Isolation of 44 polymorphic microsatellite loci in three host races of the phytopathogenic fungus Microbotryum violaceum

We report the development of 44 microsatellite markers in three host races of the fungus Microbotryum violaceum, a sexually transmitted disease of the Caryophyllaceae. An enrichment protocol was used to isolate microsatellite loci from three strains, collected, respectively, from the plant species Gypsophila repens, Dianthus sylvestris, and Silene vulgaris. Polymorphism and cross‐amplification were explored with 32 isolates of M. violaceum, collected on 12 different plant species in natural populations.     

Development and cross‐species amplification of 18 microsatellite markers in the Sumatran tiger (Panthera tigris sumatrae)

Eighteen polymorphic microsatellite loci from the highly endangered Sumatran tiger (Panthera tigris sumatrae) were isolated and characterized. Upon polymerase chain reaction amplification, 16 of these markers produced a single, sharp band in all three tiger and 10 non‐tiger felid species examined. Of the two remaining loci, 6HDZ057 and 6HDZ635 failed to amplify genomic DNA from puma (Felis concolor) and cheetah (Acinonyx jubatus), respectively. The amplification of these markers across four genera is an indication of their usefulness for population genetics studies and conservation work in a wide range of felid species.     

Isolation and characterization of microsatellite loci from the entomopathogenic hyphomycete,Paecilomyces fumosoroseus

Nine microsatellite markers were isolated from the entomopathogenic haploid fungus Paecilomyces fumosoroseus. Genetic diversity was assessed in 26 P. fumosoroseus isolates originated from the whitefly Bemisia tabaci collected in various geoclimatic areas. Eight loci were polymorphic with an observed number of alleles ranging from two to six. The loci differentiated some isolates and group of isolates according to their geographical location, showing promise for the study of gene flow. All loci failed to give clear amplifications in P. fumosoroseus isolates from hosts other than B. tabaci. These microsatellite markers provide powerful tools for ecological, epidemiological, and population genetic studies.     

Isolation of 12 polymorphic microsatellite loci in the phytopathogenic fungus Alternaria brassicicola

Twelve polymorphic microsatellite markers were isolated from the phytopathogenic fungus Alternaria brassicicola, the causal agent of black spot of crucifers. An enrichment protocol was used to isolate microsatellite loci, which were then analysed in a collection of 46 isolates sampled from seven different countries. The number of alleles detected in 12 loci ranged from two to 10 (mean 3.5). Investigation of cross‐species amplifications showed that the designed primers were specific to A. brassicicola.     

Isolation and characterisation of di and tri nucleotide microsatellite loci in Rosmarinus officinalis (Lamiaceae), using enriched genomic libraries

An enrichment protocol was used to isolate and characterise microsatellite loci in Rosmarinus officinalis, a Mediterranean chamephyte. Twelve microsatellite loci were characterised and amplified a total of 117 alleles in a sample of 30 individuals from one population, with an average of 9.75 alleles per locus. Observed heterozygosities ranged from 0.333 to 0.900. Cross-species transferability was also assayed in the two other species of the genus. The cumulated probabilities of exclusion for paternity and parentage of the 12 loci were of 0.999971 and 1, respectively, supporting the usefulness of these microsatellite loci for parentage analyses. Nine out of 12 microsatellite loci amplified in the two species and were polymorphic detecting a total of 49 and 45 in R. eriocalyx and R. tomentosus, respectively. Twenty-two alleles were exclusive of R. eriocalyx and 12 of R. tomentosus, additionally, three alleles were shared between these two species but were otherwise absent in the analysed individuals of R. officinalis. In total, this set of markers amplified 154 different microsatellite alleles, supporting their usefulness to conduct population genetic, reproductive biology and hybridisation studies in Rosmarinus.     

PRIMER NOTE. Using the incomplete genome of the ectomycorrhizal fungus Amanita bisporigera to identify molecular polymorphisms in the related Amanita phalloides

We describe the cross‐genomic isolation of 13 single nucleotide polymorphisms (SNPs) and one variable microsatellite from five loci for the death cap mushroom Amanita phalloides. Microsatellite repeats were identified by searching the partial Amanita bisporigera genome. Flanking primers were designed for 25 of these microsatellite loci and tested for cross‐amplification in A. phalloides. One locus contained an interrupted, compound microsatellite, and four loci contained one to six SNPs. These results demonstrate the usefulness of even an incomplete genome to identify molecular markers for population studies in nonmodel organisms.     

Polymorphic microsatellites for the study of newly established populations of the gastropod Cyclope neritea

Both human‐mediated introductions and climatic changes may promote the settlement of species in new areas outside of their natural geographical range. To investigate the settlement of recently established populations of the neogastropod Cyclope neritea, we developed eight microsatellite markers. Their usefulness was studied in two native populations previously found to be monomorphic with mitochondrial markers. The eight loci were found to be polymorphic in both populations, with two to 18 alleles per locus. This result shows promise for these loci in studies of recently founded populations of C. neritea.     

