Phycomycetes parasitizing the ovaries of blackflies (simuliidae)

The developmental stages of the fungus in the ovary and freshwater are discussed. Based on the evidence to date the simuliid host is infected in the early larval instars. Parasitism results in a destruction of the ovarian tissues with no or few eggs produced. The fungus replaces the ovarian tissue and produces massive numbers of spores which are released by the female fly at the time of oviposition. After oviposition, infected flies do not seek a blood meal. Longevity of the host does not appear to be affected by the presence of the fungus. The spores released by the fly during oviposition did not withstand desiccation and did not develop on the artificial media tested; development occurs only in water. Development of sporangia takes 5 to 7 days at 4°C, and 2 to 3 days at temperatures above 10°C. The developmental process in spores from S. verecundum was followed from the formation of sporangia to the opening of discharge tubes which zoospores were presumed to be released but not observed.     

Functional Response of Chrysoperla carnea (Stephens) (Neuroptera: Chrysopidae) to Sugarcane Whitefly Aleurolobus barodensis (Maskell) in Laboratory Conditions

A functional response study of Chrysoperla carnea (Stephens) larvae to different densities of sugar cane whitefly Aleurolobus barodensis (Maskell) was conducted in test tubes at 26?±?2 °C, 65?±?5 % RH. Chrysoperla carnea showed two different types of functional response in larval instars. First instar exhibits type II. However, second and third larval instars revealed type III functional response. Based on modified Holling’s disk equation, the highest searching rates (a) of 0.82?±?0.0247 h^?1 was found for first instar larva. For second and third larval instars, the attack coefficient (b) were 0.002?±?0.030 and 0.0025?±?0.0424 respectively. The shortest handling time (T_h) per prey was observed at third instar stage (1.574?±?0.0568 h) followed by second and first instar with 1.72?±?0.0411 h and 1.919?±?0.0568 h respectively.     

Host selection and survival of the parasitoidEncarsia formosa on greenhouse whitefly,Trialeurodes vaporariorum, in the presence of hosts infected with the fungusAschersonia aleyrodis

Successful control of greenhouse whitefly may be achieved by complementary activity of the parasitoidEncarsia formosa and the fungusAschersonia aleyrodis. One way to obtain an additive mortality effect of both entomopathogen and parasitoid would be achieved by the selection of healthy hosts by the parasitoid and rejection of fungus-infected hosts. Third and fourth instar larvae ofTrialeurodes vaporariorum which had been treated with a spore suspension ofA. aleyrodis 0, 4, 7, 10 or 14 days beforehand, were presented to female parasitoids. The parasitoids adopted the oviposition posture on untreated hosts as well as on treated hosts, irrespective of the different stages of infection in the hosts. However, significantly more hosts were parasitized byE. formosa in the control treatment than in the fungal treatment. The parasitoids offered treated hosts, showed rejection behaviour after probing on hosts showing detectable signs of infection (containing hyphal bodies or mycelium in the haemolymph). For instance, when hosts were offered seven days after spore treatment, the parasitoids showed an oviposition posture on a total of 83 (95.4%) out of 87 infected larvae, but laid only 4 eggs (4.6%). In contrast, on 48 (94.1%) out of 51 noninfected (or showing no detectable signs of infection) hosts an oviposition posture was adopted and 40 eggs (78.4%) were found after dissection. When infected hosts were encountered the oviposition posture lasted less than 1′40″ while rejection of non-infected hosts occurred after more than 1′40″. Other experiments were carried out offering treated hosts for 24 h to the parasitoids. The hosts were dissected afterwards. Again, significantly more eggs were laid in the non-infected hosts. When hosts were parasitized shortly after fungal spore treatment they were colonized by the fungus and the parasitoids did not develop. Transmission of the entomopathogen after probing infected hosts was observed to a limited extent. In conclusion,A. aleyrodis andE. formosa can be used together in a glasshouse situation. The parasitoid will be most effective when introduced more than seven days after application ofA. aleyrodis, because from that time onwards it is able to detect and reject fungus-infected hosts.     

