The influence of weather upon the activity of slugs

Summary The activity of slugs was measured by a catch per unit effort sampling system based upon night searching, and was related to the microclimatological conditions in the habitat by regression analysis. The activity of Arion hortensis Fér., Arion subfuscus (Drap.), Arion lusitanicus Mab., Milax budapestensis (Hazay), and of all slugs irrespective of species, was found to be best related to temperature and vapour pressure deficit. The relevance of these results to the application of effective methods for the control of slugs is discussed.     

Molecular identification of the exotic slug <Emphasis Type="Italic">Arion subfuscus</Emphasis> sensu stricto (Gastropoda: Pulmonata) in California,with comments on the source location of introduced populations

Arion subfuscus sensu lato (s. l.) refers to a cryptic slug species complex that is native to Europe and which, from there, has been introduced into the northeastern U.S. The species complex was detected in California for the first time in 2005 and recently 12 specimens were collected during statewide surveys. The genital morphology of these specimens suggested that only A. subfuscus sensu stricto (s. s.) was present and partial sequences of mitochondrial 16S rDNA (443 bp) showed that they all belonged to a single haplotype of the mitochondrial lineage, S1 sensu Pinceel et al. (Genetica 125: 155–171, 2005). This result was corroborated by a parallel analysis of a 655 bp COI DNA barcode. The 16S rDNA S1 haplotype (S1-03) of the Californian specimens is hitherto known only from the northeastern U.S. Hence, it is likely that populations may have been introduced to California from the northeastern states, rather than directly from the native area of the S1 haplogroup in Europe.     

CONCENTRATIONS OF METALS (Cu, Pb, Cd, Zn, Ca) IN SIX SPECIES OF BRITISH TERRESTRIAL GASTROPODS NEAR A DISUSED LEAD AND ZINC MINE

Heavy metal concentrations in 6 terrestrial slug species inhabitinga disused Pb/Zn mine site were measured by atomic absorptionspectrophotometry. Differences in metal accumulation patternswere such that the species could be ranked in order of overallmetal accumulation capability: Arion hortensis < Deocerasreticulatum < Milax budapestensis < Arion ater < Derocerascaruanac < Arion subfuscus. Species belonging to the samegenus were widely separated by this simple ranking scheme. Theresults are discussed in the context of inter-species competitionand resource partitioning, and cautionary comments are madeconcerning the utility of slugs as biomonitors of terrestrialheavy metal pollution. (Received 25 November 1987; accepted 2 January 1988)     

Population Genetics and Identity of an Introduced Terrestrial Slug: Arion subfuscus s.l. in the North-east USA (Gastropoda, Pulmonata, Arionidae)

Several European species of the terrestrial slug genus Arion have been introduced into North America. A case in point is the species complex A. subfuscus s.l. which has become one of the most abundant slug taxa in North America. In Europe this complex consists of at least two cryptic species, viz. A. fuscus and A. subfuscus s.s., the latter of which is further subdivided in five strongly divergent mtDNA lineages (A. subfuscus S1–S5). In order to determine which of these A. subfsucus s.l. taxa are present in the NE USA and in order to assess their population genetic structure, we compared mtDNA, nDNA and allozyme variation between populations from the NE USA and Europe. Our results show that (1) at least A. subfuscus S1 has become successfully established in the NE USA, (2) founder effects are the most likely explanation for the loss of a large amount of molecular genetic variation in populations from the NE USA (i.e. a loss of 96% of the 16S rDNA haplotypes, 67% of the ITS1 alleles and 46% of the alleles at polymorphic allozyme loci), and (3) part of the remaining genetic variation in NE USA populations was probably due to multiple introductions from the British Isles and the European mainland, and the hybrid structure of most of these source populations. Apparently, the extreme loss of molecular genetic variation in this introduced species has not prevented it from successfully establishing and spreading in novel environments.     

