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In the original article,Box 1 was not printed in its entirety, and two references were badly quoted in page 3 of the pdf and were missing in the reference list.

The use of DNA data is ubiquitous across animal sciences. DNA may be obtained from an organism for a myriad of reasons including identification and distinction between cryptic species, sex identification, comparisons of different morphocryptic genotypes or assessments of relatedness between organisms prior to a behavioural study. DNA should be obtained while minimizing the impact on the fitness, behaviour or welfare of the subject being tested, as this can bias experimental results and cause long-lasting effects on wild animals. Furthermore, minimizing impact on experimental animals is a key Refinement principle within the ‘3Rs’ framework which aims to ensure that animal welfare during experimentation is optimised. The term ‘non-invasive DNA sampling’ has been defined to indicate collection methods that do not require capture or cause disturbance to the animal, including any effects on behaviour or fitness. In practice this is not always the case, as the term ‘non-invasive’ is commonly used in the literature to describe studies where animals are restrained or subjected to aversive procedures. We reviewed the non-invasive DNA sampling literature for the past six years (380 papers published in 2013-2018) and uncovered the existence of a significant gap between the current use of this terminology (i.e. ‘non-invasive DNA sampling’) and its original definition. We show that 58% of the reviewed papers did not comply with the original definition. We discuss the main experimental and ethical issues surrounding the potential confusion or misuse of the phrase ‘non-invasive DNA sampling’ in the current literature and provide potential solutions. In addition, we introduce the terms ‘non-disruptive’ and ‘minimally disruptive’ DNA sampling, to indicate methods that eliminate or minimise impacts not on the physical integrity/structure of the animal, but on its behaviour, fitness and welfare, which in the literature reviewed corresponds to the situation for which an accurate term is clearly missing. Furthermore, we outline when these methods are appropriate to use.

Habitat loss is one of the greatest drivers for extinction worldwide. Understanding a taxon's habitat requirements is crucial in being able to protect it from decline. The Australasian trapdoor spider genus Cantuaria (Idiopidae) is widespread and diverse within the islands of New Zealand, but little is known of its ecology. We studied the habitat requirements of Cantuaria, using niche modelling, to inform conservation efforts and predict how the genus may fare under future climate conditions. Two data sets were employed. A fine‐scale data set used hand‐collected data from the vicinity of individual burrows for localised soil, vegetation, and human disturbance data directly inside and adjacent to Cantuaria populations. A spatial GIS data set at the landscape level mapped presence/absence of populations, as well as general environmental and soil conditions, and vegetation type. The two data sets were analysed separately, using linear models for the fine‐scale data set (17 presences, 17 absences). General linear models were used to analyse the spatial data set (72 presences, 17 absences), with variables selected using a random forest analysis in R. In the fine‐scale data set, no significant variables were found. In the spatial data set, rainfall explained most (71.4%) of the variation, according to the random forest analysis, followed by soil group (33.71%) and elevation (27.12%). Some variation was explained by more than one factor. The two models with the lowest AICC numbers included rainfall (P < 0.01) and elevation (P = 0.06), or rainfall only (P < 0.01), as variables. We conclude that low rainfall is desirable for populations of most Cantuaria spp., although some have been found where rainfall is up to 3000 mm/year. Climate change may cause challenges for Cantuaria conservation, particularly in areas with increased rainfall, such as Southland, Tasman, the West Coast, and Central Otago. Elevation and soil type may also be important factors in Cantuaria habitat selection, but further research with gene flow analysis, captive studies, more structured sampling, and/or a greater sample size may reveal more about the effects of elevation and soil type on Cantuaria populations. [ABSTRACT FROM AUTHOR]

Homeoviscous adaptation (HVA) is a key cellular response by which fish protect their membranes against thermal stress. We investigated evolutionary HVA (long time scale) in Antarctic and non-Antarctic fish. Membrane lipid composition was determined for four Perciformes fish: two closely related Antarctic notothenioid species (Trematomus bernacchii and Pagothenia borchgrevinki); a diversified related notothenioid Antarctic icefish (Chionodraco hamatus); and a New Zealand species (Notolabrus celidotus). The membrane lipid compositions were consistent across the three Antarctic species and these were significantly different from that of the New Zealand species. Furthermore, acclimatory HVA (short time periods with seasonal changes) was investigated to determine whether stenothermal Antarctic fish, which evolved in the cold, stable environment of the Southern Ocean, have lost the acclimatory capacity to modulate their membrane saturation states, making them vulnerable to anthropogenic global warming. We compared liver membrane lipid composition in two closely related Antarctic fish species acclimated at 0 °C (control temperature), 4 °C for a period of 14 days in T. bernacchii and 28 days for P. borchgrevinki, and 6 °C for 7 days in both species. Thermal acclimation at 4 °C did not result in changed membrane saturation states in either Antarctic species. Despite this, membrane functions were not compromised, as indicated by declining serum osmolality, implying positive compensation by enhanced hypo-osmoregulation. Increasing the temperature to 6 °C did not change the membrane lipids of P. borchgrevinki. However, in T. bernacchii, thermal acclimation at 6 °C resulted in an increase of membrane saturated fatty acids and a decline in unsaturated fatty acids. This is the first study to show a homeoviscous response to higher temperatures in an Antarctic fish, although for only one of the two species examined. [ABSTRACT FROM AUTHOR]

