Frequency of resistance to Bacillus thuringiensis toxin Cry1Ab in an Iowa population of European corn borer (Lepidoptera: Crambidae)

The refuge plus high-dose strategy for resistance management assumes that the frequency of resistance alleles is low. We used an F2 screen to estimate the frequency of resistance to transgenic corn that produces Bacillus thuringiensis Berliner Cry1Ab toxin (Bt corn) in an Iowa population of European corn borer, Ostrinia nubilalis (Hübner). We also proposed a modification to the statistical analysis of the F2 screen that extends its application for nonuniform prior distributions and for repeated sampling of a single population. Based on a sample of 188 isofemale lines derived from females caught at light traps during the 2nd flight of 1997, we show with 95% confidence that the frequency of resistance to Bt corn was <3.9 x 10(-3) in this Iowa population. These results provide weak evidence that the refuge plus high-dose strategy may be effective for managing resistance in O. nubilalis to Bt corn. Partial resistance to Cry1Ab toxin was found commonly. The 95% CI for the frequency of partial resistance were [8.2 x 10(-4), 9.4 x 10(-3)] for the Iowa population. Variable costs of the method were 14.90 dollars per isofemale line, which was a reduction of 25% compared with our initial estimate.     

Ancestral population reconstitution from isofemale lines as a tool for experimental evolution

Experimental evolution is a powerful tool to study adaptation under controlled conditions. Laboratory natural selection experiments mimic adaptation in the wild with better‐adapted genotypes having more offspring. Because the selected traits are frequently not known, adaptation is typically measured as fitness increase by comparing evolved populations against an unselected reference population maintained in a laboratory environment. With adaptation to the laboratory conditions and genetic drift, however, it is not clear to what extent such comparisons provide unbiased estimates of adaptation. Alternatively, ancestral variation could be preserved in isofemale lines that can be combined to reconstitute the ancestral population. Here, we assess the impact of selection on alleles segregating in newly established Drosophila isofemale lines. We reconstituted two populations from isofemale lines and compared them to two original ancestral populations (AP) founded from the same lines shortly after collection. No significant allele frequency changes could be detected between both AP and simulations showed that drift had a low impact compared to Pool‐Seq‐associated sampling effects. We conclude that laboratory selection on segregating variation in isofemale lines is too weak to have detectable effects, which validates ancestral population reconstitution from isofemale lines as an unbiased approach for measuring adaptation in evolved populations.     

Pedigreed crosses to estimate recessive virulence allele frequencies in natural populations of gall midges

Monitoring changes in rare, recessive allele frequencies in natural populations can be accomplished using pedigreed individuals sampled from these populations. A pedigree keeps track of and limits the mating of sampled individuals, to preserve information about the genotype of the sampled individual in the phenotypes of its descendents. To estimate allele frequencies in a natural population using pedigreed crosses, four relations must be specified: (1) a method to determine whether the pedigreed line carries the desired allele; (2) a method to estimate the phenotypic frequency of the trait among the pedigreed lines and a credibility limit for the estimate; (3) the genetic relation between the phenotype frequency among the lines and the allele frequency in the natural population; and (4) a method to estimate the probability that the first method did not detect the trait, assuming that the allele was present in the sampled individual. Knowledge about the segregation patterns of the allele enables specification of (3) and (4). Bayesian statistics were used to estimate the phenotypic frequency of the trait among the pedigreed lines. The method determining whether the pedigreed line carries the desired allele will vary with the species and trait of concern. We focused on monitoring of vGm1, a recessive autosomal allele, and vGm2, a recessive sex‐linked allele, which provide virulence against certain rice resistance genes in rice gall midge, Orseolia oryzae (Wood‐Mason) (Diptera: Cecidomyiidae). We show how three pedigrees can be used to estimate these allele frequencies. An F₁ field screen challenges the F₁ offspring of sampled individuals on the rice differentials. A P₁ test‐cross mates the sampled individual with a homozygous lab colony for the allele of interest, and evaluates their offspring on the rice differentials. A conditional F₁ test‐cross takes the offspring from pedigrees that were negative in an F₁ field screen, and test‐crosses these offspring with the homozygous laboratory colony. We also indicate how to test for independent assortment when a double (or multiple) homozygote laboratory colony is used in a test‐cross, how to test for differences among samples, and how to pool data to produce a single estimate based on a larger number of pedigreed lines. These methods may encourage the development of a variety of pedigreed monitoring strategies that could improve and prolong the use of scarce plant resistance alleles in rice and other plants.     

