Parentage versus two-generation analyses for estimating pollen-mediated gene flow in plant populations

Assessment of contemporary pollen-mediated gene flow in plants is important for various aspects of plant population biology, genetic conservation and breeding. Here, through simulations we compare the two alternative approaches for measuring pollen-mediated gene flow: (i) the NEIGHBORHOOD model--a representative of parentage analyses, and (ii) the recently developed TWOGENER analysis of pollen pool structure. We investigate their properties in estimating the effective number of pollen parents (N(ep)) and the mean pollen dispersal distance (delta). We demonstrate that both methods provide very congruent estimates of N(ep) and delta, when the methods' assumptions considering the shape of pollen dispersal curve and the mating system follow those used in data simulations, although the NEIGHBORHOOD model exhibits generally lower variances of the estimates. The violations of the assumptions, especially increased selfing or long-distance pollen dispersal, affect the two methods to a different degree; however, they are still capable to provide comparable estimates of N(ep). The NEIGHBORHOOD model inherently allows to estimate both self-fertilization and outcrossing due to the long-distance pollen dispersal; however, the TWOGENER method is particularly sensitive to inflated selfing levels, which in turn may confound and suppress the effects of distant pollen movement. As a solution we demonstrate that in case of TWOGENER it is possible to extract the fraction of intraclass correlation that results from outcrossing only, which seems to be very relevant for measuring pollen-mediated gene flow. The two approaches differ in estimation precision and experimental efforts but they seem to be complementary depending on the main research focus and type of a population studied.     

Development of a semi-automatic system for pollen recognition

A semi-automatic system for pollen recognitionis studied for the european project ASTHMA. The goal of such a system is to provideaccurate pollen concentration measurements. This information can be used as well by thepalynologists, the clinicians or a forecastsystem to predict pollen dispersion. At first,our emphasis has been put on Cupressaceae, Olea, Poaceae and Urticaceae pollen types. The system is composed of two modules: pollengrain extraction and pollen grain recognition. In the first module, the pollen grains areobserved in light microscopy and are extractedautomatically from a pollen slide coloured withfuchsin and digitized in 3D. In the secondmodule, the pollen grain is analyzed forrecognition. To accomplish the recognition, itis necessary to work on 3D images and to usedetailed palynological knowledge. Thisknowledge describes the pollen types accordingto their main visible characteristerics and tothose which are important for recognition. Somepollen structures are identified like the porewith annulus in Poaceae, the reticulum in Oleaand similar pollen types or the cytoplasm inCupressaceae. The preliminary results show therecognition of some pollen types, likeUrticaceae or Poaceae or some groups of pollentypes, like reticulate group.     

Pollen viability and pollen vigor

Summary Investigations were carried out to correlate pollen viability, assessed on the basis of a fluorochromatic reaction (FCR) test, with pollen vigor, assessed on the basis of the time taken for in vitro germination in pollen grains subjected to high humidity (>95% RH) and temperature (38 °C) or storage stress of Nicotiana tabacum, Agave sp., Tradescantia virginiana, and Iris sp. Both high RH and temperature, as well as storage stresses, affected pollen vigor before affecting pollen viability. The results are discussed in the light of available data on the viability and vigor of stressed pollen and of aged seeds. The need for consideration of pollen vigor, particularly in stored pollen, the inadequacy of the methods presently used, and some of the methods suitable to assess pollen vigor are elaborated.     

An efficient and rapid transgenic pollen screening and detection method using flow cytometry

Assaying for transgenic pollen, a major vector of transgene flow, provides valuable information and essential data for the study of gene flow and assessing the effectiveness of transgene containment. Most studies have employed microscopic screening methods or progeny analyses to estimate the frequency of transgenic pollen. However, these methods are time-consuming and laborious when large numbers of pollen grains must be analyzed to look for rare transgenic pollen grains. Thus, there is an urgent need for the development of a simple, rapid, and high throughput analysis method for transgenic pollen analysis. In this study, our objective was to determine the accuracy of using flow cytometry technology for transgenic pollen quantification in practical application where transgenic pollen is not frequent. A suspension of non-transgenic tobacco pollen was spiked with a known amount of verified transgenic tobacco pollen synthesizing low or high amounts of green fluorescent protein (GFP). The flow cytometric method detected approximately 75% and 100% of pollen grains synthesizing low and high amounts of GFP, respectively. The method is rapid, as it is able to count 5000 pollen grains per minute-long run. Our data indicate that this flow cytometric method is useful to study gene flow and assessment of transgene containment.     

