Inheritance of resistance in <Emphasis Type="Italic">Solanum nigrum</Emphasis> to <Emphasis Type="Italic">Phytophthora infestans</Emphasis>

Solanum nigrum, black nightshade, is a wild non-tuber bearing hexaploid species with a high level of resistance to Phytophthora infestans (Colon et al. 1993), the causal agent of potato late blight, the most devastating disease in potato production. However, the genetic mode of resistance in S. nigrum is still poorly understood. In the present study, two S. nigrum accessions, 984750019 (N19) and #13, resistant (R) and susceptible (S), respectively, to three different isolates of P. infestans, were sexually crossed. The various kinds of progeny including F1, F2, F3, and backcross populations (BC₁; F1 × S), as well as two populations produced by self-pollinating the R parent and S parent, were each screened for susceptibility to P. infestans isolate MP 324 using detached leaf assays. Fifty seedling plant individuals of the F1 progeny were each resistant to this specific isolate, similarly to the seedling plants resulting from self-pollination of the resistant R parent. Thirty seedling plants obtained from self-pollination of the S parent were susceptible. Among a total of 180 F2 plants, the segregation ratio between resistant and susceptible plants was approximately 3: 1. Among the 66 seedling plants of the BC₁ progeny originating from crossing an F1 plant with the susceptible S parent, there were 26 susceptible and 40 resistant plants to P. infestans. The segregation patterns obtained indicated monogenic dominant inheritance of resistance to P. infestans isolate MP 324 in S. nigrum acc. 984750019. This gene, conferring resistance to P. infestans, may be useful for the transformation of potato cultivars susceptible to late blight.     

BABA effects on the behaviour of potato cultivars infected by <Emphasis Type="Italic">Phytophthora infestans</Emphasis> and <Emphasis Type="Italic">Fusarium solani</Emphasis>

Since most plants possess resistance mechanisms which can be induced upon pre-treatment with a variety of chemical compounds, the use of β-aminobutyric acid (BABA) as a defence inducer without reported toxic effect on the environment was studied. The aim of this work was to analyse the effectiveness of BABA to induce resistance against Phytophthora infestans and Fusarium solani in potato cultivars differing in their level of resistance to late blight. The behaviour of some components of biochemical mechanisms by which BABA increases resistance against P. infestans, as well as the effect of BABA on the activity of a potential pathogenic factor of F. solani, were studied. Plants with four applications of BABA throughout the crop cycle produced tubers more resistant to P. infestans and F. solani than non-treated plants. In addition, tuber slices from treated plants, inoculated with P. infestans, showed an increase in phenol and phytoalexin content. The aspartyl protease StAP1 accumulation was also higher in tubers obtained from treated plants and inoculated with P. infestans. This result was observed only in the more resistant potato cv. Pampeana, early after infection. In the potato–F. solani interaction, infected tubers coming from BABA-treated plants showed minor fungal proteolytic activity than infected, non-treated ones. For potato cvs Pampeana and Bintje, the BABA treatment improved the yield of harvested tubers. The number of tubers per plant and total weight of harvested tubers was greater for those obtained from treated plants with two early or four applications of BABA. The results show that the BABA treatment increases the resistance of potatoes but the degree of increase depends on the original level of resistance present in each cultivar.     

Resistance to <Emphasis Type="Italic">Phytophthora infestans</Emphasis>: exploring genes required for disease resistance in Solanaceae plants

The oomycete Phytophthora infestans is the causal agent of potato late blight, one of the most destructive and historically significant pathogens in agricultural production. A virus-induced gene silencing-based screening of the solanaceous model plant N. benthamiana resulted in revealing a wide range of resistance mechanisms of solanaceous plants against this pathogen. In this article, we present an overview of the various pathways involved in the N. benthamiana–P. infestans pathosystem, including some of the follow-up work that was triggered by these findings. The purpose of this review is to assemble these findings and integrate them into our current understanding of plant pathogen defense mechanisms and discuss their potential application for the development of potato resistance to P. infestans.     

