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In this study we report the effects of fungal metabolites isolated from cultures of the fungus Trichoderma citrinoviride ITEM 4484 on the feeding preference of the aphid Rhopalosiphum padi, a major pest of cereal crops. Different phagodeterrent metabolites were purified by a combination of direct and reverse phase column chromatography and thin-layer chromatography. Chemical investigations, by spectroscopic and chemical methods, led to the identification of different long chain primary alcohols (LCOHs) of the general formula R-OH, wherein R is a long, unbranched, unsubstituted, linear aliphatic group. LCOHs have been reported as components of lepidopteran pheromone blends, but their phagodeterrent effect to aphids is herein reported for the first time. The effects of LCOHs on R. padi were studied by behavioral and electrophysiological bioassays. Feeding preference tests that were carried out with winged and wingless morphs of R. padi showed that LCOHs had high phagodeterrent activity and restrained aphids from settling on treated leaves at a concentration as low as 0.15 mM (0.036 g/l). The results of different electrophysiological analyses indicated that taste receptor neurons located on the aphid tarsomeres were involved in the LCOHs perception. Behavioral assays carried out with some commercial agrochemicals, including azadirachtin A, pyrethrum and a mineral oil-based product, in combination with 1-hexadecanol, the LCOH most abundantly produced by T. citrinoviride ITEM 4484, showed that these different active principles could be applied together, resulting in a useful increase of the phagodeterrent effect. The data shown indicate that these compounds can be profitably utilized for novel applications in biotechnical control of aphid pests. Furthermore, the tested LCOHs have no chiral centers and therefore can be obtained with good yield and at low cost through chemical synthesis, as well as from natural sources.

This study investigated the effects of a strain of the fungus Lecanicillium lecanii (Zimm.) Zare and Gams (Hypocreales: Ascomycota) on the aphid Schizaphis graminum (Rondani) (Hemiptera: Aphididae). The fungus was administered to fourth instar nymphs and to alate and apterous adult morphs as a ground rice-kernel formulation. This study showed that L. lecanii formulation affected the survival of the aphids and interacted differently with the studied morphs, the Lethal Time values being lower for alate compared to apterous morphs and nymphs. The treatment also caused a significant reduction in the fecundity of the three treated aphid groups. Histological analysis revealed that the hyphae invaded the host hemocoel of a limited number of alate and apterous morphs: the fungus only entered through the spiracles. Scanning Electron Microscope observations revealed that L. lecanii adhered to the body surface of both adults and nymphs without differentiation. In conclusion, the present study suggests that this strain might be a good candidate for a programme of biological control of S. graminum and other aphid species.

In a simple experimental system with wheat plants, interactions between the collembolan Onychiurus armatus and Trichoderma harzianum, a soil-borne fungus parasite of several plant pathogenic fungi, were studied in the presence of Gaeumannomyces graminis var. tritici , one of the most important foot and root fungal pathogens of cereals world-wide. Trichoderma harzianum was used according to two different modalities: fungal inoculum applied to seed or mixed with substrate. The isolate of T. harzianum proved to be ineffective against the pathogenic fungus, independent of the application modality, as shown in the experiments where this fungus was used alone, whereas Collembola used alone significantly reduced disease severity. However, the mode by which T. harzianum was applied significantly influenced the disease control ability of Collembola. In fact, only when T. harzianum was coated to seed were springtails able to reduce the disease caused by G. graminis var. tritici.

