Secondary leaf fall of Hevea brasiliensis: meteorological and other factors affecting infection by Colletotrichum gloeosporioides

The lower leaf surface of Hevea brasiliensis was more susceptible to infection by Colletotrichum gloeosporioides than the upper. Few lesions were produced if spore drops on susceptible leaves were allowed to dry. Lesion development after 72 h was quickest at 21 ^oC, slower at 26.5 ^oC and was stopped at 32 ^oC, probably because of bacteria in the inoculation drop. On leaflets aged 7 days from bud-burst, the effective spore dose for 50% of leaflets infected (ED₅₀) after 16 h incubation, was 260 spores and after 46 h, 120 spores/infection droplet; the minimum ED₅₀ for the upper leaf surface was about 4 spores/mm². Leaflets 15 days old, which are normally resistant, were rendered susceptible by abrading the surface with carborundum powder. Spores caught in a Hirst spore trap reached a daily maximum at 23 h, at rates of up to 440 spores/m³ air/h, but fell to low concentrations as the humidity dropped during the daytime, and also during rain. There was some correlation between disease severity and duration of 97–100% relative humidity, and moderate to severe defoliation of clone PB 86 occurred when this reached 13.5 h/day. Rainfall increases infection by prolonging the period of atmospheric saturation and leaf wetness.     

CgMFS1, a Major Facilitator Superfamily Transporter,Is Required for Sugar Transport,Oxidative Stress Resistance,and Pathogenicity of Colletotrichum gloeosporioides from Hevea brasiliensis

Colletotrichum gloeosporioides is the main causal agent of anthracnose in various plant species. Determining the molecular mechanisms underlying the pathogenicity and fungicide resistance of C. gloeosporioides could help build new strategies for disease control. The major facilitator superfamily (MFS) has multiple roles in the transport of a diverse range of substrates. In the present study, an MFS protein CgMFS1 was characterized in C. gloeosporioides. This protein contains seven transmembrane domains, and its predicted 3D structure is highly similar to the reported hexose transporters. To investigate the biological functions of CgMFS1, the gene knock-out mutant ΔCgMFS1 was constructed. A colony growth assay showed that the mutant was remarkably decreased in vegetative growth in minimal medium supplemented with monosaccharides and oligosaccharides as the sole carbon sources, whereas it showed a similar growth rate and colony morphology as wild types when using soluble starch as the carbon source. A stress assay revealed that CgMFS1 is involved in oxidative stress but not in the fungicide resistance of C. gloeosporioides. Furthermore, its pathogenicity was significantly impaired in the mutant, although its appressorium formation was not affected. Our results demonstrate that CgMFS1 is required for sugar transport, resistance to oxidative stress, and the pathogenicity of Colletotrichum gloeosporioides from Hevea brasiliensis.     

Factors affecting the germination of spores of Bacillus anthracis

Spores of Bacillus anthracis germinated poorly at high cell densities unless the alanine racemase inhibitor O-carbamyl-D-serine was added to the germination medium. Spores derived from a variety of strains of B. anthracis germinated optimally at 22 degrees C. No correlation was found between rate of spore germination and virulence or between susceptibility of animal species to anthrax and spore germination rate using sera from those animals as the germination medium.     

Factors affecting the germination of spores of Bacillus anthracis

Spores of Bacillus anthracis germinated poorly at high cell densities unless the alanine racemase inhibitor O-carbamyl-D-serine was added to the germination medium. Spores derived from a variety of strains of B. anthracis germinated optimally at 22°C. No correlation was found between rate of spore germination and virulence or between susceptibility of animal species to anthrax and spore germination rate using sera from those animals as the germination medium.     

