Ten polymorphic microsatellite markers for Scaphoideus titanus,the vector of flavescence dorée phytoplasma

The leafhopper Scaphoideus titanus is a vector of flavescence dorée phytoplasma, the causal agent of the most important grapevine yellow disease in European vineyards. Ten polymorphic microsatellite loci were developed from a genomic library enriched for AC and AG repeats. Levels of polymorphism were evaluated in 106 individuals from S. titanus European and American populations. An average of 16 alleles per locus was detected and the observed heterozygosity ranged from 0.141 to 0.813. Cross‐species amplification was successful in four other Cicadellidae species. These 10 microsatellites are valuable markers for population genetic and phylogeographical studies of S. titanus.     

Dispersal patterns and chromatic response of Scaphoideus titanus Ball (Homoptera Cicadellidae), vector of the phytoplasma agent of grapevine flavescence dorée

Abstract 1 Scaphoideus titanus Ball, a nearctic leafhopper introduced into Europe in the 1950s, is known to be the vector of the phytoplasma agent of flavescence dorée (FD), a persistent disease of grapevine. Knowledge of its dispersal patterns is thus very important to prevent disease outbreaks. 2 Yellow sticky traps were used to study the seasonal flight activity of S. titanus, its vertical flight, its movement outside the vineyard and the influence of plant density. Sticky traps of different colours (yellow, red, blue, and white) were also compared. The behaviour of males and females was tested for all those conditions. 3 Abundance was greater in normal than in low plant density conditions, and a positive relationship was found between number of plants per square metre and presence of S. titanus. Leafhoppers did not appear capable of spreading significantly outside a vineyard. Few individuals were trapped above the canopy. Red sticky traps caught more individuals than white, yellow or blue, with the latter showing a poor attractiveness. Sex ratio was almost always male biased. 4 Scaphoideus titanus is monophagous and appears incapable of great dispersal away from its host plant, and females are less likely to fly than males. Further studies on the influence of different factors on the behaviour of this leafhopper are suggested.     

Genetic Variability in European Populations of an Invasive American Crayfish: Preliminary Results

Since 1973, the red swamp crayfish, Procambarus clarkii, native to south-central United States and northeastern Mexico, has spread throughout Europe. Here, we surveyed the genetic variability of five European populations of the species using RAPD markers. Genetic variation was found to be so high as to uniquely fingerprint most of the surveyed individuals. Analysis of molecular variance (AMOVA) of the RAPD markers showed that 1) a large part of the genetic variation can be attributed to the differentiation among localities, and 2) the differentiation was mainly due to the separation of the samples from Louisiana with respect to the European set. A hypothesis emerged in which subsequent introductions of crayfish from different sources were performed. This hypothesis might explain the high genetic diversity observed within each population and the genetic differentiation among populations, as the result, respectively, of the introduction of different sets of crayfish and the casual bias of introductions. Although preliminary, our results suggest that RAPDs could be helpful in providing information about human-mediated introduced populations.     

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.     

Distinct invasion sources of common ragweed <Emphasis Type="Italic">(Ambrosia artemisiifolia)</Emphasis> in Eastern and Western Europe

The common ragweed (Ambrosia artemisiifolia L.; Asteraceae) is a North American native that is invading Eurasia. Besides its economic impact on crop yield, it presents a major health problem because of its highly allergenic pollen. The plant was imported inadvertently to Europe in the eighteenth century and has become invasive in several countries. By analyzing French and North American populations, it was previously shown that French populations were best described as a mixture of native sources and that range expansion in France probably involved sequential bottlenecks. Here, our aim was to determine whether Eastern European populations of A. artemisiifolia originated from the previously established French populations or from independent trans-Atlantic colonization events. We used nuclear microsatellite markers to elucidate the relationships among populations from Eastern and Western Europe in relation to populations from a broad survey across the native North American range. We found that A. artemisiifolia from Eastern Europe did not originate from the earlier established French populations but rather represents multiple independent introductions from other sources, or introductions from a not yet identified highly diverse native population. Eastern European populations show comparable amounts of genetic variability as do previously characterized French and North American populations, but analyses of population structure clearly distinguish the two European groups. This suggests separate introductions in Eastern and Western Europe as well as divergent sources for these two invasions, possibly as a result of distinct rules for trade and exchange for Eastern Europe during most of the twentieth century.     