The New Zealand fur seal (Arctocephalus forsteri) passed through a population bottleneck due to commercial sealing during the eighteenth to nineteenth centuries. To facilitate future management options, we reconstructed the demographic history of New Zealand fur seals in a Bayesian framework using maternally inherited, mitochondrial DNA sequences. Mitogenomic data suggested two separate clades (most recent common ancestor 5000 years ago) of New Zealand fur seals that survived large-scale human harvest. Mitochondrial haplotype diversity was high, with 45 singletons identified from 46 individuals although mean nucleotide diversity was low (0.012 ± 0.0061). Variation was not constrained geographically. Analyses of mitogenomes support the hypothesis for a population bottleneck approximately 35 generations ago, which coincides with the peak of commercial sealing. Mitogenomic data are consistent with a pre-human effective population size of approximately 30,000 that first declined to around 10,000 (due to the impact of Polynesian colonization, particularly in the first 100 years of their arrival into New Zealand), and then to 100-200 breeding individuals during peak of commercial sealing. [ABSTRACT FROM AUTHOR]

Homeoviscous adaptation (HVA) is a key cellular response by which fish protect their membranes against thermal stress. We investigated evolutionary HVA (long time scale) in Antarctic and non-Antarctic fish. Membrane lipid composition was determined for four Perciformes fish: two closely related Antarctic notothenioid species (Trematomus bernacchii and Pagothenia borchgrevinki); a diversified related notothenioid Antarctic icefish (Chionodraco hamatus); and a New Zealand species (Notolabrus celidotus). The membrane lipid compositions were consistent across the three Antarctic species and these were significantly different from that of the New Zealand species. Furthermore, acclimatory HVA (short time periods with seasonal changes) was investigated to determine whether stenothermal Antarctic fish, which evolved in the cold, stable environment of the Southern Ocean, have lost the acclimatory capacity to modulate their membrane saturation states, making them vulnerable to anthropogenic global warming. We compared liver membrane lipid composition in two closely related Antarctic fish species acclimated at 0 °C (control temperature), 4 °C for a period of 14 days in T. bernacchii and 28 days for P. borchgrevinki, and 6 °C for 7 days in both species. Thermal acclimation at 4 °C did not result in changed membrane saturation states in either Antarctic species. Despite this, membrane functions were not compromised, as indicated by declining serum osmolality, implying positive compensation by enhanced hypo-osmoregulation. Increasing the temperature to 6 °C did not change the membrane lipids of P. borchgrevinki. However, in T. bernacchii, thermal acclimation at 6 °C resulted in an increase of membrane saturated fatty acids and a decline in unsaturated fatty acids. This is the first study to show a homeoviscous response to higher temperatures in an Antarctic fish, although for only one of the two species examined. [ABSTRACT FROM AUTHOR]

. 1. Physical structure and species composition of vegetation determine spider diversity through habitat availability. Here, we assess, for the first time, the role of plant structure on spider communities in New Zealand native alpine tussock grasslands. We investigate the specific associations between spider assemblages and plant communities and test the hypothesis that spider diversity increases with plant diversity and tussock cover. 2. Spiders were sampled using density- and abundance-activity-dependent methods. Data on physical characteristics and species composition of plants revealed gradients in vegetation, driven by distinct intra-ecosystem plant communities. Effects of vegetation on spider diversity and composition were assessed through linear mixed-effects models. Redundancy analysis was used to determine and visualise the spider species-level response to gradients in plant compositions. 3. There was a positive relationship between plant diversity and spider diversity, while the effects of tussock cover varied with spider taxa. Overall, physical structure and species composition of vegetation had effects on the abundance of a similar number of spider species and families. 4. Gradients in vegetation were matched to gradients in spider communities, whose diversity and composition varied according to their habitat preference. The family Orsolobidae was associated with wetland vegetation, and Linyphiidae with shrubs. The abundance of certain spider families and species, such as Lycosidae and Anoteropsis hilaris, was consistently affected by vegetation. 5. Environmental factors, such as soil moisture, may affect plant species composition and physical structure in tussock grasslands, which in turn determine spider assemblages. Lycosidae were identified as potential indicators of structural changes in tussock grassland plants and could be valuable for ecological monitoring in conservation management. [ABSTRACT FROM AUTHOR]

