Morphological variability of achenes of some European populations of Lactuca serriola L.

This study focuses on the morphological variability of Lactuca serriola achenes in relation to eco-geographic features. Fifty L. serriola populations from four European countries, Czech Republic, Germany, the Netherlands and United Kingdom, were studied. Five morphological characters of the achenes - length and width of achene, length/width index, length of beak, and number of ribs - were evaluated. Significant differences exist in achene morphology of two leaf forms of L. serriola, forma serriola and forma integrifolia. Achenes of f. serriola are shorter, thinner, shorter beaked, lower length/width index, and higher number of ribs compared to f. integrifolia. There was significant variation in the measured characters. Statistical analysis indicated that achene length and width increased along an east-west transect from 2.95 to 3.35 mm and 0.93 to 1.00 mm, respectively. Mean beak length had a similar trend with the exception of German achenes. They had shorter beaks than achenes originating from the Czech Republic with 4.38 and Germany with 4.33 mm. The same trend was evident for L/W index from Czech with a ratio of 3.21 and Germany with 3.14. The number of ribs increased from east to the west in continental Europe, whereas the lowest number of ribs was recorded in achenes collected in Czech with 10.89 and the UK with 10.59. Achene morphology was significantly correlated with three eco-geographic features; longitude, latitude, soil texture of the habitats. The other eco-geographic factors, altitude and population size, did not significantly correlate with the studied characters of L. serriola achenes.     

Microsatellite fingerprinting in lettuce (Lactuca sativa L.) and wild relatives

Southern hybridisation with a single microsatellite probe, (TCT)₁₀, sufficed to discriminate between a representative set of cultivars/accessions of lettuce, Lactuca sativa L., and its wild relatives L. serriola, L. saligna and L. virosa. Variability within cultivars was tested in a relatively modern cultivar (Hector), where no variation was found, and in an older and morphologically more variable cultivar (Madrilene), where heterogeneity was observed in the TCT fingerprint. (TCT)₁₀ fingerprinting should be useful for variety identification and homogeneity testing in lettuce. Received: 25 July 1997 / Revision received: 5 August 1997 / Accepted: 30 August 1997     

A detailed linkage map of lettuce based on SSAP, AFLP and NBS markers

Molecular markers based upon a novel lettuce LTR retrotransposon and the nucleotide binding site-leucine-rich repeat (NBS-LRR) family of disease resistance-associated genes have been combined with AFLP markers to generate a 458 locus genetic linkage map for lettuce. A total of 187 retrotransposon-specific SSAP markers, 29 NBS-LRR markers and 242 AFLP markers were mapped in an F₂ population, derived from an interspecific cross between a Lactuca sativa cultivar commonly used in Europe and a wild Lactuca serriola isolate from Northern Europe. The cross has been designed to aid efforts to assess gene flow from cultivated lettuce into the wild in the perspective of genetic modification biosafety. The markers were mapped in nine major and one minor linkage groups spanning 1,266.1 cM, with an average distance of 2.8 cM between adjacent mapped markers. The markers are well distributed throughout the lettuce genome, with limited clustering of different marker types. Seventy-seven of the AFLP markers have been mapped previously and cross-comparison shows that the map from this study corresponds well with the previous linkage map.     

A Bayesian analysis of gene flow from crops to their wild relatives: cultivated (Lactuca sativa L.) and prickly lettuce (L. serriola L.) and the recent expansion of L. serriola in Europe