Funghi appartenenti a diverse specie del genere Trichoderma sono stati fino ad ora utilizzati per il controllo di funghi fitopatogeni e il loro impiego ha confermato l'assenza di effetti negativi per la salute umana e l'ambiente. Nostri precedenti studi hanno messo in evidenza per alcune specie del genere Trichoderma una potenziale attività di controllo degli afidi. Tali insetti rappresentano uno dei più importanti gruppi di fitomizi di molte coltivazioni di notevole interesse economico. Il danno provocato alla pianta ospite non è dovuto solo alla sottrazione di elementi nutritivi e ormonali attraverso la suzione della linfa, ma anche e soprattutto, alla capacità di trasmettere virus. Test di preferenza alimentare da noi condotti, hanno evidenziato una significativa attività fagodeterrente di colture dell'isolato di T. citrinoviride ITEM 4484 nei confronti di diverse specie di afidi tra cui il polifago Myzus persicae (Sulzer) e Rhopalosiphum padi un diffuso parassita dei cereali. Ulteriori indagini condotte mediante metodi spettroscopici (essenzialmente NMR e tecniche MS) e volte ad identificare i composti fungini con attività fagodeterrente prodotti dall'isolato di T. citrinoviride hanno permesso di isolare e caratterizzare quattro distinti gruppi di metaboliti tra di loro strutturalmente correlati. I primi due metaboliti, denominati citrantifidiene e citrantifidiol, sono rispettivamente un esa-1,3-dienil estere dell'acido acetico simmetrico ed un cicloesadien-1,3-diolo tetrasostituito. Altri due composti individuati appartengono alla famiglia dei bisorbicillinoidi, e sono il bislongiquinolide e il diidrotricodimerolo. Il quarto gruppo, il quale mostra una spiccata attività fagodeterrente, è composto da diversi alcoli primari a catena lunga: esadecanolo, octadecanolo, cis-9-octadecanolo e trans-9-octadecanolo. Questi composti non hanno centri chirali e oltre ad essere reperibili in natura possono essere facilmente sintetizzati con una buona resa. Per questi alcoli a catena lunga e per altri ad essi chimicamente correlati è stata depositata una domanda di brevetto nazionale (MI2011A000831), con successiva estensione a livello internazionale (PTC IB2012/052383) per la loro attività fagodeterrente che potrà essere utilizzata per il controllo biotecnico degli afidi.

Take-all and brown foot rot, caused respectively by Gaeumannomyces graminis var. tritici and Fusarium culmorum, are two important components of the foot and root fungal disease complex of winter cereals world-wide. These fungi persist in soil and in crop debris in the same layer of agricultural soil as Collembola, a well represented taxon of soil animals. Previous in vitro tests showed that these fungi grown on agarised medium were readily consumed by springtails. In a simplified experimental system with wheat plants and the pathogenic fungi grown on millet and wheat kernels, the severity of disease was significantly reduced by collembolan feeding activity.

Fungal strains belonging to the genera Fusarium, Paecilomyces and Trichoderma were tested in vitro in order to study their effects against Schizaphis graminum, one of the major pests of cereal crops around the world. Biological assays were performed using a solid formulation that was obtained from fungal cultures grown on rice and then finely ground ( 60%) within 10 minutes after application. As F. larvarum ITEM 2139 did not produce metabolites toxic to mammals, it might be a good candidate as a biocontrol agent of S. graminum in the field.

The morphogenesis of the gut during nauplius stages of Lernaea cyprinacea was reported to occur, without apparent morphogenetic movements, by a non‐simultaneous and spatially determined differentiation of the various regions. A developing foregut was first seen at the nauplius‐II stage, immediately after moult; on the other hand, the midgut did riot appear until the moult to nauplius III. Gut development initially involved some cells containing refractive granules in their cytoplasm. At the nauplius‐III stage, strongly Pyronin‐positive cells without granules appeared among those with granules. Subsequently, the mouth parts and the hindgut were formed. Only in the late nauplius III and early copepodid‐I stages did all parts fuse into a continuous digestive tube. The finding that the entire gut apparatus develops during the naupliar life was taken to suggest that this stage is part of the embryonic development of L. cyprinacea.

Effects of interactions of the collembolan Protaphorura armata, the arbuscular mycorrhizal fungus Glomus intraradices and the foot and root pathogenic fungus Fusarium culmorum on growth and health of wheat plants were studied in modified Leonard bottle-jars containing sterile sand and peat. The mycorrhizal colonisation, the root and shoot dry weight and the disease index were determined. Also the number of adult Collembola was determined and their gut content analysed. The presence of Collembola did not decrease the positive effect of the arbuscular mycorrhizal fungus Glomus intraradices on the plant biomass and did not reduce the biocontrol capacity of this fungus.

We report the effects of some bisorbicillinoids isolated from biomass of the fungus Trichoderma citrinoviride on settling and feeding preference of the aphid Schizaphis graminum. Purification of the fungal metabolites was carried out by a combination of column chromatography and thin-layer chromatography using direct and reverse phases. Chemical identification was performed by spectroscopic methods including nuclear magnetic resonance and mass spectrometry. The identified bisorbicillinoids appeared to be bislongiquinolide, its 16,17-dihydro derivative, trichodimerol, and dihydrotrichodimerol. A feeding preference test with alate morphs of S. graminum was used to identify the active fractions. Among the four bisorbicillinoids, dihydrotrichodimerol and bislongiquinolide influenced aphid feeding preference, restraining specimens from settling on leaves treated with metabolites. Taste neurons sensitive to these compounds, particularly to bislongiquinolide, were located on tarsi of the S. graminum alate morphs.

