High-throughput monitoring of wild bee diversity and abundance via mitogenomics

1. Bee populations and other pollinators face multiple, synergistically acting threats, which have led to population declines, loss of local species richness and pollination services, and extinctions. However, our understanding of the degree, distribution and causes of declines is patchy, in part due to inadequate monitoring systems, with the challenge of taxonomic identification posing a major logistical barrier. Pollinator conservation would benefit from a high-throughput identification pipeline. 2. We show that the metagenomic mining and resequencing of mitochondrial genomes (mitogenomics) can be applied successfully to bulk samples of wild bees. We assembled the mitogenomes of 48 UK bee species and then shotgun-sequenced total DNA extracted from 204 whole bees that had been collected in 10 pan-trap samples from farms in England and been identified morphologically to 33 species. Each sample data set was mapped against the 48 reference mitogenomes. 3. The morphological and mitogenomic data sets were highly congruent. Out of 63 total species detections in the morphological data set, the mitogenomic data set made 59 correct detections (93�7% detection rate) and detected six more species (putative false positives). Direct inspection and an analysis with species-specific primers suggested that these putative false positives were most likely due to incorrect morphological IDs. Read frequency significantly predicted species biomass frequency (R2 = 24�9%). Species lists, biomass frequencies, extrapolated species richness and community structure were recovered with less error than in a metabarcoding pipeline. 4. Mitogenomics automates the onerous task of taxonomic identification, even for cryptic species, allowing the tracking of changes in species richness and istributions. A mitogenomic pipeline should thus be able to contain costs, maintain consistently high-quality data over long time series, incorporate retrospective taxonomic revisions and provide an auditable evidence trail. Mitogenomic data sets also provide estimates of species counts within samples and thus have potential for tracking population trajectories.

Size matters: body size determines functional responses of ground beetle interactions

Understanding patterns in predator:prey systems and the mechanisms that underlie trophic interactions provides a basis for predicting community structure and the delivery of natural pest control services. The functional response of predators to prey density is a fundamental measure of interaction strength and its characterisation is essential to understanding these processes. We used mesocosm experiments to quantify the functional responses of five ground beetle species that represent common generalist predators of north-west European arable agriculture. We investigated two mechanisms predicted to be key drivers of trophic interactions in natural communities: predator:prey body size ratio and multiple predator effects. Our results show regularities in foraging patterns characteristic of similarly sized predators. Ground beetle attack rates increased and handling times decreased as the predator:prey body-mass ratio rose. Multiple predator effects on total prey consumption rates were sensitive to the identity of the interacting species but not prey density. The extent of interspecific interactions may be a result of differences in body mass between competing beetle species. Overall these results add to the growing evidence for the importance of size in determining trophic interactions and suggest that body mass could offer a focus on which to base the management of natural enemy assemblages.

The ancient Britons: groundwater fauna survived extreme climate change over tens of millions of years across NW Europe

Global climate changes during the Cenozoic (65.5–0 Ma) caused major biological range shifts and extinctions. In northern Europe, for example, a pattern of few endemics and the dominance of wide-ranging species is thought to have been determined by the Pleistocene (2.59–0.01 Ma) glaciations. This study, in contrast, reveals an ancient subsurface fauna endemic to Britain and Ireland. Using a Bayesian phylogenetic approach, we found that two species of stygobitic invertebrates (genus Niphargus) have not only survived the entire Pleistocene in refugia but have persisted for at least 19.5 million years. Other Niphargus species form distinct cryptic taxa that diverged from their nearest continental relative between 5.6 and 1.0 Ma. The study also reveals an unusual biogeographical pattern in the Niphargus genus. It originated in north-west Europe approximately 87 Ma and underwent a gradual range expansion. Phylogenetic diversity and species age are highest in north-west Europe, suggesting resilience to extreme climate change and strongly contrasting the patterns seen in surface fauna. However, species diversity is highest in south-east Europe, indicating that once the genus spread to these areas (approximately 25 Ma), geomorphological and climatic conditions enabled much higher diversification. Our study highlights that groundwater ecosystems provide an important contribution to biodiversity and offers insight into the interactions between biological and climatic processes.

