Central European Populations Research Articles

Sequential polyandry in female Tengmalm's owl (Aegolius funereus) during a poor rodent year

In birds of prey, food availability affects the entire breeding process, including reproductive success. Sequential polyandry, sometimes documented in raptors and owls, has been observed occasionally in Tengmalm's owl in years of high food availability. In this study, the effect of food supply on the reproductive strategy of the central European Tengmalm's owl population has been investigated. The availability of the main prey of owls was significantly below average in the study year 2014. This resulted in low breeding density of owls, delayed egg laying, small clutch sizes, and low reproductive success. Nevertheless, successful sequential polyandry of one Tengmalm's owl female was recorded during the breeding season. The polyandrous female laid four eggs in each nests, and 50 % and 75 % of four hatched nestlings left the nest during the first and second breeding, respectively. In both nesting attempts, the two-year old female was of a substandard body mass and she abandoned the fledglings before they left the nests. Prey delivered by males to both nests was comprised mainly of alternative prey (birds and shrews). The results of this study suggest that there are probably some other factors, in addition to food availability, that may play a role in Tengmalm's owl's decisions in matters of parental care.

Comparison of Genetic Structure of Epixylic Liverwort Crossocalyx hellerianus between Central European and Fennoscandian Populations.

Patterns of genetic variation and spatial genetic structure (SGS) were investigated in Crossocalyx hellerianus, a strictly epixylic dioicous liverwort (Scapaniaceae s.l., Marchantiophyta). Studied populations were located in Fennoscandia and Central Europe, with localities differing in availability of substrate and the population connectivity, and their populations consequently different in size, density, and prevailing reproductive mode. A set of nine polymorphic microsatellites was successfully developed and used. Identical individuals were only found within populations. Especially in large populations, the majority of the individuals were genetically unique. Resampled number of genotypes, mean number of observed alleles per locus after rarefaction, and Nei’s gene diversity in large populations reached high values and ranged between 4.41–4.97, 3.13–4.45, and 0.94–0.99, respectively. On the contrary, the values in small populations were lower and ranged between 1.00–4.42, 1.00–2.73, and 0.00–0.95, respectively. As expected, large populations were found to be more genetically diverse than small populations but relatively big diversity of genotypes was also found in small populations. This indicated that even small populations are important sources of genetic variation in bryophytes and processes causing loss of genetic variation might be compensated by other sources of variability, of which somatic mutations might play an important role. The presence of SGS was discovered in all populations. Large populations possessed less SGS, with individuals showing a pronounced decrease in kinship over 50 cm of distance. Apparent SGS of small populations even at distances up to 16 meters suggests the aggregation of similar genotypes, caused predominantly by the deposition of asexually formed gemmae. Although no strong kinship was detectable at the distances over 16 meters in both small and large populations, identical genotypes were occasionally detected at longer distances (20–80 m), suggesting effective dispersal of asexual propagules.

Inferring the genetic history of lactase persistence along the Italian peninsula from a large genomic interval surrounding the LCT gene.

Although genetic variants related to lactase persistence in European populations were supposed to have firstly undergone positive selection in farmers from the Balkans and Central Europe, demographic and evolutionary dynamics that subsequently shaped the distribution of this adaptive trait across the continent have still to be elucidated. To deepen the knowledge about potential routes of diffusion of lactase persistence to Western Europe we investigated variation at a large genomic region surrounding the LCT gene along the Italian peninsula, a geographical area that played a key role in population movements responsible for Neolithic diffusion across Europe. By genotyping 40 highly selected SNPs in more than 400 Italian individuals we described gradients of nucleotide and haplotype variation potentially related to lactase persistence and compared them with those observed in several European and Mediterranean human groups. Multiple migratory events responsible for earlier introduction of the examined alleles in Italy than in Northern European regions could be invoked. Different demic processes occurred along the western and eastern sides of the peninsula were also inferred via linkage disequilibrium and population structure analyses. The appreciable genetic continuum observed between people from Northern or Central-Western Italy and Central European populations suggested a local arrival of lactase persistence-related variants mainly via overland routes. On the contrary, diversity of Central-Eastern and Southern Italian groups entailed also gene flow from South-Eastern Mediterranean regions, in accordance to the earlier entrance of the Neolithic in Southern Italy via maritime population movements along the Mediterranean coastlines.

