Molecular diversity of arbuscular mycorrhizal fungi and patterns of host association over time and space in a tropical forest

We have used molecular techniques to investigate the diversity and distribution of the arbuscular mycorrhizal (AM) fungi colonizing tree seedling roots in the tropical forest on Barro Colorado Island (BCI), Republic of Panama. In the first year, we sampled newly emergent seedlings of the understory treelet Faramea occidentalis and the canopy emergent Tetragastris panamensis, from mixed seedling carpets at each of two sites. The following year we sampled surviving seedlings from these cohorts. The roots of 48 plants were analysed using AM fungal-specific primers to amplify and clone partial small subunit (SSU) ribosomal RNA gene sequences. Over 1300 clones were screened for random fragment length polymorphism (RFLP) variation and 7% of these were sequenced. Compared with AM fungal communities sampled from temperate habitats using the same method, the overall diversity was high, with a total of 30 AM fungal types identified. Seventeen of these types have not been recorded previously, with the remainder being similar to types reported from temperate habitats. The tropical mycorrhizal population showed significant spatial heterogeneity and nonrandom associations with the different hosts. Moreover there was a strong shift in the mycorrhizal communities over time. AM fungal types that were dominant in the newly germinated seedlings were almost entirely replaced by previously rare types in the surviving seedlings the following year. The high diversity and huge variation detected across time points, sites and hosts, implies that the AM fungal types are ecologically distinct and thus may have the potential to influence recruitment and host composition in tropical forests.     

Nuclear and Nucleomorph SSU rDNA Phylogeny in the Cryptophyta and the Evolution of Cryptophyte Diversity

The plastid-bearing members of the Cryptophyta contain two functional eukaryotic genomes of different phylogenetic origin, residing in the nucleus and in the nucleomorph, respectively. These widespread and diverse protists thus offer a unique opportunity to study the coevolution of two different eukaryotic genomes within one group of organisms. In this study, the SSU rRNA genes of both genomes were PCR-amplified with specific primers and phylogenetic analyses were performed on different data sets using different evolutionary models. The results show that the composition of the principal clades obtained from the phylogenetic analyses of both genes was largely congruent, but striking differences in evolutionary rates were observed. These affected the topologies of the nuclear and nucleomorph phylogenies differently, resulting in long-branch attraction artifacts when simple evolutionary models were applied. Deletion of long-branch taxa stabilized the internal branching order in both phylogenies and resulted in a completely resolved topology in the nucleomorph phylogeny. A comparison of the tree topologies derived from SSU rDNA sequences with characters previously used in cryptophyte systematics revealed that the biliprotein type was congruent, but the type of inner periplast component incongruent, with the molecular trees. The latter is indicative of a hidden cellular dimorphism (cells with two periplast types present in a single clonal strain) of presumably widespread occurrence throughout cryptophyte diversity, which, in consequence, has far-reaching implications for cryptophyte systematics as it is practiced today.     

Freshwater foraminiferans revealed by analysis of environmental DNA samples

Sediment-dwelling protists are among the most abundant meiobenthic organisms, ubiquitous in all types of aquatic ecosystems. Yet, because their isolation and identification are difficult, their diversity remains largely unknown. In the present work, we applied molecular methods to examine the diversity of freshwater Foraminifera, a group of granuloreticulosan protists largely neglected until now. By using specific PCR primers, we detected the presence of Foraminifera in all sediment samples examined. Phylogenetic analysis of amplified SSU rDNA sequences revealed two distinct groups of freshwater foraminiferans. All obtained sequences branched within monothalamous (single-chambered), marine Foraminifera, suggesting a repeated colonization of freshwater environments. The results of our study challenge the traditional view of Foraminifera as essentially marine organisms, and provide a conceptual framework for charting the molecular diversity of freshwater granuloreticulosan protists.     