Construction,validation, and application of nocturnal pollen transport networks in an agro‐ecosystem: a comparison using light microscopy and DNA metabarcoding

1. Moths are globally relevant as pollinators but nocturnal pollination remains poorly understood. Plant–pollinator interaction networks are traditionally constructed using either flower‐visitor observations or pollen‐transport detection using microscopy. Recent studies have shown the potential of DNA metabarcoding for detecting and identifying pollen‐transport interactions. However, no study has directly compared the realised observations of pollen‐transport networks between DNA metabarcoding and conventional light microscopy. 2. Using matched samples of nocturnal moths, we constructed pollen‐transport networks using two methods: light microscopy and DNA metabarcoding. Focussing on the feeding mouthparts of moths, we developed and provide reproducible methods for merging DNA metabarcoding and ecological network analysis to better understand species interactions. 3. DNA metabarcoding detected pollen on more individual moths, and detected multiple pollen types on more individuals than microscopy, although the average number of pollen types per individual was unchanged. However, after aggregating individuals of each species, metabarcoding detected more interactions per moth species. Pollen‐transport network metrics differed between methods because of variation in the ability of each to detect multiple pollen types per moth and to separate morphologically similar or related pollen. We detected unexpected but plausible moth–plant interactions with metabarcoding, revealing new detail about nocturnal pollination systems. 4. The nocturnal pollination networks observed using metabarcoding and microscopy were similar yet distinct, with implications for network ecologists. Comparisons between networks constructed using metabarcoding and traditional methods should therefore be treated with caution. Nevertheless, the potential applications of metabarcoding for studying plant–pollinator interaction networks are encouraging, especially when investigating understudied pollinators such as moths.     

Aerodynamics of saccate pollen and its implications for wind pollination

Pollen grains of many wind-pollinated plants contain 1-3 air-filled bladders, or sacci. Sacci are thought to help orient the pollen grain in the pollination droplet. Sacci also increase surface area of the pollen grain, yet add minimal mass, thereby increasing dispersal distance; however, this aerodynamic hypothesis has not been tested in a published study. Using scanning electron and transmission electron microscopy, mathematical modeling, and the saccate pollen of three extant conifers with structurally different pollen grains (Pinus, Falcatifolium, Dacrydium), we developed a computational model to investigate pollen flight. The model calculates terminal settling velocity based on structural characters of the pollen grain, including lengths, widths, and depths of the main body and sacci; angle of saccus rotation; and thicknesses of the saccus wall, endoreticulations, intine, and exine. The settling speeds predicted by the model were empirically validated by stroboscopic photography. This study is the first to quantitatively demonstrate the adaptive significance of sacci for the aerodynamics of wind pollination. Modeling pollen both with and without sacci indicated that sacci can reduce pollen settling speeds, thereby increasing dispersal distance, with the exception of pollen grains having robust endoreticulations and those with thick saccus walls. Furthermore, because the mathematical model is based on structural characters and error propagation methods show that the model yields valid results when sample sizes are small, the flight dynamics of fossil pollen can be investigated. Several fossils were studied, including bisaccate (Pinus, Pteruchus, Caytonanthus), monosaccate (Gothania), and nonsaccate (Monoletes) pollen types.     

Biological-based and remote sensing techniques to link vegetative and reproductive development and assess pollen emission in Mediterranean grasses

Connecting the signals of the vegetative and reproductive cycles of plants using large-scale phenological techniques is not always an easy task, and this complexity increases considerably when analysing the plant life cycle in grasses, due to the ubiquity and diversity of this taxonomic family. This work integrates remote sensing techniques (NDVI from satellite remote sensing data and greenness from near-surface imagery) and biological-based techniques (airborne pollen monitoring and field observations and sampling) to analyse phenological patterns and productivity in grass-dominated vegetation types. We aim to answer two main applied and unanswered questions; i) how are the specific phases of vegetative and reproductive cycles in grasses linked at the species and plant community level? and ii) which grass-dominant habitats are the major contributors of grass pollen emission to the atmosphere at the plant community level? The multi-scale integration and validation of large-scale methods such as satellite remote sensing data and aerobiological monitoring using high-resolution or field phenological techniques is recommended. The results clearly support the hypothesis that the highest rates of grass pollen emission are successively produced when the major grass-dominated vegetation types go through the final phases of vegetative development during their biological senescence or equivalent phases. At the plant community level, natural and semi-natural grass-dominated vegetation types, rather than grass cropland habitats, constitute the major sources of pollen emission. The major contributors to the grass pollen emission at the species level are also identified. Finally, a positive relationship between year-to-year primary productivity (measured as annual sum or maximum NDVI) and pollen production (measured as airborne pollen intensity) was observed at the community level. This is a very timely study, as the availability of remote sensing data is increasing interest in generating enhanced forecasting model of allergenic airborne pollen.     

