Conservative ecological and evolutionary patterns in liverwort–fungal symbioses

Liverworts, the most ancient group of land plants, form a range of intimate associations with fungi that may be analogous to the mycorrhizas of vascular plants. Most thalloid liverworts contain arbuscular mycorrhizal glomeromycete fungi similar to most vascular plants. In contrast, a range of leafy liverwort genera and one simple thalloid liverwort family (the Aneuraceae) have switched to basidiomycete fungi. These liverwort switches away from glomeromycete fungi may be expected to parallel switches undergone by vascular plants that target diverse lineages of basidiomycete fungi to form ectomycorrhizas. To test this hypothesis, we used a cultivation-independent approach to examine the basidiomycete fungi associated with liverworts in varied worldwide locations by generating fungal DNA sequence data from over 200 field collections of over 30 species. Here we show that eight leafy liverwort genera predominantly and consistently associate with members of the Sebacina vermifera species complex and that Aneuraceae thalloid liverworts associate nearly exclusively with Tulasnella species. Furthermore, within sites where multiple liverwort species co-occur, they almost never share the same fungi. Our analyses reveal a strikingly conservative ecological and evolutionary pattern of liverwort symbioses with basidiomycete fungi that is unlike that of vascular plant mycorrhizas.     

Soluble carbohydrates of liverworts

Five soluble carbohydrates, (−)-l-bornesitol, (−)-d-mannitol, (+)-d-glucose, (+)-sucrose and (+)-trehalose, have been isolated as their acetyl derivatives from the liverwort, Mylia taylorii. The absolute configuration of these carbohydrates is the same as that of sugars found in vascular plants, although the liverwort sesquiterpenoids are usually enantiomeric relative to those of vascular plants. The soluble carbohydrates in seven other liverwort species have also been analysed by HPLC.     

Group I introns in the liverwort mitochondrial genome: the gene coding for subunit 1 of cytochrome oxidase shares five intron positions with its fungal counterparts.

The complete nucleotide sequence of the mitochondrial DNA (mtDNA) from a liverwort, Marchantia polymorpha, contains thirty-two introns. Twenty-five of these introns possess the characteristic secondary structures and consensus sequences of group II introns. The remaining seven are group I introns, six of which happen to interrupt the gene coding for subunit 1 of cytochrome oxidase (cox1). Interestingly, the insertion sites of one group II and four group I introns in the cox1 gene coincide with those of the respective fungal mitochondrial interns. Moreover, comparison of the four group I introns with their fungal counterparts shows that group I introns inserted at identical genomic sites in different organisms are indeed related to one another, in terms of the peptide sequences generated from the complete or fragmental ORFs encoded by these introns. At the same time, the liverwort introns turned out to be more divergent from their fungal cognates than the latter are from one another. We therefore conclude that vertical transmission from a common ancestor organism is the simplest explanation for the presence of cognate introns in liverwort and fungal mitochondrial genomes.     

