Eduardo de la Peña

Physiological integration increases the survival and growth of the clonal invader Carpobrotus edulis.

Clonal growth seems to be a common trait for many of the most aggressive invasive plant species. However, little research has been conducted to determine the role of clonality in the successful invasion of new areas by exotic species. Carpobrotus edulis (L.) N.E. Br. is a mat-forming succulent plant, native to South Africa that is invasive in coastal dunes of Australia, New Zealand, USA and Southern Europe. Although Carpobrotus edulis is a clonal plant, there is no information on the role of clonality for the invasion by this species, therefore the objective of this study was to test whether or not physiological integration improves the performance of C. edulis invading coastal sand dunes. To do that, a 6-month field experiment was designed in which the stolon connections between the apical ramets and the C. edulis mats were severed to prevent physiological integration. This treatment was applied to ramets growing under high and low competition with the native species. Apical ramets with intact stolon connections were used as control. Integration improved the survivorship and growth of apical ramets, both in high and low competition. Connected ramets showed a more pronounced increase of clonal growth (estimated as stolon length) during the experimental period and a higher total biomass and number of ramets at the completion of the experiment. In terms of survivorship, the benefit of integration was greater under high competition. Physiological integration can therefore be considered an important factor in the invasiveness of C. edulis, both in open space and in direct competition with the native plants.

Stage sensitivity of medaka (Oryzias latipes) eggs and embryos to permethrin.

The effects of exposure to permethrin on gametes, fertilization and embryonic development were examined in medaka (Oryzias latipes). Following range finding (25, 50, 100, 200 or 300 microg/l) and duration of exposure (0, 120, 144, 168, 192, 216, or 240-h) assays, the relative sensitivity was studied when initiation of exposure to permethrin (100 microg/l, for 192-h) occurred at one of four different stages, i.e., unfertilized egg (0-h), late morula (5-h), early neurula (24-h), and early organogenesis (40-h). The later exposure interval proved the most sensitive. Also, differences were observed in rates of recovery in larvae initially affected following the earliest exposure treatment (0-h, gametes prior to fertilization). Permethrin (100 microg/l) did not affect fertilization success and no lethal effects were observed in embryos. Sublethal effects were primarily observed at hatch. Toxicity endpoints in larvae included: delayed swim bladder inflation; inability of hatchling to respond to stimulus; uncoordinated movements, myoskeletal defects and transient enlargement of gall bladder. These changes were characteristic for all hatchlings exposed to nominal concentrations of 50 microg/l. While certain of the above alterations were reversed within 72-h after hatching, lack of swim bladder inflation and inability to respond to stimuli were two features that persisted with significant incidences. Based on persistence of sublethal effects, results from this work indicate the importance of exposures to gametes and to embryos prior to water hardening. The approach taken herein may better reflect environmental risk conditions than assays limited to exposure of embryonated eggs.

Plant-soil feedback as a mechanism of invasion by Carpobrotus edulis

Invasive plant species have been suggested to change the composition of the soil community in a way that results in a positive feedback for them and a negative feedback for the native plant community. Carpobrotus edulis, a species native to South Africa, is one of the most aggressive exotic species in Mediterranean Europe. Although several aspects of its invasion biology have been studied, the occurrence of plant-soil feedback has been scarcely investigated. We first checked for the existence of biotic resistance in soils from two invaded sites of Mediterranean Europe and one site in the native area. Secondly, we evaluated the effects of soil conditioning on the germination and plant growth of C. edulis and two key species of native dunes. Finally, we tested the effects of short- and long-term soil conditioning on the performance and reproductive effort of C. edulis. Our results show that at first there is a natural resistance to invasion by the soil biota. Later, biotic resistance in invaded soil is suppressed by the establishment of a soil community that enhances the growth of C. edulis and that negatively influences the growth and survival of the native plants. Long-term soil conditioning in the field resulted in shifts in the balance between vegetative growth and sexual reproduction. Long-term invasion was also reflected in high levels of endophyte colonization by chytrids in roots, although the physiological consequences of this colonization remain unknown. The results obtained illustrate a mechanism that explains how C. edulis breaks the initial biotic resistance of newly-invaded landscapes. Finally, this study highlights the importance of studying plant-soil interactions on different members of the plant community and temporal stages in order to fully understand invasion.

Local adaptation of aboveground herbivores towards plant phenotypes induced by soil biota.