Population studies on <Emphasis Type="Italic">Phytophthora infestans</Emphasis> on potatoes and tomatoes in southern Germany

Fifty-seven isolates of Phytophthora infestans from blighted potato foliage were collected in 1995 in southern Germany and analysed for mating type and sensitivity to metalaxyl. Fifty-six of them were characterised as A1 and one as A2 mating types. Resistance to metalaxyl was observed frequently: 53 isolates were resistant, three were partially sensitive, and one was sensitive. In a subsequent field study in 1999, 84 isolates collected from blighted potato and tomato foliage were analysed for mating type. Seventy-two were characterised as A1 and twelve as A2 mating types. The response of 76 isolates to metalaxyl and to propamocarb was tested. The majority (42) of the 76 isolates was classified as resistant to metalaxyl; 31 were partially sensitive and only three isolates were sensitive. The results with propamocarb were less discrete; 10 isolates were classified as resistant and three were clearly sensitive. AFLP fingerprinting was used to examine the genetic structure of the southern German P. infestans population collected in 1999 and indicated that the tested population can be sub-divided into a tomato group, a potato group and a mixed group containing isolates collected from both crops. The presence of Ia and IIa mitochondrial DNA haplotypes indicates that the German P. infestans isolates belong to the new pathogen population that has also been reported in neighbouring regions of Europe. The present study indicates that at the beginning of the season only a few genotypes were present, and the population became genetically more variable at the end of the growing season.     

Races of <Emphasis Type="Italic">Phytophthora infestans</Emphasis> isolated from potato in Hokkaido,Japan

The virulence of 29 isolates of Phytophthora infestans collected in potato fields in Hokkaido, Japan, in 2013 and 2014, was tested for race identification. Thirteen different races were identified, each of which had five to eight virulence factors. All of the isolates caused a virulent reaction against plants with R1 and R7, and most of the isolates caused a virulent reaction against plants with R3, R4, R10, and R11. On the other hand, no isolate was virulent against plants with R9. These results demonstrate that the current Japanese P. infestans population is more complex than the population in the 1990s from the viewpoint of race.     

Cytomolecular aspects of rice sheath blight caused by <Emphasis Type="Italic">Rhizoctonia solani</Emphasis>

Sheath blight, caused by anastomosis group 1-IA of Rhizoctonia solani Kühn (teleomorph Thanatephorus cucumeris (Frank) Donk), is one of the most destructive rice diseases worldwide. The pathogen is able to infect plants belonging to more than 27 families, including many economically important monocots and dicots such as rice, wheat, alfalfa, bean, peanut, soybean, cucumber, papaya, corn, potato, tomato and sugar beet. It is a soil borne necrotrophic fungus that survives in plant debris as sclerotia, which are small brown-to-black, rocklike reproductive structures. The sclerotia can survive in the soil for several years and infect rice plants at the water-plant interface in the flooded field by producing mycelia. Management of rice sheath blight requires an integrated approach based on the knowledge of each stage of the disease and cytomolecular aspects of rice defence responses against R. solani. This review summarizes current knowledge on molecular aspects of R. solani pathogenicity, genetic structure of the pathogen populations, and the rice-R. solani interaction with emphasis on cellular and molecular defence components such as signal transduction pathways, various plant hormones, host defence genes and production of defence-related proteins involved in basal and induced resistance in rice against sheath blight disease.     

Transmission of <Emphasis Type="Italic">Colletotrichum coccodes via</Emphasis> tomato seeds

Anthracnose of tomato caused by Colletotrichum coccodes is a devastating disease of ripe fruits. This pathogen may also infect tomato roots, stems and leaves. In the present study, C. coccodes is shown to be capable of contaminating seeds collected from artificially inoculated tomato fruits. Seedlings germinating from these infected seeds exhibited disease symptoms and therefore may transmit the pathogen to the next crop. The proportion of infected seeds ranged between 20% and 63% in all C. coccodes isolates tested and correlated with the aggressiveness of the isolates to tomato fruits. Fungicidal treatment of the collected seeds reduced, but did not eliminate, seed infection. A transgenic C. coccodes isolate expressing green fluorescent protein was used to visualize the pathogen. Mycelium was observed both on surfaces of the seed coat and within 1% of the embryos.     