Long branch attraction effects and the status of "basal eukaryotes": phylogeny and structural analysis of the ribosomal RNA gene cluster of the free-living diplomonad Trepomonas agilis

The three taxa emerging at the base of the eukaryotic ribosomal RNA phylogenetic tree (Diplomonadida, Microspora, and Parabasalia) include a wide array of parasitic species. and some free-living organisms that appear to be derived from a parasitic ancestry. The basal position of these taxa, which lack mitochondria, has recently been questioned. I sequenced most of the ribosomal RNA gene cluster of the free-living diplomonad Trepomonas agilis and a secondary structure model was reconstructed for the SSU rRNA. I conducted a RASA matrix analysis to identify, independently from tree reconstruction, putative long branch attraction effects in the data matrix. The results show that each of the basal clades and the euglenozoan clade act, indeed, as long branches and may have been engaged in a process of accelerated rate of evolution. A nucleotide signature analysis was conducted in the conserved regions for positions defining the three great domains of life (Eubacteria, Archea, and Eukaryota). For the three basal taxa, this analysis showed the presence of a significant number of different non-eukaryotic nucleotides. A precise study of the nature and location of these nucleotides led to conclusions supporting the results of the RASA analysis. Altogether, these findings suggest that the basal placement of these taxa in the SSU ribosomal RNA phylogenetic tree is artifactual, and flawed by long branch attraction effects.     

SSU rDNA phylogeny of cladoniiform lichens

To examine phylogenetic relationships among the "cladoniiform" lichenized fungi, i.e., the families Cladoniaceae, Baeomycetaceae, Icmadophilaceae, Stereocaulaceae, and Siphulaceae, and to provide evidence for the anticipated independent origins of podetia and pseudopodetia, we conducted phylogenetic analyses of SSU (small subunit) rDNA sequences from 39 lichen-forming fungi. These fungi represent all of the major growth forms of lichen associations, fruticose (including "cladoniiform"), foliose, and crustose. Our analysis suggests that lichen-forming fungi with a "cladoniiform" morphology arose multiple times within the ascomycetes. Additionally, each of the other thallus growth forms, crustose, foliose, and fruticose, have originated multiple times. It also seems to be clear that neither all podetiate nor all pseudopodetiate taxa form a monophyletic group. Therefore the term "podetium" should be restricted to homologous structures that are most probably limited to the genera Cladonia, Cladina, Pycnothelia, and allies. The "pseudopodetia" of Stereocaulon (Stereocaulaceae) and Cladia (Cladiaceae) may represent different states of the same homologous character. Our phylogenetic hypothesis supports the monophyletic origin of the order Lecanorales sensu stricto, including representatives of five suborders Cladoniineae, Lecanorineae, Teloschistineae, Agyriineae and Peltigerineae, but excluding representatives of the suborders Acarosporineae (Acarospora schleicheri and Megaspora verrucosa), Pertusariineae (Pertusaria trachythallina), and Umbilicarineae. The suborder Cladoniineae and the family Cladoniaceae both appear to be polyphyletic assemblages.     

A multigene phylogeny of the Gelidiales including nuclear large-subunit rRNA sequence data

Partial sequences (1032 bp) of the nuclear-encoded large ribosomal RNA gene (LSU) were determined for 16 gelidialean species, and analyzed separately and in combination with plastid rbcL and nuclear SSU gene sequences. The number of informative characters and levels of sequence divergence among taxa are intermediate in LSU sequences as compared to that for rbcL and SSU. Analyses of the separate LSU, and a combined LSU, SSU, and rbcL data sets have identified early-diverging lineages within the Gelidiales including Gelidiella, Pterocladia, Pterocladiella, and a lineage including Gelidium and species classified in other genera. The relationships among most gelidialean taxa are well-resolved and well-supported by analyses of the combined data; however, the relationships of Ptilophora and Capreolia remain unclear. It is speculated that these two lineages have diverged from a common ancestor over an evolutionarily short period of time. This revised version was published online in June 2006 with corrections to the Cover Date.     

