Annua Populations Research Articles

Do seasons impact the root microbiome of wild Poa annua populations?

Low temperatures, one of the main characteristics of winters in temperate climates, are restrictive for agriculture, because they negatively affect plant growth and development. To alleviate cold stress, plants are able to establish a symbiosis with soil bacteria. Annual bluegrass (Poa annua) is adapted to low temperatures, has a global distribution, and colonizes a wide range of environments, including polar regions. Here, Poa annua plants were harvested randomly within a one square meter plot at five sites in Flanders, Belgium, during September (summer samples) and March (winter samples) over 2019 and 2020. For each time point, location and season, ten samples of bulk soil, rhizosphere soil and root endosphere were obtained, making a total of 570 samples. As specialized bacteria in the winter root microbiome may reduce the negative impact of cold, we wanted to unravel how winter temperatures affect the root microbiome. Amplicon sequencing of 16S rRNA genes revealed that location was the factor with the largest influence on the bacterial community composition, followed by compartment (root endosphere, rhizosphere, and bulk soil), season, and year. While season was not the main determining factor of the root endosphere, comparison of the summer and winter samples allowed the identification of cold-enriched root-colonizing bacterial families. Interestingly, one ASV corresponding to Massilia was enriched in all sites during the winter season and, thus, may represent an interesting taxonomic group for the development of biostimulants to help plants cope with cold conditions.

Survey of target site resistance alleles conferring resistance in Poa annua

AbstractPoa annua L. (annual bluegrass) is a common weed in turfgrass and has been reported resistant to 12 different herbicide sites of action, with various combinations of multiple‐herbicide resistance having been identified. To quantify the extent of herbicide‐resistant P. annua, the ResistPoa Project (resistpoa.org) surveyed 1349 P. annua populations for resistance to nine sites of action and one plant growth retardant. Herein, we report results from sequencing of known target site mutations found in 5‐enolpyruvylshikimate‐3 phosphate synthase (EPSPS), acetolactate synthase (ALS), photosystem II protein D (psbA), and 𝛼‐tubulin genes. Populations were sequenced using either capillary or amplicon sequencing (AmpSeq), depending on the complexity of the gene, and were analyzed for target‐site resistance. After additional resistance screening, a total of 389 suspected resistant populations were sequenced—131 for ALS, 83 for EPSPS, 93 for psbA, and 82 for 𝛼‐tubulin. From the resistant populations, 64 displayed resistance to multiple sites of action. After sequencing, it was determined that target‐site resistance was the common form of resistance for all sites of action outside of psbA with 65.6% of ALS populations, 73.5% of EPSPS, 39.8% of psbA, and 91.5% of 𝛼‐tubulin having presented a target‐site mutation.

Developing population identification tool based on polymorphism of rDNA for traditional Chinese medicine: Artemisia annua L.

BackgroundArtemisia annua, a well-known traditional Chinese medicine, is the main source for production of artemisinin, an anti-malaria drug. A. annua is distributed globally, with great diversity of morphological characteristics and artemisinin contents. Diverse traits among A. annua populations impeded the stable production of artemisinin, which needs an efficient tool to identify strains and assess population genetic homogeneity. PurposeIn this study, ribosomal DNA (rDNA), were characterized for A. annua for strains identification and population genetic homogeneity assessment. MethodsThe ribosomal RNA (rRNA) genes were identified using cmscan and assembled using rDNA unit of LQ-9 as a reference. rDNA among Asteraceae species were compared performing with 45S rDNA. The rDNA copy number was calculated based on sequencing depth. The polymorphisms of rDNA sequences were identified with bam-readcount, and confirmed by Sanger sequencing and restriction enzyme experiment. The ITS2 amplicon sequencing was used to verify the stability of ITS2 haplotype analysis. ResultsDifferent from other Asteraceae species, 45S and 5S linked-type rDNA was only found in Artemisia genus. Rich polymorphisms of copy number and sequence of rDNA were identified in A. annua population. The haplotype composition of internal transcribed spacer 2 (ITS2) region which had moderate sequence polymorphism and relative short size was significantly different among A. annua strains. A population discrimination method was developed based on ITS2 haplotype analysis with high-throughput sequencing. ConclusionThis study provides comprehensive characteristics of rDNA and suggests that ITS2 haplotype analysis is ideal tool for A. annua strain identification and population genetic homogeneity assessment.

