Raymond I. Carruthers

Influence of thermal ecology on the mycosis of a rangeland grasshopper

The clearwinged grasshopper, Camnula pellucida, exploits incident solar radiation to raise internal body temperatures above ambient levels. Under adequate tem- perature and light conditions, grasshopper body temperatures may exceed air temperatures by 1 0- 1 50C with both nymphs and adults preferring to thermoregulate near 40C, their optimal temperature for development. Comparison of age-specific phenology data collected from field sites with projections from computer simulation studies on grasshopper devel- opment, indicated that incident solar radiation can highly accelerate maturation of grass- hoppers. Also, prolonged exposure to temperatures above 3 5C is beneficial to grasshoppers, as it is detrimental to the fungal pathogen Entomophaga grylli (US pathotype I), which is a major cause of natural mortality in many species. E. grylli protoplasts grew little or not at all if incubated at a constant 350C in tissue culture medium. Studies, conducted in vivo at constant temperatures, verified that E. grylli has an upper thermal limit of survival and development near 350C. In nature, however, these organisms are exposed to fluctuating temperatures. Further in vitro studies revealed that protoplast survival and development were significantly affected by incubation at 40C for as little as 2 h per day and almost totally inhibited by 400C for 8 h each day. In vivo incubation studies also showed an increase in the incubation period and a decline in the proportion of grasshoppers dying of E. grylli mycosis with increased daily exposure to 40C. Infected grasshoppers were also held at 25`-30C and allowed to range freely between shaded and lighted areas, so body temperatures could be raised through normal basking activity. Compared to a control group held in diffuse light, E. grylli mycosis was almost totally eliminated from the basking grasshoppers. The few individuals treated with solar radiation that died from E. grylli infection survived significantly longer then infected individuals not exposed to the solar simulator. Computer simulations using field-collected temperature, moisture, and solar radiation data revealed that behaviorally regulated thermal effects on E. grylli survival and development could limit disease development in some locations and seasons. Although it is unclear that the behavioral trait of basking is an adaptive response to selection pressures caused either by the pathogen or by other selective factors, it is clear that thermoregulatory behavior provides survival advantages to C. pellucida. In addition to their specific biological significance, these results demonstrate that behavioral exploitation of a physical resource in the environment can be used to profoundly alter biological relationships between species, in this case, allowing one organism to eliminate an antagonist by causing its mortality.

A fluorescence microscopy method for determining the viability of entomophthoralean fungal spores

Abstract Conidial viability of two species of entomophthoralean fungi was assessed using parallel vital fluorochrome staining and germination studies. Fluorescein diacetate (FDA) indicates viability when cells fluoresce yellow-green while propidium iodide (PI) indicates nonviability when cells fluoresce red. Various methods were used to induce Entomophaga maimaiga and Zoophthora radicans conidial mortality, including exposure to electromagnetic radiation, ethanol, or high temperatures. Following each treatment, half of the conidia were either stained with FDA and PI or held at ideal conditions for assessment of germination. In separate experiments, resting spores of Entomophaga grylli and E. maimaiga were treated either by autoclaving or with ethanol. Vital staining was used to assess the viability of the resting spores. FDA and PI were found to assess the viability of E. maimaiga and Z. radicans conidia accurately and precisely when compared to standard germination tests. No differences in comparative test results were found whether conidia were killed with ethanol, electromagnetic radiation, or high temperatures. When assessing resting spore viability, however, FDA and PI were not effective due to limited penetration of these stains through the thick walls of the spores and also due to the autofluorescence of E. maimaiga and E. grylli resting spores.

The effect of solar radiation on the survival of Entomophaga grylli (Entomophthorales: Entomophthoraceae) conidia

Abstract Laboratory studies on the effects of simulated solar radiation on Entomophaga grylli conidial viability showed definite interactions between the duration and intensity of exposure to the light source. When either variable was increased, the viability of E. grylli conidia was reduced. An exponential decay model was developed based on combining these two factors into a single predictive variable, exposure to cummulative solar radiation in Langleys. A minimum of approximately 3 Langleys was required before conidial viability decreased, with nearly 100% noted after exposure to 20 Langleys. The exponential decay model when applied to conidial viability data collected in the field provided an excellent fit; however, parameter estimates were significantly different from laboratory estimates as conidia were found to survive different periods of time in different habitats. The model predicted over 95% mortality in the most open habitats after exposure to ca. 50 Langleys. As expected, conidia exposed in lower canopy areas and in canopies with more dense foliage survived significantly longer (ca. 95% mortality after exposure to 700 Langleys).

Simulation of insect disease dynamics: an application of SERB to a rangeland ecosystem:

An object-oriented simulation program, SERB (Simulation En vironment for Research Biologists), has been used to simulate insect disease dynamics in a rangeland ecosystem. SERB was used to construct a mechanistic simulation model of a fungal pathogen of grasshoppers by synthesizing experimentally derived compo nents. Emphasis is placed on how SERB was used to construct and evaluate the model both in a piece-wise and combined manner to aid in understanding population interactions and dynamics. The interactive nature of SERB in the ZetaLisp environ ment was found to be highly conducive to model construction, manipulation and evaluation. Detailed examples from several phases of this process are presented from the perspective of a population bioligist. The orientation of SERB allows the research er to focus on a problem in an intuitive and systematic manner, in biological terms, unconstrained by details of an unfamiliar programming environment.

