Pyrola Japonica Research Articles

Effects of fungicide treatments on mycorrhizal communities and carbon acquisition in themixotrophicPyrola japonica (Ericaceae).

Pyrola japonica, a member of the family Ericaceae, is a mixotroph that grows on forest floors and obtains carbon (C) from both its photosynthesis and its mycorrhizal fungi. Its mycorrhizal community is dominated by Russulaceae. However, the mechanism of its C acquisition and its flexibility are not well understood. Our aim was to assess the impact of disturbance of the mycorrhizal fungal communities on C acquisition by P. japonica. We repeatedly applied a fungicide (Benomyl) to soils around P. japonica plants in a broad-leaved forest of central Japan, in order to disturb fungal associates near roots. After fungicide treatment, P. japonica roots were collected and subjected to barcoding by next-generation sequencing, focusing on the ITS2 region. The rate of mycorrhizal formation and α-diversity did not significantly change upon fungicide treatments. Irrespective of the treatments, Russulaceae represented more than 80% of the taxa. Leaves and seeds of the plants were analysed for 13C stable isotope ratios that reflect fungal C gain. Leaf and seed δ13C values with the fungicide treatment were significantly lower than those with the other treatments. Thus the fungicide did not affect mycorrhizal communities in the roots, but disturbed mycorrhizal fungal pathways via extraradical hyphae, and resulted in a more photosynthetic behaviour of P. japonica for leaves and seeds.

Life history costs drive the evolution of mycoheterotrophs: Increased sprouting and flowering in a strongly mycoheterotrophic Pyrola species

Abstract Land plants are typically photosynthetic, but some species have lost the ability to photosynthesize, instead relying on mycorrhizal fungi to obtain carbon. Increasing levels of partial mycoheterotrophy, in which seemingly autotrophic plants receive fungal carbon from their fungal partners, and reduced sprouting in concert with greater reproduction when sprouting, may be intermediate steps in the evolution of this trait. We studied the microevolutionary demography of Pyrola japonica and the closely related species P. subaphylla, which are currently considered sister species. While these species are both partially mycoheterotrophic, P. subaphylla is more strongly so, indicating that it may be closer to evolving full mycoheterotrophy. We tracked individuals in two nearby populations in Fukushima, Japan from 2015 to 2020. We analysed vital rates, population trends and long‐run population structure with historical function‐based matrix projection models. We assessed how shifts in P. subaphylla demography relate to fitness using a stochastic life table response experiment (SLTRE). P. subaphylla exhibited strong costs of growth to survival and of reproduction to sprouting, size and fecundity. It sprouted more than P. japonica but to smaller stages. P. subaphylla's flowering frequency was approximately four times that of P. japonica, but after flowering it was smaller and more likely to die. These life history costs appeared to drive differences in fitness, measured as stochastic population growth rate. Relative to P. japonica, shifts in fitness in P. subaphylla were due more to shifts in means than standard deviations of matrix elements. Shifts in growth transitions had the strongest negative impacts on fitness but also had the strongest positive impacts. Increased shifts to small non‐flowering stages were associated with increased fitness, while transitions involving dormancy were associated more strongly with drops in fitness. Synthesis. As far as we are aware, we found the first evidence that costs of growth and reproduction drive the evolution of increased sprouting and smaller size as a species becomes more intensely mycoheterotrophic. Surprisingly, vegetative dormancy did not evolve with mycoheterotrophy. We suggest further studies assessing whether these seemingly maladaptive trends result from genetic drift, genetic linkage or another mechanism.

Mycorrhizal communities of two closely related species, Pyrola subaphylla and P. japonica, with contrasting degrees of mycoheterotrophy in a sympatric habitat.

