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.

Levels of mate recognition within and between two Drosophila species and their hybrids

If sexual selection is to result in speciation, traits involved in mate choice within species need to be capable of producing sexual isolation between species. We investigated the association between mate choice and sexual isolation using interspecific hybrids between two sibling species, Drosophila serrata and Drosophila birchii. A perfuming experiment demonstrated that olfaction was involved in the sexual isolation between the two species. A quantitative genetic analysis using 30 populations of hybrids between the two species indicated that mating success in hybrid individuals was predominately determined by cuticular hydrocarbons; the average genetic correlation between mating success and cuticular hydrocarbon profile was 0.84, and in some instances exceeded 0.95. Multivariate analysis of the cuticular hydrocarbons of the two species revealed that there were three independent blends of cuticular hydrocarbons that separated three levels of organization: species, sex, and sex within species. The hydrocarbons used by hybrids in mate choice included those that separated the two species, demonstrating that species-specific characters may be used in mate choice within populations. The interspecific reciprocal cross had a major effect on which cuticular hydrocarbons were associated with mating success, indicating that the expression of the cuticular hydrocarbons was strongly sex linked.

Development and evaluation of a species diagnostic polymerase chain reaction-restriction fragment-length polymorphism procedure for cryptic members of the Culex sitiens (Diptera : Culicidae) subgroup in Australia and the southwest Pacific

Members of the Culex sitiens subgroup are important vectors of arboviruses, including Japanese encephalitis virus, Murray Valley encephalitis virus and Ross River virus. Of the eight described species, Cx. annulirostris Skuse, Cx. sitiens Wiedemann, and Cx. palpalis Taylor appear to be the most abundant and widespread throughout northern Australia and Papua New Guinea (PNG). Recent investigations using allozymes have shown this subgroup to contain cryptic species that possess overlapping adult morphology. We report the development of a polymerase chain reaction-restriction fragment-length polymorphism (PCR-RFLP) procedure that reliably separates these three species. This procedure utilizes the sequence variation in the ribosomal DNA ITS1 and demonstrates species-specific PCR-RFLP profiles from both colony and field collected material. Assessment of the consistency of this procedure was undertaken on mosquitoes sampled from a wide geographic area including Australia, PNG, and the Solomon Islands. Overlapping adult morphology was observed for Cx. annulirostris and Cx. palpalis in both northern Queensland and PNG and for all three species at one site in northwest Queensland.

Mating behaviour and alternative oviposition sites for male eggs in the heteronomous hyperparasitoid Coccophagus gurneyi Compere (Hymenoptera : Aphelinidae)

The aphelinid parasitoid Coccophagus gurneyi Compere has unusual sex-related host relationships. Females are diploid and develop internally within mealybugs Pseudococcus calceolariae (Maskell). Males, in contrast, are haploid and hyperparasitic, developing on primary parasitoid larvae within the mealybugs. Furthermore, males have been claimed to be capable of either internal or external development, depending on the precise site of deposition of the haploid egg. This diversity of developmental pathways could indicate the existence of a sibling-species complex. We therefore quantified the mating and ovipositional behaviour of C. gurneyi, for comparison with that of an undescribed sibling species. We also checked whether the females deposit male eggs in alternative sites. The pattern of mating was found to be typical of mating behaviour in Coccophagus spp. and was consistent among all mating pairs, suggesting that the colony comprised one species. Further, the mating behaviour was significantly different from that of the undescribed sibling species. The site of male egg deposition varied and is apparently dictated by two factors; whether the mealybug is parasitised and, if so, the size of the parasitoid it contains. If the mealybugs were unparasitised or if the parasitoids within the mealybugs were small (< 0.53 mm), male eggs were deposited within the mealybug haemocoel. If the parasitoids were large (> 1.05 mm), male eggs were deposited within the parasitoids. These results support the claim of alternate host relationships and developmental pathways within males of C. gurneyi.