Prevalence and nonrandom distribution of exonic mutations in interferon regulatory factor 6 in 307 families with Van der Woude syndrome and 37 families with popliteal pterygium syndrome

Purpose: Interferon regulatory factor 6 encodes a member of the IRF family of transcription factors. Mutations in interferon regulatory factor 6 cause Van der Woude and popliteal pterygium syndrome, two related orofacial clefting disorders. Here, we compared and contrasted the frequency and distribution of exonic Mutations in interferon regulatory factor 6 between two large geographically distinct collections of families with Van der Woude and between one collection of families with popliteal pterygium syndrome. Methods: We performed direct sequence analysis of interferon regulatory factor 6 exons oil samples from three collections, two with Van der Woude and one with popliteal pterygium syndrome. Results: We identified mutations in interferon regulatory factor 6 exons in 68% of families in both Van der Woude collections and in 97% of families with popliteal pterygium syndrome. In sum, 106 novel disease-causing variants were found. The distribution of mutations in the interferon regulatory factor 6 exons in each collection was not random; exons 3, 4, 7, and 9 accounted for 80%. In the Van der Woude collections, the mutations were evenly divided between protein truncation and missense, whereas most mutations identified in the popliteal pterygium syndrome collection were missense. Further, the missense mutations associated with popliteal pterygium syndrome were localized significantly to exon 4, at residues that are predicted to bind directly to DNA. Conclusion: The nonrandom distribution of mutations in the interferon regulatory factor 6 exons suggests a two-tier approach for efficient mutation screens for interferon regulatory factor 6. The type and distribution of mutations are consistent with the hypothesis that Van der Woude is caused by haploinsufficiency of interferon regulatory factor 6. Oil the other hand, the distribution of popliteal pterygium syndrome-associated mutations suggests a different, though not mutually exclusive, effect oil interferon regulatory factor 6 function. Genet Med 2009:11(4):241-247.

Selective preying of the sphecid wasp Trachypus boharti on the meliponine bee Scaptotrigona postica: potential involvement of caste-specific cuticular hydrocarbons

The specialist digger wasp Trachypus boharti Rubio-Espina preys exclusively on males of the stingless bee Scaptotrigona postica Latreille 1807, although the hunting attacks involve both male and worker bees of S. postica and members of its own species. To understand the mechanism of prey selection, the cuticular hydrocarbon patterns of workers and males of S. postica are analyzed in detail, and the mandibular secretion of males is examined. The cuticular profiles of males and workers are distinctively different. The major group of cuticular compounds, heptacosene isomers, is twice as abundant in workers as in males. There is no clear distinction between worker and male mandibular secretions. Such a distinct and straightforward caste-specific difference in cuticular hydrocarbons could function as a recognition cue by which T. boharti distinguishes between workers and males of S. postica.

Nested diets: a novel pattern of individual-level resource use

Many generalist populations may actually be composed of relatively specialist individuals. This `individual specialization` may have important ecological and evolutionary implications. Although this phenomenon has been documented in more than one hundred taxa, it is still unclear how individuals within a population actually partition resources. Here we applied several methods based on network theory to investigate the intrapopulation patterns of resource use in the gracile mouse opossum Gracilinanus microtarsus. We found evidence of significant individual specialization in this species and that the diets of specialists are nested within the diets of generalists. This novel pattern is consistent with a recently proposed model of optimal foraging and implies strong asymmetry in the interactions among individuals of a population.

Feeding habits of the maned wolf, Chrysocyon brachyurus (Carnivora: Canidae), in southeast Brazil

The feeding habits of the maned wolf were studied in southeast Brazil to understand its response towards changes in the environment and in relation to its prey. By occurrence, miscellaneous fruits, small mammals and wolf`s fruit were the most consumed items. Armadillos, small mammals and wolf`s fruit ( Solanum lycocarpum) provided most of the ingested biomass. While wolf`s fruit and small mammals were mainly consumed in the dry season, other miscellaneous fruits were taken mostly in the wet season. There was selectivity in the predation on some small mammal species. The maned wolf`s diet followed patterns similar to those found in more pristine areas.