1. DNA barcoding studies use Kimura's two-parameter substitution model (K2P) as the de facto standard for constructing genetic distance matrices. Distances generated under this model then provide the basis for most downstream analyses, but uncertainty in model choice is rarely explored and could potentially affect how reliably DNA barcodes discriminate species. 2. Using information-theoretic approaches for a data set comprising 14 472 DNA barcodes from 14 published studies, we tested whether the K2P model was a good fit at the species level and whether applying a better fitting model biased error rates or changed overall identification success. 3. We report that the K2P was a poorly fitting model at the species level; it was never selected as the best model and very rarely selected as a credible alternative model. Despite the lack of support for the K2P model, differences in distance between best model and K2P model estimates were usually minimal, and importantly, identification success rates were largely unaffected by model choice even when interspecific threshold values were reassessed. 4. Although these conclusions may justify using the K2P model for specimen identification purposes, we found simpler metrics such as p distance performed equally well, perhaps obviating the requirement for model correction in DNA barcoding. Conversely, when incorporating genetic distance data into taxonomic studies, we advocate a more thorough examination of model uncertainty. [ABSTRACT FROM AUTHOR]

DNA barcoding remains a challenge when applied to diet analyses, ancient DNA studies, environmental DNA samples and, more generally, in any cases where DNA samples have not been adequately preserved. Because the size of the commonly used barcoding marker (COI) is over 600 base pairs (bp), amplification fails when the DNA molecule is degraded into smaller fragments. However, relevant information for specimen identification may not be evenly distributed along the barcoding region, and a shorter target can be sufficient for identification purposes. This study proposes a new, widely applicable, method to compare the performance of all potential 'mini-barcodes' for a given molecular marker and to objectively select the shortest and most informative one. Our method is based on a sliding window analysis implemented in the new R package SPIDER (Species IDentity and Evolution in R). This method is applicable to any taxon and any molecular marker. Here, it was tested on earthworm DNA that had been degraded through digestion by carnivorous landsnails. A 100 bp region of 16 S rDNA was selected as the shortest informative fragment (mini-barcode) required for accurate specimen identification. Corresponding primers were designed and used to amplify degraded earthworm (prey) DNA from 46 landsnail (predator) faeces using 454-pyrosequencing. This led to the detection of 18 earthworm species in the diet of the snail. We encourage molecular ecologists to use this method to objectively select the most informative region of the gene they aim to amplify from degraded DNA. The method and tools provided here, can be particularly useful (1) when dealing with degraded DNA for which only small fragments can be amplified, (2) for cases where no consensus has yet been reached on the appropriate barcode gene, or (3) to allow direct analysis of short reads derived from massively parallel sequencing without the need for bioinformatic consolidation. [ABSTRACT FROM AUTHOR]

S pider: SPecies IDentity and Evolution in R is a new R package implementing a number of useful analyses for DNA barcoding studies and associated research into species delimitation and speciation. Included are functions essential for generating important summary statistics from DNA barcode data, assessing specimen identification efficacy, and for testing and optimizing divergence threshold limits. In terms of investigating evolutionary and taxonomic questions, techniques for assessing diagnostic nucleotides and probability of reciprocal monophyly are also provided. Additionally, a sliding window function offers opportunities to analyse information across a gene, essential for marker design in degraded DNA studies. S pider capitalizes on R's extensible ethos and offers an integrated platform ideal for the analysis of both nucleotide and morphological data. The program can be obtained from the comprehensive R archive network (CRAN, ) and from the R-Forge package development site (). [ABSTRACT FROM AUTHOR]

Background: Poorly regulated international trade in ornamental fishes poses risks to both biodiversity and economic activity via invasive alien species and exotic pathogens. Border security officials need robust tools to confirm identifications, often requiring hard-to-obtain taxonomic literature and expertise. DNA barcoding offers a potentially attractive tool for quarantine inspection, but has yet to be scrutinised for aquarium fishes. Here, we present a barcoding approach for ornamental cyprinid fishes by: (1) expanding current barcode reference libraries; (2) assessing barcode congruence with morphological identifications under numerous scenarios (e.g. inclusion of GenBank data, presence of singleton species, choice of analytical method); and (3) providing supplementary information to identify difficult species. Methodology/Principal Findings: We sampled 172 ornamental cyprinid fish species from the international trade, and provide data for 91 species currently unrepresented in reference libraries (GenBank/Bold). DNA barcodes were found to be highly congruent with our morphological assignments, achieving success rates of 90-99%, depending on the method used (neighbour-joining monophyly, bootstrap, nearest neighbour, GMYC, percent threshold). Inclusion of data from GenBank (additional 157 spp.) resulted in a more comprehensive library, but at a cost to success rate due to the increased number of singleton species. In addition to DNA barcodes, our study also provides supporting data in the form of specimen images, morphological characters, taxonomic bibliography, preserved vouchers, and nuclear rhodopsin sequences. Using this nuclear rhodopsin data we also uncovered evidence of interspecific hybridisation, and highlighted unrecognised diversity within popular aquarium species, including the endangered Indian barb Puntius denisonii. Conclusions/Significance: We demonstrate that DNA barcoding provides a highly effective biosecurity tool for rapidly identifying ornamental fishes. In cases where DNA barcodes are unable to offer an identification, we improve on previous studies by consolidating supplementary information from multiple data sources, and empower biosecurity agencies to confidently identify high-risk fishes in the aquarium trade. [ABSTRACT FROM AUTHOR]