Larval diet affects number of femoral 'brush' scales in male Helicoverpa punctigera moths (Lepidoptera : Noctuidae)

Males of Helicoverpa punctigera (Wallengren) show considerable variation in the number of femoral scales on the prothoracic legs. Such intraspecific variation in adult morphology could indicate the presence of undetected sibling species, or it may be related to larval diet. Helicoverpa putactigera is polyphagous, and different host plant species are likely to represent diets of different quality. Femoral lengths and the numbers of femoral scales on the prothoracic legs were therefore determined from: (i) individuals that had been collected as larvae from various host species in the field; and (ii) individuals that had been laboratory-reared, in split-family tests, on different diets, namely cotton, lucerne, sowthistle and artificial diet. Host plant species (and therefore presumably diet quality) influenced femoral length of H. punctigera males and, perhaps in conjunction with this, the number of femoral scales on the fore leg. The rearing experiment indicated, in addition, that the effect of host plant quality varies with larval stage, and that the pattern of this variation across the immature stages is dependent on host plant species. The recorded variation in the morphology of field-collected H. punctigera males is therefore most readily explained as a consequence of different individuals developing (at least for most of their larval life) on different host plant species, with diet quality varying significantly with species. The relevance of these results for insect developmental studies and evolutionary interpretations of host relationships is outlined.

Distribution and evolution of the Anopheles punctulatus group (Diptera : Culicidae) in Australia and Papua New Guinea

The members of the Anopheles punctulatus group are major vectors of malaria and Bancroftian filariasis in the southwest Pacific region. The group is comprised of 12 cryptic species that require DNA-based tools for species identification. From 1984 to 1998 surveys were carried out in northern Australia, Papua New Guinea and on islands in the southwest Pacific to determine the distribution of the A. punctulatus group. The results of these surveys have now been completed and have generated distribution data from more than 1500 localities through this region. Within this region several climatic and geographical barriers were identified that restricted species distribution and gene flow between geographic populations. This information was further assessed in light of a molecular phylogeny derived from the ssrDNA (18S). Subsequently, hypotheses have been generated on the evolution and distribution of the group so that future field and laboratory studies may be approached more systematically. This study suggested that the ability for widespread dispersal was found to have appeared independently in species that show niche-specific habitat preference (Anopheles farauti s.s. and A. punctulatus) and conversely in species that showed diversity in their larval habitat (Anopheles farauti 2). Adaptation to the monsoonal climate of northern Australia and southwest Papua New Guinea was found to have appeared independently in A. farauti s.s., A. farauti 2 and Anopheles farauti 3. Shared or synapomorphic characters were identified as saltwater tolerance (A. farauti s.s. and Anopheles farauti 7) and elevational affinities above 1500 m (Anopheles farauti 5, Anopheles farauti 6 and A. farauti 2). (C) 2002 Australian Society for Parasitology Inc. Published by Elsevier Science Ltd. All rights reserved.

Cuticular hydrocarbons of Drosophila birchii and D-serrata: Identification and role in mate choice in D-serrata

The cuticular hydrocarbon compositions of two sympatric species of Australian Drosophila in the montium subgroup of the melanogaster group that use cuticular hydrocarbons in mate recognition have been characterized. Drosophila birchii has 34 components in greater than trace amounts, with a carbon number range of C-20 to C-33. Drosophila serrata has 21 components above trace level and a carbon number range of C-24 to C-31. These two species share eight hydrocarbon components, with all but two of them being monoenes. For both species, the (Z)-9-monoenes are the predominant positional isomer. The hydrocarbons of D. birchii are n-alkanes, n-alkenes (Z)-5-, (Z)-7-, (Z)-9-, and (Z)-11-), low to trace levels of homologous (Z,Z)-7,11- and (Z,Z)-9,13-dienes; and trace amounts of (Z,Z)-5,9- C-25:2, a major component of D. serrata. Only one methyl branched hydrocarbon was detected (2-methyl C-28), and it occurred at very low levels. The hydrocarbons of D. serrata are dominated by a homologous series of (Z,Z)-5,9-dienes, and notably, are characterized by the apparent absence of n-alkanes. Homologous series of (Z)-5-, (Z)-7-, and (Z)-9- alkenes are also present in D. serrata as well as 2-methyl alkanes. Drosophila serrata females display strong directional mate choice based on male cuticular hydrocarbons and prefer D. serrata males with higher relative abundances of the 2-methyl alkanes, but lower relative abundances of (Z,Z)-5,9- C-24:2 and (Z)-9-C-25:1.