Reproductive ecology of dipsadine snakes, with emphasis on South American species

A relatively large amount of variation occurs in the reproductive ecology of tropical snakes, and this variation is generally regarded as being a consequence of seasonality in climate and prey availability. In some groups, even closely related species may differ in their reproductive ecology; however, in others it seems to be very conservative. Here we explore whether characters related to reproduction are phylogenetically constrained in a monophyletic group of snakes, the subfamily Dipsadinae, which ranges from Mexico to southern South America. We provide original data on reproduction for Leptodeira annulata, Imantodes cenchoa, and three species of Sibynomorphus from southern, southeastern and central Brazil, and data from literature for other species and populations of dipsadines. Follicular cycles were seasonal in Atractus reticulatus, Dipsa, albifrons, Hypsiglena torquata, Leptodeira maculata, L. punctata, Sibynomorphus spp. and Sibon sanniola from areas where climate is seasonal. In contrast, extended or continuous follicular cycles were recorded in Dipsas catesbyi, D. neivai, Imantodes cenchoa, Leptodeira annulata, and Ninia maculata from areas with seasonal and aseasonal climates. Testicular cycles also varied from seasonal (in H. torquiata) to continuous (in Dipsa,5 spp., Leptodeira annulata, L. maculata, N. maculata, and Sibynomorphus spp.). Most dipsadines are small (less than 500 rum SVL), and females attain sexual maturity with similar relative body size than males. Sexual dimorphism occurred in terms of SVL and tail length in most species, and clutch size tended to be small (less than five eggs). Combat behavior occurs in Imantodes cenchoa, which did not show sexual size dimorphism. Reproductive timing, for both females and males, varied among species but in general there were no differences between the tribes of Dipsadinae in most of the reproductive characteristics, such as mean body size, relative size at sexual maturity, sexual size and tail dimorphism, duration of vitellogenesis or egg-carrying in oviducts.

Proteomics of the neurotoxic fraction from the sea anemone Bunodosoma cangicum venom: Novel peptides belonging to new classes of toxins

In contrast to the many studies on the venoms of scorpions, spiders, snakes and cone snails, tip to now there has been no report of the proteomic analysis of sea anemones venoms. In this work we report for the first time the peptide mass fingerprint and some novel peptides in the neurotoxic fraction (Fr III) of the sea anemone Bunodosoma cangicum venom. Fr III is neurotoxic to crabs and was purified by rp-HPLC in a C-18 column, yielding 41 fractions. By checking their molecular masses by ESI-Q-Tof and MALDI-Tof MS we found 81 components ranging from near 250 amu to approximately 6000 amu. Some of the peptidic molecules were partially sequenced through the automated Edman technique. Three of them are peptides with near 4500 amu belonging to the class of the BcIV, BDS-I, BDS-II, APETx1, APETx2 and Am-II toxins. Another three peptides represent a novel group of toxins (similar to 3200 amu). A further three molecules (similar to similar to 4900 amu) belong to the group of type 1 sodium channel neurotoxins. When assayed over the crab leg nerve compound action potentials, one of the BcIV- and APETx-like peptides exhibits an action similar to the type 1 sodium channel toxins in this preparation, suggesting the same target in this assay. On the other hand one of the novel peptides, with 3176 amu, displayed an action similar to potassium channel blockage in this experiment. In summary, the proteomic analysis and mass fingerprint of fractions from sea anemone venoms through MS are valuable tools, allowing us to rapidly predict the occurrence of different groups of toxins and facilitating the search and characterization of novel molecules without the need of full characterization of individual components by broader assays and bioassay-guided purifications. It also shows that sea anemones employ dozens of components for prey capture and defense. (C) 2008 Elsevier Inc. All rights reserved.