Evaluation of biocontrol preparations and plant extracts for the control of <Emphasis Type="Italic">Phytophthora infestans</Emphasis> on potato leaves

The potential of biocontrol products and plant extracts for control of late blight on potato plants, caused by Phytophthora infestans was evaluated in detached leaf assays and on potted plants. Based on an initial screening of 22 preparations and plant extracts, the 10 most active treatments were selected for further investigation. In the detached leaf assays the commercial preparations Elot-Vis, Serenade and Trichodex, and plant extracts of Rheum rhabarbarum and Solidago canadensis showed a significant effect on the level of infestation by P. infestans. However, none of the treatments was as effective as copper. In the case of Serenade, the metabolites produced by its active micro-organism, Bacillus subtilis, were demonstrated to be the effective component of the formulation, and not the micro-organism itself. In order to take curative and protective modes of action into account, the test substances were applied 24 h before, or 90 min after inoculation with P. infestans. Generally, better effects were obtained when the applications were made 24 h before inoculation. For defining the optimum time of application, potted plants were treated 72 or 24 h before, and 1 and 24 h after inoculation with P. infestans. In these tests, Trichodex showed no activity, while Elot-Vis gave best results when applied 1 day before inoculation. Serenade and the extracts of R. rhabarbarum and S. canadensis (all at 5% concentration) however, were effective when applied up to 3 days before and just after inoculation with P. infestans. The results of the experiments on potted plants indicated direct effects on the pathogen for all agents except the extract of S. canadensis, but other mode of actions, e.g. induced resistance, could not be ruled out. None of the treatments had a curative effect.     

Comparison of the development <Emphasis Type="Italic">in planta</Emphasis> of a pyrrolnitrin-resistant mutant of <Emphasis Type="Italic">Botrytis cinerea</Emphasis> and its sensitive wild-type parent isolate

Botrytis cinerea is able to build-up resistance to pyrrolnitrin, an antibiotic produced by diverse biocontrol agents, possibly compromising the durability of this method of disease control. The development of two near-isogenic lines of B. cinerea differing in their level of resistance to pyrrolnitrin was compared in tomato plants and on PDA medium. In tomato plants, significant differences in the percentage of infected petioles 1 day after inoculation and in symptom progression on petioles and stems were observed between the resistant mutant and the sensitive wild-type parent, suggesting a difference in their level of aggressiveness. Cytohistological investigations revealed that conidia of both near-isogenic lines germinated 6 h after inoculation and mycelium developed within petiole tissues 12 h after inoculation. However, while the wild-type parent isolate spread throughout the petiole and rapidly invaded the stem tissues via the leaf-abscission zone 72 h after inoculation, the pyrrolnitrin-resistant mutant failed to extend beyond petiole tissues to invade the stem. Moreover, 72 h after inoculation, the mycelial development of the pyrrolnitrin-resistant mutant was accompanied by abnormal glycogen accumulation and chlamydospore-like cell formation. In contrast, wild-type parent mycelium was normally structured with intensive colonization of stem tissues. Additionally, on PDA medium the mycelium of the pyrrolnitrin-resistant mutant was less vigorous than the wild-type isolate. These results suggest that the acquisition of pyrrolnitrin-resistance in B. cinerea is accompanied by changes in mycelial structure and reduction in mycelial growth, leading to a noticeable loss of aggressiveness on tomato plants.     