Two novel metabolites with potential antifeedant activity were isolated from cultures of the fungus Trichoderma citrinoviride strain ITEM 4484 grown in solid-state fermentation on sterile rice kernels. The producing strain was identified at species level by sequence analysis of the internal transcribed spacer regions ITS-1 and ITS-2 of the nuclear rDNA and a fragment of the translation elongation factor gene TEF-1alpha. Fractionation by column chromatography and TLC of the culture organic extract, followed by feeding preference tests on the aphid pest Schizaphis graminum (Rondani), allowed the purification of 5.8 and 8.9 mg/kg of culture of two bioactive metabolites, which were named citrantifidiene and citrantifidiol ( 1 and 2). Citrantifidiene and citrantifidiol, whose structures were determined by spectroscopic methods (NMR and MS) are a symmetrical disubstituted hexa-1,3-dienyl ester of acetic acid and a tetrasubstituted cyclohexane-1,3-diol, respectively. The pure metabolites influenced the feeding preference of S. graminum restraining individuals from feeding on wheat leaves dipped in 5% aqueous methanol solution containing 0.57 mg/mL of citrantifidiene or 0.91 mg/mL of citrantifidiol.

The feeding preferences of alate and apterous morphs of the aphid Schizaphis graminum (Rondani) (Homoptera: Aphididae) were evaluated using leaves treated with powdered rice cultures of four fungal isolates belonging to different species of the genus Trichoderma (Deuteromycotina: Hyphomycetes). All of the fungal isolates restrained alate morphs of S. graminum from visiting treated leaves, but only Trichoderma citrinoviride Bisset also influenced the preference of apterous morphs. Trials carried out with supernatants obtained by centrifuging aqueous suspensions of the fungal cultures showed that the feeding preference of aphids was maintained in the absence of fungal spores and mycelia, supporting the hypothesis that at least part of the fungal metabolites responsible for this effect were water-soluble compounds. Electrophysiological studies showed that the structures involved in the perception of the fungal metabolites are located on the aphid tarsomeres.

Summary Effects of compost products, enriched or not-enriched with a strain of the mycoparasitic fungus Trichoderma atroviride, on the survival of the collembolan Protaphorura armata and the viability of fungal conidia after the transit through the springtail gut were investigated. The effect of compost enriched with Ca-Lignosulphonate (Ca-Ls), a low cost by-product of the acid sulphite pulping process, with lignin-like structure, on P. armata was also evaluated. All compost products enriched or not with the mycoparasitic fungus or Ca-Ls did not affect P. armata survival. No statistical differences were found in animal survival for different types of product or in enriched and not-enriched products. In addition to adults, live juveniles were also observed in all compost products. The gut content of animals, collected at the end of the survival test from compost enriched with T. atroviride, was examined under the light microscope, and in a few cases observations revealed the presence of some T. atroviride conidia. Subsequent tests carried out to study the viability of conidia after the transit through the springtail gut showed that colonies of the fungus developed from all faecal pellets produced by adult and juveniles specimens of P. armata previously fed on conidia of T. atroviride. These results suggest compatibility between Collembola and Trichoderma or Ca-Ls in the composts.

Summary The feeding preference of the collembolan Protaphorura armata in the presence of Fusarium culmorum and Gaeumannomyces graminis var. tritici, two soil-borne fungi pathogenic for winter cereals, was studied in a simplified experimental system including wheat seedlings. Analysis of gut content of all animals from microcosms containing inoculum of both fungi showed that F. culmorum was clearly preferred but that G. graminis var. tritici was also fed. At microscopic examination the majority of F. culmorum conidia present in the gut lacked cytoplasmic content, and only few conidial cells were intact. The feeding preference of P. armata favoured G. graminis var. tritici over F. culmorum in the competition for infection sites on wheat plants; in fact, the former resulted the prevalent cause of plant disease. The viability of fungal propagules after passage through the gut of P. armata was also studied. No colonies of G. graminis var. tritici and only a few colonies of F. culmorum developed from faecal pellets set on agar medium. Fungal propagules dispersed by springtails were not sufficient to induce disease, as demonstrated by introducing animals, previously fed on fungal cultures separately, into microcosms containing a sterile substrate where wheat kernels were seeded.

