A test of the ash-free dry weight technique on the developmental stages of Patiriella spp. (Echinodermata : Asteroidea)

Determination of the ash-free dry weight (AFDW) of marine specimens requires samples to be rinsed, soaked, and centrifuged. Problems associated with this technique were examined with the developmental stages of seastar species (Patiriella) with different modes of development. The influence of three rinsing solutions (ammonium formate [AF], filtered seawater [FSW], and reverse osmosis water [RO]) was assessed. The hypothesis that the AFDW technique is a measure of organic material was addressed by drying inorganic salts. Developmental stages of Patiriella calcar rinsed in FSW were twice as heavy as those rinsed in RO or AE indicating that samples should be rinsed in RO or AF before weighing. Soaking treatments had a significant effect on the AFDW of samples of P. calcar (planktonic developer), indicating that the rinsing period should be brief. Zygotes of Patiriella re gularis (planktonic developer) were significantly heavier than ova or gastrulae, regardless of treatment. In contrast, there were no significant differences in the AFDW of any stages or treatments of Patiriella exigua (benthic developer). This may be due to the presence of a modified fertilization envelope, which protects these benthic embryos. Inorganic salts with water of crystallization and FSW lost 20-75% and 14% of their dry weight, respectively, after ashing. We propose that salt ions may retain water, which does not evaporate during drying but is lost during ashing, resulting in the overestimation of sample AFDW. If a similar process occurs in the developmental stages of marine invertebrates, changes in the intracellular ionic composition through development may result in inaccurate estimates of biomass.

Field test of a model of migration of moths (Lepidoptera : Noctuidae) in inland Australia

A migration of Helicoverpa punctigera (Wallengren), Heliothis punctifera (Walker) and Agrotis munda Walker was tracked from Cameron Corner (29degrees00'S, 141degrees00'E) in inland Australia to the Wilcannia region, approximately 400 km to the south-east. A relatively isolated source population was located using a distribution model to predict winter breeding, and confirmed by surveys using sweep netting for larvae. When a synoptic weather pattern likely to produce suitable conditions for migration developed, moths were trapped in the source region. The next morning a simulation model of migration using wind-field data generated by a numerical weather-prediction model was run. Surveys using sweep netting for larvae, trapping and flush counts were then conducted in and around the predicted moth fallout area, approximately 400 km to the south-east. Pollen carried on the probosces of moths caught in this area was compared with that on moths caught in the source area. The survey data and pollen comparisons provided evidence that migration had occurred, and that the migration model gave accurate estimation of the fallout region. The ecological and economic implications of such migrations are discussed.

Laboratory evaluation of two native fishes from tropical North Queensland as biological control agents of subterranean Aedes aegypti

The ability of 2 freshwater fishes, eastern rainbow fish Melanotaenia splendida splendida and fly-specked hardyhead Craterocephalus stercusmuscarum stercusmuscarum. native to North Queensland to prey on immature Aedes aegypti was evaluated under laboratory conditions. The predation efficiency of the 2 species was compared to the exotic guppy, Poecilia reticulata, which is commonly used as a biological control agent of mosquito larvae. Of the 3 fish species tested, M. s. splendida was shown to be the most promising agent for the biological control of Ae. aegypti that breed in wells. Melanotaenia s. splendida consumed significantly greater numbers of immature Ae. aegypti than P. reticulata, irrespective of developmental stage or light conditions. Unlike C. s. stercusmuscarum, M, s. splendida could be handled, transported, and kept in captivity for extended periods with negligible mortality. However, M. s. splendida was also an efficient predator of Litoria caerulea tadpoles, a species of native frog found in wells during the dry season. This result may limit the usefulness of M. s. splendida as a biological control agent of well-breeding Ae. aegypti and suggests that predacious copepods, Mesocyclops spp., are more suitable. However, the use of M. s. splendida as a mosquito control agent in containers that are unlikely to support frog populations (e.g., aquaculture tanks and drinking troughs) should be given serious consideration.

