Sustained high levels of foliar herbivory of the mangrove Rhizophora stylosa by a moth larva Doratifera stenosa (Limacodidae) in north-eastern Australia

The moth larva, Doratifera stenosa (Lepidoptera: Limacodidae), was observed feeding voraciously in great numbers on mature leaves of Rhizophora stylosa in mangroves at Port Curtis in Central Queensland, NE Australia. This behaviour was considered unusual since mangroves, and the Rhizophora species in particular, reportedly harbour few herbivores and have relatively low levels of herbivory, less than 10%. During a two year period (1996-1998), larvae were observed consuming around 30-40% of leaves in the canopy each year, and the mangroves appeared able to sustain these high levels of herbivory. The impact on trees was assessed in conjunction with a study of the herbivore, its behaviour and life history, in an attempt to explain the occurrence. Larvae were 1-2 cm in length, bright green and gregarious, with numerous small, stinging hairs along their upper bodies. Feeding was in small cohort groups of 5-70 individuals that broke up immediately prior to each moult after which they regrouped in much larger numbers of mixed cohorts to form single-file processions across branches, stems and prop roots. In this way, they moved to neighbouring trees with less affected foliage. One of the outstanding characteristics of this herbivore was its ability to desist from killing host trees although it appeared quite capable of doing so had it remained on individual trees. By moving from tree to tree, the herbivore was able to heavily crop Rhizophora foliage in an apparently sustainable manner. These findings demonstrate the role and importance of foliar herbivory in severely affected forests and how such instances best not be ignored or treated as curiosities in future assessments of herbivory and forest turnover in mangrove ecosystems.

Exotic vs endemic biocontrol agents: Would the real stratiolaelaps miles (Berlese) (Acari : Mesostigmata : Laelapidae), please stand up?

The ability of introduced organisms to invade undisturbed native habitats is a major concern in conservation biology and has resulted in a re-evaluation of the introduction of exotic biocontrol agents, especially of generalist predators. One such agent is Stratiolaelaps miles (Berlese), a predatory mite described from Italy, known from throughout the Holarctic, and apparently accidentally introduced to other areas of the world, including Australia. Initial investigations revealed that putative S. miles could be found in both disturbed and relatively pristine habitats in Queensland, Australia. However, analysis of the mitochondrial DNA of five populations showed most to be highly divergent genetically. Subsequent morphological analysis established two species groups: the lamington-group from cool-temperate to subtropical rainforests in Eastern Australia and the more eurytopic miles-group with a cosmopolitan distribution. We describe two new species from each of these complexes (Stratiolaelaps womersleyi, Stratiolaelaps lamington; Stratiolaelaps marilyn, Stratiolaelaps lorna, respectively), and resurrect Stratiolaelaps scimitus (Womersley), a species which often appears to have been confused with S. miles. Additionally, the large genetic distances among morphologically homogenous species in the miles-group suggest that the apparently cosmopolitan S. miles may be composed of a suite of cryptic species of potentially varying utility in biological control. (C) 2002 Elsevier Science (USA). All rights reserved.

Application of quantitative analytical electron microscopy to the mineral content of insect cuticle

Quantification of calcium in the cuticle of the fly larva Exeretonevra angustifrons was undertaken at the micron scale using wavelength dispersive X-ray microanalysis, analytical standards, and a full matrix correction. Calcium and phosphorus were found to be present in the exoskeleton in a ratio that indicates amorphous calcium phosphate. This was confirmed through electron diffraction of the calcium-containing tissue. Due to the pragmatic difficulties of measuring light elements, it is not uncommon in the field of entomology to neglect the use of matrix corrections when performing microanalysis of bulk insect specimens. To determine, firstly, whether such a strategy affects the outcome and secondly, which matrix correction is preferable, phi-rho (z) and ZAF matrix corrections were contrasted with each other and without matrix correction. The best estimate of the mineral phase was found to be given by using the phi-rho (z) correction. When no correction was made, the ratio of Ca to P fell outside the range for amorphous calcium phosphate, possibly leading to flawed interpretation of the mineral form when used on its own.

