Unusual presentations of type 2 idiopathic macular telangiectasia.

PURPOSE: To report unusual presentations of type 2A idiopathic macular telangiectasia (IMT). METHODS: A retrospective analysis of disease presentation was conducted in 32 patients with type 2A IMT. All patients underwent a complete ophthalmological examination, including spectral domain optical coherence tomography (SD-OCT), fluorescein angiography (FA) and indocyanine green angiography (ICGA). RESULTS: Three out of 32 study patients showed the simultaneous presentation of type 2 IMT and other retinal diseases. In the first patient, the ophthalmological examination revealed a proliferative IMT associated with late-onset Stargardt disease. The second patient presented bilateral nonproliferative IMT and chronic serous chorioretinopathy in the left eye. The examination of the third patient revealed basal laminar drusen and soft drusen associated with IMT in both eyes. CONCLUSIONS: Type 2 IMT may represent an unusual presentation of Stargardt disease, chronic serous chorioretinopathy and basal laminar drusen. These presentations are most likely coincidental and highlight the importance of FA, ICGA and SD-OCT in the diagnosis and treatment of such cases.

Influence of Pythium oligandrum on the bacterial communities that colonize the nutrient solutions and the rhizosphere of tomato plants.

The influence exerted by the biocontrol oomycete Pythium oligandrum on the bacterial populations proliferating in the rhizosphere of tomato plants grown in a hydroponic system and in the circulating solutions is studied in the present experiment. Quantitative PCR and single-strand conformation polymorphism were used to investigate the genetic structure and dynamics of the bacterial communities colonizing the root systems and the various circulating solutions. Quantitative PCR assays showed that bacteria heavily colonized the rhizosphere of tomato plants with, however, no significant density changes throughout the cultural season (April-September). Single strand conformation polymorphism fingerprints revealed the occurrence of transient perturbations in the rhizospheric indigenous bacterial communities following P. oligandrum introduction in the root system of plants. This effect was, however, transient and did not persist until the end of the cropping season. Interestingly, the genetic structure of the bacterial microflora colonizing either the roots or the nutrient solutions evolved throughout the cropping season. This temporal evolution occurred whatever the presence and persistence of P. oligandrum in the rhizosphere. Evidence is also provided that bacterial microflora that colonize the root system are different from the ones colonizing the circulating solutions. The relationships between these 2 microflora (at the root and solution levels) are discussed.

Accumulation of an acidic dehydrin in the vicinity of the plasma membrane during cold acclimation of wheat

Expression of the acidic dehydrin gene wcor410 was found to be associated with the development of freezing tolerance in several Gramineae species. This gene is part of a family of three homologous members, wcor410, wcor410b, and wcor410c, that have been mapped to the long arms of the homologous group 6 chromosomes of hexaploid wheat. To gain insight into the function of this gene family, antibodies were raised against the WCOR410 protein and affinity purified to eliminate cross-reactivity with the WCS120 dehydrin-like protein of wheat. Protein gel blot analyses showed that the accumulation of WCOR410 proteins correlates well with the capacity of each cultivar to cold acclimate and develop freezing tolerance. Immunoelectron microscope analyses revealed that these proteins accumulate in the vicinity of the plasma membrane of cells in the sensitive vascular transition area where freeze-induced dehydration is likely to be more severe. Biochemical fractionation experiments indicated that WCOR410 is a peripheral protein and not an integral membrane protein. These results provide direct evidence that a subtype of the dehydrin family accumulates near the plasma membrane. The properties, abundance, and localization of these proteins suggest that they are involved in the cryoprotection of the plasma membrane against freezing or dehydration stress. We propose that WCOR410 plays a role in preventing the destabilization of the plasma membrane that occurs during dehydrative conditions.

Analysis of retinal flecks in fundus flavimaculatus using optical coherence tomography.

