The genetic structure of the plant pathogenic fungus Melampsora larici-populina on its wild host is extensively impacted by host domestication

Wild and cultivated plants represent very different habitats for pathogens, especially when cultivated plants bear qualitative resistance genes. Here, we investigated to what extent the population genetic structure of a plant pathogenic fungus collected on its wild host can be impacted by the deployment of resistant cultivars. We studied one of the main poplar diseases, poplar rust, caused by the fungus Melampsora larici‐populina. A thousand and fifty individuals sampled from several locations in France were phenotyped for their virulence profile (ability to infect or not the most deployed resistant cultivar ‘Beaupré’), and a subset of these was genotyped using 25 microsatellite markers. Bayesian assignment tests on genetic data clustered the 476 genotyped individuals into three genetic groups. Group 1 gathered most virulent individuals and displayed evidence for selection and drastic demographic changes resulting from breakdown of the poplar cultivar ‘Beaupré’. Group 2 comprised individuals corresponding to ancestral populations of M. larici‐populina naturally occurring in the native range. Group 3 displayed the hallmarks of strict asexual reproduction, which has never previously been demonstrated in this species. We discuss how poplar cultivation has influenced the spatial and genetic structure of this plant pathogenic fungus, and has led to the spread of virulence alleles (gene swamping) in M. larici‐populina populations evolving on the wild host.     

Temporal variation in coat colour (genotypes) supports major changes in the Nordic cattle population after Iron Age

Variation in coat colour genotypes of archaeological cattle samples from Finland was studied by sequencing 69 base pairs of the extension locus (melanocortin 1‐receptor, MC1R) targeting both a transition and a deletion defining the three main alleles, such as dominant black (E^D), wild type (E⁺) and recessive red (e). The 69‐bp MC1R sequence was successfully analysed from 23 ancient (1000–1800 AD) samples. All three main alleles and genotype combinations were detected with allele frequencies of 0.26, 0.17 and 0.57 for E^D, E⁺ and e respectively. Recessive red and dominant black alleles were detected in both sexes. According to the best of our knowledge, this is the first ancient DNA study defining all three main MC1R alleles. Observed MC1R alleles are in agreement with calculated phenotype frequencies from historical sources. The division of ancient Finnish cattle population into modern Finnish breeds with settled colours was dated to the 20th century. From the existing genotyped populations in Europe (43 breeds, n = 2360), the closest match to ancient MC1R genotype frequencies was with the Norwegian native multicoloured breeds. In combined published genotype data of ancient (n = 147) and genotypes and phenotypes of modern Nordic cattle (n = 738), MC1R allele frequencies showed temporal changes similar to neutral mitochondrial DNA and Y‐chromosomal haplotypes analysed earlier. All three markers indicate major change in genotypes in Nordic cattle from the Late Iron Age to the Medieval period followed by slower change through the historical periods until the present.     

Genetic Structure and Kinship Patterns in a Population of Black Howler Monkeys, Alouatta pigra, at Palenque National Park, Mexico

We investigated the genetic structure and kinship patterns of black howler monkeys (Alouatta pigra) at Palenque National Park, Mexico. Fecal samples from 49 individuals residing in eight social groups were successfully genotyped for 19 polymorphic microsatellite markers known to be variable in other ateline primates. Overall, genetic diversity was low (H_o = 0.588) with an average of 4.2 alleles per loci (range = 2–8). We found that intergroup genetic variation among adults was relatively high (mean between‐group F_ST = 0.119), largely due to the genetic divergence of one study group from the others. Intragroup kinship patterns showed that in most social groups, either adult males, adult females, or individuals of both sexes resided with same‐sexed adult kin, suggesting that some black howler males and females may not disperse from their natal group or may disperse with related individuals. Of the six sampled immigrant males, two males joined established groups by themselves, and four males formed two pairs that each took over the social group they joined after evicting the resident males. Males in both these coalitions were genetically closely related, while the two solitary immigrants were not closely related to any of the resident males present in the group they joined. Am. J. Primatol. 74:948‐957, 2012. © 2012 Wiley Periodicals, Inc.     