咸海湖泊退缩对岸边土壤真菌和植物内生真菌的影响研究

[目的] 研究咸海岸边不同暴露时期土壤带的土壤真菌和植物内生真菌群落构成及其对湖泊干涸的响应。[方法] 从咸海湖岸远端（土壤带的暴露时间最长）到湖岸近端（土壤带的暴露时间最短）的不同土壤带采集土壤样品,对其进行地球化学和矿物学分析。同时也采集各土壤带的土壤样品和优势植物,通过ITS基因高通量测序方法分析土壤真菌和植物内生真菌群落构成,进而探讨其如何响应湖泊干涸（如盐度升高、矿物组分变化、植物种类丰富度变化等）过程。[结果] 持续暴露的咸海湖床从湖泊远岸到湖泊近岸形成了一个连续的盐度梯度：E48（暴露于1970年之前,总可溶解盐0.5±0.5 g/L）;E38（暴露于1980年之前,总可溶解盐0.4±0.2 g/L）;E28（暴露于1990年之前,总可溶解盐23.3±2.1 g/L）;E18（暴露于2000年之前,总可溶解盐23.7±7.5 g/L）;E9（暴露于2009年之前,总可溶解盐71.3±6.1 g/L）;E1（暴露于2017年之前,总可溶解盐62.9±10.7 g/L）和E0（2018年湖岸线附近沉积物样品,总可溶解盐69.9±8.3 g/L）。咸海岸边不同土壤带分布着不同的植物：梭梭（Haloxylon ammodendron）在E48和E38区域中占优势地位;滨藜（Chenopodium album）在E28、E18和E9区域占优势;而在E1和E0区域无可见植物物种分布。另外,咸海岸边不同土壤带的主要矿物成分也存在差异：粘土矿物和蒸发岩的含量从咸海湖岸远端到湖岸近端逐渐增加,而碳酸盐矿物含量逐渐减少。咸海岸边不同土壤带土壤样品优势真菌类群（>5%）为散囊菌纲（Eurotiomycetes）、粪壳菌纲（Sordariomycetes）、锤舌菌纲（Leotiomycetes）、座囊菌纲（Dothideomycetes）、黑粉菌亚门（Ustilaginomycotina）和伞菌纲（Agaricomycetes）,且按植物种类丰富度进行聚类。而植物样品优势真菌类群为未知真菌门类（>97.8%）,且按植物种类进行聚类。线性回归结果显示,咸海岸边不同土壤带土壤样品真菌群落差异性与暴露时间距离具有显著相关（R²=0.32,P<0.05）,而与总可溶解盐差异则无明显相关性。而植物内生真菌群落差异性与暴露时间距离/总可溶解盐差异之间均无显著相关。Mantel检验结果显示,咸海岸边不同土壤带土壤真菌群落与植物种类丰富度和矿物成分组成（如白云石、方解石、微斜长石和石膏）呈显著相关（P<0.05）,其中植物种类丰富度和方解石含量的相关性系数最大;植物内生真菌群落与方解石含量之间呈显著相关（P<0.05）。[结论] 咸海岸边不同暴露时期土壤带的土壤真菌和植物内生真菌种群结构具有时空差异,与植物种类丰富度和特定矿物组成相关,而与总可溶解盐无显著相关。     

Identification of a novel nifH-like (frxC) protein in chloroplasts of the liverwort Marchantia polymorpha

The frxC gene, one of the unidentified open reading frames present in liverwort chloroplast DNA, shows significant homology with the nifH genes coding for the Fe protein, a component of the nitrogenase complex (Ohyama et al., 1986, Nature 322: 572–574). A truncated form of the frxC gene was designed to be over-expressed in Escherichia coli and an antibody against this protein was prepared using the purified product as an antigen. This antibody reacted with a protein in the soluble fraction of liverwort chloroplasts, which had an apparent molecular weight of 31 000, as revealed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, in good agreement with a putative molecular weight of 31945 deduced from the DNA sequence of the frxC gene. In a competitive inhibition experiment, the antigenicity of this protein was indicated to be similar to that of the over-expressed protein in E. coli. Therefore, we concluded that the frxC gene was expressed in liverwort chloroplasts and that its product existed in a soluble form. The molecular weight of the frxC protein was approximately 67 000, as estimated by gel filtration chromatography, indicating that the frxC protein may exist as a dimer of two identical polypeptides analogous to the Fe protein of nitrogenase. The results obtained from affinity chromatography supported the possibility that the frxC protein, which possesses a ATP-binding sequence in its N-terminal region that is conserved among various other ATP-binding proteins, has the ability to bind ATP.     

Sebacinales are associates of the leafy liverwort Lophozia excisa in the southern maritime Antarctic

The leafy liverwort Lophozia excisa, which is colonised by basidiomycete fungi in other biomes and which evidence suggests may be colonised by mycorrhizal fungi in Antarctica, was sampled from Léonie Island in the southern maritime Antarctic (67°36′ S, 68°21′ W). Microscopic examination of plants indicated that fungal hyphae colonised 78% of the rhizoids of the liverwort, apparently by entering the tips of rhizoids prior to growing into their bases, where they formed hyphal coils. Extensive colonisation of stem medullary cells by hyphae was also observed. DNA was extracted from surface-sterilised liverwort tissues and sequenced following nested PCR, using the primer set ITS1F/TW14, followed by a second round of amplification using the ITSSeb3/TW13 primer set. Neighbour-joining analyses showed that the sequences obtained nested in Sebacinales clade B as a 100% supported sister group to Sebacinales sequences from the leafy liverworts Lophozia sudetica, L. incisa and Calypogeia muelleriana sampled from Europe. Direct PCR using the fungal specific primer set ITS1F/ITS4 similarly identified fungi belonging to Sebacinales clade B as the principal colonists of L. excisa tissues. These observations indicate the presence of a second mycothallus in Antarctica and support the previous suggestion that the Sebacinales has a wide geographical distribution.     

