Isolation of microsatellite loci from spotted gum (Corymbia variegata), and cross-species amplification in Corymbia and Eucalyptus

Corymbia variegata (spotted gum) is an important commercial hardwood timber species in Australia. Fourteen polymorphic microsatellite loci were isolated from C. variegata, with 3-5 alleles amplified in three individuals examined. Cross-species amplification in Corymbia was successful for all primer pairs, while 10 loci (71%) were successfully transferred to at least one species in the closely related genus Eucalyptus.

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.

Factors affecting the establishment and dispersal of nymphs of Pristhesancus plagipennis Walker (Hemiptera : Reduviidae) when released onto soybean, cotton and sunflower crops

A problem with augmenting predatory bugs through mass release is the logistical difficulty of delivering nymphs onto the foliage of field crops. In this paper we examine postrelease establishment and dispersal of the nymphs of the predatory bug Pristhesancus plagipennis on soybean, cotton and sunflower in an effort to devise an appropriate strategy for field release. The effects of predator stadia and release rates on field establishment and within-crop-canopy dispersal after hand release were recorded in soybean, cotton and sunflower. Field establishment improved with the release of more-developed nymphs, with third instars providing the most appropriate compromise between field hardiness and rearing cost. Increased nymphal density at the point of release had little effect on nymphal dispersal throughout the crop canopy. The patterns of nymphal dispersal observed on the three crops suggest that crop-canopy architecture may have an effect on the ability of nymphs to spread out postrelease, as nymphs dispersed poorly in cotton and sunflower compared to soybean. To overcome poor dispersal of nymphs after release, a mechanical release method, where nymphs were mixed with vermiculite and delivered onto a target crop through a spinning disk fertiliser spreader, was tested, and provided similar nymph establishment rates and dispersal patterns as releasing nymphs individually by hand. The implications of nymph dispersal and field hardiness in regard to inundative field release techniques are discussed.

Wolbachia density and virulence attenuation after transfer into a novel host

The factors that control replication rate of the intracellular bacterium Wolbachia pipientis in its insect hosts are unknown and difficult to explore, given the complex interaction of symbiont and host genotypes. Using a strain of Wolbachia that is known to over-replicate and shorten the lifespan of its Drosophila melanogaster host, we have tracked the evolution of replication control in both somatic and reproductive tissues in a novel host/Wolbachia association. After transinfection (the transfer of a Wolbachia strain into a different species) of the over-replicating Wolbachia popcorn strain from D. metanogaster to Drosophila simulans, we demonstrated that initial high densities in the ovaries were in excess of what was required for perfect maternal transmission, and were likely causing reductions in reproductive fitness. Both densities and fitness costs associated with ovary infection rapidly declined in the generations after transinfection. The early death effect in D. simulans attenuated only slightly and was comparable to that induced in D. metanogaster. This study reveals a strong host involvement in Wolbachia replication rates, the independence of density control responses in different tissues, and the strength of natural selection acting on reproductive fitness.

First record of the mite Hirstiella diolii Baker (Prostigmata : Pterygosomatidae) from Australia, with a review of mites found on Australian lizards

We report the First occurrence in Australia of an exotic mite parasite of lizards, Hirstiella diolii Baker (Prostigmata, Pterygosomatidae), and its association with iguanas at the Taronga Zoo in Sydney. We also report on the spread of the exotic snake mite Ophionyssus natricis (Gervais) to native populations of skinks, provide a key to the mites associated with Australian lizards and review how to distinguish chiggers from pterygosomatid mites.

Mating aggregations and mating success in the flower thrips, Frankliniella schultzei (Thysanoptera: Thripidae), and a possible role for pheromones

Aggregations of Frankliniella schultzei males were observed on the corollas of Hibiscus rosasinensis and Gossypium hirsutum flowers in southeast Queensland. Aggregations were seen only on the upper surfaces of corollas but may have occurred on other flower parts, which were hidden from view. Conspecific females entered aggregations and a small proportion of them mated [18% (n = 163), H. rosasinensis; 30% (n = 181), G. hirsutum]. Most females (87 and 72%, respectively) that did not mate in aggregations walked to other flower parts. Behavior was difficult to observe on these parts, but mating was sometimes observed there. The number of females that landed within aggregations on the upper surfaces of both H. rosasinensis and G. hirsutum corollas was highly correlated with the number of males (r = 0.88, r = 0.93, respectively; P < 0.001). Significantly more mating pairs were observed in high-density aggregations (mean +/- SE, 1.10 +/- 0.22 and 4.44 +/- 0.48, respectively) than in low-density aggregations (0.37 +/- 0.11 and 1.67 +/- 0.29, respectively) (P < 0.05) on flowers of both species. More F. schultzei females were attracted to sticky traps baited with live conspecific males set among flowering Ipomoea indica (mean +/- SE, 8.83 +/- 0.32) and G. hirsutum (10.90 +/- 0.79) plants than to control traps (0.10 +/- 0.05 and 0.70 +/- 0.25, respectively)( P < 0.05), presumably in response to male-produced pheromones. Significantly more females were attracted to traps with high male densities than to traps with low densities. We found no statistical evidence that aggregation size influenced mating success (proportion males that mated). Mating success, however, should be evaluated with respect to mating on all flower parts and not just the upper surfaces of corollas. The results of this study constitute the first behavioral evidence for an attractant sex pheromone in thrips.

