Studies on the diversity of the distinct phylogenetic lineage encompassing Glomus claroideum and Glomus etunicatum

Morphological and molecular characters were analysed to investigate diversity within isolates of the Glomus claroideum/Glomus etunicatum species group in the genus Glomus. The inter- and intra-isolate sequence diversity of the large subunit (LSU) rRNA gene D2 region of eight isolates of G. claroideum and G. etunicatum was studied using PCR-single strand conformational polymorphism (SSCP)-sequencing. In addition, two isolates recently obtained from Southern China were included in the analysis to allow for a wider geographic screening. Single spore DNA isolation confirmed the magnitude of gene diversity found in multispore DNA extractions. An apparent overlap of spore morphological characters was found between G. claroideum and G. etunicatum in some isolates. Analysis of the sequence frequencies in all G. etunicatum and G. claroideum isolates (ten) showed that four LSU D2 sequences, representing 32.1% of the clones analysed for multispore extraction (564) were found to be common to both species, and those sequences were the most abundant in four of the ten isolates analysed. The frequency of these sequences ranged between 23.2% and 87.5% of the clones analysed in each isolate. The implications for the use of phenotypic characters to define species in arbuscular mycorrhizal fungi are discussed. The current position of G. claroideum/G.etunicatum in the taxonomy of the Glomeromycota is also discussed.     

The effect of host mycorrhizal status on host plant–parasitic plant interactions

Two pot experiments were conducted to examine three-level interactions between host plants, mycorrhizal fungi and parasitic plants. In a greenhouse experiment, Poa annua plants were grown in the presence or absence of an AM fungus (either Glomus lamellosum V43a or G. mosseae BEG29) and in the presence or absence of a root hemiparasitic plant (Odontites vulgaris). In a laboratory experiment, mycorrhizal infection (Glomus claroideum BEG31) of Trifolium pratense host plants (mycorrhizal versus non-mycorrhizal) was combined with hemiparasite infection (Rhinanthus serotinus) of the host (parasitized versus non-parasitized). Infection with the two species of Glomus had no significant effect on the growth of P. annua, while hemiparasite infection caused a significant reduction in host biomass. Mycorrhizal status of P. annua hosts (i.e. presence/absence of AM fungus) affected neither the biomass nor the number of flowers produced by the attached O. vulgaris plants. Infection with G. claroideum BEG31 greatly increased the biomass of T. pratense, but hemiparasite infection had no effect. The hemiparasitic R. serotinus plants attached to mycorrhizal hosts had higher biomass and produced more flowers than plants growing with non-mycorrhizal hosts. Roots of T. pratense were colonized by the AM fungus to an extent independent of the presence or absence of the hemiparasite. Our results confirm earlier findings that the mycorrhizal status of a host plant can affect the performance of an attached root hemiparasite. However, improvement of the performance of the parasitic plant following attachment to a mycorrhizal host depends on the extent to which the AM fungi is able to enhance the growth of the host. Accepted: 23 February 2001     

Population studies in Goodyera (Orchidaceae) with emphasis on the hybrid origin of G. Tesselata

Morphological, cytological, and paper Chromatographic studies of populations from northern Michigan and examination of herbarium specimens from throughout North America were used to clarify the relationships ofGoodyera oblongifolia, G. repens var.ophioides, andG. tesselata. A canonical analysis of morphological data from mixed populations of these three species depictsG. tesselata as intermediate betweenG. oblongifolia andG. repens var.ophioides. The latter two species are diploid (2n = 30) andG. tesselata is tetraploid (2n = 60). Triploids (2n = ca. 45) were found in two mixed-species populations in northern Michigan.Goodyera tesselata produces three phenolic compounds present inG. oblongifolia and five different compounds present inG. repens var.ophioides. The range ofG. tesselata is confined to glaciated territory (except for two stations) in northeastern North America where the postglacially produced ranges ofG. oblongifolia andG. repens var.ophioides overlap. However,G. tesselata is quite abundant in areas outside the region of sympatry of the other two species. Based on this evidence, it is postulated thatG. tesselata is an allotetraploid species which resulted from hybridization betweenG. oblongifolia andG. repens var.ophioides during early post-Pleistocene. The slightly earlier blooming season ofG. tesselata may have been selected for to provide a measure of reproductive isolation between the tetraploid and its parents and to adapt the new species to the rather short growing season of northeastern North America.     