C9ORF72 hexanucleotide repeat expansion in ALS patients from the Central European Russia population

Cohorts of amyotrophic lateral sclerosis (ALS) patients and control individuals of Caucasian origin from the Central European Russia (Moscow city and region) were analyzed for the presence of hexanucleotide repeat GGGGCC expansion within the first intron of the C9ORF72 gene. The presence of a large (>40) repeat expansion was found in 15% of familial ALS cases (3 of 20 unrelated familial cases) and 2.5% of sporadic ALS cases (6 of 238) but in none of control cases. These results suggest that the frequency of C9ORF72 hexanucleotide repeats expansions in the Central European Russian ALS patients is significantly lower than in Western European or Northern American ALS patients of Caucasian origin but higher than in Asian ALS patients.

Within-range translocations and their consequences in European larch.

In contrast to biological invasions, translocations of individuals within a species range are understudied, due to difficulties in systematically detecting them. This results in limited knowledge about the corresponding processes and uncertainties regarding the status of extant populations. European larch, a forest tree whose fragmented native distribution is restricted to the Alps and to other Central European mountains, has been massively planted for at least 300 years. Here we focus on the genetic characterization of translocations having taken place within its native range. Microsatellite variation at 13 nuclear loci and sequence data of two mitochondrial DNA fragments were analyzed on the basis of a comprehensive range-wide population sample. Two complementary methods (Geneclass and Structure) were used to infer translocation events based on nuclear data whereas mitochondrial data were used for validation of these inferences. Using Geneclass, we found translocation events in a majority of populations. Additional cases of translocation and many instances of admixture were identified using Structure, thanks to the clear-cut ancestral genetic structure detected in this species. In particular, a strong divide between Alpine and Central European populations, also apparent at mitochondrial markers, helped uncover details on translocation events and related processes. Translocations and associated admixture events were found to be heterogeneously distributed across the species range, with a particularly high frequency in Central Europe. Furthermore, translocations frequently involved multiple geographic sources, some of which were over-represented. Our study illustrates the importance of range-wide investigations for tracing translocations back to their origins and for revealing some of their consequences. It provides some first clues for developing suitable conservation and management strategies.

Rapid decline of an isolated population of the black grouse Tetrao tetrix: the crisis at the southern limit of the range

Over the last century, the numbers and range of black grouse Tetrao tetrix have declined dramatically over most of Europe. The Central-European population is at present divided into small, isolated subpopulations, and Orawa (Carpathians, Poland-Slovakia) was considered until recently to be the most abundant one in the region. In 2002, it was estimated at 146–152 males. Surveys conducted in 2011–2014 showed that only 30–40 males were still present in this region. The total disappearance of leks (13 of 24 known leks have been abandoned) and the decline in overall numbers of males at existing leks (some leks were used by single males only) were recorded. In consequences, the number of lekking males decline by 76 % between 2002 and 2014. The nearest neighbour distance between active leks did not change, but the contraction of the total area used by the population (from 187.3 km2 in 2002 to 71.5 km2 in 2014) was recorded. The disappearance of leks was related to the size of the nearest peat bog; leks located close to small peat bogs were abandoned, while those located close to the larger ones remained active. Along with the decline in the numbers of lekking birds and the shrinkage of the range, a decrease in the size of wintering flocks was noted. This rapid fall over a short period of time in the number of black grouse and in the area used by the region’s largest population is indicative of a real threat of extinction of isolated populations. Rapid extinction risk should be taken into consideration when planning conservation measures applicable to its entire range of distribution.

Development of 18 polymorphic microsatellite markers for Vinca minor (Apocynaceae) via 454 pyrosequencing.

Premise of the study:Polymorphic microsatellite markers were developed in Vinca minor (Apocynaceae) to evaluate the level of clonality, population structure, and genetic diversity of the species within its native and introduced range.Methods and Results:A total of 1371 microsatellites were found in 43,565 reads from 454 pyrosequencing of genomic V. minor DNA. Additional microsatellite loci were mined from publicly available cDNA sequences. After several rounds of screening, 18 primer pairs flanking di-, tri-, or tetranucleotide repeats were identified that revealed high levels of genetic diversity in two native Italian populations, with two to 11 alleles per locus. Clonal growth predominated in two populations from the introduced range in Germany. Five loci successfully cross-amplified in three additional Vinca species.Conclusions:The novel polymorphic microsatellite markers are promising tools for studying clonality and population genetics of V. minor and for assessing the historical origin of Central European populations.