The effect of mode of exposure to Beauveria bassiana on conidia acquisition and host mortality of Colorado potato beetle, Leptinotarsa decemlineata

The effects of the mode of exposure of second instar Colorado potato beetles to Beauveria bassiana on conidia acquisition and resulting mortality were investigated in laboratory studies. Larvae sprayed directly with a B, bassiana condial suspension, larvae exposed to B, bassiana-treated foliage, and larvae both sprayed and exposed to treated foliage experienced 76, 34, and 77% mortality, respectively. The total number of conidia and the proportion of germinating conidia were measured over time for four sections of the insect body: the ventral surface of the head (consisting mostly of ventral mouth parts), the ventral abdominal surface, the dorsal abdominal surface, and the legs. From observations at 24 and 36 h posttreatment, mean totals of 161.1 conidia per insect were found on sprayed larvae, 256.1 conidia on larvae exposed only to treated foliage, and 408.3 conidia on larvae both sprayed and exposed to treated foliage, On sprayed larvae, the majority of conidia were found on the dorsal abdominal surface, whereas conidia were predominantly found in the ventral abdominal surface and mouth parts on larvae exposed to treated foliage, Between 24 and 36 h postinoculation the percentage of conidia germinating on sprayed larvae increased slightly from 80 to 84%), On the treated foliage, the percentage of germinated conidia on larvae increased from 35% at 24 h to 50% at 36 h posttreatment, Conidia germination on sprayed larvae on treated foliage was 65% at 24 h and 75% at 36 h posttreatment, It is likely that the gradual acquisition of conidia derived from the continuous exposure to B. bassiana inoculum on the foliar surface was responsible for the increase in germination over time on larvae exposed to treated foliage, The density and germination of conidia were observed 0, 4, 8, 12, 16, 20, and 24 h after being sprayed with or dipped in conidia suspensions or exposing insects to contaminated foliage, Conidia germinated twice as fast on sprayed insects as with any other treatment within the first 12 h, This faster germination may be due to the pressure of the sprayer enhancing conidial lodging on cuticular surfaces. (C) 2001 Academic Press.

Descriptions of the Anopheles (Cellia) farauti complex of sibling species (Diptera : Culicidae) in Australia

Descriptions of the three sibling species of the Anopheles farauti complex in Australia, A. farauti Laveran (formerly A. farauti No. 1), A. hinesorum Schmidt sp.n. (formerly A. farauti No. 2) and A. torresiensis Schmidt sp.n. (formerly A. farauti No. 3) are provided. These species form a part of the punctulatus group, which contains the major malaria vectors in the southwest Pacific. Morphological markers are described for adult females, fourth instar larvae and pupae which identify most specimens, and are presented in keys.

Potential of Cactoblastis cactorum as a vector for fungi pathogenic to pricklypear, Opuntia inermis

In Australia, fungi associated with larvae of the biological control agent Cactoblastis cactorum may contribute to the control of the exotic weed pricklypear (Opuntia inermis), C, cactorum larvae were assessed for their ability to vector pathogenic fungi into O, inermis by the infestation of larvae with fungal suspensions. Six fungal isolates caused disease after being carried into the host on external surfaces of larvae, and propagules of one isolate (UQ5109) initiated disease after being transferred from the cladode epidermis into the host by larvae feeding on the plant. Scanning electron microscopy revealed extensive hyphal growth on the external surfaces of larvae infested with several of the isolates. Fungi isolated from field-grown O, inermis cladodes were tested for pathogenicity to this plant in an in vivo plant assay. In total, 152 isolates were screened, 22 of which infected the host in pathogenicity tests. Only 1 (UQ5115) infected undamaged host tissue, whereas the remainder required the host to be wounded before infection could proceed. The majority of isolates were only weakly pathogenic, even when inoculated via wounds, suggesting that most were either saprophytes or weak parasites. This study demonstrates that it is possible for larvae of C, cactorum to transmit fungal pathogens into O, inermis tissue and it has provided a sound basis for future field work to determine the contribution that fungi make to the control of O. inermis, (C) 2001 Academic Press.