Multiple introductions, polyploidy and mixed reproductive strategies are linked to genetic diversity and structure in the most widespread invasive plant across Southern Ocean archipelagos.

Biological invasions in remote areas that experience low human activity provide unique opportunities to elucidate processes responsible for invasion success. Here we study the most widespread invasive plant species across the isolated islands of the Southern Ocean, the annual bluegrass, Poa annua. To analyse geographical variation in genome size, genetic diversity and reproductive strategies, we sampled all major sub-Antarctic archipelagos in this region and generated microsatellite data for 470 individual plants representing 31 populations. We also estimated genome sizes for a subset of individuals using flow cytometry. Occasional events of island colonization are expected to result in high genetic structure among islands, overall low genetic diversity and increased self-fertilization, but we show that this is not the case for P. annua. Microsatellite data indicated low population genetic structure and lack of isolation by distance among the sub-Antarctic archipelagos we sampled, but high population structure within each archipelago. We identified high levels of genetic diversity, low clonality and low selfing rates in sub-Antarctic P. annua populations (contrary to rates typical of continental populations). In turn, estimates of selfing declined in populations as genetic diversity increased. Additionally, we found that most P. annua individuals are probably tetraploid and that only slight variation exists in genome size across the Southern Ocean. Our findings suggest multiple independent introductions of P. annua into the sub-Antarctic, which promoted the establishment of genetically diverse populations. Despite multiple introductions, the adoption of convergent reproductive strategies (outcrossing) happened independently in each major archipelago. The combination of polyploidy and a mixed reproductive strategy probably benefited P. annua in the Southern Ocean by increasing genetic diversity and its ability to cope with the novel environmental conditions.

Inheritance and mechanism of glyphosate resistance in annual bluegrass (Poa annua L.).

In initial screening, glyphosate was ineffective in controlling five Poa annua populations. These populations were tested for resistance, and studies undertaken to determine resistance mechanisms and inheritance pattern. Dose-response studies conducted at 16/12°C and 27/20°C on the five putative resistant populations showed low-level resistance (1.4- to 2.5-fold) to glyphosate. Shikimic acid accumulation in response to glyphosate confirmed differences among the populations, with greater shikimic acid accumulation in the susceptible population. The EPSPS gene copy number was 0.5- to 5.2-fold greater in one resistant population (HT) than in the susceptible (S) population, but not in the others. EPSPS gene expression was five- to tenfold higher in HT compared with the susceptible population. Target site mutations, differences in glyphosate absorption or translocation or altered expression of aldo-keto reductase (AKR) were not identified in any of the resistant populations. Crosses were successful between one resistant population and the susceptible population (P262-16♂ ✕ S♀) and inheritance of glyphosate resistance appears to be controlled by a single, nuclear dominant gene in this population. Our study identified EPSPS gene amplification in a South Australian glyphosate-resistant P. annua population (HT). This mechanism of resistance was not identified in the other four glyphosate-resistant populations, and other common mechanisms were excluded. Although the resistance mechanism in some P. annua populations remains unknown, inheritance studies with one population suggest the involvement of a single dominant gene. © 2021 Society of Chemical Industry.

Retrotransposon-based genetic variation of Poa annua populations from contrasting climate conditions.

