Consequences of genetic incompatibility on fitness and mate choice: the male point of view

Hymenopterans under single‐locus complementary sex determination (sl‐CSD) face inbreeding costs due to this sex determination mode. Under sl‐CSD, homozygote eggs at the sl‐CSD locus usually develop into unviable or sterile diploid males. Production of such costly males increases when sib‐mating happens because related individuals share half of their genome. In the hymenopteran Venturia canescens (a solitary parasitoid wasp), diploid males are sterile, leading to fitness costs through genetic incompatibility between parents. Whereas the costs of producing diploid males and behavioural strategies that would reduce such costs have been studied in females, the potential fitness costs faced by males have not. Here, we aimed to investigate fitness costs that males endure after a single sib‐mating and tested whether they have the ability to avoid sib‐mating through kin recognition. Our results show that males have a reduced fitness (i.e. they produce fewer daughters) when mating with their sibs. We also show that males have the ability to distinguish between non‐sib and sib females (i.e. kin). They use chemical marks emitted by the females to discriminate kin from non‐kin. We discuss the evolution of kin recognition in males in the context of mate choice for genetic compatibility. © 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2015, 114 , 279–286.     

Inbreeding in a natural population of the gregarious parasitoid wasp Cotesia glomerata

Inbreeding occurs in numerous animal and plant species. In haplodiploid hymenopterans with the widespread single locus complementary sex determination, the frequency of diploid males, which are produced at the expense of females, is increased under inbreeding. Diploid males in species of bees, ants and wasps are typically either unviable or effectively sterile and thus impose a severe genetic load on populations. However, a recent study indicated that diploid males can be reproductive in the gregarious parasitoid wasp Cotesia glomerata, effectively reducing the diploid male load. To understand the role of inbreeding as a potential selective pressure towards the evolution of diploid male fertility, we genotyped specimens collected in the field at four locations using microsatellite markers to estimate the ratio of sibling matings under natural conditions. Results show that more than half of all matings involved siblings. We argue that the frequent occurrence of inbreeding has driven the evolution of diploid male fertility.     

Insects and incest: Sib‐mating tolerance in natural populations of a parasitoid wasp

Sib‐mating avoidance is a pervasive behaviour that is expected to evolve in species subject to inbreeding depression. Although laboratory studies provide elegant demonstrations, small‐scaled bioassays minimize the costs of mate finding and choice, and thus may produce spurious findings. We therefore combined laboratory experiments with field observations to examine the existence of inbreeding avoidance using the parasitoid wasp Venturia canescens. In the laboratory, our approach consisted of mate‐choice experiments to assess kin discrimination in population cages with competitive interactions. A higher mating probability after sib rejections suggested that females could discriminate their sibs; however, in contrast to previous findings, sib‐mating avoidance was not observed. To compare our laboratory results to field data, we captured 241 individuals from two populations. Females laid eggs in the lab, and 226 daughters were obtained. All individuals were genotyped at 18 microsatellite loci, which allowed inference of the genotype of each female's mate and subsequently the relatedness within each mating pair. We found that the observed rate of sib‐mating did not differ from the probability that sibs encountered one another at random in the field, which is consistent with an absence of sib‐mating avoidance. In addition, we detected a weak but significant male‐biased dispersal, which could reduce encounters between sibs. We also found weak fitness costs associated with sib‐mating. As such, the sex‐biased dispersal that we found is probably sufficient to mitigate these costs. These results imply that kin discrimination has probably evolved for purposes other than mate choice, such as superparasitism avoidance.     