The complete mitochondrial genome of three mustelid species, stoats (Mustela erminea), weasels (Mustela nivalis) and ferrets (Mustela furo), and the New Zealand fur seal (Arctocephalus forsteri) were sequenced using direct mitochondrial DNA extraction and overlapping long PCRs. The usual 37 mammalian mitochondrial genes (13 protein coding genes, 22 t-RNA and 2 r-RNA) were identified in all four mitogenomes. The divergence of stoats from other members of the sub-family Mustelinae was dated 4.5 million years ago. The mitogenomic data were consistent with a bear-like origin of seals. [ABSTRACT FROM AUTHOR]

The Philoceanus complex is a large assemblage of lice that parasitise procellariiform seabirds (petrels, albatrosses, and their relatives). We obtained mitochondrial 12S rRNA and cytochrome oxidase I DNA sequences from 39 species from diverse hosts and localities. Resolution of deeper relationships between genera was limited, however there is evidence for two major clades, one hosted by albatrosses, the other by petrels. Based on our results, the genera hosted by albatrosses are excellent candidates for detailed analysis of cospeciation. Our results also suggest that a previous estimate of a 5-fold difference in the relative rate of sequence evolution in lice and their avian hosts is an artefact of limited taxonomic sampling. [Copyright &y& Elsevier]

Few estimates of relative substitution rates, and the underlying mutation rates, exist between mitochondrial and nuclear genes in insects. Previous estimates for insects indicate a 2–9 times faster substitution rate in mitochondrial genes relative to nuclear genes. Here we use novel methods for estimating relative rates of substitution, which incorporate multiple substitutions, and apply these methods to a group of insects (lice, Order: Phthiraptera). First, we use a modification of copath analysis (branch length regression) to construct independent comparisons of rates, consisting of each branch in a phylogenetic tree. The branch length comparisons use maximum likelihood models to correct for multiple substitution. In addition, we estimate codon-specific rates under maximum likelihood for the different genes and compare these values. Estimates of the relative synonymous substitution rates between a mitochondrial (COI) and nuclear (EF-1α) gene in lice indicate a relative rate of several 100 to 1. This rapid relative mitochondrial rate (>100 times) is at least an order of magnitude faster than previous estimates for any group of organisms. Comparisons using the same methods for another group of insects (aphids) reveals that this extreme relative rate estimate is not simply attributable to the methods we used, because estimates from aphids are substantially lower. Taxon sampling affects the relative rate estimate, with comparisons involving more closely related taxa resulting in a higher estimate. Relative rate estimates also increase with model complexity, indicating that methods accounting for more multiple substitution estimate higher relative rates. [Copyright &y& Elsevier]

The delineation of species is important to the fields of evolution, ecology and conservation. The use of only a single line of evidence, e.g., morphology or a single gene sequence, may underestimate or overestimate the level of diversity within a taxon. This problem often occurs when organisms are morphologically similar but genetically different, i.e., for cryptic species. The Hadramphus genus contains four endangered, morphologically similar species of weevils, each endemic to a specific New Zealand region (Hadramphus spinipennis Chatham Islands, H. stilbocarpae Fiordland, H. tuberculatus McKenzie Country, H. pittospori Poor Knights Islands). The systematic relationships among these species are unclear. We used samples from these species and a closely related genus, Lyperobius huttoni, to obtain data from the mitochondrial gene cytochrome c oxidase subunit I and the nuclear gene internal transcribe spacer 2. In addition to the multi-locus coalescent approach, we modelled morphological characteristics combined with the genetic data. We found that H. spinipennis, H. tuberculatus and H. stilbocarpae were a closely related clade. Despite a strong morphological similarity, Hadramphus pittospori was found to be genetically distinct from the other Hadramphus species, which supports the resurrection of the monotypic genus Karocolens for this species. [ABSTRACT FROM AUTHOR]

The author discusses the alleged failure of researchers in biological science to include taxonomic identifications in the publication of their research. Topics include the use of morphologically-based and DNA-sequence identification methods, ways that an increase in citations would increase the impact factors and h-indices of taxonomic journals, and the burden of implementation resting with the editorial and peer-review practices of biology journals.