BackgroundPoa annua L. is an example of a plant characterized by abundant, worldwide distribution from polar to equatorial regions. Due to its high plasticity and extraordinary expansiveness, P. annua is considered an invasive species capable of occupying and surviving in a wide range of habitats including pioneer zones, areas intensively transformed by human activities, remote subarctic meadows and even the Antarctic Peninsula region.MethodsIn the present study, we evaluated the utility of inter-primer binding site (iPBS) markers for assessing the genetic variation of P. annua populations representing contrasting environments from the worldwide range of this species. The electrophoretic patterns of polymerase chain reaction products obtained for each individual were used to estimate the genetic diversity and differentiation between populations.ResultsiPBS genotyping revealed a pattern of genetic variation differentiating the six studied P. annua populations characterized by their different climatic conditions. According to the analysis of molecular variance, the greatest genetic variation was recorded among populations, whereas 41.75% was observed between individuals within populations. The results of principal coordinates analysis (PCoA) and model-based clustering analysis showed a clear subdivision of analyzed populations. According to PCoA, populations from Siberia and the Kola Peninsula were the most different from each other and showed the lowest genetic variability. The application of STRUCTURE software confirmed the unique character of the population from the Kola Peninsula.DiscussionThe lowest variability of the Siberia population suggested that it was subjected to genetic drift. However, although demographic expansion was indicated by negative values of Fu’s FS statistic and analysis of mismatch distribution, it was not followed by significant traces of a bottleneck or a founder effect. For the Antarctic population, the observed level of genetic variation was surprisingly high, despite the observed significant traces of bottleneck/founder effect following demographic expansion, and was similar to that observed in populations from Poland and the Balkans. For the Antarctic population, the multiple introduction events from different sources are considered to be responsible for such an observation. Moreover, the results of STRUCTURE and PCoA showed that the P. annua from Antarctica has the highest genetic similarity to populations from Europe.ConclusionsThe observed polymorphism should be considered as a consequence of the joint influence of external abiotic stress and the selection process. Environmental changes, due to their ability to induce transposon activation, lead to the acceleration of evolutionary processes through the production of genetic variability.

Response of Poa annua to extreme conditions: comparison of morphological traits between populations from cold and temperate climate conditions

Poa annua is an expansive species that has developed a stable breeding population on the west shore of Admiralty Bay, King George Island (Antarctica). We investigated whether the colonization success of this species in extreme climatic conditions is associated with morphological variability. We compared the differences in 12 traits among P. annua populations thriving in Admiralty Bay, Tatra Mountains and Warsaw. Our expectations that plants occurring in maritime Antarctic and mountain conditions should exhibit similar morphological characteristics were not confirmed. Comparison of individual morphometric traits indicated high variability within as well as between the studied populations. Plants from the Admiralty Bay population differed significantly from plants from the Warsaw and Tatra populations in 9 of the 12 studied traits. We discovered more similarities between the Polish populations (Warsaw and Tatra) than between the populations from harsh environments (maritime Antarctic and Tatra). The Tatra population exhibited intermediate morphological characteristics in relation to plants from the other two studied populations. In parallel, the climatic conditions expressed in mean monthly air temperature were intermediate in the Tatra location. Four traits analyzed by other authors in the sub-Antarctic populations and by us in the maritime Antarctic population were consistently lower than for the Tatra and Warsaw populations. This finding is in accordance with our working hypothesis (i.e., plants growing in harsh cold conditions exhibit similar morphological characteristics). Our results might suggest that the morphological response to environmental stress of plants occurring in mountain and polar conditions may be similar.

Historical and experimental evidence for enhanced concentration of artemesinin, a global anti-malarial treatment, with recent and projected increases in atmospheric carbon dioxide

