Direct and indirect top‐down effects of previous contact with an enemy on the feeding behavior of blowfly larvae

We investigated the addition of a trophic level to a simple food web. Direct and indirect effects caused by the presence of a new species in the food web were quantified by estimating survival and consumption rates on the basal resource. We focused on a blowfly intraguild prey–predator system with various ecological interactions taking place during the larval period. The experiments were designed to set Chrysomya megacephala (Fabricius) (Diptera: Calliphoridae) as the intraguild prey and Chrysomya albiceps (Wiedemann) as the intraguild predator and/or cannibal. The generalist pupal parasitoid Nasonia vitripennis (Walker) (Hymenoptera: Pteromalidae) was introduced into the system during a non‐susceptible life stage of the interacting blowfly species. The cascading parasitoid effects induced behavioral changes in the blowfly larvae, increasing the impact of intraguild predation and cannibalism on blowfly survival. The results suggest that blowfly larvae can change their feeding behavior in response to the presence of a parasitoid.     

Temperature adaptation in Chrysomya megacephala and Chrysomya pinguis,two blow fly species of forensic significance

Chrysomya megacephala (Fabricius) (Diptera: Calliphoridae) is a common and forensically important blow fly species in the Oriental region. However, in the higher mountain regions and on winter days, its habitats are occupied by a closely related species, Chrysomya pinguis (Walker). The resources that the two species employ to survive are very similar and competition between the species may be one of the factors that trigger differentiation of their behaviors. We conducted experiments to examine how these two closely related species may have adapted to different temperature regimes to avoid competition. Several adult and immature parameters were assessed, such as fecundity, locomotor ability, hatching ratio, larval survivorship, and eclosion ratio. Results indicate that species show specific diapause at high temperature (38 °C), larval survivorship of Ch. megacephala was significantly better than that of Ch. pinguis. Conversely, at low temperature (15 °C), adult locomotor ability was better for Ch. pinguis than for Ch. megacephala. The results indicate that the two species may have evolved different temperature adaptation strategies to avoid competition. In mixed‐species larval rearing experiments, competition between Ch. pinguis and Ch. megacephala was observed: at higher temperature (30 °C), the immature performance index of Ch. megacephala was significantly increased when compared to that in single‐species culture, whereas the index of Ch. pinguis was decreased. These data are consistent with the idea that tolerance for higher temperature conditions would allow larvae of Ch. megacephala to gain a competitive advantage over Ch. pinguis in certain habitats. These results may help to explain their current distribution in the environment and provide more biological information on these forensically important species.     

Dispersal and Predation Behavior in Larvae of <Emphasis Type="Italic">Chrysomya albiceps</Emphasis> and <Emphasis Type="Italic">Chrysomya megacephala</Emphasis> (Diptera: Calliphoridae)

Chrysomya albiceps and Chrysomya megacephala are exotic blowfly species known by producing myiasis in humans and other animals and by transmitting pathogens mechanically. C. albiceps stand out by being a facultative predator of other dipteran larvae. In this paper we investigated the influence of larval predation on the dispersal of larvae of C. albiceps and C. megacephala single and double species for three photophases. An experimental acrylic channel graduated and covered with wood shavings was used to observe the larval dispersal. The results showed that C. albiceps attacks C. megacephala larvae during dispersal and keeps an aggregated pattern close to the release point, in single and double species, independently of the different photophases. Chrysomya megacephala single species exhibited the same pattern, but in double species this was changed to a random distribution.     