Association of slugs with the fungal pathogen Epichloë typhina (Ascomycotina: Clavicipitaceae): potential role in stroma fertilisation and disease spread

Epichloë spp. are endophytes of grasses, and form epiphytic external stromata on flowering tillers. E. typhina was first noticed infecting Dactylis glomerata (= orchardgrass, cocksfoot) stands in the Willamette Valley in 1996, and soon became the primary factor limiting the longevity of seed production fields. Several species of slugs are present in these fields, and we investigated their role in E. typhina biology. Pre‐dawn surveys of D. glomerata fields in 2009 and 2010 found Prophysaon andersoni and Arion subfuscus slugs feeding on the fungal stromata. When unfertilised and fertilised immature stromata predominated, approximately 80% of the individuals of these two species that were seen on plants were found on the stromata. As the majority of stromata reached maturity the presence of these species on stromata declined to between 20–40%. The common agricultural slug pest, Deroceras reticulatum, was on stromata only 20% of the time early in the season, and declined to <5% at stromata maturity. Observations of frass from slugs determined that the most common constituent was the food sources upon which the slug species was usually found during these surveys. Typically 100% of the frass from P. andersoni and A. subfuscus contained stroma material, compared to 25% for D. reticulatum. Spermatia, and ascospores later in the season, were commonly seen in the frass of slugs that consumed stromata. Some slugs that had no stroma material in their frass appeared to have consumed spermatia and ascospores from the leaf surface. A multiple‐choice laboratory test confirmed the different proportional preferences of P. andersoni and D. reticulatum for stroma (0.72 vs 0.20) and leaf (0.07 vs 0.38), respectively. Two laboratory multiple‐choice tests, and a field survey, found that P. andersoni preferred unfertilised and immature stroma over mature stroma. D. reticulatum is the most common and abundant slug in Willamette Valley grass seed fields, yet it is the least likely to move spermatia between unfertilised stromata, or ascospores to uninfected plants. P. andersoni and A. subfuscus are mycophagous, frequently transport viable spermatia and ascospores in their frass; yet they are generally confined to field edges. Data and observations suggest the role of slugs in the epidemiology of E. typhina is small compared to other factors.     

Phylogeny of slug species of the genus Arion: evidence of monophyly of Iberian endemics and of the existence of relict species in Pyrenean refuges

The Iberian Peninsula contains the majority of the Paleartic land slug species of the genus Arion, which exhibits diverse taxonomic problems. The present study investigated Arion taxonomy on the basis of analyses of the mitochondrial ND1 gene and nuclear internal transcribed spacer 1 (ITS1) sequences. The Iberian endemic species were monophyletically clustered in two divergent sister clades. The topotype specimens of Arion lusitanicus and the closely related species Arion nobrei and Arion fuligineus, as well as Arion hispanicus and Arion flagellus, were grouped into an ‘Atlantic’ clade, whereas Arion baeticus, Arion gilvus, Arion anguloi, Arion wiktori and Arion paularensis were included in a ‘Continental–Mediterranean’ clade. Calibration of mutation rate in the ND1 gene suggested that the divergence of these two clades occurred around the Pliocene–Pleistocene boundary, with subsequent speciation events during the Pleistocene. A group of ancestral and divergent endemic species with distribution centred in the Pyrenean mountain range (Arion molinae, Arion lizarrusti, Arion antrhacius and Arion iratii) arose in the Pliocene and survived through the Pleistocene in geographically confined small populations. Arion lusitanicus showed up to be polyphyletic: specimens, sampled outside the geographic range of the topotype in the north‐western Iberian Peninsula, were included in a non‐monophyletic clade together with the widely distributed species Arion ater and Arion rufus. The divergent species with a wide European distribution (Arion subfuscus, Arion hortensis, Arion fagophilus and Arion intermedius) were located in basal positions in all topologies. The evolutionary history of these slug species (highly sensitive to climatic factors, with capacity for both outcrossing and selfing, and with low dispersal ability) appears to have been moulded by Pliocene–Pleistocene climate events and by the rugged topography of southern Europe, giving rise to repeated cycles of population isolation during periods of glaciation alternating with interglacial expansions limited by geographic barriers.     