Background Soil biota may trigger strong physiological responses in plants and consequently induce distinct phenotypes. Plant phenotype, in turn, has a strong impact on herbivore performance. Here, we tested the hypothesis that aboveground herbivores are able to adapt to plant phenotypes induced by soil biota. Methodology and Principal Findings We bred spider mites for 15 generations on snap beans with three different belowground biotic interactions: (i) no biota (to serve as control), (ii) arbuscular mycorrhizal fungi and (ii) root-feeding nematodes. Subsequently, we conducted a reciprocal selection experiment using these spider mites, which had been kept on the differently treated plants. Belowground treatments induced changes in plant biomass, nutrient composition and water content. No direct chemical defence through cyanogenesis was detected in any of the plant groups. Growth rates of spider mites were higher on the ecotypes on which they were bred for 15 generations, although the statistical significance disappeared for mites from the nematode treatment when corrected for all multiple comparisons. Conclusion/Significance These results demonstrate that belowground biota may indeed impose selection on the aboveground insect herbivores mediated by the host plant. The observed adaptation was driven by variable quantitative changes of the different separately studied life history traits (i.e. fecundity, longevity, sex-ratio, time to maturity).

Genotoxicity of the insecticide rotenone in cultured human lymphocytes

We have investigated the genotoxic activity of rotenone on three genetic endpoints, sister-chromatid exchanges (SCE), chromosome aberrations (CA) and micronuclei (MN) in human lymphocyte cultures in the presence and absence of a metabolic activation system (S9 mix). Our results indicate that rotenone increases the frequency of binucleated micronucleated (BNMN) cells and causes a delay in the cell cycle but does not increase the frequency of CA and SCE at the concentrations used. The presence of S9 mix reduces the genotoxic activity of rotenone.

Analysis of the specificity of three root-feeders towards grasses in coastal dunes

Among the root-feeding nematodes that accumulate in the rhizosphere of grasses in European dunes, the genus Pratylenchus is of special relevance given its diversity and distribution. Although different species of Pratylenchus have been reported in dune grasses, the specificity towards dune plants, a fundamental aspect of the biology of the species, has hitherto not been studied. Two inoculation experiments using different combinations of grasses and nematodes were performed. The multiplication and the effect on plant growth of P. dunensis and P. brzeskii, two species which only occur in dune areas was compared with that of P. penetrans, a broad host-range species. The three Pratylenchus spp. could multiply under all hosts; however, there was a clear host-dependent response. The species-specific response observed might account for the shift of Pratylenchus spp. detected in the field. Although, a negative effect on the growth of A. arenaria was demonstrated for the three nematode species, different densities were needed to observe the same effects in plant biomass which point at nematode-specific tolerance. While the typical dune species needed very high densities to produce damage, P. penetrans needed very few specimens. The results obtained indicate that species with similar feeding adaptations show very different multiplication abilities on co-occurring hosts, an aspect that is usually overlooked for belowground herbivores in natural systems. The obtained results might suggest a coevolutionary relationship between specific nematode species and Ammophila arenaria.

Relative Importance of Biotic and Abiotic Soil Components to Plant Growth and Insect Herbivore Population Dynamics

Background Plants are affected by several aspects of the soil, which have the potential to exert cascading effects on the performance of herbivorous insects. The effects of biotic and abiotic soil characteristics have however mostly been investigated in isolation, leaving their relative importance largely unexplored. Such is the case for the dune grass Ammophila, whose decline under decreasing sand accretion is argued to be caused by either biotic or abiotic soil properties. Methodology/Principal Findings By manipulating dune soils from three different regions, we decoupled the contributions of region, the abiotic and biotic soil component to the variation in characteristics of Ammophila arenaria seedlings and Schizaphis rufula aphid populations. Root mass fraction and total dry biomass of plants were affected by soil biota, although the latter effect was not consistent across regions. None of the measured plant properties were significantly affected by the abiotic soil component. Aphid population characteristics all differed between regions, irrespective of whether soil biota were present or absent. Hence these effects were due to differences in abiotic soil properties between regions. Although several chemical properties of the soil mixtures were measured, none of these were consistent with results for plant or aphid traits. Conclusions/Significance Plants were affected more strongly by soil biota than by abiotic soil properties, whereas the opposite was true for aphids. Our results thus demonstrate that the relative importance of the abiotic and biotic component of soils can differ for plants and their herbivores. The fact that not all effects of soil properties could be detected across regions moreover emphasizes the need for spatial replication in order to make sound conclusions about the generality of aboveground-belowground interactions.