Host switching in a generalist parasitoid: contrasting transient and transgenerational costs associated with novel and original host species

Parasitoids face challenges by switching between host species that influence survival and fitness, determine their role in structuring communities, influence species invasions, and affect their importance as biocontrol agents. In the generalist parasitoid, Venturia canescens (Gravenhorst) (Hymenoptera: Ichneumonidae), we investigated the costs in encapsulation, survival, and body size on juveniles when adult parasitoids switched from their original host, Plodia interpunctella (Hübner) (Lepidotera, Pyralidae) to a novel host, Ephestia kuehniella (Zeller) (Lepidoptera, Pyralidae), over multiple generations. Switching had an initial survival cost for juvenile parasitoids in the novel host, but increased survival occurred within two generations. Conversely, mortality in the original host increased. Body size, a proxy for fecundity, also increased with the number of generations in the novel host species, reflecting adaptation or maternal effects due to the larger size of the novel host, and therefore greater resources available to the developing parasitoid. Switching to a novel host appears to have initial costs for a parasitoid, even when the novel host may be better quality, but the costs rapidly diminish. We predict that the net cost of switching to a novel host for parasitoids will be complex and will depend on the initial reduction in fitness from parasitizing a novel host versus local adaptations against parasitoids in the original host.     

Isolation and characterization of twelve polymorphic microsatellite markers in the endangered Hopea hainanensis (Dipterocarpaceae)

Microsatellite markers were isolated and characterized for Hopea hainanensis Merrill & Chun, an endangered tree species with scattered distribution in Hainan Island and northern Vietnam. Twenty‐six microsatellite markers were developed based on next‐generation sequencing data and were genotyped by capillary electrophoresis on an ABI 3730xl DNA Analyzer. Twelve markers were found to be polymorphic in H. hainanensis. GENODIVE analyses indicated that the number of alleles ranged from 2 to 6 per locus, and the observed and expected heterozygosity varied from 0 to 0.755 and from 0.259 to 0.779, respectively. Primer transferability was tested with Hopea chinensis Hand.‐Mazz. and Hopea reticulata Tardieu, in which 3 and 7 microsatellite markers were found to be polymorphic, separately. The results showed that H. reticulata and H. hainanensis had similar levels of genetic diversity. A neighbor joining dendrogram clustered all individuals into two major groups, one of which was exclusively constituted by H. hainanensis, while the other consisted of two subgroups, corresponding to H. reticulata and H. chinensis, respectively. The 12 polymorphic microsatellite markers could be applied to study genetic diversity, population differentiation, mating system, and fine‐scale spatial genetic structures of H. hainanensis as well as its close relatives, facilitating the conservation and restoration of these endangered but valuable Hopea species.     

利用17个微卫星标记分析鳙鱼的遗传多样性

选用本实验室克隆的17个鳙鱼微卫星分子标记分析四川泸州和江西鄱阳湖的两个种群鳙鱼的遗传多样性及种质特性,计算和统计了杂合度、多态信息含量（PIC）、有效等位基因数、等位基因频率、遗传距离、遗传相似系数、Hardy-Weinberg平衡偏离指数等方面内容。结果表明：选择使用17个微卫星标记,其中有4个为单态标记,13个为多态标记。江西和四川鳙鱼群体每个微卫星位点的平均等位基因数分别为3.325及3.882,平均有效等位基因数分别为3.531及2.676,多态位点百分率分别为82.4及70.5, 17个微卫星标记共有等位基因71个,多态微卫星位点的PIC在0.114~0.960之间变动,平均为0.417 ,两群体位点平均观测杂合度为0.385和0.452,平均期望杂合度为0.360和0.422,两个群体间的遗传相似系数为0.897,群体间的遗传距离为0.109。     

Deacetylation biocatalysis and elicitation by immobilized Penicillium canescens in Astragalus membranaceus hairy root cultures: towards the enhanced and sustainable production of astragaloside IV

A novel biotechnology approach by combining deacetylation biocatalysis with elicitation of immobilized Penicillium canescens (IPC) in Astragalus membranaceus hairy root cultures (AMHRCs) was proposed for the elevated production of astragaloside IV (AG IV). The highest AG IV accumulation was achieved in 36‐day‐old AMHRCs co‐cultured with IPC for 60 h, which resulted in the enhanced production of AG IV by 14.59‐fold in comparison with that in control (0.193 ± 0.007 mg/g DW). Meanwhile, AG IV precursors were almost transformed to AG IV by IPC deacetylation. Moreover, expression of genes involved in AG IV biosynthetic pathway was significantly up‐regulated in response to IPC elicitation. Also, FTIR and SEM showed that cell wall lignification was enhanced following IPC treatment and root surface was likely to be IPC deacetylation site. Overall, dual roles of IPC (biocatalyst and elicitor) offered an effective and sustainable way for the mass production of AG IV in AMHRCs.     