Insect chemistry and chirality

Examination of the chemistry of a number of Australian insect species provided examples of unusual structures and encouraged determinations of their absolute stereochemistry by stereocontrolled syntheses and chromatographic comparisons. Inter alia, studies with the fruit-spotting bug (Amblypelta nitida), certain parasitic wasps (Biosteres sp.), the aposematic shield bug (Cantao parentum), and various species of scarab grubs are summarized. The determination of enantiomeric excesses (ee's) for component epoxides, lactones, spiroacetals, and allenes are described. Stereochemical and related aspects of the biosynthesis of spiroacetals in certain fruit-fly species (Bactrocerae sp.) are also presented.

Purification and characterisation of functional early pregnancy factor expressed in Sf9 insect cells and in Escherichia coli

Early pregnancy factor (EPF) is a secreted protein with growth regulatory and immunomodulatory properties. It is an extracellular form of the mitochondrial matrix protein chaperonin 10 (Cpn10), a molecular chaperone. An understanding of the mechanism of action of EPF and an exploration of therapeutic potential has been limited by availability of purified material. The present study was undertaken to develop a simple high-yielding procedure for preparation of material for structure/function studies, which could be scaled up for therapeutic application. Human EPF was expressed in Sf9 insect cells by baculovirus infection and in Escherichia coli using a heat inducible vector. A modified molecule with an additional N-terminal alanine was also expressed in E coli. The soluble protein was purified from cell lysates via anion exchange (negative-binding mode), cation exchange, and hydrophobic interaction chromatography, yielding similar to42 and 36 mg EPF from 300 ml bacterial and I L Sf9 cultures, respectively. The preparations were highly purified ( greater than or equal to99% purity on SDS-PAGE for the bacterial products and greater than or equal to97% for that of insect cells) and had the expected mass and heptameric structure under native conditions, as determined by mass spectrometry and gel permeation chromatography, respectively. All recombinant preparations exhibited activity in the EPF bioassay, the rosette inhibition test, with similar potency both to each other and to the native molecule. In two in vivo assays of immuno suppressive activity, the delayed-type hypersensitivity reaction and experimental autoimmune encephalomyelitis, the insect cell and modified bacterial products, both with N-terminal additions (acetylation or amino acid), exhibited similar levels of suppressive activity, but the bacterial product with no N-terminal modification had no effect in either assay. Studies by others have shown that N-terminal addition is not necessary for Cpn10 activity. By defining techniques for facile production of molecules with and without immunosuppressive properties, the present studies make it possible to explore mechanisms underlying the distinction between EPF and Cpn10 activity. (C) 2003 Elsevier Inc. All rights reserved.

Towards the on-farm conservation of the assassin bug Pristhesancus plagipennis (Walker) (Hemiptera : Reduviidae) during winter using crop plants as refuges

Habitat instability associated with seasonal crop succession in broad-acre farming systems presents a problem for the conservation and utilisation of beneficial insects in annual field crops. The present paper describes two experiments used to measure the potential of seven plant species to be utilised as winter refuges to support and conserve the predatory bug Pristhesancus plagipennis (Walker). In the first experiment, replicated plots of canola (Brassica napus ), red salvia (Salvia coccinea ), niger (Guizotia abyssinica ), linseed (Linum usitatissimum ), lupins (Lupinus angustifolius ), and lucerne (Medicago falcata ) were planted in a randomized experiment during Autumn 1998. Upon crop establishment, adults and nymphs of P. plagipennis were released into treatment plots and their numbers were assessed, along with those of their potential prey, throughout the ensuing winter months. Post-release sampling suggested that canola and niger retained a proportion of adult P. plagipennis , while niger, lucerne and canola retained some nymphs. The other plant species failed to support P. plagipennis nymphs and adults postrelease. In the second experiment, niger was compared with two lines of sunflower (Helianthus annus ). Both sunflower lines harboured significantly higher (P < 0.05) densities of P. plagipennis nymphs than did niger. The more successful refuge treatments (sunflower, niger and canola) had an abundance of yellow flowers that were attractive to pollinating insects, which served as supplementary prey on which P. plagipennis were observed to feed. Sunflower and niger also supported high densities of the prey insect Creontiades dilutus (Stal) and provided protective leafy canopies which supplied shelter during the winter months. The potential and limitations for using each plant species as a winter refuge to retain P. plagipennis during winter are discussed.

