Infectivity of two members of theEntomophthora muscae complex [Zygomycetes: Entomophthorales] forMusca domestica [Dipt.: Muscidae]

Dose-mortality studies were conducted with 2 members of theEntomophthora muscae (Cohn) Fresenius complex from southern California (CA) and Denmark (DA) infecting house flies,Musca domestica L., from southern California. Primary conidia of the DA form were significantly more infective (LC₅₀=34 conidia/mm²) than primary conidia of the CA form (LC₅₀=67 conidia/mm²) for 2–7 days old (‘young’) flies. Infectivity (single experimental trial) for 10–14 day old (‘old’) flies of primary conidia of the CA form (LC₅₀=34 conidia/mm²) was similar to that of the DA form for young flies. Secondary conidia of the CA form were markedly more infective (LC₅₀=0.36 conidia/mm²) than primary conidia of either form. Dose had no significant effect on incubation period for primary conidia of either form, but increased doses resulted in significantly shorter incubation periods for flies exposed to secondary conidia of the CA form.      

Effect of the entomopathogenic fungus,Entomophthora muscae (Zygomycetes: Entomophthoraceae), on sex pheromone and other cuticular hydrocarbons of the house fly,Musca domestica

House fly (Musca domestica) males are highly attracted to dead female flies infected with the entomopathogenic fungus Entomophthora muscae. Because males orient to the larger abdomen of infected flies, both visual and chemical cues may be responsible for the heightened attraction to infected flies. Our behavioral assays demonstrated that the attraction is sex-specific-males were attracted more to infected females than to infected males, regardless of cadaver size. We examined the effect of E. muscae on the main component of the house fly sex pheromone, (Z)-9-tricosene, and other cuticular hydrocarbons including n-tricosane, n-pentacosane, (Z)-9-heptacosene, and total hydrocarbons of young (7 days old) and old (18 days old) virgin females. Young E. muscae-infected female flies accumulated significantly less sex pheromone and other hydrocarbons on their cuticular surface than uninfected females, whereas the cuticular hydrocarbons of older flies were unaffected by fungus infection. These results suggest that chemical cues other than (Z)-9-tricosene, visual cues other than abdomen size, or a combination of both sets of cues might be responsible for attraction of house fly males to E. muscae-infected females.     

Seasonal Prevalence ofEntomophthora muscaeand Introduction ofEntomophthora schizophorae(Zygomycotina: Entomophthorales) inMusca domestica(Diptera: Muscidae) Populations on California Dairies

Two entomopathogenic fungi in theEntomophthora muscaespecies complex that infect house flies were used in this study:E. muscaeFresenius (16–17 nuclei/conidium) occurred naturally at four southern California dairies, whileEntomophthora schizophoraeKeller and Wilding (4–8 nuclei/conidium) did not. During the first year of the study, onset of measurableE. muscaeinfections occurred between September and November but varied among sites. At least 20% of the flies at all four dairies were infected by November, and infection at one site exceeded 70%. During the fall epizootic period, infection levels were inversely related to temperature. Average weekly temperatures higher than 17–20°C and maximum daily temperatures higher than 26–28°C were statistically correlated with low infection levels. In the second year,E. schizophoraewas introduced by releasing diseased flies at two dairies (four times at one dairy and three times at the other).E. schizophoraewas recovered for a brief time in the house fly population after the first two releases at one site but not at the second site.     

Entomophthora ferdinandii (Zygomycetes: Entomophthorales) Causing Natural Infections of Musca domestica (Diptera: Muscidae) in Argentina

The identity and activity of an entomopathogenic fungus belonging to the Entomophthora muscae species complex and infecting Musca domestica in poultry houses from La Plata, Argentina, is reported. Entomophthora caused natural infections between September 2001 and September 2003. Primary conidia of this fungus were on average 29.5 ± 1.2 × 23.4 ± 2.4 μm and contained, on average, 10.5 ± 0.1 nuclei (range: 7–15) with an average diameter of 4.8 ± 0.1 μm. This fungus is identified as E. ferdinandii Keller (this specific epithet includes a nomenclaturally required spelling correction); this is a first record of E. ferdinandii in South America and of any member of the E. muscae species complex from flies in Argentina.     

