Comparative study of genotoxicity and antimutagenicity of methanolic extracts from Teucrium chamaedrys and Teucrium montanum in human lymphocytes using micronucleus assay

Since Teucrium chamaedrys and Teucrium montanum are the most popular plants used in the treatment of many diseases, we evaluated genotoxic potential of their methanolic extracts on cultured human peripheral blood lymphocytes (PBLs) using cytokinesis-block micronucleus (MN) assay. Cultures were treated with four concentrations of both plants (125, 250, 500 and 1,000 μg/ml), both separately and in combination with mitomycin C (MMC). The results revealed that extract of T. chamaedrys administered at the tested concentrations did not significantly affect the mean MN frequency in comparison to untreated cells. Methanolic extract of T. montanum increased the mean MN frequency in PBL at the tested concentrations, but significantly only at the concentration of 1,000 μg/ml. In all tested concentrations, the extract of T. chamaedrys significantly reduced the MMC-induced MN frequency, in a dose dependent manner (r = − 0.687, p < 0.01). The extract of T. montanum decreased the MMC-induced MN frequency at the tested concentrations, but statistically only at 125 μg/ml. Both extracts administered alone did not significantly affect the nuclear division index (NDI) at the tested concentrations. In the combined treatments with MMC, the extract obtained from T. chamaedrys in the concentrations of 500 and 1,000 μg/ml significantly decreased NDI values in comparison to MMC-treated cells alone, while the extract of T. montanum significantly decreased NDI at all tested concentrations. Both extracts nonsignificantly decreased NDI at all tested concentrations in comparison to untreated cells. Our results suggest the important function of T. chamaedrys extract in cancer therapy, this methanolic extract may prevent genotoxic effects of chemotherapy in PBLs.     

The interplay of nurse and target plant traits influences magnitude and direction of facilitative interactions under different combinations of stress and disturbance intensities in Andean dry grassland

Aims Facilitation is a key process in vegetation dynamics, driving the response to natural and anthropogenic pressures. In harsh-grazed systems, palatable plants mainly survive when nested under unpalatable tussocks and shrubs. The magnitude and direction of positive interactions are driven by resource availability, extent of herbivory and type of nurse species. We hypothesized that different combinations of disturbance and environmental stress affect community composition in the dry Puna (southern Peruvian Andes) by modifying nurse types and plant interactions in magnitude and specific associations. We investigated whether different combinations of stress and disturbance influence species richness, type and frequency of occurrence of nurse and beneficiary species and magnitude and patterns of plant interactions; whether nurse species influence these interactions and target species change their interactions under different combinations of stress and disturbance and whether plant functional traits differ in the studied communities and influence the pattern of spatial interactions.Methods We selected three plant communities subject to different precipitation and management regimes: in each we laid a number of transects proportional to its extension. Data collected include species presence/absence, type of spatial interactions with nurse species and functional traits. We calculated species richness and rarefaction patterns, described the patterns of plant–plant spatial interactions and investigated the associations between nurse and other species in the three communities using indicator species analysis (ISA). We performed ISA and correlation analysis to investigate whether plant functional traits influenced facilitative interactions.Important findings We found that different combinations of stress and disturbance shaped a complex set of responses, including changes in the nurse species set. Nurse composition influenced magnitude and direction of plant interactions under different stress intensities. Heavy disturbance increased the relative importance of facilitation, even if the overall number of facilitated species decreased. Under equivalent disturbance regimes, increased abiotic stress led to a greater importance of facilitation. Different combinations of stress and disturbance affected the community assemblage also by changing the behaviour of some non-nurse species. Both heavy disturbance and strong stress led to a decrease of trait states; with certain combinations of stress and disturbance, preferential distribution of these states was observed. We also found that plant traits were of key importance in determining facilitative interactions. Some traits were mainly associated with one type of spatial interaction: plant architecture, life cycle and root type influenced the type of interaction between nurses and beneficiaries under different combinations of stress and disturbance. Our results also demonstrate that in plant interaction research the object of observations (species per se, species percentage, etc.) might influence outputs, and to effectively assess the impact of different stress and disturbance intensities on plant interactions it is necessary to work at the community level to consider the whole species pool.     