Regeneration capacity of oceanic-montane liverworts: implications for community distribution and conservation

Abstract

Scotland’s mountains are home to a rare and unique liverwort community, ‘the oceanic-montane liverwort-rich heath’, but its component species are absent from regions where they could potentially thrive. Many biological characteristics of these species are unknown, making it impossible to explain the reasons for their rarity; however, they have not been observed to produce sporophytes within Britain. We use ex situ cultivation of whole liverworts and fragments, and in situ cultivation of fragments, to assess the growth rate and the potential for vegetative reproduction of several species. Most of the species grew from both fragments and as whole plants, indicating that the rarity of the liverwort heath is not due to poor powers of regeneration. We propose that growth rate and the potential to regenerate from fragments are important factors structuring the liverwort heath community, at least locally. Furthermore, this study demonstrates that there is potential for ex situ conservation of rare liverwort species, in situ enhancement of existing populations, and creation of new ones.     

Airborne sperm of Conocephalum conicum (Conocephalaceae)

We have obtained the first momentary photographs of sperms just as they are discharged from the antheridium of a liverwort, Conocephalum conicum, and have succeeded in monitoring the airborne sperms of bryophytes under field conditions. Airborne sperm of liverworts seems to be an effective strategy for raising the efficiency of fertilization between male and female plants separated in a drought environment. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Discovery of bacteria in the cell wall of the aquatic liverwort Jungermannia vulcanicola Steph. in an acid stream with pH 4.2-4.6

Abstract

T.E.M.s of the leaves of the aquatic liverwort Jungermannia vulcanieola showed the existence of rod-shaped bacteria and numerous holes in the cell walls. This suggests that bacteria playa role in the decomposition of the liverwort in an aquatic ecosystem.     

Culture studies on Radula flaccida Lindenb. & Gottsche

Abstract

Radula flaccida has been grown on defined inorganic media. Its growth varies with the type and concentration of the media, lower concentrations proving better than higher ones. Basile's medium supported a better growth of the liverwort than that of Diller et al., Müller's medium being least effective. The liverwort was successfully established on leaves and bark of some higher plants, thus proving that it is not ‘host’ specific with respect to leaf types and that it is not exclusively epiphyllous. Its less frequent occurrence on bark may be due to other habitat factors.     

Athalamia hyalina (Sommerf.) S.Hatt. in Scotland,new to the British Isles

Abstract

The liverwort Athalamia hyalina (Sommerf.) S.Hatt. and the family Cleveaceae are reported as new to the British Isles from the eastern Scottish Highlands. The Scottish plants are described and illustrated and the significance of the discovery is assessed. In Scotland, Athalamia hyalina occurs on thin soil on eroding limestone ledges in a montane habitat with a continental climate. It is in active growth in the winter months, producing spores in spring. Its range is restricted but it occurs within a protected area and is not threatened at present. The species is widespread in the northern hemisphere.     

Phytogeography of the liverwort flora of the Atlantic Forest of southeastern Brazil