Although the role of rising atmospheric carbon dioxide concentration [CO2] on plant growth and fecundity is widely acknowledged as important within the scientific community; less research is available regarding the impact of [CO2] on secondary plant compounds, even though such compounds can play a significant role in human health. At present, Artemisia annua, an annual plant species native to China, is widely recognized as the primary source of artemesinin used in artemesinin combination therapies or ACTs. ACTs, in turn, are used globally for the treatment of simple Plasmodium falciparum malaria, the predominant form of malaria in Africa. In this study, artemesinin concentration was quantified for multiple A. annua populations in China using a free-air CO2 enrichment (FACE) system as a function of [CO2]-induced changes both in situ and as a function of the foliar ratio of carbon to nitrogen (C:N). The high correlation between artemesinin concentration and C:N allowed an historical examination of A. annua leaves collected at 236 locations throughout China from 1905 through 2009. Both the historical and experimental data indicate that increases in artemesinin foliar concentration are likely to continue in parallel with the ongoing increase in atmospheric [CO2]. The basis for the [CO2]-induced increase in artemesinin is unclear, but could be related to the carbon: nutrient hypothesis of Bryant et al. (1983). Overall, these data provide the first evidence that historic and projected increases in atmospheric [CO2] may be associated with global changes in artemesinin chemistry, potentially allowing a greater quantity of drug available for the same area of cultivation.

Utilizing next-generation sequencing to study homeologous polymorphisms and herbicide-resistance-endowing mutations in Poa annua acetolactate synthase genes.

Detection of single nucleotide polymorphisms (SNPs) related to herbicide resistance in non-model polyploid weed species is fraught with difficulty owing to the gene duplication and lack of reference sequences. Our research seeks to overcome these obstacles by Illumina HiSeq read mapping, SNP calling and allele frequency determinations. Our focus is on the acetolactate synthase (ALS) gene, the target site of ALS-inhibiting herbicides, in Poa annua, an allotetraploid weed species originating from two diploid parents, P. supina and P. infirma. ALS contigs with complete coding regions of P. supina, P. infirma and P. annua were assembled and compared with ALS genes from other plant species. The ALS infirma-homeolog of P. annua showed higher levels of nucleotide sequence variability than the supina-homeolog. Comparisons of read mappings of P. annua and a simulated P. supina × P. infirma hybrid showed high resemblance. Two homeolog-specific primer pairs were designed and used to amplify a 1860 bp region covering all resistance-conferring codons in the ALS gene. Four P. annua populations, GN, RB, GW and LG, showed high resistance to two ALS inhibitors, bispyribac-sodium and foramsulfuron, and two populations, HD and RS, showed lower resistance in the rate-response trial. Mutations conferring Trp-574-Leu substitution were observed in the infirma-homeolog of GN and RB and in the supina-homeolog of GW and LG, but no resistance-conferring mutation was observed in the two populations of lower resistance, HD and RS. In this study we have demonstrated the use of NGS data to study homeologous polymorphisms, parentage and herbicide resistance in an allotetraploid weed species, P. annua. Complete coding sequences of the ALS gene were assembled for P. infirma, P. supina, infirma-homeolog and supina-homeolog in P. annua. A pipeline consisting of read mapping, SNP calling and allele frequency calculation was developed to study the parentage of P. annua, which provided a new perspective on this topic besides the views of morphology, karyotype and phylogeny. Our two homeolog-specific primer pairs can be utilized in future research to separate the homeologs of the ALS gene in P. annua and cover all the codons that have been reported to confer herbicide resistance.

Genetic structure along a gaseous organic pollution gradient: a case study with Poa annua L.

The population genetic composition of Poa annua L. was studied by starch electrophoresis along a transect running NE from an organic reagents factory at Shanghai, China. Five enzyme systems were stained. We have reached the following preliminary conclusions: (1) Organic pollution has dramatically changed genotypic frequencies at some loci of Poa annua populations. At polluted sites, significant deviations from Hardy–Weinberg equilibrium were observed on loci Sod-1 and Me due to the excess of heterozygote. Especially in the two nearest sites to pollution source, all the individuals were heterozygous at locus Sod-1. The data suggests that heterozygotes were more tolerant to organic pollution than homozygotes, indicating the fitness superiority of heterozygotes. (2) A tendency towards clinal changes of allele frequencies was found at some polymorphic loci. Frequencies of the common alleles at loci Sod-1, Me and Fe-1 increased as the distance to the pollution source increased. (3) The effective number of alleles per locus, and the observed and expected heterozygosity were much higher in the pollution series than in the clear control site (Botanic Park population), but genetic multiplicity (number of alleles per locus) was lower than for the control. (4) Most genetic variability was found within populations, and only 2.56% were among populations of the polluted series. However, 9.48% of the total genetic variation occurred among populations when including the Botanic Park population. The genetic identity between populations of the pollution series (0.9869–1.0000, mean 0.9941) was higher than those between the pollution series and the Botanic Park population. UPGMA divided the five populations into two groups. One contained the four polluted populations, and the other only contained the Botanic Park population.