Evaluation of synergistic interactions between the Colorado potato beetle (Coleoptera : Chrysomelidae) pathogen Beauveria bassiana and the insecticides, imidacloprid, and cyromazine

Laboratory studies investigated the interaction between the fungal entomopathogen Beauveria bassiana (Balsamo) Vuillemin and sublethal doses of the insecticides imidacloprid and cyromazine when applied to larvae of the Colorado potato beetle, Leptinotarsa decemlinenta (Say). When second instars were fed potato leaf discs treated with sublethal doses of imidacloprid and a range of doses of B. bassiana, a synergistic action was demonstrated. Similar results were observed when larvae were sprayed directly with B. bassiana conidia and immediately fed leaf discs treated with imidacloprid. No synergistic interaction was detected when larvae were fed leaf discs treated with sublethal doses of imidacloprid 24 h after application of R. bassiana conidia to larvae. However, a synergistic interaction was detected when larvae were fed leaf discs treated with imidacloprid and sprayed with B, bassiana conidia 24 h later. Although sublethal doses of both imidacloprid and the triazine insect growth regulator (IGR) cyromazine prolonged the duration of the second instar, only imidacloprid interacted with B. bassiana to produce a synergistic response in larval mortality. In leaf consumption studies, the highest dose of B, bassiana tested promoted feeding in inoculated second instars. Feeding was inhibited when larvae were fed foliage treated with sublethal doses of imidacloprid and significantly reduced when fed foliage treated with a sublethal dose of cyromazine. Starvation of larvae for 24 h immediately after B. bassiana treatment produced a similar result to the combined treatment of B. bassiana and imidacloprid and increased the level of mycosis when compared with B. bassiana controls. Imidacloprid treatment affected neither the rate of germination of B. bassiana conidia on the insect cuticle nor the rate at which conidia were removed from the integument after application. The statistical analysis used to detect synergism and the possible role of starvation-induced stress factors underlying the observed synergistic interactions are discussed.

Horizontal transmission of entomopathogenic fungi by the diamondback moth

The relative potential of the pathogenic fungi Beauveria bassiana and Zoophthora radicans for use as autodisseminated biological control agents of the diamondback moth (Plutella xylostella) was compared. The LC50 of B. bassiana conidia to third instar larvae was 499 conidia/mm(2) of leaf surface and individual cadavers of mycosed fourth instar larvae yielded a mean of 67.5 X 10(6) (+/- 7.5 x 10(6)) conidia. All concentrations of B. bassiana tested in inoculation chambers (0.24, 2.4, and 6.2 mug/mm(2)) induced 100% mortality in adult male moths within 7 days. The times to death and sporulation were concentration and exposure duration dependent. A standard procedure for inoculating male moths resulted in > 85% mortality from Z. radicans and > 93% mortality from B. bassiana. Pairing of inoculated males with clean moths of both sexes yielded higher rates of passive transmission of B. bassiana than Z. radicans, but there was no evidence for sexual transmission of either pathogen. Similarly, B. bassiana was more effectively transmitted from inoculated male moths to larvae foraging on whole plants. Single sporulating cadavers producing B. bassiana or Z. radicans conidia placed on plants infested with larvae resulted in a similar rate of transmission for both pathogens. However, an increase of the density of sporulating cadavers from one to three/plant increased Z. radicans transmission (greater than fourfold) but had no effect on B. bassiana transmission. Simultaneous inoculations of larvae with conidia of both fungi reduced the mortality induced by each pathogen, the reduction being most acute for B. bassiana-induced mortality. Inoculation of adults with both fungi showed that, at concentrations required for effective passive transmission to larvae, B. bassiana severely inhibited Z. radicans mycosis in adults. (C) 2001 Academic Press.

Cytological basis of photoresponsive behavior in a sponge larva

Ontogenetic changes in the photoresponse of larvae from the demosponge Reneira sp. were studied by analyzing the swimming paths of individual larvae exposed to diffuse white light. Larvae swam upward upon release from the adult, but were negatively phototactic until at least 12 hours after release. The larval photoreceptors are presumed to be a posterior ring of columnar monociliated epithelial cells that possess 120-mum-long cilia and pigment-filled protrusions. A sudden increase in light intensity caused these cilia to become rigidly straight. If the light intensity remained high, the cilia gradually bent over the pigmented vesicles in the adjacent cytoplasm, and thus covered one entire pole of the larva. The response was reversed upon a sudden decrease in light intensity. The ciliated cells were sensitive to changes in light intensity in larvae of all ages. This response is similar to the shadow response in tunicate larvae or the shading of the photoreceptor in Euglena and is postulated to allow the larvae to steer away from brighter light to darker areas, such as under coral rubble-the preferred site of the adult sponge on the reef flat. In the absence of a coordinating system in cellular sponges, the spatial organization and autonomous behavior of the pigmented posterior cells control the rapid responses to light shown by these larvae.