BACKGROUND/AIM: Retinal flecks are commonly observed in both Stargardt disease and fundus flavimaculatus (FFM). The aim was to determine the precise localisation of these flecks within the retinal layers using Stratus optical coherence tomography (OCT). METHODS: A prospective observational case series. A complete ophthalmological examination, including autofluorescence, fluorescein angiography (FA), and Stratus OCT (Carl Zeiss) was performed in 49 eyes of 26 consecutive patients with FFM. Six to 12 Stratus OCT linear scans focused on the retinal flecks were performed in each eye. RESULTS: The age at presentation ranged from 23 years to 71 years and visual acuity ranged from 20/20 to 20/400. Hyper-reflective deposits classified into two types were observed on Stratus OCT: type 1 lesions (94% of eyes) presented as dome-shaped deposits located in the inner part of the retinal pigment epithelium (RPE) layer and type 2 lesions (86% of eyes) presented as small linear deposits located at the level of the outer nuclear layer and clearly separated from the RPE layer. CONCLUSIONS: Stratus OCT is a non-invasive instrument that provides new information on the location of flecks in FFM. The location of type 2 lesions is quite unusual among macular dystrophies; OCT may therefore be useful in the diagnosis of retinal flecks in some cases of FFM.

Improvement of texture and structure of reduced-fat Cheddar cheese by exopolysaccharide-producing lactococci.

The objective of this study was to evaluate the effect of capsular and ropy exopolysaccharide (EPS)-producing strains of Lactococcus lactis ssp. cremoris on textural and microstructural attributes during ripening of 50%-reduced-fat Cheddar cheese. Cheeses were manufactured with added capsule- or ropy-forming strains individually or in combination. For comparison, reduced-fat cheese with or without lecithin added at 0.2% (wt/vol) to cheese milk and full-fat cheeses were made using EPS-nonproducing starter, and all cheeses were ripened at 7 degrees C for 6 mo. Exopolysaccharide-producing strains increased cheese moisture retention by 3.6 to 4.8% and cheese yield by 0.28 to 1.19 kg/100 kg compared with control cheese, whereas lecithin-containing cheese retained 1.4% higher moisture and had 0.37 kg/100 kg higher yield over the control cheese. Texture profile analyses for 0-d-old cheeses revealed that cheeses with EPS-producing strains had less firm, springy, and cohesive texture but were more brittle than control cheeses. However, these effects became less pronounced after 6 mo of ripening. Using transmission electron microscopy, fresh and aged cheeses with added EPS-producing strains showed a less compact protein matrix through which larger whey pockets were dispersed compared with control cheese. The numerical analysis of transmission electron microscopy images showed that the area in the cheese matrix occupied by protein was smaller in cheeses with added EPS-producing strains than in control cheese. On the other hand, lecithin had little impact on both cheese texture and microstructure; after 6 mo, cheese containing lecithin showed a texture profile very close to that of control reduced-fat cheese. The protein-occupied area in the cheese matrix did not appear to be significantly affected by lecithin addition. Exopolysaccharide-producing strains could contribute to the modification of cheese texture and microstructure and thus modify the functional properties of reduced-fat Cheddar cheese.

Benzothiadiazole-mediated induced resistance to fusarium oxysporum f. sp. radicis-lycopersici in tomato

Benzo-(1,2,3)-thiadiazole-7-carbothioic acid S-methyl ester (BTH), a synthetic chemical, was applied as a foliar spray to tomato (Lycopersicon esculentum) plants and evaluated for its potential to confer increased resistance against the soil-borne pathogen Fusarium oxysporum f. sp. radicis-lycopersici (FORL). In nontreated tomato plants all root tissues were massively colonized by FORL hyphae. Pathogen ingress toward the vascular stele was accompanied by severe host cell alterations, including cell wall breakdown. In BTH-treated plants striking differences in the rate and extent of fungal colonization were observed. Pathogen growth was restricted to the epidermis and the outer cortex, and fungal ingress was apparently halted by the formation of callose-enriched wall appositions at sites of fungal penetration. In addition, aggregated deposits, which frequently established close contact with the invading hyphae, accumulated in densely colonized epidermal cells and filled most intercellular spaces. Upon incubation of sections with gold-complexed laccase for localization of phenolic-like compounds, a slight deposition of gold particles was observed over both the host cell walls and the wall appositions. Labeling was also detected over the walls of fungal cells showing signs of obvious alteration ranging from cytoplasm disorganization to protoplasm retraction. We provide evidence that foliar applications of BTH sensitize susceptible tomato plants to react more rapidly and more efficiently to FORL attack through the formation of protective layers at sites of potential fungal entry.

Induction of Defense-Related Ultrastructural Modifications in Pea Root Tissues Inoculated with Endophytic Bacteria.