Genetic and chaetal variation in Nais worms (Annelida,Clitellata, Naididae)

The genus Nais is a group of oligochaetous clitellates, common in eutrophic freshwater habitats. About 30 species are described. Species identification is based primarily on chaetal characters, which are often subtle, inconsistent, and even overlapping between nominal species. We investigated the correlation between genetic variation and chaetal morphology in this genus. Eighty‐one individuals from Europe, North America, and China were included in the study. Seventy‐five of these were preserved as vouchers. They were scrutinized with regard to chaetal morphology, and ten different morphotypes were identified. Three molecular markers, two mitochondrial (the COI gene and 16S rDNA) and one nuclear (the ITS region), were used to establish the genetic lineages in the material. Genetic variation was found to be largely congruent with chaetal character patterns. However, at least nine separately evolving lineages (all supported by mitochondrial as well as nuclear data) correspond to at most six nominal species. Four morphotypes/lineages are recognized as Nais barbata, Nais christinae, Nais elinguis, and Nais stolci, respectively, whereas five, or possibly more, lineages represent a morphological continuum covering the variation of the Nais communis/variabilis complex. Thus, cryptic speciation is revealed. Our results indicate that a taxonomic revision of the genus will be needed in the future.     

Effects of seed-bed conditions on slug numbers and damage to winter wheat in a clay soil

In a field experiment three levels of consolidation were combined with three seed-bed tilths, to produce nine different types of seed-bed. The soil properties of each combination of tilth and consolidation were measured and effects on the numbers and biomass of slugs in the top 10 cm of soil and on damage to wheat seeds and seedlings were assessed. Eight species of slug were present, with three species predominating (Deroceras reticulatum, Arion distinctus and Arion subfuscus). Between 3% and 33% of seeds and seedlings were killed by slugs (recorded at Growth Stage 12). Contrary to expectation, most damage occurred on consolidated seed-beds of fine or medium tilth, least on loose seed-beds of the same texture; intermediate damage occurred on cloddy seed-beds, where consolidation had little effect. The level of damage occurring on different types of seed-bed was directly related to the biomass of slugs in the top 10 crn of soil, and was inversely related to depth of sowing and the percentage of fine soil in the seed-bed. These three factors together accounted for 94–97% of the variance in slug damage. Consolidation was associated with increased slug damage probably because of its effects on these three factors: slug biomass was greater and seed was at shallower depth in consolidated than loose seed-beds and consolidation, whether before or after drilling, failed to break down clods of soil into finer particles. This is the first experimental evidence in the field of the effects of seedbed conditions on slug numbers and damage to winter wheat and represents a significant step towards forecasting and avoiding slug damage in this crop.     

Chromosomal Study of Colostethus brunneus from the Type Locality and Two Related Species (Anura – Dendrobatidae)

In this paper, we report a chromosomal study of three Brazilian species of Colostethus, C. brunneus from the type locality, Colostethus sp. (aff. trilineatus), and Colostethus sp., which is morphologically similar to C. brunneus. The diploid number for C. brunneus was 2n = 24 chromosomes, in agreement with that previously described for specimens from Peru. Colostethus sp. (aff. trilineatus) and Colostethus sp. showed a very similar karyotype with 22 chromosomes. The NOR was located on pair 3 in C. brunneus, on pair 4 in Colostethus sp. (aff. trilineatus), and on pair 2 in Colostethus sp. In one specimen of Colostethus sp., an additional NOR site was located on pair 7 in only one of the homologs. This extra Ag-NOR site was confirmed by FISH using an rDNA probe. In addition to the NOR location, the C-banding pattern was also species-specific, despite the similar chromosomal morphology of the species. These results indicate that although these species may be closely related, there is a clear dichotomy in their chromosome number.     

Ecological Differentiation of Cryptic Species within an Asexual Protist Morphospecies: A Case Study of Filamentous Green Alga Klebsormidium (Streptophyta)

Taxa of microbial eukaryotes defined on morphological basis display a large degree of genetic diversity, implying the existence of numerous cryptic species. However, it has been postulated that genetic diversity merely mirrors accumulation of neutral mutations. As a case taxon to study cryptic diversity in protists, we used a widely distributed filamentous genus, Klebsormidium, specifically the lineage E (K. flaccidum/K. nitens complex) containing a number of morphologically similar strains. Fourteen clades were recognized in the phylogenetic analysis based on a concatenated ITS rDNA + rbcL data set of more than 70 strains. The results of inferred character evolution indicated the existence of phylogenetic signal in at least two phenotypic characters (production of hydro‐repellent filaments and morphology of zoosporangia). Moreover, the lineages recovered exhibited strong ecological preferences to one of the three habitat types: natural subaerial substrata, artificial subaerial substrata, and aquatic habitats. We interpret these results as evidence of existence of a high number of cryptic species within the single morphospecies. We consider that the permanent existence of genetically and ecologically well‐defined cryptic species is enabled by the mechanism of selective sweep.     