Differences in virulence of <Emphasis Type="Italic">Phytophthora capsici</Emphasis> isolates from a worldwide collection on host fruits

Phytophthora capsici causes root, crown, and fruit rot of vegetable and tropical hosts. Cucumber, zucchini, tomato, and pepper fruits were inoculated using 6-mm-diameter agar plugs of P. capsici, incubated in clear plastic boxes at room temperature (25 ± 2°C and 100% relative humidity), and virulence was estimated by measuring the lesion diameter, pathogen growth diameter, and pathogen sporulation density three (cucumber, zucchini) or four (tomato, pepper) days later. When isolates were grouped by genetic cluster, significant differences in virulence were observed on cucumber and zucchini, with isolates belonging to genetic cluster five causing larger lesions than isolates from genetic cluster six. On tomato, no significant differences were observed for isolates grouped by genetic cluster, but isolates from vegetable crops were generally more virulent than isolates from tropical hosts. Isolates from fabaceous hosts sporulated better on cucumber fruits than isolates from solanaceous hosts. Isolates from vegetable hosts sporulated better on zucchini than isolates from tropical hosts. No significant differences in lesion diameter were noted on pepper when isolates were grouped by host family of origin or genetic cluster, but differences in pathogen sporulation were apparent by host family. Our findings suggest that isolate characteristics such as host family of origin and genetic cluster membership may be used to guide initial isolate selection for cucurbit fruit resistance screening. Final isolate selection should incorporate the phenotypic and genetic diversity of P. capsici, including isolates with differing virulence to the host organ of interest.     

Post-infection activity of fungicides against <Emphasis Type="Italic">Phytophthora infestans</Emphasis> on tomato (<Emphasis Type="Italic">Solanum lycopersicum</Emphasis> L.)

Late blight caused by Phytophthora infestans is the most devastating disease of tomato (Solanum lycopersicum L.) and causes important economically losses if not properly controlled. Control is achieved mainly by preventive fungicide applications. However, even if curative applications are discouraged because they increase the risk of resistance development in the target pathogens, in practice fungicides may be applied also when the disease is already present, a situation that commonly occurs in the field. The aim of this work was to study the curative activity of several fungicides toward P. infestans to determine their efficacy when applied after the infection process. Nine trials were performed in greenhouse using potted tomato plants that were treated 24 h after inoculation. Disease severity was assessed three times from the development of the symptoms on the untreated plants and data analysed using a linear mixed model. Differences in post-infection control between the different chemical classes were found. Metalaxyl-M and cymoxanil showed the best curative activity while among the CAA fungicides, a good efficacy was expressed by dimethomorph. Interestingly, evidence of synergy between active ingredients having different modes of action was observed such as in the mixtures containing dimethomorph?+?ametoctradin, dimethomorph?+?pyraclostrobin and fosetyl-Al?+?propamocarb. This study provided useful information on the post-infection activity of some fungicides used to control tomato late blight and should be taken into account to perform more in depth studies at the field level and to improve the management strategies of the disease.     

Host-delivered RNAi-mediated root-knot nematode resistance in <Emphasis Type="Italic">Arabidopsis</Emphasis> by targeting <Emphasis Type="Italic">splicing factor</Emphasis> and <Emphasis Type="Italic">integrase</Emphasis> genes

Root-knot nematodes (RKNs) are one of the most important biotic factors limiting crop productivity in many crop plants. The major RKN control strategies include development of resistant cultivars, application of nematicides and crop rotation, but each has its own limitations. In recent years, RNA interference (RNAi) has become a powerful approach for developing nematode resistance. The two housekeeping genes, splicing factor and integrase, of Meloidogyne incognita were targeted for engineering nematode resistance using a host-delivered RNAi (HD-RNAi) approach. Splicing factor and integrase genes are essential for nematode development as they are involved in RNA metabolism. Stable homozygous transgenic Arabidopsis lines expressing dsRNA for both genes were generated. In RNAi lines of splicing factor gene, the number of galls, females and egg masses was reduced by 71.4, 74.5 and 86.6%, respectively, as compared with the empty vector controls. Similarly, in RNAi lines of the integrase gene, the number of galls, females and egg masses was reduced up to 59.5, 66.8 and 63.4%, respectively, compared with the empty vector controls. Expression analysis revealed a reduction in mRNA abundance of both targeted genes in female nematodes feeding on transgenic plants expressing dsRNA constructs. The silencing of housekeeping genes in the nematodes through HD-RNAi significantly reduced root-knot nematode infectivity and suggests that they will be useful in developing RKN resistance in crop plants.     