Presence of a novel small RNA-containing virus in a laboratory culture of the endoparasitic wasp Venturia canescens (Hymenoptera: Ichneumonidae)

Parasitoid wasps use a variety of mechanisms to alter their host's physiology to the benefit of the developing endoparasite inside the host larva. Association of certain wasps with viruses and virus-like particles (VLPs) that contribute to their success in parasitism is one of the fascinating evolutionary adaptations conferring active or passive protection for the endoparasite from the host immune system. Venturia canescens has been shown to produce VLPs that provide protection for the developing parasitoid egg inside the host, Ephestia kuehniella. Here, we report on the presence of a novel small RNA-containing virus from V. canescens, designated as VcSRV, occurring in the ovaries of the wasp. The virus particles are found together with VcVLPs in the lumen of the calyx region of the ovaries and are injected together with the egg and VcVLPs into E. kuehniella larvae where they enter hemocytes. Alignment of the RNA-dependent RNA polymerase gene of VcSRV indicates that the virus most likely belongs to the recently described genus Iflavirus.     

Heterozygosity correlates with body size,nest site quality and productivity in a colonial waterbird,the whiskered tern (Chlidonias hybrida,Aves: Sternidae)

Positive effects of individual heterozygosity on naturally selected traits have been reported in wild populations of many animal taxa. The aim of this study was to test whether heterozygosity predicts the quality of acquired nest sites and productivity in a colonially breeding waterbird, the whiskered tern (Chlidonias hybrida). For this purpose, 40 adult terns from a small, recently established population in Central Poland were typed at eight microsatellite loci. We demonstrate that individual heterozygosity is positively related to hatching success. We hypothesize that this association could be mediated by direct effects of heterozygosity on the competitive abilities of individuals. We found that more heterozygous terns tended to breed in better protected central parts of the colony, suggesting that they had capabilities of outcompeting less heterozygous individuals and relegating them to the less attractive peripheries of the colony. It was also demonstrated that the link between heterozygosity and individual abilities to acquire more attractive nest site could be mediated by the larger size of heterozygous individuals. Although no correlations between heterozygosity and different components of condition were found, there was a positive association between female heterozygosity and both clutch size and egg size. We suggest that demonstrated heterozygosity‐fitness correlations could be primarily caused by inbreeding depression in the studied whiskered tern population.     

Evolution of life‐history traits and mating strategy in males: a case study on two populations of a Drosophila parasitoid

Abiotic and biotic factors affect life‐history traits and lead populations to exhibit different behavioural strategies. Due to the direct link between their behaviour and fitness, parasitoid females have often been used to test the theories explaining these differences. In male parasitoids, however, such investigations are vastly understudied, although their mating strategy directly determines their fitness. In this study, we compared the pattern of life history traits and the mating strategy of males in two populations of the Drosophila parasitoid Asobara tabida, exposed to different biotic and abiotic conditions, with the major difference being that one of them was recently exposed to strong competition with the dominant competitor Leptopilina boulardi after recent climate change, the other population being settled in a location where L. boulardi has not been recorded. The results showed that individuals of both populations have a different reproductive strategy: in one population, females produced a more female‐biased sex ratio, while males accumulated more lipids during their larval development, invested more energy in reproduction and decreased their locomotor activity, suggesting a higher proportion of matings on their emergence patch, all of these factors being possibly linked to the new competition pressure. In both populations, mating without sperm transfer may persist for several days after males become sperm‐depleted, and may be more frequent than mating with sperm transfer over their whole lifespan. This point is discussed from an evolutionary point of view.     