The Royal Entomological Society International Symposium on Insects and Disease was held at the University of Aberdeen on 10–12 September 2001. The symposium was run alongside the society''s Annual Meeting (the first time this meeting has been held outside England) in association with the International Union of Forest Research Organisations and the Entomological Society of The Netherlands. The symposium was divided into three sessions: The Insect–Host–Disease Interface; Ecology, Epidemiology and Modelling, and Insect Control in the New Millennium. [Copyright &y& Elsevier]

Predation is often difficult to observe or quantify for species that are rare, very small, aquatic or nocturnal. The assessment of such species’ diet can be conducted using molecular methods that target prey DNA remaining in predators’ guts and faeces. These techniques do not require high taxonomic expertise, are applicable to soft-bodied prey and allow for identification at the species level. However, for generalist predators, the presence of mixed prey DNA in guts and faeces can be a major impediment as it requires development of specific primers for each potential prey species for standard (Sanger) sequencing. Therefore, next generation sequencing methods have recently been applied to such situations. In this study, we used 454-pyrosequencing to analyse the diet of Powelliphanta augusta, a carnivorous landsnail endemic to New Zealand and critically endangered after most of its natural habitat has been lost to opencast mining. This species was suspected to feed mainly on earthworms. Although earthworm tissue was not detectable in snail faeces, earthworm DNA was still present in sufficient quantity to conduct molecular analyses. Based on faecal samples collected from 46 landsnails, our analysis provided a complete map of the earthworm-based diet of P. augusta. Predated species appear to be earthworms that live in the leaf litter or earthworms that come to the soil surface at night to feed on the leaf litter. This indicates that P. augusta may not be selective and probably predates any earthworm encountered in the leaf litter. These findings are crucial for selecting future translocation areas for this highly endangered species. The molecular diet analysis protocol used here is particularly appropriate to study the diet of generalist predators that feed on liquid or soft-bodied prey. Because it is non-harmful and non-disturbing for the studied animals, it is also applicable to any species of conservation interest. [ABSTRACT FROM AUTHOR]

We investigated the genetic structure of mitochondrial DNA (COI) and nuclear DNA (actin 5C) for variation within and among populations of two nurseryweb spider species: Dolomedes aquaticus Goyen, 1888 and D. minor Koch, 1876. Specimens were collected from intermediately disturbed braided rivers located in southern South Island, New Zealand. The genetic variation was compared against morphological characteristics to identify traits, both genetically and phenotypically, that indicate past occurrences of introgression. Haplotypes clearly assignable to D. aquaticus were also present in specimens of D. minor, supporting previous research suggesting introgression of mtDNA from D. aquaticus to D. minor. No evidence was found to indicate introgression from D. minor to D. aquaticus, suggesting that the introgression is asymmetrical and that the isolation mechanisms that may be in place to prevent such an occurrence are more successful within one species. In addition, the distribution patterns of identical haplotypes were found to provide an indication for when and where introgression took place. [ABSTRACT FROM AUTHOR]

Abstract: Several species of sap beetles in the genus Carpophilus are minor pests of fresh produce and stored products, and are frequently intercepted in biosecurity operations. In the South Pacific region, the superficially similar species C. maculatus and C. oculatus are frequently encountered in these situations. Three subspecies of C. oculatus have been described, and the complex of these four taxa has led to inaccurate identification and questions regarding the validity of these taxa. A molecular phylogenetic study using the mitochondrial gene cytochrome c oxidase I (COI) and two nuclear markers comprising the rDNA internal transcribed spacer 2 (ITS2) and the D1–D2 region of the large (28S) ribosomal RNA subunit showed that C. maculatus, and C. o. cheesmani were easily differentiated from the two other subspecies of C. oculatus. COI also showed differentiation between C. o. gilloglyi and C. o. oculatus, but this was not shown when third codon positions were removed and when RY-coding analyses were conducted. Generalised mixed Yule-Coalescent (GMYC) models were fitted to trees estimated from the COI data and were analysed using a multimodel approach to consider the evidence for three taxonomic groupings of the C. oculatus group. While the arrangement with the highest cumulative weight was not the arrangement ultimately accepted, the accepted taxonomy also had an acceptable level of support. ITS2 showed structure within C. oculatus, however C. o. oculatus was resolved as paraphyletic with respect to C. o. gilloglyi. COI showed evidence of sequence saturation and did not adequately resolve higher relationships between species represented in the dataset. 28S resolved higher relationships, but did not perform well at the species level. This study supports the validity of C. maculatus as a separate species, and provides sufficient evidence to raise C. o. cheesmani to the level of species. This study also shows significant structure within and between C. o. gilloglyi and C. o. oculatus, giving an indication of recent speciation events occurring. To highlight the interesting biology between these two taxa, C. o. gilloglyi is retained as a subspecies of C. oculatus. These results give clarity regarding the taxonomic status of C. maculatus and the subspecies of C. oculatus and provide a platform for future systematic research on Carpophilus. [Copyright &y& Elsevier]

