Effect on germination and post-germination development of Colletotrichum dematium var circinans due to light and dark incubation and coverslip placement

Light regime was not a major factor in germination of spores of Colletotrichum dematium var circinans. Number of germ-tubes and appressoria formed were affected by incubation regime and coverslip placement. Most germ-tubes and appressoria were produced on spores under shimmed coverslips. Light incubation favored germ-tube production and dark incubation favored appressoria production. Predicted time to 100% germination was affected by treatments. Shortest times were predicted for light incubated spores under shimmed coverslips. Longest times were predicted for non-coverslipped spores. A significant number of appressoria were produced sessile on spores. Most sessile appressoria were produced on dark incubated spores. Approximately 56 % of all spores germinated, and approximately 60% of all germ-tubes and appressoria were produced on these spores which were situated closest to coverslip or droplet edges. Placement of germ-tubes on spores was not affected by treatments. However, percentages of germ-tubes, initiated from various areas of spores, were significantly different. This variation appears to be due to factors internal to spores.     

Germination and post-germination development of Colletotrichum dematium f. circinans on Allium cepa

p6e production of aflatoxins by Aspergillus flavus SRRC-1000 growing on soybean (Forrest) and glandless cottonseed (Deltapine 16) meals was examined with respect to effects of zinc and phytate. Aflatoxins were not produced on unautoclaved soybean meal. Addition of zinc (as zinc sulfate) to autoclaved meal inhibited aflatoxin production and supplementation with sodium phytate relieved this inhibition. Addition of sodium phytate alone promoted production. When cottonseed meal was treated to release native phytate into the meal from phytate-sequestering globoids, aflatoxin production increased. However, the largest production on cottonseed meal occurred upon dialysis of the meal without releasing phytate, implying removal of a small molecular weight inhibitor.     

Effect of prism generated discreet wavelengths of light on germination and post-germination development of Colletotrichum dematium var circinans (Berk.) v. Arx

Spores, harvested from 8 da old colonies ofColletotrichum dematium varcircinans (Berk.) v. Arx, were placed on slides under shimmed coverslips and subjected to 10 min irradiation with wavelengths of light ranging from 400 to 750 nm. Controls consisted of spores exposed to 10 min of fluorescent or tungsten light, or to continuous dark. Germination, and production of sessile and non-sessile appressoria, and germ-tubes was monitored. In addition position of germ-tubes on spores was noted and germination types were classified by numbers of germ-tubes and sessile and non-sessile appressoria produced. When compared to dark controls certain wavelengths, 400, 450, 600 and 650 nm, appeared to be inhibitory to germination and post-germination development, while others, 500, 550 and 750 nm, appeared to be stimulatory. Spores irradiated with some of the wavelengths (550, 700, 750 nm) that affected other responses produced the smallest percentage of sessile appressoria (approx. 10%). Placement of germ-tubes was not affected by treatment. More germ-tubes were produced from spore bottoms (53%) than tops (32%), and tips (15%). The fewest number of germ-tubes was produced from spore centers and from one tip. Thirteen germination types were noted. About 53% of spores germinated with one germ-tube, one non-sessile appressoria, and no sessile appressoria. The remaining 47% were distributed throughout the remaining 12 types, and ranged from 21% to less than 1% of the total. Treatments did not affect this distribution.     

Visualization of halos in the epidermal cell wall of Allium cepa caused by Colletotrichum dematium f. circinans and Botrytis allii using fluorochromes

Halos were detected with epifluorescence microscopy around penetration sites of Colletotrichum dematium f. circinans and Botrytis allii in onion epidermal cell walls as areas of less intense fluorescence or negatively stained areas in fluorescing cell walls following treatments with berberin sulphate and acridine orange but not with brilliant sulphaflavine (which stained the cell wall), ninhydrin, dansylchloride, or analine blue. Since pectin, pectic acid, avacil (microcrystaline cellulose super fine), filter paper, and Sephadex G-100–120 fluoresced with acridine orange and berberin sulphate, it was inferred that the halos were negatively stained or appeared as areas with less intense fluorescence because enzymes from these pathogens degraded cell wall pectin and cellulose at the point of penetration. Spores of both pathogens fluoresced when stained with brilliant sulphaflavine, acridine orange, ninhydrin, and dansylchloride. These stains and berberin sulphate caused germ tubes, appressoria, and primary infection mycelia to fluoresce. Nuclei in these fungal structures fluoresced when stained with acridine orange and brilliant sulphaflavine.     