Mycoheterotrophic plants typically form associations with a narrow range of mycorrhizal fungi. Consequently, mycorrhizal specialization is often considered to be an important step in mycoheterotrophic evolution. However, it remains unclear whether such specialization is likely to occur in plants of the genus Pyrola, which are generally associated with fungi in multiple ectomycorrhizal families. Here, we investigated the mycorrhizal communities of a nearly fully mycoheterotrophic Pyrola species (Pyrola subaphylla), a closely related partially mycoheterotrophic Pyrola species (Pyrola japonica), and a co-occurring autotrophic ectomycorrhizal tree, Quercus crispula, which is their potential carbon source, in a cool-temperate Japanese forest. High-throughput DNA sequencing revealed that numerous common ectomycorrhizal OTUs interact with the two Pyrola species and Q. crispula, thereby providing an opportunity to exploit a certain amount of carbon from common mycorrhizal networks. In addition, not only P. japonica but also P. subaphylla exhibited exceptionally high alpha mycobiont diversity, with 52 ectomycorrhizal OTUs belonging to 12 families being identified as P. subaphylla mycobionts and 69 ectomycorrhizal OTUs in 18 families being detected as P. japonica mycobionts. Nonetheless, the beta mycobiont diversity of P. subaphylla and P. japonica individuals was significantly lower than that of Q. crispula. Moreover, the beta mycobiont diversity of P. subaphylla was found to be significantly lower than that of P. japonica. Therefore, despite their seemingly broad mycorrhizal interactions, the two Pyrola species (particularly P. subaphylla) showed consistent fungal associations, suggesting that mycorrhizal specialization may have developed during the course of mycoheterotrophic evolution within the genus Pyrola.

Communities of mycorrhizal fungi in different trophic types of Asiatic Pyrola japonica sensu lato (Ericaceae).

Mixotrophic plants obtain carbon by their own photosynthetic activity and from their root-associated mycorrhizal fungi. Mixotrophy is deemed a pre-adaptation for evolution of mycoheterotrophic nutrition, where plants fully depend on fungi and lose their photosynthetic activity. The aim of this study was to clarify mycorrhizal dependency and heterotrophy level in various phenotypes of mixotrophic Pyrola japonica (Ericaceae), encompassing green individuals, rare achlorophyllous variants (albinos) and a form with minute leaves, P. japonica f. subaphylla. These three phenotypes were collected in two Japanese forests. Phylogenetic analysis of both plants and mycorrhizal fungi was conducted based on DNA barcoding. Enrichment in 13C among organs (leaves, stems and roots) of the phenotypes with reference plants and fungal fruitbodies were compared by measuring stable carbon isotopic ratio. All plants were placed in the same clade, with f. subaphylla as a separate subclade. Leaf 13C abundances of albinos were congruent with a fully mycoheterotrophic nutrition, suggesting that green P. japonica leaves are 36.8% heterotrophic, while rhizomes are 74.0% heterotrophic. There were no significant differences in δ13C values among organs in both albino P. japonica and P. japonica f. subaphylla, suggesting full and high mycoheterotrophic nutrition, respectively. Among 55 molecular operational taxonomic units (OTUs) detected as symbionts, the genus Russula was the most abundant in each phenotype and its dominance was significantly higher in albino P. japonica and P. japonica f. subaphylla. Russula spp. detected in P. japonica f. subaphylla showed higher dissimilarity with other phenotypes. These results suggest that P. japonica sensu lato is prone to evolve mycoheterotrophic variants, in a process that changes its mycorrhizal preferences, especially towards the genus Russula for which this species has a marked preference.

Effects of Pyrola japonica Extracts on Osteoclast Differentiation and Bone Resorption

Effects of Pyrola japonica Extracts on Osteoclast Differentiation and Bone Resorption

Development of microsatellite markers for partially and putative fully mycoheterotrophic varieties of Pyrola japonica sensu lato (Ericaceae).