Developmental expression of a class IV POU gene in the gastropod Haliotis asinina supports a conserved role in sensory cell development in bilaterians

POU-IV genes regulate neuronal development in a number of deuterostomes (chordates) and ecdysozoans (arthropods and nematodes). Currently their function and expression in the third bilaterian clade, the Lophotrochozoa, comprising molluscs, annelids and. their affiliates, is unclear. Herein we characterise the developmental expression of HasPOU-IV in the gastropod mollusc, Haliotis asinina. The POU-IV gene is transiently expressed in I I distinct larval territories during the first 3 days of development. HasPOU-IV is first expressed in sets of ventral epidermal cells in the newly hatched trochophore larvae. As larval morphogenesis proceeds, we observe HasPOU-IV transcripts in cells that putatively form a range of sensory systems including chemo- and mechanosensory cells in the foot, cephalic tentacles, the ctenidia. the geosensory statocyst and the eyes. By comparing HasPOU-IV expression with POU-IV genes in other bilaterians we infer that this class of POU-domain genes had an ancestral role in regulating sensory cell development.

Ecological regulation of development: Induction of marine invertebrate metamorphosis

In the marine environment a wide range of invertebrates have a pelagobenthic lifecycle that includes planktonic larval and benthic adult phases. Transition between these morphologically and ecologically distinct phases typically occurs when the developmentally competent larva comes into contact with a species-specific environmental cue. This cue acts as a morphogenetic signal that induces the completion of the postlarval/juvenile/adult developmental program at metamorphosis. The development of competence often occurs hours to days after the larva is morphologically mature. In the non-feeding - lecithotrophic - larvae of the ascidian Herdmania curvata and the gastropod mollusc Haliotis asinina, gene expression patterns in pre-competent and competent stages are markedly different, reflecting the different developmental states of these larval stages. For example, the expression of Hemps, an EGF-like signalling peptide required for the induction of Herdmania metamorphosis, increases in competent larvae. Induction of settlement and metamorphosis results in further changes in developmental gene expression, which apparently is necessary for the complete transformation of the larval body plan into the adult form.

Embryogenesis and metamorphosis in a haplosclerid demosponge: gastrulation and transdifferentiation of larval ciliated cells to choanocytes

Early development and metamorphosis of Reniera sp., a haplosclerid demosponge, have been examined to determine how gastrulation occurs in this species, and whether there is an inversion of the primary germ layers at metamorphosis. Embryogenesis occurs by unequal cleavage of blastomeres to form a solid blastula consisting micro- and macromeres; multipolar migration of the micromeres to the surface of the embryo results in a bi-layered embryo and is interpreted as gastrulation. Polarity of the embryo is determined by the movement of pigment-containing micromeres to one pole of the embryo; this pole later becomes the posterior pole of the swimming larva. The bi-layered larva has a fully differentiated monociliated outer cell layer, and a solid interior of various cell types surrounded by dense collagen. The pigmented cells at the posterior pole give rise to long cilia that are capable of responding to environmental stimuli. Larvae settle on their anterior pole. Fluorescent labeling of the monociliated outer cell layer with a cell-lineage marker (CMFDA) demonstrates that the monociliated cells resorb their cilia, migrate inwards, and transdifferentiate into the choanocytes of the juvenile sponge, and into other amoeboid cells. The development of the flagellated choanocytes and other cells in the juvenile from the monociliated outer layer of this sponge's larva is interpreted as the dedifferentiation of fully differentiated larval cells-a process seen during the metamorphosis of other ciliated invertebrate larvae-not as inversion of the primary germ layers. These results suggest that the sequences of development in this haplosclerid demosponge are not very different than those observed in many cnidarians.