Isolation and characterization of simple sequence repeat loci in Rubus hochstetterorum and their use in other species from the Rosaceae family

The identification of microsatellite loci in Rubus hochstetterorum provides an important tool for the characterization and conservation of wild populations of this species. Cross‐species amplification of markers may be of particular interest for the study of other Rubus species. In this study, 41 simple sequence repeat markers were identified in a genomic library of R. hochstetterorum. Fifteen of the identified microsatellite loci were characterized in a set of 30 samples and revealed to be polymorphic with three to 19 alleles per locus. All the identified markers allowed cross‐species amplification in at least one of the other three tested species from the Rosaceae family.     

Isolation of 21 new polymorphic microsatellite loci in the phytopathogenic fungus Venturia inaequalis

Twenty‐one new polymorphic microsatellite markers were isolated in the phytopathogenic fungus Venturia inaequalis, the causal agent of apple scab. An enrichment protocol was used to isolate microsatellite loci and the level of polymorphism was assessed on 44 European isolates. All loci were polymorphic with an average of 9.1 alleles per locus (range 2–24). Tests of cross‐species amplifications suggest that at least some of these microsatellites could be used in different species, mainly Spilocaea pyracanthae and S. eriobotryae.     

Microsatellite loci from the ectomycorrhizal basidiomycete Suillus pictus associated with the genus Pinus subgenus Strobus

Seven microsatellite loci were isolated and characterized from ectomycorrhizal basidiomycete Suillus pictus associated with soft pine species (Pinus subgenus Strobus) using a dual‐suppression‐PCR technique. Microsatellite variation was assessed using 38 isolates of S. pictus sampled from three different local populations in Japan. The number of alleles per locus ranged from three to 12 within populations. These microsatellite loci can be used for studying the allelic variation and genetic structure in/among local populations of S. pictus.     

Seventeen novel microsatellite markers from an enriched library of the pest species Bactrocera dorsalis sensu stricto

Seventeen novel microsatellite markers were isolated and characterized from an enriched DNA library of the pest species Bactrocera dorsalis sensu stricto. The polymorphism of these loci was tested in individual fruit flies from a Thai mass‐rearing strain and three wild Asian populations: two from Thailand and one from Myanmar. Allele numbers per locus ranged from two to 15. In the natural populations, a high level of polymorphism was detected in all loci suggesting the usefulness of these markers to quantify the genetic variation in this highly invasive insect.     

Isolation and characterization of five microsatellite loci in Dolichopoda cave crickets (Orthoptera Rhaphidophoridae)

Five microsatellite loci are described for the cave cricket genus Dolichopoda. Preliminary data on allelic variation of these loci are presented for one population of D. schiavazzii and one population of D. laetitiae to test their usefulness in fine‐scale studies of the genetic aspect of cave colonization. Cross‐species amplifications were carried out in four other Dolichopoda species and in two species belonging to another cave cricket genus (Troglophilus) to test the potential use of these microsatellite markers in studies of both congeneric species and species belonging to the same family.     

Isolation of polymorphic microsatellite loci from Eurasian woodcock (Scolopax rusticola) and their cross‐utility in related species

We isolated one trinucleotide and seven tetranucleotide microsatellite loci for the Eurasian woodcock (Scolopax rusticola). We describe polymerase chain reaction conditions and primers for the successful amplification of these loci and report the results obtained from their use in 42 specimens from two populations in Europe. The number of alleles per locus ranged from three to 15, observed heterozygosity was comprised between 0.11 and 1.00 and expected heterozygosity ranged between 0.10 and 0.91. Cross‐specific amplification experiments highlighted the potential usefulness of these molecular markers for the study of three related scolopacid waders.     

Isolation of twelve microsatellite loci,using an enrichment protocol,in the phytopathogenic fungus Puccinia striiformis f.sp. tritici

We report the characterization of 12 microsatellite markers in the biotrophic fungus Puccinia striiformis f.sp. tritici, responsible for yellow rust on wheat. An enrichment protocol was used to isolate microsatellite loci, and polymorphism was explored with 96 isolates from natural populations collected from several French and Chinese locations. Eight primers (67%) showed cross‐amplification when tested with eight isolates of P. triticina.     