Tracing the genetic origin of Europe's first farmers reveals insights into their social organization.

Farming was established in Central Europe by the Linearbandkeramik culture (LBK), a well-investigated archaeological horizon, which emerged in the Carpathian Basin, in today's Hungary. However, the genetic background of the LBK genesis is yet unclear. Here we present 9 Y chromosomal and 84 mitochondrial DNA profiles from Mesolithic, Neolithic Starčevo and LBK sites (seventh/sixth millennia BC) from the Carpathian Basin and southeastern Europe. We detect genetic continuity of both maternal and paternal elements during the initial spread of agriculture, and confirm the substantial genetic impact of early southeastern European and Carpathian Basin farming cultures on Central European populations of the sixth-fourth millennia BC. Comprehensive Y chromosomal and mitochondrial DNA population genetic analyses demonstrate a clear affinity of the early farmers to the modern Near East and Caucasus, tracing the expansion from that region through southeastern Europe and the Carpathian Basin into Central Europe. However, our results also reveal contrasting patterns for male and female genetic diversity in the European Neolithic, suggesting a system of patrilineal descent and patrilocal residential rules among the early farmers.

Genotypic diversity of apomictic microspecies of the Taraxacum scanicum group (Taraxacum sect. Erythrosperma)

Populations of polyploid apomictic dandelions consist of a mixture of clonal genotypes that are nearly phenotypically and genetically uniform. Some apomictic taxa are widespread, but many other taxa occur locally or at a single locality. Additionally, certain Central European dandelion populations consist of both diploid sexuals and polyploid apomicts, and the gene flow expected in mixed populations obscures clear genetic and morphological borders among apomictic taxa. In the present study, we investigated genotypic diversity among seven apomictic microspecies coexisting in Central Europe. Using microsatellites, amplified fragment length polymorphism and chloroplast DNA markers, we focused on the clonal structure of apomictic taxa, i.e., whether the studied apomictic taxa are represented by one or a few related genotypes or whether they are represented by many different and unrelated genotypes. The pattern of genotypic diversity within the studied microspecies suggests both intra- and inter-specific genetic diversification. At the intra-specific level, the studied apomictic taxa consist of several related genotypes, and difference among these genotypes is the result of somatic mutations and/or the consequence of repeated origin from different lineages or recent hybridisation. The inter-specific genetic diversity is greater and may reflect a different evolutionary origin of apomictic taxa. Our results show that despite their increased genetic diversity, apomictic dandelions form definable clusters that may be characterised by both molecular markers and morphology.

Characteristics of Morphological Parameters of Donkeys in the Czech Republic

The donkey population in Central Europe and Czech Republic is very variation, which is a consequence of the donkeys originating from various areas of Europe. This has been proved to affect their body conformation. In this work, we are focusing on the population of donkeys in the Czech Republic and its analysis.The main aim of our work was to determine the values morphological parameters in the donkey population in the Czech Republic. Altogether, 23 body dimensions were taken and further processed to calculate 6 hippo metric indexes. The sample group we analyzed consisted by a total of 70 individuals, out of there were 23 stallions and 47 mares. The results were then processed using the methods of general linear model (GLM) and multiple comparisons.We managed to prove a statistically significant influence of the sex factor for the following body measurements: shin length of the front limbs, chest width, withers height measured by tape, shin circumference on both front and pelvic limbs and also for weight index. Also, we have found a statistically significant difference in the scapula length when considering the age factor.

Ancient DNA in the Cantabrian fringe populations: A mtDNA study from Prehistory to Late Antiquity

The present mtDNA study on human remains of fourteen archaeological sites from Cantabria, Basque Country and Navarra provided a diachronic overview from Paleolithic–Mesolithic to Late Antiquity period of some communities settled in the Cantabrian fringe. Ancient DNA studies in European human remains indicated a genetic discontinuity between the hunter–gatherers and later populations. However, some of the mtDNA lineages found in the Cantabrian fringe in Paleolithic–Mesolithic times persist in present-day populations.The mtDNA variability observed in hunter–gatherers and farmers in Europe denoted a complex pattern for the Neolithic transition, occurring along several different routes into and across Europe. The mtDNA lineages found in the Cantabrian fringe indicated that the dispersion of Neolithic farmers had a different genetic impact in this area with respect to Central and Mediterranean regions of Europe. The differences in mtDNA variability were also apparent after the Neolithic, as shown by the genetic distance between the Chalcolithic populations from the Cantabrian fringe and the Bell Beaker Culture (BBC) populations of Central Europe.

