Divergent selection along climatic gradients in a rare central European endemic species,Saxifraga sponhemica

Background and Aims The effects of habitat fragmentation on quantitative genetic variation in plant populations are still poorly known. Saxifraga sponhemica is a rare endemic of Central Europe with a disjunct distribution, and a stable and specialized habitat of treeless screes and cliffs. This study therefore used S. sponhemica as a model species to compare quantitative and molecular variation in order to explore (1) the relative importance of drift and selection in shaping the distribution of quantitative genetic variation along climatic gradients; (2) the relationship between plant fitness, quantitative genetic variation, molecular genetic variation and population size; and (3) the relationship between the differentiation of a trait among populations and its evolvability.Methods Genetic variation within and among 22 populations from the whole distribution area of S. sponhemica was studied using RAPD (random amplified polymorphic DNA) markers, and climatic variables were obtained for each site. Seeds were collected from each population and germinated, and seedlings were transplanted into a common garden for determination of variation in plant traits.Key Results In contrast to previous results from rare plant species, strong evidence was found for divergent selection. Most population trait means of S. sponhemica were significantly related to climate gradients, indicating adaptation. Quantitative genetic differentiation increased with geographical distance, even when neutral molecular divergence was controlled for, and Q_ST exceeded F_ST for some traits. The evolvability of traits was negatively correlated with the degree of differentiation among populations (Q_ST), i.e. traits under strong selection showed little genetic variation within populations. The evolutionary potential of a population was not related to its size, the performance of the population or its neutral genetic diversity. However, performance in the common garden was lower for plants from populations with reduced molecular genetic variation, suggesting inbreeding depression due to genetic erosion.Conclusions The findings suggest that studies of molecular and quantitative genetic variation may provide complementary insights important for the conservation of rare species. The strong differentiation of quantitative traits among populations shows that selection can be an important force for structuring variation in evolutionarily important traits even for rare endemic species restricted to very specific habitats.     

Little or no gene flow despite F1 hybrids at two interspecific contact zones

Hybridization can create the selective force that promotes assortative mating but hybridization can also select for increased hybrid fitness. Gene flow resulting from hybridization can increase genetic diversity but also reduce distinctiveness. Thus the formation of hybrids has important implications for long‐term species coexistence. This study compares the interaction between the tree wētā Hemideina thoracica and its two neighboring species; H. crassidens and H. trewicki. We examined the ratio of parent and hybrid forms in natural areas of sympatry. Individuals with intermediate phenotype were confirmed as first generation hybrids using nine independent genetic markers. Evidence of gene flow from successful hybridization was sought from the distribution of morphological and genetic characters. Both species pairs appear to be largely retaining their own identity where they live in sympatry, each with a distinct karyotype. Hemideina thoracica and H. trewicki are probably reproductively isolated, with sterile F₁ hybrids. This species pair shows evidence of niche differences with adult size and timing of maturity differing where Hemideina thoracica is sympatric with H. trewicki. In contrast, evidence of a low level of introgression was detected in phenotypes and genotypes where H. thoracica and H. crassidens are sympatric. We found no evidence of size divergence although color traits in combination with hind tibia spines reliably distinguish the two species. This species pair show a bimodal hybrid zone in the absence of assortative mating and possible sexual exclusion by H. thoracica males in the formation of F₁ hybrids.     

Leaf shape evolution has a similar genetic architecture in three edaphic specialists within the Mimulus guttatus species complex

Background and Aims The genetic basis of leaf shape has long interested botanists because leaf shape varies extensively across the plant kingdom and this variation is probably adaptive. However, knowledge of the genetic architecture of leaf shape variation in natural populations remains limited. This study examined the genetic architecture of leaf shape diversification among three edaphic specialists in the Mimulus guttatus species complex. Lobed and narrow leaves have evolved from the entire, round leaves of M. guttatus in M. laciniatus, M. nudatus and a polymorphic serpentine M. guttatus population (M2L).Methods Bulk segregant analysis and next-generation sequencing were used to map quantitative trait loci (QTLs) that underlie leaf shape in an M. laciniatus × M. guttatus F₂ population. To determine whether the same QTLs contribute to leaf shape variation in M. nudatus and M2L, F₂s from M. guttatus × M. nudatus and lobed M2L × unlobed M. guttatus crosses were genotyped at QTLs from the bulk segregant analysis.Key Results Narrow and lobed leaf shapes in M. laciniatus, M. nudatus and M. guttatus are controlled by overlapping genetic regions. Several promising leaf shape candidate genes were found under each QTL.Conclusions The evolution of divergent leaf shape has taken place multiple times in the M. guttatus species complex and is associated with the occupation of dry, rocky environments. The genetic architecture of elongated and lobed leaves is similar across three species in this group. This may indicate that parallel genetic evolution from standing variation or new mutations is responsible for the putatively adaptive leaf shape variation in Mimulus.     