Studies in search of a suitable experimental insect model for xenodiagnosis of hosts with Chagas' disease: 3 - On the interaction of vector species and parasite strain in the reaction of bugs to infection by Trypanosoma cruzi

The reaction of nine vector species of Chagas' disease to infection by seven different Trypanosoma cruzi strains; Berenice, Y, FL, CL, S. Felipe, Colombiana and Gávea, are examined and compared. On the basis of the insects' ability to establish and maintain the infection, vector species could be divided into two distinct groups which differ in their reaction to an acute infection by T. cruzi. While the proportion of positive bugs was found to be low in Triatoma infestans and Triatoma dimidiata it was high, ranging from 96.9% to 100% in the group of wild (Rhodnius neglectus, Triatoma rubrovaria)and essentially sylvatic vectors in process of adaptation to human dwellings, maintained under control following successful insecticidal elimination of Triatoma infestans (Panstrongylus megistus, Triatoma sordida and Triatoma pseudomaculata). An intermediate position is held by Triatoma brasiliensis and Rhodnius prolixus. This latter has been found to interchange between domestic and sylvatic environments. The most important finding is the strikingly good reaction between each species of the sylvatic bugs and practically all T. cruzi strains herein studied, thus indicating that the factors responsible for the excellent reaction of P.megistus to infection by Y strain, as previously reported also come into operation in the reaction of the same vector species to acute infections by five of the remaining T.cruzi strains. Comparison or data reported by other investigators with those herein described form the basis of the discussion of Dipetalogaster maximus as regards its superiority as a xenodiagnostic agent.

Studies in search of a suitable experimental insect model for xenodiagnosis of hosts with Chagas' disease: 4 - The reflection of parasite stock in the responsiveness of different vector species to chronic infection with different Trypanosoma cruzi stocks

Previous studies (1982,1987) have emphasized the superiority of sylvatic vector species over domestic species as xenodiagnostic agents in testing hosts with acute or chronic infections by T. cruzi "Y" stock. The present study, which is unique in that it contains data on both infectivity rates produced by the same stock in 11 different vector species and also the reaction of the same vector species to seven different parasite stocks, establishes the general validity of linking efficiency of xenodiagnosis to the biotope of its agent. For example, infectivity rates produced by "São Felipe" stock varied from 82.5% to 98.3% in sylvatic vectors but decreased to 42.5% to 71.3% in domestic species. "Colombiana" stock produced in the same sylvatic vectors infectivity rates ranging from 12.5% to 45%. These shrank to 5%-22.5% in domestic bugs. The functional role of the biotope in the vector-parasite interaction has not been eluddated. But since this phenomenon has been observed to be stable and easy to reproduce, it leads us to believe that the results obtained are valid. Data presented also provide increasing evidence that the infectivity rates exhibited by bugs from xenodiagnosis in chronic hosts, are parasite stock specific. For example, infectivity rates produced by "Berenice", "Y", "FL" and "CL" varied in R. neglectus from 26.3% to 75%; in P. megistus from 56.3% to 83.8%; in T. sordida from 28.8% to 58.8% in T. pseudomaculata from 41.3% to 66.3% and in T. rubrovaria from 48.8% to 85%. Data from xenodiagnosis in the same hosts, carrying acute infections by the same parasite stocks, gave the five sylvatic vectors a positive rating of approximately 100%, thus suggesting that the heavy loads of parasites circulating in the acute hosts obscured the characteristic interspecific differences for the parasite stock. Nonetheless these latter were revealed in the same hosts with chronic infections stimulated by very low numbers of the same parasite stocks. Certain observations here described lead us to speculate as to the possibility of further results from other parasite stocks, allowing the association of the infectivity rates produced in bugs by different parasite stocks with the isoenzymic patterns revealed by these stocks.