Genetic diversity of Poa annua in western Oregon grass seed crops

The genetic diversity of Poa annua L.populations collected from western Oregon grass-seed fields was surveyed using 18 randomly amplified polymorphic DNA (RAPD) markers. Markers from 1357 individual plants from 47 populations collected at three sampling dates (fall, winter, and spring) for 16 sites were used to measure genetic diversity within and among populations. Site histories varied from low to high herbicide selection pressure, and some sites were subdivided by 3 years of differing post-harvest residue management. Gene diversity statistics, simple frequency of haplotype occurrence, and analysis of molecular variance (AMOVA) revealed the presence of significant variability in P. annua among sites, among collection dates within sites, and within collection dates. Nei gene-diversity statistics and population-differentiation parameters indicated that P. annua populations were highly diverse. Mean Nei gene diversity (h) for all 47 populations was 0.241 and total diversity (HT) was 0.245. A greater proportion of this diversity, however, was within (HS=0.209) rather than among (GST=0.146) populations. When populations were grouped by season of collection, within-group diversity was HS=0.241, while among-group diversity was GST=0.017. When populations were grouped by site, within-group diversity was HS=0.224, while among-group diversity was GST=0.087. The diversity among populations within season for fall, winter, and spring collections was GST=0.121, 0.142, and 0.133, respectively. Populations collected from fields with histories of high herbicide selection pressure showed low differentiation among collection dates, with GST as low as 0.016, whereas those collected from fields with low herbicide selection pressure showed greater differentiation among collection dates, with GST as high as 0.125. At high selection-pressure sites, populations were also lower in gene diversity (as low as h=0.155), while at low selection-pressure sites there was higher gene diversity (as high as h=0.286). The site to site variability was greater for the high selection-pressure sites (GST=0.107 or 69% of the total among-population variance), while the season of germination variability was greater at sites of low herbicide-selection pressure (GST=0.067, or 70% of the total among-population variance). High initial diversity coupled with a long-term re-supply of genotypes from the seed bank must have been factors in maintaining the genetic diversity of this weed despite the intensive use of herbicides. Knowledge of the genetic diversity of Willamette Valley P. annua should help in formulating more effective strategies for managing this weed.

Spontaneous hybridization of the putative ancestors of the allotetraploid Poa annua.

Spontaneous hybrids between the two diploids Poa infirma Kunth and P. supina Schrader were obtained at the rate of 0.2%. No reciprocal maternal effect was observed. Almost all hybrids survived, grew normally but were sterile. Hybrids were diploid, and resembled in morphology the tetraploid P. annua L. (annual meadow grass). In their isozymes, they showed additivity of the parental patterns, and were very similar to those found in P. annua populations. The ease with which hybrids were obtained and the similarity of their morphological and isozyme pattern to P. annua suggest that the two diploid species are closely related to the allotetraploid species, and are perhaps its parent species.

Immunoquantitative analysis of artemisinin from Artemisia annua using polyclonal antibodies