The potential of plant viruses to promote genotypic diversity via genotype x environment interactions

Background and Aims

Genotype by environment (G × E) interactions are important for the long-term persistence of plant species in heterogeneous environments. It has often been suggested that disease is a key factor for the maintenance of genotypic diversity in plant populations. However, empirical evidence for this contention is scarce. Here virus infection is proposed as a possible candidate for maintaining genotypic diversity in their host plants.

Methods

The effects of White clover mosaic virus (WClMV) on the performance and development of different Trifolium repens genotypes were analysed and the G × E interactions were examined with respect to genotype-specific plant responses to WClMV infection. Thus, the environment is defined as the presence or absence of the virus.

Key Results

WClMV had a negative effect on plant performance as shown by a decrease in biomass and number of ramets. These effects of virus infection differ greatly among host genotypes, representing a strong G × E interaction. Moreover, the relative fitness and associated ranking of genotypes changed significantly between control and virus treatments. This shift in relative fitness among genotypes suggests the potential for WClMV to provoke differential selection on T. repens genotypes, which may lead to negative frequency-dependent selection in host populations.

Conclusions

The apparent G × E interaction and evident repercussions for relative fitness reported in this study stress the importance of viruses for ecological and evolutionary processes and suggest an important role for viruses in shaping population dynamics and micro-evolutionary processes.     

The shape of things to come: woodland herb niche contraction begins during recruitment in mesic forest microhabitat

Natural abundance is shaped by the abiotic requirements and biotic interactions that shape a species' niche, yet these influences are rarely decoupled. Moreover, most plant mortality occurs during early life stages, making seed recruitment critical in structuring plant populations. We find that natural abundance of two woodland herbs, Hexastylis arifolia and Hepatica nobilis, peaks at intermediate resource levels, a pattern probably formed by concurrent abiotic and biotic interactions. To determine how this abundance patterning reflects intrinsic physiological optima and extrinsic biotic interactions, we translocate adults and seeds to novel locations across experimentally extended abiotic gradients. These experiments indicate that the plant distributions probably reflect biotic interactions as much as physiological requirements, and that adult abundance provides a poor indication of the underlying niche requirements. The positive response exhibited by adult transplants in the wettest conditions is offset by increased fungal attack on buried seeds consistent with peak natural abundance where soil moisture is intermediate. This contraction of niche space is best described by Connell's model--species are limited by physiological tolerances where resources are low and biotic interactions where resources are high.     

Oxidative stress induces distinct physiological responses in the two Trebouxia phycobionts of the lichen Ramalina farinacea

Background and Aims

Most lichens form associations with Trebouxia phycobionts and some of them simultaneously include genetically different algal lineages. In other symbiotic systems involving algae (e.g. reef corals), the relative abundances of different endosymbiotic algal clades may change over time. This process seems to provide a mechanism allowing the organism to respond to environmental stress. A similar mechanism may operate in lichens with more than one algal lineage, likewise protecting them against environmental stresses. Here, the physiological responses to oxidative stress of two distinct Trebouxia phycobionts (provisionally named TR1 and TR9) that coexist within the lichen Ramalina farinacea were analysed.

Methods

Isolated phycobionts were exposed to oxidative stress through the reactive oxygen species propagator cumene hydroperoxide (CuHP). Photosynthetic pigments and proteins, photosynthesis (through modulated chlorophyll fluorescence), the antioxidant enzymes superoxide dismutase (SOD) and glutathione reductase (GR), and the stress-related protein HSP70 were analysed.