Abstract: Restoration programs for human-disturbed ecosystems rely on a good understanding of how recovery occurs. This requires elucidating the underlying succession process, which depends on species adaptations, their interactions, and the spatiotemporal characteristics of the disturbance. Using spiders, we aim to identify the drivers of succession after burning, commonly used in New Zealand native tussock grasslands, test the hypothesis of post-burning dominance of generalists over specialists, and test the presumption that managed summer burns are more detrimental than spring burns. We established a 7-year experiment, with spring and summer burn treatments and unburned control plots, and sampled annually before and after the burning. We identified changes in spider assemblages and their drivers using clustering and indicator value analyses, and we analysed the response of spider diversity and taxa through linear mixed-effect models. In both spring and summer burns, functional groups and diversity decreased and the density of individuals of generalist species increased. However, spring and summer burn treatments showed no differences in spider diversity and composition. We found evidence that adaptations, such as habitat specificity and dispersal ability, determine the recovery of spider communities, and we suggest a set of indicators for monitoring programs. Time of burning may interact with factors like flammability and fire intensity, reducing differences in the effects of burning between seasons. We recommend planning burns for wet seasons as they are easier to control. To facilitate recolonisation by rare species of spiders of tussock grasslands, we suggest maintaining unburned areas that represent different vegetation types. [Copyright &y& Elsevier]

Abstract: Genetic diversity of 50 isolates of Neofusicoccum parvum, the predominant species of the Botryosphaeriaceae recovered from grapevines displaying symptoms of dieback and decline in New Zealand, was compared to that of isolates from Australia, South Africa, and California. The eight universally primed polymerase chain reaction (UP-PCR) primers distinguished 56 genotypes, with only four clonal pairs found. Seven main groups were identified in a neighbour-joining (NJ) tree with isolates from different regions and vineyards of New Zealand, Australia, and California distributed in different groups, indicating a high level of intra and intervineyard genetic variation. All of the South African isolates were positioned in a separate UP-PCR group, indicating that these isolates were less related to the other N. parvum isolates. When compared to fungi that reproduce sexually the genetic diversity and Shannon diversity indices were low (0.076–0.249; 0.109–0.367, respectively), genetic identity levels were high (0.76–0.95), and genetic distance levels were low (0.04–0.27). The large number of genotypes and the low number of clones in the New Zealand N. parvum populations may be explained by parasexual recombination as anastomosis was observed between nonself pairings. Pathogenicity tests using isolates from different UP-PCR groups inoculated onto either green shoots or 1-y-old grapevines detected virulence diversity, indicating intra and intervineyard variation between isolates, however, no correlation was detected between UP-PCR group and virulence. [Copyright &y& Elsevier]

Abstract: Ecosystem services provided by earthworms include increasing soil fertility, recycling organic material and provision of food for numerous predatory species. Estimating the role of earthworms in food webs is essential for a thorough evaluation of the ecosystem services they provide. Although their soft bodies are rapidly digested, earthworm remains can be detected via their chaetae in the faeces of predators. This study proposes a method for the identification of predated earthworm species using only predator faeces. We conducted morphological and molecular analyses of earthworm remains in the faeces of a highly endangered carnivorous land snail (Powelliphanta augusta) endemic to a restricted area of New Zealand. Microscopic examination of faecal strings from P. augusta showed the presence of earthworm chaetae in 47 of the 49 strings examined, suggesting an important role for earthworms in snail diets. Although earthworm tissue was not detectable in snail faeces, earthworm DNA was still present in sufficient quantity for molecular identification. Results showed that this land snail feeds on at least five species of New Zealand endemic earthworms. The method developed here is particularly appropriate for investigating the diet of potential invertebrate and vertebrate predators for which feeding behaviour is difficult to observe and gut content analysis is not possible. Such studies can help to develop a more complete understanding of the ecosystem services provided by earthworm communities. In this case it also helps the conservation of a rare and endangered predatory snail species. [Copyright &y& Elsevier]