The Crossveinless Polygenes in an Iowa Population

A natural population of Drosophila melanogaster was tested for the presence of crossveinless (cve) alleles over the course of a summer. Approximately 8% of the wild-caught females tested carried enough cve alleles to produce true-breeding crossveinless lines. In some cases, different isofemale lines produced identical cve strains. Also, differences were sometimes observed among independently selected cve sublines of a given isofemale line.-The cv-2 allele was found to occur with a frequency of about 1% to 2%.     

Associated LAH Numbers,Factorial Series Distributions and Unbiased Estimation of a Size Parameter

A finite population consists of kN individuals of N different categories with k individuals each. It is required to estimate the unknown parameter N, the number of different classes in the population. A sequential sampling scheme is considered in which individuals are sampled until a preassigned number of repetitions of already observed categories occur in the sample. Corresponding fixed sample size schemes were considered by Charalambides (1981). The sequential sampling scheme has the advantage of always allowing unbiased estimation of the size parameter N. It is shown that relative to Charalambides' fixed sample size scheme only minor adjustments are required to account for the sequential scheme. In particular, MVU estimators of parametric functions are expressible in terms of the C-numbers introduced by Charalambides.     

Planning and Assessment of Activity Budget Studies Employing Instantaneous Sampling

To obtain accurate estimates of activity budget parameters, samples must be unbiased and precise. Many researchers have considered how biased data may affect their ability to draw conclusions and examined ways to decrease bias in sampling efforts, but few have addressed the implications of not considering estimate precision. We propose a method to assess whether the number of instantaneous samples collected is sufficient to obtain precise activity budget parameter estimates. We draw on sampling theory to determine the number of observations per animal required to reach a desired bound on the error of estimation based on a stratified random sample, with individual animals acting as strata. We also discuss the optimal balance between the number of individuals sampled and the number of observations sampled per individual for a variety of sampling conditions. We present an empirical dataset on pronghorn (Antilocapra americana) as an example of the utility of the method. The required numbers of observation to reach precise estimates for pronghorn varied between common and rare behaviors, but precise estimates were achieved with <255 observations per individual for common behaviors. The two most apparent factors affecting the required number of observations for precise estimates were the number of individuals sampled and the complexity of the activity budget. This technique takes into account variation associated with individual activity budgets and population variation in activity budget parameter estimates, and helps to ensure that estimates are precise. The method can also be used for planning future sampling efforts.     

Purging of inbreeding depression and fitness decline in bottlenecked populations of Drosophila melanogaster

Drastic reductions in population size, or bottlenecks, are thought to significantly erode genetic variability and reduce fitness. However, it has been suggested that a population can be purged of the genetic load responsible for reduced fitness when subjected to bottlenecks. To investigate this phenomenon, we put a number of Drosophila melanogaster isofemale lines known to differ in inbreeding depression through four ‘founder‐flush’ bottleneck cycles with flush sizes of 5 or 100 pairs and assayed for relative fitness (single‐pair productivity) after each cycle. Following the founder‐flush phase, the isofemale lines, with a large flush size and a history of inbreeding depression, recovered most of the fitness lost from early inbreeding, consistent with purging. The same isofemale lines, with a small flush size, did not regain fitness, consistent with the greater effect of genetic drift in these isofemale lines. On the other hand, the isofemale lines that did not show initial inbreeding depression declined in fitness after repeated bottlenecks, independent of the flush size. These results suggest that the nature of genetic variation in fitness may greatly influence the way in which populations respond to bottlenecks and that stochastic processes play an important role. Consequently, an attempt intentionally to purge a population of detrimental variation through inbreeding appears to be a risky strategy, particularly in the genetic management of endangered species.     