Key Results

Photosynthetic performance was severely impaired by CuHP in phycobionts, as indicated by decreases in the maximal PSII photochemical efficiency (F_v/F_m), the quantum efficiency of PSII (Φ_PSII) and the non-photochemical dissipation of energy (NPQ). However, the CuHP-dependent decay in photosynthesis was significantly more severe in TR1, which also showed a lower NPQ and a reduced ability to preserve chlorophyll a, carotenoids and D1 protein. Additionally, differences were observed in the capacities of the two phycobionts to modulate antioxidant activities and HPS70 levels when exposed to oxidative stress. In TR1, CuHP significantly diminished HSP70 and GR but did not change SOD activities. In contrast, in TR9 the levels of both antioxidant enzymes and those of HSP70 increased in response to CuHP.

Conclusions

The better physiological performance of TR9 under oxidative conditions may reflect its greater capacity to undertake key metabolic adjustments, including increased non-photochemical quenching, higher antioxidant protection and the induction of repair mechanisms.     

The relative importance of positive and negative interactions for pollinator attraction in a plant community

Plant–pollinator interactions provide ideal frameworks for studying interactions in plant communities. Despite the large potential influence of such interactions on plant community structure, biodiversity and evolutionary processes, we know surprisingly little about the relative importance of positive and negative interactions among plant species for pollinator attraction. Therefore, we explored the relationships between conspecific and heterospecific floral densities and the flower visitation rates of nine plant species mainly visited by bumble bees, and six plant species mainly visited by flies, in a temperate grassland, through stepwise multiple regressions. Significant relationships were interpreted as interactions for pollinator attraction. Our results revealed that positive intra- and interspecific interactions for pollinator attraction were far more frequent than negative ones. Seventeen interspecific interactions were revealed of which 14 were significantly positive, whereas three of four significant intraspecific interactions were positive. Seven species experienced only positive interactions and two species experienced only negative interactions. The results presented here indicate that negative interactions are not necessarily the dominant ecological interaction for pollination among plants within a community, and the study represents a straightforward approach to study intra- and interspecific interactions among multiple species within a community. We discuss which mechanisms may drive the positive interactions for pollinator attraction and whether this may result in facilitative effects on reproductive success. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

The roles of stochasticity and biotic interactions in the spatial patterning of plant species in alpine communities

Questions

Plant community composition can be influenced by multiple biotic, abiotic, and stochastic factors acting on the local species pool to determine their establishment success and abundance and subsequently the diversity of the community. We asked if the influences of biotic interactions on the composition of plant species in communities, as indicated by patterns of plant species spatial associations (independent, positive or negative), vary across a productivity gradient within a single ecosystem type. Do dominant species of communities show spatial patterning suggestive of competitive interactions with interspecific neighbors? Do species that span multiple community types exhibit the same heterospecific interactions with neighbours in each community?

Location

Three alpine communities in the southern Rocky Mountains.

Methods

We measured the occurrence of species in a 1‐cm spatial grid within 2 m × 2 m plots to determine the spatial patterns of species pairs in the three communities. A null model of independent species spatial arrangements was used to determine whether species pairs were positively, negatively or independently associated, and how these patterns differed among the communities across the gradient of resource supply and environmental stress.

Results

Positive associations, indicative of facilitation between species, were most common in the most resource‐poor and least productive community. However negative associations, suggestive of competitive interactions among species, were not more common in the two more resource‐rich, productive communities. The dominant species of these communities did exhibit higher negative than positive associations with neighbours relative to positive patterning. Independent interspecific patterning was equally common relative to positive and negative patterns in all communities. Species that previously were shown to either facilitate other species or compete with neighbours exhibited spatial patterning consistent with the earlier experimental work.

Conclusions

A large number of species exhibit a lack of net biotic interactions, and stochastic factors appear to be as important as competition and facilitation in shaping the structure of the three alpine plant communities we studied.