Microstructure of the silk apparatus in the coelotine spider Paracoelotes spinivulva (Araneae: Amaurobiidae)

The microstructural characteristics of the silk‐spinning apparatus and its ecological significance in the coelotine spider Paracoelotes spinivulva were examined by field emission scanning electron microscopy, with the goal of understanding the properties and the evolutionary origins of these silk constructs. The silk apparatuses of this spider were composed of four basic types of silk‐spinning spigot (ampullate, pyriform, aciniform and tubuliform), which connected with typical silk glands in the abdominal cavity. Of the three pairs of spinnerets, the posterior pairs were highly elongated along the body axis. Anterior spinnerets comprised two pairs of ampullate glands and approximately 70–80 pairs of pyriform glands in both sexes. Middle spinnerets had one to two pairs of ampullate spigots, three pairs of tubuliform spigots in females, and 50–60 (female) or 80–90 (male) pairs of aciniform spigots. An additional two pairs of tubuliform spigots in females and 70–80 (female) or 100–120 (male) pairs of aciniform spigots were counted on the spinning surfaces of the posterior spinnerets in both sexes. Although the coelotine spiders use their silk to catch prey, P. spinivulva characteristically do not have a typical “triad” spigot, including a flagelliform and two aggregate spigots, for capture thread production.     

Characterization of microsatellite loci in Primula modesta Bisset et Moore (Primulaceae)

Eleven microsatellite loci were isolated from alpine plant Primula modesta. Enriched repeat libraries were constructed and screened by colony hybridization. Forty‐eight polymerase chain reaction (PCR) primer pairs were designed, of which 11 pairs producted successful amplification. A total of 31–35 adult individuals were genotyped for allelic diversity. The polymorphism ranged from three to 14 alleles, and the heterozygosity ranged from 0.161 to 0.828. No linkage disequilibrium was found between these 11 loci.     

Genome‐wide association study for semen traits of the bulls in Chinese Holstein

A genome‐wide association study (GWAS) was performed to identify markers and candidate genes for five semen traits in the Holstein bull population in China. The analyzed dataset consisted of records from 692 bulls from eight bull stations; each bull was genotyped using the Illumina BovineSNP50 BeadChip. Association tests between each trait and the 41 188 informative high‐quality SNPs were achieved with gapit software. In total, 19 suggestive significant SNPs, partly located within the reported QTL regions or within or close to the reported candidate genes, associated with five semen traits were detected. By combining our GWAS results with the biological functions of these genes, eight novel promising candidate genes, including ETNK1, PDE3A, PDGFRB, CSF1R, WT1, DSCAML1, SOD1 and RUNX2, were identified that potentially relate to semen traits. Our findings may provide a basis for further research on the genetic mechanism of semen traits and marker‐assisted selection of such traits in Holstein bulls.     

Isolation and characterization of microsatellite loci from yellowtail Seriola quinqueradiata and cross‐species amplification within the genus Seriola

We developed five microsatellite primer pairs for the yellowtail Seriola quinqueradiata. The loci were highly polymorphic, with eight to 14 alleles per locus, and can be used to study kinship and/or population structure. Many of these primer pairs amplified polymorphic loci in cross‐species amplification tests for two other Seriola species (S. lalandi and S. dumerili).     

Characterization of polymorphic microsatellite loci for the invasive monk parakeet (Myiopsitta monachus)

Microsatellite loci were characterized for the monk parakeet (Myiopsitta monachus) from a GT_n‐enriched genomic library. Twelve of 14 microsatellite loci were polymorphic, averaging 6.7 alleles per locus across the 20 individuals genotyped. Mean expected heterozygosity was 0.72, with locus‐specific values ranging from 0.53 to 0.90. An equally high multilocus probability of identity (2.48 × 10^?12) was revealed for this set of loci. In addition, all 12 loci were demonstrated to cross‐amplify to varying extents within three additional parrot genera suggesting their potential utility for population‐level studies in a broad range of Neotropical psittacines.     

Genetic variation in piñon pine, Pinus edulis, associated with summer precipitation

Three previous reports of microgeographical variation of glycerate dehydrogenase (Gly) frequencies in piñon, Pinus edulis, established the hypothesis that Gly frequencies contribute to adaptation to heterogeneous environments, specifically to variation in soil moisture. In each of these studies, the frequency of the Gly‐3 allele or of Gly‐33 homozygotes was higher on dry sites than on nearby moist sites. Here we attempt to extend these observations by testing the hypothesis that Gly frequencies respond to soil moisture variation on a range‐wide scale. Gly frequencies were surveyed in 11 natural populations, and the frequency of the Gly‐3 allele varied from 0.27 to 0.65 among the sample sites. Elevation varied from 1650 to 3100 m, and summer precipitation, defined as precipitation from April to August, varied from 13.7 to 26.4 cm. The soil types at the collection sites were schist, quaternary volcanic or a mixture of shale and sandstone. Logistic regression revealed that Gly frequencies did not respond to either elevation or soil type, but were related to summer precipitation (P < 0.01). The correlation between summer precipitation and the frequency of the Gly‐3 allele was r = ?0.92 (P < 0.001). Thus, the patterns of differentiation on microgeographical scales are consistent with greater differentiation on a range‐wide scale.     