Reduced efficacy of fluazinam against <Emphasis FontCategory="SansSerif" Type="Italic">Phytophthora infestans</Emphasis> in the Netherlands

Phytophthora infestans is the causal organism of potato late blight, the most important disease in potato, the second most important arable crop in Europe. The P. infestans population in Europe is well known for its sudden changes in composition. Currently it is composed of a wide variety of genotypes, some of which are dominant clonal lines while others are rare or even unique to a year or location. Fungicides play a crucial role in the integrated control of late blight. Since its introduction in the Netherlands in 1992, fluazinam has been used in late blight control strategies in ware and starch potatoes. It has a broad spectrum of activity and is effective against a range of diseases including potato late blight. Fluazinam interrupts the pathogen cell’s energy production process by an uncoupling effect on oxidative phosphorylation. It is considered to have a low resistance risk. Until recently, reduced efficacy against fluazinam was not detected in P. infestans surveys in Europe. In this paper we present the finding of a new clonal lineage (EU_33_A2) of P. infestans in the Netherlands and the reduced efficacy of fluazinam to control one of the EU_33_A2 isolates in field experiments carried out in 2011 and 2015 under high disease pressure. The potential effects of this finding on practical late blight control strategies are discussed.     

Cytological events in tomato leaves inoculated with conidia of <Emphasis Type="Italic">Blumeria graminis</Emphasis> f. sp. <Emphasis Type="Italic">hordei</Emphasis> and <Emphasis Type="Italic">Oidium neolycopersici</Emphasis> KTP-01

Leaves of tomato and barley were inoculated with conidia of Blumeria graminis f. sp. hordei race 1 (R1) or Oidium neolycopersici (KTP-01) to observe cytological responses in search of resistance to powdery mildew. Both conidia formed appressoria at similar rates on tomato or barley leaves, indicating that no resistance was expressed during the prepenetration stage of these fungi. On R1-inoculated tomato leaves, appressoria penetrated the papillae, but subsequent haustorium formation was inhibited by hypersensitive necrosis in the invaded epidermal cells. On the other hand, KTP-01 (pathogenic to tomato leaves) successfully developed functional haustoria in epidermal cells to elongate secondary hyphae, although the hyphal elongation from some conidia was later suppressed by delayed hypersensitive necrosis in some haustorium-harboring epidermal cells. Thus, the present study indicated that the resistance of tomato to powdery mildew fungi was associated with a hypersensitive response in invaded epidermal cells but not the prevention of fungal penetration through host papilla.     

Characterization of Egyptian <Emphasis Type="Italic">Phytophthora infestans</Emphasis> population using simple sequence repeat markers

The plant pathogenic oomycete, Phytophthora infestans, is the causal agent of late blight disease in tomato and potato. For characterizing Egyptian P. infestans isolates by DNA marker analysis, 40 isolates of P. infestans were collected from different locations in Egypt during two growing seasons (2012/2013 and 2013/2014). The 40 isolates were grouped into seven genotypes, in which 24 alleles were detected. The identified genotypes were not completely associated with geographic location and sample collection years. These results provide genetic and geographical information for developing a program to manage late blight disease.     