Seed bank dynamics govern persistence of Brassica hybrids in crop and natural habitats

Background and Aims Gene flow from crops to their wild relatives has the potential to alter population growth rates and demography of hybrid populations, especially when a new crop has been genetically modified (GM). This study introduces a comprehensive approach to assess this potential for altered population fitness, and uses a combination of demographic data in two habitat types and mathematical (matrix) models that include crop rotations and outcrossing between parental species.Methods Full life-cycle demographic rates, including seed bank survival, of non-GM Brassica rapa × B. napus F₁ hybrids and their parent species were estimated from experiments in both agricultural and semi-natural habitats. Altered fitness potential was modelled using periodic matrices including crop rotations and outcrossing between parent species.Key Results The demographic vital rates (i.e. for major stage transitions) of the hybrid population were intermediate between or lower than both parental species. The population growth rate (λ) of hybrids indicated decreases in both habitat types, and in a semi-natural habitat hybrids became extinct at two sites. Elasticity analyses indicated that seed bank survival was the greatest contributor to λ. In agricultural habitats, hybrid populations were projected to decline, but with persistence times up to 20 years. The seed bank survival rate was the main driver determining persistence. It was found that λ of the hybrids was largely determined by parental seed bank survival and subsequent replenishment of the hybrid population through outcrossing of B. rapa with B. napus.Conclusions Hybrid persistence was found to be highly dependent on the seed bank, suggesting that targeting hybrid seed survival could be an important management option in controlling hybrid persistence. For local risk mitigation, an increased focus on the wild parent is suggested. Management actions, such as control of B. rapa, could indirectly reduce hybrid populations by blocking hybrid replenishment.     

Spatial and temporal genetic structure in a hybrid cordgrass invasion

Invasive hybrids and their spread dynamics pose unique opportunities to study evolutionary processes. Invasive hybrids of native Spartina foliosa and introduced S. alterniflora have expanded throughout San Francisco Bay intertidal habitats within the past 35 years by deliberate plantation and seeds floating on the tide. Our goals were to assess spatial and temporal scales of genetic structure in Spartina hybrid populations within the context of colonization history. We genotyped adult and seedling Spartina using 17 microsatellite loci and mapped their locations in three populations. All sampled seedlings were hybrids. Bayesian ordination analysis distinguished hybrid populations from parent species, clearly separated the population that originated by plantation from populations that originated naturally by seed and aligned most seedlings within each population. Population genetic structure estimated by analysis of molecular variance was substantial (F_ST=0.21). Temporal genetic structure among age classes varied highly between populations. At one population, the divergence between adults and 2004 seedlings was low (F_ST=0.02) whereas at another population this divergence was high (F_ST=0.26). This latter result was consistent with local recruitment of self-fertilized seed produced by only a few parental plants. We found fine-scale spatial genetic structure at distances less than ∼200 m, further supporting local seed and/or pollen dispersal. We posit a few self-fertile plants dominating local recruitment created substantial spatial genetic structure despite initial long-distance, human dispersal of hybrid Spartina through San Francisco Bay. Fine-scale genetic structure may more strongly develop when local recruits are dominated by the offspring of a few self-fertile plants.     