Polytene chromosome sof monogenic and amphogenic Chrysomya species (Calliphoridae,diptera): analysis of banding patterns and in situ hybridization with Drosophila sex determining gene sequences

Standard maps for the five banded polytene chromosomes found in trichogen cell nuclei of the monogenic blowfly Chrysomya rufifacies and the amphogenic Chrysomya pinguis are presented. The chromosomes are highly homologous in the two species; differences in banding patterns are predominantly caused by one pericentric and ten paracentric inversions. In chromosome 5 of the amphogenic Chrysomya phaonis, also analysed in this paper, an additional paracentric inversion was observed. The distribution of species specific inversions indicates that the monogenic C. rufifacies is phylogenetically older than the amphogenic species. The maternal sex realizer locus F'/f on polytene chromosome 5 of C. rufifacies is not associated with a structural heterozygosity. Chromosome pair 6 of C. rufifacies and the sex chromosome pair of C. pinguis are under-replicated in polytene nuclei; they consist of irregular chromatin granules, frequently associated with nucleolus material. Evolution of heteromorphic sex chromosomes in Chrysomya is probably correlated with heterochromatin accumulation. A search for sex determining genes in Chrysomya was initiated using sex determining sequences from Drosophila melanogaster for in situ hybridization. The polytene band 41A1 on chromosome 5 of monogenic and amphogenic Chrysomya species contains sequences homologous to the maternal sex determining gene daughterless (da). Homology to the zygotic gene Sex-lethal (Sxl) of Drosophila is detected in band 39A1 on chromosome 5 of C. rufifacies. The findings reported here are the first evidence for a possible homology between the da gene of Drosophila and the maternal sex realizer F of C. rufifacies. An hypothesis for the evolution of the maternal effect sex determination of C. rufifacies is proposed.Dedicated to Professor Dr. Fritz-Helmut Ullerich on the occasion of his 60th birthday     

Models of development for blowfly sister species Chrysomya chloropyga and Chrysomya putoria

Abstract Developmental curves for the sister species Chrysomya chloropyga (Wiedemann, 1818) and Chrysomya putoria (Wiedemann, 1830) (Diptera: Calliphoridae) were established at eight and 10 different constant temperatures, respectively, using developmental landmarks and body length as measures of age. The thermal summation constants (K) and developmental threshold (D₀) were calculated for five developmental landmarks using a previously described method. Isomorphen and isomegalen diagrams were also constructed for the purpose of estimating postmortem intervals (PMIs). Chrysomya chloropyga had an average developmental threshold value (D₀) of 10.91 °C (standard error [SE] = 0.94 °C, n = 5), significantly lower than that of C. putoria (13.42 °C, SE = 0.45 °C, n = 5) (paired t‐test: t = ? 4.63, d.f. = 8, P < 0.00). Similarly, K values for C. chloropyga were larger than those for C. putoria for all developmental events except onset of the wandering phase. These are the first data that can be used to calculate minimum PMIs and predict population growth of C. chloropyga and C. putoria in Africa; the data indicate that developmental data for one of these species cannot be used as surrogate data for the sister species.     

Foraging behaviour by an intraguild predator blowfly, <Emphasis Type="Italic">Chrysomya albiceps</Emphasis> (Diptera: Calliphoridae)

Optimal foraging theory assumes that predators use different prey types to maximize their rate of energetic gain. Studies focusing on prey preference are important sources of information to understand the foraging dynamics of Chrysomya albiceps. The purpose of this investigation is to determine the influence of larval starvation in C. albiceps on the predation rate of different prey blowfly species and instars under laboratory conditions. Our results suggest that C. albiceps prefers Cochliomyia macellaria larvae to Chrysomya megacephala under non-starvation and starvation conditions. Nevertheless, predators gained more weight consuming C. macellaria. This result suggests that C. albiceps profit more in consuming C. macellaria rather than C. megacephala. The foraging behaviour displayed by C. abiceps on their prey and the consequences for the blowfly community are also discussed.     