Abstract

This work analyzes the phytogeographic patterns of the liverwort flora of the Atlantic Forest of the Rio de Janeiro State, south-eastern Brazil. The analysis was based on inventories made in fragments of Atlantic Forest in the state, collections from the RB herbarium, and information from the checklist and database of the bryoflora of Rio de Janeiro. The phytogeographic pattern for each taxon was characterized based on its current distribution. Three hundred and sixty liverwort taxa are recognized for Rio de Janeiro. The liverwort flora varies along an altitudinal gradient within the Atlantic Forest, with the montane belt having the greatest species richness (238), the highest number of exclusive taxa (63) and the greatest number of endemic species (23). The predominant phytogeographic element is of neotropical species (49%), followed by disjunct liverworts (18%). Forty-one taxa (11%) are endemic to the country, of which 34 are restricted to the Atlantic Forest. In the lowland and submontane formations species are wide-ranging, whereas the montane and upper montane formations are characterized by endemic species or those disjunct with the Andes. The liverwort flora emphasizes the importance of the fragments of Atlantic Forest in Rio de Janeiro as a centre of diversity and endemism, supporting 50% of the total liverwort species known in Brazil, 72% of those recorded from Atlantic Forest and 55% of the endemic species of the country. Species with an Afro-American and Andean disjunction make up a characteristic part of the liverwort flora, probably reflecting the effects of long-distance dispersal by air currents, migration over land before the breakup of the continents and climatic similarities between the high-altitude grasslands and the northern Andes.     

Fungal cerebral abscess in a diabetic patient successfully treated with surgery followed by prolonged antifungal therapy

BackgroundIntracranial fungal masses are uncommon diseases, but their incidence is increasing, most often due to the prolonged survival of patients with different immunodeficiencies. The management of patients with intracranial fungal masses included stereotactic biopsy for diagnosis, partial or radical surgery excision and prolonged antifungal therapy.AimsWe report the case of a 51-year-old diabetic man with a history of psoas abscess due to Candida albicans 1 year before the onset of neurological symptoms, including headache and generalized tonoclonic seizures.MethodsMagnetic resonance imaging showed a single lesion located in the right parietal lobe with mass effect, surrounding edema and enhancement after injection of gadolinium. The material was purulent.ResultsDirect microscopic examination showed hyaline, branched and septate hyphae compatible with fungal elements.ConclusionsFungal infections, especially due to Candida species, should be considered in diabetic patients with parenchymal brain abscesses. Radical excision followed by prolonged antifungal therapy based on fluconazole or amphotericin B is necessary to improve the prognosis of this type of patients.     

Aerial and underground biomasses of three Sphagnum-dwelling liverworts

Abstract

The few studies that have investigated levels of genetic variation in liverworts have found very little polymorphism. Our electrophoretic data show, however, that the leafy liverwort Porella platyphylla maintains high levels of genetic variation in at least some natural populations from the southeastern United States. Within a single population from southwestern North Carolina, we detected 26 distinct multilocus genotypes and more than 80% of the enzyme loci we surveyed were polymorphic. It seems likely that earlier studies of mostly thalloid species from glaciated regions of Europe have presented a biased picture of levels of variation in liverwort populations.     

The Habitat of Zygodon forsteri (Brid) Mitt. in the New Forest,Hants

Abstract

Seven species are reported here for the first time for South Georgia. Two of these were already known from Tierra del Fuego, Acrolophozia fuegiana Schust. and Marsupella sprucei (Limpr.) H. Bern., but Acrolophozia sulcata Hässel, Marsupella austrogeorgica Hässel, and Marsupella minutula Hässel are described as new. The additional two species which are described, Austrolembidium crassum Hässel and Greeneothallus gemmiparus Hässel, also required the creation of two new genera. The total number of liverwort species now known to be present on South Georgia is sixty-six.     

Homologues of the green algal gidA gene and the liverwort frxC gene are present on the chloroplast genomes of conifers

Strong hybridization signals were obtained from total DNA of two conifers, lodgepole pine (Pinus contorta) and Norway spruce (Picea abies), in a Southern blot analysis using a probe derived from the chloroplast gidA gene of the green alga Chlamydomonas reinhardtii. The pine fragments detected by the probe were found to originate from the chloroplast genome and, as judged by the signal intensity, this was also true for the spruce fragments. Sequence analysis of the hybridizing pine chloroplast DNA region revealed an open reading frame potentially encoding a 459 amino acid polypeptide, highly homologous to that deduced from the algal gene and to ORF465 of liverwort chloroplast DNA. Upstream of the gidA sequence, we found a trnN(GUU) gene and an open reading frame of 291 codons which was 78% identical to the frxC gene of liverwort. Since ORF465 is located immediately downstream of trnN and frxC in liverwort, the genetic organization of this region is very similar in the two plants. In contrast, neither the gidA nor the frxC gene is present in the chloroplast DNA of tobacco or rice. It was recently reported that deletions in the gidA region of the chloroplast genome of Chlamydomonas reinhardtii abolish the light-independent pathway of chlorophyll synthesis which exists in many algae and lower plants. The presence of the gidA gene on the chloroplast genomes of conifers may therefore be of significance with respect to the ability of these plants to synthesize chlorophyll in the dark.     