Mating system and kin relationship between adults and young in the shell-brooding cichlid fish <Emphasis Type="Italic">Neolamprologus meeli</Emphasis> in Lake Tanganyika

Mating system and parental behavior of ten monogamous pairs and two polygynous groups of the Tanganyikan cichlid Neolamprologus meeli were observed in their natural habitat. The home ranges of males and females overlapped with each other. Most groups included one to six young. Paternal and maternal relationships were determined for 22 young from DNA microsatellite markers. Three types of kinship were found: (I) kinship to both the male and female; (II) kinship to females only; and (III) non-kinship to both sexes. In the groups with type II young, step-fathering or sneaking may have occurred. Type III young were larger than type I, suggesting that the former were of sufficient size to leave their birth nest and settle in the territories of foster parents. Both males and females drove out potential predators of young (including three species of Lepidiolamprologus) as a parental behavior. Adults with type III young attacked approaching predators with as much frequency as those with type I young only, indicating that they provided alloparental care. Adults and young swam together, but, a significant difference existed in the frequencies of interactions between adults versus kin young and adults versus non-kin young. The results suggest that both adults and young recognized kin. Electronic Publication     

Development and characterization of microsatellite markers in a clonal plant,Dioscorea japonica Thunb.

We developed 10 microsatellite loci from genomic DNA of a dioecious clonal plant, Dioscorea japonica. Out of 384 clones, 148 contained microsatellite repeats. Polymerase chain reaction primer pairs were designed for 95 of these clones from their sequence data, of which, 10 pairs produced successful amplification. Thirty‐eight individuals were genotyped for allelic diversity. We detected three to nine alleles per locus, and the expected heterozygosity ranged from 0.461 to 0.851.     

Detection of Laribacter hongkongensis using species-specific duplex PCR assays targeting the 16S rRNA gene and the 16S-23S rRNA intergenic spacer region (ISR)

Aims: For the rapid detection of Laribacter hongkongensis, which is associated with human community‐acquired gastroenteritis and traveller’s diarrhoea, we developed a duplex species‐specific PCR assay. Methods and Results: Full‐length of the 16S–23S rRNA intergenic spacer region (ISR) sequences of 52 L. hongkongensis isolates were obtained by PCR‐based sequencing. Two species‐specific primer pairs targeting 16S rRNA gene and ISR were designed for duplex PCR detection of L. hongkongensis. The L. hongkongensis species‐specific duplex PCR assay showed 100% specificity, and the minimum detectable level was 2·1 × 10^?2 ng μl^?1 genomic DNA which corresponds to 5000 CFU ml^?1. Conclusions: The high specificity and sensitivity of the assay make it suitable for rapid detection of L. hongkongensis. Significance and Impact of the Study: This species‐specific duplex PCR method provides a rapid, simple, and reliable alternative to conventional methods to identify L. hongkongensis and may have applications in both clinical and environmental microbiology.     

Using microsatellite diversity in wild Anegada iguanas (<Emphasis Type="Italic">Cyclura pinguis</Emphasis>) to establish relatedness in a captive breeding group of this critically endangered species

Awareness of the genealogical relationships between founder animals in captive breeding programs is essential for the selection of mating pairs that maintain genetic diversity. If captive founder relationships are unknown they can be inferred using genetic data from wild populations. Here, we report the results of such an analysis for six Cyclura pinguis (Sauria: Iguanidae) acquired as adults in 1999 by the San Diego Zoo Institute for Conservation Research to begin a captive breeding program for this critically endangered species. The six founder animals were reportedly hatched in captivity from eggs collected on Anegada in 1985. No records exist, however, as to where on Anegada the eggs were collected or from how many nests they originated. To assist determination of genealogical relationships, we genotyped the six captive founders, their first six offspring, and 33 wild adult iguanas from Anegada at 23 informative microsatellite loci. With these data, we estimated allele frequencies among the wild samples and then estimated the relatedness of the captive population. Using likelihood inference, we determined that three closely related pairs exist among the six captive founders and that each pair is not closely related to the other two. In addition, we were able to assign parentage for all six of the founders’ offspring tested, one of which had been previously misdiagnosed. Using the assigned parentage and inferred relatedness of the six founders, we calculated mean kinship for each of the six founders and their five living offspring. Finally, based on the allelic diversity of the wild iguanas sampled, we conclude that the C. pinguis population on Anegada is not excessively inbred; however, further investigation is warranted.     