Differential diagnosis of LTP‐sensitised patients in Central Europe

Lipid transfer protein (LTP) is a major allergen in peach often causing anaphylactic reactions with a high prevalence of sensitization in the Mediterranean area. The prevalence of LTP sensitization in Central Europe is currently unknown, where the diagnosis is complicated by a high background sensitization to the major birch pollen allergen, Bet v 1, and Bet v 1 cross-reactive food allergens, which cause only mild reactions. LTP and Bet v 1-related proteins are both distributed throughout the plant-kingdom. In this study, we aimed to characterize the sensitization-patterns of patients with severe reactions to plant-derived food in Austria. Thirteen patients reporting strong reactions after consumption of fruits, vegetables or herbs were invited to skin prick and prick to prick tests. For visualizing patients' IgE-reactivity patterns, immunoblots were performed, which allowed the determination of specific IgE to raw and boiled parsley, raspberry, apricot, and peach. These immunoblots showed that cooking did not influence IgE binding to LTPs, while cooking reduced IgE binding to all Bet v 1-related proteins. We further observed that LTP-sensitized patients showed variable IgE reactivity to different LTPs, however, the strongest reactions were seen to fruits from the Rosaceae family, especially to peach, suggesting that peach LTP (Pru p 3) contains the majority of IgE epitopes also for a Central European population. Analysis of IgE reactivity to single molecules (including Bet v 1 and Pru p 3) using an allergen micro-array (ISAC) and single ImmunoCAP measurements enabled to determine the following sensitization patterns: 1 patient was monosensitized to Bet v 1, 3 were monosensitized to LTP, whereas 8 showed a co-sensitization to Bet v 1 and LTP and 1 patient was monosensitized to profilin. Based on this study, we conclude that analysis of specific IgE to the LTP Pru p 3 from peach is also sufficient for diagnosis of reactions to Rosacea fruits even in regions where frequent co-sensitization to Bet v 1 makes the interpretation of extracts-based skin prick tests difficult. In addition, also other plant-related panallergens, such as profilin, should be included in such a differential diagnostic workup scheme.

Genetic variants in interleukin 7 receptor α chain (IL-7Ra) are associated with multiple sclerosis risk and disability progression in Central European Slovak population.

In this study, we determined the association between rs6897932 in interleukin 7 receptor α (IL7Ra) gene and the risk and disability progression of multiple sclerosis (MS) in 270 MS patients and 303 controls. We found allele C to be associated with the risk of MS and minor allele T to be protective against MS development. Moreover, we revealed for the first time that rs6897932 in IL7Ra gene is associated with the progression of MS, evaluated by MSSS scores. The minor allele T and genotype TT are protective against a rapid disability progression in MS in the Central European Slovak population.

Antioxidant vitamin intake and mortality in three Central and Eastern European urban populations: the HAPIEE study.

PurposeThe aim of the study was to assess the relationships between individual-level dietary intakes of antioxidant vitamins C, E and beta-carotene with all-cause and cause-specific mortality in three Central and Eastern European (CEE) populations.MethodsData from the Health, Alcohol and Psychosocial factors in Eastern Europe cohort study were used. At the baseline survey, between 2002 and 2005, 28,945 men and women aged 45–69 years were examined in Novosibirsk (Russia), Krakow (Poland) and seven Czech towns. Deaths in the cohorts were identified through mortality registers. Cox regression was used to estimate the association between vitamin consumption and all-cause, cardiovascular (CVD) disease and cancer mortality.ResultsIn multivariable-adjusted analyses, there were no clear inverse associations between antioxidant vitamin intakes and mortality, although in some groups, several hazard ratios (HRs) were significant. For example, in men, compared with the lowest quintile of vitamin C intake, all-cause mortality in the third and fourth quintiles was lower by 28 % (HR 0.72; 95 % CI 0.61–0.85) and by 20 % (HR 0.80; 95 % CI 0.68–0.95), respectively. CVD mortality was lower by 35 % (HR 0.65; 95 % CI 0.50–0.84) and by 23 % (HR 0.77; 95 % CI 0.59–0.99) in third and fourth quintile of vitamin C intake, respectively. In women, the third and fourth quintiles of dietary intake of vitamin E were associated with reduced risk of all-cause death by 33 % (HR 0.67; 95 % CI 0.53–0.84) and by 23 % (HR 0.77; 95 % CI 0.61–0.97), respectively. Consumption of vitamin C, vitamin E and beta-carotene was not related to CVD mortality in women and to cancer mortality in either gender.ConclusionThis large prospective cohort study in CEE populations with low prevalence of vitamin supplementation did not find a strong, dose–response evidence for protective effects of antioxidant vitamin intake.Electronic supplementary materialThe online version of this article (doi:10.1007/s00394-015-0871-8) contains supplementary material, which is available to authorized users.

