Eukaryotic translation elongation factor 1 alpha (eEF1A) inhibits Siniperca chuatsi rhabdovirus (SCRV) infection through two distinct mechanisms.

IMPORTANCE: Although a virus can regulate many cellular responses to facilitate its replication by interacting with host proteins, the host can also restrict virus infection through these interactions. In the present study, we showed that the host eukaryotic translation elongation factor 1 alpha (eEF1A), an essential protein in the translation machinery, interacted with two proteins of a fish rhabdovirus, Siniperca chuatsi rhabdovirus (SCRV), and inhibited virus infection via two different mechanisms: (i) inhibiting the formation of crucial viral protein complexes required for virus transcription and replication and (ii) promoting the ubiquitin-proteasome degradation of viral protein. We also revealed the functional regions of eEF1A that are involved in the two processes. Such a host protein inhibiting a rhabdovirus infection in two ways is rarely reported. These findings provided new information for the interactions between host and fish rhabdovirus.

High expression of eukaryotic elongation factor 1-alpha-2 in lung adenocarcinoma is associated with poor prognosis.

Eukaryotic elongation factor 1 alpha 2 (eEF1A2) encodes an isoform of the alpha subunit of the elongation factor 1 complex and is responsible for the enzymatic delivery of aminoacyl tRNA to the ribosome. Our proteomic analysis has identified eEF1A2 as one of the proteins expressed during malignant progression from adenocarcinoma in situ (AIS) to early invasive lung adenocarcinoma. The expression level of eEF1A2 in 175 lung adenocarcinomas was examined by immunohistochemical staining in relation to patient prognosis and clinicopathological factors. Quantitative PCR analysis and fluorescence in situ hybridization (FISH) were performed to evaluate the amplification of the eEF1A2 gene. Relatively high expression of eEF1A2 was observed in invasive adenocarcinoma (39/144 cases) relative to minimally invasive adenocarcinoma (1/10 cases) or AIS (0/21 cases). Among invasive adenocarcinomas, solid-type adenocarcinoma (15/32 cases, 47%) showed higher expression than other histological subtypes (23/92, 25%). Patients with eEF1A2-positive tumors had a significantly poorer prognosis than those with eEF1A2-negative tumors. Of the five tumors that were eEF1A2-positive, two cases showed amplified genomic eEF1A2 DNA, which was confirmed by both qPCR and FISH. These findings indicate that eEF1A2 overexpression occurs in the course of malignant transformation of lung adenocarcinomas and is partly due to eEF1A2 gene amplification.

Fusarium Species Associated with Fusarium Head Blight in Hungarian Wheat Fields.

The species composition of the genus Fusarium associated with Fusarium head blight (FHB) in wheat fields of Hungary in the year 2019 was assessed. Symptomatic wheat heads were collected at 20 geographical locations representing different ecosystems. A total of 256 Fusarium strains were isolated and identified by partial sequences of the translation elongation factor 1-alpha gene and, where required, the second-largest subunit of the DNA-directed RNA polymerase gene. Overall, Fusarium graminearum (58.2%) proved to be the dominant species, followed by F. annulatum (formerly F. proliferatum) (17.2%) and F. verticillioides (7.4%). The presence of all other species, including F. culmorum, in the population was less than 5%. F. graminearum was identified as the main species associated with FHB at 14 sampling sites. Fumonisin-producing F. annulatum, primarily known as the pathogen of maize in Hungary, was detected nearly as frequently as F. graminearum at three locations and dominated at two other sites. F. poae was not found during the survey. F. vorosii, a species that is believed to be of Asian origin and was already found in Hungary in 2002, was identified at two locations.

Novel Enhanced Mammalian Cell Transient Expression Vector via Promoter Combination.

During the emergence of infectious diseases, evaluating the efficacy of newly developed vaccines requires antigen proteins. Available methods enhance antigen protein productivity; however, structural modifications may occur. Therefore, we aimed to construct a novel transient overexpression vector capable of rapidly producing large quantities of antigenic proteins in mammalian cell lines. This involved expanding beyond the exclusive use of the human cytomegalovirus (CMV) promoter, and was achieved by incorporating a transcriptional enhancer (CMV enhancer), a translational enhancer (woodchuck hepatitis virus post-transcriptional regulatory element), and a promoter based on the CMV promoter. Twenty novel transient expression vectors were constructed, with the vector containing the human elongation factor 1-alpha (EF-1a) promoter showing the highest efficiency in expressing foreign proteins. This vector exhibited an approximately 27-fold higher expression of enhanced green fluorescent protein than the control vector containing only the CMV promoter. It also expressed the highest level of severe acute respiratory syndrome coronavirus 2 receptor-binding domain protein. These observations possibly result from the simultaneous enhancement of the transcriptional activity of the CMV promoter and the human EF-1a promoter by the CMV enhancer. Additionally, the synergistic effect between the CMV and human EF-1a promoters likely contributed to the further enhancement of protein expression.