Morphology ofEntomophthora muscae protoplasts grownin vitro

Summary Entomophthora muscae (C.) Fres. can be grownin vitro as protoplasts. Light and electron microscopical studies of thein vitro developed protoplasts have demonstrated the absence of an organized wall over the protoplasmic Con A-positive membrane at all stages of growth. The cytological organization is typical of the Entomophthorales with condensed chromatin in the interphase nuclei and small eccentric metaphase spindles. Long strands of endoplasmic reticulum, microubules and vesicles surrounding the plasmalemma may be involved in maintaining the precise shape ofE. muscae protoplast. Starvation of the fungus induces the formation of hyphal bodies after deposition of Con A- and WGA-positive wall material at the plasmalemma surface.Abbreviations Con A concanavalin A - DH Drosophila cell culture medium - FITC fluorescein isothiocyanate - GLEN glucose-lactal-bumin-yeast extract-NaCl culture medium for protoplasts - HBL hyphal body-like protoplasts - MM Mitsuhashi and Maramorosch' insect cell culture medium - PATAg periodic acid-thiocarbohydrazide-silver proteinate technique - PBN phosphate buffer with NaCl - S spherical protoplasts - WGA wheat germ agglutinin     

Field Studies of Entomophthora (Zygomycetes: Entomophthorales)—Induced Behavioral Fever in Musca domestica (Diptera: Muscidae) in Denmark

House flies were collected over 3 days (three to five times per day) from specific sites on a dairy farm with a range of high to low temperatures. Flies were held individually to determine whether the distribution of fungus-infected (Entomophthora muscae and E. schizophorae) house flies differed according to the stage of infection and temperature. All but 2 of 396 infected flies (99.5%) had E. muscae. More E. muscae-infected flies collected from cool areas were in later stages of infection (i.e., dying 0–2 days after capture), whereas flies collected on sun-exposed surfaces tended to be in earlier stages of infection (i.e., dying 6–8 days after capture). Most flies died 3–5 days after capture and were consequently in the middle stages of infection. A mark and release experiment was conducted to determine whether E. schizophorae-inoculated flies frequented surfaces with higher temperatures than did uninfected control flies. About 3000 yellow-marked house flies inoculated with E. schizophorae and 3000 blue-marked control flies were released in an enclosed swine farrowing barn. Significantly more inoculated flies were recorded on the heat lamps than flies in the control group. The results suggest that behavioral fever occurs in the field for flies infected with both E. muscae and E. schizophorae and that flies can cure themselves of infection through the use of artificial heat sources.     

Entomophthora brevinucleata sp. nov. [Zygomycetes,Entomophthoraceae], a pathogen of gall midges [Dip.: Cecidomyiidae]

Entomophthora brevinucleata, a fungal pathogen of gall midges, is described. The species occurs on at least 3 host species, including the wheat blossom midgeSitodiplosis mosellana Géhin andMycodiplosis sp.E. brevinucleata produces rhizoids, either monohyphal or comprising bundles of a few hyphae, without holdfasts. The hyphal bodies are subspherical with mean dimensions of 27.5×22.1 μm. The primary conidia are campanulate with mean dimensions from individual hosts of 11.2–18.9×8.7–15.8 μm. They contain 3–13 nuclei with a mean diameter of between 2.7 and 3.7 μm stained with lactophenol-aceto-orcein. The secondary conidia are of similar shape but smaller. Cystidia and resting spores were not observed.     

Abnormal egg-laying behaviour of female carrot flies (Psila rosae) induced by the fungus Entomophthora muscae

Female carrot flies (Psila rosae F.) infected with Entomophthora muscae (C.) Fres. on days 1 to 4 of adult life were unable to lay their eggs near the food plant. Flies infected on day 5 of adult life showed an egg-laying behaviour similar to that of uninfected flies. E. muscae-infected female carrot flies caught in the field showed an egg-laying behaviour comparable to that of the flies infected in the laboratory during the first day of their live.Eggs laid by infected females were equally as fertile as eggs laid by uninfected females, and developed into normal flies.It is concluded that female carrot flies infected with E. muscae in the field do not in general contribute to the development of the carrot fly population, due to their abnormal egg-laying behaviour.

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Zusammenfassung Wenn Weibchen der echten Möhrenfliegen (Psila rosae) zwischen den ersten und der vierten Tag in dem Laboratorium mit E. muscae infiziert worden waren, könnten sie ihre Eier an den Futterpflanze nicht ablegen. E. muscae-infizierte Weibchen, die ins Feld gefangen worden waren, zeigten auch ein abweichendes Eiablageverhalten. Ihr Behalten war mit dem Verhalten von Fliegen vergleichbar, die an ihrem ersten Lebenstag im Laboratorium infiziert worden waren.Von infizierten Weibchen gelegte Eier waren ebenso fruchtbar wie die von nicht infizierten Weibchen gelegten Eiern. Es wird konkludiert, dass E. muscae-infizierten Weibchen der echten Möhrenfliegen der Entwicklung der Population keinen Beitrag leisten.