Polish Glomales

The morphological features of spores of Glomus pustulatum found in Poland are described and illustrated, and the worldwide occurrence of this species is characterized. Spores of G. pustulatum from Poland do not differ from those originally found in Canada and the United States of America. They were found in only one of more than 300 soil samples taken from over 100 localities in Poland. G. pustulatum was associated with roots of Ammophila arenaria colonizing maritime sand dunes of the Sowinski National Park, with a mean spore density of 12 in 100 g dry soil. Associated species of G. pustulatum were Acaulospora dilatata and Scutellospora dipurpurascens. G. pustulatum was found in Poland for the first time and is probably a species new to Europe.     

Presence of different arbuscular mycorrhizal infection patterns in roots of<Emphasis Type="Italic"> Lotus glaber</Emphasis> plants growing in the Salado River basin

Morphological types of arbuscular mycorrhizal (AM) fungi associated with Lotus glaber in sodic soils of the Salado River basin were studied. At least eight colonization patterns (IP) of AM fungi in roots of L. glaber were observed after 30 plants were analyzed. Arum- and Paris-type infection were found in the same plant species. This result supports the idea that AM morphology is not solely under plant control, but is also influenced by fungal identity. One infection pattern, presumably corresponding to Glomus intraradices, and a second, possibly assignable to Glomus tenue, were the most commonly found. Our results reinforce previous suggestions that G. intraradices is well adapted to sodic-saline conditions and may play a role in the resistance of L. glaber to these soils.     

Photonastic and thermonastic opening of capitulum in dandelion,Taraxacum officinale andTaraxacum japonicum

The capitula ofTaraxacum officinale andT. japonicum open in response to temperature rise at lower temperatures (thermonasty), and in response to light at higher temperatures (photonasty), as was the case inT. albidum. The capitula ofT. officinale could respond to the same temperature rise more sensitively than those ofT. albidum orT. japonicum. The minimum temperature for photonastic opening is as low as 13 C forT. officinale, while that forT. albidum andT. japonicum is about 18 C. That is why the capitula ofT. officinale opened earlier than those ofT. albidum andT. japonicum in the morning in April under natural conditions. The capitulum continued to be open for about 13–14 hr inT. officinale and about 8–11 hr inT. japonicum and inT. albidum both under natural conditions in April and even under constant light-temperature conditions, suggesting that the time of capitula-closing in these three species is not controlled by changes in environmental factors (light and/or temperature).     

Genetic diversity in the top onion,Allium ×proliferum (Alliaceae), analysed by isozymes

Polymorphisms for six enzyme systems (GPI, IDH, PDG, PGM, SKD, and TPI) were analysed in the top onion,Allium ×proliferum. Five multilocus isozyme genotypes were found. The banding patterns of top onions were compared with those ofA. ×wakegi, A. cepa, A. fistulosum, A. altaicum, and artificial hybrids between these three species. One top onion type and one artificial hybrid had identical banding patterns. Shallots andA. altaicum, the wild progenitor ofA. fistulosum, cannot be distinguished from the common onion andA. fistulosum, respectively; these species are also potential contributors to the top onion's gene pool.     

Arbuscular mycorrhizal fungi on adjacent semi-natural grasslands with different vegetation in Japan

Arbuscular mycorrhizal (AM) fungi on Japanese semi-natural grasslands were investigated at three adjacent sites with different vegetation. The predominant grasses at the three sites were 1)Pleioblastus chino, 2)Miscanthus sinensis andArundinella hirta (M. sinensis/A. hirta), and 3)Zoysia japonica, respectively. The degree of colonization was higher inM. sinensis/A. hirta than inP. chino andZ. japonica. AM fungi were recovered by spore extraction and by pot cultures started from soil inoculum or from transplanting of field plants. Total spore number obtained by the spore extraction method was highest in the rhizosphere ofM. sinensis/A. hirta and lowest in that ofP. chino. AGlomus sp. resemblingG. geosporum predominated in association withM. sinensis/A. hirta andP. chino. FromZ. japonica, three species,Acaulospora gerdemannii, Glomus leptotichum, and a species resemblingG. clarum, were isolated by pot culture from soil and two species,A. longula andScutellospora cerradensis, by pot culture from transplanting ofZ. japonica. FromM. sinensis/A. hirta, one species,A. longula, was found by pot culture from soil. FromP. chino, no AM fungus was detected by either method. Single-spore culture confirmed thatG. leptotichum andA. gerdemannii are conspecific.     