A morphological study of the Anopheles punctulatus group (Diptera: Culicidae) in the Solomon Islands, with a description of Anopheles (Cellia) irenicus Schmidt, sp.n.

A description of Anopheles (Cellia) irenicus Schmidt, sp.n. (formerly A. farauti No. 7) is provided. This species is one of six recorded from the Solomon Islands within the A. punctulatus group, which contains the major vectors of the causative agents of malaria and lymphatic filariasis in the southwest Pacific. Morphological markers are described for adult females, fourth-instar larvae and pupae that identify most specimens of A. irenicus. Keys are presented to distinguish members of the A. punctulatus group in the Solomon Islands.

A rapid single-step multiplex method for discriminating between Trichogramma (Hymenoptera: Trichogrammatidae) species in Australia

Inaccurate species identification confounds insect ecological studies. Examining aspects of Trichogramma ecology pertinent to the novel insect resistance management strategy for future transgenic cotton, Gossypium hirsutum L., production in the Ord River Irrigation Area (ORIA) of Western Australia required accurate differentiation between morphologically similar Trichogramma species. Established molecular diagnostic methods for Trichogramma identification use species-specific sequence difference in the internal transcribed spacer (ITS)-2 chromosomal region; yet, difficulties arise discerning polymerase chain reaction (PCR) fragments of similar base pair length by gel electrophoresis. This necessitates the restriction enzyme digestion of PCR-amplified ITS-2 fragments to readily differentiate Trichogramma australicum Girault and Trichogramma pretiosum Riley. To overcome the time and expense associated with a two-step diagnostic procedure, we developed a “one-step” multiplex PCR technique using species-specific primers designed to the ITS-2 region. This approach allowed for a high-throughput analysis of samples as part of ongoing ecological studies examining Trichogramma biological control potential in the ORIA where these two species occur in sympatry.

Local adaptation and species segregation in two mussel (Mytilus edulis X M. trossulus) hybrid zones

Few marine hybrid zones have been studied extensively, the major exception being the hybrid zone between the mussels Mytilus edulis and M. galloprovincialis in southwestern Europe. Here, we focus on two less studied hybrid zones that also involve Mytilus spp.; M. edulis and M. trossulus are sympatric and hybridize on both western and eastern coasts of the Atlantic Ocean. We review the dynamics of hybridization in these two hybrid zones and evaluate the role of local adaptation for maintaining species boundaries. In Scandinavia, hybridization and gene introgression is so extensive that no individuals with pure M. trossulus genotypes have been found. However, M. trossulus alleles are maintained at high frequencies in the extremely low salinity Baltic Sea for some allozyme genes. A synthesis of reciprocal transplantation experiments between different salinity regimes shows that unlinked Gpi and Pgm alleles change frequency following transplantation, such that post-transplantation allelic composition resembles native populations found in the same salinity. These experiments provide strong evidence for salinity adaptation at Gpi and Pgm (or genes linked to them). In the Canadian Maritimes, pure M. edulis and M. trossulus individuals are abundant, and limited data suggest that M. edulis predominates in low salinity and sheltered conditions, whereas M. trossulus are more abundant on the wave-exposed open coasts. We suggest that these conflicting patterns of species segregation are, in part, caused by local adaptation of Scandinavian M. trossulus to the extremely low salinity Baltic Sea environment.