Development of diagnostic microsatellite markers from whole‐genome sequences of Ammodramus sparrows for assessing admixture in a hybrid zone

Studies of hybridization and introgression and, in particular, the identification of admixed individuals in natural populations benefit from the use of diagnostic genetic markers that reliably differentiate pure species from each other and their hybrid forms. Such diagnostic markers are often infrequent in the genomes of closely related species, and genomewide data facilitate their discovery. We used whole‐genome data from Illumina HiSeqS2000 sequencing of two recently diverged (600,000 years) and hybridizing, avian, sister species, the Saltmarsh (Ammodramus caudacutus) and Nelson's (A. nelsoni) Sparrow, to develop a suite of diagnostic markers for high‐resolution identification of pure and admixed individuals. We compared the microsatellite repeat regions identified in the genomes of the two species and selected a subset of 37 loci that differed between the species in repeat number. We screened these loci on 12 pure individuals of each species and report on the 34 that successfully amplified. From these, we developed a panel of the 12 most diagnostic loci, which we evaluated on 96 individuals, including individuals from both allopatric populations and sympatric individuals from the hybrid zone. Using simulations, we evaluated the power of the marker panel for accurate assignments of individuals to their appropriate pure species and hybrid genotypic classes (F1, F2, and backcrosses). The markers proved highly informative for species discrimination and had high accuracy for classifying admixed individuals into their genotypic classes. These markers will aid future investigations of introgressive hybridization in this system and aid conservation efforts aimed at monitoring and preserving pure species. Our approach is transferable to other study systems consisting of closely related and incipient species.     

Characterization of new microsatellite loci in Pieris amamioshimensis (Ericaceae), a species nearly extinct in the wild

A set of 11 polymorphic microsatellite markers has been developed and characterized for the critically endangered species Pieris amamioshimensis. Fifty‐nine individuals of an ex‐situ population were used to identify these markers. The total number of alleles for each locus ranged from 3 to 9, with an average of 5.4. The expected heterozygosities (H_S) and observed heterozygosities (H_O) ranged from 0.47 to 0.77 and 0.22 to 0.88, respectively. In total, four loci exhibited significant deviations from Hardy–Weinberg equilibrium: two loci showed significant heterozygosity excess and the other two loci showed significant heterozygosity deficit. The polymorphism information content (0.43 ≤ PIC ≤ 0.73), the probability of exclusions (PE₁ = 0.9565, PE₂ = 0.9969 and PE₃ = 0.9999) and probabilities for identity (PI = 3.78 × 10^?9 and PI‐Sib = 2.35 × 10^?4) suggest that these markers are useful for estimating not only genetic diversity but also parentage, for the ex‐situ conservation management of populations.     

Population size and time since island isolation determine genetic diversity loss in insular frog populations

Understanding the factors that contribute to loss of genetic diversity in fragmented populations is crucial for conservation measurements. Land‐bridge archipelagoes offer ideal model systems for identifying the long‐term effects of these factors on genetic variations in wild populations. In this study, we used nine microsatellite markers to quantify genetic diversity and differentiation of 810 pond frogs (Pelophylax nigromaculatus) from 24 islands of the Zhoushan Archipelago and three sites on nearby mainland China and estimated the effects of the island area, population size, time since island isolation, distance to the mainland and distance to the nearest larger island on reduced genetic diversity of insular populations. The mainland populations displayed higher genetic diversity than insular populations. Genetic differentiations and no obvious gene flow were detected among the frog populations on the islands. Hierarchical partitioning analysis showed that only time since island isolation (square‐root‐transformed) and population size (log‐transformed) significantly contributed to insular genetic diversity. These results suggest that decreased genetic diversity and genetic differentiations among insular populations may have been caused by random genetic drift following isolation by rising sea levels during the Holocene. The results provide strong evidence for a relationship between retained genetic diversity and population size and time since island isolation for pond frogs on the islands, consistent with the prediction of the neutral theory for finite populations. Our study highlights the importance of the size and estimated isolation time of populations in understanding the mechanisms of genetic diversity loss and differentiation in fragmented wild populations.     

