Transgenic potato plants overexpressing the pathogenesis-related STH-2 gene show unaltered susceptibility to Phytophthora infestans and potato virus X

The STH-2 gene is rapidly activated in potato leaves and tubers following elicitation or infection by Phytophthora infestans. However, its biochemical function remains unknown. In order to ascertain if STH-2 protein is directly involved in the defense of potato against pathogens, the STH-2 coding sequence under the control of the CaMV 35S promoter was introduced into potato plants. Transgenic plants expressing the STH-2 gene were analyzed for an altered pattern of susceptibility to a compatible race of P. infestans and to potato virus X. Results indicate that constitutive expression of the STH-2 gene did not reduce susceptibility of potato to these pathogens.     

Susceptibility of potato tubers to infection by Phytophthora infestans

Phytophthora infestans infects King Edward potato tubers more readily through inoculated eyes than through lenticels, but more lenticels than eyes became infected when whole tubers were sprayed with inoculum. The resistance of lenticels but not of eyes increased as tubers aged. The spores did not infect through intact periderm. The likelihood of tubers on plants grown in pots becoming infected by sporangial suspension poured on to the soil increased the nearer the tubers were to the soil surface, the stem, or the side of the pot. Naturally infected tubers, and those sprayed with sporangial suspension, had most eyes infected at the rose end, and most lenticels infected on the middle region of the tuber. Of naturally infected tubers, on which the site of infection could be identified, most were infected through eyes at the rose end.     

Potato phytoalexin elicitors in Phytophthora infestans spore germination fluids

Media from germinating spores of Phytophthora infestans contain substances that elicit accumulation of the phytoalexin rishitin in potato tuber slices. Gel permeation chromatography of media extracts indicates the presence of several substancec. The active substances can be precipitated with ammonium sulfate, are heat labile and pronase-sensitive, which suggests that they are proteins.     

Effect of wounding or infection by Phytophthora infestans on the contents of terpenoids in potato tubers

Inoculation of the potato cut surfaces with an incompatiblerace of Phytophthora infestans induced an accumulation of rishitin,but only a trace occurred when infected by a compatible race.When the tuber was cut, a large amount of steroid-glycoalkaloids(solanine) accumulated in the cut surface tissue, although onlya trace was found in intact tissue. Only a small amount of solanine,if any, was contained in the wound periderm tissue. Most ofthe solanine seemed to be distributed in tissue neighbouringthe newly formed meristematic tissue zone. Distribution of solanineas a function of distance from the cut surface was exponential.Infection of the cut surface by an incompatible race of Phytophthorainfestans reduced the accumulation of solanine. The higher theconcentration of zoospore used, the less the solanine content.It has been reported that the higher the concentration of zoosporesused for inoculation of the cut surface, the less the numberof renewed meristematic cells in the wound tissue. In experimentsusing fresh and aged tubers, a good correlation between thenumber of renewed meristematic cells and solanine content wasfound. The accumulation of solanine in the wound tissue andits reduction due to infection by an incompatible race may berelated to renewed meristematic cells formation and its reductioncaused by the infection. No drastic change in carotenoids or sterol contents was found2 days after cutting or inoculation, when the tubers were cut,or cut and then infected by the incompatible race. ¹ Studies on the phytoalexin (No. 10). (6) in References constitutes"Studies on the phytoalexin No. 9". ² Present adress: Plant Pathology Laboratory, Faculty of Agriculture,Nagoya University, Chikusa-ku, Nagoya, 464, Japan. (Received April 1, 1972; )     

Biochemical reactions of different tissues of potato (Solanum tuberosum) to zoospores or elicitors from Phytophthora infestans

The time courses of sesquiterpenoid phytoalexin accumulation were examined in compatible and incompatible interactions of leaves and tubers from five different R genotypes of potato (Solanum tuberosum) with corresponding pathotypes of Phytophthora infestans, as well as in non-host interactions of all five potato cultivars with Phytophthora megasperma f. sp. glycinea and in elicitor-treated tubers from five, and cell suspension cultures from two, of the cultivars. In tubers, rishitin and several structurally related sesquiterpene derivatives accumulated rapidly in non-host incompatible interactions, less rapidly in host incompatible interactions, and more slowly in compatible interactions. Treatment of tubers or cell cultures with fungal culture filtrate or arachidonic acid elicited in most cases a transient accumulation of the sesquiterpenoid phytoalexins. None of these compounds was detectable under any of the applied conditions either in infected or in elicitortreated leaves. Sesquiterpenoid phytoalexins might therefore be helpful, but appear not to be essential, in disease resistance of potato.Abbreviations CF concentrated culture filtrate of Pi - cv. cultivar - Pi Phytophthora infestans (numbering indicates pathotypes corresponding to R genes in potato) - Pmg Phytophthora megasperma f. sp. glycinea     