We developed microsatellite markers to compare the genetic variation and reproductive biology between the partially mycoheterotrophic Pyrola japonica var. japonica and the putative fully mycoheterotrophic P. japonica var. subaphylla. Fifteen primer pairs were developed for P. japonica sensu lato and they were tested on 77 ramets from three populations of the two varieties. Thirteen loci were polymorphic in at least one of the two var. japonica populations, whereas only four loci were polymorphic in the var. subaphylla population. The considerably lower genetic variation of the var. subaphylla population may be attributed to frequent selfing and/or inbreeding. The markers developed in this study will be useful for comparing the genetic diversity of P. japonica s. l. populations and measuring gene flow within and between populations and varieties.

Effect of antioxidant activity of Pyrola japonica on H2O2-induced oxidative stress in HSEC cells

Effect of antioxidant activity of Pyrola japonica on H2O2-induced oxidative stress in HSEC cells

Variation in vegetative morphology tracks the complex genetic diversification of the mycoheterotrophic species Pyrola japonica sensu lato

Although the evolution of full mycoheterotrophy has attracted many plant researchers, molecular phylogenetic studies that focus on the transition from partial to full mycoheterotrophy are limited to a few taxa. Pyrola japonica sensu lato is an ideal model for examining the evolution of mycoheterotrophy, owing to its variable leaf size, which suggests that the species comprises several transitional stages. To elucidate the molecular and morphological changes that occur during the evolutionary transition between partial and full mycoheterotrophy in P. japonica s.l. from 18 populations in Japan, we estimated a parsimony network of plastid haplotypes based on three noncoding regions, measured the leaf size and scape color of the shoots, and compared morphology among haplotypes. The seven haplotypes exhibited star-like relationships, and at least three divergent haplotypes were associated with differences in morphology. The first was mainly observed in large-leaved and green-scaped populations, whereas the second was observed in extremely small-leaved and reddish-scaped populations, which indicated a high degree of mycoheterotrophy, and the last was detected among mixed populations with both green- and reddish-scaped shoots with intermediate leaf sizes. In addition, the inconsistent association between the haplotypes and morphology suggests a complex relationship. Pyrola japonica s.l. has at least three separate genetic lineages that have different leaf morphologies. The genetic lineages and their coexistence could have led to the variable leaf size and suggest the possibility that gene flow from partial to full mycoheterotrophs could reverse the evolutionary transition to full mycoheterotrophy.

Pyrola japonica, a partially mycoheterotrophic Ericaceae, has mycorrhizal preference for russulacean fungi in central Japan

Mycorrhizal symbiosis often displays low specificity, except for mycoheterotrophic plants that obtain carbon from their mycorrhizal fungi and often have higher specificity to certain fungal taxa. Partially mycoheterotrophic (or mixotrophic, MX) plant species tend to have a larger diversity of fungal partners, e.g., in the genus Pyrola (Monotropoideae, Ericaceae). Preliminary evidence however showed that the Japanese Pyrola japonica has preference for russulacean fungi based on direct sequencing of the fungal internal transcribed spacer (ITS) region from a single site. The present study challenges this conclusion using (1) sampling of P. japonica in different Japanese regions and forest types and (2) fungal identification by ITS cloning. Plants were sampled from eight sites in three regions, in one of which the fungal community on tree ectomycorrhizal (ECM) tips surrounding P. japonica was also analyzed. In all, 1512 clone sequences were obtained successfully from 35 P. japonica plants and 137 sequences from ECM communities. These sequences were collectively divided into 74 molecular operational taxonomic units (MOTUs) (51 and 33 MOTUs, respectively). MOTUs from P. japonica involved 36 ECM taxa (96% of all clones), and 17 of these were Russula spp. (76.2% of all clones), which colonized 33 of the 35 sampled plants. The MOTU composition significantly differed between P. japonica and ECM tips, although shared species represented 26.3% of the ECM tips community in abundance. This suggests that P. japonica has a preference for russulacean fungi.