A Potential Role for the Gsdf-eEF1α Complex in Inhibiting Germ Cell Proliferation: A Protein-Interaction Analysis in Medaka (Oryzias latipes) From a Proteomics Perspective.

Gonadal soma-derived factor (gsdf) has been demonstrated to be essential for testicular differentiation in medaka (Oryzias latipes). To understand the protein dynamics of Gsdf in spermatogenesis regulation, we used a His-tag "pull-down" assay coupled with shotgun LC-MS/MS to identify a group of potential interacting partners for Gsdf, which included cytoplasmic dynein light chain 2, eukaryotic polypeptide elongation factor 1 alpha (eEF1α), and actin filaments in the mature medaka testis. As for the interaction with transforming growth factor ß-dynein being critical for spermatogonial division in Drosophila melanogaster, the physical interactions of Gsdf-dynein and Gsdf-eEF1α were identified through a yeast 2-hybrid screening of an adult testis cDNA library using Gsdf as bait, which were verified by a paired yeast 2-hybrid assay. Coimmunoprecipitation of Gsdf and eEF1α was defined in adult testes as supporting the requirement of a Gsdf and eEF1α interaction in testis development. Proteomics analysis (data are available via ProteomeXchange with identifier PXD022153) and ultrastructural observations showed that Gsdf deficiency activated eEF1α-mediated protein synthesis and ribosomal biogenesis, which in turn led to the differentiation of undifferentiated germ cells. Thus, our results provide a framework and new insight into the coordination of a Gsdf (transforming growth factor ß) and eEF1α complex in the basic processes of germ cell proliferation, transcriptional and translational control of sexual RNA, which may be fundamentally conserved across the phyla during sexual differentiation.

Cryptosporidium parvum Elongation Factor 1α Participates in the Formation of Base Structure at the Infection Site During Invasion.

BACKGROUND: Cryptosporidium is a genus of apicomplexan parasites, the causative agents of cryptosporidiosis in humans and/or animals. Although most apicomplexans parasitize within the host cell cytosols, Cryptosporidium resides on top of host cells, but it is embraced by a double-layer parasitophorous vacuole membrane derived from host cell. There is an electron-dense band to separate the parasite from host cell cytoplasm, making it as an intracellular but extracytoplasmic parasite. However, little is known on the molecular machinery at the host cell-parasite interface. METHODS: Cryptosporidium parvum at various developmental stages were obtained by infecting HCT-8 cells cultured in vitro. Immunofluorescence assay was used to detect CpEF1α with a polyclonal antibody and host cell F-actin with rhodamine-phalloidin. Recombinant CpEF1α protein was used to evaluate its effect on the invasion by the parasite. RESULTS: We discovered that a C parvum translation elongation factor 1α (CpEF1α) was discharged from the invading sporozoites into host cells, forming a crescent-shaped patch that fully resembles the electron-dense band. At the same time, host cell F-actin aggregated to form a globular-shaped plug beneath the CpEF1α patch. The CpEF1α patch remained for most of the time but became weakened and dissolved upon the completion of the invasion process. In addition, recombinant CpEF1α protein could effectively interfere the invasion of sporozoites into host cells. CONCLUSIONS: CpEF1α plays a role in the parasite invasion by participating in the formation of electron-dense band at the base of the parasite infection site.

Recombinant elongation factor 1 alpha of Haemonchus contortus affects the functions of goat PBMCs.

Excretory/secretory proteins of Haemonchus contortus (HcESPs) intermingle comprehensively with host immune cells and modulate host immune responses. In this study, H contortus ES antigen named as elongation factor 1 alpha (HcEF-1α) was cloned and expressed. The influences of recombinant HcEF-1α on multiple functions of goat peripheral blood mononuclear cells (PBMCs) were observed in vitro. Immunoblot analysis revealed that rHcEF-1α was recognized by the serum of goat infected with H contortus. Immunofluorescence analysis indicated that rHcEF-1α was bound on surface of PBMCs. Moreover, the productions of IL-4, TGF-ß1, IFN-γ and IL-17 of cells were significantly modulated by the incubation with rHcEF-1α. The production of interleukin IL-10 was decreased. Cell migration, cell proliferation and cell apoptosis were significantly increased; however, nitric oxide production (NO) was significantly decreased. The MHC II molecule expression of cells incubated with rHcEF-1α was increased significantly, whereas MHC-I was not changed as compared to the control groups (PBS control and pET32a). These findings indicated that rHcEF-1α protein might play essential roles in functional regulations of HcESPs on goat PBMC and mediate the immune responses of the host during host-parasite relationship.