The stimulation exerted by the endophytic bacterium Bacillus pumilus strain SE34 in plant defense reactions was investigated at the ultrastructural level using an in vitro system in which root-inducing T-DNA pea (Pisum sativum L.) roots were infected with the pea root-rotting fungus Fusarium oxysporum f. sp. pisi. In nonbacterized roots, the pathogen multiplied abundantly through much of the tissue including the vascular stele, whereas in prebacterized roots, pathogen growth was restricted to the epidermis and the outer cortex In these prebacterized roots, typical host reactions included strengthening the epidermal and cortical cell walls and deposition of newly formed barriers beyond the infection sites. Wall appositions were found to contain large amounts of callose in addition to being infiltrated with phenolic compounds. The labeling pattern obtained with the gold-complexed laccase showed that phenolics were widely distributed in Fusarium-challenged, bacterized roots. Such compounds accumulated in the host cell walls and the intercellular spaces as well as at the surface or even inside of the invading hyphae of the pathogen. The wall-bound chitin component in Fusarium hyphae colonizing bacterized roots was preserved even when hyphae had undergone substantial degradation. These observations confirm that endophytic bacteria may function as potential inducers of plant disease resistance.

Histochemical and immunochemical study of the fate of Candida albicans inside human neutrophil phagolysosomes.

To further define the ultrastructural events associated with the killing of Candida albicans by human neutrophils, four methods were used: (1) the periodate-thiocarbohydrazide-silver proteinate (PA-TCH-SP) staining of vicinal-glycol-containing complex carbohydrates; (2) the localization of thermostable immunodeterminants of the yeast cell wall, mannans or mannoproteins, using monospecific antibodies and a protein A-gold complex (monAb-gold); (3) the localization of mannose residues with concanavalin A labeled with gold particles (Con A-gold); (4) the localization of chitin oligomers using wheat germ agglutinin and ovomucoid labeled with gold particles (WGA-gold). The mannan-rich cell wall layers were progressively lost as shown by altered PA-TCH-SP reactivity and a diffuse pattern of staining with Con A-gold and monAb-gold. The de novo appearance of conspicuous amounts of glycogen-like particles near the plasmalemma and in the cell wall was interpreted as evidence of a reparative process of the yeast cell wall. Chitin was seemingly unaltered and readily demonstrated by the WGA-gold in the wall remnants of ghost cells.

Ultrastructural localization of glucoside residues on tissue sections by applying the enzyme-gold approach.

The enzyme-gold approach was applied for ultrastructural localization of glucoside residues in animal and plant tissues. A beta-glucosidase-gold complex was prepared and used on thin tissue sections to reveal the corresponding substrate molecules by electron microscopy. Conditions for preparation of the complex, as well as for its application, were determined. Once applied on thin tissue sections, the glucosidase-gold complex yielded labeling over the rough endoplasmic reticulum, mainly on the ribosomal side of the membranes, and over the dense chromatin in the nucleus. Mitochondria, Golgi apparatus, and secretory granules in liver and pancreatic cells were free of gold particles. In plant cells, the labeling pattern was similar. In addition, the stroma regions of chloroplasts were densely labeled. In the extracellular space, labeling was found over the basal laminae of cells in animal tissues and over the fibrillar wall material bordering the intercellular space in plant tissues. Fungal cell cytoplasm was also labeled, as well as the membrane delineating mycoplasma-like organisms. Control conditions confirmed these labelings, demonstrating the possibility of revealing glucoside residues on tissue sections with high resolution and specificity.

Ultrastructural localization of glycoconjugates in the fungus Ascocalyx abietina, the Scleroderris canker agent of conifers, using lectin-gold complexes.

Different glycoconjugates were revealed in the fungus Ascocalyx abietina (Lagerberg.) Schlaepfer-Bernhard, by using various lectin-gold complexes. N-acetylglucosamine, N-acetylgalactosamine, and D-mannose were specifically localized in cell walls of fungal cells. N-acetylneuraminic acid (sialic acid) and L-fucose were detected in structures corresponding to lipid bodies, whereas they were totally absent in the cell wall. This is the first report on the occurrence of sialic acid in fungi and of fucose in Ascomycetes. The great advantage of using lectin-gold complexes for ultrastructural localization of sugars in phytopathogenic fungi, as well as in studies concerning host-pathogen interactions, is discussed.

A new lectin-gold complex for ultrastructural localization of galacturonic acids.