Cryptic population genetic structure: the number of inferred clusters depends on sample size

Clustering methods have been used extensively to unravel cryptic population genetic structure. We investigated the effect of the number of individuals sampled in each location on the resulting number of clusters. Our study was motivated by recent results in Arabidopsis thaliana: studies in which more than one individual was sampled per location apparently have led to a much higher number of clusters than studies where only one individual was sampled in each location, as is generally done in this species. We show, using computer simulations and microsatellite data in A. thaliana, that the number of sampled individuals indeed has a strong impact on the number of resulting clusters. This effect is smaller if the sampled populations have a hierarchical structure. In most cases, sampling 5–10 individuals per population should be enough. The results argue for abandoning the concept of ‘accessions’ in partially selfing organisms.     

Statistics of natural populations. III. Sequential sampling plans for the estimation of gene frequencies

In previous work several models have been developed for genetic surveys of natural populations. Parents of unknown genotype are collected from a natural population, polymorphic at a single genetic locus. From each of these N cryptic parents a number of offspring are identified for their genotype. Our problem is to select an efficient offspring sampling plan for estimating the frequency of an allele in the cryptic adult population based on the N family profiles of juvenile genotypes. A criterion called the information per unit cost of observation is introduced to evaluate sequential sampling plans, in which the number of offspring per family examined is random. Some simple, practical schemes for stopping the sampling of offspring from a collected parent are introduced; one example is stopping when: (i) the offspring are definitive about the parental genotype(s) for the first time; (ii) a fixed number of one genotype only is seen; or (iii) a fixed maximum feasible number of offspring have been genotyped. This sampling scheme is recommended. For each sampling scheme, the best linear unbiased estimator and the sequential maximum likelihood estimator of the allele frequency are characterized. From the moments of these estimators, it is then possible to tabulate efficient sequential sampling plans, which are better (in the sense of information per unit cost), just as simple, and less costly than corresponding fixed sampling plans in use.     

Sampling strategies for using female gametophytes to estimate heterozygosity in conifers

Summary Unbiased estimators of genotype and allele frequencies and their respective variances are obtained for loci identified by mendelian segregation in haploid female gametophytes from individual trees. By a minimum sampling variance criterion, the allocation of experimental effort between the number of female gametophytes analysed per tree and the number of trees sampled per population is examined for a fixed total amount of experimental effort. For estimating heterozygosity, the optimum sampling design for many (generally most) cases is three female gametophytes per tree, but may be more than three depending upon the true genotype frequencies in the population. For estimating allele frequencies, the optimum sampling design is one female gametophyte per tree except in cases where a strong negative correlation exists between alleles within genotpyes. Guidelines are discussed for determining a suitable number of female gametophytes to be analysed per tree in order to estimate heterozygosity.     

Identifying the source of species invasions: sampling intensity vs. genetic diversity

Population geneticists and community ecologists have long recognized the importance of sampling design for uncovering patterns of diversity within and among populations and in communities. Invasion ecologists increasingly have utilized phylogeographical patterns of mitochondrial or chloroplast DNA sequence variation to link introduced populations with putative source populations. However, many studies have ignored lessons from population genetics and community ecology and are vulnerable to sampling errors owing to insufficient field collections. A review of published invasion studies that utilized mitochondrial or chloroplast DNA markers reveals that insufficient sampling could strongly influence results and interpretations. Sixty per cent of studies sampled an average of less than six individuals per source population, vs. only 45% for introduced populations. Typically, far fewer introduced than source populations were surveyed, although they were sampled more intensively. Simulations based on published data forming a comprehensive mtDNA haplotype data set highlight and quantify the impact of the number of individuals surveyed per source population and number of putative source populations surveyed for accurate assignment of introduced individuals. Errors associated with sampling a low number of individuals are most acute when rare source haplotypes are dominant or fixed in the introduced population. Accuracy of assignment of introduced individuals is also directly related to the number of source populations surveyed and to the degree of genetic differentiation among them ( F _ST). Incorrect interpretations resulting from sampling errors can be avoided if sampling design is considered before field collections are made.     