Seasonal and environmental changes of mycorrhizal associations and heterotrophy levels in mixotrophicPyrola japonica(Ericaceae) growing under different light environments

Mixotrophy is a strategy whereby plants acquire carbon both through photosynthesis and heterotrophic exploitation of mycorrhizal fungi. In Euro-American Pyroleae species studied hitherto, heterotrophy levels vary according to species, sites of study, and possibly light conditions. We investigated mycorrhizal association and mixotrophy in the Asiatic forest species Pyrola japonica, and their plasticity under different light conditions. Pyrola japonica was sampled bimonthly in sunny and shaded conditions from a deciduous broadleaf forest. We microscopically assessed the rate of fungal colonization and sequenced the ITS to identify the mycorrhizal fungi. We measured (13)C and (15)N isotopic abundances in P. japonica as compared with neighboring autotrophic and mycoheterotrophic plants, to evaluate P. japonica's heterotrophy level. Pyrola japonica formed arbutoid mycorrhizas devoid of fungal mantles, with intracellular hyphal coils and a Hartig net. It tended to be more colonized by mycorrhizal fungi in spring and summer. Most associated fungi belonged to ectomycorrhizal taxa, and 84% of identified fungi were Russula spp. Rate of mycorrhizal colonization and Russula frequency tended to be higher in shaded conditions. Both δ(13)C and δ(15)N values of P. japonica were significantly higher in autotrophic plants, showing that about half of the carbon on average was received from mycorrhizal fungi. Both isotopic values negatively correlated with light availability, suggesting higher heterotrophy levels in shaded conditions. The mixotrophic P. japonica undergoes changes in mycorrhizal symbionts and carbon nutrition according to light availability. Our results suggest that during Pyroleae evolution, a tendency to increased heterotrophy emerged in the Pyrola/Orthilia clade.

노루발풀(Pyrola japonica) 추출물의 Bacillus subtilis에 대한 항균활성

우리나라 전국 산지에서 쉽게 구할 수 있는 노루발풀을 건조시켜 80% ethanol 추출물과 여러 가지 용매 분획물의 미생물 증식 억제 효과에 대해서 검색하였다. 노루발풀의 80% ethanol 추출물은 그람 양성균 B. subtilis에서 농도 의존적으로 항균활성을 나타내는 반면 그람 음성균 E. coli에서는 활성이 나타나지 않았다. 노루발풀의 80% ethanol 추출물을 용매 분획하여 얻은 분획물 중 hexane층에서 항균활성이 나타나는 것을 확인하였다. 노루발풀의 항균활성 물질을 검색하기 위한 목적으로 hexane 분획물을 silica gel column chromatography, prep. TLC 및 HPLC를 실시하였으며 그 결과, peak I의 단일 물질이 노루발풀의 B. subtilis에 대한 항균활성을 나타내는 물질임을 확인할 수 있었다. The antimicrobial substance from Pyrola japonica were extracted and isolated. Eighty percent ethanol extract of dried Pyrola japonica was fractionated to hexane, diethyl ether, ethyl acetate and aqueous layer. The hexane-soluble fraction showed the highest inhibitory activity against Bacillus subtilis. Moreover the hexane layer was fractionated into 5 groups by silica gel column chromatography. From the results, group No. 2 (<TEX>$18{\sim}40$</TEX> fractions) showed the highest antimicrobial activity. The group was re-separated to 10 fractions by preparative thin layer chromatography and the peak I as active fraction was isolated by HPLC.

Phenolic Glycosides from Pyrola japonica.

AbstractFor Abstract see ChemInform Abstract in Full Text.

Phenolic glycosides from Pyrola japonica.