We report the development of a cytochemical affinity technique for detection of galacturonic acids at the ultrastructural level. The highly purified gonad lectin from Aplysia depilans (AGL) was tagged with colloidal gold particles and used for labeling carbohydrates in resin-embedded sections of various plant and fungal tissues. Patterns of AGL binding sites were compared to those obtained with a D-galactose-specific lectin, Ricinus communis agglutinin I. Differences in labeling patterns were noted, indicating that the lectins exhibited differential carbohydrate binding. In addition, the considerable loss of labeling over isolated wheat coleoptile walls treated for removal of pectin, after incubation with the AGL-gold complex, strongly suggested an affinity of AGL for pectic substances. A series of cytochemical controls, including sugar inhibition tests, has proven the specificity of the technique and the high affinity of AGL towards galacturonic acids. The potential value of this new lectin for ultrastructural studies on cell wall pectic substances in plant biology and pathology is demonstrated.

A novel ABCR nonsense mutation responsible for late-onset fundus flavimaculatus.

PURPOSE: To report the ophthalmologic features of a novel truncating mutation in the ABCR gene in a patient affected with late-onset fundus flavimaculatus (FFM). METHODS: A complete ophthalmologic examination was performed in a 70-year-old patient, including best-corrected visual acuity measurement, slit lamp and fundus examination, fundus photographs, frequent fluorescein and indocyanine green angiographies, visual field testing, color vision analysis, electroretinogram, and electro-oculogram. The 50 exons of the ABCR gene were analyzed using direct sequencing. RESULTS: Fluorescein and indocyanine green angiographies confirmed the diagnosis of FFM. A heterozygous base change was found, resulting in the substitution of an arginine to a stop at codon 152 of the ABCR gene. CONCLUSIONS: A heterozygous nonsense ABCR gene mutation was found in a patient affected with FFM. No other mutation has been identified in the entire coding sequence and the promoter region, suggesting that a heterozygous severe ABCR mutant may be responsible for a mild and delayed FFM phenotype, different from that of age-related macular degeneration.

Induction of defense responses in cucumber plants (Cucumis sativus L. ) By the biocontrol agent trichoderma harzianum

The potential of the biocontrol agent Trichoderma harzianum T-203 to trigger plant defense responses was investigated by inoculating roots of cucumber seedlings with Trichoderma in an aseptic, hydroponic system. Trichoderma-treated plants were more developed than nontreated plants throughout the experiment. Electron microscopy of ultrathin sections from Trichoderma-treated roots revealed penetration of Trichoderma into the roots, restricted mainly to the epidermis and outer cortex. Strengthening of the epidermal and cortical cell walls was observed, as was the deposition of newly formed barriers. These typical host reactions were found beyond the sites of potential fungal penetration. Wall appositions contained large amounts of callose and infiltrations of cellulose. The wall-bound chitin in Trichoderma hyphae was preserved, even when the hyphae had undergone substantial disorganization. Biochemical analyses revealed that inoculation with Trichoderma initiated increased peroxidase and chitinase activities within 48 and 72 h, respectively. These results were observed for both the roots and the leaves of treated seedlings, providing evidence that T. harzianum may induce systemic resistance mechanisms in cucumber plants.

Immunocytochemistry of plant defense mechanisms induced upon microbial attack.

During the past few years, cyto- and immunocytochemical techniques have been developed and widely used for locating and identifying various molecules in plant cell compartments. The last decade has witnessed tremendous improvements in molecular cytology, thus allowing an accurate in situ detection of various components thought to play important biological functions in the plant metabolism. The use of immunocytochemistry to investigate resistance mechanisms of plants upon pathogen attack has provided key information on the defense strategy that plants elaborate during a host-pathogen interaction. Of the various proteins induced in response to infection, chitinases and beta-1,3-glucanases have been the focus of particular attention due to their believed antimicrobial activity through the hydrolysis of the main fungal wall components, chitin and beta-1,3-glucans. Attention has also been paid to beta-fructosidase, the enzyme that hydrolyzes sucrose into glucose and fructoside. The marked accumulation of this enzyme upon pathogen infection has led to the consideration that infection may greatly influence the metabolic activity of colonized tissues by creating alterations of source-sink relationships. Another facet of the plant's defense strategy that has been the focus of considerable interest is related to the accumulation of structural compounds, such as hydroxyproline-rich glycoproteins and callose, to reinforce the wall architecture, thus decreasing vulnerability to microbial enzymes. A number of alternatives designed to improve plant protection towards pathogen invasion have been suggested. Among these, the production of transgenic plants expressing constitutively a foreign resistance gene and the pretreatment of plants with elicitors of defense reactions have been the subject of intensive studies at the molecular, biochemical, and cytological levels. Results of such studies clearly demonstrate the important contribution that cyto- and immunocytochemical approaches can make to our knowledge of how plants defend themselves and how plant disease resistance can be directly enhanced. These approaches will undoubtedly be active areas for future research in the development of biological control alternatives in which the mode of action of the product used is of key importance.