Distribution of Meloidogyne chitwoodi in potato tubers and comparison of extraction methods

Meloidogyne chitwoodi and M. fallax are quarantine organisms in Europe. One measure to restrict the spread of these nematodes is careful inspection of potato tubers. The distribution of M. chitwoodi in heavily infected tubers was studied and several methods for extraction of these nematodes from tubers were compared. The majority of the nematodes (96%) were found in the first 5.25 mm of the tuber, corresponding to the depth of the vascular ring. About half of them were found between 1.75 and 3.50 mm deep. Incubation of small pieces of tuber on Baermann funnels in the misting chamber during 36 days yielded about 12 times fewer juveniles than mixing potato tissues and extracting them using zonal centrifugation, a process that took about 1 h. Enzymatic maceration of potato tissues for 24 or 48 h did not liberate more nematodes than 2 min of blending the tissues at high speed. More nematodes, with 95% consisting of eggs, were extracted by zonal centrifuging than by pouring the macerated suspension over a set of 500 μm, 250 μm and 20 μm sieves.

Beyond plant–soil feedbacks: mechanisms driving plant community shifts due to land‐use legacies in post‐agricultural forests

Summary Although biotic legacies of past agricultural practices are widespread and increasing in contemporary ecosystems, our understanding of the mechanisms driving such legacies is still poor. Forest understories on former agricultural land show low frequencies and abundance of typical woodland species when compared with ancient forests. These community shifts have been ascribed to the effects of dispersal limitation. A rarely considered mechanism is that post-dispersal processes driven by plant-associated communities determine the poor performance and recruitment of woodland indicators. Given the strong alterations in soil conditions due to former agricultural practices, we hypothesized that (abiotic) plant–soil feedbacks could be a major factor in community shifts. We addressed this hypothesis by comparing plant-associated communities in the soil and above the ground in ancient and post-agricultural alluvial forests; then, we experimentally tested whether the changes in biotic and abiotic soil properties could affect above-ground herbivore abundance and pressure and plant performance. Ancient and post-agricultural communities clearly differed in composition at different levels of the food web. Besides the plant community, we also observed the differences in the microbial and nematode community with increased abundance of root-feeding nematodes in post-agricultural soils. The composition of the above-ground invertebrate community did not differ in ancient and post-agricultural forest parcels; however, plants growing in post-agricultural sites showed higher abundance of invertebrate herbivores and suffered more herbivory. Nutrient analyses of soil and plants showed that increased levels of phosphorus (and to a lesser extent, nitrogen) made plants more nutritious for insect herbivores. Laboratory experiments further pointed to this mechanism as an explanation of the poorer performance of woodland indicators in post-agricultural woodlands. Our results point to biotic and abiotic plant–soil feedbacks coupled with herbivory as a new mechanism to explain the legacy effects in temperate forests. The modification of the below-ground community and soil abiotic characteristics by previous agricultural activity affects not only the plant growth but also the plant nutrient content in the compared understorey species, making them more susceptible to above-ground herbivory. Our results provide one of the first examples of integrating plant–soil feedback and above- and below-ground interactions to explain land-use legacies.

Contrasting covariation of above- and belowground invertebrate species across plant genotypes.

1. Invertebrate species generally do not respond independently to genotypic variation in plants, giving rise to clusters of species that naturally associate with or avoid certain genotypes. This covariation causes coevolution to be diffuse rather than pairwise. Studies on this topic, however, have never considered the belowground invertebrate community, leaving a critical gap in our understanding. 2. We investigated the covariation among naturally colonising above- and belowground invertebrate species across six genetically distinct populations of the dune grass Ammophila arenaria. After having grown from seed in a common garden, plants were randomised in a single field site to exclude all but broad-sense genetic variation. 3. Strong positive covariation across genotypes among both above- and belowground invertebrates was detected, while correlations between these two groups were negative. This clustering of above- and belowground species matched well with order level taxonomy. Host range, trophic level and food type on the other hand did not correspond well with the clusters. Within the cluster of aboveground fauna, subsequent groupings were not related to any phylogenetic or ecological characteristic, although correlations within these subgroups were very high. We furthermore demonstrated significant differences in multiple invertebrate species occurrence between plant genotypes, in general as well as at the above- and belowground level. 4. The observed strong covariation suggests diffuse coevolution between A. arenaria and its associated invertebrate species. The trade-off between root and shoot invertebrates could however hamper directional selection on resistance to either group. 5. Our results clearly demonstrate the need for studies of plant-animal interactions to include the belowground fauna, as this might drastically alter our general conception of how plants and their associated animal communities interact and how these interactions shape the process of evolution.