Simulation and object-oriented programming: the development of SERB:

An object-oriented programming language (OOPL), Flavors, was used to implement a modeling language and ancillary simula tion tools designed specifically to meet the needs of research biologists. A biological system is typically considered as a set of interacting entities, an abstraction which closely matches the pro gramming paradigm of Flavors. Some of the important charac teristics of Flavor programming are presented. Then SERB (the Simulation Environment for Research Biologists) is described. SERB uses Flavor objects to represent biological entities, resulting in a high degree of transparency between model and simuland. The simulationist creates model components from a library of component types and defines a model as some interacting set of these components. These models can then be used in simula tions or by higher level functions, such as sensitivity analysis. Flavors was found to be an ideal language for implementing modeling and simulation software for this domain.

Description of a Zoophthora radicans (Zygomycetes: Entomophthoraceae) epizootic in a population of Empoasca kraemeri (Homoptera: Cicadellidae) on beans in central Brazil

Development of a natural epizootic of Zoophthora radicans in an Empoasca kraemeri population on beans near Goiânia, Goias, Brazil, was monitored over a 6-week period. At the initiation of monitoring on 23 April 1985, disease prevalence in the leafhopper population was 12.8%. Over the course of the epizootic, infection of second-, third-, fourth-, and fifth-instar nymphs was similar and reached a peak of approximately 55% on 13 May, while infection of adult leafhoppers never exceeded 19%. Fungal inoculum expressed as the number of host cadavers with active (sporulating) fungus attached to the bean foliage was the variable most closely correlated to disease prevalence. Variables quantifying moisture or moisture combined with temperature (degree hours under moist conditions) were the most important abiotic predictors. A moisture variable incorporating a quantitative measure of moisture (level of wetness of a leaf wetness sensor) and a variable based only on the presence or absence of free moisture (dew) were equivalent predictors of disease prevalence. Epizootic development appeared to be inhibited when foliage was wet for less than 9 hr during the night. Infection trends in relation to fungus inoculum levels indicated an inoculum density threshold for epizootic development of approximately 0.8 leafhopper cadavers per plant.

Age Estimation of Mexican Fruit Fly (Diptera: Tephritidae) Based on Accumulation of Pterins

Abstract A common method of aging adult flies, fluorescence spectrometry, was used to monitor the increase of overall pterine titer in head extracts of Anastrepha ludens (Loew). Accumulation of fluorescent compounds was measured as a function of chronological age of flies maintained at 17 and 27°C. Although relative fluorescence increased with age, field studies revealed that this phenomenon could not be used for accurate age estimation, as relative fluorescence did not increase predictably with age over the entire life span. Accumulation of individual pterins, deoxysepiapterin and sepiapterin, were studied in a similar manner. These two specific compounds were separated by high-pressure liquid chromatography and their accumulation was followed at 15 and 30°C in the laboratory and under caged field conditions. While titer of deoxysepiapterin increased steadily in a curvilinear fashion, sepiapterin quickly reached a maximum and then maintained a constant level for the rest of the life of the flies. Based on the physiological response of deoxysepiapterin to chronological time and ambient thermal conditions, this compound was determined to be an age specific biological parameter for the Mexican fruit fly and should allow age estimation in field-collected flies.

Temperature-dependent germination and host penetration of the entomophthoralean fungus Zoophthora radicans on the leafhopper Empoasca kraemeri

Zoophthora radicans primary and secondary ovoid conidia inoculated onto Empoasca kraemeri fifth-instar nymphs germinated in three alternative modes, forming either germ-tubes, capilliconidiophores or replicative conidiophores. Production of germ-tubes was positively correlated, and production of capilliconidiophores inversely correlated, to temperatures over 5–32°C. Significant numbers of replicative conidiophores were produced only at 25–30°. The temperatures inducing the fastest rates of germination as capilliconidiophores, germ-tubes, and replicative conidiophores were 16·5, 22, and 26°, successively. Germ-tube formation was the most rapid mode of germination, occurring in ca 0·7 h at 22°; germination as capilliconidiophores or replicative conidiophores required 1·9 and 3·2 h, respectively, under optimal temperature conditions. Total germination (all modes combined) was optimum at 22°. High-temperature inhibition of germination was evident at 28° and severe at 30°; however, most germ-tubes produced at these temperatures were able to complete the processes of appressorium formation and penetration of the host cuticle. At 32°, only 6% of conidia germinated and no appressoria were produced. AT 10, 20, and 25°, 40, 20, and 4%, of all penetrations, successively, originated from capilliconidia produced by the ovoid conidia. The optimum temperature for infection (23°) was found to be similar to the optimum for vegetative germination (22°). The median time from inoculation to infection of the leafhoppers at 23° was 5.6 h. At temperatures near the upper threshold, lack of germination of the inoculum was the most important factor limiting host infection.