Molecular circumscription of the blackfly Simulium oyapockense from South America

Problems related to the identity of Simuliidae species are impediments to effective disease control in Amazonia. Some of these species, such as Simulium oyapockense Floch & Abonnenc, 1946 (Diptera: Simuliidae), are vectors of the organisms that cause onchocerciasis and mansonellosis diseases. This blackfly species has a wide distribution in South America, and it is suspected of being a complex of cryptic species. The aim of this study is to characterize the nominal species S. oyapockense using partial COI gene sequences. Seven populations of S. oyapockense (morphologically identified) were analysed, including one from its type-locality. The other six populations were collected in Brazil and in Argentina. A taxon collected in Amazonas state, Brazil, with adults similar to S. oyapockense but with distinct pupae, was also included in the analysis (Simulium ‘S’). The nominal species S. oyapockense is circumscribed, and its geographical distribution is restricted to areas north of the Amazon River. Populations of S. oyapockense s.l. collected south of the Amazon River comprise a species complex that needs to be evaluated using integrative taxonomy. Simulium ‘S’ represents a species with unique morphological and molecular characteristics. Distinguishing cryptic species is a prerequisite for reducing the taxonomic impediment, especially in medically important taxa.     

The palatability of 78 wildflower strip plants to the slug Arion lusitanicus

As Arion lusitanicus has been observed in high densities in wildflower strips, the palatability of the foliage of 78 species of sown and naturally occurring plants found in such strips was investigated. In the laboratory, the slugs were offered a surplus of fresh, mature leaves and, after 16 h, a consumption index was calculated to rank the plants according to their palatability to the slugs. Arion lusitanicus showed clear preferences for particular plant species, but more than two thirds of all plants investigated were scarcely eaten. Brassica napus was most eaten, followed by Papaver rhoeas, Capsella bursa-pastoris and Lamium purpureum. Overall, A. lusitanicus preferred sown species to naturally occurring ones and preferred annuals to perennials.     

A polyphasic approach to the delimitation of diatom species: a case study for the genus Pinnularia (Bacillariophyta)

Diatoms are one of the most abundant and arguably the most species‐rich group of protists. Diatom species delimitation has often been based exclusively on the recognition of morphological discontinuities without investigation of other lines of evidence. Even though DNA sequences and reproductive experiments have revealed several examples of (pseudo)cryptic diversity, our understanding of diatom species boundaries and diversity remains limited. The cosmopolitan pennate raphid diatom genus Pinnularia represents one of the most taxon‐rich diatom genera. In this study, we focused on the delimitation of species in one of the major clades of the genus, the Pinnularia subgibba group, based on 105 strains from a worldwide origin. We compared genetic distances between the sequences of seven molecular markers and selected the most variable pair, the mitochondrial cox1 and nuclear encoded LSU rDNA, to formulate a primary hypothesis on the species limits using three single‐locus automated species delimitation methods. We compared the DNA‐based primary hypotheses with morphology and with other available lines of evidence. The results indicate that our data set comprised 15 species of the P. subgibba group. The vast majority of these taxa have an uncertain taxonomic identity, suggesting that several may be unknown to science and/or members of (pseudo)cryptic species complexes within the P. subgibba group.     

The rhabditid nematode Phasmarhabditis hermaphrodita as a potential biological control agent for slugs

A nematode, Phasmarhabditis hermaphrodita, known to be associated with slugs but not previously thought to be parasitic, was shown to be a parasite capable of killing the pest slug Deroceras reticulatum. The parasite infects slugs in the area beneath the mantle surrounding the shell, causing a disease with characteristic symptoms, particularly swelling of the mantle. Infection leads to death of the slug, usually between seven and 21 days afterwards. The nematode then spreads and multiplies in the cadaver. In an experiment where individual D. reticulatum were exposed to different numbers of P. hermaphrodita, a significant positive relationship was found between nematode dose and slug mortality. In two experiments on host range, the nematode was found to infect and kill all pest slug species tested: Deroceras caruanae, Arion distinctus, Arion silvaticus, Arion intermedius, Arion ater, Tandonia sowerbyi and T. budapestensis, in addition to D. reticulatum.     