Interspecific gene flow can lead to the formation of hybrid populations that have a competitive advantage over the parental populations, even for hybrids from a cross between crops and wild relatives. Wild prickly lettuce (Lactuca serriola) has recently expanded in Europe and hybridization with the related crop species (cultivated lettuce, L. sativa) has been hypothesized as one of the mechanisms behind this expansion. In a basically selfing species, such as lettuce, assessing hybridization in natural populations may not be straightforward. Therefore, we analysed a uniquely large data set of plants genotyped with SSR (simple sequence repeat) markers with two programs for Bayesian population genetic analysis, STRUCTURE and NewHybrids. The data set comprised 7738 plants, including a complete genebank collection, which provided a wide coverage of cultivated germplasm and a fair coverage of wild accessions, and a set of wild populations recently sampled across Europe. STRUCTURE analysis inferred the occurrence of hybrids at a level of 7% across Europe. NewHybrids indicated these hybrids to be advanced selfed generations of a hybridization event or of one backcross after such an event, which is according to expectations for a basically selfing species. These advanced selfed generations could not be detected effectively with crop‐specific alleles. In the northern part of Europe, where the expansion of L. serriola took place, the fewest putative hybrids were found. Therefore, we conclude that other mechanisms than crop/wild gene flow, such as an increase in disturbed habitats and/or climate warming, are more likely explanations for this expansion.     

An Association Between High Peroxidase Activity in Lettuce (Lactuca sativa) and Field Resistance to Downy Mildew (Bremia lactucae)

The association between variation for pre-infection peroxidase activity and levels of field resistance-susceptibility to downy mildew (Bremia lactucae) was investigated in lettuce (Lactuca sativa) cultivars, accessions of L. serriola (prickly lettuce), segregating F₂ populations and selected F₃ families from a cross between field resistant and susceptible lettuce cultivars. A trend was apparent in this series of experiments indicating that one component of field resistance could be related to a high level of peroxidase activity prior to infection. The data suggest that in breeding programmes there could be merit in imposing primary selection for high peroxidase activity prior to field selection for resistance.     

Molecular diversity at the major cluster of disease resistance genes in cultivated and wild Lactuca spp.

Diversity was analyzed in wild and cultivated Lactuca germplasm using molecular markers derived from resistance genes of the NBS-LRR type. Three molecular markers, one microsatellite marker and two SCAR markers that amplified LRR-encoding regions, were developed from sequences of resistance gene homologs at the main resistance gene cluster in lettuce. Variation for these markers were assessed in germplasm including accessions of cultivated lettuce, Lactuca sativa L. and three wild Lactuca spp., L. serriola L., L. saligna and L. virosa L. Diversity was also studied within and between natural populations of L. serriola from Israel and California; the former is close to the center of diversity for Lactuca spp. while the latter is an area of more recent colonization. Large numbers of haplotypes were detected indicating the presence of numerous resistance genes in wild species. The diversity in haplotypes provided evidence for gene duplication and unequal crossing-over during the evolution of this cluster of resistance genes. However, there was no evidence for duplications and deletions within the LRR-encoding regions studied. The three markers were highly correlated with resistance phenotypes in L. sativa. They were able to discriminate between accessions that had previously been shown to be resistant to all known isolates of Bremia lactucae. Therefore, these markers will be highly informative for the establishment of core collections and marker-aided selection. A hierarchical analysis of the population structure of L. serriola showed that countries, as well as locations, were significantly differentiated. These differences may reflect local founder effects and/or divergent selection. Received: 7 March 1999 / Accepted: 25 March 1999     

Host specificity and virulence variation in populations of lettuce powdery mildew pathogen (Golovinomyces cichoracearum s. str.) from prickly lettuce (Lactuca serriola)

Fifty-four isolates of Golovinomyces cichoracearum (GC) sensu stricto were collected during the period 2008–2010 from natural populations of prickly lettuce (Lactuca serriola) growing in the Czech Republic. Population variation in virulence was assessed with a previously developed race differential set, comprising 13 accessions of four Lactuca species (L. serriola, L. sativa, L. saligna, L. virosa, and L. sativa ‘Hilde’ × L. serriola hybrid). GC isolates differed in their host specificity and virulence; most were able to infect most Lactuca spp. accessions (i.e., complete compatibility), but some showed only moderate virulence (i.e., incomplete compatibility). ‘Avirulent’ reactions were the least common, and these were most frequently observed on L. virosa (LVIR/50) and L. saligna (09-H58-1013). Furthermore, a temporal shift in virulence variability was recorded for the Czech GC populations. Generally, virulence of GC to the Lactuca spp. accessions increased during the study period. Across individual years, the most variable were reactions of GC to L. serriola accession (PI 273617), L. sativa cv. ‘Colorado’, L. saligna (09-H58-1013) and L. virosa (LVIR/50). A broad spectrum of virulence (v)-phenotypes was determined among the 54 isolates; however, differences in the reaction patterns of individual isolates were mostly small and related to differentiation between moderately and completely compatible interactions. However, most of the v-phenotypes characterized were unique to a given population in one year. Altogether, 16 v-phenotypes were identified among 16 isolates tested in 2008, 10 v-phenotypes were determined among 10 isolates tested in 2009, and 24 v-phenotypes among 28 isolates tested in 2010; i.e., different v-phenotypes were identified in different localities.     