Molecular characterisation of fungal endophytic morphospecies associated with the indigenous forest tree, Theobroma gileri, in Ecuador

Fungal endophytes were isolated from healthy stems and pods of Theobroma gileri, an alternative host of the frosty pod rot pathogen of cacao. Non-sporulating isolates were grouped into 46 different morphological species according to their colony morphology. Many of these morphospecies were assumed to be basidiomycetes and, therefore, were of particular interest. Basidiomycetous endophytes have received far less attention than ascomycetes and also have potential as biological control agents of the basidiomycetous pathogens of T. cacao: Moniliophthora roreri (frosty pod rot pathogen) and M. perniciosa (witches' broom disease). The morphospecies were further characterised by molecular analyses. Amplification of the nuLSU was undertaken for phylogenetic placement of these non-sporulating cultures and revealed a total of 31 different taxa of which 15 were basidiomycetes belonging to the class Agaricomycetes, and 16 ascomycetes primarily belonging to the Sordariomycetes.     

Moniliophthora roreri,causal agent of cacao frosty pod rot

Taxonomy : Moniliophthora roreri (Cif.) H.C. Evans et al. 1978 ; Phylum Basidiomycota; Class Agaricomycetes; Order Agaricales; Family Marasmiaceae; Genus Moniliophthora. Biology : Moniliophthora roreri attacks Theobroma and Herrania species causing frosty pod rot. Theobroma cacao (cacao) is the host of major economic concern. Moniliophthora roreri is a hemibiotroph with a long biotrophic phase (45–90 days). Spore masses, of apparent asexual origin, are produced on the pod surface after initiation of the necrotrophic phase. Spores are spread by wind, rain and human activity. Symptoms of the biotrophic phase can include necrotic flecks and, in some cases, pod malformation, but pods otherwise remain asymptomatic. Relationship to Moniliophthora perniciosa : Moniliophthora roreri and Moniliophthora perniciosa, causal agent of witches’ broom disease of cacao, are closely related. Their genomes are similar, including many of the genes they carry which are considered to be important in the disease process. Moniliophthora perniciosa, also a hemibiotroph, has a typical basidiomycete lifestyle and morphology, forming clamp connections and producing mushrooms. Basidiospores infect meristematic tissues including flower cushions, stem tips and pods. Moniliophthora roreri does not form clamp connections or mushrooms and infects pods only. Both pathogens are limited to the Western Hemisphere and are a threat to cacao production around the world. Agronomic importance : Disease losses caused by frosty pod rot can reach 90% and result in field abandonment. Moniliophthora roreri remains in the invasive phase in the Western Hemisphere, not having reached Brazil, some islands within the Caribbean and a few specific regions within otherwise invaded countries. Disease management : The disease can be managed by a combination of cultural (for example, maintenance of tree height and removal of infected pods) and chemical methods. These methods benefit from regional application, but can be cost prohibitive. Breeding for disease resistance offers the greatest potential for frosty pod rot management and new tolerant materials are becoming available.     

Characterization of nine microsatellite loci in the sea star Astropecten aranciacus and cross-species amplification for related taxa

So far, only few microsatellite markers have been developed and extensively tested for echinoderms. To study the population genetic structure of the sea star Astropecten aranciacus, we developed primers for nine polymorphic microsatellite loci and tested them on two populations from Faro in Portugal (N = 25) and from La Herradura in Spain (N = 20). Within populations, allele numbers varied from four to 20, while expected and observed heterozygosities ranged from 0.593 to 0.936 and from 0.222 to 0.900, respectively. Additional cross‐species amplifications were polymorphic at some loci, indicating their potential usefulness for related taxa.     

Characterization and cross‐species amplification of microsatellite loci in the plant pathogenic fungus Leptosphaeria maculans

Seven polymorphic microsatellite markers suitable for population genetic studies and genetic mapping were developed for Leptosphaeria maculans, a fungal pathogen of canola (Brassica napus). Polymorphism was evaluated using 14 isolates from diverse geographical locations. Each locus had either two or three alleles. Cross‐species amplification was observed for almost all loci in L. biglobosa ‘brassicae’ and L. maculans ‘lepidii’.     

Development of novel microsatellite markers for the grassland species Lolium multiflorum,Lolium perenne and Festuca pratensis

Twelve microsatellite markers were isolated from Lolium multiflorum. Allelic variability and cross‐species amplification were assessed on 16 individuals of each of the three grassland species L. multiflorum, Lolium perenne and Festuca pratensis. Cross‐species amplification success was 100% for L. perenne and 83% for F. pratensis. The number of alleles detected ranged from one to 14 with an average of 3.4. While three microsatellite loci were polymorphic in all three species, one marker produced species‐specific alleles in all three species. These microsatellite markers provide a valuable tool for population genetic studies within and among species of the Festuca–Lolium complex.     

Isolation of 12 microsatellite loci,using an enrichment protocol,in the phytopathogenic fungus Puccinia triticina

We report the characterization of 12 polymorphic microsatellite markers in the biotrophic fungus Puccinia triticina, the causal agent of leaf rust on wheat. An enrichment protocol was used to isolate microsatellite loci and polymorphism was explored with 15 European isolates. Significant level of cross‐amplification (44% of the loci) was found in P. striiformis.