Stocking rate and organic waste type affect development of three Chrysomya species and Lucilia sericata (Diptera: Calliphoridae): Implications for bioconversion

Fly larvae can be used effectively to reduce various organic waste types and produce value-added products, including protein as an ingredient in livestock feeds and oil for biodiesel production. However, fly development on different waste types may cause differences in growth rate and the body composition, which can further be influenced by fly species and their stocking rate. This study explored the impact of different waste types (kitchen waste, abattoir waste and swine manure) and larval stocking rate on growth and body composition of four blowfly species, Chrysomya chloropyga (Wiedemann), Chrysomya megacephala (Fabricius), Chrysomya putoria (Wiedemann) and Lucilia sericata (Meigen). First-instar larvae (20, 50 or 100), less than 3 hr old, were placed on 100 g of each waste type. Pre-pupal mass at commencement of post-feeding larval dispersal, time to onset of dispersal, survival and nutrient reserves were determined for each species, stocking rate and waste type. Our results revealed that larvae fed kitchen and abattoir waste had significantly higher dry mass, crude protein and lipid content compared with those fed swine manure. Higher survival rate was observed with increasing larval stocking rate. We provide important information to guide the mass production of high-quality nutrient-rich larvae and recommend C. putoria, which is versatile and effective on a range of waste products, as well as high in protein and lipids. The implications for waste management are discussed.     

Molecular phylogeny of echinometrid sea urchins: more species of Heliocidaris with derived modes of reproduction

Abstract. Pachechinus bajulus is an endemic Australian sea urchin with an unusual mode of brooded larval development. We used mitochondrial and nuclear gene sequences to estimate the phylogenetic relationships among Pachechinus and other Echinometridae, including well‐studied species of Heliocidaris with planktonic development. We found strong evidence for the planktotrophic species Heliocidaris tuberculata as the sister group to a clade of three closely related species in which development is known (Heliocidaris erythrogramma, P. bajulus) or suspected (Pachechinus australiae) to be lecithotrophic. Clade support values and likelihood ratio tests rejected monophyly of Heliocidaris species. The sister group to H. erythrogramma is most likely the two Pachechinus species. We resolve the paraphyly problem by reassigning the Pachechinus species to the genus Heliocidaris (the senior synonym), which has six extant species including Heliocidaris australiae and Heliocidaris bajulus. The phylogeny has potentially important implications for comparative studies of developmental morphology and genetics that have assumed a close sister‐group relationship between H. erythrogramma and H. tuberculata, and highlights the need for such data from H. bajulus and other Heliocidaris species.     

Temperature‐dependant development of Nasonia vitripennis on five forensically important carrion fly species

The effects of temperature and host species on the development of Nasonia vitripennis Walker (Hymenoptera: Pteromalidae), a forensically important parasitoid of carrion flies, were studied under laboratory conditions. Development time of N. vitripennis on five species of Calliphoridae (Diptera), Calliphora albifrontalis Malloch, Calliphora dubia Macquart, Lucilia sericata Meigen, Chrysomya rufifacies Macquart, and Chrysomya megacephala Fabricius, were determined under eight constant temperatures (15, 18, 21, 24, 27, 30, 33, and 36 °C). Thermal requirements for development (developmental thresholds, thermal constant, and optimum temperature) of N. vitripennis in each host species were estimated using linear and nonlinear models. Upper and lower developmental thresholds ranged between 36.6–38.4 and 9.6–11.1 °C, respectively. The optimum temperature for development was estimated at between 30.6 and 31.8 °C. Statistical differences in the development time of N. vitripennis on the various calliphorid host species were evident within all temperature treatments, particularly at the upper and lower temperature range investigated. As such, it is recommended that insect‐based estimates of time since death in forensic investigations relying on parasitoid evidence should use host‐specific development data where available.     

Specific detection of the Old World screwworm fly,Chrysomya bezziana,in bulk fly trap catches using real‐time PCR