A sponge allelochemical induces ascidian settlement but inhibits metamorphosis

Secondary metabolites synthesised by sessile invertebrates appear to play a role in creating and maintaining space on hard substrata by repelling competitors. In this study, we investigated the responses of the larvae of the ascidian Herdmania curvata to haliclonacyclamine A (HA), the major component of a suite of cytotoxic alkaloids extracted from the sponge Haliclona sp. 628. Both Haliclona sp. 628 and Herdmania curvata inhabit the crest and slope of Heron Island Reef. High rates of settlement were induced in competent H. curvata larvae by a range of concentrations of HA, all lower than that naturally occurring in the sponge. HA did not induce precompetent larvae to settle. Although early metamorphosis of HA-induced larvae was normal, larvae exposed to all but the lowest concentration of HA were developmentally arrested after completion of tail resorption, at about 4 h after the initiation of metamorphosis. These postlarvae underwent extensive cellular necrosis within 24 h. We also demonstrate that the addition of a transcriptional inhibitor, actinomycin D, to larvae also causes inhibition of metamorphosis after tail resorption is completed. Analyses of incorporation of radiolabelled nucleotides to measure levels of transcription during normal development and after the addition of the transcriptional inhibitor indicate that there is a significant burst of transcriptional activity just after tail resorption is completed. Despite inhibiting metamorphosis at the same stage as actinomycin D, HA increases initial rates of RNA synthesis after induction of metamorphosis in a manner similar to that observed in normal postlarvae until the onset of cellular necrosis. We conclude that HA initially induces H. curvata larvae to settle and progress through early metamorphosis possibly by engaging the same pathway as other artificial and environmental cues but subsequently inhibits completion of metamorphosis, resulting in death of the postlarvae. Since HA does not affect overall transcription rates, it appears to disrupt another important developmental process during early metamorphosis.

Stream-bank shade and larval distribution of the Philippine malaria vector Anopheles flavirostris

The principal malaria vector in the Philippines, Anopheles flavirostris (Ludlow) (Diptera: Culicidae), is regarded as 'shade-loving' for its breeding sites, i.e. larval habitats. This long-standing belief, based on circumstantial observations rather than ecological analysis, has guided larval control methods such as 'stream-clearing' or the removal of riparian vegetation, to reduce the local abundance of An. flavirostris . We measured the distribution and abundance of An. flavirostris larvae in relation to canopy vegetation cover along a stream in Quezon Province, the Philippines. Estimates of canopy openness and light measurements were obtained by an approximation method that used simplified assumptions about the sun, and by hemispherical photographs analysed using the program hemiphot(C) . The location of larvae, shade and other landscape features was incorporated into a geographical information system (GIS) analysis. Early larval instars of An. flavirostris were found to be clustered and more often present in shadier sites, whereas abundance was higher in sunnier sites. For later instars, distribution was more evenly dispersed and only weakly related to shade. The best predictor of late-instar larvae was the density of early instars. Distribution and abundance of larvae were related over time (24 days). This pattern indicates favoured areas for oviposition and adult emergence, and may be predictable. Canopy measurements by the approximation method correlated better with larval abundance than hemispherical photography, being economical and practical for field use. Whereas shade or shade-related factors apparently have effects on larval distribution of An. flavirostris , they do not explain it completely. Until more is known about the bionomics of this vector and the efficacy and environmental effects of stream-clearing, we recommend caution in the use of this larval control method.

Augmentation of the assassin bug Pristhesancus plagipennis (Walker) (Hemiptera : Reduviidae) as a biological control agent for Helicoverpa spp. in cotton

Third-instar nymphs of the Australian assassin bug, Pristhesancus plagipennis (Walker), were released into cotton plots at two release densities and two crop growth stages to test their biological control potential. Release rates of 2 and 5 nymphs per metre row resulted in field populations of 0.51 and 1.38 nymphs per metre row, respectively, indicating that over 70% of nymphs died or emigrated within two weeks of release. Effective release rates of 1.38 nymphs per metre row reduced the number of Helicoverpa spp. larvae in the plots for a 7-week period. Crop yields were significantly greater in the plots to which P. plagipennis nymphs were released, with the effective release rate of 1.38 nymphs per metre row providing equivalent yields as insecticide treated plots. The data suggest that P. plagipennis has the capacity to reduce Helicoverpa spp. larvae densities in cotton crops when augmented through inundative release.