Separating fluorescent species of aqueous PbS semiconductor nanocrystals using micro-emulsions

Colloidal PbS nanocrystals over-coated with CdS are prepared in aqueous solutions and exhibit strong photoluminescence with two distinct peaks in the visible regime. A photoluminescence peak is observed at 640 nm, which is attributed to the band edge recombination in the PbS nanocrystals, and another peak at 510 nm, which is above the band edge of the PbS nanocrystals. The two PL peaks are isolated by extracting separate species of nanocrystal based upon their surface morphology. Micro-emulsions of hexane:PVA are used to remove the species containing the PL peak at 640 nm from the solution, leaving a singular peak at 510 nm. We show conclusively that the double-peaked structure observed in the photoluminescence spectra of PbS nanocrystals over-coated with CdS is due to the presence of two distinctly different nanocrystal species.

Wolbachia infections of tephritid fruit flies: molecular evidence for five distinct strains in a single host species

Endosymbiotic bacteria of the genus Wolbachia are widespread among arthropods and can induce cytoplasmic incompatibility, thelytokous parthenogenesis, male-killing or feminization in their hosts. Here, we report phylogenetic relationships of Wolbachia in tephritid fruit flies based on wsp gene sequences. We also report, for the first time, five distinct strains of Wolbachia in Bactrocera ascita sp. B. Four of the five Wolbachia strains found in this species were in the same groups as those found in other tephritid fruit flies, suggesting possible horizontal transmission of Wolbachia from other fruit flies into B. ascita sp. B. The unreliability of wsp-specific group primers demonstrated in this study suggests that these primers might be useful only for preliminary identification of Wolbachia. Final determination of group affiliation needs to be verified with wsp sequence data.

Positive selection on an acrosomal sperm protein, M7 lysin, in three species of the mussel genus Mytilus

Marine invertebrate sperm proteins are particularly interesting because they are characterized by positive selection and are likely to be involved in prezyogotic isolation and, thus, speciation. Here, we present the first survey of inter and intraspecific variation of a bivalve sperm protein among a group of species that regularly hybridize in nature. M7 lysin is found in sperm acrosomes of mussels and dissolves the egg vitelline coat, permitting fertilization. We sequenced multiple alleles of the mature protein-coding region of M7 lysin from allopatric populations of mussels in the Mytilus edulis species group (M. edulis, M. galloprovincialis, and M. trossulus). A significant McDonald-Kreitman test showed an excess of fixed amino acid replacing substitutions between species, consistent with positive selection. In addition, Kolmogorov-Smirnov tests showed significant heterogeneity in polymorphism to divergence ratios for both synonymous variation and combined synonymous and non-synonymous variation within M. galloprovincialis. These results indicate that there has been adaptive evolution at M7 lysin and, furthermore, shows that positive selection on sperm proteins can occur even when post-zygotic reproductive isolation is incomplete.

The Indotriradites dolianitii Morphon, a distinctive group of miospore species from the Lower Carboniferous of Gondwana

An emended diagnosis and generic reallocation are proposed for the trilete miospore Indotriradites dolianitii (Daemon, 1974) Loboziak et al., comb. nov. A new species, I. daemonii Loboziak et al., sp. nov., from Viséan strata of Western Gondwana, is erected. These two species, together with I. zosteriformis (Playford et Satterthwait) Playford, 1991 from the Viséan of Australia, belong to a cohesive morphological miospore category, here termed the Indotriradites dolianitii Morphon, which is evidently restricted to the Lower Carboniferous of Gondwana.

Radiizonates arcuatus, a distinctive new miospore species from the Lower Carboniferous of Western Gondwana

A new species of trilete zonate miospores, Radiizonates arcuatus, is established for Lower Carboniferous Western Gondwanan forms hitherto ascribed misguidedly to Radiizonates genuinus (Jushko) Loboziak and Alpern (1978), a Russian Lower Carboniferous species. The latter binomen is, moreover, not a valid combination and is more correctly designated as Vallatisporites genuinus (Jushko) Byvsheva, 1980. R. arcuatus is, from records to date, confined to westerly parts of Gondwana (Brazil, North Africa and Middle East), in which it is characteristic of Early Carboniferous strata, albeit with some slightly older and slightly younger occurrences.