Mating behavior of Trissolcus basalis (Wollaston) (Hymenoptera : Scelionidae): Potential for outbreeding in a predominantly inbreeding species

The mating behavior of the quasi-gregarious egg parasitoid Trissolcus basalis (Wollaston) was investigated under field conditions. Trissolcus basalis has female-biased sex ratios and is a protandrous species, with males emerging 1-2 days before females. Males competed aggressively for control of the egg mass, with one male assuming dominance and control of the egg mass, although changes in dominance occurred at least once on each egg mass observed. Typical mating behavior involved the dominant male mating his sisters immediately upon their emergence from the egg mass. These behaviors are characteristic of an inbreeding species that manifests local mate competition. However, several aspects of the mating behavior of T. basalis are inconsistent with that of an inbreeding species. Over 18% of emerging females were not mated by the dominant male upon emergence, 13% of females were not observed to be mated at all and may have left their natal site as virgins, 25% of females were mated multiple times and sometimes by multiple males, females remained near the natal site for up to several hours after emergence before emigrating, and males dispersed away from the natal site during female emergence. Trissolcus basalis may be a predominantly inbreeding species but its emergence and mating behavior suggest that low-frequency outbreeding is also likely to occur.

Development and survival of the diamondback moth (Lepidoptera : Plutellidae) at constant and alternating temperatures

Survival and development time from egg to adult emergence of the diamondback moth, Plutella xylostella (L.), were determined at 19 constant and 14 alternating temperature regimes from 4 to 40degreesC. Plutella xylostella developed successfully front egg to adult emergence at constant temperatures from 8 to 32degreesC. At temperatures from 4 to 6degreesC or from 34 to 40degreesC, partial or complete development of individual stages or instars was possible, with third and fourth instars having the widest temperature limits. The insect developed successfully from egg to adult emergence under alternating regimes including temperatures as low as 4degreesC or as high as 38degreesC. The degree-day model, the logistic equation, and the Wang model were used to describe the relationships between temperature and development rate at both constant and alternating temperatures. The degree-day model described the relationships well from 10 to 30degreesC. The logistic equation and the Wang model fit the data well at temperatures 32degreesC. Under alternating regimes, all three models gave good simulations of development in the mid-temperature range, but only the logistic equation gave close simulations in the low temperature range, and none gave close or consistent simulations in the high temperature range. The distribution of development time was described satisfactorily by a Weibull function. These rate and time distribution functions provide tools for simulating population development of P. xylostella over a wide range of temperature conditions.

Specificity of PCR and in situ hybridization assays designed for detection of Marteilia sydneyi and M-refringens

Primers and DNA probes designed for use in the specific detection of the paramyxean parasites Marteilia sydneyi and Marteilia refringens were tested for their potential to cross-react with closely related species in Polymerase Chain Reaction (PCR) and in situ hybridization. PCR primers and a DNA probe designed within the ITS1 rRNA of M. sydneyi were specific for M. sydneyi when compared with related species of Marteilia and Marteilioides. PCR primers designed within the 18S rRNA of M. refringens were specific in the detection of this species in PCR while a DNA probe (named Smart 2) designed on the same gene cross-reacted with M. sydneyi in tissue sections of Saccostrea glomerata as well as Marteilioides sp. infecting Striostrea mytiloides. Though not species specific, the Smart 2 probe provided a stronger signal in detection of all stages of M. sydneyi than the ITS1 probe. The ITS probe is proposed for use as a confirmatory diagnostic too] for M. sydneyi.

