Formation Of Wall Appositions Research Articles

Molecular cytology ofPhytophthora–plant interactions

Phytophthora diseases cause widespread economic and environmental losses worldwide. Thousands of plant species are susceptible. Disease is typically initiated through the activity of motile, biflagellate zoospores. Plant penetration and colonisation are achieved through the secretion of a diverse range of cell wall-degrading enzymes and effector proteins. Effector proteins are especially important during biotrophic growth; they function to suppress host defence and regulate host metabolism to favour pathogen growth. Plants can detect the presence of Phytophthora cells and rapidly mount a basal defence response that often successfully inhibits disease development.A key aspect of basal defence is the formation of wall appositions that constitute a physical and chemical barrier to pathogen growth. Components of basal defence can be triggered by chemical and physical signals produced by invading Phytophthora cells. If basal defence fails to inhibit pathogen ingress, the plant cell under attack can undergo hypersensitive cell death. In Phytophthora-plant interactions, hypersensitive cell death can be triggered by elicitors or effectors. In the continuing arms race between pathogen and plant host, Phytophthora species have evolved a range of counter-defence mechanisms that include suppression of hypersensitive cell death, inhibition of plant degradative enzymes and protection against reactive oxygen species. This brief article provides an overview of the contribution of modern molecular cytology to our understanding of Phytophthora-plant interactions.

Biological control of take-all in wheat by endophytic Bacillus subtilis E1R-j and potential mode of action

The bacterial strain E1R-j, isolated as an endophyte from wheat roots, exhibited high antifungal activity to Gaeumannomyces graminis var. tritici ( Ggt). Strain E1R-j was identified as Bacillus subtilis based on morphological, physiological and biochemical methods as well as on 16S rDNA analysis. This strain inhibited mycelium growth in vitro of numerous plant pathogenic fungi, especially of Ggt, Coniothyrium diplodiella, Phomopsis sp . and Sclerotinia sclerotiorum. In greenhouse experiments, soil drenches with cell densities of 10 6, 10 9 and 10 12 CFU ml −1 E1R-j reduced significantly take-all disease, caused by Ggt, in wheat seedling by 62.6%, 68.6% and 70.7%, respectively, compared to the inoculated control, 4 weeks after sowing. Growth parameters such as lengths and fresh weights of roots and shoots of Ggt-inoculated control plants were significantly lower compared to Ggt-inoculated and E1R-j treated plants. Field experiments in the season 2006/2007, heights of wheat plants in the Ggt inoculated plots were significantly reduced compared to the non inoculated treatments. Yield parameters such as kernels per head and thousand kernel weight (TKW) in inoculated control plants were lower compared to the other treatments. In the experimental year 2007/2008, independent treatments with the bacterial strain E1R-j and the fungicide Triadimefon reduced take-all disease in wheat roots by 55.3% and 61.9%, compared to the inoculated control plants. In this season plant height in inoculated control was significantly lower and also the yield parameters seeds per head and especially TKW were drastically reduced compared to the other treatments. E1R-j treatment alleviated the detrimental effects of take-all on grain yield parameters to a similar extent as Triadimefon application. SEM studies revealed that in the presence of E1R-j, hyphae of Ggt showed leakage, appeared ruptured, swollen and shriveled. Following root drench, strain E1R-j was able to colonize endophytically roots and leaves of wheat seedlings. While the population of the bacterial strain in wheat roots steadily increased from the second to the fourth leaf stage, in the leaf tissue the population of the strain rapidly declined. TEM studies also showed that cells of E1R-j were present in roots of wheat seedlings and effectively retarded infection and colonization of Ggt in root tissue; suppression of Ggt by E1R-j was accompanied by disintegration of hyphal cytoplasm. In addition, in the presence of E1R-j cells in Ggt-infected root tissue morphological defense reactions were triggered such as formation of wall appositions and papillae. The results presented indicate that the endophytic strain E1R-j of B. subtilis meets demands required for biocontrol of take-all.

Loss of pectin is an early event during infection of cocoyam roots by Pythium myriotylum

Cocoyam (Xanthosoma sagittifolium) is an important tuber crop in most tropical zones of Africa and America. In Cameroon, its cultivation is hampered by a soil-borne fungus Pythium myriotylum which is responsible for root rot disease. The mechanism of root colonisation by the fungus has yet to be elucidated. In this study, using microscopical and immunocytochemical methods, we provide a new evidence regarding the mode of action of the fungus and we describe the reaction of the plant to the early stages of fungal invasion. We show that the fungal attack begins with the colonisation of the peripheral and epidermal cells of the root apex. These cells are rapidly lost upon infection, while cortical and stele cells are not. Labelling with the cationic gold, which binds to negatively charged wall polymers such as pectins, is absent in cortical cells and in the interfacial zone of the infected roots while it is abundant in the cell walls of stele cells. A similar pattern of labelling is also found when using the anti-pectin monoclonal antibody JIM5, but not with anti-xyloglucan antibodies. This suggests that early during infection, the fungus causes a significant loss of pectin probably via degradation by hydrolytic enzymes that diffuse and act away from the site of attack. Additional support for pectin loss is the demonstration, via sugar analysis, that a significant decrease in galacturonic acid content occurred in infected root cell walls. In addition, we demonstrate that one of the early reactions of X. sagittifolium to the fungal invasion is the formation of wall appositions that are rich in callose and cellulose.