South American leaf blight of Hevea brasiliensis: spore dispersal of Microcyclus ulei

Trapping of ascospores and conidia of Microcyclus ulei among young trees of Hevea brasiliensis in Trinidad from May 1973 to May 1975 snowed that ascospores occurred throughout the year whilst conidia were present only during the wet season. Peak ascospore concentrations occurred in August and November during the wet season, the latter peak being more marked and the former coinciding with the period of maximum conidium liberation. In dry weather the number of ascospores increased during the night to a maximum at 06.00 h, and decreased to a low level during the day. On rainy days heavy ascospore discharge also occurred during the day. Ascospore concentration decreased significantly after dawn on sunny days whilst on overcast days the concentration remained high most of the day. Conidium production was highest around 10.00 h and decreased towards the evening to a low level during the night, reaching a minimum at 07.00 h.     

Identification and characterization of Neopestalotiopsis fungi associated with a novel leaf fall disease of rubber trees (Hevea brasiliensis) in Thailand

A novel leaf fall disease of rubber trees (Hevea brasiliensis) has been recently noted in Thailand. The fungal pathogens of this disease were identified based on both morphological and molecular characteristics as Neopestalotiopsis cubana and N. formicarum. Portions of internal transcribed spacer (ITS) region and the large subunit (LSU), translation elongation factor 1-α (TEF1-α) and β-tubulin (TUB) genes were PCR amplified with the primer pairs ITS1/ITS4, LR0R/LR5, EF1-728F/EF2 and T1/Bt2b, respectively. Sequencing of the PCR products and a phylogenetic tree based on the combined ITS, TEF1-α and TUB confirmed these pathogens as N. cubana and N. formicarum. Pathogenicity test results showed that the pathogens cause leaf spot and leaf fall similar to that observed in natural infections. This is the first report on the novel leaf fall disease of rubber trees in Thailand, with the results demonstrating that it is associated with N. cubana and N. formicarum.     

Regulation of pathogenic spore germination by CgRac1 in the fungal plant pathogen Colletotrichum gloeosporioides

Colletotrichum gloeosporioides is a facultative plant pathogen: it can live as a saprophyte on dead organic matter or as a pathogen on a host plant. Different patterns of conidial germination have been recognized under saprophytic and pathogenic conditions, which also determine later development. Here we describe the role of CgRac1 in regulating pathogenic germination. The hallmark of pathogenic germination is unilateral formation of a single germ tube following the first cell division. However, transgenic strains expressing a constitutively active CgRac1 (CA-CgRac1) displayed simultaneous formation of two germ tubes, with nuclei continuing to divide in both cells after the first cell division. CA-CgRac1 also caused various other abnormalities, including difficulties in establishing and maintaining cell polarity, reduced conidial and hyphal adhesion, and formation of immature appressoria. Consequently, CA-CgRac1 isolates were completely nonpathogenic. Localization studies with cyan fluorescent protein (CFP)-CgRac1 fusion protein showed that the CgRac1 protein is abundant in conidia and in hyphal tips. Although the CFP signal was equally distributed in both cells of a germinating conidium, reactive oxygen species accumulated only in the cell that produced a germ tube, indicating that CgRac1 was active only in the germinating cell. Collectively, our results show that CgRac1 is a major regulator of asymmetric development and that it is involved in the regulation of both morphogenesis and nuclear division. Modification of CgRac1 activity disrupts the morphogenetic program and prevents fungal infection.     

Role of dipicolinic acid in the germination, stability, and viability of spores of Bacillus subtilis

Spores of Bacillus subtilis spoVF strains that cannot synthesize dipicolinic acid (DPA) but take it up during sporulation were prepared in medium with various DPA concentrations, and the germination and viability of these spores as well as the DPA content in individual spores were measured. Levels of some other small molecules in DPA-less spores were also measured. These studies have allowed the following conclusions. (i) Spores with no DPA or low DPA levels that lack either the cortex-lytic enzyme (CLE) SleB or the receptors that respond to nutrient germinants could be isolated but were unstable and spontaneously initiated early steps in spore germination. (ii) Spores that lacked SleB and nutrient germinant receptors and also had low DPA levels were more stable. (iii) Spontaneous germination of spores with no DPA or low DPA levels was at least in part via activation of SleB. (iv) The other redundant CLE, CwlJ, was activated only by the release of high levels of DPA from spores. (v) Low levels of DPA were sufficient for the viability of spores that lacked most alpha/beta-type small, acid-soluble spore proteins. (vi) DPA levels accumulated in spores prepared in low-DPA-containing media varied greatly between individual spores, in contrast to the presence of more homogeneous DPA levels in individual spores made in media with high DPA concentrations. (vii) At least the great majority of spores of several spoVF strains that contained no DPA also lacked other major spore small molecules and had gone through some of the early reactions in spore germination.     