Evidence of a genetic bottleneck in an El Nino affected population of South American fur seals, Arctocephalus australis

The South American fur seal, Arctocephalus australis, was one of the earliest otariid seals to be exploited by humans: at least 6000 years ago on the Atlantic coast and 4000 on the Pacific coast of South America. More than 750,000 fur seals were killed in Uruguay until 1991. However, a climatological phenomenon-the severe 1997-1998 El Nino Southern Oscillation (ENSO)-was responsible for the decline of 72% Of the Peruvian fur seal population due to starvation as a consequence of warming of sea-surface temperatures and primary productivity reduction. Currently, there is no precise information on global population size or on the species` conservation status. The present study includes the first bottleneck test for the Pacific and Atlantic populations of A. australis based on the analysis of seven microsatellite loci. Genetic bottleneck compromises the evolutionary potential of a population to respond to environmental changes. The perspective becomes even more alarming due to current global warming models that predict stronger and more frequent ENSO events in the future. Our analysis found moderate support for deviation from neutrality-equilibrium for the Pacific population of fur seals and none for the Atlantic population. This difference among population reflects different demographic histories, and is consistent with a greater reduction in population size in the Pacific. Such an event could be a result of the synergic effects of recurrent ENSO events and the anthropogenic impact (sealing and prey overfishing) on this population.

The hidden diversity of Coleodactylus amazonicus (Sphaerodactylinae, Gekkota) revealed by molecular data

Coleodactylus amazonicus, a small leaf-litter diurnal gecko widely distributed in Amazon Basin has been, considered a single species with no significant morphological differences between populations along its range. A recent molecular study, however, detected large genetic differences between populations of central Amazonia and those in the easternmost part of the Amazon Basin, suggesting the presence of taxonomically unrecognised diversity. In this study, DNA sequences of three mitochondrial (165, cytb, and ND4) and two nuclear genes (RAG-1, c-mos) were used to investigate whether the species currently identified as C. amazonicus contains morphologically cryptic species lineages. The present phylogenetic analysis reveals further genetic subdivision including at least five potential species lineages, restricted to northeastern (lineage A), southeastern (lineage B), central-northern (lineage E) and central-southern (lineages C and D) parts of Amazon Basin. All clades are characterized by exclusive groups of alleles for both nuclear genes and highly divergent mitochondrial haplotype clades, with corrected pairwise net sequence divergence between sister lineages ranging from 9.1% to 20.7% for the entire mtDNA dataset. Results of this study suggest that the real diversity of ""C. amazonicus"" has been underestimated due to its apparent cryptic diversification. (C) 2009 Elsevier Inc. All rights reserved.

Comparing methods for sampling large- and medium-sized mammals: camera traps and track plots

Activities involving fauna monitoring are usually limited by the lack of resources; therefore, the choice of a proper and efficient methodology is fundamental to maximize the cost-benefit ratio. Both direct and indirect methods can be used to survey mammals, but the latter are preferred due to the difficulty to come in sight of and/or to capture the individuals, besides being cheaper. We compared the performance of two methods to survey medium and large-sized mammal: track plot recording and camera trapping, and their costs were assessed. At Jatai Ecological Station (S21 degrees 31`15 ``- W47 degrees 34`42 ``-Brazil) we installed ten camera traps along a dirt road directly in front of ten track plots, and monitored them for 10 days. We cleaned the plots, adjusted the cameras, and noted down the recorded species daily. Records taken by both methods showed they sample the local richness in different ways (Wilcoxon, T=231; p;;0.01). The track plot method performed better on registering individuals whereas camera trapping provided records which permitted more accurate species identification. The type of infra-red sensor camera used showed a strong bias towards individual body mass (R(2)=0.70; p=0.017), and the variable expenses of this method in a 10-day survey were estimated about 2.04 times higher compared to track plot method; however, in a long run camera trapping becomes cheaper than track plot recording. Concluding, track plot recording is good enough for quick surveys under a limited budget, and camera trapping is best for precise species identification and the investigation of species details, performing better for large animals. When used together, these methods can be complementary.

Head shape evolution in Tropidurinae lizards: does locomotion constrain diet?