Laboratory and field evaluations of the susceptibility of immature Choristoneura rosaceana (Lepidoptera: Tortricidae) to Beauveria bassiana (Hypocreales: Cordycipitaceae)

Abstract

The susceptibility of immature Choristoneura rosaceana (Harris) to Beauveria bassiana-GHA (BotaniGard® 22WP) was evaluated under laboratory and field conditions. Although egg masses ≤24 h old were susceptible to infection by topically sprayed B. bassiana spores in the laboratory and resulting mycosis significantly reduced the percent of neonates able to emerge, no significant egg mass infections resulted from orchard treatment of ≤24–48 h eggs. Exposure to high levels of the B. bassiana product on apple leaf surfaces in the laboratory caused significant dose-related mortality in first to fourth instar C. rosaceana. First instar C. rosaceana were the most susceptible of the larval stages assayed in the laboratory although only 36% of neonates introduced into the orchard prior to Beauveria treatment and 27% of the neonates emerging from orchard treated egg masses became infected. Fourth instar mortalities were similar but moderate when treated with 1×10⁸ spores mL^?1 in the orchard and on leaf surfaces in the laboratory. Beauveria bassiana-induced larval mortalities were significantly higher when the spores were applied directly to the larval integument as opposed to the leaf surface where the insect would encounter the pathogen in a treated orchard. Feeding of C. rosaceana larvae on antibiotic containing meridic diet prior to their use in trials did not impact the susceptibility of the larvae to B. bassiana. Antibiotic containing meridic diet significantly reduced larval C. rosaceana mortality when treated larvae or the spores were placed directly on the diet as opposed to leaf tissue.     

Influence ofNosema bordati (Microsporida: Nosematidae) on the biological cycle ofChilo partellus (Lep: Pyralidae), a stem borer of cultivated graminae

The biological cycle ofChilo partellus (Swinhoe) was described on artificial diet. From egg to adult, it lasted 32 to 49 days with an average of 36.6 days. About 2,000 larvae from the 2^nd to the 5^th instars were artificially infected by ingestion with doses ofNosema bordati Goudegnon, varying from 2×10² to 2×10⁷ spores per ml. Only 72 survived (7.66 %) of these infected larvae.N. bordati, when present in the larvae, continued to multiply in the resulting pupae. The parasite affected the adults of this Pyralid reducing in a proportion of 5 the productivity of infected females and increasing the production of sterile eggs in the proportion of 8.     

Performance of Microplitis tuberculifer (Hymenoptera: Braconidae) parasitizing Mythimna separata (Lepidoptera: Noctuidae) in different larval instars

The solitary parasitoid Microplitis tuberculifer (Wesmael) is an important biological control agent of various lepidopteran pests in Asia. We examined the preference of M. tuberculifer for different instars of its common host, Mythimna separata (Walker), host instar effects on parasitoid development, and the consequences of parasitism in different stages for growth and consumption of host larvae. The wasp successfully parasitized the first four larval instars of M. separata, but not the fifth, which appeared to be behaviorally resistant. First and second instars were parasitized at higher rates compared to thirds and fourths in no-choice situations, ostensibly due to longer handling times for the latter, but second instars were most preferred in a choice test that presented all stages simultaneously. Although later instar hosts yielded heavier cocoons, the fastest parasitoid development was obtained in second instars. Lower sex ratios were obtained from first instars as females appeared to lay a smaller proportion of fertilized eggs in small hosts. Both weight gain and food consumption of parasitized larvae were reduced significantly within 24 h of parasitism, regardless of the stage parasitized, and final body weights were less than 10% those of unparasitized larvae. Thus, M. tuberculifer has good potential as a biological control agent of M. separata, successfully parasitizing the first four larval instars and dramatically reducing plant consumption by the host in all cases.     