Detection of arbuscular mycorrhizal fungi (Glomales) in roots by nested PCR and SSCP (Single Stranded Conformation Polymorphism)

PCR amplification of a region of the large subunit ribosomal DNA sequence with Glomus specific primers was used to detect arbuscular mycorrhizal fungi in root tissue of four plant species. The primers were specific to Glomus mosseae, Glomus caledonium, Glomus geosporum, Glomus coronatum, Glomus fragilistratum and Glomus constrictum, and did not recognise sequences from Glomus claroideum. Sequence differences between isolates were detected by Single Stranded Conformation Polymorphisms (SSCPs) in polyacrylamide gels under non-denaturing conditions. Isolates of G. mosseae, G. caledonium and G. coronatum could be separated by their SSCP patterns, while three isolates of G. geosporumshowed no variation. Specific SSCP patterns from isolates of G. mosseae and G. caledonium allowed detection of both fungi in the same root segment. Sequence differences leading to variations in SSCP patterns were confirmed by direct sequencing. This revised version was published online in June 2006 with corrections to the Cover Date.     

Taxon-specific oligonucleotide primers for detection of Glomus etunicatum

The 5.8S subunit and flanking internal transcribed spacer (ITS) regions in nuclear ribosomal DNA (rDNA) from spores of Glomus etunicatum MD107, MD127, TN101, and FL329 were amplified by polymerase chain reaction (PCR) using ITS1Kpn and ITS4Pst as primers. The amplification products (597, 599, 598, and 613 bp, respectively) were cloned and sequenced. The similarity among ITS region sequences from MD107, MD127, and TN101 was 99%, whereas the sequence similarity between the ITS regions of these three DNAs and that from FL329 was 91%. The 5.8S rDNA sequences of all four G. etunicatum isolates were identical. In contrast, major dissimilarities in the corresponding rDNA sequence regions of other glomalean taxa were observed. Oligonucleotide sequences unique to G. etunicatum were tested for their specificity in PCR amplification of genomic DNA from spores of 55 isolates comprising 29 glomalean fungi: 18 isolates of G. etunicatum, five G. intraradices, three G. claroideum, 16 other Glomus isolates, and 11 other glomalean taxa from each of four other genera. The G. etunicatum isolates were from a broad range of geographic regions and soils. The oligonucleotide pair GETU1:GETU2 primed specific amplification of an oligonucleotide sequence (approximately 400 bp) present in all G. etunicatum. This primer pair did not prime PCR when template consisted of DNA from any of the other glomalean fungi or any of the non-mycorrhizal controls, including roots of corn (Zea mays). In addition, the pair successfully detected G. etunicatum in nested PCR using a primary PCR product amplified from highly diluted extracts of colonized corn roots using modified ITS1:ITS4 primers. In the phylogenetic analysis of Glomus 5.8S and ITS2 rDNA region sequences, which included 500 bootstrap data sets, confidence in the G. etunicatum branch was very strong (90%) and clearly independent of G. claroideum and G. intraradices, to which it is very closely related. Accepted: 15 October 2000     

Two Arbuscular Mycorrhizal Fungi Alleviates Drought Stress and Improves Plant Growth in Cinnamomum migao Seedlings