播种季节和种植方式对多年生苦荞主要农艺性状的影响

该研究选取六个多年生苦荞新品系,对春季、秋季直播与秋季再生其主要农艺性状进行调查。结果表明:(1)不同播种季节对多年生苦荞新品系主花序的花粉可育率、总结实率、有效结实率、植株株高、主茎粗、主茎分枝数、主茎节数、籽粒百粒重、单株粒数、单株产量的影响均达到显著或极显著水平;秋播主花序花粉可育率、总结实率、有效结实率、植株主茎分枝数、籽粒百粒重、单株粒数、单株产量均极显著高于春播;植株株高、主茎粗、主茎节数均极显著低于春播;主花序花朵大小、籽粒种子长宽比无显著差异。(2)不同种植方式对主花序花粉可育率、有效结实率、植株主茎节数及籽粒百粒重的影响达到显著或极显著水平;秋季再生主花序花粉可育率、籽粒单株粒数显著高于秋季直播;主花序有效结实率、植株主茎粗、主茎节数、籽粒百粒重显著低于秋季直播;主花序花朵大小、总结实率、植株株高、主茎分枝数、籽粒种子长宽比、单株产量无显著差异;相关分析表明,各生长季节下主花序有效结实率及单株粒数与单株产量的相关系数均最高。(3)所有参试品系中,1612-241秋季直播的单株产量显著高于其他品系; 1612-16、1612-33秋季再生单株产量较正季优势显著。该研究结果有助于筛选出适宜一季播种两季收获的优良品系,为今后多年生苦荞的选择育种提供线索基础。     

Designing new automatically generated pollen calendars for the public in Switzerland

Pollen calendars are one of the most comprehensible means to inform allergy sufferers or medical professionals about the mean presence of allergenic pollen during the course of the year. They have been produced with a variety of methods and were distributed with great success since the beginning of pollen monitoring. Current technologies, longer data series and changing user demands allow to develop new calculation methods. For designing the new pollen calendars of Switzerland, the following requirements were formulated: The pollen load levels in the calendars should correspond to the levels used in pollen forecasts. A pollen load level in the calendar should show the time window in which it potentially occurs. Further requirements were an automatic generation of the calendars, a regular update and the possibility to provide calendars for single stations, regions or specific pollen species. The data analysis is based on mean daily pollen concentrations of the last 20 years of all 14 pollen-monitoring stations in Switzerland for the 15 pollen types most relevant for allergies. For each day of the year, the 90% quantile of the daily pollen concentrations is determined in a moving 9-day time window over 20 years of data. The calculated concentrations are converted afterward into a pollen load level. The new method is flexible because various parameters can be selected freely: the reference period, the size of the moving time window, the quantile value and the thresholds for pollen load levels. Adjusting these parameters, also pollen calendars for fewer than 20 years can be calculated. However, a sensitivity analysis showed that a reference period of 20 years provides much more stable pollen calendars than shorter reference periods.     

A new procedure to asses pollen viability

We tested pollen viability of eight species using four vital dyes, a new peroxidase test together with three other established methods (MTT, Baker’s and X-Gal), to determine their potential to differentiate fresh pollen from pollen heated for 2 h and 24 h at 80°C (killed pollen) and compared the results with in vitro germination. We found that two of three dyes previously employed to determine viability also stained killed pollen, while the new peroxidase test and MTT did not. We suggest that the latter two are the best methods to test pollen viability, since they do not normally stain either killed or aborted pollen. Received: 7 May 1999 / Revision accepted: 25 October 1999     