A note on the measurement of genetic diversity within genebank accessions of lettuce (Lactuca sativa L.) using AFLP markers

This paper discusses a statistical approach for measuring genetic diversity within genebank accessions of a self-fertilising species. This approach is applied to lettuce (Lactuca sativa L.), using AFLP marker data on a set of 1,390 accessions, representing six different lettuce types. Knowledge of the within-accession genetic diversity is important for decisions about the way accessions have to be maintained by genebanks. It is argued that if the within-accession diversity is small, as can be expected for a self-fertilising species like L. sativa, the best approach is to sample as many accessions as possible with only two plants per accession and estimate the within-accession diversity by the proportion of accessions of which the individuals are different.     

Soundness of in situ lipid biomarker analysis: Early effect of heavy metals on leaf fatty acid composition of Lactuca serriola

In previous studies we proposed the plant leaf fatty acid composition, and more precisely the ratio C18:3/(C18:0 + C18:1 + C18:2) as a tool to diagnose soil contamination ex situ. These studies led us to normalize this parameter as a biomarker of soil quality using Lactuca sativa growing under controlled conditions as the plant model species. The evaluation of the toxic effects of pollutants is more difficult to grasp in a real and more complex environment. Hence we developed a study to test the hypothesis that plant leaf fatty acid composition could be used also as a reliable tool to diagnose soil contamination in situ. We tested whether Lactuca serriola (prickly lettuce) could be used as a plant model species for such studies, primarily because it is widely distributed in various European countries and recorded at different habitats, and regarded as an ancestor of the cultivated lettuce L. sativa that was used for ex situ experiments. The results show that the lipid biomarker values were significantly lower in leaves of L. serriola grown on a metallurgic landfill soil than in leaves of plants harvested on surrounding countryside areas. To determine whether the value of the lipid biomarker in L. serriola leaves could be correlated to the soil metal content, we also measured the metal content of the soil corresponding to the rhyzospheric areas from where the prickly lettuce leaves were harvested. The results of the Principal Component Analysis (PCA) show that the lipid biomarker is negatively correlated with both Ni and Cr content. Moreover, these results show that the lipid biomarker values measured in situ were repeatable from year to year, confirm that this biomarker measured in situ would provide an early indication of a plant's exposure to metal, and could be therefore used to facilitate or strengthen the diagnosis of soil contamination.     

Biodiversity of Lactuca aculeata germplasm assessed by SSR and AFLP markers,and resistance variation to Bremia lactucae

In total, seventy two Lactuca aculeata and three Lactuca serriola samples originating from natural populations of these species in Turkey, Jordan, and Israel were analysed by eight microsatellite and 287 amplified fragment length polymorphism (AFLP) markers. Neighbor–Network and Bayesian clustering were used for visualisation of the differences among the analysed L. aculeata and L. serriola samples, and to confirm hybrid origin (L. aculeata × L. serriola) of three samples (343-8A, 343-8B, 54/07) previously indicated by their morphological traits. Molecular data reflect the geographical origin, i.e., the clustering of samples according to their country of origin. Samples from neighbouring parts of Jordan and Israel expressed similar genetic characteristics, indicating the possibility of migration or artificial introduction of plant material. Forty-one L. aculeata samples were screened for their response to five Bremia lactucae races (Bl: 17, Bl: 18, Bl: 24, Bl: 27, and Bl: 28). Susceptible reactions of L. aculeata prevailed. L. aculeata samples were most frequently susceptible to races Bl: 18, Bl: 24, Bl: 27, Bl: 28; and least susceptible to Bl: 17. No highly efficient source of resistance was detected; however, race-specific reaction patterns were frequently recorded, indicating the possible presence of some race-specific resistance factors/genes in the studied samples of L. aculeata. Conservation and exploitation of this material in lettuce breeding is discussed.     