     

Biological seed priming mitigates the effects of water stress in sunflower seedlings

The sunflower (Helianthus annuus L. cv. PAC 36) seedlings were inoculated with plant growth promoting rhizobacteria (PGPR), viz. Azotobacter chroococcum (A+), Bacillus polymyxa (B+), separately and in combination of the two (AB+). Relative water content and seedling growth were maximum in AB+ seedlings under control. Water stress significantly decreased the RWC, growth and dry mass of non-inoculated seedlings. However, inoculated seedlings maintained higher growth even under water stress. Pigments and protein contents decreased under water stress, but higher amount of the same was observed in stressed AB+ seedlings. Enhanced activity of nitrate reductase was recorded in AB+ seedlings with maximum in control. Water stress significantly decreased the nitrate reductase activity. A significant increase in the activity of superoxide dismutase (SOD) in leaves was recorded under water stress except in B+ with maximum increase in non-inoculated seedlings. Catalase (CAT) activity decreased in stressed non-inoculated seedlings while increased in the leaves of A+ and AB+ seedlings. Almost similar trends were recorded for both leaves and cotyledons. PGPR improved the water status in stressed seedlings and thereby physiological and biochemical parameters and thus ameliorated the severe effects of water stress.     

Effects of geographical heterogeneity in species interactions on the evolution of venom genes

Geographical heterogeneity in the composition of biotic interactions can create a mosaic of selection regimes that may drive the differentiation of phenotypes that operate at the interface of these interactions. Nonetheless, little is known about effects of these geographical mosaics on the evolution of genes encoding traits associated with species interactions. Predatory marine snails of the family Conidae use venom, a cocktail of conotoxins, to capture prey. We characterized patterns of geographical variation at five conotoxin genes of a vermivorous species, Conus ebraeus, at Hawaii, Guam and American Samoa, and evaluated how these patterns of variation are associated with geographical heterogeneity in prey utilization. All populations show distinct patterns of prey utilization. Three ‘highly polymorphic’ conotoxin genes showed significant geographical differences in allelic frequency, and appear to be affected by different modes of selection among populations. Two genes exhibited low levels of diversity and a general lack of differentiation among populations. Levels of diversity of ‘highly polymorphic’ genes exhibit a positive relationship with dietary breadth. The different patterns of evolution exhibited by conotoxin genes suggest that these genes play different roles in prey capture, and that some genes are more greatly affected by differences in predator–prey interactions than others. Moreover, differences in dietary breadth appear to have a greater influence on the differentiation of venoms than differences in the species of prey.     

Lack of evidence for intermolecular epistatic interactions between adiponectin and resistin gene polymorphisms in Malaysian male subjects

Epistasis (gene-gene interaction) is a ubiquitous component of the genetic architecture of complex traits such as susceptibility to common human diseases. Given the strong negative correlation between circulating adiponectin and resistin levels, the potential intermolecular epistatic interactions between ADIPOQ (SNP+45T > G, SNP+276G > T, SNP+639T > C and SNP+1212A > G) and RETN (SNP-420C > G and SNP+299G > A) gene polymorphisms in the genetic risk underlying type 2 diabetes (T2DM) and metabolic syndrome (MS) were assessed. The potential mutual influence of the ADIPOQ and RETN genes on their adipokine levels was also examined. The rare homozygous genotype (risk alleles) of SNP-420C > G at the RETN locus tended to be co-inherited together with the common homozygous genotypes (protective alleles) of SNP+639T > C and SNP+1212A > G at the ADIPOQ locus. Despite the close structural relationship between the ADIPOQ and RETN genes, there was no evidence of an intermolecular epistatic interaction between these genes. There was also no reciprocal effect of the ADIPOQ and RETN genes on their adipokine levels, i.e., ADIPOQ did not affect resistin levels nor did RETN affect adiponectin levels. The possible influence of the ADIPOQ gene on RETN expression warrants further investigation.     