The cross-talk between neurons and glia is receiving increased attention because of its potential role in information processing in the nervous system. We choose the cluster of supramedullary neurons (SN) and glial cells of pufferfish as a suitable model for studying neuron-glial interactions, identifying the implicated cell types and the signalling involved. In particular, among proopiomelanocortin (POMC)-derived peptides, adrenocorticotrope hormone (ACTH)-immunopositivity was found both in SN and in microglial cells. The present results for the first time show the presence of ACTH in microglia of a vertebrate. The role of ACTH is discussed, including its possible neuroprotective function. Moreover, SN immunoreactivity supports the idea that ACTH participates in neurotransmission and/or neuromodulation. In addition to these possible functions, the hypothesis is put forward that ACTH represents a common language by which neurons and glial cells communicate with each other.

The effects of beauvericin, a toxic fungal metabolite common contaminant of maize and wheat, on aphid fitness were studied in three consecutive generations of females. Aphids were reared on wheat leaves inserted into a sandy substratum wetted with a solution of beauvericin. Ingestion of this solution through leaves did not significantly decrease the lifespan of females of all generations as compared to controls. However, the mean number of offspring from the third generation of treated females was significantly smaller than those in controls. Furthermore, treated second and third generation females produced a greater number of abortive embryos. Histological analysis revealed abundant DAPI and Feulgen positive material in the cytoplasm of some bacteriocytes of treated third generation females. This material was attributed to the endosymbionts of bacteriocytes. Tests by contact were also carried out and revealed a significantly lower survival of treated first instar aphids as compared to controls 18 h after the start of the trial.

Interactions between springtails (Onychiurus armatus, O. tuberculatus, Folsomia candida) and plant pathogenic fungi (Gaeumannomyces graminis var. tritici, Fusarium culmorum, Bipolaris sorokiniana and Rhizoctonia cerealis) living in the same soil layer were investigated under laboratory conditions. The fungi are responsible for the foot and root disease complex of winter cereals and can cause serious reduction in yield. Since these species coexist in agricultural soils, feeding preference tests were performed in such a way that the springtails were allowed to interact simultaneously with all the tested fungi. O. armatus, O. tuberculatus and F. candida fed on the mycelia of G. graminis var. tritici, F. culmorum and R. cerealis: in the first hours of the trials individuals of all springtail species distributed themselves among the colonies, though moving from one to the other. Subsequently F. culmorum mycelium became the preferred food of all species tested; however, the other two fungi continued to be consumed. Mycelia of G. graminis var. tritici, F. culmorum and R. cerealis were shown to be an adequate food source for reproduction of Folsomia candida. Bipolaris sorokiniana mycelium had both repellent and lethal effects on F. candida and O. armatus, whereas this fungus was lethal, although not repellent, for O. tuberculatus. Long-term experiments on F. candida indicate that conidia of B. sorokiniana were eaten and proved to be a sufficient diet for reproduction.

The short-term effects on soil microarthropods of the herbicide triasulfuron, belonging to the chemical class of sulfonylureas, were evaluated in two fields which had never been treated with sulfonylureas, and were cultivated with winter wheat. In particular, the effects of single applications at rates corresponding to two- (rate 2) and sixfold (rate 6) the recommended agricultural rate (7.5 g active ingredient ha–1) were analysed and compared with controls. The changes in the populations of the main groups of microarthropods were evaluated. Rate 2 had very low effects, whereas rate 6 produced a significant decrease in the number of microarthropods, Acarina and Collembola in the surface soil layer (0–7.5 cm). The Collembola were analysed at the species level. Statistical analysis revealed significant differences only for a few species, and only after treatment with the highest rate of triasulfuron. Finally, the results of the field tests were compared to those of laboratory tests carried out previously, which examined the effects of the same herbicide on a collembolan species.

Triasulfuron, a member of the sulfonylureas class of herbicides, was tested under laboratory conditions on the collembolan Onychiurus pseudogranulosus. Pure triasulfuron and commercial formulations at different rates (starting from a dose about 5 times the recommended agricultural rate) were tested separately on one-week old juveniles and adults reared in the laboratory. The persistence of the herbicide at the end of the trials lasting 30 and 60 days was verified by HPLC analysis. Laboratory tests indicated that only the rates exceeding 500 times the soil concentration expected soon after field application of the herbicide directly affected the tested species.