Ultrastructural and Cytochemical Investigation of Asymptomatic Infection by Pythium spp.

ABSTRACT The influence exerted by Pythium group F (a minor pathogen ubiquitous in soilless cultures) and P. uncinulatum (a nonpathogenic species) colonization on tomato roots was investigated. In both interactions, infected roots did not exhibit obvious symptoms; however, major physiological changes occurred within the host tissues colonized by Pythium group F compared to P. uncinulatum. According to our cytological observations, Pythium group F colonization involved a series of events: first, development and growth of the fungus in the epidermis and outer cortex tissues, which was associated with marked host cell disorganization and even breakdown. In colonized roots, symptoms were not easily discernible because alterations were restricted to the epidermis and outer cortex tissues. Second, pathogen ingress in the inner cortex and stele tissues was associated with massive induction of host defense reactions and alteration of invading hyphae. In a complex interaction that involved major metabolic changes in root cells, an array of defense-related reactions was produced, as exemplified by the formation of wall appositions and plugging of host cells with osmiophilic, electron-dense, granular, or fibrillar materials. P. uncinulatum growth was restricted to the epidermis and outer cortex tissues and associated with relatively minor damage to host cells. P. uncinulatum colonization of root tissues did not result in defense events similar to those observed in Pythium group F-infected inner cortex and stele tissues. Moreover, most of the invading P. uncinulatum hyphae were moribund. The results of the current study indicated that symptomless infections can be insignificant in terms of pathology for the plant (i.e., P. uncinulatum) or can be potentially dangerous (i.e., Pythium group F). Our results clearly showed a striking difference between nonpathogenic and minor pathogenic Pythium spp. attacks. The description of the Pythium group F infection process suggests that this interaction is unique, because the fungus behaves as a necrotrophic fungus in the epidermis and outer cortex tissues, whereas it is a potential inducer of plant defense reactions in the inner cortex and stele tissues. To our knowledge, cytological events similar to those observed with Pythium group F have not been described until now, even though descriptions of these events could help elucidate several aspects of the relationships between plants and minor pathogens.

Ultrastructure of compatible and incompatible interactions of soybean roots infected with the plant pathogenic oomycete Phytophthora sojae

Compatible and incompatible interactions of two soybean isolines containing either Rps1a or Rps1b resistance genes with races 2 and 8 of Phytophthora sojae were examined by light and electron microscopy. Phytophthora sojae race 2 is virulent on Rps1b plants and avirulent on Rps1a plants. Race 8 shows the reverse reaction; it is avirulent on Rps1b plants, but virulent on Rps1a plants. All combinations of races and cultivars were examined at times ranging from 30 min to 20 h postinoculation. Zoospore encystment, germination, and infection occurred within 30 min in all interactions. No evidence of appressorium formation was found. Wall appositions in epidermal cells adjacent to hyphae were very frequent by 30 min postinoculation. Differences between compatible and incompatible interactions became evident as early as 4 h postinoculation. The major difference appeared to relate to timing of host responses, which lead to two different types of relationships. In compatible interactions, P. sojae exhibited a short biotrophic phase with the establishment of many haustoria without triggering visible plant responses in cortical cells until approximately 10 h postinoculation. By 15 h postinoculation, almost the entire root was necrotic, wall appositions were abundant, and vascular tissue was colonized. The incompatible interaction was characterized by a nearly complete absence of haustoria, rapid host cell necrosis, and formation of many wall appositions by 4 h postinoculation. The pathogen rarely penetrated beyond the endodermis of the resistant host and colonization of vascular tissue was rare. Overall there were clear ultrastructural differences between compatible and incompatible interactions of soybean with P. sojae. These data support a strong correlation of resistance with host cell death, formation of wall appositions, and absence of root stele colonization. Key words: Phytophthora sojae, Glycine max, host–pathogen interaction, ultrastructure.