The declining Spadefoot toad, Pelobates fuscus (Pelobatidae): paleo and recent environmental changes as a major influence on current population structure and status

Populations of the European Spadefoot toad (Pelobates fuscus) have experienced recent declines all over Europe, but these appear to be more intense in north and western Europe. Due to the toad’s fossorial nature and specific habitat requirements, environmental conditions have played a major role in structuring current populations. We examined the phylogeographic structure in P. fuscus from 16 localities throughout Europe using mitochondrial cytochrome b gene sequence analysis. Sequence divergence among haplotypes was low (0.54±0.15%). Three very closely related haplotypes occupy northern and western parts of Europe whereas 12 others were observed among samples from south-eastern Europe, including the Balkans. Our results suggest that toads only recently colonized the northern and western parts of Europe following glacial retreat. This expansion probably took place in steppic-like areas during the younger Dryas cold interval, about 12,900–11,500 years ago. Restricted gene flow with an isolation-by-distance population structure characterises a major part of its distribution range. Based on our results we suggest that the northern and western lineages should be considered as distinct conservation units, while the south-eastern populations from the refugial areas, where nearly all genetic polymorphism occurs and populations appear less vulnerable, should receive special attention.     

The effects of heavy metal pollution on genetic diversity in zinc/cadmium hyperaccumulator Sedum alfredii populations

The genetic diversity and population structure of seven populations of Sedum alfredii growing in lead/zinc (Pb/Zn) mine spoils or in uncontaminated soils from eastern and southern China were investigated using random amplified polymorphic DNA (RAPD) technology. Four of the sampled sites were heavily contaminated with heavy metals (Zn, Cd, Pb), and extremely high concentrations of Zn, Cd, and Pb were found among these corresponding populations. A significant reduction of genetic diversity was detected in the mining populations. The reduction of genetic diversity could be derived from a bottleneck effect and might also be attributed to the prevalence of vegetative reproduction of the mining populations. Analysis of molecular variance (AMOVA) and the unweighted pair group method with arithmetic mean (UPGMA) tree derived from genetic distances further corroborated that the genetic differentiation between mine populations and uncontaminated populations was significant. Polymorphism with the heavy metal accumulation capability of S. alfredii probably due to the genetic variation among populations and heavy metal contamination could have more impact on the genetic diversity and population structure of S. alfredii populations than geographic distance.     

Genetic biogeography of the rate “copper moss”,Scopelophila cataractae (Pottiaceae)

Scopelophila cataractae, one of the so-called copper mosses, has a broad geographic distribution that includes North, Central, and South America, Europe, and Asia, but is rare throughout its range. A genetic analysis of 32 populations from the United States, Europe, and Asia based on 15 putative allozyme loci indicates that levels of genetic diversity vary among geographic regions. Six European populations are fixed for the same alleles at all 15 loci, consistent with the hypothesis thatS. cataractae is a recent immigrant in that region. The species is more diverse in the U.S., where it appears to be native. Five populations collected on copper-enriched soils around shrines and temples in Tokyo are genetically monomorphic, but Asian populations from another Japanese site, India, and Nepal are exceptionally diverse in terms of numbers of alleles and multilocus haplotypes, total gene diversity (H_T), and in the degree of differentiation among populations (measured as Nei'sI andD). Long-distance dispersal has probably played an important role in the geographic history ofS. cataractae, but the species appears to be native in both the New and Old Worlds. Gene flow between plants disjunct on different continents is insufficient to explain the lack of geographically correlated morphological and genetic differentiation inS. cataractae.     