Quantitative trait loci associated with seed and seedling traits in Lactuca

Seed and seedling traits related to germination and stand establishment are important in the production of cultivated lettuce (Lactuca sativa L.). Six seed and seedling traits segregating in a L. sativa cv. Salinas x L. serriola recombinant inbred line population consisting of 103 F8 families revealed a total of 17 significant quantitative trait loci (QTL) resulting from three seed production environments. Significant QTL were identified for germination in darkness, germination at 25 and 35°C, median maximum temperature of germination, hypocotyl length at 72 h post-imbibition, and plant (seedling) quality. Some QTL for germination and early seedling growth characteristics were co-located, suggestive of pleiotropic loci regulating these traits. A single QTL (Htg6.1) described 25 and 23% of the total phenotypic variation for high temperature germination in California- and Netherlands-grown populations, respectively, and was significant between 33 and 37°C. Additionally, Htg6.1 showed significant epistatic interactions with other Htg QTL and a consistent effect across all the three seed production environments. L. serriola alleles increased germination at these QTL. The estimate of narrow-sense heritability (h²) of Htg6.1 was 0.84, indicating potential for L. serriola as a source of germination thermotolerance for lettuce introgression programs.     

LSGermOPA,a custom OPA of 384 EST-derived SNPs for high-throughput lettuce (<Emphasis Type="Italic">Lactuca sativa</Emphasis> L.) germplasm fingerprinting

To deploy a high-throughput genotyping platform in germplasm management, we designed and tested a custom OPA (Oligo Pool All), LSGermOPA, for assessing the genetic diversity and population structure of the USDA cultivated lettuce (Lactuca sativa L.) germplasm collection using Illumina’s GoldenGate assay. This OPA contains 384 EST (expressed sequence tag)-derived SNP (single nucleotide polymorphism) markers selected from a large set of SNP markers experimentally validated and mapped by the Compositae Genome Project. Used for genotyping were DNA samples prepared from bulked leaves of five randomly-selected seedlings from each of 380 lettuce accessions. High-quality genotype data were obtained from 354 of the 384 SNPs. The reproducibility of automatic genotype calls was 99.8% as calculated from the four pairs of duplicated DNA samples in the assay. An unexpectedly high percentage of heterozygous genotypes at the polymorphic loci for most accessions indicated a high level of heterogeneity within accessions. Only 148 homogenous accessions, collectively comprising all five horticultural types, were used in subsequent analyses to demonstrate the usefulness of LSGermOPA. The results of phylogenetic relationship, population structure and genetic differentiation analyses were consistent with previous reports using other marker systems. This suggests that LSGermOPA is capable of revealing sufficient levels of polymorphism among lettuce cultivars and is appropriate for rapid assessment of genetic diversity and population structure in the lettuce germplasm collection. Challenges and strategies for effective genotyping and managing lettuce germplasm are discussed.     

An integrated interspecific AFLP map of lettuce (Lactuca) based on two L. sativa × L. saligna F2 populations

AFLP markers were obtained with 12 EcoRI/ MseI primer combinations on two independent F₂ populations of Lactuca sativa ×Lactuca saligna. The polymorphism rates of the AFLP products between the two different L. saligna lines was 39%, between the two different L. sativa cultivars 13% and between the L. sativa and L. saligna parents on average 81%. In both F₂ populations segregation distortion was found, but only Chromosome 5 showed skewness that was similar for both populations. Two independent genetic maps of the two F₂ populations were constructed that could be integrated due to the high similarity in marker order and map distances of 124 markers common to both populations. The integrated map consisted of 476 AFLP markers and 12 SSRs on nine linkage groups spanning 854 cM. The AFLP markers on the integrated map were randomly distributed with an average spacing between markers of 1.8 cM and a maximal distance of 16 cM. Furthermore, the AFLP markers did not show severe clustering. This AFLP map provides good opportunities for use in QTL mapping and marker-assisted selection. Received: 13 July 2000 / Accepted: 19 January 2001     