Isolation of Beauveria species from Lebanon and evaluation of its efficacy against the cedar web-spinning sawfly, Cephalcia tannourinensis

Larvae of the cedar web-spinning sawfly, Cephalcia tannourinensis Chevin (Hymenoptera: Pamphiliidae), infected with a white fungus were collected from the Tannourine-Hadath El-Jebbeh cedar forest. Macro- and micro-morphological data based on the examination of colonies, conidiophores, and conidial shape of the fungus suggested a Beauveria species. Sequence analysis of the internal transcribed spacer regions of the isolated fungus showed that it is most closely related to isolates of B. bassiana Clade C. The present study showed that the isolated B. bassiana is a naturally occurring entomopathogenic fungus parasitizing the larvae of C. tannourinensis in Lebanon. Laboratory bioassays showed that B. bassiana caused high mortality of eggs and larvae. The infected eggs turned brownish in color, while larvae of the first instar ceased feeding and showed immobility and rigidity within 5 days and before sporulating conidial mat appeared on their cuticle. Second and third larval instars took longer time to show fungal sporulation: mortalities ranged between 85 and 100% within 7 days when treated with different conidial concentrations. The efficacy of control of C. tannourinensis using B. bassiana was higher or equal to the reference insect growth regulator, diflubenzuron, suggesting the possibility of its success as a biological control agent.     

Effects of environmental humidity on the survival and development of pine caterpillars, Dendrolimus tabulaeformis (Lepidoptera: Lasiocampidae)

The pine caterpillar, Dendrolimus tabulaeformis is one of the most important pests on Pinus tabulaeformis and other pine species in North China. In the present study, effects of relative humidity (RH) on the development and survival of pine caterpillars and soil moisture (SM) on their diapausing larvae were investigated. Low RH (20%) deferred the development of eggs and larvae, reduced egg hatching and larval surviving compared to 40%, 60% and 80% RH. Both low (20%) and high (100%) RH reduced egg hatching, but only 20% RH deferred the development of larvae, prolonged developmental duration and reduced the body mass and body length of larvae. The SM influenced the survival of diapausing larvae significantly. The dry treatment significantly reduced the supercooling points (SCPs), whereas increased the mortality and reduced body mass from 56.9 to 36.5 mg and body water content from 78% to 63% after 2 weeks' exposure. Therefore, higher RH is more favorable for the development of early instars and survival of diapausing larvae of the pine caterpillars.     

Quantitative studies on the pathogenicity of Nosema carpocapsae,a microsporidian pathogen of the codling moth,Cydia pomonella,in New Zealand

The pathogenicity of Nosema carpocapsae for codling moth was studied using dose-infectivity experiments. The IC₅₀ (median infective concentration) was similar for the five larval instars (range 4.0 × 10³ to 6.7 × 10⁴ spores/ml). Spore loads in moths ranged from 6.0 × 10⁶ to 7.1 × 10⁷ spores per moth and varied with dose and with larval age at infection. The infection does not cause mortality but does reduce the fecundity and fertility of infected moths. Nosema carpocapsae is transmitted transovarially as well as horizontally. Infected eggs were not produced by healthy females mated with infected males, although such pairs generally produced fewer eggs than healthy pairs.     

In vitro bioassay for the effect of Ajuga reptans phytoecdysteroids on Trialeurodes vaporariorum larval development

A rapid and reliable in vitro bioassay method to test the effects of synthetic and natural compounds on the development of greenhouse whitefly Trialeurodes vaporariorum Westwood (Homoptera Aleyrodidae) is described.Leaf fragments of Nicotiana glauca Graham infested with eggs of whitefly were cultured under aseptic conditions on half-strength Murashige-Skoog medium. The development of the insect in vitro and in vivo were comparable except for a higher mortality rate in the first larval instar.Ecdysteroids from Ajuga reptans L. were applied to leaf fragments by immersion in aqueous solution of the compounds or by addition to the culture medium. A high mortality rate of first larval instars was detected with 29-norsengosterone and ajugalactone treatments. This effect was specially pronounced when the products were incorporated in the medium. A significant increase in mortality was also recorded in the pupal state with a 20-hydroxyecdysone treatment. This effect was never observed before, because all whitefly larvae on A. reptans leaves die in the first larval instar.     