     

Some observations onStrongwellsea castrans [Zygomycetes: Entomophthorales], a parasite of root flies [Delia spp.], in the south of Scotland

Root flies,Delia radicum (L.) andD. floralis (Fallén) (Diptera: Anthomyiidae), trapped in swede crops in the south of Scotland, were infected withStrongwellsea castrans Batko and Weiser (Zygomycetes: Entomophthorales). The pathogen was associated mainly with females of the 1st and 2nd generations ofD. radicum and of the single generation ofD. floralis. Disease incidence showed time-lagged density dependence. Greatest sustained infection levels were recorded in the coolest wettest season. Higher proportions ofD. radicum were infected in the headland than in the crop. Unbaited yellow water traps caught greater proportions of infected flies than traps baited with mustard oil (allyl isothiocyanate). The reasons for high levels of infection in Scotland are discussed.

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Résumé Les mouches des racines,Delia radicum (L.) etD. floralis (Fallén) [Diptera: Anthomyiidae] attrapées dans la culture du rutabaga dans le sud de l'écosse, étaient contaminées parStrongwellsea castrans Batko & Weiser [Zygomycetes: entomophthorales]. Le pathogène était surtout associé aux fe nelles de la lère et de la 2ème générations deD. radicum et de la seule génération deD. floralis. Les cas de maladie montraient avec retard leur dépendance avec la densité des mouches. Les niveaux d'infection les plus grands étaient enregistrés pendant la saison la plus froide et la plus humide. Les bordures en friche des parcelles hébergeaient de plus grandes proportions deD. radicum infectés que les cultures. Des pièges jaunes sans appat ont attrapé de plus grandes proportions de mouches que lorsqu'ils étaient appatés avec de l'huile de moutarde (allyl-isothiocyanate). Les raisons qui expliquent pourquoi les niveaux d'infection en écosse sont plus élevés qu'ailleurs sont discutées.

     

Étude de l'évolution d'une mycose aNeozygites fumosa [Zygomycetes,Entomophthorales] dans une population de la cochenille du manioc,Phenacoccus manihoti [Hom.: Pseudococcidae]

Résumé L'action régulatrice d'un champignon du groupe des Entomophthorales,Neozygites fumosa (Speare) Remaudière et Keller, est mise en évidence pour la lère fois dans une population de la cochenille du maniocPhenacoccus manihoti Matile-Ferrero (Homoptères: Pseudococcidae) en République Populaire du Congo en 1982. Le pathogène est de loin l'ennemi naturel qui intervient le plus significativement dans la régulation des effectifs. L'extension de la maladie dans la population h?te appara?t conditionnée par l'apparition simultanée d'une humidité relative supérieure ou égale à 90% et d'une température journalière minimale supérieure à 20°C. Elle semble également liée à la densité du ravageur. On notera enfin que les formes adultes sont systématiquement plus infectées que les formes larvaires.

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Summary The regulatory action of the entomophthoraceous fungusNeozygites fumosa (Speare) Remaudière & Keller (Zygomycetes) is shown for the ist time in a population of the Cassava mealybug,Phenacoccus manihoti (Hom.: Pseudococcidae), in the People's Republic of the Congo in 1982. This pathogen is by far the natural enemy which intervenes the most significantly in the regulation of this insect. The development of the epizootic appears to be influenced by a relative humidity of at least 90% along with a minimal daily temperature greater than 20°C. The spread of the disease also seems to be affected by the host density. Throughout the entire duration of this study, the adult forms were more infected than the larval forms.

     

The culture ofEntomophthora muscae (C) Fres. in carrot flies (Psila rosae F.) and the effect of temperature on the pathology of the fungus

A method for maintaining anin vivo culture ofEntomophthora muscae (C) Fres. on its original host, adult carrot flies (Psila rosae F.), is described. The lethal time for adult carrot flies was greatly influenced by temperature, both for infected and for uninfected flies. In the range 8.2°C–20.2°C the LT₅₀ for infected flies was about 5.4 times shorter than the estimated average life-span for uninfected flies. The discharge of primary spores was also strongly dependent on temperature. The total number of primary spores discharged per fly at 100% RH and in darkness ranged between 1.2×10⁴ and 9.6×10⁴ with a mean of 5.1×10⁴.      