Salicylic Acid and Phenylalanine Ammonia-Lyase in Potato Plants Infected with the Causal Agent of Late Blight

In tuber tissues of potato (Solanum tuberosum L.) infected with an incompatible race of Phytophthora infestans (Mont.) de Bary, the activity of phenylalanine ammonia-lyase and the contents of free and bound salicylic acid (SA) considerably exceeded the corresponding indices in the tissues infected with a compatible race of the oomycete. The accumulation of the free form of SA apparently resulted from both enhanced SA biosynthesis and the liberation from the bound SA forms. SA accumulation in the incompatible host-pathogen combination presumes that SA participated in the local potato resistance to late blight.__________Translated from Fiziologiya Rastenii, Vol. 52, No. 4, 2005, pp. 573–577.Original Russian Text Copyright © 2005 by Panina, Gerasimova, Chalenko, Vasyukova, Ozeretskovskaya.     

Expression of the Phytophthora infestans ipiB and ipi0 genes in planta and in vitro

The ipiB and ipiO genes of the potato late blight fungus Phytophthora infestans (Mont.) de Bary were isolated from a genomic library in a screen for genes induced in planta. Expression of these genes was studied during pathogenesis on various host tissues and different host plants, some of which show specific resistance against P. infestans infection. During pathogenesis on leaves and tubers of the fully susceptible potato cultivar (cv.) Ajax and on leaves of the fully susceptible tomato cv. Moneymaker, the P. infestans ipiB and ipiO genes show a transient expression pattern with highest mRNA levels in the early stages of infection. During the interaction with leaves of the partially resistant potato cv. Pimpernel, the expression is also transient but accumulation and disappearance of the mRNAs is delayed. Also in P. infestans inoculated onto a race-specific resistant potato cultivar and onto the nonhost Solanum nigrum, ipiB and ipiO mRNA is detectable during the initial stages of infection. Apparently, the expression of the ipiB and the ipiO genes is activated in compatible, incompatible and nonhost interactions. In encysted zoospores, ipiB and ipiO mRNA accumulation was not detectable, but during cyst germination and appressorium formation on an artificial surface the genes are highly expressed. Expression studies in mycelium grown in vitro revealed that during nutrient starvation the expression of the ipiB and ipiO genes is induced. For ipiO gene expression, carbon deprivation appeared to be sufficient. The ipiO gene promoters contain a sequence motif that functions as a glucose repression element in yeast and this motif might be involved in the regulation of ipiO gene expression.     

Temporal and spatial patterns of 1, 3-β-glucanase and chitinase induction in potato leaves infected by Phytophthora infestans

Infection of potato (Solarium tuberosum L.) leaves with the fungal pathogen Phytophthora infestans caused a similar, strong and coordinated induction of 1, 3-β-glucanases and chitinases in compatible (plant susceptible) and incompatible (plant resistant) interactions of two selected plant cultivars with appropriate races of the fungus. The temporal and spatial patterns of 1, 3-β-glucanase induction were studied in further detail by immunohistochemical and in-situ hybridization methods. Accumulation of the protein was preceded by progressive activation of the corresponding gene, commencing near infection sites and spreading rapidly throughout the whole infected leaf as well as to adjacent, non-infected leaves. Protein and mRNA distribution patterns were nearly identical in compatible and incompatible interactions. In comparison with 1, 3-β-glucanase mRNA, phenyl-alanine ammonia-lyase mRNA accumulated more rapidly and remained restricted to the vicinity of fungal infection sites, in addition to its constitutive occurrence in the vascular bundles. Even more rapid than any detectable mRNA induction was the accumulation of auto fluorescing material in plant cells immediately surrounding fungal structures, particularly and invariably in incompatible interactions and less frequently in compatible interactions. It is concluded that cultivar-race-specific resistance is established early in the interaction of potato leaves with P. infestans and hence the observed massive accumulation of 1, 3-β-glucanase and chitinase is presumably not involved in determining this specificity.     