Relationships of the Insect-Pathogenic Order Entomophthorales (Zygomycota, Fungi) Based on Phylogenetic Analyses of Nuclear Small Subunit Ribosomal DNA Sequences (SSU rDNA)

We sequenced the nuclear small subunit of ribosomal DNA (SSU rDNA) from seven species within the insect-pathogenic order Entomophthorales. These sequences were aligned with other published SSU rDNA sequences and phylogenies were inferred using phenetic and cladistic methods. Based on three different phylogenetic methods the Entomophthorales (excludingBasidiobolus ranarum) is monophyletic;B. ranarumwas more closely related to chytrids from Chytridiales and Neocallimasticales than to Entomophthorales, as was proposed by Nagahamaet al.(Mycologia87:203–209, 1995). Nuclear characters (large nuclei containing conspicuous condensed chromatin and lack of a prominent nucleolus) were of predictive value for the monophyly of the family Entomophthoraceae. Conidial characters separate the Entomophthoraceae, which only includes obligate pathogens, into at least two lineages: one lineage with uninucleate conidia and another with multinucleate conidia. The two species ofConidiobolusstudied were paraphyletic in our analyses and only distantly related to each other. This information may prove to be important in the use of these fungi as biocontrol agents.     

2型猪链球菌MocR家族转录调控因子SSU0562基因敲除突变体的构建及毒力分析

目的:构建2型猪链球菌强毒株05ZYH33中MocR家族转录调控因子SSU0562基因敲除的突变株,探索SSU0562基因缺失对细菌基本生物学特性和毒力的影响。方法:构建左右两侧为SSU0562基因上下游的同源序列,中间部分为壮观霉素抗性基因(Spcr)的基因敲除质粒,通过同源重组的方法筛选SSU0562基因敲除突变株Δ0562。对突变株与野生株的基本生物学特性进行系统的比较分析,并且将小鼠作为动物感染的模型来研究突变株的毒力。结果:组合PCR的分析及基因测序结果均表明Spcr完全取代了S.suis2中SSU0562基因位点,表明基因敲除突变体Δ0562构建成功,反转录PCR(RT-PCR)证实了突变株Δ0562中SSU0562基因在转录水平的缺失;在溶血活性、生长速率及对小鼠的致病力方面,突变株Δ0562与野生株05ZYH33相比均无显著差别,然而革兰染色实验显示突变株Δ0562的成链能力明显减弱。结论:猪链球菌强毒株05ZYH33的毒力并未因SSU0562基因的缺失而发生显著性改变,表明SSU0562基因并非猪链球菌的毒力决定因子,但很有可能参与猪链球菌成链能力的调控。     

A Description of a New "Amoebozoan" Isolated from the American Lobster, Homarus americanus

ABSTRACT. Our knowledge of the diversity of amoeboid protists is rapidly expanding as new and old habitats are more fully explored. In 2003, while investigating the cause of an amoeboid disease afflicting lobsters on the East Coast, samples were examined for the presence of amoebae from the carapace washings of the American lobster, Homarus americanus . During this survey a unique community of gymnamoebae was discovered. Among the new taxa discovered was a small Thecamoeba -like organism with a single posteriorly directed pseudopodium. Although resembling Parvamoeba rugata , this amoeba displayed distinctive morphology from that isolate or any other amoebozoan. Phylogenetic analysis shows this amoeba is distantly related to the Thecamoebidae. In this paper we describe the unique morphology of a second species of Parvamoeba and discuss its phylogenetic position with respect to the "Amoebozoa."     

Redescription of Favella ehrenbergii (Claparède and Lachmann, 1858) Jörgensen, 1924 (Ciliophora: Choreotrichia), with Phylogenetic Analyses Based on Small Subunit rRNA Gene Sequences

ABSTRACT. The identification of Favella ehrenbergii, a marine planktonic ciliate, has largely been based on its lorica features. This approach is potentially problematic given the polymorphic lorica during this organism's life cycle. We isolated a population of F. ehrenbergii from the coastal waters of Incheon, Korea, and revealed its infraciliature using the protargol staining method. Phylogenetic analysis based on small subunit rRNA gene sequences was also performed. Results showed that this population possessed 16 collar membranelles (CM) and about 100 somatic kineties. These features are highly conserved, even in later dividers. As such, the number of CM and somatic kineties can be used as key characteristics for identification of Favella species.