Pure species in a continuum of genetic and morphological variation: sympatric oaks at the edge of their range

Background and Aims Studies on oaks (Quercus spp.) have often been hampered by taxonomic confusion, a situation further compounded by the occurrence of extensive interspecific hybridization. In the present study, a combination of genetic and morphological analyses was used to examine sympatric populations of Q. petraea and Q. robur at the north-western edge of their ranges in Northern Ireland, since it had previously been suggested that hybridization could facilitate the apparent rapid, long-distance dispersal of oaks following the glaciations.Methods Samples were collected from 24 sites across Northern Ireland that had been previously designated as ancient or semi-natural woodland. Genotypes were obtained from a total of 950 trees using 12 nuclear microsatellite loci, and admixture coefficients were calculated based on a Bayesian clustering approach. Individuals were also classified as Q. petraea, Q. robur or hybrids based on two objective morphometric characters shown previously to delineate pure individuals effectively. Genetically ‘pure’ individuals of both species, as defined by the Bayesian clustering, were also genotyped for five chloroplast microsatellites.Key Results Genetic and morphological analyses both indicated the presence of pure individuals of both species, as well as a continuum of intermediates. There was a good agreement between the molecular and morphological classification, with a generally clear separation between pure individuals.Conclusions Despite millennia of hybridization and introgression, genetically and morphologically pure individuals of both Q. petraea and Q. robur can be found at the edge of their range, where both species occur sympatrically. The high proportion of individuals exhibiting introgression compared with previous studies may reflect the historical role of hybridization in facilitating dispersal following the glaciations. This is further supported by the significantly higher chloroplast diversity in Q. robur compared with Q. petraea.     

High molecular diversity in the true service tree (Sorbus domestica) despite rareness: data from Europe with special reference to the Austrian occurrence

Background and Aims Sorbus domestica (Rosaceae) is one of the rarest deciduous tree species in Europe and is characterized by a scattered distribution. To date, no large-scale geographic studies on population genetics have been carried out. Therefore, the aims of this study were to infer levels of molecular diversity across the major part of the European distribution of S. domestica and to determine its population differentiation and structure. In addition, spatial genetic structure was examined together with the patterns of historic and recent gene flow between two adjacent populations.Methods Leaf or cambium samples were collected from 17 populations covering major parts of the European native range from north-west France to south-east Bulgaria. Seven nuclear microsatellites and one chloroplast minisatellite were examined and analysed using a variety of methods.Key Results Allelic richness was unexpectedly high for both markers within populations (mean per locus: 3·868 for nSSR and 1·647 for chloroplast minisatellite). Moreover, there was no evidence of inbreeding (mean F_is = –0·047). The Italian Peninsula was characterized as a geographic region with comparatively high genetic diversity for both genomes. Overall population differentiation was moderate (F_ST = 0·138) and it was clear that populations formed three groups in Europe, namely France, Mediterranean/Balkan and Austria. Historic gene flow between two local Austrian populations was high and asymmetric, while recent gene flow seemed to be disrupted.Conclusions It is concluded that molecular mechanisms such as self-incompatibility and high gene flow distances are responsible for the observed level of allelic richness as well as for population differentiation. However, human influence could have contributed to the present genetic pattern, especially in the Mediterranean region. Comparison of historic and recent gene flow may mirror the progress of habitat fragmentation in eastern Austria.     

Essential oils from two Eucalyptus from Tunisia and their insecticidal action on Orgyia trigotephras (Lepidotera,Lymantriidae)

Background

Essential oils extracted from aromatic and medicinal plants have many biological properties and are therefore an alternative to the use of synthetic products. The chemical composition of essential oils from two medicinal plants (Eucalyptus globulus and E. lehmannii) was determined and, their insecticidal effects on the third and fourth larval stages of Orgyia trigotephras were assessed.

Results

Larvae were collected from Jebel Abderrahmane (North-East of Tunisia), conserved in groups of 50/box (21 × 10 × 10 cm) at a temperature of 25°C. Larvae were tested for larvicidal activities of essential oils. Each oil was diluted in ethanol (96%) to prepare 3 test solutions (S1 = 0.05%, S2 = 0.10% and S3 = 0.50%). Essential oils were used for contact, ingestion and Olfactory actions and compared to reference products (Bacillus thuringiensis and Decis). Olfactory action of essential oils shows that larvae mortality is higher than contact action, lower than ingestion action. MTM and FTM of S3 of E. lehmannii were respectively 1 h 32 min and 1 h 39 min are higher than those of E. globulus (MTM = 51 min and FTM = 1 h 22 min 34 sec). Contact action of E. lehmannii oil shows low insecticidal activity compared to E. globulus. MTM are respectively (1 min 52 sec and 1 min 7 sec), FTM are (2 min 38 sec, 1 min 39 sec), are the shortest recorded for S3, on the third stage of larvae. The fourth stage of larvae, MTM are (2 min 20 sec and 2 min 9 sec), FTM are (3 min 25 sec, 3 min 19 sec). Ingestion action of essential oils is longer than the contact action, since the time of death exceeds 60 minutes for all species.