The occurrence of axillary antheridia in dioecious mosses

Abstract

Antheridia that are undoubtedly axillary occur in species of Bryum, Mnium and Aulacomnium. It is suggested that this is a primitive feature and that the typical moss antheridial head, with all the antheridia apparently terminal, has been derived from species with axillary antheridia, either by restriction of the antheridia to the axils of the uppermost bracts, as in the liverwort Haplomitrium blumei, or by the suppression of the upper bracts.     

象山港硅藻赤潮过程中真菌群落演替及互作特征

[背景] 近岸海域赤潮的发生能显著改变包括真菌在内的微生物群落组成,进而影响海洋中碳氮元素循环。真菌是海洋中重要的分解者,但赤潮过程对真菌群落的影响少有报道。[目的] 探明赤潮过程对真菌群落的影响,对进一步阐释赤潮对生态系统中物质循环的影响具有重要意义。[方法] 针对2017年春季象山港硅藻赤潮,对真菌内源转录间隔区1（Internally Transcribed Spacer 1,ITS1）序列进行高通量测序,研究赤潮过程中真菌群落响应及共现性特征。[结果] 真菌群落的α多样性指数在赤潮暴发的各阶段差异显著,而且均与磷酸盐、硝酸盐和亚硝酸盐有显著相关性（P<0.05）。子囊菌门（Ascomycota,90.2%）和担子菌门（Basidiomycota,8.27%）是本次硅藻赤潮的优势菌门,其中子囊菌门遍布赤潮暴发的4个阶段;锤舌菌纲（Leotiomycetes,16.1%）和散囊菌纲（Eurotiomycetes,9.3%）是纲水平上的主要优势类群。赤潮暴发过程中硝酸盐、亚硝酸盐、磷酸盐和温度等环境因素的改变,驱动真菌群落组成及结构发生显著变化。Helotiales、Eurotiales、Xylariales、Sacc haromycetales和Agaricostilbales构成了真菌群落共现网络的主体。[结论] 真菌群落在赤潮的各阶段具有显著的演替特征,赤潮藻生消是引起真菌群落演替的主要因素,但是环境因子也扮演着重要角色。赤潮过程中,真菌群落同类群内的互作要强于不同类群间的互作,这对维持群落的稳定性具有重要意义。     

Two cases of sporotrichosis of the right upper extremity in right-handed patients with diabetes mellitus

BackgroundSporothrix species have proved to show high degrees of endemicity. Sporothrix globosa is the only pathogenic Sporothrix species that has till date been reported from China, where it is endemic in the northeastern provinces.AimsWe report two cases of lymphocutaneous sporotrichosis with diabetes mellitus as underlying disease in patients from the non-endemic area of China.MethodsA 59-year-old farmer and a 60-year-old gardener were admitted in February and June 2014, respectively. Both patients were right-handed men and presented with progressive plaques and nodules, which they had for several years, involving the right upper extremity. Skin biopsy from the granuloma was taken and cultured on Sabouraud medium, and molecular identification based on the calmodulin region was performed. Antifungal susceptibility testing was also performed with the microdilution method.ResultsBiopsy of the lesions showed the presence of infectious granuloma. The fungal cultures were identified as Sporothrix globosa by conventional methods, and confirmed by molecular identification. A subsequent course of oral antifungal therapy with low dosage of itraconazole was well tolerated and resolved the infection.ConclusionsIdentification of fungal species and antifungal susceptibility testing are mandatory for epidemiological and therapeutic reasons. Early diagnosis of sporotrichosis is essential to prevent those sequelae when the disease progresses and provides highly effective methods for treating this emerging disease. Avoiding the exposure to plant material potentially contaminated with fungal spores should be recommended, especially in immunocompromised patients.