Population genetic structure and distribution of introduced American mink (<Emphasis Type="Italic">Mustela vison</Emphasis>) in Spain,based on microsatellite variation

The population genetic structure of an invasive species in Spain, the American mink (Mustela vison), was investigated using microsatellite DNA markers. This semi-aquatic carnivore, originating from North America, was imported into Europe for fur farming since the beginning of the 20th century. Due to massive escapes, farm damages, deliberate releases and/or accidents, feral mink populations were established in the aquatic ecosystems of many European countries, including Spain. We genotyped 155 American mink originating from the Spanish regions Basque Country, Catalonia, Castilla-Leon, Aragon, Valencia and Galicia using 10 polymorphic microsatellite loci to highlight population genetic structure, distribution and dispersal. M. vison populations in Spain appear differentiated and not yet connected by gene flow. Bayesian clustering analyses and spatial analyses of molecular variance detected four inferred clusters, overall coinciding with the sampled geographical localities. Preliminary testing shows moderate to large estimated effective population sizes. Molecular analyses result useful to provide baseline data for further research on the evolution of invasive mink populations, as well as support local management strategies and indirectly benefit the conservation of threatened species in Spain, such as the endangered European mink (Mustela lutreola), and the polecat (Mustela putorius), which share the habitat with the American mink. This paper is dedicated to the memory of Xavier Domingo-Roura.     

Preliminary genetic data on some Caribbean Artemia franciscana strains based on RAPD's

A total of fourteen Artemia samples from Colombia, Venezuela, Curaçao (Netherlands Antilles), Puerto Rico, and reference samples from U.S.A. (San Francisco Bay, SFB) belonging to the superspecies Artemia franciscana, and Argentina (A. persimilis), were analysed with the RAPD technique in order to demonstrate genetic dissimilarities. Pearson's correlation coefficients between the DNA banding patterns were calculated. They served as input values for the construction of UPGMA dendrograms. The results indicate that, within the collection of Colombian, Venezuelan and the two Netherlands Antilles Artemiacyst samples examined, two different groups seem to exist. Geographically, the mountainous area of Sierra Nevada de Santa Marta separates these two groups (lower Caribbean to the South and middle Caribbean to the North). Although the Caribbean, North and South American populations belong to A. franciscana, genetic discontinuities are to be expected due to habitat differences and geographic isolation. The Sierra Nevada (with an altitude of about 5800 m) emerges as the barrier very likely to explain the observed RAPD differences. Little genetic variability was present in the Colombian samples from Manaure that were collected almost every ten years, nor in the samples from Galerazamba collected almost two decades apart, although these samples were more likely subjected to different prevailing environmental conditions. The SFB population did not show a very close relation with all Caribbean populations analyzed, including the Puerto Rican. All A. franciscana populations analyzed were divergent from A. persimilis(Argentina).     

Latitudinal variability of Drosophila melanogaster: Allozyme frequencies divergence between European and Afrotropical populations

Allelic frequencies at five polymorphic loci were determined in seven European and six Afrotropical populations of Drosophila melanogaster. African populations, which may be considered as ancestral for the species, showed a greater genetic diversity as measured by the number of alleles found. Within each geographic group (Europe or tropical Africa) genetic distances between local populations were very small (D=0.027). By contrast, the average distance between European and African populations (D=0.389) was more than 12 times bigger. It was previously known that various morphological or physiological differences, which probably reflect genetic adaptations to different environments, exist between these temperate and tropical populations. Data presented here suggest that the divergence in allozyme frequencies also reflects some selective mechanisms.     