Cytological Analysis of Defense-Related Mechanisms Induced in Pea Root Tissues in Response to Colonization by Nonpathogenic Fusarium oxysporum Fo47.

The ability of nonpathogenic Fusarium oxysporum, strain Fo47, to trigger plant defense reactions was investigated using Ri T-DNA-transformed pea roots. Cytological investigations of strain Fo47-inoculated roots showed that the fungus grew actively at the root surface and colonized a number of epidermal and cortical cells, inducing marked host cell metabolic changes. In roots inoculated with pathogenic F. oxysporum f. sp. pisi, the pathogen multiplied abundantly through much of the tissues, whereas in Fo47-inoculated roots, fungal growth was restricted to the epidermis and the outer cortex. Invading cells of strain Fo47 suffered from serious alterations, a phenomenon that was not observed in control roots in which F. oxysporum f. sp. pisi grew so actively that the vascular stele was invaded within a few days. Strain Fo47 establishment in the root tissues resulted in a massive elaboration of hemispherical wall appositions and in the deposition of an electron-opaque material frequently encircling pathogen hyphae and accumulating in the noninfected xylem vessels. This suggests that the host roots were signaled to defend themselves through the rapid stimulation of a general cascade of nonspecific defense responses. The specific relationship established between strain Fo47 and the root tissues is discussed in relation to other types of plant-fungus interactions, including pathogenic and symbiotic associations.

Evidence for Antibiosis and Induced Host Defense Reactions in the Interaction Between Verticillium lecanii and Penicillium digitatum, the Causal Agent of Green Mold.

ABSTRACT Chronological events of the intercellular interaction between Verticillium lecanii and the postharvest pathogen Penicillium digitatum were investigated by transmission electron microscopy and gold cytochemistry. Growth inhibition of P. oligandrum as a response to V. lecanii attack correlated with striking host changes including retraction of the plasma membrane and cytoplasm disorganization. Such changes were associated with the deposition on the inner host cell surface of a chitin- and cellulose-enriched material which appeared to be laid down as a structural defense reaction. The accumulation of chitin in the newly formed material correlated with a decrease in the amount of wallbound chitin. However, the deposition of cellulose appeared to correspond to a de novo synthesis, as evidenced by the occurrence of cellulose-containing vesicles which released their content in the space between the invaginated plasma membrane and the host cell wall. Results of the present study provide the first ultrastructural and cytochemical evidence that antagonism, triggered by V. lecanii, is a multifaceted process in which antibiosis, with alteration of the host hyphae prior to contact with the antagonist, appears to be the key process in the antagonism against P. digitatum.

Ultrastructural and Cytochemical Investigations of the Antagonistic Effect of Verticillium lecanii on Cucumber Powdery Mildew.

ABSTRACT Chronological events of the intercellular interaction between Verticillium lecanii and cucumber powdery mildew, caused by Sphaerotheca fuliginea, were investigated at different times after inoculation by transmission electron microscopy. V. lecanii hyphae colonized host structures by tight binding, apparently mediated by a thin mucilaginous matrix. As early as 24 h after application of the antagonist, increased vacuolation and disorganization of the cytoplasm of the pathogen hyphae were easily detected. By 36 h after treatment, plasmalemma retraction and local cytoplasm aggregation were typical features of damage. Labeling chitin with the wheat germ agglutinin (WGA)/ovomucoid-gold complex showed that intracellular invasion of S. fuliginea by V. lecanii did not cause extensive host cell wall alterations, except in the area of hyphal penetration. By 48 h after inoculation, further cytoplasm disorganization was observed, as evidenced by the loss of cell turgor and contortion of the cell wall. Such deformation suggests that penetration of the antagonist results from mechanical pressure or localized enzymatic hydrolysis through the action of chitinases, as confirmed by the pattern of labeling obtained with the WGA/ovomucoid-gold complex. By 72 h after contact between the fungi, S. fuliginea cells were markedly collapsed, depleted of their protoplasm due to extensive multiplication of the antagonist, and totally encircled by the antagonist. Based on the current observations, the antagonism of S. fuliginea by V. lecanii appears to involve the following events: (i) attachment of the antagonist to the powdery mildew fungus; (ii) mechanical pressure and production of cell-wall degrading enzymes such as chitinases; (iii) penetration and active growth of the antagonist inside the pathogen hyphae; and (iv) digestion of host tissues and release of the antagonist from dead cells of S. fuliginea. The interaction between V. lecanii and S. fuliginea also affected the morphological and structural features of the haustorial bodies, as shown by increased vacuolation, distortion, and necrotization of the haustorial lobes. These observations provide the first experimental evidence that V. lecanii, primarily known as an entomopathogenic fungus, also has the potential to colonize mycelial structures of S. fuliginea. V. lecanii, therefore, may become a valuable alternative to current management of cucumber powdery mildew in greenhouses.