Detection of the initial infective stages of the protozoan parasite Marteilia sydneyi in Saccostrea glomerata and their development through to sporogenesis

DNA probes were used in in situ hybridisation on histological sections of oysters exposed for defined intervals to Marteilia sydneyi infection to reveal the early development of the parasite in the oyster host, Saccostrea glomerata. The initial infective stages enter through the palps and gills whereupon extrasporogonic proliferation results in the liberation of cells into surrounding connective tissue and haemolymph spaces. Following systemic dissemination, the parasite infiltrates the digestive gland and becomes established as a nurse cell beneath the epithelial cells ill a digestive tubule. Here, cell-within-cell proliferation results in the eventual liberation of daughter cells from the nurse cell into spaces between adjacent epithelial cells. None of these stages had previously been described. Proliferation is associated with host responses, including haemocytic infiltration of the connective tissue and diapedesis across tubule epithelia. The responses cease as sporogenesis begins. (C) 2002 Australian Society for Parasitology Inc. Published by Elsevier Science Ltd. All rights reserved.

Analysis of the sequence and expression of a second putative acetylcholinesterase cDNA from organophosphate-susceptible and organophosphate-resistant cattle ticks

The cattle tick, Boophilus microplus, is a major pest of cattle in Australia, Central and South America, and parts of Africa and Asia. Control of ticks with organophosphates (OPs) and carbamates, which target acetylcholinesterases (AChE), led to evolution of resistance to these pesticides. Alleles at the locus studied here, AChE2, from OP-susceptible female ticks from Australia and Mexico differed at 46 of 1689 nucleotide positions (20 putative amino acid differences) whereas alleles from three strains of OP-resistant ticks from Australia differed with the allele from the Australian susceptible ticks at six to 13 nucleotide positions (three to six putative amino acid differences). However, the role, if any, of these polymorphisms in the OP-resistance phenotype is unknown. Certainly none of the polymorphisms correspond to sites in ACK that are involved in catalysis or binding of acetylcholine in other organisms. Both of the AChE loci of B. microplus, AChE1 and AChE2, are apparently expressed in synganglia; AChE1 is also expressed in salivary glands and ovaries, in OP-susceptible and OP-resistant ticks. This seems to contradict studies of enzyme kinetics, which indicated that only one form of AChE was present in the synganglia, the site of the action of OPs, in this species of tick. (C) 2002 Elsevier Science Ltd. All rights reserved.

Synonymy of Perkinsus olseni Lester & Davis, 1981 and Perkinsus atlanticus Azevedo, 1989 and an update on the phylogenetic position of the genus Perkinsus

Sequences of the rRNA nontranscribed spacer (NTS) were determined for six isolates of Perkinsus olseni, seven isolates of Perkinsus sp. from Anadara trapezia and one isolate of Perkinsus sp. from Austrovenus stutchburyi. These sequences were compared with previously published NTS sequences for R atlanticus, P. marinus and P. andrewsi. Consensus sequences for Perkinsus olseni, the Perkinsus isolates and P. atlanticus were approximately 98-99% similar to each other but only 65-79% similar to P. marinus and P. andrewsi sequences. Some individual P. olseni sequences were less similar to each other (97.4%) than they were to P. atlanticus sequences (97.8-98.2%), therefore NTS provides further evidence that P. atlanticus, P. olseni, Perkinsus sp. from Anadara trapezia and Perkinsus sp. from Austrovenus stutchburyi are conspecific. We propose that P. atlanticus be synonymised with P. olseni Lester & Davis, 1981 which has taxonomic priority, and that Perkinsus sp. from Anadara trapezia and Perkinsus sp. from Austrovenus stutchburyi belong to R olseni sensu lato as well. A phylogenetic analysis of SSU rDNA, incorporating recently published Perkinsus sequences, supports the placement of the Perkinsus species with Parvilucifera infectans within the Dinoflagellata.

In vivo and in vitro toxicities of pithraj and neem against rice green leafhopper (Nephotettix cincticeps Uhler)

Seed extracts of Aphanamixis polystachya Wall et Parker (pithraj) and Azadirachta indica A. Juss (neem) were evaluated for their in vivo and in vitro toxicity to Nephotettix cincticeps Uhler (rice green leafhopper). Crude extracts from both plants showed toxicity to leafhopper. Among them, the methanol extract of pithraj (MCX) was most toxic and showed 95% mortality effects at 72 h after treatment (HAT), followed by neem (74%). When LD50's were compared, it was found that the neem extract possessed the highest toxicity (LD50 16.59 μg/insect) at 72 HAT. Both the pithraj (MCX) and neem extracts showed their enzyme inhibition effectiveness against rice green leafhopper. The highest inhibition rate (IR) was caused by neem (60%) at the concentration of 2.0 mg/ml, followed by MCX (47%). The lowest IR50 value (0.97 mg/ml) was observed in neem at 30 min.