The New Zealand fur seal (Arctocephalus forsteri) passed through a population bottleneck due to commercial sealing during the eighteenth to nineteenth centuries. To facilitate future management options, we reconstructed the demographic history of New Zealand fur seals in a Bayesian framework using maternally inherited, mitochondrial DNA sequences. Mitogenomic data suggested two separate clades (most recent common ancestor 5000 years ago) of New Zealand fur seals that survived large-scale human harvest. Mitochondrial haplotype diversity was high, with 45 singletons identified from 46 individuals although mean nucleotide diversity was low (0.012 ± 0.0061). Variation was not constrained geographically. Analyses of mitogenomes support the hypothesis for a population bottleneck approximately 35 generations ago, which coincides with the peak of commercial sealing. Mitogenomic data are consistent with a pre-human effective population size of approximately 30,000 that first declined to around 10,000 (due to the impact of Polynesian colonization, particularly in the first 100 years of their arrival into New Zealand), and then to 100-200 breeding individuals during peak of commercial sealing.

The taxonomy of the damselfly genus Xanthocnemis is revised, with particular focus on populations inhabiting the North Island of New Zealand. Earlier studies revealed two species: X. sobrina, restricted to cool, shaded streams in kauri forests and other forested areas, and X. zealandica, a common species throughout New Zealand except the Chatham and subantarctic islands. A field study encompassing aquatic habitats throughout the whole North Island was carried out to establish the relationship between morphological variation (body size and various morphological traits over the entire body) observed by previous researchers with ecological conditions and/or geographical location. The main aim was to propose reliable diagnostic features that could be used in future studies. Morphological and molecular variation was assessed. Morphological examination included assigning landmarks for all body parts corresponding to the external morphological features that are usually used in Odonata taxonomy. Molecular analysis targeted fragments of the 28S and 16S rRNA genes. Congruence was sought between both types of data, statistical support for two morphological types previously described as different species and a maximum likelihood phylogenetic tree in conjunction with a pairwise genetic distance matrix constructed from the DNA sequences obtained from the sampled specimens. Geometric morphometrics revealed statistically significant differentiation between specimens identified as X. zealandica and X. sobrina for four traits: (1) dorsal view of the head for both sexes as well as male appendages from (2) dorsal, (3) ventral and (4) lateral views. Wings appeared different when analysed for males only. Molecular analysis, however, grouped all specimens into a single undifferentiated cluster with very low mean pairwise distance (&lt;0.01) between them showing almost no variation at the molecular level among the sampled populations on the North Island. Therefore, an additional analysis of the mitochondrial cytochrome c-oxidase I gene was carried out comparing randomly selected North Island specimens to Xanthocnemis specimens targeted in other molecular studies (Nolan et al. 2007, Amaya-Perilla et al. 2014). The analysis of the COI gene confirmed that all North and South Island isolates of Xanthocnemis cluster together in a well-supported clade with pairwise identity &gt;96% and ~93% pairwise identity with X. tuanuii sequences obtained from the Chatham Island specimens. A careful investigation of the thin plate spline deformations generated for the geometric morphometric landmarks showed that the significant variations in the appendages of the Xanthocnemis specimens appeared to be the result of size, rather than shape, differences. Therefore, X. sobrina is proposed as a synonym of X. zealandica. Recently Amaya-Perilla et al. (2014) synonymised X. sinclairi with X. zealandica and confirmed the status of the Chatham Island X. tuanuii as a distinct species. It is therefore proposed that the genus Xanthocnemis consists of two species only: zealandica occurring all over the North, South and Stewart Islands, and tuanuii, endemic to Chatham and Pitt islands. Considering several statistical tests involving body measurements and ecological variables recorded during the field study, as well as various discussion points from similar studies of other species of Odonata, two alternative hypotheses are proposed for future testing. The first hypothesis synonymises X. sobrina with X. zealandica and suggests a possible explanation for the evolution of the two morphological traits that have previously been considered diagnostic for these species. The second hypothesis suggests that as typical X. sobrina were not sampled during this study this could represent a species that is now extinct, unless future studies prove it otherwise.

Human alteration of islands has made restoration a key part of conservation management. As islands are restored to their original state, species interactions change and some populations may be impacted. In this study we examine the coxella weevil, (Hadramphus spinipennis Broun) and its host-plant Dieffenbach's speargrass (Aciphylla dieffenbachii Kirk), which are both open habitat specialists with populations on Mangere and Rangatira Islands, Chathams, New Zealand. Both of these islands were heavily impacted by the introduction of livestock; the majority of the forest was removed and the weevil populations declined due to the palatability of their host-plant to livestock. An intensive reforestation program was established on both islands over 50 years ago but the potential impacts of this restoration project on the already endangered H. spinipennis are poorly understood. We combined genetic and population data from 1995 and 2010-2011 to determine the health and status of these species on both islands. There was some genetic variation between the weevil populations on each island but little variation within the species as a whole. The interactions between the weevil and its host-plant populations appear to remain intact on Mangere, despite forest regeneration. A decline in weevils and host-plant on Rangatira does not appear to be caused by canopy regrowth. We recommend that (1) these populations be monitored for ongoing effects of long-term reforestation, (2) the cause of the decline on Rangatira be investigated, and (3) the two populations of weevils be conserved as separate evolutionarily significant units.