Declining tibial curvature parallels ∼6150 years of decreasing mobility in Central European agriculturalists.

Long bones respond to mechanical loading through functional adaptation in a suite of morphological characteristics that together ensure structural competence to in vivo loading. As such, adult bone structure is often used to make inferences about past behavior from archaeological remains. However, such biomechanical approaches often investigate change in just one aspect of morphology, typically cross-sectional morphology or trabecular structure. The relationship between longitudinal bone curvature and mobility patterns is less well understood, particularly in the tibia, and it is unknown how tibial curvature and diaphyseal cross-sectional geometry interact to meet the structural requirements of loading. This study examines tibial curvature and its relationship with diaphyseal cross-sectional geometry (CSG) and body size in preindustrial Central Europeans spanning ∼6150 years following the introduction of agriculture in the region. Anteroposterior centroid displacement from the proximo-distal longitudinal axis was quantified at nine diaphyseal section locations (collectively representative of diaphyseal curvature) in 216 tibial three-dimensional laser scans. Results documented significant and corresponding temporal declines in midshaft centroid displacement and CSG properties. Significant correlations were found between mid-diaphyseal centroid displacement and all mobility-related CSG properties, while the relationship weakened toward the diaphyseal ends. No significant relationship was found between centroid displacement and body size variables with the exception of the most distal section location. Results support a relationship between tibial curvature and cross-sectional geometry among prehistoric Central European agricultural populations, and suggest that changes in mechanical loading may have influenced a suite of morphological features related to bone adaptation in the lower limb.

Adaptation of flowering phenology and fitness-related traits across environmental gradients in the widespread Campanula rotundifolia

Plant populations need to adjust to climate warming through phenotypic plasticity or evolution of trait means. We performed a common-garden experiment with European populations of Campanula rotundifolia to investigate current adaptation in fitness-related traits and the potential for future adaptation. The common garden was situated in Switzerland and contained plants from 18 populations from four regions: Central Europe, The Netherlands, Scandinavia and the Swiss Alps. We assessed current adaptation with trait–environment correlations, and we compared molecular marker with trait differentiation to investigate past selection. How traits may change under future climate warming was investigated via selection analysis. Trait–trait correlations were performed to reveal genetic constraints. The majority of analysed phenotypic traits showed regional differentiation and all traits showed indications of past selection. Flowering duration decreased with latitude and elevation, suggesting adaptation to growing season length. The Central European populations performed best, indicating home-site advantage. Selection analysis showed positive selection on fitness-related traits whereas phenological traits showed less clear patterns. Trait–trait correlations were mostly neutral or favourable to selection. This study suggests that flowering phenology and other fitness-related traits of C. rotundifolia are adapted to the current climatic conditions and have the potential to evolve under climate change.

Estrogen receptor 2 polymorphism in idiopathic scoliosis

Objectives In estrogen receptor 2 (ESR2) gene the rs1256120 polymorphism was described to be associated with predisposition to and severity of idiopathic scoliosis (IS) in Chinese population. This observation has not been confirmed in Japanese population. The ESR2 AluI and RasI site polymorphisms were described to present association with breast cancer, rheumatoid arthritis and bone mineral density, however the association with IS has not been evaluated. The purpose of the study was to investigate associations of the ESR2 polymorphisms with either predisposition to or progression of IS in Central European population.

Ancient mitochondrial DNA from the northern fringe of the Neolithic farming expansion in Europe sheds light on the dispersion process.

