The influence of a single-spore isolate and repeated subculturing on the pathogenicity of conidia of the entomophagous fungus Beauveria bassiana

The potential of monospore isolations for detecting strains of the entomophagous fungus Beauveria bassiana with increased virulence was investigated as well as the effect of repeated subculturing of both multispore parent strains and 28 newly obtained single-spore isolates on virulence. The use of the singlespore method enabled the disclosure of six spontaneous mutants surpassing the mother strain in pathogenicity, i.e., in a quicker effect and a higher virulence. The use of an optimal medium enabled an enhancement of virulence of all types of isolates. Differences among spontaneous mutants were indicated by the slope of the probit line. Our newly formed positive mutants retained their high pathogenicity both in repeated subculturing and in materials subcultured after a 12-month storage.     

Current understanding on Villosiclava virens,a unique flower‐infecting fungus causing rice false smut disease

Villosiclava virens (Vv) is an ascomycete fungal pathogen that causes false smut disease in rice. Recent reports have revealed some interesting aspects of the enigmatic pathogen to address the question of why it specifically infects rice flowers and converts a grain into a false smut ball. Comparative and functional genomics have suggested specific adaptation of Vv in the colonization of rice flowers. Anatomical studies have disclosed that Vv specifically infects rice stamen filaments before heading and intercepts seed formation. In addition, Vv can occupy the whole inner space of a spikelet embracing all floral organs and activate the rice grain‐filling network, presumably for nutrient acquisition to support the development of the false smut ball. This profile provides a general overview of the rice false smut pathogen, and summarizes advances in the Vv life cycle, genomics and genetics, and the molecular Vv–rice interaction. Current understandings of the Vv–rice pathosystem indicate that it is a unique and interesting system which can enrich the study of plant–pathogen interactions. Taxonomy: Ustilaginoidea virens is the anamorph form of the pathogen (Kingdom Fungi; Phylum Ascomycota; Class Ascomycetes; Subclass Incertae sedis; Order Incertae sedis; Family Incertae sedis; Genus Ustilaginoidea). The teleomorph form is Villosiclava virens (Kingdom Fungi; Phylum Ascomycota; Class Ascomycetes; Subclass Sordariomycetes; Order Hypocreales; Family Clavicipitaceae; Genus Villosiclava). Disease symptoms: The only visible symptom is the replacement of rice grains by ball‐shaped fungal mycelia, namely false smut balls. When maturing, the false smut ball is covered with powdery chlamydospores, and the colour changes to yellowish, yellowish orange, green, olive green and, finally, to greenish black. Sclerotia are often formed on the false smut balls in autumn. Identification and detection: Vv conidia are round to elliptical, measuring 3–5 μm in diameter. Chlamydospores are ornamented with prominent irregularly curved spines, which are 200–500 nm in length. The sclerotia are black, horseshoe‐shaped and irregular oblong or flat, ranging from 2 to 20 mm. Nested polymerase chain reaction (PCR) and quantitative PCR have been developed to specifically detect Vv presence in rice tissues and other biotic and abiotic samples in fields. Host range: Rice is the primary host for Vv. Natural infection by Vv has been found on several paddy field weeds, including Digitaria marginata, Panicum trypheron, Echinochloa crusgalli and Imperata cylindrica. However, the occurrence of infection in these potential alternative hosts is very rare. Life cycle: Vv infects rice spikelets at the late rice booting stage, and produces false smut balls covered with dark‐green chlamydospores. Occasionally, sclerotia form on the surface of false smut balls in late autumn when the temperature fluctuates greatly between day and night. Both chlamydospores and sclerotia may serve as primary infection sources. Rainfall at the rice booting stage is a major environmental factor resulting in epidemics of rice false smut disease. Disease control: The use of fungicides is the major approach for the control of Vv. Several fungicides, such as cuproxat SC, copper oxychloride, tebuconazole, propiconazole, difenoconazole and validamycin, are often applied. However, the employment of resistant rice cultivars and genes has been limited, because of the poor understanding of rice resistance to Vv. Useful websites: Villosiclava virens genome sequence: http://www.ncbi.nlm.nih.gov/Traces/wgs/?val=JHTR01#contigs     