基于微卫星标记的中国枣食芽象甲地理种群遗传多样性分析

【目的】枣食芽象甲Scythropus yasumatsui是我国北方枣树Zizyphus jujuba上的一种重要的灾害性害虫,在陕西、山西、河北、河南等枣树主产区普遍发生。本研究旨在揭示我国枣食芽象甲不同种群间的遗传分化和基因交流规律。【方法】利用枣食芽象甲转录组测序的SSR序列,使用荧光标记PCR及毛细管电泳分型方法,筛选出8个微卫星位点,对我国5个省份(山西、陕西、宁夏、河北和河南)10个地理种群共308头枣食芽象甲样本进行种群遗传多样性分析。【结果】8个SSR位点均存在无效等位基因且偏离哈迪 温伯格平衡。各位点的有效等位基因数(Ne)为2.113~8.016,多态信息含量(PIC)为0.561~0.908,期望杂合度(He)为0.476~0.865;种群间遗传分化系数(Fst)平均值为0.151;基因流(Nm)平均值为1.594。枣食芽象甲种群间遗传分化系数与地理距离之间显著正相关(r=0.596, P=0.0035),基于Nei’s遗传距离和Cavalli-Sforza & Edwards余弦遗传距离的系统进化树将10个地理种群均聚为3个相同的分支。【结论】结果说明,枣食芽象甲种群遗传多样性较高,不同地理种群间存在高度的遗传分化,且有一定的基因交流;地理隔离是影响枣食芽象甲地理种群遗传分化和基因交流的重要因素。     

Intraspecific genetic variation in the common midwife toad (Alytes obstetricans): subspecies assignment using mitochondrial and microsatellite markers

The common midwife toad (Alytes obstetricans), widely distributed in the northern half of the Iberian Peninsula and part of Western Europe, is currently subdivided into four subspecies: A. o. obstetricans, A. o. boscai, A. o. pertinax and A. o. almogavarii. However, the delimitation of these subspecies and their ranges are still under discussion because strong discordances have been found between morphological and molecular data, and especially among different genetic markers. Here, we screen a set of novel microsatellite loci and mtDNA sequences of A. obstetricans populations representative of all currently recognized subspecies to investigate the correspondence between genetic groupings inferred from clustering analysis of microsatellite genotypes and the described subspecies and test whether patterns of mtDNA variation are concordant with those genetic clusters. Our results confirm previous expectations of extremely high intraspecific diversity in A. obstetricans in Iberia. Analyses of microsatellite and mtDNA data were concordant in recovering five well‐defined groups, of which three correspond to previously defined subspecies, while the two additional clusters correspond to populations of subspecies A. o. boscai separated by the Douro River. Our results suggest the occurrence of two distinct genetic units within A. o. boscai that likely result from a long independent evolutionary history, thus deserving special attention from a conservation point of view.     

Development and characterization of 20 polymorphic microsatellite markers for Livistona chinensis R. Br. var. subglobosa Becc., a fan palm in the Ryukyu Archipelago,Japan

Livistona chinensis R. Br. var. subglobosa Becc. is a subtropical fan palm distributed mainly in the islands of the Ryukyu Archipelago, Japan. Some small populations, separated by a few 100 km from the main species range, are also established on islands around northern Kyushu and Cape Ashizuri, Shikoku. Most of those are designated as national or prefectural natural treasures. We developed 20 microsatellite markers for this species and characterized them for polymorphisms within six populations. The number of alleles per locus ranged from two to nine, with an average of 4.2; the expected heterozygosity ranged from 0.16 to 0.61, with an average of 0.46. These markers will be useful for investigating disjunct range formation within L. chinensis var. subglobosa and the origins of the remote populations, providing crucial information for conservation of the populations.     

Changes in genetic diversity and differentiation in Red‐cockaded woodpeckers (Dryobates borealis) over the past century