Frequency of alleles conferring resistance to a Bacillus thuringiensis toxin in a Philippine population of Scirpophaga incertulas (Lepidoptera: Pyralidae)

Using the F2 screen methodology, we estimated the frequency of alleles conferring resistance to the Cry1Ab toxin of Bacillus thuringiensis Berliner in a Philippine population of the stem borer Scirpophaga incertulas (Walker). Evaluation of >450 isofemale lines for survival of F2 larvae on cry1Ab plants did not detect the presence of an allele conferring a high level of resistance. The frequency of such an allele in the sampled population was conservatively estimated to be <3.6 x 10(-3) with 95% confidence and a detection probability of 94%. However, there was evidence of the presence of alleles conferring partial resistance to Cry1Ab. The frequency of alleles for partial resistance was estimated as 4.8 x 10(-3) with a 95% CI between 1.3 x 10(-3) and 1.04 x 10(-2) and a detection probability of 94%. Our results suggest that the frequency of alleles conferring resistance to Cry1Ab in the population of S. incertulas sampled is not too high to preclude successful implementation of the high dose/refuge resistance management strategy.     

Cost‐effective development of highly polymorphic microsatellite in Japanese quail facilitated by next‐generation sequencing

Next‐generation sequencing technologies permit rapid and cost‐effective identification of numerous putative microsatellite loci. Here, from the genome sequences of Japanese quail, we developed microsatellite markers containing dinucleotide repeats and employed these for characterisation of genetic diversity and population structure. A total of 385 individuals from 12 experimental and one wild‐derived Japanese quail lines were genotyped with newly developed autosomal markers. The maximum number of alleles, expected heterozygosity and polymorphic information content (PIC) per locus were 10, 0.80 and 0.77 respectively. Approximately half of the markers were highly informative (PIC ≥ 0.50). The mean number of alleles per locus and observed heterozygosity within a line were in the range of 1.3–4.1 and 0.11–0.53 respectively. Compared with the wild‐derived line, genetic diversity levels were low in the experimental lines. Genetic differentiation (F_ST) between all pairs of the lines ranged from 0.13 to 0.83. Genetic clustering analyses based on multilocus genotypes of individuals showed that most individuals formed clearly defined clusters corresponding to the origins of the lines. These results suggest that Japanese quail experimental lines are highly structured. Microsatellite markers developed in this study may be effective for future genetic studies of Japanese quail.     

Estimating the proportion of identifiable individuals and group sizes in photographic identification studies

Simple Bayesian statistical models are introduced to estimate the proportion of identifiable individuals and group sizes in photographic identification, or photo‐ID, studies of animals that are found in groups. The models require a simple random photographic sampling of animals, where the photographic captures are treated as sampling with replacement within each group. The total number of images, including those that cannot be identified, and the number of images that contain identifiable individuals are used to make inference about the proportion of identifiable individuals within each group and as the population when a number of groups are sampled. The numbers of images for individuals within each group are used to make inference about the group size. Based on analyses of simulated and real data, the models perform well with respect to accuracy and precision of posterior distributions of the parameters. Widths of posterior intervals were affected by the number of groups sampled, sampling duration, and the proportion of identifiable individuals in each group that was sampled. The structure of the models can accommodate covariates, which may affect photographic efficiency, defined in this study as the probability of photographically capturing individuals.     

CHROMOSOMAL POLYMORPHISM IN ISOFEMALE LINES AND CAGE POPULATIONS OF DROSOPHILA MELANOGASTER

Drosophila melanogaster populations in nature usually carry inversion polymorphisms. When they were transferred to and maintained in the laboratory as large cage populations, frequencies of polymorphic inversions were drastically decreased and finally eliminated. This “cage effect” was observed irrespective of the geographical origin of the population or the initial frequency of each inversion. The decrease and elimination of inversions in the cage was not overcome by changing conditions such as medium, temperature, or the number of isofemale lines (40-600) introduced. On the other hand, in the sets of isofemale lines derived from the same geographical origins as the cage populations, each of which was maintained as a small vial population, the inversion frequencies, though decreased from the initial frequencies, were kept at significantly high levels. The cage populations initiated with one or two isofemale lines also maintained the inversion polymorphisms that were as high as vial populations.     