Free and conjugated forms of salicylic acid: Their content and role in the potato

Hydrolysis of conjugated forms of salicylic acid and accumulation of its free form was observed after infection of potato tubers (Solanum tuberosum L.) with an incompatible race of phytophthora or treatment with an elicitor (chitosan). Infection of tubers with a compatible race of the pathogen or treatment with a suppressor (laminarin) decreased both the degree of hydrolysis of conjugated forms of salicylic acid and the accumulation of its free form.__________Translated from Prikladnaya Biokhimiya i Mikrobiologiya, Vol. 41, No. 3, 2005, pp. 354–357.Original Russian Text Copyright © 2005 by Panina, Vasyukova, Ozeretskovskaya.     

Free and conjugated forms of salicylic acid: content and role in potato

Hydrolysis of conjugated forms of salicylic acid and accumulation of its free form was observed after infection of potato tubers (Solanum tuberosum L.) with an incompatible race of phytophthora or treatment with an elicitor (chitosan). Infection of tubers with a compatible race of the pathogen or treatment with a suppressor (laminarin) decreased both the degree of hydrolysis of conjugated forms of salicylic acid and the accumulation of its free form.     

Tuber surface microorganisms influence the susceptibility of potato tubers to late blight

Potato tubers (cvs Cara and Bintje) were grown in compost in a glasshouse and immature tubers harvested 57, 68 and 78 days after planting. Two moisture levels were imposed after the first harvest by disconnecting the water supply to one of the treatments and allowing the soil in that treatment to dry naturally. Tubers from wetter compost (59.4% moisture holding capacity) were more resistant to Phytophthora infestans than those from drier compost (14.7% moisture holding capacity) 78 days after planting. The potential causes of this difference were investigated. Aqueous extracts of wet compost did not inhibit the growth of P. infestans. The susceptibility of the internal tuber tissue, from which the periderm had been removed, was different to whole tuber susceptibility. The internal tissue of tubers from wet compost was more susceptible (cv. Cara), or as susceptible (cv. Bintje) as that of tubers from dry compost 78 days after planting. Fungi were isolated from the surface of whole tubers and there were no differences between the populations of potentially antagonistic fungal genera on tubers from wet and dry compost. As the experiment progressed, the number of bacteria per gram fresh weight on tubers grown in wet compost increased, whereas that on tubers from drier compost decreased (cv. Bintje) or remained similar (cv. Cara). There were significantly (P= 0.008) more bacteria on the surface of tubers from wet compost 78 days after planting. When P. infestans was co-cultured in Petri dishes with randomly selected tuber surface bacteria, some isolates (≤ 16.7%) inhibited the growth of the fungus. The percentage of the total bacterial population that was antagonistic to P. infestans was not significantly affected by soil moisture level (P= 0.368). The greater numbers of bacteria, of which a proportion are antagonistic to P. infestans, on the surface of tubers grown in wet compost may account for the greater resistance to tuber blight in that instance.     

Expression of the Phytophthora infestans ipiB and ipi0 genes in planta and in vitro

The ipiB and ipiO genes of the potato late blight fungus Phytophthora infestans (Mont.) de Bary were isolated from a genomic library in a screen for genes induced in planta. Expression of these genes was studied during pathogenesis on various host tissues and different host plants, some of which show specific resistance against P. infestans infection. During pathogenesis on leaves and tubers of the fully susceptible potato cultivar (cv.) Ajax and on leaves of the fully susceptible tomato cv. Moneymaker, the P. infestans ipiB and ipiO genes show a transient expression pattern with highest mRNA levels in the early stages of infection. During the interaction with leaves of the partially resistant potato cv. Pimpernel, the expression is also transient but accumulation and disappearance of the mRNAs is delayed. Also in P. infestans inoculated onto a race-specific resistant potato cultivar and onto the nonhost Solanum nigrum, ipiB and ipiO mRNA is detectable during the initial stages of infection. Apparently, the expression of the ipiB and the ipiO genes is activated in compatible, incompatible and nonhost interactions. In encysted zoospores, ipiB and ipiO mRNA accumulation was not detectable, but during cyst germination and appressorium formation on an artificial surface the genes are highly expressed. Expression studies in mycelium grown in vitro revealed that during nutrient starvation the expression of the ipiB and ipiO genes is induced. For ipiO gene expression, carbon deprivation appeared to be sufficient. The ipiO gene promoters contain a sequence motif that functions as a glucose repression element in yeast and this motif might be involved in the regulation of ipiO gene expression.     