Conclusion

Results shows that essential oils have a toxic action on nerves leading to a disruption of vital system of insects. High toxic properties make these plant-derived compounds suitable for incorporation in integrated pest management programs.     

Phylogeography of the endangered rosewood Dalbergia nigra (Fabaceae): insights into the evolutionary history and conservation of the Brazilian Atlantic Forest

The Brazilian rosewood (Dalbergia nigra) is an endangered tree endemic to the central Brazilian Atlantic Forest, one of the world''s most threatened biomes. The population diversity, phylogeographic structure and demographic history of this species were investigated using the variation in the chloroplast DNA (cpDNA) sequences of 185 individuals from 19 populations along the geographical range of the species. Fifteen haplotypes were detected in the analysis of 1297 bp from two non-coding sequences, trnV-trnM and trnL. We identified a strong genetic structure (F_ST=0.62, P<0.0001), with a latitudinal separation into three phylogeographic groups. The two northernmost groups showed evidence of having maintained historically larger populations than the southernmost group. Estimates of divergence times between these groups pointed to vicariance events in the Middle Pleistocene (ca. 350 000–780 000 years ago). The recurrence of past climatic changes in the central part of the Atlantic forest, with cycles of forest expansion and contraction, may have led to repeated vicariance events, resulting in the genetic differentiation of these groups. Based on comparisons among the populations of large reserves and small, disturbed fragments of the same phylogeographic group, we also found evidence of recent anthropogenic effects on genetic diversity. The results were also analysed with the aim of contributing to the conservation of D. nigra. We suggest that the three phylogeographic groups could be considered as three distinct management units. Based on the genetic diversity and uniqueness of the populations, we also indicate priority areas for conservation.     

In glucose-limited continuous culture the minimum substrate concentration for growth,smin, is crucial in the competition between the enterobacterium Escherichia coli and Chelatobacter heintzii,an environmentally abundant bacterium

The competition for glucose between Escherichia coli ML30, a typical copiotrophic enterobacterium and Chelatobacter heintzii ATCC29600, an environmentally successful strain, was studied in a carbon-limited culture at low dilution rates. First, as a base for modelling, the kinetic parameters μ_max and K_s were determined for growth with glucose. For both strains, μ_max was determined in batch culture after different precultivation conditions. In the case of C. heintzii, μ_max was virtually independent of precultivation conditions. When inoculated into a glucose-excess batch culture medium from a glucose-limited chemostat run at a dilution rate of 0.075 h⁻¹ C. heintzii grew immediately with a μ_max of 0.17±0.03 h⁻¹. After five transfers in batch culture, μ_max had increased only slightly to 0.18±0.03 h⁻¹. A different pattern was observed in the case of E. coli. Inoculated from a glucose-limited chemostat at D=0.075 h⁻¹ into glucose-excess batch medium E. coli grew only after an acceleration phase of ∼3.5 h with a μ_max of 0.52 h⁻¹. After 120 generations and several transfers into fresh medium, μ_max had increased to 0.80±0.03 h⁻¹. For long-term adapted chemostat-cultivated cells, a K_s for glucose of 15 μg l⁻¹ for C. heintzii, and of 35 μg l⁻¹ for E. coli, respectively, was determined in ¹⁴C-labelled glucose uptake experiments. In competition experiments, the population dynamics of the mixed culture was determined using specific surface antibodies against C. heintzii and a specific 16S rRNA probe for E. coli. C. heintzii outcompeted E. coli in glucose-limited continuous culture at the low dilution rates of 0.05 and 0.075 h⁻¹. Using the determined pure culture parameter values for K_s and μ_max, it was only possible to simulate the population dynamics during competition with an extended form of the Monod model, which includes a finite substrate concentration at zero growth rate (s_min). The values estimated for s_min were dependent on growth rate; at D=0.05 h⁻¹, it was 12.6 and 0 μg l⁻¹ for E. coli and C. heintzii, respectively. To fit the data at D=0.075 h⁻¹, s_min for E. coli had to be raised to 34.9 μg l⁻¹ whereas s_min for C. heintzii remained zero. The results of the mathematical simulation suggest that it is not so much the higher K_s value, which is responsible for the unsuccessful competition of E. coli at low residual glucose concentration, but rather the existence of a significant s_min.     