Occurrence and characterization of Colletotrichum dematium (Fr.) grove

Colletotrichum dematium was isolated from caraway for the first time in Poland in 2005. Isolations of this fungus were repeated in 2006 and 2007. The cultures of fungus were obtained from superficially disinfected leaves, root necks, roots, stems and umbels. The isolates were identified on culture media: PDA and malt agar with addition of pieces of caraway stems and on the base of macro and microscopic structures. Studies on the biotic effect between C. dematium and other species of phyllosphere fungi of caraway showed that the majority of the studied species limited the growth and development of C. dematium, but the size of the limiting effect was different. The species from Trichoderma and Gliocladium genera were the most effective against C. dematium, causing degeneration and lysis of hyphae and precluded the formation of the pathogen's acervuli and conidia. C. dematium in dual culture with E. purpurascens, A. radicina, S. sclerotiorum, B. cinerea and R. solani produced an inhibition zone which indicated its capacity for antibiosis.     

Spore Yield and Microcycle Conidiation of Colletotrichum gloeosporioides in Liquid Culture

The effect of V8 juice concentration (5 to 40%, vol/vol), spore inoculum density (10⁵ and 10⁷ spores per ml), and liquid batch or fed-batch culture condition on mycelium and spore production by Colletotrichum gloeosporioides was evaluated. The amount of mycelium produced, the time required for initiation of sporulation following attainment of maximum mycelium, and the time for attainment of maximum spore concentration increased with increasing V8 juice concentration in batch culture. Cultures containing V8 juice at >10% achieved a similar spore density (apparent spore-carrying capacity) of about 0.8 mg of spores per ml (1 × 10⁷ to 2 × 10⁷ spores per ml) independent of inoculum density and V8 juice concentration. The relative spore yield decreased from a high of 64% of the total biomass for the low-inoculum 5% V8 culture, through 13% for the analogous 40% V8 culture, to a low of 2% for the high-inoculum 27% V8 culture. Fed-batch cultures were used to establish conditions of high spore density and low substrate availability but high substrate flux. The rate of addition of V8 juice was adjusted to approximate the rate of substrate utilization by the (increasing) biomass. The final spore concentration was about four times higher (3.0 mg of spores per ml) than the apparent spore-carrying capacity in batch culture. This high spore yield was obtained at the expense of greatly reduced mycelium, resulting in a high relative spore yield (62% of the total biomass). Microcycle conidiation occurred in the fed-batch but not batch systems. These data indicate that substrate-limited, fed-batch culture can be used to increase the amount and efficiency of spore production by C. gloeosporioides by maintaining microcycle conidiation conditions favoring allocation of nutrients to spore rather than mycelium production.     

Foliar disease of Clitorea ternatea due to Colletotrichum dematium and its effect on secondary metabolite production

Clitorea ternatea L. finds extensive application in therapeutic medicine. Forest regions of the Western Ghats, India were surveyed for foliar disease(s) caused in C. ternatea. The incidence and severity of foliar disease were determined during August 2006 to July 2009. The distribution of foliar disease incidence was determined by Taylor’s power law. The pathogen was tested for its seed-borne nature and seed transmission. Fungicides, like Bavistin, Antrocol, Hyzeb M-45 and Captra, were used for managing seed-borne pathogens in C. ternatea. Diseased foliages were tested for secondary metabolite content. Results indicated that the foliar disease in C. ternatea is caused by Colletotrichum dematium. The disease incidence and severity was moderately high. The disease is distributed homogeneously and heterogeneously in two forest regions. The pathogen is seedborne and transmitted from seed to plant. Among fungicides, Antrocol and Bavistin reduced seed mycoflora incidence. The disease caused increased production of alkaloid, flavonoid, phenol and sterol in foliages.     