Life cycle of a new species of Amblyospora (Microspora: Amblyosporidae) in the mosquito Aedes taeniorhynchus

A new species of Microspora, Amblyospora polykarya, is described from the mosquito Aedes taeniorhynchus. The parasite is transovarially transmitted for one generation only. Spores in adult females extrude binucleate sporoplasms which infect developing eggs. Merogony occurs in larval oenocytes with diplokaryotic stages in early instars giving rise to plasmodia with many diplokarya. Plasmodia undergo cytokinesis to form diplokaryotic sporonts. In fat body cells, these sporonts secrete pansporoblastic membranes and undergo two nuclear divisions to form octonucleate sporonts. Cytokinesis and differentiation result in uninucleate spores in packets of eight. These spores are not transmissible per os and are of different morphotype from those in adult females. Infected larvae die in the fourth stadium.     

Protection of potato crop against Phthorimaea operculella (Zeller) infestation using frass extract of two noctuid insect pests under laboratory and storage simulation conditions

The oviposition deterrent effect of water extract of Spodoptera littoralis and Agrotis ipsilon larval frass on Phthorimaea operculella adult females was studied using two types of larval food “Natural host and Semi-artificial diet” under laboratory and storage simulation (semi-field) conditions. Extracted frass of fed larvae on semi-artificial diet showed complete oviposition deterrent effect at treatments with 4th, 5th and 6th instars of S. littoralis, also at treatments with 1st–3rd and 6th instars of A. ipsilon, while the same effect was observed when the larvae fed on castor oil leaves as a natural host only at treatment with frass extract of A. ipsilon 6th instar larvae. Presence of low amounts of phenols and flavonoids in water extract of A. ipsilon larval frass resulted in relatively more effect as oviposition deterrent to fertile adult females on treated oviposition sites, while the opposite effect was obtained in S. littoralis larval frass experiments. At semi-field experiments, the percentage reduction of laid eggs reached 100% after two?days at treatments with frass extracts of 4th and 5th S. littoralis larval instars and A. ipsilon 6th instar larvae fed on semi-artificial diet and/or castor oil leaves. Percentage reduction of laid eggs for untreated sacks reached 93.24 and 48.95% after 2 and 30?days, respectively, when placed between treated sacks, in comparison with the mean number of laid eggs for isolated control.     

Assessment of insecticidal activity of red pigment produced by the fungus <Emphasis Type="Italic">Beauveria bassiana</Emphasis>

Insecticidal activity of the red pigment produced by a strain of the fungus Beauveria bassiana that was locally isolated from infected whitefly, Bemisia tabaci (Genn.) was assessed. The pigment is produced extracellularly and is a water-soluble. This makes it easy and simple to be recovered from fermentation broth and used in pathogenicity experiments. When applied alone to Bm. tabaci nymphs, mortality percentages of 18% was recorded. For nymphs treated with Bv. bassiana spore suspension, mortality was 60%. The best results were obtained when red pigment was combined with fungal spores with the mortality percentage being increased to up to 92%. The highest insecticidal activity against adults emerging later on from the surviving larvae of Bm. tabaci was recorded also with treatment combining pigment and fungal spores with the longest days to pupation.     

Effect of leaf toughness and temperature on development in the lilac pyralid,Palpita nigropunctalis (Bremer) (Lepidoptera: Crambidae)

This study focuses on three factors that affect the survival of the lilac pyralid, Palpita nigropunctalis (Lepidoptera:Crambidae): (1) the effect of leaf toughness on survival rate to clarify the availability of leaves as food, (2) the effect of temperature on immature development to determine the lower thermal threshold, and (3) the effect of temperature on head capsule width to clarify whether head capsule width can be used to discriminate among field-collected larval instars. Larvae could develop on Osmanthus fragrans var. aurantiacus leaves collected in April, but not on leaves collected in June or September which were too tough to eat. More than 80% of the larvae on the leaves of Ligustrum lucidum, Ligustrum japonicum, Ligustrum obtusifolium and Syringa vulgaris completed development, regardless of the collection time. P. nigropunctalis completed development on L. lucidum at temperatures from 15 to 27.5 °C with a photoperiod of either 15 L:9D or 16 L:8D, but not at 30 °C, at which temperature no eggs hatched. The lower thermal threshold and thermal constant for total development from egg to adult were estimated at about 7 °C and 450–460 degree-days. Most of the larvae were 5-instar type larvae (passed through 5 instars) regardless of the temperature, but a few 6-instar type larvae (4 of 355) were noted at temperatures of 22.5 °C and higher. No overlap of the ranges of head capsule widths was detected for the 5-instar type larvae, indicating that head capsule width can be used to discriminate among field-collected larval instars.     