Herbivory and intraspecific competition in a stream caddisfly population

Summary Field experiments were conducted to assess the effects of different population densities of the herbivorous caddisfly Helicopsyche borealis on periphyton biomass and on its own growth rate in Big Sulphur Creek, a third-order stream in northern California, USA. Stream enclosures were used to vary grazer density from one-eighth to twice natural density (1/8X–2X) in two experiments (35 d and 60 d), which spanned the period of most rapid larval growth. Periphyton biomass and chlorophyll a were inversely related to grazer density. Grazer densities of 1/8X–1/2X moderately reduced periphyton when compared to an ungrazed control, whereas densities of 1X–2X greatly depleted periphyton. The growth rate of H. borealis larvae declined with increasing larval density. Growth rate was highest at densities of 1/8X–1/2X; larvae grew more slowly at 1X and showed no growth at 2X. Weekly supplementation of periphyton to 1/4X, 1X, and 2X densities significantly increased the final body mass of larvae at 1X and 2X (by 26% and 50%, respectively, compared to unsupplemented larvae) but did not change the body mass of larvae at 1/4X. These results suggest that periphyton is a limiting resource in Big Sulphur Creek and that H. borealis larvae compete exploitatively for that resource. Intraspecific competition may be an important, but often overlooked, feature of many herbivore populations in streams.     

Climatic conditions, diapause and migration in a troglophile caddisfly

1. Factors explaining the end of diapause include environmental conditions such as daily photoperiod, temperature and humidity. However, because all these factors are almost constant inside deep caves, they would hardly affect diapause termination in many animal taxa that use such habitats in which to aestivate or hibernate (such as bats and some insects, snakes or frogs).
2. An innate biological signal, based on body reserves (mass), could determine the end of diapause in cave-dwelling animals. Another possibility is that the diapausing animals may use as a stimulus convective air circulation, produced by a fall in temperature outside the cave to a value below that inside (i.e. temperature inversions). Here, we explored these two non-exclusive hypotheses explaining the end of diapause and the start of migration in the caddis Mesophylax aspersus . This species aestivates as an adult in caves, as a physiological adaptation to seasonal drought and stream drying.
3. The variation in body mass of individuals entering and leaving the cave was similar, suggesting no role for the 'reserve level' hypothesis in breaking diapause. However, the onset of a temperature inversion in autumn coincided with the breaking of diapause followed after a few days by the migration of the caddisflies.
4. We conclude that the seasonal air currents, produced by thermal inversions, could be the migration stimulus for caddisflies and many other animals that spend a period of diapause in caves.     

Nematocide effect of entomophilic and entomophagous fungi

Study of nematocide effect of 48 species of entomophilic and entomophagous fungi from the class Fungi imperfecti against Panagrellus redivivus and Rhabditis oxycerca nematodes revealed a high frequency of fungi with nematocide effect. A close correlation between these two effects was disclosed by measuring the intensity of the nematocide and insecticide effect of fungal extracts.     

Sub-lethal effects of disturbance on a predatory net-spinning caddisfly

1. The sub-lethal effects of hydrologic disturbances on stream invertebrates are poorly understood, but integral to some models of how disturbances influence population and community dynamics. Carnivorous larvae of a net-spinning caddisfly, Plectrocnemia conspersa , have a strong predatory impact in some streams. Their silken nets, however, are vulnerable to high flow disturbance and the consequent destruction of nets could reduce predatory impacts and have life history consequences.
2. In a laboratory experiment, we manipulated the frequency of disturbances that destroyed the nets of P. conspersa , in the presence and absence of potential prey. Animals were housed individually and each trial lasted 8 days. We estimated net size, cumulative mass of silk produced, net allocation (net mass expressed as a proportion of body mass), per capita prey consumption and growth or mass loss of larvae.
3. In the absence of prey, increased disturbance frequency was accompanied by increased loss of body mass, a reduction of net size and an increase in the cumulative mass of silk produced. At the highest disturbance frequency, larvae eventually gave up producing nets. The ratio of net mass to body mass decreased with increasing disturbance, suggesting a trade-off in the allocation of resources, with a decreasing proportion of resources available for foraging. In the presence of prey, increased disturbance frequency was accompanied by a reduction in per capita prey consumption. Although foraging success offset the costs of silk production, growth rate decreased with increasing disturbance and could eventually lead to reduced body size and fecundity of adults.
4. These sub-lethal effects suggest that hydrologic disturbances could impose metabolic costs and reduce foraging efficiency of this predator. Thus, disturbances may reduce predator impact on prey populations and reduce predator population size without any direct mortality or loss of individuals.     

Ultrastructural studies on the fungus,Nomuraea rileyi,infecting the velvetbean caterpillar,Anticarsia gemmatalis

Combined scanning and transmission electron microscopy was used to study the fine structure of the developmental stages of Nomuraea rileyi infecting host larvae of Anticarsia gemmatalis. Larvae were dusted with large numbers of fungal conidia, which germinated and penetrated the cuticle within 6 hr post-treatment. Within 24 hr, penetration hyphae had reached the cuticular epidermis and, via a budding process, initiated the hyphal body stage in the hemocoel. The hyphal bodies, suspended in hemolymph, multiplied and spread throughout the host larvae. By 6–7 days post-treatment, the majority of larvae were mummified. Within 12 hr postmortem numerous conidiophores emerged producing a confluent mycelial mat over the entire cuticular surface. Numerous hydrophobic conidia were formed on phialides present on the aerial conidiophores.     