Interference between parasitoids [Hym.: Aphidiidae] and fungi [Entomophthorales] attacking cereal aphids

In field populations of cereal aphids parasitism levels declined through the season as fungal infection increased. In laboratory trials the fungusErynia neoaphidis Remaudiere & Hennebert took 3 to 4 days to kill the rose-grain aphid,Metopolophium dirhodum (Walker), whereas the parasitoidAphidius rhopalosiphi De Stefani-Perez took 8 to 9 days at 20°C. When aphids were infected by the fungus less than 4 days after being parasitized the parasitoids were prevented from completing their development. Conversely, when infection occurred more than 4 days after parasitization development of the fungus was significantly impaired. There was no histological evidence that the fungus invaded the tissues of the parasitoid when both attacked the same aphid. Interference between parasitoids and fungal pathogens must be taken into account when estimating the impact of these mortality agents on pest populations.     

An immunofluorescence study of mitosis in a mite-pathogen,Neozygites sp. (Zygomycetes: Entomophthorales)

Summary Mitosis in a mite-pathogenic species ofNeozygites (Zygomycetes: Entomophthorales) was investigated by indirect immunofluorescence microscopy using an antibody against -tubulin for visualization of microtubules (MTs). DAPI and rhodamine-conjugated phalloidin were used to stain chromatin and actin, respectively. Salient features of mitosis inNeozygites sp. are (1) a strong tendency for mitotic synchrony in any given cell, (2) conical protrusions at the poles of metaphase and anaphase nuclei revealed by actin staining, (3) absence of astral and other cytoplasmic MTs, (4) a spindle that occupies most of the nuclear volume at metaphase, (5) a spindle that remains symmetrical throughout most of mitosis, (6) kinetochore MTs that shorten during anaphase A, (7) a central spindle that elongates during anaphase B, pushing the daughter nuclei into the cell apices, and (8) interpolar MTs that continue to elongate even after separation of the daughter nuclei. Cortical cytoplasmic MTs are present in a few interphasic and post-cytokinetic cells. The data presented show thatNeozygites possesses features unique to this genus and support the erection of theNeozygitaceae as a separate family in theEntomophthorales.Abbreviations DAPI 4,6-diamidino-2-phenylindole - MT microtubule - SPB spindle pole body     

Isolationin vitro ofStrongwellsea castrans [Fungi: Entomophthorales] a pathogen of adult cabbage root flies,Delia radicum [Dipt.: Anthomyiidae]

The entomogenous fungusStrongwellsea castrans was isolatedin vitro for the first time, by incubating conidia projected from infected cabbage root flies (Delia radicum) in a simple, semi-defined liquid medium comprising dextrose, yeast extract and lactalbumin hydrolysate buffered to pH 7. The fungus grew as long unitunicate hyphae. After transfer to a solid nutrient medium, multinucleate hyphal bodies were formed which developed a thick, laminated wall. Neither conidia nor resting spores developed in liquid or on solid media and the fungus survived successive sub-culturing only in liquid media. Using the API-ZYM system, tests on extracts on hyphae ofS. castrans were positive for 11 enzymes but there were no consistent differences in enzyme profiles betweenS. castrans and fungi of the related genusErynia.      

Pathogenicity and specificity of Neozygites tanajoae and Neozygites floridana (Zygomycetes: Entomophthorales) isolates pathogenic to the cassava green mite

The cassava green mite (CGM), Mononychellus tanajoa, a native of South America was accidentally introduced into Africa where it causes serious crop losses. The possibility of introducing classical biological agents from the native home of CGM into Africa was investigated. Thus, we conducted a series of laboratory assays of the native fungal pathogens, Neozygites tanajoae from Brazil and Neozygites floridana from Colombia and Brazil, and compared them with N. tanajoae isolates from Benin. Infectivity of both fungal species, was assayed against the twospotted spider mite, Tetranychus urticae, and against the red mite, Oligonychus gossypii. Pathogenicity against CGM and host range studies were conducted by transferring adult females of each mite species to leaf discs containing sporulated cadavers with a halo of conidia of each fungal isolate. All isolates caused some degree of infectivity to CGM. None of the isolates of N. floridana and N. tanajoae tested were pathogenic to O. gossypii, and only two isolates infected T. urticae. Most isolates from Brazil were highly virulent and infected only CGM. Sixteen N. tanajoae isolates caused more than 89% mortality and more than 62% of the CGM became mummified. A mummified CGM is characteristically a swollen, brown fungus-killed mite that has great potential to produce conidia. However, high mortality was not always associated with high mummification. The median mummification time ranged from 4.4 to 6.7 days. Five Brazilian isolates caused >75% mummification with a median mummification time <5 days. Isolates that cause high mummification in a short period of time would be more likely to cause epizootics and to establish in the new environment. Therefore, these isolates would be the best candidates for introduction to Africa.     