Red‐cockaded woodpeckers (RCW; Dryobates borealis) declined after human activities reduced their fire‐maintained pine ecosystem to <3% of its historical range in the southeastern United States and degraded remaining habitat. An estimated 1.6 million RCW cooperative breeding groups declined to about 3,500 groups with no more than 10,000 birds by 1978. Management has increased RCW population abundances since they were at their lowest in the 1990s. However, no range‐wide study has been undertaken since then to investigate the impacts of this massive bottleneck or infer the effects of conservation management and recent demographic recoveries. We used mitochondrial DNA sequences (mtDNA) and nine nuclear microsatellite loci to determine if range‐wide demographic declines resulted in changes to genetic structure and diversity in RCW by comparing samples collected before 1970 (mtDNA data only), between 1992 and 1995 (mtDNA and microsatellites), and between 2010 and 2014 (mtDNA and microsatellites). We show that genetic diversity has been lost as detected by a reduction in the number of mitochondrial haplotypes. This reduction was apparent in comparisons of pre‐1970 mtDNA data with data from the 1992–1995 and 2010–2014 time points, with no change between the latter two time points in mtDNA and microsatellite analyses. The mtDNA data also revealed increases in range‐wide genetic differentiation, with a genetically panmictic population present throughout the southeastern United States in the pre‐1970s data and subsequent development of genetic structure that has remained unchanged since the 1990s. Genetic structure was also uncovered with the microsatellite data, which like the mtDNA data showed little change between the 1992–1995 and 2010–2014 data sets. Temporal haplotype networks revealed a consistent, star‐like phylogeny, suggesting that despite the overall loss of haplotypes, no phylogenetically distinct mtDNA lineages were lost when the population declined. Our results may suggest that management during the last two decades has prevented additional losses of genetic diversity.     

Exploring the role of asexual multiplication in poplar rust epidemics: impact on diversity and genetic structure

Fungal plant pathogens, especially rust fungi (Pucciniales), are well known for their complex life cycles, which include phases of sexual and asexual reproduction. The effect of asexual multiplication on population genetic diversity has been investigated in the poplar rust fungus Melampsora larici‐populina using a nested hierarchical sampling scheme. Four hierarchical levels were considered: leaf, twig, tree and site. Both cultivated and wild poplar stands were sampled at two time points at the start and end of rust epidemics. A total of 641 fungal isolates was analysed using nine microsatellite markers. This study revealed that the genetic signature of asexual multiplication in the wild poplar stand was seen only at lower hierarchical levels (leaf and twig). Moreover, we observed an erosion of clonal structure through time, with an increase in both gene and genotypic diversity. New genotypes contributed to host infection over time, which demonstrates the importance of allo‐infection in the epidemic process in this host‐pathogen system. Compared with the wild stands, the nearly lack of detection of clonal structure in the cultivated stands reflects the higher infection level on cultivated poplars. More generally, this genetic analysis illustrates the utility of population genetics approach for elucidating the proportion of asexual reproduction in the multiplication of isolates during an epidemic, and for proper quantification of asexual dispersal in plant pathogens.     

Using the knowns to discover the unknowns: MS‐based dereplication uncovers structural diversity in 17‐hydroxygeranyllinalool diterpene glycoside production in the Solanaceae

Exploring the diversity of plant secondary metabolism requires efficient methods to obtain sufficient structural insights to discriminate previously known from unknown metabolites. De novo structure elucidation and confirmation of known metabolites (dereplication) remain a major bottleneck for mass spectrometry‐based metabolomic workflows, and few systematic dereplication strategies have been developed for the analysis of entire compound classes across plant families, partly due to the complexity of plant metabolic profiles that complicates cross‐species comparisons. 17‐hydroxygeranyllinalool diterpene glycosides (HGL‐DTGs) are abundant defensive secondary metabolites whose malonyl and glycosyl decorations are induced by jasmonate signaling in the ecological model plant Nicotiana attenuata. The multiple labile glycosidic bonds of HGL‐DTGs result in extensive in‐source fragmentation (IS‐CID) during ionization. To reconstruct these IS‐CID clusters from profiling data and identify precursor ions, we applied a deconvolution algorithm and created an MS/MS library from positive‐ion spectra of purified HGL‐DTGs. From this library, 251 non‐redundant fragments were annotated, and a workflow to characterize leaf, flower and fruit extracts of 35 solanaceous species was established. These analyses predicted 105 novel HGL‐DTGs that were restricted to Nicotiana, Capsicum and Lycium species. Interestingly, malonylation is a highly conserved step in HGL‐DTG metabolism, but is differentially affected by jasmonate signaling among Nicotiana species. This MS‐based workflow is readily applicable for cross‐species re‐identification/annotation of other compound classes with sufficient fragmentation knowledge, and therefore has the potential to support hypotheses regarding secondary metabolism diversification.