The development of lettuce backcross inbred lines (BILs) for exploitation of the<Emphasis Type="Italic"> Lactuca saligna</Emphasis> (wild lettuce) germplasm

Backcross inbred lines (BILs) were developed in which chromosome segments of Lactuca saligna (wild lettuce) were introgressed into L. sativa (lettuce). These lines were developed by four to five backcrosses and one generation of selfing. The first three generations of backcrossing were random. Marker-assisted selection began in the BC₄ generation and continued until the final set of BILs was reached. A set of 28 lines was selected that together contained 96% of the L. saligna genome. Of these lines, 20 had a single homozygous introgression (BILs), four had two homozygous introgressions (doubleBILs) and four lines had a heterozygous single introgression (preBILs). Segregation ratios in backcross generations were compared to distorted segregation ratios in an F₂ population, and the results indicated that most of the distorted segregations can be explained by genetic effects on pollen- or egg-cell fitness. By means of BIL association mapping we were able to map 12 morphological traits and hundreds of additional amplified fragment length polymorphic (AFLP) markers. The total AFLP map now comprises 757 markers. This set of BILs is very useful for future genetic studies.Communicated by F. Salamini     

Towards new sources of resistance to the currant-lettuce aphid (<Emphasis Type="Italic">Nasonovia ribisnigri</Emphasis>)

Domesticated lettuce varieties encompass much morphological variation across a range of crop type groups, with large collections of cultivars and landrace accessions maintained in genebanks. Additional variation not captured during domestication, present in ancestral wild relatives, represents a potentially rich source of alleles that can deliver to sustainable crop production. However, these large collections are difficult and costly to screen for many agronomically important traits. In this paper, we describe the generation of a diversity collection of 96 lettuce and wild species accessions that are amenable to routine phenotypic analysis and their genotypic characterization with a panel of 682 newly developed expressed sequence tag (EST)-linked KASP? single nucleotide polymorphism (SNP) markers that are anchored to the draft Lactuca sativa genome assembly. To exemplify the utility of these resources, we screened the collection for putative sources of resistance to currant-lettuce aphid (Nasonovia ribisnigri) and carried out association analyses to look for potential SNPs linked to resistance.     

Genetic diversity of Dioscorea dumetorum (Kunth) Pax using Amplified Fragment Length Polymorphisms (AFLP) and cpDNA

We have utilized Amplified Fragment Length Polymorphisms (AFLP) in conjunction with chloroplast DNA (cpDNA) sequence data to study the genetic diversity in 53 accessions of Dioscorea dumetorum from six countries in West and Central Africa. Our results provide a comparison of the two marker systems with regards to their applicability to differentiate intraspecific genotypes and the grouping of the accessions based on localities of collection. A total of 1052 AFLP fragments (of which 94.1% were polymorphic) produced from twelve primer combinations indicate a relatively high level of polymorphism among the accessions. Three major genetic groups that do not strictly follow a geographic distribution pattern were identified using Neighbour-joining and the principal coordinate (PCo) analyses. Accessions from Togo showed higher numbers of private fragments and the highest percentage polymorphism (59.4%). The detection of highest genetic diversity in accessions from Nigeria and Togo and their relationship to other accessions suggest that these countries are the centre of origin and diversity of D. dumetorum. The moderately high genetic diversity (average of 61%) is suggesting great influence on the D. dumetorum germplasms through exchange and transfer of cultivars among local farmers in the sub-region. In contrast, DNA sequence data from the psbA-trnH and the rpoB-trnC chloroplast regions revealed no variation among accessions from the different localities and clearly differentiated by AFLP patterns. The results demonstrate the usefulness of the AFLP marker in generating high polymorphism in the D. dumetorum accessions from West and Central Africa and hence may be used for agronomic purposes.     