Effect of mechanical abrasion on the viability, disruption and germination of spores of Bacillus subtilis

AIMS: To elucidate the factors influencing the sensitivity of Bacillus subtilis spores in killing and disrupting by mechanical abrasion, and the mechanism of stimulation of spore germination by abrasion. METHODS AND RESULTS: Spores of B. subtilis strains were abraded by shaking with glass beads in liquid or the dry state, and spore killing, disruption and germination were determined. Dormant spores were more resistant to killing and disruption by abrasion than were growing cells or germinated spores. However, dormant spores of the wild-type strain with or without most coat proteins removed, spores of strains with mutations causing spore coat defects, spores lacking their large depot of dipicolinic acid (DPA) and spores with defects in the germination process exhibited essentially identical rates of killing and disruption by abrasion. When spores lacking all nutrient germinant receptors were enumerated by plating directly on nutrient medium, abrasion increased the plating efficiency of these spores before killing them. Spores lacking all nutrient receptors and either of the two redundant cortex-lytic enzymes behaved similarly in this regard, but the plating efficiency of spores lacking both cortex-lytic enzymes was not stimulated by abrasion. CONCLUSIONS: Dormant spores are more resistant to killing and disruption by abrasion than are growing cells or germinated spores, and neither the complete coats nor DPA are important in spore resistance to such treatments. Germination is not essential for spore killing by abrasion, although abrasion can trigger spore germination by activation of either of the spore's cortex-lytic enzymes. SIGNIFICANCE AND IMPACT OF THE STUDY: This work provides new insight into the mechanisms of the killing, disruption and germination of spores by abrasion and makes the surprising finding that at least much of the spore coat is not important in spore resistance to abrasion.     

Ultrastructure and Differentiation of Protein-storing Cells in Secondary phloem of Hevea brasiliensis Stem

The ultrastructure and differentiation of the protein-storingcells in secondary phloem of terminal branchlets of Hevea brasiliensisMull. Arg. were studied using electron microscopy. The cellsare parenchyma tissue in the axial system and characterizedby the presence of a large amount of proteinaceous fibrils inthe central vacuole. The fibrils of the protein-storing cellsare straight, about 9 nm in diameter and arranged in a directionroughly parallel to the longitudinal axis of the stem. At theearly differentiation stage of the protein-storing cells, amass of proteinaceous fibrils appears in the cytoplasm, thenis separated from the peripheral cytoplasm by the endomembranesystem derived from endoplasmic reticulum, resulting in theformation of the central vacuole with the fibrils inside asthe vacuolar content The peripheral cytoplasm may continue toproduce proteinaceous fibrils with which the fibrils of thecentral vacuole is supplemented after the protein-storing cellsare formed. Hevea brasiliensis, storage protein, proteinaceous fibrils, vacuole, secondary phloem     

Hevains: Serine-centred proteases from the latex of Hevea brasiliensis

Hevains b and l, isolated respectively from the serum and lutoids of freeze-dried latex from Hevea brasiliensis, were purified to homogeneity and compared with hevain a from commercial, ammonia-treated latex. The M_rs of hevains a and b are 69 000 and 58 000, respectively, and both exist in several charged forms. The amino acid compositions of the two enzymes differ significantly, but the reactivities to a variety of ester and protein substrates are similar, as are the pH optima. Hevain l is a distinct protease of M_r 80 000 and unique amino acid composition. It displays esterolytic activity and will digest insulin B chain, but is not proteolytic to azocollagen, azocasein, bovine serum albumen or haemoglobin. The activities of all three enzymes are dependent on the presence of serine and histidine residues.