Different components of complex integrated systems may be specialized for different functions, and thus the selective pressures acting on the system as a whole may be conflicting and can ultimately constrain organismal performance and evolution. The vertebrate cranial system is one of the most striking examples of a complex system with several possible functions, being associated to activities as different as locomotion, prey capture, display and defensive behaviours. Therefore, selective pressures on the cranial system as a whole are possibly complex and may be conflicting. The present study focuses on the influence of potentially conflicting selective pressures (diet vs. locomotion) on the evolution of head shape in Tropidurinae lizards. For example, the expected adaptations leading to flat heads and bodies in species living on vertical structures may conflict with the need for improved bite performance associated with the inclusion of hard or tough prey into the diet, a common phenomenon in Tropidurinae lizards. Body size and six variables describing head shape were quantified in preserved specimens of 23 species, and information on diet and substrate usage was obtained from the literature. No phylogenetic signal was observed in the morphological data at any branch length tested, suggesting adaptive evolution of head shape in Tropidurinae. This pattern was confirmed by both factor analysis and independent contrast analysis, which suggested adaptive co-variation between the head shape and the inclusion of hard prey into the diet. In contrast to our expectations, habitat use did not constrain or drive head shape evolution in the group.

Genomic imbalances associated with mullerian aplasia

Background: Aplasia of the mullerian ducts leads to absence of the uterine corpus, uterine cervix, and upper (superior) vagina. Patients with mullerian aplasia (MA) often exhibit additional clinical features such as renal, vertebral and cardiac defects. A number of different syndromes have been associated with MA, and in most cases its aetiology remains poorly understood. Objective and methods: 14 syndromic patients with MA and 46, XX G-banded karyotype were screened for DNA copy number changes by similar to 1 Mb whole genome bacterial artificial chromosome (BAC) array based comparative genomic hybridisation (CGH). The detected alterations were validated by an independent method and further mapped by high resolution oligo-arrays. Results: Submicroscopic genomic imbalances affecting the 1q21.1, 17q12, 22q11.21, and Xq21.31 chromosome regions were detected in four probands. Presence of the alterations in the normal mother of one patient suggests incomplete penetrance and/or variable expressivity. Conclusion: 4 of the 14 patients (29%) were found to have cryptic genomic alterations. The imbalances on 22q11.21 support recent findings by us and others that alterations in this chromosome region may result in impairment of mullerian duct development. The remaining imbalances indicate involvement of previously unknown chromosome regions in MA, and point specifically to LHX1 and KLHL4 as candidate genes.

Tenacity and silk investment of two orb weavers: considerations about diversification of the Araneoidea

Orbiculariae consists of two major clades: the cribellate Deinopidea and the much more diverse ecribellate Araneoidea. It has been hypothesized that the higher diversity of Araneoidea is a consequence of the superiority of the viscid orb web. However, this explanation seems incomplete: for example, cribellate silk may perform better than viscid silk in some contexts. Here, we consider the hypothesis that the diversification of Araneoidea was facilitated by changes in microhabitat occupation behavior due to the cheaper viscid orb web. In the present work we investigate the idea that the reduction in site tenacity caused by the emergence of the viscid orb web has led to an increase in the exploration of different resources and to a greater diversification of the Arancoidea through the evolutionary time. To test this idea, we evaluated the response of one cribellate orb web spider (Zosis geniculata Olivier 1789, Uloboridae) and one ecribellate orb web spider (Metazygia rogenhoferi Keyserling 1878, Arancidae) to abrupt prey absence. The changes in site tenacity and the day-to-day investment in web silk were evaluated. Spiders with three-dimensional webs tend to exhibit greater site tenacity than spiders making orb webs. Zosis geniculata and M. rogenhoferi show similar site tenacity when prey is ample. When prey is unavailable, the tenacity of the cribellate species increases while the tenacity of the ecribellate remains unchanged, and the silk investment of both species decreases. However, this decrease in silk investment is more extensive in Z. geniculata. These results coincide with the idea that a less costly ecribellate orb web leads to a lower tenacity and suggest that more frequent microhabitat abandonment in a context of insect radiation (Neiptera) leads to more diverse and opportunistic exploration of microhabitats that, in the long term, may be one explanation for the greater Araneoidea diversification.