Photosynthetic electron transport is differentially affected during early stages of cultivar/race-specific interactions between potato andPhytophthora infestans

The influence of the early stages of fungal infection on chloroplast metabolism was studied in cultivar/race-specific interactions between potato (Solanum tuberosum L. cv. Datura) and the late-blight fungusPhytophthora infestans. The accumulation of several mRNAs encoding components of the photosynthetic apparatus was not affected, either in compatible or in incompatible interactions. However, within 3 h after inoculation of potato leaves with fungal spores, a change in the photochemistry of photosystem II was detectable by measuring chlorophylla fluorescence. Characteristic fluorescence parameters, such as maximum fluorescence yield (F_m), variable fluorescence yield (F_v) and photochemical efficiency (F_v/F_m), were specifically reduced in the compatible host/pathogen interaction. Analyses of photochemical and nonphotochemical fluorescence quenching showed an increase in the photochemical fraction. The amounts of two selected thylakoid membrane proteins and of total chlorophyll remained unchanged during this process, suggesting that the functional modification of the electron-transport system was not correlated with a change in the composition of the photosynthetic apparatus. The alterations of photosynthetic electron transport represent a rapidly detectable and sensitive physiological marker for compatible interactions in the potato/Phytophthora infestans pathosystem.     

The effects of infection by Phytophthora infestans on the control of phenylpropanoid metabolism in wounded potato tissue

During the first 24 h of in vitro incubation of excised potato tuber (Solanum tuberosum L.) discs, the appearance of phenylalanine ammonia-lyase (PAL; EC 3.4.1.5) and the accumulation of chlorogenic acid are both stimulated by infection with Phytophthora infestans (Mont.) de Bary. Whereas in control tissue the level of PAL reached a stable plateau value after 40 h, in infected tissue it subsequently rose again, in one experiment, as the fungal mycelium developed. In the infected but not the control tissue, the level of chlorogenic acid subsequently fell to about to about 20% of its maximum after 50 h. The time courses of increases in cinnamic acid 4-hydroxylase (CA4H; EC 1.14.13.11; 0–60 h) and of caffeic acid acid o-methyltransferase (COMT; EC 2.1.1.42; 0–160 h) are not altered by fungal infection. If the discs are restored to the tuber environment immediately after excision, by placing them inside a host tuber, the activity of PAL as well as those of CA4H and COMT remained at the constant low endogenous level for at least 60 h, irrespective of whether the discs had first been inoculated with P. infestans. The increase in PAL may not be an obligatory feature of the P. infestans/potato compatible interaction but dependent on an underlying wound response. The experiments provide further evidence that PAL is the rate limiting step of chlorogenic acid biosynthesis in potato tuber discs.Abbreviations PAL phenylalanine ammonia-lyase - CA4H cinnamic acid 4-hydroxylase - COMT caffeic acid o-methyltransferase - CGA chlrogenic acid (5-o-caffeoylquinic acid) - gfwt gram fresh weight     

Effect of jasmonic acid on Ca<Superscript>+2</Superscript> transport through the plasmalemma of potato tuber cells

The rate of Ca²⁺ accumulation in plasmalemma vesicles isolated from quiescent and sprouting potato (Solanum tuberosum L.) tubers and the effect of 10^?5–10^?10 M jasmonic acid on the accumulation of Ca⁺² in plasmalemma vesicles and its efflux were studied. It was found that potato tuber plasmalemma contains a Ca⁺²,Mg⁺²-ATPase whose activity decreases upon the transition from forced quiescence to growth. The direction of the effect of jasmonic acid on Ca⁺²,Mg⁺²-ATPase (stimulation or suppression) depends on the physiological state of tubers and the phytohormone concentration.     

Methods of measuring the prevalence of Phoma exigua on potatoes and in soil

Methods are described for assessing inoculum of Phoma exigua by directly wounding potato test tubers and by inoculating soil into Arran Banner test tubers or slices. Large samples of tubers were wounded by dropping onto perforated metal sheet or grading on farm riddles, but standardization was achieved by inflicting four uniform wounds with shaped brass teeth. Tubers were then incubated for 12 wk at 5 ^oC. Soil samples to be tested were inoculated into crush wounds on surface-sterilized test tubers or tuber slices which were incubated for 8 wk at 5 ^oC. Assessments were speeded by immersing wounded slices in a mixture of Agrimycin and isopropylphenyl carbamate (IPC) solutions before inoculating, and then incubating for 2–3 wk at 10 ^oC. Although the sensitivity of test tubers is limited by their latent infection, comparisons of the tests in 3–5 years showed that the test tuber methods were frequently more sensitive than direct wounding, especially when amounts of inoculum were small.