Genetic variation and population structure in black-grass (Alopecurus myosuroides Huds.), a successful, herbicide-resistant, annual grass weed of winter cereal fields

Black‐grass (Alopecurus myosuroides) is an allogamous grass weed common in cereal fields of northern Europe, which developed resistance to a widely used family of herbicides, the ACCase‐inhibiting herbicides. Resistance is caused by mutations at the ACCase gene and other, metabolism‐based, mechanisms. We investigated the genetic structure of 36 populations of black‐grass collected in one region of France (Côte d’Or), using 116 amplified fragment length polymorphism (AFLP) loci and sequence data at the ACCase gene. The samples were characterized for their level of herbicide resistance and genotyped for seven known ACCase mutations conferring resistance. All samples contained herbicide‐resistant plants, and 19 contained ACCase mutations. The genetic diversity at AFLP loci was high (H_T = 0.246), while differentiation among samples was low (F_ST = 0.023) and no isolation by distance was detected. Genetic diversity within samples did not vary with the frequency of herbicide resistance. A Bayesian algorithm was used to infer population structure. The two genetic clusters inferred were not associated with any geographical structure or with herbicide resistance. A high haplotype diversity (H_d = 0.873) and low differentiation (G_ST = 0.056) were observed at ACCase. However, haplotype diversity within samples decreased with the frequency of ACCase‐based resistance. We suggest that the genetic structure of black‐grass is affected by its recent expansion as a weed. Our data demonstrate that the strong selection imposed by herbicides did not modify the genome‐wide genetic structure of an allogamous weed that probably has large effective population sizes. Our study gives keys to a better understanding of the evolution of successful, noxious weeds in modern agriculture.     

Can chilling tolerance of C4 photosynthesis in Miscanthus be transferred to sugarcane?

The goal of this study was to investigate whether chilling tolerance of C₄ photosynthesis in Miscanthus can be transferred to sugarcane by hybridization. Net leaf CO₂ uptake (A_sat) and the maximum operating efficiency of photosystem II (Ф_PSII) were measured in warm conditions (25 °C/20 °C), and then during and following a chilling treatment of 10 °C/5 °C for 11 day in controlled environment chambers. Two of three hybrids (miscanes), ‘US 84‐1058’ and ‘US 87‐1019’, did not differ significantly from the chilling tolerant M. ×giganteus ‘Illinois’ (Mxg), for A_sat, and Φ_PSII measured during chilling. For Mxg grown at 10 °C/5 °C for 11 days, A_sat was 4.4 μmol m^?2 s^?1, while for miscane ‘US 84‐1058’ and ‘US 87‐1019’, A_sat was 5.7 and 3.5 μmol m^?2 s^?1, respectively. Miscanes ‘US 84‐1058’ and ‘US 87‐1019’ and Mxg had significantly higher rates of A_sat during chilling than three tested sugarcanes. A third miscane showed lower rates than Mxg during chilling, but recovered to higher rates than sugarcane upon return to warm conditions. Chilling tolerance of ‘US 84‐1058’ was further confirmed under autumn field conditions in southern Illinois. The selected chilling tolerant miscanes have particular value for biomass feedstock and biofuel production and at the same time they can be a starting point for extending sugarcane's range to colder climates.     

Maintenance of genetic and morphological identity in two sibling Syrrhopodon species (Calymperaceae,Bryopsida) despite extensive introgression

Bryophytes are a group of land plants in which the role of hybridization has long been challenged. Using genotyping by sequencing to circumvent the lack of molecular variation at selected loci previously used for phylogeny and morphology, we determine the level of genetic and morphological divergence and reproductive isolation between the sibling Syrrhopodon annotinus and S. simmondsii (Calymperaceae, Bryopsida) that occur in sympatry but in different habitats in lowland Amazonian rainforests. A clear morphological differentiation and a low (0.06), but significant Fst derived from the analysis of 183 single nucleotide polymorphisms were observed between the two species. Conspecific pairs of individuals consistently exhibited higher average kinship coefficients along a gradient of geographic isolation than interspecific pairs. The weak, but significant genetic divergence observed is consistent with growing evidence that ecological specialization can lead to genetic differentiation among bryophyte species. Nevertheless, the spatial genetic structures of the two species were significantly correlated, as evidenced by the significant slope of the Mantel test based on kinship coefficients between pairs of interspecific individuals and the geographic distance separating them. Interspecific pairs of individuals are thus more closely related when they are geographically closer, suggesting that isolation‐by‐distance is stronger than the interspecific reproductive barrier and pointing to interspecific gene flow. We conclude that interspecific introgression, whose role has long been questioned in bryophytes, may take place even in species wherein sporophyte production is scarce due to dioicy, raising the question as to what mechanisms maintain differentiation despite weak reproductive isolation.     