Molecular phylogeny of Glossodoris (Ehrenberg, 1831) nudibranchs and related genera reveals cryptic and pseudocryptic species complexes

Chromodorid nudibranchs (Chromodorididae) are brightly coloured sea slugs that live in some of the most biodiverse and threatened coral reefs on the planet. However, the evolutionary relationships within this family have not been well understood, especially in the genus Glossodoris. Members of Glossodoris have experienced large‐scale taxonomic instability over the last century and have been the subject of repeated taxonomic changes, in part due to morphological characters being the sole traditional taxonomic sources of data. Changing concepts of traditional generic boundaries based on morphology also have contributed to this instability. Despite recent advances in molecular systematics, many aspects of chromodorid taxonomy remain poorly understood, particularly at the traditional species and generic levels. In this study, 77 individuals comprising 32 previously defined species were used to build the most robust phylogenetic tree of Glossodoris and related genera using mitochondrial genes cytochrome c oxidase subunit I and 16S, and the nuclear gene 28S. Bayesian inference, maximum likelihood, and maximum parsimony analyses verify the most recent hypothesized evolutionary relationships within Glossodoris. Additionally, a pseudocryptic and cryptic species complex within Glossodoris cincta and a pseudocryptic complex within Glossodoris pallida emerged, and three new species of Doriprismatica are identified.     

Phylogeny of macrophagous leeches (Hirudinea,Clitellata) based on molecular data and evaluation of the barcoding locus

Oceguera‐Figueroa, A., Phillips, A. J., Pacheco‐Chaves, B., Reeves, W. K. & Siddall, M. E. (2010). Phylogeny of macrophagous leeches (Hirudinea, Clitellata) based on molecular data and evaluation of the barcoding locus. —Zoologica Scripta, 40, 194–203. The phylogenetic relationships of macrophagous leech species are studied using two mitochondrial [cytochrome c oxidase subunit I (COI) and 12S rDNA] and two nuclear (28S rDNA and 18S rDNA) markers. The complete dataset analysed in this study included 49 terminals and 5540 aligned characters. Phylogenetic analyses were performed under two optimality criteria: Maximum Parsimony and Maximum Likelihood. The monophyly of the two currently recognized families (i.e. Erpobdellidae and Salifidae) is confirmed and well supported. The phylogenetic position of Gastrostomobdellidea is studied for the first time and found to be sister to family Salifidae nested well within Erpobdelliformes. Previously recognized taxonomic arrangements were evaluated and discarded through successive constraint analyses. Correlation between morphology and phylogeny was notable in Salifidae but not in Erpobdellidae. Variability of COI, the barcoding locus, was examined across species leading to the recognition of the invasive Barbronia weberi in Mexico, Costa Rica, Germany, South Africa and Taiwan.     

Species delimitation methods reveal cryptic diversity in the Hypnea cornuta complex (Cystocloniaceae,Rhodophyta)

The simple and convergent morphologies of many red algae make these species difficult to identify using traditional morphological characters. Many cryptic species have been described in recent years based on molecular datasets, and this has led to the application of an integrative taxonomy approach in species delimitation. Here, we performed several species delimitation methods (mBGD, ABGD, SPN, PTP, GMYCs and GMYCm) based on two different loci (COI-5P and rbcL) in species of the Hypnea cornuta complex. These methods were combined with morphological and phylogenetic data, extensive sampling, analysis of topotype material, and historically relevant herbarium samples. Our findings demonstrate that the groups morphologically assigned to H. cornuta and H. stellulifera consist of five different cryptic species. H. cornuta is a polyphyletic taxon composed of three well-separated lineages, thus requiring sequencing of type or topotype specimens to determine which one is Hypnea cornuta sensu stricto. We have revealed that the distribution of H. stellulifera is limited to Asia, while the Brazilian specimens initially assigned to this species were clarified as a new endemic species: Hypnea cryptica sp. nov. Our results indicated that only an integrative approach combining several lines of evidence, including morphology, nomenclature history, molecular data, biogeography and ecology can correctly solve the taxonomic status of widely distributed cryptic species.