Significance of the fungus balansia cyperi infecting medicinal species of cyperus (Cyperaceae) from Amazonia

At least two species of Amazonian sedges (piripiri, i.e., Cyperus articulatus andC. prolixus, Cyperaceae) are widely utilized for various medicinal purposes, including birth control and induction of labor, and in hallucinogenic preparations. Examination of herbarium specimens collected from throughout South America and of living specimens collected in Ecuador has revealed that plants are infected by the systemic, ascomycetous fungusBalansia cyperi. This fungus is classified in the Clavicipitaceae and is known to infect only six species ofCyperus, including both North and South American species. Pure cultures of the fungus were obtained from two species of piripiri from Ecuador andC. virens from southern USA, and grown in submerged liquid culture. Extracts of the liquid medium revealed several unidentified ergot alkaloids. Current and past pharmacological uses of ergot alkaloids are consistent with uses of piripiri, suggesting a fungal basis for the ethnobotanical utilization of piripiri in Amazonia.     

Toxins of the entomophagous fungus Beauveria bassiana. II. Effect of nitrogen sources on formation of the toxic protease in submerged culture

A series of 24 nitrogen sources including inorganic, organic nonprotein, proteins and complex natural media were examined to determine their stimulatory effects on the production of a toxic proteolytic complex in Beauveria bassiana in submerged cultures. It was found that this effect is enhanced by the sources in the order presented. The best sources are maize meal, yeast extract, and beef extract. The production optimum on these sources occurs on the third day of fermentation. The composition of the protease complex may be influenced by the type of nitrogen source.     

An alternative route for infecting armadillos with Mycobacterium leprae

A nine-banded armadillo was inoculated with Mycobacterium leprae in both hind footpads. The animals were usually inoculated intravenously, or intradermally in the abdominal skin. Profuse multiplication of the bacilli occurred at the injection sites after more than two years. Eventually bacteraemia developed, and large numbers of the organisms were found in skin biopsies and in lymph nodes. There was limited dissemination of the bacteria into the spleen and the liver, and peripheral nerve invasion by the bacilli was also detected. M. leprae remained viable in the liver tissue, kept frozen at -80 degrees C for three years. This experimental system would be useful in testing the effects of certain immunological and chemotherapeutic agents against M. leprae by injecting them directly at the infection site.     

Phenotypic plasticity of larval retreat design in a net-spinning caddisfly

Larval Macrostemum carolina caddisflies construct silken catchnetswithin protective retreats, often on submerged trees and branches(i.e., snags). In the Savannah River, M. carolina larvae constructthree distinct retreats that differ in the configuration ofthe water entrance hole: (1) at the end of a silken tube, (2)flush with the top of the retreat, and (3) backed by a 180-degreesilken backstop. To identify the proximate mechanism mediatingthis retreat polymorphism, we removed larvae of known phenotypefrom their original retreats and brought them into the laboratory,allowing them to construct new retreats. We found that (1) larvaecan construct more than one type of retreat, so variation inthis behavior is not under strict genetic control; (2) larvaedo not preferentially reconstruct their original retreat design,so these alternative behaviors apparently exhibit little heritability;and (3) larvae primarily construct each phenotype in a particularmicrohabitat (i.e., "tube" and "backstop" retreats are principallyconstructed on the downstream half of the snag, and "flush"retreats on the upstream–bottom quadrant). Therefore,the retreat polymorphism in M. carolina is phenotypically plasticand is controlled by microhabitat location or a correlated environmentalvariable. Most net-spinning caddisflies construct their netsin fairly specific microhabitats. However, behavioral plasticityallows M. carolina larvae to construct retreats all around asnag, thereby reducing potentially intense competition for spacewith other net-spinning caddisflies. Consequently, this maybe the ultimate reason this polymorphism evolved.     

Coprophilin: An anticoccidial agent produced by a dung inhabiting fungus

Coprophilin, a decalin pentanedienoic acid methyl ester, was isolated from an unidentified fungus by bioassay guided separation. It inhibited (MIC = 1.5 μM) the growth of Eimeria tenella in an in vitro assay. The isolation, structure elucidation, absolute stereochemistry and biology are described.     