Five new phenolic glycosides, 2-beta-D-glucopyranosyloxy-5-hydroxyphenylacetic acid methyl ester (4), 4-hydroxy-2-[3-hydroxy-3-methylbutyl]-5-methylphenyl beta-D-glucopyranoside (5), 4-hydroxy-2-[(E)-4-hydroxy-3-methyl-2-butenyl]-5-methylphenyl beta-D-glucopyranoside (7), 4-hydroxy-2-[(2E,6Z)-8-beta-D-glucopyranosyloxy-3,7-dimethylocta-2,6-dien-1-yl]-5-methylphenyl beta-D-glucopyranoside (8), and 2,7-dimethyl-1,4-dihydronaphthalene-5,8-diol 5-O-beta-D-xylopyranosyl(1-->6)-beta-D-glucopyranoside (10), were isolated from the whole plants of Pyrola japonica (Pyrolaceae), together with androsin, (-)-syringaresinol glucoside, homoarbutin, pirolatin, hyperin, monotropein and chimaphilin.

A new naphthoquinone from Pyrola japonica.

A new naphthoquinone, 5,8-dihydro-2,7-dimethyl-[1,4]naphthoquinone (1), which was named 5,8-dihydrochimaphilin, isolated from an ethyl acetate soluble fraction from the root of Pyrola japonica, together with chimaphilin (2). Compound 1 was transformed rapidly to 2 upon exposure to air by HPLC analysis. This fact supported that chimaphilin (2) may be an artifact from 1.

ALLOZYME GENETIC AND CLONAL DIVERSITY WITHIN POPULATIONS OF CHIMAPHILA JAPONICA AND PYROLA JAPONICA (PYROLACEAE)

Chimapliila japonica Miq., a rhizomatous evergreen subshrub, and Pyrola japonica Klenze, an evergreen herbaceous perennial, occur mainly in conifer forests in East Asia. The levels of genetic variability within populations of C. japonica are fairly low, whereas those of P. japonica are less so. Mean percentage of polymorphic loci within populations (P p), mean number of alleles per locus across populations (A p), and mean population genetic diversity (He p) were 14.8%, 1.15, and 0.030 for C. japonica and 18.4%, 1.35, and 0.087 for P. japonica. The degree of population differentiation observed in C. japonica (mean G ST = 0.028) and P. japonica (mean G ST = 0.147) was lower than that in other plants with similar life history traits. Considering the habit and pollen and seed dispersal mechanisms, it is suggested that the low G ST of C. japonica is caused by unifying selection. For C. japonica, the mean number of multilocus genotypes per population sample (G/N = 0.19) and mean genotypic diversity index (DG = 0.69) were relatively low. On the other hand, relatively high values of clonal diversity were observed in P. japonica (G/N = 0.39 and DG = 0.93). For C. japonica and P. japonica, 14% (3/21) and 6% (5/81) of multilocus genotypes were “widespread genotypes” (genotypes occurring in more than 75% of the populations), whereas the proportions of “local genotypes” (genotypes occurring in only one population) were 33% (7/21) and 40% (22/81). About 50% and 20% of the total samples examined were homozygous multilocus-genotypes for C. japonica and P. japonica, respectively. Relatively high numbers of these genotypes and low levels of clonal diversity in C. japonica are responsible for the low levels of expected heterozygosity within populations of the species. The results indicate that, for C. japonica, clonal reproduction may act as an enhancer of genetic drift by reducing the effective size of local populations.

Light environment and carbon gain of understory herbs associated with sunflecks in a warm temperate deciduous forest in Japan

AbstractSeasonal variation in the light environment on the forest floor of a deciduous forest was investigated with special reference to sunflecks. Diurnal variations and seasonal changes in frequency and irradiation period of the sunflecks (sunfleck duration) were measured. The hourly total sunfleck duration varied seasonally; that is, 30–40 min in spring and autumn and about 15–20 min in summer. There was no large variation in the hourly sunfleck duration during daytime hours (from 9.00 to 15.00 h). The emergence frequency of sunflecks was 1.3–4.8 per h with two peaks, one in the morning and one in the afternoon. The mean duration of a sunfleck, however, showed a characteristic daily pattern with a peak around noon. Sunfleck duration was long around noon, ranging from 12 to 18 min, and short around 10.00 and 14.00 h, ranging from 6 to 10 min. Using the light photosynthesis curves of Pyrola japonica and Syneilesis palmata (Koizumi &amp; Oshima 1985), the contribution of sunflecks to the dry matter production of these understory species was evaluated. It was shown that the sunflecks contributed 7–10% of the carbon gain in S. palmata, but only 2–3% of that in P. japonica.