Patterns of Genetic Variation in Swertia przewalskii, an Endangered Endemic Species of the Qinghai-Tibet Plateau

Swertia przewalskii Pissjauk. (Gentianaceae) is a critically endangered and endemic plant of the Qinghai-Tibet Plateau in China. RAPD and ISSR analyses were carried out on a total of 63 individuals to assess the extent of genetic variation in the remaining three populations. Percentage of polymorphic bands was 94% (156 bands) for RAPD and 96% (222 bands) for ISSR. A pairwise distance measure calculated from the RAPD and ISSR data was used as input for analysis of molecular variance (AMOVA). AMOVA indicated that a high proportion of the total genetic variation (52% for RAPD and 56% for ISSR) was found among populations; pairwise Φ _ST comparisons showed that the three populations examined were significantly different (p < 0.001). Significant genetic differentiation was found based on different measures (AMOVA and Hickory θ_B) in S. przewalskii (0.52 on RAPD and 0.56 on ISSR; 0.46 on RAPD and 0.45 on ISSR). The differentiation of the populations corresponded to low average gene flow (0.28 based on RAPD and 0.31 based on ISSR), whereas genetic distance-based clustering and coalescent-based assignment analyses revealed significant genetic isolation among populations. Our results indicate that genetic diversity is independent of population size. We conclude that although sexual reproduction and gene flow between populations of S. przewalskii are very limited, they have preserved high levels of genetic diversity. The main factors responsible for the high level of difference among populations are the isolation and recent fragmentation under human disturbance.     

Structure and genetic diversity in Colliguaja odorifera Mol. (Euphorbiaceae), a shrub subjected to Pleisto-Holocenic natural perturbations in a mediterranean South American region

Aim Colliguaja odorifera Mol., a Euphorbiaceous shrub of central Chile, inhabits the matorral formation, growing at low altitudes on both Andean and coastal mountain range slopes. In the recent geological past, this region was subjected to climatic changes and geological disturbances that most probably caused population shrinkages on the Andean mountain slopes. This study tested the hypothesis that under such a scenario, existing populations should show lower genetic diversity in the Andean than in the coastal areas; these coastal populations being the potential source populations for recolonization. Location The study was carried out in central Chile by comparing the genetic diversity between the Andean and coastal areas, each represented by five localities distributed from 32°30′ S to 34° S. Methods Genetic diversity was estimated by DNA analysis using 18 dominant multilocus Random Amplified Polymorphic DNA (RAPD) loci, characterizing 73 genetic phenotypes. Results The comparison of the two matorral areas showed that Andean populations of C. odorifera have a subset of the genetic diversity found in the coastal populations. Andean populations also show a consistently lower genetic diversity, lower genetic distances and higher genetic structure, coincident with expectations based on the Pleisto‐Holocenic perturbation regime. Main conclusions This first genetic analysis for South American mediterranean populations confirms the findings of previous floristic and palynological studies that identified refuge zones in the coastal mountain range of central Chile, a situation analogous with that occurring during periods of inter‐glacial northward migration in Southern Europe.     

High regional differentiation in a North American crab species throughout its native range and invaded European waters: a phylogeographic analysis

Rhithropanopeus harrisii (Gould 1841) has a native distribution from New Brunswick (Canada) to Veracruz (Mexico) and is considered an invasive species in northwestern North American (Oregon and California), South American (Brazil) and European estuaries and rivers. In Europe, it was observed for the first time in 1874, in The Netherlands. We sequenced and analyzed part of the cytochrome oxidase subunit I gene (mitochondrial DNA) of eight populations, three from the east coast of the United States of America (USA) and five from Europe, in order to assess their genetic diversity and to determine a potential founder population. European populations are characterized by a lower number of haplotypes than the whole native region of the eastern USA, suggesting that genetic bottlenecks occurred during the European colonisation. Along the North American East Coast, there is evidence of clearcut genetic heterogeneity, New Jersey being the most similar population in its genetic structure to the postulated Europe-founding population. Also the different European populations are heterogeneous and there is a tendency of higher genetic diversity in the populations founded earlier. R. harrisii is still in the process of expansion in Europe and may have been introduced once or repeatedly by different invasion mechanisms. The pronounced lack of gene flow among populations is of great ecological significance, since it may facilitate rapid adaptation and specialization to local conditions within single estuarine systems.     