Cinnamomum migao plants often face different degrees of drought in karst habitats, which can lead to plants’ death, especially in the seedling stage. Widespread of arbuscular mycorrhizal (AM) fungi in karst soils have the potential to address this drought, which is a threat to C. migao seedlings. We inoculated C. migao seedlings with spores from Glomus lamellosum and Glomus etunicatum, two AM fungi widely distributed in karst soils, to observe seedling growth response after simulated drought. Our results showed that 40 g of G. lamellosum and G. etunicatum significantly promoted the growth of C. migao seedlings, 120 days after inoculation. Following a 15-day drought treatment, root colonization of the seedlings with G. lamellosum or G. etunicatum had lower the accumulation of malondialdehyde (MDA) and increased the accumulation of enzymes and osmotic substances in the seedlings. The relative water content in different organs (roots, stems, and leaves) of the drought-stressed seedlings was higher in plants with G. lamellosum or G. etunicatum than in plants without AM fungi colonization. Our results showed that inoculation with AM fungi was an effective means to improve the drought resistance of C. migao seedlings.     

Enhanced Pb Absorption by Hordeum vulgare L. and Helianthus annuus L. Plants Inoculated with an Arbuscular Mycorrhizal Fungi Consortium

The effect of an arbuscular mycorrhizal fungi (AMF) consortium conformed by (Glomus intraradices, Glomus albidum, Glomus diaphanum, and Glomus claroideum) on plant growth and absorption of Pb, Fe, Na, Ca, and ³²P in barley (Hordeum vulgare L.) and sunflower (Helianthus annuus L.) plants was evaluated. AMF-plants and controls were grown in a substrate amended with powdered Pb slag at proportions of 0, 10, 20, and 30% v/v equivalent to total Pb contents of 117; 5,337; 13,659, and 19,913 mg Pb kg^?1 substrate, respectively. Mycorrhizal root colonization values were 70, 94, 98, and 90%, for barley and 91, 97, 95, and 97%, for sunflower. AMF inoculum had positive repercussions on plant development of both crops. Mycorrhizal barley absorbed more Pb (40.4 mg Pb kg^?1) shoot dry weight than non-colonized controls (26.5 mg Pb kg^?1) when treated with a high Pb slag dosage. This increase was higher in roots than shoots (650.0 and 511.5 mg Pb kg^?1 root dry weight, respectively). A similar pattern was found in sunflower. Plants with AMF absorbed equal or lower amounts of Fe, Na and Ca than controls. H. vulgare absorbed more total P (1.0%) than H. annuus (0.9%). The arbuscular mycorrizal consortium enhanced Pb extraction by plants.     

Phylogenetic conclusions from Giemsa banding and NOR staining in top onions (Liliaceae)

The chromosomes of different strains of the top or tree onion, ofAllium cepa andA. fistulosum, as well as of cloned progenies from reciprocal crosses between these two taxa have been studied by application of Feulgen- or aceto carmine-, Giemsa- and silver staining. It was possible to differentiate between the satellite chromosomes and 2–4 other chromosome pairs ofA. cepa andA. fistulosum. The phylogenetic origin of the top onions [A. ×proliferum (Moench)Schrad.] from hybridization ofA. cepa andA. fistulosum is substantiated, taking into consideration the variability in size and position of satellites and of active NORs.     

Ultrastructure and Secretion of the Secretory Cells of Two Species ofFagoniaL. (Zygophyllaceae)

The ultrastructure and secretion of the secretory cells of theglandular trichomes ofFagonia mollisandF. glutinosawere studied.The most important finding of this study is that two speciesof the same genus produce the lipophilic component of the secretorymaterial in completely different ways and at different siteswithin the cell. In the early stages of development of secretorycells ofF. mollis,numerous mitochondria, containing myelin-likestructures, occur in the basal part of the cell. Above them,highly-elongate elements, which are suspected to develop frommitochondria with myelin-like structures, are present. Thesehave been termed ‘modified mitochondria’. It issuggested that the myelin-like structures are precursors ofthe lipophilic material ofF. mollis.InF. glutinosa,the lipophilicmaterial appears first in the plastids as plastoglobuli. Polysaccharidesappear to be produced by dictyosomes in both species. Secretionof the secretory substance to the outside of the protoplastappears to be granulocrine.Copyright 1998 Annals of Botany Company Fagonia mollis,Fagonia glutinosa,glandular trichomes, secretory cell, mitochondria, modified mitochondria, plastids, dictyosomes, lipophilic material, myelin-like structures, polysaccharides     