Analysis of population genetic structure with RAPD markers

Recent advances in the application of the polymerase chain reaction make it possible to score individuals at a large number of loci. The RAPD (random amplified polymorphic DNA) method is one such technique that has attracted widespread interest. The analysis of population structure with RAPD data is hampered by the lack of complete genotypic information resulting from dominance, since this enhances the sampling variance associated with single loci as well as induces bias in parameter estimation. We present estimators for several population-genetic parameters (gene and genotype frequencies, within- and between-population heterozygosities, degree of inbreeding and population subdivision, and degree of individual relatedness) along with expressions for their sampling variances. Although completely unbiased estimators do not appear to be possible with RAPDs, several steps are suggested that will insure that the bias in parameter estimates is negligible. To achieve the same degree of statistical power, on the order of 2 to 10 times more individuals need to be sampled per locus when dominant markers are relied upon, as compared to codominant (RFLP, isozyme) markers. Moreover, to avoid bias in parameter estimation, the marker alleles for most of these loci should be in relatively low frequency. Due to the need for pruning loci with low-frequency null alleles, more loci also need to be sampled with RAPDs than with more conventional markers, and some problems of bias cannot be completely eliminated.     

Sample design effects in landscape genetics

An important research gap in landscape genetics is the impact of different field sampling designs on the ability to detect the effects of landscape pattern on gene flow. We evaluated how five different sampling regimes (random, linear, systematic, cluster, and single study site) affected the probability of correctly identifying the generating landscape process of population structure. Sampling regimes were chosen to represent a suite of designs common in field studies. We used genetic data generated from a spatially-explicit, individual-based program and simulated gene flow in a continuous population across a landscape with gradual spatial changes in resistance to movement. Additionally, we evaluated the sampling regimes using realistic and obtainable number of loci (10 and 20), number of alleles per locus (5 and 10), number of individuals sampled (10–300), and generational time after the landscape was introduced (20 and 400). For a simulated continuously distributed species, we found that random, linear, and systematic sampling regimes performed well with high sample sizes (>200), levels of polymorphism (10 alleles per locus), and number of molecular markers (20). The cluster and single study site sampling regimes were not able to correctly identify the generating process under any conditions and thus, are not advisable strategies for scenarios similar to our simulations. Our research emphasizes the importance of sampling data at ecologically appropriate spatial and temporal scales and suggests careful consideration for sampling near landscape components that are likely to most influence the genetic structure of the species. In addition, simulating sampling designs a priori could help guide filed data collection efforts     

遗传多样性的取样策略

合理取样是生物多样性有效保护、利用和研究所面临的最基本问题 ,它在很大程度上受到植物自身的生物学特性、环境条件和取样目的的影响。遗传多样性的取样策略是指对一定地理分布范围内的生物个体取样时 ,使样本具有代表性和包含尽可能多的遗传变异的最佳取样方法 ,包括了取样数目 (一个给定区域的居群数和一个居群的个体数 )以及取样方式。包括“哈迪 温伯格平衡 (Hardy WeinbergEquilibrium)”定律在内的居群遗传学基本原理是研究取样策略的理论基础 ,在此基础上可以对居群内的取样个体数及应获取的居群数进行理论计算 ,同时还可以根据物种居群的遗传结构特点和环境条件的异质性来决定取样的方式。因此 ,应该依据研究对象本身的特点和取样的目的来确定某一特定区域的居群取样数 ,以及某一居群内的样本数及取样方式。     

Intraspecific variation ofDrosophila buzzatii larval breeding success onOpuntia: A yeast-plant-insect relationship

The cactophilic species,Drosophila buzzatii, normally breeds in decaying pockets ofOpuntia cladodes, in which there is a complex interaction with the microbial flora, especially yeast species. Isofemale lines were used to estimate genetic variation among larvae reared on their natural feeding substrate. Four naturally occurring cactophilic yeast species isolated from the same Tunisian oasis as theDrosophila population were used. Two fitness components were studied for each line, viability and developmental time. Genetic variations amongD. buzzatii lines were observed for both traits. A significant yeast species x isofemale line interaction for viability was also evidenced, suggesting the occurrence of specialized genotypes for the utilization of breeding substrates. This genetic heterogeneity in the natural population may favor a better adaptation to the patchily distribution of yeasts.