A few north Appalachian populations are the source of European black locust

The role of evolution in biological invasion studies is often overlooked. In order to evaluate the evolutionary mechanisms behind invasiveness, it is crucial to identify the source populations of the introduction. Studies in population genetics were carried out on Robinia pseudoacacia L., a North American tree which is now one of the worst invasive tree species in Europe. We realized large‐scale sampling in both the invasive and native ranges: 63 populations were sampled and 818 individuals were genotyped using 113 SNPs. We identified clonal genotypes in each population and analyzed between and within range population structure, and then, we compared genetic diversity between ranges, enlarging the number of SNPs to mitigate the ascertainment bias. First, we demonstrated that European black locust was introduced from just a limited number of populations located in the Appalachian Mountains, which is in agreement with the historical documents briefly reviewed in this study. Within America, population structure reflected the effects of long‐term processes, whereas in Europe it was largely impacted by human activities. Second, we showed that there is a genetic bottleneck between the ranges with a decrease in allelic richness and total number of alleles in Europe. Lastly, we found more clonality within European populations. Black locust became invasive in Europe despite being introduced from a reduced part of its native distribution. Our results suggest that human activity, such as breeding programs in Europe and the seed trade throughout the introduced range, had a major role in promoting invasion; therefore, the introduction of the missing American genetic cluster to Europe should be avoided.     

Genetic structure analysis of natural <Emphasis Type="Italic">Sargassum muticum</Emphasis> (Fucales,Phaeophyta) populations using RAPD and ISSR markers

Sargassum muticum is important in maintaining the structure and function of littoral ecosystems, and is used in aquaculture and alginate production, however, little is known about its population genetic attributes. In this study, random amplified polymorphic DNA (RAPD) and inter-simple sequence repeat (ISSR) markers were used to investigate the genetic structure of four populations of S. muticum and one outgroup of S. fusiforme (Harv.) Setchell from Shandong peninsula of China. The selected 24 RAPD primers and 19 ISSR primers amplified 164 loci and 122 loci, respectively. Estimates of genetic diversity with different indicators (P%, percentage of polymorphic loci; H, the expected heterozygosity; I, Shannon’s information index) revealed low or moderate level of genetic variations within each S. muticum population, and a high level of genetic differentiations were determined with pairwise unbiased genetic distance (D) and fixation index (F _ST ) among the populations. The Mantel test showed that two types of matrices of D and F _ST were highly correlated whether from RAPD (r = 0.9706, P = 0.009) or ISSR data (r = 0.9161, P = 0.009). Analysis of molecular variance (AMOVA) was conducted to apportion the variations among and within the S. muticum populations. It indicated that variations among populations were higher than those within populations, being 55.82% verse 44.18% by RAPD and 55.21% verse 44.79% by ISSR, respectively. Furthermore, the Mantel test suggested that genetic differentiations among populations were related to the geographical distances (r > 0.6), namely, conformed to the IBD (isolation by distance) model, as expected from UPGMA (unweighted pair group method with arithmetic averages) cluster analysis. On the whole, the high genetic structuring among the four S. muticum populations along the distant locations was clearly indicated in RAPD and ISSR analyses (r > 0.9, P < 0.05) in our study.     

Examining the genetic diversity of the orange blossom wheat midge,Sitodiplosis mosellana (Géhin), across North America

The orange blossom wheat midge, Sitodiplosis mosellana (Géhin) (Diptera: Cecidomyiidae), is a significant pest of wheat (Triticum spp.) grown in the Northern Hemisphere. It was accidently introduced to North America over 200 years ago and has subsequently spread throughout the northern Great Plains. Since 2010, several Canadian spring wheat varieties containing the resistance gene Sm1 have been released. Due to the potential of wheat midge populations to evolve virulent biotypes to Sm1, cultivars containing Sm1 are grown with a susceptible cultivar in an interspersed refuge. An understanding of the genetic diversity of wheat midge populations could provide important information on the potential development of resistance to Sm1. In the current study, we used two mitochondrial genes (CO1 and ND4) from wheat midge collected across the northern Great Plains and Québec in North America to assess population structure and genetic diversity. We found limited genetic diversity and population structure across the sampled North American populations. We also assessed North American haplotype similarity to wheat midge collected from Europe and China and found high similarity between North American and European populations, although sampling in Europe was limited. This supports the hypothesis that North American populations originated from Europe.     