Demographic factors shaped diversity in the two gene pools of wild common bean Phaseolus vulgaris L.

Wild common bean (Phaseolus vulgaris L.) is distributed throughout the Americas from Mexico to northern Argentina. Within this range, the species is divided into two gene pools (Andean and Middle American) along a latitudinal gradient. The diversity of 24 wild common bean genotypes from throughout the geographic range of the species was described by using sequence data from 13 loci. An isolation–migration model was evaluated using a coalescent analysis to estimate multiple demographic parameters. Using a Bayesian approach, Andean and Middle American subpopulations with high percentage of parentages were observed. Over all loci, the Middle American gene pool was more diverse than the Andean gene pool (π_sil=0.0089 vs 0.0068). The two subpopulations were strongly genetically differentiated over all loci (F_st=0.29). It is estimated that the two current wild gene pools diverged from a common ancestor ∼111 000 years ago. Subsequently, each gene pool underwent a bottleneck immediately after divergence and lasted ∼40 000 years. The Middle American bottleneck population size was ∼46% of the ancestral population size, whereas the Andean was 26%. Continuous asymmetric gene flow was detected between the two gene pools with a larger number of migrants entering Middle American gene pool from the Andean gene pool. These results suggest that because of the complex population structure associated with the ancestral divergence, subsequent bottlenecks in each gene pool, gene pool-specific domestication and intense selection within each gene pool by breeders; association mapping would best be practised within each common bean gene pool.     

A genome-guided analysis of energy conservation in the thermophilic,cytochrome-free acetogenic bacterium Thermoanaerobacter?kivui

Background

Acetogenic bacteria are able to use CO₂ as terminal electron acceptor of an anaerobic respiration, thereby producing acetate with electrons coming from H₂. Due to this feature, acetogens came into focus as platforms to produce biocommodities from waste gases such as H₂ + CO₂ and/or CO. A prerequisite for metabolic engineering is a detailed understanding of the mechanisms of ATP synthesis and electron-transfer reactions to ensure redox homeostasis. Acetogenesis involves the reduction of CO₂ to acetate via soluble enzymes and is coupled to energy conservation by a chemiosmotic mechanism. The membrane-bound module, acting as an ion pump, was of special interest for decades and recently, an Rnf complex was shown to couple electron flow from reduced ferredoxin to NAD⁺ with the export of Na⁺ in Acetobacterium woodii. However, not all acetogens have rnf genes in their genome. In order to gain further insights into energy conservation of non-Rnf-containing, thermophilic acetogens, we sequenced the genome of Thermoanaerobacter kivui.

Results

The genome of Thermoanaerobacter kivui comprises 2.9 Mbp with a G + C content of 35% and 2,378 protein encoding orfs. Neither autotrophic growth nor acetate formation from H₂ + CO₂ was dependent on Na⁺ and acetate formation was inhibited by a protonophore, indicating that H⁺ is used as coupling ion for primary bioenergetics. This is consistent with the finding that the c subunit of the F₁F_O ATP synthase does not have the conserved Na⁺ binding motif. A search for potential H⁺-translocating, membrane-bound protein complexes revealed genes potentially encoding two different proton-reducing, energy-conserving hydrogenases (Ech).

Conclusions

The thermophilic acetogen T. kivui does not use Na⁺ but H⁺ for chemiosmotic ATP synthesis. It does not contain cytochromes and the electrochemical proton gradient is most likely established by an energy-conserving hydrogenase (Ech). Its thermophilic nature and the efficient conversion of H₂ + CO₂ make T. kivui an interesting acetogen to be used for the production of biocommodities in industrial micobiology. Furthermore, our experimental data as well as the increasing number of sequenced genomes of acetogenic bacteria supported the new classification of acetogens into two groups: Rnf- and Ech-containing acetogens.     