The role of multiple partners in a digestive mutualism with a protocarnivorous plant

Background and aims

The protocarnivorous plant Paepalanthus bromelioides (Eriocaulaceae) is similar to bromeliads in that this plant has a rosette-like structure that allows rainwater to accumulate in leaf axils (i.e. phytotelmata). Although the rosettes of P. bromelioides are commonly inhabited by predators (e.g. spiders), their roots are wrapped by a cylindrical termite mound that grows beneath the rosette. In this study it is predicted that these plants can derive nutrients from recycling processes carried out by termites and from predation events that take place inside the rosette. It is also predicted that bacteria living in phytotelmata can accelerate nutrient cycling derived from predators.

Methods

The predictions were tested by surveying plants and animals, and also by performing field experiments in rocky fields from Serra do Cipó, Brazil, using natural abundance and enriched isotopes of ¹⁵N. Laboratory bioassays were also conducted to test proteolytic activities of bacteria from P. bromelioides rosettes.

Key Results

Analyses of ¹⁵N in natural nitrogen abundances showed that the isotopic signature of P. bromelioides is similar to that of carnivorous plants and higher than that of non-carnivorous plants in the study area. Linear mixing models showed that predatory activities on the rosettes (i.e. spider faeces and prey carcass) resulted in overall nitrogen contributions of 26·5 % (a top-down flux). Although nitrogen flux was not detected from termites to plants via decomposition of labelled cardboard, the data on ¹⁵N in natural nitrogen abundance indicated that 67 % of nitrogen from P. bromelioides is derived from termites (a bottom-up flux). Bacteria did not affect nutrient cycling or nitrogen uptake from prey carcasses and spider faeces.

Conclusions

The results suggest that P. bromelioides derive nitrogen from associated predators and termites, despite differences in nitrogen cycling velocities, which seem to have been higher in nitrogen derived from predators (leaves) than from termites (roots). This is the first study that demonstrates partitioning effects from multiple partners in a digestion-based mutualism. Despite most of the nitrogen being absorbed through their roots (via termites), P. bromelioides has all the attributes necessary to be considered as a carnivorous plant in the context of digestive mutualism.     

Assessing the regulation of leaf redox status under water stress conditions in Arabidopsis thaliana: Col-0 ecotype (wild-type and vtc-2), expressing mitochondrial and cytosolic roGFP1

Using Arabidopsis plants Col-0 and vtc2 transformed with a redox sensitive green fluorescent protein, (c-roGFP) and (m-roGFP), we investigated the effects of a progressive water stress and re-watering on the redox status of the cytosol and the mitochondria. Our results establish that water stress affects redox status differently in these two compartments, depending on phenotype and leaf age, furthermore we conclude that ascorbate plays a pivotal role in mediating redox status homeostasis and that Col-0 Arabidopsis subjected to water stress increase the synthesis of ascorbate suggesting that ascorbate may play a role in buffering changes in redox status in the mitochondria and the cytosol, with the presumed buffering capacity of ascorbate being more noticeable in young compared with mature leaves. Re-watering of water-stressed plants was paralleled by a return of both the redox status and ascorbate to the levels of well-watered plants. In contrast to the effects of water stress on ascorbate levels, there were no significant changes in the levels of glutathione, thereby suggesting that the regeneration and increase in ascorbate in water-stressed plants may occur by other processes in addition to the regeneration of ascorbate via the glutathione. Under water stress in vtc2 lines it was observed stronger differences in redox status in relation to leaf age, than due to water stress conditions compared with Col-0 plants. In the vtc2 an increase in DHA was observed in water-stressed plants. Furthermore, this work confirms the accuracy and sensitivity of the roGFP1 biosensor as a reporter for variations in water stress-associated changes in redox potentials.     