Rice <Emphasis Type="Italic">OsPBL1</Emphasis> (ORYZA SATIVA ARABIDOPSIS PBS1-LIKE 1) enhanced defense of <Emphasis Type="Italic">Arabidopsis</Emphasis> against <Emphasis Type="Italic">Pseudomonas syringae</Emphasis> DC3000

The receptor-like cytoplasmic kinases (RLCK family VII) are required for plant defense against various pathogens. Previously, OsPBL1 (ORYZA SATIVA ARABIDOPSIS PBS1-LIKE 1) was isolated from rice as a potential RSV (rice stripe virus) resistant factor, but its physiological roles in plant defense are yet to be investigated. In this study, we demonstrated that OsPBL1increased defense against P. syringae in transgenic Arabidopsis. To ascertain the role of OsPBL1 gene in plant defense, OsPBL1 tagged with HA (i.e. Hemagglutinin) was overexpressed in Arabidopsis and examined for the resistance against Pseudomonas syringae pv. tomato DC3000 (i.e. Pst DC3000). At 3 dpi of Pst DC3000, transgenic Arabidopsis lines exhibited the reduced chlorotic lesion and propagation of P. syringae, compared to wild type. Elevated pathogen resistance of transgenic lines was correlated with increased H₂O₂ accumulation and callose deposition on the infected leaves. It was also revealed that expression levels of salicylic acid dependent genes such as PR1, PR2, and PR5, were induced higher in transgenic lines than wild type. Taken together, our data suggested that OsPBL1 exerted the role in defense against pathogen attacks in plant via mainly facilitating salicylic acid dependent pathway.     

GC-MS metabolite profiling of <Emphasis Type="Italic">Phytophthora infestans</Emphasis> resistant to metalaxyl

Phytophthora infestans is the most important potato pathogen worldwide. Various alternatives have been used to control the pathogen, including continuous applications of phenylamide fungicides which has caused a rapid development of resistance in populations of P. infestans. Despite the importance of the disease, metabolite profiling of fungicide-resistant P. infestans has not been reported. In vitro resistance of Phytophthora infestans isolates to metalaxyl was characterized and metabolic changes in resistant isolates were evaluated at low (0.5 mg/L) and high (100 mg/L) concentrations of the fungicide. About 70% of the isolates tested showed resistance to metalaxyl and a total of 49 metabolites were differently expressed in resistant isolates growing in the presence of the fungicide. Principal components analysis revealed a distinct metabolite profile of resistant isolates exposed to both low and high levels of metalaxyl. The main metabolites responsible for the clustering in both fungicide concentrations included fatty acids such as hexadecanoic and octadecanoic acids, sugars such as glucose and fructose, aminoacids such as proline and valine, and organic acids such as butanedioic and propanoic acids. Potential resistance-related metabolic pathways are mostly involved in the regulation of the pathogen’s membrane fluidity and included the fatty acid biosynthesis as well as the glycerophospholipid metabolism pathways. This is the first metabolomic-based characterization of fungicide resistance in plant pathogens.     

Effects of garlic (<Emphasis Type="Italic">Allium sativum</Emphasis>) juice containing allicin on <Emphasis Type="Italic">Phytophthora infestans</Emphasis> and downy mildew of cucumber caused by <Emphasis Type="Italic">Pseudoperonospora cubensis</Emphasis>

The volatile antimicrobial substance allicin (diallylthiosulphinate) is produced in garlic when the tissues are damaged and the substrate allicin (S-allyl-l-cysteine sulphoxide) mixes with the enzyme alliin-lyase (E.C.4.4.1.4). Allicin undergoes thiol-disulphide exchange reactions with free thiol groups in proteins and it is thought that this is the basis of its antimicrobial action. At 50 μg ml^-1, allicin in garlic juice inhibited the germination of sporangia and cysts and subsequent germ tube growth by Phytophthora infestans both in vitro and in vivo on the leaf surface. Disease severity in P. infestans-infected tomato seedlings was also reduced by spraying leaves with garlic juice containing allicin over the range tested (55–110 μg ml⁻¹) with an effectiveness ranging from approximately 45–100%. Similarly, in growth room experiments at concentrations from 50–1,000 μg ml⁻¹, allicin in garlic juice reduced the severity of cucumber downy mildew caused by Pseudoperonospora cubensis by approximately 50–100%. These results suggest a potential for developing preparations from garlic for use in specialised aspects of organic farming, e.g. for reducing pathogen inoculum potential and perhaps for use under glass in horticulture.