The Old World screwworm fly (OWS), Chrysomya bezziana Villeneuve (Diptera: Calliphoridae), is a myiasis‐causing blowfly of major concern for both animals and humans. Surveillance traps are used in several countries for early detection of incursions and to monitor control strategies. Examination of surveillance trap catches is time‐consuming and is complicated by the presence of morphologically similar flies that are difficult to differentiate from Ch. bezziana, especially when the condition of specimens is poor. A molecular‐based method to confirm or refute the presence of Ch. bezziana in trap catches would greatly simplify monitoring programmes. A species‐specific real‐time polymerase chain reaction (PCR) assay was designed to target the ribosomal DNA internal transcribed spacer 1 (rDNA ITS1) of Ch. bezziana. The assay uses both species‐specific primers and an OWS‐specific Taqman^® MGB probe. Specificity was confirmed against morphologically similar and related Chrysomya and Cochliomyia species. An optimal extraction protocol was developed to process trap catches of up to 1000 flies and the assay is sensitive enough to detect one Ch. bezziana in a sample of 1000 non‐target species. Blind testing of 29 trap catches from Australia and Malaysia detected Ch. bezziana with 100% accuracy. The probability of detecting OWS in a trap catch of 50 000 flies when the OWS population prevalence is low (one in 1000 flies) is 63.6% for one extraction. For three extractions (3000 flies), the probability of detection increases to 95.5%. The real‐time PCR assay, used in conjunction with morphology, will greatly increase screening capabilities in surveillance areas where OWS prevalence is low.     

Using COI barcodes to identify forensically and medically important blowflies

Abstract.  The utility of cytochrome oxidase I (COI) DNA barcodes for the identification of nine species of forensically important blowflies of the genus Chrysomya (Diptera: Calliphoridae), from Australia, was tested. A 658-bp fragment of the COI gene was sequenced from 56 specimens, representing all nine Chrysomya species and three calliphorid outgroups. Nucleotide sequence divergences were calculated using the Kimura-two-parameter distance model and a neighbour-joining (NJ) analysis was performed to provide a graphic display of the patterns of divergence among the species. All species were resolved as reciprocally monophyletic on the NJ tree. Mean intraspecific and interspecific sequence divergences were 0.097% (range 0–0.612%, standard error [SE] = 0.119%) and 6.499% (range 0.458–9.254%, SE = 1.864%), respectively. In one case, a specimen that was identified morphologically was recovered with its sister species on the NJ tree. The hybrid status of this specimen was established by sequence analysis of the second ribosomal internal transcribed spacer (ITS2). In another instance, this nuclear region was used to verify four cases of specimen misidentification that had been highlighted by the COI analysis. The COI barcode sequence was found to be suitable for the identification of Chrysomya species from the east coast of Australia.     

Genetic and environmental influences on the cuticular hydrocarbon profiles of Goniozus wasps

The cuticular hydrocarbon (CHC) profiles of insects are well known to be variable. This variation may be due to genetic influences, environmental influences, or both. Most prior studies have focused on social insects, mainly those in the Hymenoptera, and have shown that hydrocarbons play an important role mediating social behaviour, particularly via kin recognition. Here, we assess the CHC profiles of three species of parasitoid wasps in the genus Goniozus (Hymenoptera: Bethylidae), some of which are known to attune their behaviour according to both environmentally based and genetically based recognition of kin. We find that CHC profiles vary according to both the genetic background (wasp species) and the developmental environment (host species) of individual parasitoids. This indicates that kin recognition could be based on CHC profiles in these parasitoids, as it is in social Hymenoptera. Because the CHC profiles of species within the genus Goniozus are dissimilar, we also conclude that chemical analysis could be used as a taxonomic tool alongside morphological and molecular genetic identification for Goniozus and other species.     

The sex-lethal gene homologue in Chrysomya rufifacies is highly conserved in sequence and exon-intron organization

A great variety of sex determination mechanisms exists in insect species. In Drosophila melanogaster sex is determined by the ratio between X chromosomes and autosomes, while in the blowfly Chrysomya rufifacies it is maternally determined. A cascade of genes which are involved in sex determination has been identified in D. melanogaster with the Sex-lethal gene (Sxl) as the key gene. We screened genomic libraries of C. rufifacies with a probe of the Sxl gene from D. melanogaster and isolated a genomic region that included most of the homologous gene. DNA- and protein-sequence comparison showed a high percent identity between the Chrysomya and the Drosophila gene. Up to 90% identity of the amino acid sequences was found in the region that contained the RNA-binding domains. The degree of identity is much lower outside of this functionally important region (18% identity). cDNA analysis showed a highly conserved exon-intron structure between the two species, although sex-specific splicing as used in D. melanogaster for the regulation of Sxl activity, could not be detected in C. rufifacies.     