Variation for host range within and among populations of the parasitic plant Striga hermonthica

Striga hermonthica is an angiosperm parasite that causes substantial damage to a wide variety of cereal crop species, and to the livelihoods of subsistence farmers in sub-Saharan Africa. The broad host range of this parasite makes it a fascinating model for the study of host-parasite interactions, and suggests that effective long-term control strategies for the parasite will require an understanding of the potential for host range adaptation in parasite populations. We used a controlled experiment to test the extent to which the success or failure of S. hermonthica parasites to develop on a particular host cultivar (host resistance/compatibility) depends upon the identity of interacting host genotypes and parasite populations. We also tested the hypothesis that there is a genetic component to host range within individual S. hermonthica populations, using three rice cultivars with known, contrasting abilities to resist infection. The developmental success of S. hermonthica parasites growing on different rice-host cultivars (genotypes) depended significantly on a parasite population by host-genotype interaction. Genetic analysis using amplified fragment length polymorphism (AFLP) markers revealed that a small subset of AFLP markers showed 'outlier' genetic differentiation among sub-populations of S. hermonthica attached to different host cultivars. We suggest that, this indicates a genetic component to host range within populations of S. hermonthica, and that a detailed understanding of the genomic loci involved will be crucial in understanding host-parasite specificity and in breeding crop cultivars with broad spectrum resistance to S. hermonthica.     

Twofold cost of reproduction: an increase in parental effort leads to higher malarial parasitaemia and to a decrease in resistance to oxidative stress

Parental effort is usually associated with high metabolism that could lead to an increase in the production of reactive oxidative species giving rise to oxidative stress. Since many antioxidants involved in the resistance to oxidative stress can also enhance immune function, an increase in parental effort may diminish the level of antioxidants otherwise involved in parasite resistance. In the present study, we performed brood size manipulation in a population of great tits (Parus major) to create different levels of parental effort. We measured resistance to oxidative stress and used a newly developed quantitative PCR assay to quantify malarial parasitaemia. We found that males with an enlarged brood had significantly higher level of malarial parasites and lower red blood cell resistance to free radicals than males rearing control and reduced broods. Brood size manipulation did not affect female parasitaemia, although females with an enlarged brood had lower red blood cell resistance than females with control and reduced broods. However, for both sexes, there was no relationship between the level of parasitaemia and resistance to oxidative stress, suggesting a twofold cost of reproduction. Our results thus suggest the presence of two proximate and independent mechanisms for the well-documented trade-off between current reproductive effort and parental survival.     

The potential roles of strigolactones and brassinosteroids in the autoregulation of nodulation pathway

Background and Aims

The number of nodules formed on a legume root system is under the strict genetic control of the autoregulation of nodulation (AON) pathway. Plant hormones are thought to play a role in AON; however, the involvement of two hormones recently described as having a largely positive role in nodulation, strigolactones and brassinosteroids, has not been examined in the AON process.

Methods

A genetic approach was used to examine if strigolactones or brassinosteroids interact with the AON system in pea (Pisum sativum). Double mutants between shoot-acting (Psclv2, Psnark) and root-acting (Psrdn1) mutants of the AON pathway and strigolactone-deficient (Psccd8) or brassinosteroid-deficient (lk) mutants were generated and assessed for various aspects of nodulation. Strigolactone production by AON mutant roots was also investigated.

Key Results

Supernodulation of the roots was observed in both brassinosteroid- and strigolactone-deficient AON double-mutant plants. This is despite the fact that the shoots of these plants displayed classic strigolactone-deficient (increased shoot branching) or brassinosteroid-deficient (extreme dwarf) phenotypes. No consistent effect of disruption of the AON pathway on strigolactone production was found, but root-acting Psrdn1 mutants did produce significantly more strigolactones.

Conclusions

No evidence was found that strigolactones or brassinosteroids act downstream of the AON genes examined. While in pea the AON mutants are epistatic to brassinosteroid and strigolactone synthesis genes, we argue that these hormones are likely to act independently of the AON system, having a role in the promotion of nodule formation.