Cultures of isolated neuronal populations from the central ganglia of the gastropod mollusc Planorbarius corneus were used for testing the effects of inorganic lead. The examined parameters were cell survival, neurite outgrowth and cytoskeletal morphology. In large heterogeneous neuronal populations as obtained from a whole cerebral or pedal ganglion, the different sensitivity to lead is reflected mainly on the cell survival. The neurons belonging to the homogeneous E cluster population are more sensitive; in fact a higher percentage of them do not survive in the presence of lead. Moreover, in this neuronal cell type the neurite outgrowth is dramatically affected by lead only when the neurons are cultured on conditioned substrate. Possibly, membrane mechanisms activated for the neurite outgrowth represent a target for inorganic lead. The few neurites sprouted in presence of lead do not evidence changes in the cytoskeletal components.

Six nervous system drugs which inhibited vegetative reproduction of S. cerevisiae arrested also mitotic division of C. utilis, C. albicans and C. tropicalis. Chlorpromazine and chlorpheniramine which proved to be the most effective, affected respiration and cytochrome biosynthesis. Electrophoretic bands with MW congruent to 100 K were faint in silver-stained electrophoregrams of proteins from cells grown in the presence of a sub-inhibitory concentration of chlorpromazine.

A homogeneous population of gram-negative elongated bacteria grows within the canal system of the calcareous sponge Clathrina cerebrum (Haeckel). The location of symbionts is probably related to the ascon organization of the host sponge. Bacteria have outer and inner membranes in the cell envelope and a number of lamellar mesosomes in the cytoplasm. They show a rough surface and peduncles at both extremities, due to a complex folding of the outer membrane. Most bacteria live in close association with the choanocytes, cells lining sponge canals. After in vitro sponge dissociation, bacteria still embrace isolated sponge cells, and show gliding motility and flexibility. Owing to motile properties and surface topography, these bacteria are tentatively referred to the genus Cytophaga , a group related to flexibacteria. A survey of three Clathrina species, C. cerebrum , C. coriacea (Montagu), and C. clathrus (Schmidt), living in the same or in neighbouring habitats, shows that only C. cerebrum can host a well-developed population of bacterial symbionts. This association is probably based on some species-specific mechanism, and shows distinctive features with respect to those which have been described to date in sponges.

Microarthropod populations were studied in alfalfa fields treated with the following organic fertilizers: poultry manure, sheep manure, worm compost, urban sludge, sterilized urban sludge, straw and vetch green manure. The manures were added to enrich the soil to the same level of 4% organic matter. Two different doses of mineral fertilizers were also tested. The experiment was carried out on randomized block field plots on two adjacent areas, one of which was irrigated by sprinklers. Irrigation strongly influences the effects of organic and mineral fertilizers on microarthropod population densities. The effects are different depending on the taxon considered and may even be species specific.

We report preliminary findings on the development of Lernaea cyprinacea from cleavage to hatching. The eggs are centrolecithal with superficial cleavage. Nucleoli were found in the nuclei of the perilecithal syncytium. Gastrulation occurs by in situ aggregation of cells, without evident morphogenetic movements.

E.U.E. cells (general population) were submitted to biochemical and cytoenzymatic tests to compare the enzymatic profile of E.U.E. cells (controls) with that of E.U.E. adapted to hypertonic medium. The adapted cells are characterized by very high oxoreductase activity (LDH, HBDH, G-6-P DH) and very high alkaline-phosphatase activity. Clones derived from general population were also submitted to biochemical tests to characterize those more strictly related to the enzymatic profile of adapted cells. The profile of clone N. 13 resembles on this respect that of the adapted cells. The high redox activity is a prerequisite supporting energy supply for osmotic work. The increased activity of plasma membrane enzymes of the adapted cells is also demonstrable in cells exposed for short time to salinity.

In five species of Onychiurus (Collembola Onychiuridae) the diploid chromosome number is higher in animals of narrower width. In three species of Acerentomon (Protura Acerentomidae) the number of the chromosome arms (FN), not the chromosome number, is inversely correlated to the length of the foretarsus and consequently to the narrowness of the passages they can frequent. These correlations are discussed from the point of view of the animal's habitat, of the karyotypes of other species of the same family, and of some morphological characters of the adults.