Biochemistry of Esterases Associated with Organophosphate Resistance in Lucilia cuprina with Comparisons to Putative Orthologues in Other Diptera

Esterase activities associated with organophosphate insecticide resistance in the Australian sheep blowfly, Lucilia cuprina, are compared with similar activities in other Diptera. The enzymes making the major contribution to methyl butyrate hydrolysis (ali-esterase) in L. cuprina, M. domestica, and D. melanogaster comigrate during electrophoresis. The enzymes in L. cuprina and D. melanogaster correspond to the naphthyl acetate hydrolyzing E3 and EST23 isozymes of those species. These and previously published data suggest that the ali-esterases of all three species are orthologous. Strains of L. cuprina fall into four groups on the basis of quantitative determinations of their ali-estesterase, OP hydrolase, and malathion carboxylesterase activities and these groups correspond to their status with respect to two types of OP resistance. Strains susceptible to OPs have high ali-esterase, low OP hydrolase, and intermediate MCE activities; those resistant to malathion but not diazinon have low ali-esterase, intermediate OP hydrolase, and high MCE activities; those resistant to diazinon but not malathion have low ali-esterase, high OP hydrolase, and low MCE activities; those resistant to both OPs have low ali-esterase, high OP hydrolase, and high MCE activities. The correlated changes among the three biochemical and two resistance phenotypes suggest that they are all properties of one gene/enzyme system; three major allelic variants of that system explain OP susceptibility and the two types of OP resistance. Models are proposed to explain the joint contribution of OP hydrolase and MCE activities to malathion resistance and the invariant association of low ali-esterase and elevated OP hydrolase activities in either type of resistance.     

A device for preventing houseflies from copulating

The construction and use of a device the virgin chamber, for preventing houseflies, Musca domestica L., from copulating are described. The flies emerge between two horizontal plates kept at a distance of 5 or 4 mm and may be kept in this chamber for 1–3 days before the sexes are separated. The method is regularly used at the Danish Pest Infestation Laboratory for providing unmated flies for selection for resistance to insecticides. Original tests showed that all eggs produced by flies that stayed 1–3 days in the device were infertile, but in recent experiments a small proportion of fertile eggs were found.

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Résumé La construction et l'utilisation d'une chambre de virginité, pour empêcher la copulation de M. domestica L. est décrite. Les mouches émergent entre deux plaques horizontales séparées par 4 à 5 mm et sont maintenues dans cette chambre pendant 1 à 3 jours jusqu'à la séparation des sexes. Il s'agit d'une méthode de routine du Laboratoire Danois des Animaux Nuisibles qui fournit des mouches vierges en vue de la sélection pour la résistance aux insecticides. Les tests originels ont montré que tous les ufs pondus par les mouches qui avaient séjourné de 1 à 3 jours dans le dispositif étaient stériles, mais dans des expériences récentes une faible proportion d'ufs fertiles a été observée dans quelques tests.

     

广东省侵染摇蚊的库蚊虫霉研究

在中国首次报道了侵染摇蚊的库蚊虫霉(Entomophtora culichs)的形态特征、侵染症状以及发生动态等。库蚊虫霉初生分生孢子,钟罩形,顶部有一明显小尖突,(14.6±1.5)μm×(10.8±1.2)μm。次生分生孢子形似初生分生孢子,但略小,(11.0±1.1)μm×(8.0±0.5)μm。休眠孢子,球形至亚球形,直径(29.3±1.9)μm。被侵染摇蚊尸体以假根附着于基物上,通体覆盖着绿色的真菌子实层。库蚊虫霉侵染率在不同植物上明显不同。库蚊虫霉侵染率随季节不同波动很大,春季最高达90.2%,其次为秋季和冬季,夏季最低为0。2006—2009年库蚊虫霉侵染率为93.5%～95.7%,年际之间变动不大。