Model risk in the pricing of exotic options

The growth experimented in recent years in both the variety and volume of structured products implies that banks and other financial institutions have become increasingly exposed to model risk. In this article we focus on the model risk associated with the local volatility (LV) model and with the Variance Gamma (VG) model. The results show that the LV model performs better than the VG model in terms of its ability to match the market prices of European options. Nevertheless, both models are subject to significant pricing errors when compared with the stochastic volatility framework.

Resistance of mice immunized with killed culture trypomastigotes against infection by insect-derived trypomastigotes of Trypanosoma cruzi

Mice immunized with heat or merthiolate-killed culture trypomastigotes of the non-virulent G strain were resistant to the challenge by insect-derived trypomastigotes of the CL strain of Trypanosoma cruzi. No parasitemia was detected, by direct microscopic examination of blood samples, in 90% of immunized mice while all control animals developed a high parasitemia. Trypsinization before heat-inactivation, or fixation with paraformaldehyde, apparently reduced the immunogenicity of the G strain trypomastigotes. Mice immunized with trypomastigotes treated by either of these procedures were not protected against infection by virulent T. cruzi. Analysis of the 13I-labeled surface proteins of G strain trypomastigotes inactivated by the various methods suggests that these components are involved in eliciting protective immunity against T. cruzi infection.

Complex Dynamics of defective interfering baculoviruses during serial passage in insect cells

Defective interfering (DI) viruses are thought to cause oscillations in virus levels, known as the ‘Von Magnus effect’. Interference by DI viruses has been proposed to underlie these dynamics, although experimental tests of this idea have not been forthcoming. For the baculoviruses, insect viruses commonly used for the expression of heterologous proteins in insect cells, the molecular mechanisms underlying DI generation have been investigated. However, the dynamics of baculovirus populations harboring DIs have not been studied in detail. In order to address this issue, we used quantitative real-time PCR to determine the levels of helper and DI viruses during 50 serial passages of Autographa californica multiple nucleopolyhedrovirus (AcMNPV) in Sf21 cells. Unexpectedly, the helper and DI viruses changed levels largely in phase, and oscillations were highly irregular, suggesting the presence of chaos. We therefore developed a simple mathematical model of baculovirus-DI dynamics. This theoretical model reproduced patterns qualitatively similar to the experimental data. Although we cannot exclude that experimental variation (noise) plays an important role in generating the observed patterns, the presence of chaos in the model dynamics was confirmed with the computation of the maximal Lyapunov exponent, and a Ruelle-Takens-Newhouse route to chaos was identified at decreasing production of DI viruses, using mutation as a control parameter. Our results contribute to a better understanding of the dynamics of DI baculoviruses, and suggest that changes in virus levels over passages may exhibit chaos.

Systems biotechnology of baculovirus-producing insect cells

Dissertation presented to obtain the Ph.D degree in Ciências da Engenharia e Tecnologia, especialidade Biotecnologia

Characterization of Pinus spp. needles by gas chromatography and mass spectrometry: Application to plant-insect interactions

Dissertação apresentada para obtenção do Grau de Doutor em Ciências do Ambiente pela Universidade Nova de Lisboa, Faculdade de Ciências e Tecn

Genetic Characterization of an Insect-Specific Flavivirus Isolated from Culex Theileri Mosquitoes Collected in Southern Portugal

We describe the full genetic characterization of an insect-specific flavivirus (ISF) from Culex theileri (Theobald) mosquitoes collected in Portugal. This represents the first isolation and full characterization of an ISF from Portuguese mosquitoes. The virus, designated CTFV, for Culex theileri flavivirus, was isolated in the C6/36 Stegomyia albopicta (=Aedes albopictus) cell line, and failed to replicate in vertebrate (Vero) cells in common with other ISFs. The CTFV genome encodes a single polyprotein with 3357 residues showing all the features expected for those of flaviviruses. Phylogenetic analyses based on all ISF sequences available to date, place CTFV among Culex-associated flaviviruses, grouping with recently published NS5 partial sequences documented from mosquitoes collected in the Iberian Peninsula, and with Quang Binh virus (isolated in Vietnam) as a close relative. No CTFV sequences were found integrated in their host’s genome using a range of specific PCR primers designed to the prM/E, NS3, and NS5 region.