New insights on Flavescence dorée phytoplasma ecology in the vineyard agro‐ecosystem in southern Switzerland

Phytoplasmas associated with Flavescence dorée (FDp) grapevine disease are quarantine pathogens controlled through mandatory measures including the prompt eradication and destruction of diseased plants, and the insecticide treatments against the insect vector, the ampelophagous leafhopper Scaphoideus titanus. In the present study, a multidisciplinary approach has been applied to investigate the FDp ecological cycle in a test vineyard agro‐ecosystem in Canton Ticino, south Switzerland. Despite the scarce population density of S. titanus, a regular trend of new infections (3.4% of the total vines) through the years was observed. The leafhopper Orientus ishidae was found as the most abundant among the captured insect species known as phytoplasma vectors (245 out of 315 specimens). The population of O. ishidae was evidenced prevalently (167 specimens) in the south‐western side of the vineyard and within the neighbouring forest constituted mainly by hazel (Corylus avellana) and willow (Salix spp.). These plant species were found infected by FDp related strains (30% of analysed trees) for the first time in this study. Interestingly, O. ishidae was found to harbour FDp related strains in high percentage (26% of the analysed pools). In addition, 16SrV phytoplasma group was detected for the first time in the insect Hyalesthes obsoletus and a FDp related strain in Thamnotettix dilutior, present in low populations within the test vineyard. Molecular characterisation and phylogenetic analyses of methionine aminopeptidase (map) gene sequences of FDp and related strains, here identified, revealed the great prevalence of the map‐type FD2 in grapevines (97%) and in O. ishidae pools (72%). Such a map‐type was found also in hazel and in T. dilutior, but not in S. titanus. Moreover, map‐types FD1 and FD3 were identified for the first time in Switzerland in several host plants and phytoplasma vectors, including grapevine (FD1), S. titanus (FD1) and O. ishidae (FD1 and FD3). Based on the data obtained in this study, it is reasonable to hypothesise that the ecological cycle of FDp could be related not exclusively to the grapevine‐specific feeding diet of S. titanus, but it could include other insect vector(s) and/or plant host(s). Further studies will be needed to prove the role of O. ishidae as vector able to transmit FDp from wild plants (e.g. hazel) to grapevine.     

Molecular genetic structure and evolution in native and colonized populations of the Chinese mitten crab, Eriocheir sinensis

Chinese mitten crab (Eriocheir sinensis), a native species in China, has populated Continental Europe and North America since 1912. In this paper, the nucleotide variation in the fragments of mitochondrial COII (693 bp), Cytb (766 bp), and nucleotide ITS (706 bp) was analyzed in native (Yantgze, Yellow, and Liaohe Rivers in China) and colonized (Elbe, Rhine, and Thames Rivers in Europe, and San Francisco Bay in North America) populations of the Chinese mitten crab. The major findings are as follows. First, the genetic variability in the native populations is higher than that in the colonized European and North American populations, with the exception of the Elbe River population, which possesses a similar level of variability with the native populations. Second, a remarkable loss of singletons has been associated with the colonization of Chinese mitten crabs. Third, the AMOVA and F _ST results demonstrate that there are no significant genetic differentiations among the populations from the three continents, but there is a significant differentiation between pairwise populations within and among continents. Fourth, it is found that expansion-drift and gene flow pattern are involved in the European populations. The neutrality test and R ₂ statistics suggest that a moderate founder population exists in the colonized populations, and only the Yangtze River population has undergone a recent population expansion. Finally, the results demonstrate that the European populations originate from multiple rivers in China on multiple occasions. The San Francisco population originates from both the native Chinese populations and the colonized European populations, most likely the Thames population.