GENE EXCHANGE IN LOBLOLLY PINE: THE RELATION BETWEEN POLLINATION MECHANISM,FEMALE RECEPTIVITY AND POLLEN AVAILABILITY

The pollination process in loblolly pine has been examined over several years, both in the field (seed orchards) and experimentally on greenhouse-grown material. Female strobili are receptive to pollination for periods of a wk or more. Initially, background pollen from outside the seed orchard is the main source of pollen but as peak receptivity approaches, pollen from the stand itself predominates especially in older orchards. Consequently, strobili can receive pollen both from outside the orchard as well as from within. The pollen lands on the micropylar horns where it is transferred through the micropyle onto the nucellus by either rainfall or the pollen drop, whichever comes first. Since the pollen drop does not occur until the latter part of the receptive period, rainfall is the most likely transfer agent and pollen flotation is vital if rain occurs. Early arriving pollen does not appear to have an advantage over later arriving pollen for uptake onto the nucellus, even if rain follows the first pollination immediately. Therefore, total pollination of the strobilus can result from both distant and nearby pollen sources.     

A comparison between modern pollen spectra of moss cushions and Cundill pollen traps

Pollen spectra of 23 Cundill pollen traps from 23 different sampling sites in Southwest Turkey are compared with the corresponding pollen spectra of moss cushions from the same sites. The Cundill pollen traps represent the modern pollen rain data from one year whereas the moss cushions represent the pollen rain of several years. The comparative study reveals some main differences between the two pollen entrapment media. The one-year entrapment medium (pollen trap) appears to be more sensitive for local (releve area 10x10 m) and regional (100-500 m, or a few kilometres for Olea europaea) vegetation. Pollen spectra of moss cushions are dominated by high pine pollen percentage values and hardly sense fine vegetation structures. The conclusion of this comparative modern pollen study allows to interpret fossil sediment spectra from the Near East in a more critical way. It is concluded that one should preferably sample rapidly deposited sediments for palynological analyses, as the resulting highresolution pollen diagrams will be most informative about the former vegetation patterns.     

花粉破壁法对花粉蜂蜜酒中氨基酸含量的影响

以蜂蜜为主料,加入花粉,经过调配、发酵、过滤、陈酿后获得氨基酸含量丰富的酒精饮料。花粉破壁分别采用超声破碎法、酶法和温差破壁法这几种不同方式处理,并添加于蜂蜜发酵液中,经酒精发酵,制得富含活性成分的花粉蜂蜜酒。最后利用毛细管电泳法来测定成品酒中的氨基酸,通过分析优化出最佳的花粉破壁方法。研究结果表明花粉经过温差破壁法处理后加入发酵液中,其成品的氨基酸含量最高。该产品不仅营养价值高,风味独特,而且还具有良好的保健作用。     

Forensic Palynology as Classroom Inquiry

This activity introduces the science of forensic palynology: the use of microscopic pollen and spores (also called palynomorphs) to solve criminal cases. Plants produce large amounts of pollen or spores during reproductive cycles. Because of their chemical resistance, small size, and morphology, pollen and spores can be used to link individuals or objects to specific locations where the parent plants grow. Students will use a digital pollen database and Google Earth to link pollen trace evidence to a specific crime scene. The methods presented are based on those used in criminal cases and palynological techniques used by forensic scientists. Step-by-step instructions for a hands-on investigation and a case simulation are presented.     

东北长白山垂直林带下现代表土花粉与植被关系

在长白山北坡5个垂直植被带的林下采集29个表土(苔藓)样品进行孢粉分析，同时将其结果作对应分析。结果表明，除个别样品，在长白山垂直植被带的孢粉组合中乔木花粉占优势，其中以松属和桦木属最多，在各植被带均有分布。松属花粉最高含量出现在以红松为主的针阔混交林带的孢粉组合中；桦木属花粉则大量见于亚高山岳桦林带和以栎桦为主的阔叶落叶林带，但前者伴有耐寒、旱的小灌从杜鹃花属花粉，而在后者中则见有大量蕨类孢子。阔叶落叶林带的孢粉组合是以类型多、百分含量高的落叶乔木花粉和大量蕨类饱子为特征。亚高山针叶林带的孢粉组合中出现数量众多的云杉属花粉。在高山苔原带孢粉组合中虽然乔木花粉略占优势，但极少或不见蕨类孢子，与其他植被带比较，草本植物和小灌丛花粉明鲜较多，其中杜鹃属花粉含量在草本中占优势，其中出现苔原带的指示花粉——仙女木属。     