Physiological and molecular aspects of aspartate-derived amino acid metabolism during germination and post-germination growth in two maize genotypes differing in germination efficiency

The Asp-derived amino acid pathway has been studied during the early stages of development in two maize genotypes, Io and F2, differing in germination efficiency and post-germination growth. In both genotypes expression of Ask2 (monofunctional Asp-kinase-2), Akh1 and Akh2 (bifunctional Asp-kinase-homo-Ser dehydrogenase-1 and 2), increased throughout germination and post-germination growth, suggesting a developmental regulation, whereas Ask1 (monofunctional Asp-kinase-1) was expressed constitutively. The major difference between Io and F2 concerned genes encoding bifunctional enzymes, particularly Akh2, the expression of which was dramatically low in F2. 15N-Asp labelling showed differences in in vivo Asp-kinase activities between the genotypes studied. Asp flux through the Met/Thr branches was higher in Io than in F2, while the latter exhibited a higher flux of Asp through the Lys branch. Physiological results, together with the higher Akh2 expression in Io, suggest that bifunctional enzyme activity, favourable to Met/Thr, was higher in Io than in F2 and that the monofunctional pathway was boosted in F2 because of the lower competition by the bifunctional pathway, thus allowing for higher flux of Asp through the Lys branch. In conclusion, it is suggested that F2 germination and post-germination growth might have been partially inhibited due to a limitation in Met and Thr availability. A negative physiological effect related to Lys accumulation in F2 is also discussed.     

Physiological and molecular aspects of aspartate-derived amino acid metabolism during germination and post-germination growth in two maize genotypes differing in germination efficiency

Fabiola Anzala, Marie-Christine Morère-Le Paven, Sylvie     

Effects of Calcium and Calmodulin on Spore Germination and Appressorium Development in Colletotrichum trifolii

Spore germination and appressorium formation are important steps in the process of fungal development and pathogenesis. These prepenetration events, which begin with spore attachment and culminate with appressorium maturation, a common scheme for many pathogenic fungi, are prerequisites for penetration of host external barriers and subsequent colonization. Conditions for in vitro spore germination and appressorium development in Colletotrichum trifolii are described. In addition, effects of Ca(sup2+) and calmodulin on these processes have been examined. Results indicate that, as for other pathogenic fungi, appressorium development is induced on a hard surface. The data suggest that disturbance of calcium homeostasis, by ethylene-bis(oxy-ethylenenitrolo)tetraacetic acid (EGTA) or calcium channel blockers, impairs appressorium development. Moreover, calmodulin inhibitors affect both germination and differentiation, implying that the Ca(sup2+)/calmodulin signal transduction pathway is important in the early development of C. trifolii on the plant host surface.     

Spore germination and mycelial growth of streptomycetes at different humidity levels

This study is the first to show the ability of streptomycetes to develop at a very low humidity level. All of the streptomycetes studied produced growth at low humidity (aw 0.86 and 0.67). This capacity was most markedly pronounced in Streptomyces odorifer, whose spores were capable of germinating, and mycelial germs increased in length, at the air humidity aw 0.50. The formation of lateral branches (mycelium branching) at this humidity was noted only in single S. odorifer germs and only after 72 h of incubation. Study of streptomycete growth on an agarized medium with different osmotic pressures, created by various glycerol concentrations in the medium, showed that, at aw 0.67, the spores of all the streptomycetes studied germinate, producing mycelial germs but not microcolonies. The ecological significance of mycelial prokaryotes in soil microbial communities that develop and function under conditions of extremely low humidity is discussed.     