Ultrastructural and cytochemical aspects of the effect of chitosan on decay of bell pepper fruit

Effect of chitosan treatment on the infection process of bell pepper fruit by Botrytis cinerea was investigated at the ultrastructural level. In inoculated control tissues, fungal colonization proceeded rapidly, causing extensive degradation of host walls and middle lamellae. Such alterations were also observed over walls distant from invading hyphae. In chitosan-treated tissue, however, fungal cells were mainly restricted to wound cavities and ruptured epidermal cells. Host walls, even when appressed against invading hyphae, appeared well preserved and showed no sign of alteration. While in inoculated control tissues, invading hyphae appeared normal, the fungal cells detected in chitosan-treated tissue displayed various degrees of cellular disorganization from wall loosening to protoplasm degradation. Structural host defence responses such as formation of wall appositions and plugging of intercellular spaces with fibrillar material were frequently observed in chitosan-treated tissues. Such structure reacted intensely with an Aplysia gonad lectingold complex, indicating the presence of pectic residues.

Formation of wall appositions in leaves and lateral roots of an iron-accumulating pea mutant

E107 is a mutant of pea (Pisum sativum L. cv. Sparkle) characterized by necrotic spots on its leaves, high Fe accumulation in its shoot, and decreased nodulation. Fresh, cleared, and fixed tissues of the mutant and its parent were observed by light and electron microscopy. E107 has cell wall appositions in its basal leaflets and in the endodermis of its lateral roots. The wall deposits in the leaves are layered and contain callose and phenols. The appositional walls in the lateral root endodermis, made of lignin or of phenols, are not organised. We hypothesize that the appositions are stress responses to metal toxicity. Key words: pea mutant, wall appositions, histochemistry, iron toxicity.

Wall appositions induced by ionophore A 23187, CaCl2, LaCl3, and nifedipine in characean cells

The formation of wall appositions (plugs) by ionophore A 23187, CaCl2, LaCl3, and nifedipine was studied in mature internodal cells of characeaen algae. CaCl2 at concentrations above 10−2M induces thick fibrillar plugs without callose inNitella flexilis. InChara corallina andNitella flexilis ionophore A 23187 (1.25×10−5 to 5×10−5M) and LaCl3 (7.5×10−5 to 2.5×10−4M) cause flat appositions which contain callose and have a more granular structure. Plug formation by ionophore A 23187, CaCl2, and LaCl3 is pH-dependent and occurs beneath the alkaline regions of the cell. Nifedipine (10−4 to 10−5M) induces plugs inNitella flexilis after previous injury. These callose-containing wall appositions consist of a heterogeneous granular core which is covered by a fibrillar layer. The results of this work are compared with previous studies on wound wall formation and chlortetracycline (CTC)-induced plug formation which reveal that abundant coated vesicles occur only when a thick fibrillar wall layer is formed. Neither LaCl3 nor nifedipine inhibit the formation of CaCl2- or CTC-plugs. The unusual effects of these substances, which normally act as Ca2+ antagonists and therefore should prevent and not induce plug formation, are discussed. It is suggested that La3+ mimicks the effects of calcium and that nifedipine binding to the Ca2+ channels is altered in the alkaline regions of characean internodes and allows an influx of Ca2+.

An ultrastructural study of root invasion in three vascular wilt diseases

Transmission electron-microscopy was used to study root invasion in resistant and susceptible cultivars of tomato infected with Fusarium oxysporum f. sp. lycopersici or Verticillium albo-atrum and pea infected with F. oxysporum f. sp. pisi. There were many similarities in invasion by all pathogens of root epidermis and cortex in resistant and susceptible cultivars. Hyphae of tomato pathogens adhered to host root cell walls of susceptible cultivars but remained separated by extracellular electron-opaque material; hyphae of F. oxysporum f. sp. pisi on pea roots were embedded in a mucilaginous sheath. Both forms of adhesion were associated with a localized disruption of adjacent host wall structure. Initial penetration was by intercellular hyphae; subsequent intercellular colonization frequently resulted in the formation of wall appositions and papillae within epidermal and cortical cells. Penetration of host walls was achieved by constricted hyphae and localized wall degradation; continued deposition of material occurred around penetration hyphae with consequent formation of elongated penetration papillae. These were lignified, resistant to enzymic and chemical degradation and apparently effective in preventing further hyphal growth, although the frequency of their formation was similar in both resistant and susceptible cultivars. Papillae did not form in the moribund cells often seen even before penetration by F. oxysporum ff. sp.; in contrast V. albo-atrum was occasionally present within apparently healthy cells surrounded by invaginated host plasmalemma. A differential expression of varietal resistance occurred at the endodermis of both plant species. Accumulation of electron-opaque material in tomato endodermal cells and deposition of suberin and autofluorescent material in pea appeared to limit colonization of the stele. Pathogens circumvented this putative line of resistance either by intercellular growth between endodermal cells or by longitudinal growth from the root tip, which is devoid of mature endodermis. The diffuse and extensive intercellular wall structure of root tip cells is particularly amenable to initial colonization from which longitudinal growth allows entry into immature unlignified xylem elements and subsequently into mature vessels; hyphae were never seen to enter vessels from mature stelar cells.