The European Neolithization process started around 12 000 years ago in the Near East. The introduction of agriculture spread north and west throughout Europe and a key question has been if this was brought about by migrating individuals, by an exchange of ideas or a by a mixture of these. The earliest farming evidence in Scandinavia is found within the Funnel Beaker Culture complex (Trichterbecherkultur, TRB) which represents the northernmost extension of Neolithic farmers in Europe. The TRB coexisted for almost a millennium with hunter-gatherers of the Pitted Ware Cultural complex (PWC). If migration was a substantial part of the Neolithization, even the northerly TRB community would display a closer genetic affinity to other farmer populations than to hunter-gatherer populations. We deep-sequenced the mitochondrial hypervariable region 1 from seven farmers (six TRB and one Battle Axe complex, BAC) and 13 hunter-gatherers (PWC) and authenticated the sequences using postmortem DNA damage patterns. A comparison with 124 previously published sequences from prehistoric Europe shows that the TRB individuals share a close affinity to Central European farmer populations, and that they are distinct from hunter-gatherer groups, including the geographically close and partially contemporary PWC that show a close affinity to the European Mesolithic hunter-gatherers.

Omega‐3 fatty acid supplementation candidates can be selected using fatty acid profiling

The aim of this study was to evaluate the fatty acid (FA) profile in the plasma and erythrocyte membranes of apparently healthy men compared to male patients with various cardiovascular diseases (CVD). The plasma FA profiles were analyzed in five groups of subjects aged 37–57 years: groups 1 and 2 were male voluntary blood donors with a body mass index (BMI) 25.1 ± 1.6, (Group 1, N = 12) and a BMI 32.9 ± 4.5, (Group 2, N = 11) respectively, male patients with chronic heart failure due to dilated cardiomyopathy (Group 3, N = 10), chronic atrial fibrillation without heart failure or coronary heart disease (Group 4, N = 10) and patients after myocardial infarction (Group 5, N = 11). The FA profile in plasma and erythrocytes was measured by gas chromatography (GC) with flame ionization detection and basic plasma lipid parameters were then estimated. Deficiencies of eicosapentaenoic acid and docosahexaenoic acid were found in all the groups of CVD patients and in Group 2 (BMI 32.9 ± 4.5) of this Central European population. Unfavorable alterations in FA composition and clinical biochemistry parameters were found in patients with dilated cardiomyopathy (Group 3). Further, 75% of Group 1 (blood donors) had an average plasma PUFA level in the worst quartile for relative risk of cardiovascular events. Tailored n‐3 PUFA supplementation should be recommended not only for secondary prevention in patients suffering from CVD but also for primary prevention in overweight and obese persons with a proven deficiency.Practical applications: Plasma and erythrocyte membrane n‐3 polyunsaturated fatty acid (PUFA) levels are an important parameter of diet composition as well as a marker of cardiovascular risk. Regular sea fish intake or alternative supplementation with the n‐3 PUFA recommendation is in all probability insufficiently realized, particularly in inland countries. In addition to contemporary guidelines and practice, research is providing data on the use of FA analysis for revealing people with an n‐3 PUFA deficiency. This approach could benefit people suffering from CVD as well as for overweight and obese populations with demonstrated deficiency.Tailored n‐3 PUFA supplementation should be recommended not only for secondary prevention in patients suffering from CVD but also for primary prevention in overweight and obese persons with a proven deficiency.

The occurrence of lines tolerant to the causal agent of bleeding canker, Pseudomonas syringae pv. aesculi, in a natural horse chestnut population in Central Europe

Pseudomonas syringae pv. aesculi (Pae) causes bleeding canker, a destructive disease of horse chestnut trees (Aesculus hippocastanum) that limits the breeding of horse chestnut genotypes. The susceptibility of 322 F1 progeny from 20 horse chestnut parent tree sources to the pathogen Pae was evaluated using different inoculation methods. The resulting infection rates, in decreasing order, were caused by inoculation of apical buds, inoculation of annual stems, insertion of bacteria into cut wounds on trunks and foliar spraying of the pathogen. According to the resistance index (RI) results of the field pathogenicity tests, at 2 or 3 years after inoculation 40, 40 and 20 % of horse chestnut parent tree sources were resistant, tolerant and susceptible, respectively, to Pae. Microbiological analyses of plant tissues from buds, phloem in the main trunks and roots of infected F1 progeny regardless of parent tree source showed long-term survival of Pae bacteria in 70 % of tolerant F1 progeny plants that lacked bleeding canker symptoms. Initial RI results and microbial analyses indicated that there was some frequency of resistance in the natural horse chestnut population. Altogether, 41 F1 progeny were the most promising lines based on their viability and the absence of Pae in their tissues.