Survival of the parasitoidEncarsia formosa after treatment of parasitized greenhouse whitefly larvae with fungal spores ofAschersonia aleyrodis

The interaction between the entomopathogenic fungusAschersonia aleyrodis and the parasitoidEncarsia formosa on greenhouse whitefly as a host organism was studied, in particular, the survival of the parasitoid after treatment of parasitized hosts with fungal spores. The mean number of parasitized black pupae per parasitoid produced at 25°C was significantly reduced after spore treatment in the first three days following parasitization. Spore treatment four, seven or ten days after parasitization resulted in a mean number of parasitized pupae not significantly different from the number of black pupae in the control. The rather sudden change from low to high survival of parasitized hosts when treated with spores four days after parasitization in spite of high numbers of infected unparasitized larvae, coincided with the hatching of the parasitoid larva from the egg inside the host. Possible reasons for this decrease in susceptibility to infection after parasitoid egg hatch, such as induced changes in host cuticle or haemolymph, are discussed. Parasitoids emerged from treated hosts did not show differences in reproduction compared with parasitoids emerging from untreated hosts. Both natural enemeies of whitefly are compatible to a great extent.     

Evaluation of the pathogenicity of Aschersonia aleyrodis on Bemisia tabaci in the laboratory and greenhouse

The entomopathogenic fungus Aschersonia aleyrodis (Webber) is a promising fungal species against whiteflies. In this work, the pathogenicity of A. aleyrodis isolate Aa005 against MEAM1 Bemisia tabaci (Gennadius) was evaluated under laboratory and greenhouse conditions. The bioassay results indicated that the percentage of larval mortalities was concentration and age dependent. A. aleyrodis showed high pathogenicity against second and third instars and pupae with LC₅₀ values of 7.93?×?10⁶, 1.08?×?10⁷, and 1.56?×?10^7?conidia?mL^?1, respectively. The median lethal time (LT₅₀) was lower (4.60 days) for second instars and was the highest (6.17 days) for pupae when inoculated with a concentration of 1?×?10^7?conidia?mL^?1. Weekly sampling of immatures showed that the per cent mortality caused by A. aleyrodis at a conidial concentration of 1?×?10^7?conidia?mL^?1 was 71.21% in small nymphs, 69.31% in large nymphs and 53.36% in pupae. The dispersion index (DI) and Lloyd’s Index of Patchiness (LIP) values indicated that the infected immatures had a tendency to aggregate. The study demonstrated that A. aleyrodis isolate A005 is an effective biocontrol agent for B. tabaci control under laboratory and greenhouse conditions.     

Mass production,infectivity, and field application studies with the entomogenous fungus Paecilomyces farinosus

Hyphal bodies (HB) of the entomogenous fungus Paecilomyces farinosus mass produced in shaken liquid cultures are infective per os and topically to Spodoptera exigua larvae. Pathogenicity of HB was greatest at pH 5.5. It was reduced by exposure for more than one-half hour to temperatures above 30°C. P. farinosus infected, killed, and produced mycelia on the surface of first-instar S. exigua larvae at a temperature—relative humidity combination as low at 15°C and 70% RH. Using two types of microdroplet applicators (droplet in the size range 1–30 and 30–90 μm, respectively) under field conditions, HB were delivered to distances of 76 m from the site of spraying. The number of HB deposited on the leaf surfaces of treated crops caused mortality of 70 to 90% in bioassayed larvae.     