Morphological variation among members of theStellaria longipes complex:S. longipes,S. longifolia,andS. porsildii (Caryophyllaceae)

Morphological relationships were investigated among diploidStellaria porsildii, polyploidS. longipes, and diploidS. longifolia. Canonical discriminant analysis, based on a priori assumptions to maximize differences among groups, showed thatS. longipes clusters equally distant between the two diploid species along an axis connecting the diploids' centroids, but it differs along an axis perpendicular to this axis. The intermediacy along the former axis is evidence thatS. longipes is an amphiploid derived fromS. longifolia andS. porsildii. The divergence along the latter axis may be attributable to adaptively valuable heterotic traits which were retained following amphidiploidization. The only morphological discontinuity occurred between the two diploids, whereas the morphological range ofS. longipes overlapped the range of both diploids forming a continuum. The lack of discrete clusters is likely due to hybridization and introgression withS. longifolia on one hand, and convergence of traits betweenS. longipes var.monantha andS. porsildii on the other. High a posteriori assignments in classificatory discriminant analysis supports the separation ofS. longipes var.monantha from otherS. longipes specimens. AlthoughS. longipes var.monantha grouped close toS. porsildii, the two groups separate based on leaf shape traits. Overall results support, firstly, the hypothesis thatS. porsildii is a diploid parent species which by hybridizing withS. longifolia gave rise to polyploidS. longipes. Secondly, results suggest thatS. longipes var.monantha converged morphologically towardsS. porsildii relatively recently due to ecological specialization, and merits distinction at least as a variety ofS. longipes.     

Traits related to differences in function among three arbuscular mycorrhizal fungi

Diversity in phosphorus (P) acquisition strategies was assessed among three species of arbuscular mycorrhizal fungi (AMF) isolated from a single field in Switzerland. Medicago truncatula was used as a test plant. It was grown in a compartmented system with root and root-free zones separated by a fine mesh. Dual radioisotope labeling (³²P and ³³P) was employed in the root-free zone as follows: ³³P labeling determined hyphal P uptake from different distances from roots over the entire growth period, whereas ³²P labeling investigated hyphal P uptake close to the roots over the 48 hours immediately prior to harvest. Glomus intraradices, Glomus claroideum and Gigaspora margarita were able to take up and deliver P to the plants from maximal distances of 10, 6 and 1 cm from the roots, respectively. Glomus intraradices most rapidly colonized the available substrate and transported significant amounts of P towards the roots, but provided the same growth benefit as compared to Glomus claroideum, whose mycelium was less efficient in soil exploration and in P uptake and delivery to the roots. These differences are probably related to different carbon requirements by these different Glomus species. Gigaspora margarita provided low P benefits to the plants and formed dense mycelium networks close to the roots where P was probably transiently immobilized. Numerical modeling identified possible mechanisms underlying the observed differences in patterns of mycelium growth. High external hyphal production at the root-fungus interface together with rapid hyphal turnover were pointed out as important factors governing hyphal network development by Gigaspora, whereas nonlinearity in apical branching and hyphal anastomoses were key features for G. intraradices and G. claroideum, respectively.     

Mycorrhiza does not alter low temperature impact on <Emphasis Type="Italic">Gnaphalium norvegicum</Emphasis>