Trichoderma panacis sp. nov., an endophyte isolated from Panax notoginseng.

An endophytic member of the genus Trichoderma was isolated from the root of a healthy 3-year-old Panax notoginseng in Yunnan province, PR China. The results of phylogenetic analyses based on a combined of ITS, tef1 and rpb2 indicated that this isolate was distinct from other species of the genus Trichoderma and closely related to Trichoderma songyi. It can be distinguished from T. songyi by its slower growth rates on PDA and colony morphology. The novel isolate formed conidia in thick white pustules scattered mostly at the margin. Its conidiophores tended to be regularly verticillium-like, little branched, sometimes substituted by phialides singly or in whorls. Conidia are smooth, mostly broadly subglobose to ellipsoidal. In combination with the genotypic and phenotypic characteristics, all data demonstrated that the fungus studied represented a unique and distinguishable novel species of the genus Trichoderma, for which the name Trichoderma panacis sp. nov. is proposed.

Early defects in translation elongation factor 1α levels at excitatory synapses in α-synucleinopathy.

Cognitive decline and dementia in neurodegenerative diseases are associated with synapse dysfunction and loss, which may precede neuron loss by several years. While misfolded and aggregated α-synuclein is recognized in the disease progression of synucleinopathies, the nature of glutamatergic synapse dysfunction and loss remains incompletely understood. Using fluorescence-activated synaptosome sorting (FASS), we enriched excitatory glutamatergic synaptosomes from mice overexpressing human alpha-synuclein (h-αS) and wild-type littermates to unprecedented purity. Subsequent label-free proteomic quantification revealed a set of proteins differentially expressed upon human alpha-synuclein overexpression. These include overrepresented proteins involved in the synaptic vesicle cycle, ER-Golgi trafficking, metabolism and cytoskeleton. Unexpectedly, we found and validated a steep reduction of eukaryotic translation elongation factor 1 alpha (eEF1A1) levels in excitatory synapses at early stages of h-αS mouse model pathology. While eEF1A1 reduction correlated with the loss of postsynapses, its immunoreactivity was found on both sides of excitatory synapses. Moreover, we observed a reduction in eEF1A1 immunoreactivity in the cingulate gyrus neuropil of patients with Lewy body disease along with a reduction in PSD95 levels. Altogether, our results suggest a link between structural impairments underlying cognitive decline in neurodegenerative disorders and local synaptic defects. eEF1A1 may therefore represent a limiting factor to synapse maintenance.

Mating analyses of Trichophyton benhamiae offspring reveals linkage of genetic markers used in taxonomy.

Mating experiments were conducted with four clinical Trichophyton benhamiae isolates, genetically similar to the Trichophyton benhamiae CBS 112371, featuring the plus mating type and with two minus type strains. One minus type strain belonged to the white subgroup, and the other minus type strain, DSM 6916, showed genetic kinship to the yellow subgroup. Only two plus type strains were able to form mature, pigmented gymnothecia with DSM 6916. These two plus type strains demonstrated dark pigmentation and powdery mycelium on Takashio agar, whereas the other three strains exhibited a low degree of pigmentation on the same medium. All five plus strains were able to mate with the minus type strain of their own white subgroup. Cultures from single ascospore isolates showed highly variable morphology and pigmentation. Three genetic markers (ITS, mating type, EF1 alpha) were analyzed in polymerase chain reaction (PCR) experiments with optimized primers and PCR conditions to discriminate between subgroups. Furthermore, RAPD-PCR was used to generate a DSM 6916-specific DNA-fragment which served as an additional genetic marker. Assessing the isolates with recombinant genotypes, it was found that three genetic markers behave like linked genes. The recombination of plus mating type went together with ITS, EF1 alpha and RAPD marker of the DSM 6916 parental strain and was most frequently isolated, whereas plus types recombinants in this case were completely missing. This shows a high imbalance in mating type distribution of recombinants.