A laboratory evaluation of the pathogenicity of Bacillus popilliae var. rhopaea, the agent of milky disease in Rhopaea verreauxi (Coleoptera: Scarabaeidae)

Two methods of infection, i.e., feeding known numbers of spores and rearing larvae in contaminated peat, were used to bioassay the susceptibility of Rhopaea verreauxi to Bacillus popilliae var. rhopaea at 23°C. The susceptibility of the three larval instars was similar as measured by the ID₅₀ and IC₅₀ values. However, within an instar, newly molted larvae were less susceptible than mature larvae when infected by the contaminated peat method. It is suggested that this was due to reduced food intake. The range of ID₅₀ values for all bioassays with R. verreauxi larvae were 1.1 × 10⁷ to 4.0 × 10⁷ spores per larva, and IC₅₀ values were 3.4 × 10⁶ to 5.0 × 10⁷ spores per g of contaminated peat. The slope of the probit line was always low (0.6 to 1.8) except for young first-instar larvae infected by contaminated peat when the slope was 4.0. Disease per se did not affect food intake, though intake was reduced at high doses of contaminated peat. Young larvae often died without developing symptoms but, with increasing age, infected larvae were more likely to develop symptoms. Bioassays with Othnonius batesi and Rhopaea morbillosa indicated a much lower susceptibility per os than for R. verreauxi. It is concluded that the potential for using B. popilliae var. rhopaea to control R. verreauxi is high, but the bacillus is unlikely to be of value in control of O. batesi or R. morbillosa.     

Stored Bacillus popilliae spores and their infectivity against Popillia japonica larvae

Bacillus popilliae spores were stored for about 7 years under three separate conditions: frozen in sterile distilled water, smeared on glass microscope slides, and stored in loam soil at room temperature. In separate experiments, each of the 7-year-old preparations was fed to Popilla japonica larvae at concentrations of 10³, 10⁵, 10⁷, and 10⁹ spores/g of soil. A significant decrease in the percentage of larvae infected occurred in all of the aged spore tests. B. popilliae spores stored in soil, for the extended period, produced 3% larval infection only at the 10⁹ spores concentration; similar results were obtained from frozen spores. When P. japonica larvae were fed spores stored dried on slides, about 20% of the larvae developed milky disease. When aged frozen spores were artificially injected into larvae, 12% became infected at concentrations of 1 × 10⁶ spores/larvae; dried spores at the same concentration infected about 38% of the insect larvae. We conclude from these data that aged B. popilliae spores are significantly less infective against P. japonica larvae than young spores.     

Steinernema carpocapsae (Rhabditida: Steinernematidae) as a biological control agent against the fungus Gnat Bradysia agrestis (Diptera: Sciaridae) in propagation houses

A recently introduced fungus gnat, Bradysia agrestis, has caused serious problems in Korean propagation houses where vegetable seedlings are produced for transplant into the fields. Although chemical insecticides are available against this pest, alternate control measures are needed. A Korean isolate of the entomopathogenic nematode, Steinernema carpocapsae Pocheon strain, was tested against this insect in the laboratory and propagation house. In the laboratory, S. carpocapsae affected oviposition, with the untreated females laying an average of 121±25 eggs, whereas the treated females averaged 7±2 eggs. The infectivity of S. carpocapsae to the fungus gnat was affected by the developmental stage and temperature, with highest mortality observed with the third and fourth instars and pupal stage. Nematode mortality in the second instar fungus gnat ranged between 23 and 35%, but showed no significant differences among the temperatures tested. The egg and first instar were not infected by the nematode. In nematode dispersal studies, adult female fungus gnats alone dispersed S. carpocapsae from the nematode-treated area to the control area at a higher rate than male and female gnats or male gnats alone. In the propagation house experiments with watermelon seedlings, no significant difference was observed in fungus gnat larval reduction at S. carpocapsae concentrations of 5, 10, or 20 infective juveniles (IJs)/g of soil at 7, 14, and 21 days after treatment. In comparison with the control, the S. carpocapsae treatments significantly reduced B. agrestis larval numbers. When the watermelon seed was treated with S. carpocapsae at sowing, the larval density of B. agrestis was significantly reduced, that is, the number of B. agrestis larvae ranged from four to eight and from five to eight in the nematode-treated plots compared with 26 and 30 in the control plots on the 17th and 34th day post-treatment, respectively. In the chemical insecticide treatments, diflubenzuron and chlorpyrifos were significantly more effective than S. carpocapsae and diazinon in reducing larval populations of the fungus gnat. Our data show that, although some of the chemical insecticides were more effective than S. carpocapsae Pocheon strain, the nematode was still an effective tool for management of the fungus gnat larvae and in protecting seedlings from damage in propagation houses.