Artemisinin was derivatized to dihydroartemisinin carboxymethylether in three steps, without disturbing the peroxide bridge, and then linked to either thyroglobulin (TGB) or bovine serum albumin (BSA). The artemisinin-TGB and -BSA conjugates were injected in female New Zealand rabbits but only the artemisinin-TGB conjugate generated polyclonal antibodies. An enzyme-linked immunosorbent assay (ELISA) was developed and the specificity of the antibodies was confirmed by comparison with pre-immune serum and by competitive assays using different dilutions of artemisinin standards. Although anti-artemisinin antibodies cross-reacted with artemisitene and dihydroartemisinin at all dilutions used, cross-reaction with deoxyartemisinin, artemisinic acid, and arteannuin B occurred only at high concentrations. ELISA successfully detected artemisinin from crude extracts in concentrations as low as 1.5 ng ml −1; and was ε 400-fold more sensitive than the HPLC-EC. The ELISA successfully detected and quantified artemisinin in different organs of greenhouse-grown plants and in eight clones of Artemisia annua grown in tissue culture but artemisinin was overestimated owing to cross-reactivity of the antibodies with artemisinin-related compounds present in the samples. Despite overestimation of artemisinin content, the correlations between ELISA and HPLC-EC were r = 0.92 ∗∗ when samples were diluted 100 times, and r = 0.90 ∗∗ when samples were diluted 500 times, indicating that ELISA is a potential tool for screening large A. annua populations.

RAPD Characterization of Poa annua L. Populations in Golf Course Greens and Fairways

Although identical populations of Poa annua L. may colonize golf course greens and fairways, management practices may cause strong selection pressure and cause genetic shifts in adjacent populations. The objective of our research was to assess genetic differences in adjacent P. annua populations with RAPD markers. Cores were collected from greens and fairways of three golf course holes and RAPD markers used to evaluate genetic differences between the two environments. Eighty‐six primers were screened to evaluate differences between bulk samples of greens and fairways. Nine primers produced 12 RAPD markers. Five of these were used to evaluate 54 individual plant samples from each environment. Statistical evaluation indicated significant differences (P = 0.01) between the green and fairway populations for four of the RAPD markers and among holes within fairways for three markers. These differences may indicate limited gene flow between adjacent populations of P. annua.

ECOTYPIC DIFFERENTIATION IN POA ANNUA: INTERPOPULATION DIFFERENCES IN RESPONSE TO COMPETITION AND CUTTING

SUMMARYCompetition between seed‐derived populations of Poa annua from three open and three closed habitats, and Lolium perenne cv. S23 was examined in a glasshouse experiment. Pure stands of P. annua and L. perenne, and 50: 50 mixtures of the two species were compared after 13 weeks growth when uncut, and when cut at 5, 7, 9, 11 and 13 weeks, to a height of 15 cm. Yields of both the P. annua populations and L. perenne were reduced by both competition and cutting; cutting causing the greatest yield reduction. Mean yield of the three populations from open habitats and of the three populations from closed habitats were regressed on overall mean yields of the four treatment combinations ‐ pure stand uncut, mixed stand uncut, pure stand cut, mixed stand cut. Results indicated that populations from closed habitats performed better in the less favourable conditions imposed by cutting or competition or both these conditions than those from open habitats, whereas this situation was reversed in their absence. Under the cutting and competition treatment the P. annua populations from closed habitats significantly outyielded their Lolium associate. No comparable difference was found for the P. annua populations from open habitats.With competition, uncut P. annua populations from open habitats significantly outyielded Lolium whilst two of the populations from closed habitats, one adjacent to one of the open populations, were significantly outyielded by Lolium. The results suggest that population differentiation in Poa annua has occurred over short distances in response to competition or grazing or mowing and that competitive ability is not a fixed characteristic of a species.

ECOLOGICAL STUDIES ON POA ANNUA IN TURFGRASSES

Ecological studies of Poa annua populations in turf using the core sampling method conducted over a period of several years showed that Poa annua was able to produce seed under frequent mowing at 1/2 to 3/4 of an inch. The population the second winter after seeding into Bermuda Grass turf was as high as that of the first.High soil moisture maintained by frequent light irrigations favoured its persistence in Bermuda Grass turf during the summer months.Matted Bermuda Grass turf which was lightly renovated in November contained a higher winter population of Poa annua than the non‐renovated. Significantly denser populations were found in turf mowed at 1/2 to 3/4 of an inch than in turf mowed at 3 inches.Perennial prostrate growing strains were selected which could be readily propagated vegetatively and persisted in field plots for two to three years when clipped at 1/4 inch.