Shoot and root physiological responses to localized zones of soil moisture in cultivated and wild lettuce (Lactuca spp.)

Cultivated crisphead lettuce (Lactuca sativa L.) has a shallower root system than its wild relative, Lactuca serriola L. The effects of localized soil water, at depth, on plant water relations, gas exchange and root distribution were examined in the two species using soil columns with the soil hydraulic-ally separated into two layers, at (0–20 cm and 20–81) cm, but permitting root growth between the layers. Three treatments were imposed on 7-week-old plants, and maintained for 4 weeks: (i) watering, both layers to field capacity; (ii) drying the upper layer while watering the lower layer to field capacity, and (iii) drying both layers. Drying only 0–20 cm of soil had no effect on leaf water status, net photosynthesis, stomatal conductance or biomass production in L. serriola compared to a well-watered control, but caused a short-term reduction (10 d) in leaf water status and photosynthesis in L. sativa that reduced final shoot production. The different responses may be explained by differences in root distribution. Just before the treatments commenced, L. serriola had 50% of total root length at 20–80 cm compared to 35% in L. sativa. Allocation of total biomass to roots in L. serriola was approximately double that in L. sativa. The wild species could provide germplasm for cultivated lettuces to extract more soil water from depth, which may improve irrigation efficiency.     

Crossing experiments of lettuce cultivars and species (Lactuca sect.Lactuca,Compositae)

The degree of relationships withinLactuca sativa and three wild relativesL. serriola, L. saligna, andL. virosa was studied by observing the performance, vigour and fertility of the F 1 hybrids obtained from crosses made in and between the four species. The crosses ofL. saligna ×L. virosa and the reciprocal crosses produced no hybrids.L. saligna andL. virosa are the least related of the four species.L. sativa ×L. serriola and the reciprocal crosses were successful and produced fertile hybrids These two species are genetically very closely related.L. saligna is known to produce, as a female parent, hybrids withL. sativa andL. serriola. Now the reciprocal cross was successful for the first time, so the unability to obtain hybrids in the past was based on the choice of accessions and not caused by unilateral incompatibility.L. virosa ×L. sativa and the reciprocal combination produced hybrids. The combinationL. serriola ×L. virosa produced hybrids with very limited fertility. In contrast to earlier reports (sterile hybrids) one combination of the reciprocal cross too produced hybrids with very limited fertility.—Some of theL. saligna ×L. sativa (and reciprocal) hybrids were found to look strikingly likeL. serriola. This adds evidence for the descent ofL. serriola andL. sativa:L. saligna also made part of the ancestral complex of the cultivated lettuce.     

Effect of plant–soil feedbacks on the growth and competition of <Emphasis Type="Italic">Lactuca</Emphasis> species

Plant species generate specific soil communities that feedback on plant growth and competition. These feedbacks have been implicated in plant community composition and dispersion. We used Lactuca sativa and its wild progenitor Lactuca serriola to test the hypotheses that separate Lactuca species generate unique soil communities and that these soil communities differentially influence host, and neighboring, plant growth and competition. We grew each Lactuca in competition with the other, in sterile and non-sterile soils. We then examined the growth of each Lactuca species in sterile, non-sterile, and preconditioned soil. Finally, we used TRFLP techniques to explore whether the two Lactuca species generate significantly different bacterial communities in their rhizosphere soils. L. sativa proved to be the stronger competitor of the two species. However, sterilization increased the competitive effect of L. serriola background competitors. The growth experiment showed a significant effect on plant species, soil treatment, and the interaction of the two. Preconditioning soil caused reduced growth in both Lactuca species. Only L. serriola showed significantly increased growth in sterile soils. Our TRFLP analysis showed that the L. sativa soil community was significantly less diverse and that soil preconditioning had the largest impact on the community composition. These results show that Lactuca serriola’s rhizosphere communities generate a stronger negative feedback for plant growth than do the communities associated with L. sativa. Our study suggests that selection for plants that are able to grow in dense monoculture may have released Lactuca from species-specific negative soil feedbacks. This has important implications for both agriculture and the evolution of invasive plant species.