Diversity and relationships of Crocus sativus and its relatives analysed by inter-retroelement amplified polymorphism (IRAP)

Background and Aims: Saffron (Crocus sativus) is a sterile triploid (2n = 3x = 24) cultivated species, of unknown origin from other diploid and polyploid species in the genus Crocus (Iridaceae). Species in the genus have high morphological diversity, with no clear phylogenetic patterns below the level of section Crocus series Crocus. Using DNA markers, this study aimed to examine the diversity and relationships within and between species of Crocus series Crocus.Methods: Eleven inter-retroelement amplified polymorphism (IRAP) primers were used in 63 different combinations with 35 single-plant accessions of C. sativus and related Crocus species in order to determine genetic variability and to conduct phylogenetic analysis.Key Results: A total of 4521 distinct polymorphic bands from 100 bp to approx. 4 kb were amplified; no fragment specific to all accessions of a single species was amplified. The polymorphic information content (PIC) values varied from approx. 0·37 to approx. 0·05 (mean 0·17 ± 0·1) and the major allele frequency had a mean of 0·87. High levels of polymorphism were identified between accessions of the six species of Crocus series Crocus related to C. sativus, with further variation between the species. In contrast, no polymorphisms were seen among 17 C. sativus accessions obtained in the region from Kashmir through Iran to Spain.Conclusions In contrast to the intraspecific variability seen in other Crocus species, C. sativus has minimal genetic variation, and it is concluded that the triploid hybrid species has most probably arisen only once. The data show that saffron is an allotriploid species, with the IRAP analysis indicating that the most likely ancestors are C. cartwrightianus and C. pallasii subsp. pallasii (or close relatives). The results may facilitate resynthesizing saffron with improved characteristics, and show the need for conservation and collection of wild Crocus.     

Identification of a common risk haplotype for canine idiopathic epilepsy in the ADAM23 gene

Background

Idiopathic epilepsy is a common neurological disease in human and domestic dogs but relatively few risk genes have been identified to date. The seizure characteristics, including focal and generalised seizures, are similar between the two species, with gene discovery facilitated by the reduced genetic heterogeneity of purebred dogs. We have recently identified a risk locus for idiopathic epilepsy in the Belgian Shepherd breed on a 4.4 megabase region on CFA37.

Results

We have expanded a previous study replicating the association with a combined analysis of 157 cases and 179 controls in three additional breeds: Schipperke, Finnish Spitz and Beagle (p_c = 2.9e–07, p_GWAS = 1.74E-02). A targeted resequencing of the 4.4 megabase region in twelve Belgian Shepherd cases and twelve controls with opposite haplotypes identified 37 case-specific variants within the ADAM23 gene. Twenty-seven variants were validated in 285 cases and 355 controls from four breeds, resulting in a strong replication of the ADAM23 locus (p_raw = 2.76e–15) and the identification of a common 28 kb-risk haplotype in all four breeds. Risk haplotype was present in frequencies of 0.49–0.7 in the breeds, suggesting that ADAM23 is a low penetrance risk gene for canine epilepsy.

Conclusions

These results implicate ADAM23 in common canine idiopathic epilepsy, although the causative variant remains yet to be identified. ADAM23 plays a role in synaptic transmission and interacts with known epilepsy genes, LGI1 and LGI2, and should be considered as a candidate gene for human epilepsies.

Electronic supplementary material

The online version of this article (doi:10.1186/s12864-015-1651-9) contains supplementary material, which is available to authorized users.     

Unidirectional hybridization and reproductive barriers between two heterostylous primrose species in north-west Yunnan,China

Background and Aims

Heteromorphy in flowers has a profound effect on breeding patterns within a species, but little is known about how it affects reproductive barriers between species. The heterostylous genus Primula is very diverse in the Himalaya region, but hybrids there have been little researched. This study examines in detail a natural hybrid zone between P. beesiana and P. bulleyana.

Methods

Chloroplast sequencing, AFLP (amplified fragment length polymorphism) markers and morphological comparisons were employed to characterize putative hybrids in the field, using synthetic F₁s from hand pollination as controls. Pollinator visits to parent species and hybrids were observed in the field. Hand pollinations were conducted to compare pollen tube growth, seed production and seed viability for crosses involving different morphs, species and directions of crossing.