Molecular phylogeny of the blowfly genus Chrysomya

Chrysomya Robineau-Desvoidy (Diptera: Calliphoridae) is a genus of blowfly commonly observed in tropical and subtropical countries of the Old World. Species in this genus are vectors of bacteria, protozoans and helminths, cause myiasis, are predators of other carrion insects, and are important forensic indicators. Hypotheses concerning the evolution of sex determination, larval anatomy and genome size in Chrysomya have been difficult to evaluate because a robust phylogeny of the genus was lacking. Similarly, the monophyly of subgenera was uncertain. The phylogeny of Chrysomya spp. was reconstructed based on 2386 bp of combined mitochondrial cytochrome oxidase subunit I (COI) and nuclear carbamoylphosphate synthetase (CPS) genes. Maximum parsimony (MP), maximum likelihood (ML) and Bayesian analysis (BA) differed only slightly in the resulting tree topology. Chrysomya was monophyletic. Monogenic reproduction is almost certainly derived rather than, as has been suggested, primitive within the genus, and tuberculate larvae probably evolved twice. Genome size is more likely to have decreased over evolutionary time rather than, as has been suggested, increased within the genus, but its correlation with developmental time was not observed. The subgenera Microcalliphora, Eucompsomyia and Achoetandrus were recovered as monophyletic.     

Mitochondrial genomes of the sheep blowfly, Lucilia sericata, and the secondary blowfly, Chrysomya megacephala

This paper presents complete mitochondrial genomes for the sheep blowfly, Lucilia sericata (Meigen), and the secondary blowfly, Chrysomya megacephala (Fabricius). Both L. sericata and C. megacephala had standard dipteran-type mitochondrial genome architectures and lengths of 15 945 bp and 15 831 bp, respectively. Additionally, C. megacephala possessed a tRNA duplication either side of the D-loop, as previously reported in another Chrysomya species, C. putoria; this duplication appears to be synapomorphic for the genus Chrysomya. As in other insect mitochondrial genomes, base compositions had a high AT content, with both genomes more than 76% AT-rich.     

How do invasive species affect native species? Experimental evidence from a carrion blowfly (Diptera: Calliphoridae) system

1. The necrobiome is a unique microcosm in which various organisms interact and compete for access to an ephemeral resource, such as carrion, that ultimately determines the structure and composition of these assemblages. 2. Blowfly species exhibit different competitive abilities which, when associated with other types of behaviour, such as predation or cannibalism, influence coexistence. Knowledge of the effects of competition between native and invasive species on development and survival is essential to understanding the dynamics of insect communities and to assess biological invasions. 3. Laboratory experiments were performed to evaluate the effect of interspecific competition on the bionomics and survival of a native (Cochliomyia macellaria) and an invasive (Chrysomya rufifacies) blowfly species at different population densities. 4. The deleterious effect of competition on the larval parameters of C. macellaria increased proportionally with increases in the larval density of C. rufifacies. When exposed to increased densities of C. rufifacies, larvae of C. macellaria accelerated their development and, as a trade‐off for this strategy, surviving adults were smaller and had reduced wing size, which were likely to reduce dispersal and reproductive capacity. 5. Larval competition – both as species‐dependent and density‐dependent phenomena – influences morphological and biological traits of surviving individuals. The impact of the invasive species has consequences at the population level, such as displacement or local population depletion of native species, a phenomenon likely to occur in other systems involving insects and ephemeral resources.     

Stochastic dynamics in exotic and native blowflies: an analysis combining laboratory experiments and a two-patch metapopulation model

In this study we explored the stochastic population dynamics of three exotic blowfly species, Chrysomya albiceps, Chrysomya megacephala and Chrysomya putoria, and two native species, Cochliomyia macellaria and Lucilia eximia, by combining a density-dependent growth model with a two-patch metapopulation model. Stochastic fecundity, survival and migration were investigated by permitting random variations between predetermined demographic boundary values based on experimental data. Lucilia eximia and Chrysomya albiceps were the species most susceptible to the risk of local extinction. Cochliomyia macellaria, C. megacephala and C. putoria exhibited lower risks of extinction when compared to the other species. The simultaneous analysis of stochastic fecundity and survival revealed an increase in the extinction risk for all species. When stochastic fecundity, survival and migration were simulated together, the coupled populations were synchronized in the five species. These results are discussed, emphasizing biological invasion and interspecific interaction dynamics.     