Plastid development in seedlings of Echinochloa crus-galli var. oryzicola under anoxic germination conditions

Plastid development in the primary leaf of Echinochloa crus-galli (L.) Beauv. var. oryzicola (Vasing.) Ohwi was followed during 5 d of anoxic germination and growth. Plastids develop slowly from simple spheroidal proplastids into larger pleomorphic plastids with several stromal membranes and many peripheral membrane vesicles. A small prolamellar body is present at 96 h with perforated (pro)thylakoids extending into the stroma. Changes in starch grains and plastoglobuli are evidence of carbohydrate and lipid metabolism. Plastid division is indicated by dumbbell plastid profiles after 4 d of anoxia. These results demonstrate that plastids not only maintain their integrity during anaerobic germination but also show developmental changes involving an increase in internal membrane complexity, although to a lesser extent than in etiolated shoots.Abbreviation PLB prolamellar body Scientific paper No. 6167. College of Agriculture, Washington State University, Pullman     

Spore germination inCylindrospermum sp.: Influence of gases and growth conditions

Influence of different gases on spore germination ofCylindrospermum sp. revealed that acetylene and ethylene suppressed the germination which was more pronounced at their higher concentrations. Germination was completely inhibited under the atmosphere of hydrogen and argon even when supplied with 1% CO₂. Carbon dioxide, both in air and in N₂, stimulated the process. However, higher concentrations of CO₂ inhibited the germination in a concentration-dependent manner, possibly due to a decrease in pH. Germination was completely arrested in the absence of light and nutrients. The energetic requirement for germination was not efficiently fulfilled by photosystem I (PS II was blocked by 3-(3,4-dichlorophenyl)-1,1-dimethylurea) or under conditions of anoxygenic photosynthesis (medium supplemented with sodium sulfide).     

Deletion of a cyclic AMP receptor protein homologue diminishes germination and affects morphological development of Streptomyces coelicolor

Open reading frame SCO3571 of Streptomyces coelicolor encodes a protein of the cyclic AMP (cAMP) receptor protein (CRP) superfamily of regulatory proteins. A mutant revealed a dramatic defect in germination, followed by growth delay and earlier sporulation. This phenotype correlates with those of an adenylate cyclase (cya) mutant that cannot synthesize cAMP. This finding suggests that S. coelicolor may use a Cya-cAMP-CRP system to trigger complex physiological processes such as morphogenesis.     

Effects of fungicides on the growth and conidial germination of Colletotrichum coccodes and on the development of black dot disease of potatoes

The effects of 10 fungicides on the growth of Colletotrichum coccodes in agar culture and on the germination of conidia was investigated. In field experiments in 1990 and 1991 the extent to which treating black dot-affected potato seed tubers with fungicides affected the development of the disease on stem bases, roots and tubers was assessed. Black dot was also assessed on plants from field trials in 1990 which were designed to investigate the efficacy of the soil sterilant 1,3-dichloropropene (Telone) and two nematicides, aldicarb (Temik) and ethoprophos (Mocap). Prochloraz and fenpiclonil were the most effective fungicides in decreasing the size of C. coccodes colonies on agar. Imazalil, propiconazole and dichlorophen were also effective, but at higher concentrations, whereas tolclofos-methyl, thiabendazole and benomyl were only moderately effective. Resistant sectors developed from inhibited colonies on agar containing fenpiclonil and tolclofos-methyl. Conidial germination was prevented at 1 mg/litre fenpiclonil and 5 mg/ litre dichlorophen; imazalil, benomyl and thiabendazole were also moderately effective. Fenpiclonil and propiconazole seed tuber treatments consistently decreased black dot infection on roots, stem bases and daughter tubers early in the season, but only fenpiclonil decreased disease on tubers at harvest in October. Propiconazole also delayed emergence and decreased stem numbers. Soil treatment with 1,3-dichloropropene, aldicarb or ethoprophos had no effect on black dot but Rhizoctonia solani tuber infection and black scurf were increased.