Key Results

Molecular data revealed all hybrid derivatives examined to be backcrosses of first or later generations towards P. bulleyana: all had the chloroplast DNA (cpDNA) of this species. Some individuals had morphological traits suggesting they were hybrids, but they were genetically similar to P. bulleyana; they might have been advanced generation backcrosses. Viable F₁s could not be produced with P. bulleyana pollen on P. beesiana females, irrespective of the flower morphs used. Within-morph crosses for each species had very low (<10 %) seed viability, whereas crosses between pin P. bulleyana (female) and pin P. beesiana had a higher seed viability of 30 %. Thus genetic incompatibility mechanisms back up mechanical barriers to within-morph crosses in each species, but are not the same between the two species. The two species share their main pollinators, and pollinators were observed to fly between P. bulleyana and hybrids, suggesting that pollinator behaviour may not be an important isolating factor.

Conclusions

Hybridization is strongly asymmetric, with P. bulleyana the only possible mother and all detected hybrids being backcrosses in this direction. Partial ecological isolation and inhibition of heterospecific pollen, and possibly complete barriers to F₁ formation on P. beesiana, may be enough to make F₁ formation very rare in these species. Therefore, with no F₁ detected, this hybrid zone may have a finite life span as successive generations become more similar to P. bulleyana.     

Novel Clostridium populations involved in the anaerobic degradation of Microcystis blooms

Understanding the microbial degradation of Microcystis biomass is crucial for determining the ecological consequences of Microcystis blooms in freshwater lakes. The purpose of this study was to identify bacteria involved in the anaerobic degradation of Microcystis blooms. Microcystis scum was anaerobically incubated for 90 days at three temperatures (15 °C, 25 °C and 35 °C). We used terminal restriction fragment length polymorphism (T-RFLP) analysis of bacterial 16S rRNA genes, followed by cloning and sequencing of selected samples, to reveal the community composition of bacteria and their dynamics during decomposition. Clostridium spp. were found to be the most dominant bacteria in the incubations, accounting for 72% of the sequenced clones. Eight new clusters or subclusters (designated CLOS.1–8) were identified in the Clostridium phylogenetic tree. The bacterial populations displayed distinct successions during Microcystis decomposition. Temperature had a strong effect on the dynamics of the bacterial populations. At 15 °C, the initial dominance of a 207-bp T-RF (Betaproteobacteria) was largely substituted by a 227-bp T-RF (Clostridium, new cluster CLOS.2) at 30 days. In contrast, at 25 °C and 35 °C, we observed an alternating succession of the 227-bp T-RF and a 231-bp T-RF (Clostridium, new cluster CLOS.1) that occurred more than four times; no one species dominated the flora for the entire experiment. Our study shows that novel Clostridium clusters and their diverse consortiums dominate the bacterial communities during anaerobic degradation of Microcystis, suggesting that these microbes'' function in the degradation process.     

Polyphenol oxidase-mediated protection against oxidative stress is not associated with enhanced photosynthetic efficiency

Background and Aims Polyphenol oxidases (PPOs) catalyse the oxidation of monophenols and/or o-diphenols to highly reactive o-quinones, which in turn interact with oxygen and proteins to form reactive oxygen species (ROS) and typical brown-pigmented complexes. Hence PPOs can affect local levels of oxygen and ROS. Although the currently known substrates are located in the vacuole, the enzyme is targeted to the thylakoid lumen, suggesting a role for PPOs in photosynthesis. The current study was designed to investigate the potential involvement of PPOs in the photosynthetic response to oxidative stress.Methods Photosynthesis (A, F_v/F_m, ΦPSII, q_N, q_P, NPQ) was measured in leaves of a wild-type and a low-PPO mutant of red clover (Trifolium pratense ‘Milvus’) under control conditions and under a stress treatment designed to induce photooxidative stress: cold/high light (2 °C/580 µmol m²s^–1) or 0–10 µm methyl viologen. Foliar protein content and oxidation state were also determined.Key Results Photosynthetic performance, and chlorophyll and protein content during 4 d of cold/high light stress and 3 d of subsequent recovery under control growth conditions showed similar susceptibility to stress in both lines. However, more extensive oxidative damage to protein in mutants than wild-types was observed after treatment of attached leaves with methyl viologen. In addition, PPO activity could be associated with an increased capacity to dissipate excess energy, but only at relatively low methyl viologen doses.Conclusions The presence of PPO activity in leaves did not correspond to a direct role for the enzyme in the regulation or protection of photosynthesis under cold stress. However, an indication that PPO could be involved in cellular protection against low-level oxidative stress requires further investigation.