Quality control in aerobiology: comparison different slide reading methods

The use of standard methodology allows us to compare and to evaluate the data generated from samplers under general use. In aerobiological pollen monitoring, the most frequently slide sampling methods used are based on the selection of different number of longitudinal or transverse traverses and random fields, which represents a small proportion of the entire slide. The aim of this study is to evaluate possible methodological errors produced when the different pollen count methods are used in relation to the results obtained if the whole slide area was quantified. Moreover, the optimization of the counting method by selecting the best longitudinal or transverses traverses that reflect the more accurate counting in relation to the total pollen obtained when the total tape surface was done. Therefore, 113 slides recorded in 2008 at Ourense (NW Spain) differing in its pollen content, recollected time of the year and representation of the different pollen types were selected in this survey. A comparison between the 4 longitudinal traverses, the 12 transverse traverses, and the 493 random fields methods was evaluated. The average relative error and squared error were calculated for both, longitudinal and transverse traverses and the most accurate lines number for counting were selected. Finally, the three counting techniques were compared and significant differences were detected.     

蜘蛛抱蛋亚族的花粉形态

本文研究并报道了百合科蜘蛛抱蛋亚族的花粉形态,主要结论如下：1．开口箭属含两种花粉类型即：(1)单槽、椭圆形;(2)无萌发孔、球形。具单槽、椭圆形花粉的种类含单型性核型,花形较小,柱头小而花柱短,雄蕊高于柱头;无萌发孔、球形花粉的种类则为单型性核型,花形较大,柱头常高度膨大,雄蕊常着生在花被筒基部。2．万年青属花粉为椭圆形,具单槽;蜘蛛抱蛋属花粉则为球形,无萌发孔;开口箭属花粉一部分与万年青属相似,另一部分则与蜘蛛抱蛋属雷同。3．蜘蛛抱蛋亚族中最原始的花粉类型为单槽椭圆形,具孔状或网状外壁纹饰。球形,无萌发孔,外壁具皱波状、瘤状或芽孢状纹饰的花粉为派生类型。4．蜘蛛抱蛋亚族种下花粉形状、萌发孔类型和外壁纹饰基本稳定,可作为分种特征。     

Cautions on direct gene flow estimation in plant populations

Through simulations we have investigated the statistical properties of two of the main approaches for directly estimating pollen gene flow (m) in plant populations: genotypic exclusion and mating models. When the assumptions about accurately known background pollen pool allelic frequencies are met, both methods provide unbiased results with comparable variances across a range of true m values. However, when presumed allelic frequencies differ from actual ones, which is more likely in research practice, both estimators are biased. We demonstrate that the extent and direction of bias largely depend on the difference (measured as genetic distance) between the presumed and actual pollen pools, and on the degree of genetic differentiation between the local population and the actual background pollen sources. However, one feature of the mating model is its ability to estimate pollen gene flow simultaneously with background pollen pool allelic frequencies. We have found that this approach gives nearly unbiased pollen gene flow estimates, and is practical because it eliminates the necessity of providing independent estimates of background pollen pool allelic frequencies. Violations of the mating model assumptions of random mating within local population affect the precision of the estimates only to a limited degree.     

东北长白山垂直林带下现代表土花粉与植被关系

在长白山北坡5个垂直植被带的林下采集29个表土(苔藓)样品进行孢粉分析,同时将其结果作对应分析.结果表明,除个别样品,在长白山垂直植被带的孢粉组合中乔木花粉占优势,其中以松属和桦木属最多,在各植被带均有分布.松属花粉最高含量出现在以红松为主的针阔混交林带的孢粉组合中;桦木属花粉则大量见于亚高山岳桦林带和以栎桦为主的阔叶落叶林带,但前者伴有耐寒、早的小灌丛杜鹃花属花粉,而在后者中则见有大量蕨类孢子.阔叶落叶林带的孢粉组合是以类型多、百分含量高的落叶乔木花粉和大量蕨类孢子为特征.亚高山针叶林带的孢粉组合中出现数量众多的云杉属花粉.在高山苔原带孢粉组合中虽然乔木花粉略占优势,但极少或不见蕨类孢子,与其他植被带比较,草本植物和小灌丛花粉明鲜较多,其中杜鹃属花粉含量在草本中占优势,其中出现苔原带的指示花粉--仙女木属.