Cytochemical Investigation of the Antagonistic Interaction Between a Microsphaeropsis sp. (Isolate P130A) and Venturia inaequalis.

ABSTRACT In an attempt to better understand the mode of action of the antagonistic fungus Microsphaeropsis sp., the interaction between this fungus and Venturia inaequalis was studied, using both light and electron microscopy. Cytological observations indicated that the antagonistic interaction between the two fungi likely involves a sequence of events, including (i) attachment and local penetration of Microsphaeropsis sp. into V. inaequalis hyphae; (ii) induction of host structural response at sites of potential antagonist entry; (iii) alteration of host cytoplasm; and (iv) active multiplication of antagonistic cells in pathogen hyphae, leading to host cell breakdown and release of the antagonist. The interaction was investigated further by gold cytochemistry. The use of gold-complexed beta-1,4-exoglucanase and wheat germ agglutinin/ovomucoid-gold complex to localize cellulosic beta-1,4-glucans and chitin monomers, respectively, resulted in regular labeling of V. inaequalis cell walls. This finding supports other studies refuting the classification of ascomycetes as only a glucan-chitin group. At an advanced state of parasitism, the labeling pattern of cellulose and chitin, which clearly showed that the level of integrity of these compounds was affected, suggested the production of cellulolytic and chitinolytic enzymes by Microsphaeropsis sp. Wall appositions formed in V. inaequalis in response to the antagonist's attack contained both cellulose and chitin. However, penetration of this newly formed material frequently succeeded. This study provides the first detailed picture of the cytological events associated with mycoparasitism in V. inaequalis.

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.

Effect of Microsphaeropsis sp. Strain P130A on Germination and Production of Sclerotia of Rhizoctonia solani and Interaction Between the Antagonist and the Pathogen.

Microsphaeropsis sp. strain P130A was evaluated for the control of tuber-borne inoculum of Rhizoctonia solani based on the viability of sclerotia produced in vitro and on both the viability and production of tuber-borne sclerotia. The interactions between the antagonist and the pathogen, as well as the effect of the toxins produced by the antagonist on mycelial growth of R. solani were studied using transmission electron microscopy. On sclerotia produced in vitro, for all incubation periods (1 to 42 days), Microsphaeropsis sp. significantly reduced germination. Percent germination of sclerotia treated with Microsphaeropsis sp. decreased with increasing incubation period from an average of 82.0% after 1 day to stabilize at an average of 5.8% after 35 days. Similarly, percent germination of tuber-borne sclerotia was significantly lower when tubers were treated with Microsphaeropsis sp. Both 2% formaldehyde and Microsphaeropsis sp. treatments significantly reduced sclerotia germination to approximately 10% after 42 days of incubation at 4 degrees C. Furthermore, on tubers treated with the antagonist, the number of sclerotia per square centimeter decreased from 1.6 to 0.5 during the 8 months of storage at 4 degrees C, whereas an increase from 1.2 to 7.8 sclerotia per square centimeter was observed on untreated tubers. Microsphaeropsis sp. (strain P130A) colonized hyphae of R. solani within 4 days after contact on culture media. Transmission electron microscopic observations showed that the antagonist induced a rupture of the pathogen plasma membrane and that a chitin-enriched matrix was deposited at sites of potential antagonist penetration. Host penetration was not associated with pathogen cell wall alterations, which occurred at the time of progress of the antagonist in the pathogen cytoplasm. In the presence of a crude extract of Microsphaeropsis sp., cells of R. solani showed cytoplasm disorganization and breakdown of plasma membranes. Antibiosis and mycoparasitism were involved in the antagonism of R. solani by Microsphaeropsis sp., but the sequence by which these events occur, as well as the significance of wall appositions produced by R. solani, is yet to be established.