Barcode haplotype variation in north American agroecosystem lady beetles (Coleoptera: Coccinellidae)

DNA barcodes have proven invaluable in identifying and distinguishing insect pests, most notably for determining the provenance of exotic invasives, but relatively few insect natural enemies have been barcoded. We used Folmer et al.’s (1994) universal invertebrate primers and Hebert et al.’s (2004) for Lepidoptera, to amplify 658 bp at the 5′ end of the mitochondrial cytochrome oxidase c subunit I (COI) gene in five species of lady beetles from crop fields in six states in the US Mid‐Atlantic, Plains and Midwest: three native species, Hippodamia convergens Guérin‐Méneville, H. parenthesis (Say) and Coleomegilla maculata (De Geer); and two exotic species, Harmonia axyridis (Pallas) and Coccinella septempunctata Linnaeus. Sequence divergences within species were low, never exceeding 0.9% (Kimura 2‐parameter distances). Sequence divergences between the two Hippodamia species ranged from 14.7 to 16.4%, mirroring the relationships found for other arthropod taxa. Among the exotic species, C. septempunctata sequences were as variable as those of the three native species, while H. axyridis populations comprised a single haplotype. Limited data on two Coleomegilla subspecies, C. m. lengi Timberlake and C. m. fuscilabris (Mulsant), are consistent with their belonging to the same species, although morphological and reproductive data indicate that they represent separate species. Our results support the general utility of COI barcodes for distinguishing and diagnosing coccinellid species, but point to possible limitations in the use of barcodes to resolve species assignments in recently divergent sibling species.     

Stereology and computer assisted three-dimensional reconstruction as tools to study probiotic effects of Aeromonas hydrophila on the digestive tract of germ-free Artemia franciscana nauplii

Aims: Validation of stereology and three‐dimensional reconstruction for monitoring the probiotic effect of Aeromonas hydrophila on the gut development of germ‐free Artemia franciscana nauplii. Methods and Results: Germ‐free Artemia nauplii were cultured using Baker’s yeast and dead Aer. hydrophila. Live Aer. hydrophila were added on the first day to the treatment group. The gut length and volume were monitored on days two and four using stereology and three‐dimensional reconstruction. Both methods showed comparable results. Stereology was least labour intensive to estimate volumes, while three‐dimensional reconstructions rendered architectural and topographical data of the gut. Moreover, a positive effect of probiotic bacterium, Aer. hydrophila is likely. Conclusion: Slight increment in the growth of the digestive tract of A. franciscana nauplii exerted by probiotic bacteria could be detected using stereology and three‐dimensional reconstruction. Significance and Impact of the Study: The gnotobiotic Artemia rearing system is unique to investigate the effects of micro‐organisms on the development of nauplii. However, in the base of this model system, only survival counts and length measurements exist as monitoring tools. Therefore, additional tools such as stereology and three‐dimensional reconstruction are prerequisite to obtain more powerful analysis.     

Population genetic correlates of declining transmission in a human pathogen

Pathogen control programs provide a valuable, but rarely exploited, opportunity to directly examine the relationship between population decline and population genetics. We investigated the impact of an ~12‐fold decline in transmission on the population genetics of Plasmodium falciparum infections (n = 1731) sampled from four clinics on the Thai–Burma border over 10 years and genotyped using 96 genome‐wide SNPs. The most striking associated genetic change was a reduction in the frequency of infections containing multiple parasite genotypes from 63% in 2001 to 14% in 2010 (P = 3 × 10^?15). Two measures of the clonal composition of populations (genotypic richness and the β‐parameter of the Pareto distribution) declined over time as more people were infected by parasites with identical multilocus genotypes, consistent with increased selfing and a reduction in the rate at which multilocus genotypes are broken apart by recombination. We predicted that the reduction in transmission, multiple clone carriage and outbreeding would be mirrored by an increased influence of genetic drift. However, geographical differentiation and expected heterozygosity remained stable across the sampling period. Furthermore, N_e estimates derived from allele frequencies fluctuation between years remained high (582 to ∞) and showed no downward trend. These results demonstrate how genetic data can compliment epidemiological assessments of infectious disease control programs. The temporal changes in a single declining population parallel to those seen in comparisons of parasite genetics in regions of differing endemicity, strongly supporting the notion that reduced opportunity for outbreeding is the key driver of these patterns.