Preliminary notes on the genus Aeschnosoma Selys 1870 (Odonata: Anisoptera: Corduliidae s. str.)

Three new species of the genus Aeschnosoma are briefly described and illustrated. A. pseudoforcipula n. sp. and A. heliophila n. sp., both from the Brazilian Central Plateau are respectively related to the two Amazonian species A. forcipula Hagen in Selys 1871, and A. auripennis Geijskes 1970. A. louissiriusi n. sp. from Northern Brazil is not closely related to any known species. Based on larval and adult derived characters, the genus Aeschnosoma appears closely related to the Australian genus Pentathemis Karsch 1890, and also to the Madagascan genus Libellulosoma Martin 1907. The clade Aeschnosomata nov. is erected to receive the three genera. Some putative plesiomorphies would place this clade sister group of the remaining Corduliidae s.str.     

Phylogeny and biogeography of Muusoctopus (Cephalopoda: Enteroctopodidae)

Deep‐sea octopuses of the genus Muusoctopus are thought to have originated in the Pacific Northern Hemisphere and then diversified throughout the Pacific and into the rest of the World Ocean. However, this hypothesis was inferred only from molecular divergence times. Here, the ancestral distribution and dispersal routes are estimated by Bayesian analysis based on a new phylogeny including 38 specimens from the south‐eastern Pacific Ocean. Morphological data and molecular sequences of three mitochondrial genes (16S rRNA, COI and COIII) are presented. The morphological data confirm that specimens newly acquired from off the coast of Chile comprise two species: Muusoctopus longibrachus and the poorly described species, Muusoctopus eicomar. The latter is here redescribed and is clearly distinguished from M. longibrachus and other closely related species in the region. A gene tree was built using Bayesian analysis to infer the phylogenetic position of these species within the species group, revealing that a large genetic distance separates the two sympatric Chilean species. M. longibrachus is confirmed as the sister species of Muusooctopus eureka from the Falkland Islands; while M. eicomar is a sister species of Muusoctopus yaquinae from the North Pacific, most closely related to the amphi‐Atlantic species Muusoctopus januarii. Molecular divergence times and ancestral distribution analyses suggest that genus Muusoctopus may have originated in the North Atlantic: one lineage dispersed directly southward to the Magellan region and another dispersed southward along the Eastern Pacific to the Southern Ocean and Antarctica. The Muusoctopus species in the Southern Hemisphere have different phylogenetic origins and represent independent invasions of this region.     

Towards the automated identification of Chrysomya blow flies from wing images

The Old World screwworm fly (OWSF), Chrysomya bezziana (Diptera: Calliphoridae), is an important agent of traumatic myiasis and, as such, a major human and animal health problem. In the implementation of OWSF control operations, it is important to determine the geographical origins of such disease‐causing species in order to establish whether they derive from endemic or invading populations. Gross morphological and molecular studies have demonstrated the existence of two distinct lineages of this species, one African and the other Asian. Wing morphometry is known to be of substantial assistance in identifying the geographical origin of individuals because it provides diagnostic markers that complement molecular diagnostics. However, placement of the landmarks used in traditional geometric morphometric analysis can be time‐consuming and subject to error caused by operator subjectivity. Here we report results of an image‐based approach to geometric morphometric analysis for delivering wing‐based identifications. Our results indicate that this approach can produce identifications that are practically indistinguishable from more traditional landmark‐based results. In addition, we demonstrate that the direct analysis of digital wing images can be used to discriminate between three Chrysomya species of veterinary and forensic importance and between C. bezziana genders.