Studies on the life history of the population of an evergreen herb,Pyrola japonica, on the forest floor in a warm temperate region

Photosynthetic and respiratory activities and gross production in relation to temperature conditions were investigated in the population of an evergreen herb,Pyrola japonica, growing on the floor of a deciduous forest in the warm temperate region of central Japan. Analysis of the temperature-photosynthesis relationship ofP. japonica leaves during the growing season indicated distinct seasonal changes in the temperature optimum for photosynthesis. This population was found to be acclimatable to ambient air temperatures exceeding 15C, but this acclimation became less pronounced under thermal conditions below 15 C. This plant possessed narrow photosynthetic optima in the warm season but wide optima in the cold season. The shape of the temperature-respiration curve did not vary significantly with the months except for April. The Q10 for respiration between 10 C and 20 C was calculated to be 1.93–2.65. Annual dry matter loss associated with respiration was estimated to amount to 159.1 g d.w.m−2 based on the measurements of the seasonal changes in the respiratory activity of each organ. Gross production of this population was estimated to be 219.3 g d.w.m−2 year−1 as the sum total of the net production (60.2 g d.w.m−2year−1) and the respiration. Monthly gross production was high in the early growing season, and low and stable in winter.

Studies on the life history of an evergreen herb,Pyrola japonica, population on a forest floor in a warm temperate region

The life history and matter economy were studied on an evergreen herb,Pyrola japonica Klenze, population growing on the floor of a deciduous forest in a warm temperate region, central Japan. Seasonal changes in standing crop, bulk density and reserve substance of each organ and in leaf area were investigated over a year. Monthly and annual net production of the population were estimated based on the growth in dry weight and the seasonal dynamics of reserve substance. Seasonal peak value of the monthly net production was 16.7 g d.w.m−2 in May. The annual net production was estimated to be 60.2 g d.w.m−2. Two phases were recognized in the annual pattern of development and matter economy of this plant. Phase I, from April to June, was characterized by the development of aboveground vegetative organs and the consumption of reserve substance rrom old organs. Phase II, from July to March, was characterized by the development of reproductive and underground organs, and the accumulation of reserve substance. Phase II was further divided into two sub-phases according to the behavior of the reserve substance and the rate of net production. The production process of the population was compared with those of other evergreen herbs growing on the forest floor in warm temperate regions.

Structure of renifolin and reconfirmation of the structure of pirolatin

The 1H and 13C NMR spectral analysis of the glucoside renifolin, isolated from Pyrola renifolia, demonstrated the so far unknown binding site of the glucose moiety to be at C-8 and hence its structure as 8-β- D-glucosyloxy-2,7-dimethyl-1,4-dihydronaphthalen-5-ol. The earlier reported structure for the glucoside pirolatin, isolated from Pyrola japonica, was also reconfirmed by the 13C NMR spectral analysis.

The oxidation of arbutin by isolated chloroplasts of arbutin-containing plants

A new explanation is proposed to explain the blackening of Pyrus leaves. The phenolic glucoside arbutin present in the leaves is oxidized directly by enzymes in the chloroplasts, but not present in chromoplasts, leucoplasts or cytoplasm. This enzyme is also present in the chloroplasts of other arbutin-containing plants, for instance, Pyrola japonica and Saxifraga stolonifera. The oxidation produces successively six chromatographically-detected glucosides by two independent pathways. One of the first products is shown to be 3,4-dihydroxyphenyl-β- D-glucoside. The two independent pathways are controlled by two different parts of chloroplasts ; the one being a protein fraction dissociable with cholate, while the other is firmly bound to the structure of the chloroplasts.