The role of species' interactions in structuring biological communities remains unclear. Mutualistic symbioses, involving close positive interactions between two distinct organismal lineages, provide an excellent means to explore the roles of both evolutionary and ecological processes in determining how positive interactions affect community structure. In this study, we investigate patterns of co-diversification between fungi and algae for a range of New Zealand lichens at the community, genus, and species levels and explore explanations for possible patterns related to spatial scale and pattern, taxonomic diversity of the lichens considered, and the level sampling replication. We assembled six independent datasets to compare patterns in phylogenetic congruence with varied spatial extent of sampling, taxonomic diversity and level of specimen replication. For each dataset, we used the DNA sequences from the ITS regions of both the fungal and algal genomes from lichen specimens to produce genetic distance matrices. Phylogenetic congruence between fungi and algae was quantified using distance-based redundancy analysis and we used geographic distance matrices in Moran's eigenvector mapping and variance partitioning to evaluate the effects of spatial variation on the quantification of phylogenetic congruence. Phylogenetic congruence was highly significant for all datasets and a large proportion of variance in both algal and fungal genetic distances was explained by partner genetic variation. Spatial variables, primarily at large and intermediate scales, were also important for explaining genetic diversity patterns in all datasets. Interestingly, spatial structuring was stronger for fungal than algal genetic variation. As the spatial extent of the samples increased, so too did the proportion of explained variation that was shared between the spatial variables and the partners' genetic variation. Different lichen taxa showed some variation in their phylogenetic congruence and spatial genetic patterns and where greater sample replication was used, the amount of variation explained by partner genetic variation increased. Our results suggest that the phylogenetic congruence pattern, at least at small spatial scales, is likely due to reciprocal co-adaptation or co-dispersal. However, the detection of these patterns varies among different lichen taxa, across spatial scales and with different levels of sample replication. This work provides insight into the complexities faced in determining how evolutionary and ecological processes may interact to generate diversity in symbiotic association patterns at the population and community levels. Further, it highlights the critical importance of considering sample replication, taxonomic diversity and spatial scale in designing studies of co-diversification.

Predation is often difficult to observe or quantify for species that are rare, very small, aquatic or nocturnal. The assessment of such species' diet can be conducted using molecular methods that target prey DNA remaining in predators' guts and faeces. These techniques do not require high taxonomic expertise, are applicable to soft-bodied prey and allow for identification at the species level. However, for generalist predators, the presence of mixed prey DNA in guts and faeces can be a major impediment as it requires development of specific primers for each potential prey species for standard (Sanger) sequencing. Therefore, next generation sequencing methods have recently been applied to such situations. In this study, we used 454-pyrosequencing to analyse the diet of Powelliphantaaugusta, a carnivorous landsnail endemic to New Zealand and critically endangered after most of its natural habitat has been lost to opencast mining. This species was suspected to feed mainly on earthworms. Although earthworm tissue was not detectable in snail faeces, earthworm DNA was still present in sufficient quantity to conduct molecular analyses. Based on faecal samples collected from 46 landsnails, our analysis provided a complete map of the earthworm-based diet of P. augusta. Predated species appear to be earthworms that live in the leaf litter or earthworms that come to the soil surface at night to feed on the leaf litter. This indicates that P. augusta may not be selective and probably predates any earthworm encountered in the leaf litter. These findings are crucial for selecting future translocation areas for this highly endangered species. The molecular diet analysis protocol used here is particularly appropriate to study the diet of generalist predators that feed on liquid or soft-bodied prey. Because it is non-harmful and non-disturbing for the studied animals, it is also applicable to any species of conservation interest.

Spider: SPecies IDentity and Evolution in R is a new R package implementing a number of useful analyses for DNA barcoding studies and associated research into species delimitation and speciation. Included are functions essential for generating important summary statistics from DNA barcode data, assessing specimen identification efficacy, and for testing and optimizing divergence threshold limits. In terms of investigating evolutionary and taxonomic questions, techniques for assessing diagnostic nucleotides and probability of reciprocal monophyly are also provided. Additionally, a sliding window function offers opportunities to analyse information across a gene, essential for marker design in degraded DNA studies. Spider capitalizes on R's extensible ethos and offers an integrated platform ideal for the analysis of both nucleotide and morphological data. The program can be obtained from the comprehensive R archive network (CRAN, http://cran.r-project.org) and from the R-Forge package development site (http://spider.r-forge.r-project.org/)., (© 2012 Blackwell Publishing Ltd.)