An epiphyllous fungus from the Lower Cretaceous

A fungus believed to be a member of the Microthyriales is described from the English Wealden. It occurs on the shoots of an undescribed conifer resembling Frenelopsis Schenk. From the structure of the thyrothecium and the presence of an epicuticular mycelium, the fungus is classified in the family Micropeltaceae, subfamily Stomiopeltoideae, and is described under the name Stomiopeltites cretacea gen. et sp. nov. It is the earliest fossil record of the Microthyriales.     

An epiphyllous fungus from the Lower Cretaceous

A fungus believed to be a member of the Microthyriales is described from the English Wealden. It occurs on the shoots of an undescribed conifer resembling Frenelopsis Schenk. From the structure of the thyrothecium and the presence of an epicuticular mycelium, the fungus is classified in the family Micropeltaceae, subfamily Stomiopeltoideae, and is described under the name Stomiopeltites cretacea gen. et sp. nov. It is the earliest fossil record of the Microthyriales.     

A capture-rate model of net-spinning caddisfly communities

Summary Empirical research suggests that net-spinning caddisflies require two basic resources, suspended particulate foods, and the currents which deliver them. I present a theoretical model of caddisfly communities based on quantitative differences in the capture rate produced by different catchnet designs. It assumes that catchnet architecture reflects a tradeoff between water filtration rate (flux through the net) and capture efficiency (the proportion of suspended items retained), and that the marginal resource concentration required by species with different catchnet morphologies should reflect the product of these parameters. The model hypothesizes a) that downstream changes in the physical morphology of the stream channel cause a shift in the relative importance of population limitations imposed by food and current-substrate availability, b) that the interaction of these physical changes with the filtering biota results in a seston resource gradient, and c) that the distribution of each taxon along this resource gradient reflects a marginal resource requirement determined by the functional morphology of its catchnet.     

Flow rate mediates the competitive influence of a grazing caddisfly on mayflies

The Glossosomatid caddisflies, which are stream-insect grazers, have been found to negatively affect the densities of mayfly grazers through intensive periphyton exploitation. The grazing efficiency of a Glossomatid caddisfly, however, is known to decrease with increasing current velocity. A manipulative field experiment was conducted to test the hypotheses that the presence of Glossosoma sp. decreased the densities of three mayfly grazers, Baetis thermicus, Cinygmula sp. and Paraleptophlebia chocorata, and that the decrease was stronger in slower stream-flow conditions. The experiment followed a two-factorial design with flow conditions and Glossosoma densities as the main factors. The periphyton biomass in conditions of reduced Glossosoma density was greater than in natural Glossosoma densities, but was not affected by flow conditions. In partial contradiction of the experimental hypothesis, the experimental reduction in Glossosoma density resulted in a higher density of Baetis, irrespective of flow conditions. The grazing activity of Glossosoma would not be affected by the flow variations that were employed in the present experiment. The reduction in Glossosoma density resulted in higher densities of Cinygmula and Paraleptophlebia only in fast-flow conditions, a result that did not support the experimental hypotheses. The competitive influence of Glossosoma on these mayflies might be altered by flow conditions causing differences in the resource structure.     

Survival of inoculum of the leaf blight fungus Phytophthora colocasiae infecting taro, Colocasia esculenta in the Solomon Islands

Phytophthora colocasiae was successfully isolated by baiting with detergent-treated taro leaf discs 8 cm diameter placed on water slurries of soil, on suspensions of macerated infected leaf lesions or on the washings from petioles of harvested plants. Taro root tips, detached or left on corms, were not susceptible to zoospores of P. colocasiae nor were detached root tips of Lupinus angustifolius. Cubes of taro corm used as baits, and agar selective for Phycomycetes which was inoculated directly with soil, both became too heavily overrun by Phythium splendens to allow detection of P. colocasiae. Investigations indicated that inoculum on lesions of detached leaves and in soil remains viable for only a few days. Petiole bases which comprise the bulk of the ‘tops’ used for vegetative propagation, lost detectable natural inoculum rapidly (2 days) if stored dry, but less rapidly (14 days) if planted immediately in the field. Artifically augmenting surface inoculum with naturally produced sporangia considerably extended the periods of detectability, probably by increasing the chances that a few propagules would survive, especially during dry storage. Incubation of inoculated tops in high humidity led to active infection and sporulation on petioles, especially on cut ends, a situation that might be paralleled under suitable moisture conditions in the field. Of several aroid species tested by artificial inoculation only Alocasia macrorrhiza was susceptible. Natural infection of this plant has not been seen, making it an unlikely alternate host of P. colocasiae under field conditions. Thus perennation between taro crops is effected by shortlived surface propagules and possibly also by mycelium within lesions on petioles. Reduction of the former and prevention of the latter might be achieved by dry storage of tops for 2 to 3 wk.