of different convolvulaceous plants. Molecular biological studies and microscopic investigations led to the observation that the trichomes do not only secrete sesquiterpenes and palmitic acid but also seem to absorb ergot alkaloids from the epibiotic fungal species of the genus Periglandula. Thus, the trichomes are likely to have a dual and key function in a metabolic dialogue between fungus and host plant.