Extreme arctic-alpine vegetation has relatively low affinity to form mycorrhizal symbiosis. We asked whether the mycorrhizal growth benefit for the host plant is lower at low temperatures. We investigated the role of two root-associated fungi and temperature in growth, carbon–nitrogen relations and germination of an arctic-alpine herb. Seeds of Gnaphalium norvegicum were germinated at 8° or 15°C with or without arbuscular mycorrhizal (AM, Glomus claroideum) and dark septate endophytic (DSE, Phialocephala fortinii) inocula in a climate chamber. We found that germination percentage, shoot and root biomass, shoot N% and root AM colonization were lower at 8°C than at 15°C. P. fortinii inoculation had a positive impact on germination at both temperatures, whereas G. claroideum produced no effect. N% was lower in AM plants at both temperatures. Plant biomass and shoot N content were higher in AM plants than in control plants at 15°C, but not at 8°C. DSE inoculation tended also to have positive effects on plant biomass and N content at 15°C. At 15°C, rate of photosynthesis, photosynthetic nutrient use efficiency and specific leaf area were positively affected by G. claroideum, which suggests that G. claroideum formed a carbon sink and possibly enhanced the seedling water economy. The positive effects of P. fortinii were probably due to its saprotrophic function in the substrate because it did not colonize the roots. These results suggest that the effects of AM and DSE on plant growth are affected by temperature and that the mycorrhizal benefit for the host plant was lower at the lower temperature. Low saprotrophic activity and decreased mycorrhiza-mediated nutrient acquisition may thus constrain plant nutrient acquisition in cold environments. Decreased mycorrhizal benefit may be related to the comparatively low mycotrophy of cold environment vegetation.     

Development and activity of Glomus intraradices as affected by co-existence with Glomus claroideum in one root system

The co-existence of two arbuscular mycorrhizal fungal (AMF) species, Glomus intraradices and Glomus claroideum, in the root systems of plants was investigated in a greenhouse experiment aimed at reconstructing interactions during an early stage of primary succession on a coal-mine spoil bank in Central Europe. Two plant species, Tripleurospermum inodorum and Calamagrostis epigejos, were inoculated either with one or both AMF species. Fungal development, determined by trypan blue and alkaline phosphatase staining as well as by PCR amplification of rRNA genes with species-specific primers, and the expression of five genes with different metabolic functions in the intraradical structures of G. intraradices were followed after 6 and 9 weeks of cultivation. The two AMF closely co-existed in the root systems of both plants possibly through similar colonisation rates and competitivity. Inoculation with the two fungi, however, did not bring any additional benefit to the host plants in comparison with single inoculation; moreover, plant growth depression observed after inoculation with G. claroideum persisted also in mixed inoculation. The expression of all the assayed G. intraradices genes was affected either by host plant or by co-inoculation with G. claroideum. The effects of both factors depended on the time of sampling, which underlines the importance of addressing this topic in time-course studies.     

Contribution of the saprobic fungi Trametes versicolor and Trichoderma harzianum and the arbuscular mycorrhizal fungi Glomus deserticola and G. claroideum to arsenic tolerance of Eucalyptus globulus

The presence of high concentrations of arsenic (As) decreased the shoot and root dry weight, chlorophyll and P and Mg content of Eucalyptus globulus colonized with the arbuscular mycorrhizal (AM) fungi Glomus deserticola or G. claroideum, but these parameters were higher than in non-AM plants. As increased the percentage of AM length colonization and succinate dehydrogenase (SDH) activity in the root of E. globulus. Trichoderma harzianum, but not Trametes versicolor, increased the shoot and root dry weight, chlorophyll content, the percentage of AM root length colonization and SDH activity of E. globulus in presence of all As concentrations applied to soil when was inoculated together with G. claroideum. AM fungi increased shoot As and P concentration of E. globulus to higher level than the non-AM inoculated controls. The contribution of the AM and saprobe fungi to the translocation of As from root to shoot of E. globulus is discussed.     

Genetic mapping of the polycystic kidney gene,pcy, on mouse chromosome 9

The murine polycystic kidney disease gene,pcy, is an autosomal recessive trait located on chromosome 9. To determine the genetic locus ofpcy, 222 intraspecific backcross mice were obtained by mating C57BL/6FG-pcy andMus molossinus. Restriction fragment length polymorphism analysis of 70 of the 222 backcross progeny showed thatpcy, dilute coat color (d), and cholecystokinin (Cck) were located in the orderd—pcy—Cck from the centromere. Simple sequence repeat length polymorphism analysis of DNA of all 222 backcross mice was carried out using four markers which were located near the central regions ofd andCck. One and eight recombinations were detected betweenD9Mit24 andpcy and betweenD9Mit16 andpcy, respectively. However, no recombinant was observed amongpcy, D9Mit14, andD9Mit148. These findings strongly suggest thatD9Mit14 andD9Mit148 are located near thepcy gene and are good markers for chromosomal walking to this gene.