The toad fly Lucilia bufonivora: its evolutionary status and molecular identification.

The blow fly genus Lucilia is composed largely of saprophages and facultative myasis agents, including the economically important species Lucilia cuprina (Wiedemann) (Diptera: Calliphoridae) and Lucilia sericata (Meigen). Only one species is generally recognized as an obligate agent of myiasis, Lucilia bufonivora Moniez, and this is an obligate parasite of toads. Lucilia silvarum (Meigen), a sister species, behaves mainly as a carrion breeder; however, it has also been reported as a facultative parasite of amphibians. Morphologically, these species are almost identical, and historically this has led to misidentification, taxonomic ambiguity and a paucity of studies of L. bufonivora. In this study, dipterous larvae were analysed from toad myiasis cases from the U.K., The Netherlands and Switzerland, together with adult specimens of fly species implicated in amphibian parasitism: L. bufonivora, L. silvarum and Lucilia elongata Shannon (from North America). Partial sequences of two genes, cox1 and ef1α, were amplified. Seven additional blow fly species were analysed as outgroups. Bayesian inference trees of cox1, ef1α and a combined-gene dataset were constructed. All larvae isolated from toads were identified as L. bufonivora and no specimens of L. silvarum were implicated in amphibian myiasis. This study confirms L. silvarum and L. bufonivora as distinct sister species and provides unambiguous molecular identification of L. bufonivora.

Regulation of eukaryotic elongation factor 1 alpha (eEF1A) by dynamic lysine methylation.

Lysine methylation is a frequent post-translational protein modification, which has been intensively studied in the case of histone proteins. Lysine methylations are also found on many non-histone proteins, and one prominent example is eukaryotic elongation factor 1 alpha (eEF1A). Besides its essential role in the protein synthesis machinery, a number of non-canonical functions have also been described for eEF1A, such as regulation of the actin cytoskeleton and the promotion of viral replication. The functional significance of the extensive lysine methylations on eEF1A, as well as the identity of the responsible lysine methyltransferases (KMTs), have until recently remained largely elusive. However, recent discoveries and characterizations of human eEF1A-specific KMTs indicate that lysine methylation of eEF1A can be dynamic and inducible, and modulates mRNA translation in a codon-specific fashion. Here, we give a general overview of eEF1A lysine methylation and discuss its possible functional and regulatory significance, with particular emphasis on newly discovered human KMTs.

Molecular phylogeny of Miltogramminae (Diptera: Sarcophagidae): Implications for classification, systematics and evolution of larval feeding strategies.

Miltogramminae is one of the phylogenetically most poorly studied taxa of the species-rich family Sarcophagidae (Diptera). Most species are kleptoparasites in nests of solitary aculeate wasps and bees, although parasitoids and saprophagous species are also known, and the ancestral miltogrammine life habit remains unsettled. Here, we present for the first time a comprehensive phylogenetic tree consisting of 58 representatives of Miltogramminae, reconstructed using sequence data from three mitochondrial (COI, cytB, ND4) and one nuclear (Ef-1α) genes. Our phylogenetic hypothesis suggests that: (1) Miltogramminae are sister to Paramacronychiinae, (2) Miltogramminae can be divided into the "lower miltogrammines" containing two clades of mainly saprophages and a clade of "higher miltogrammines" with mainly kleptoparasitic species, (3) only three genera turn out to be non-monophyletic: Miltogramma, Senotainia and Pterella and (4) the genus Sarcotachina, which traditionally has been considered as belonging to the Paramacronychiinae, is placed in one of the clades of "lower miltogrammines". Ancestral state reconstruction of larval feeding strategy and five larval characters reveals that the ancestor of Miltogramminae was likely a saprophage retaining plesiomorphic oral ridges and a cephaloskeleton with sclerotized dorsal bridge. Synapomorphies like large pseudocephalic sensory organs and well-developed cuticular sculpture suggest that the ancestral first instar larva actively searched for a buried food supply.

SPL33, encoding an eEF1A-like protein, negatively regulates cell death and defense responses in rice.

Lesion-mimic mutants are useful to dissect programmed cell death and defense-related pathways in plants. Here we identified a new rice lesion-mimic mutant, spotted leaf 33 (spl33) and cloned the causal gene by a map-based cloning strategy. SPL33 encodes a eukaryotic translation elongation factor 1 alpha (eEF1A)-like protein consisting of a non-functional zinc finger domain and three functional EF-Tu domains. spl33 exhibited programmed cell death-mediated cell death and early leaf senescence, as evidenced by analyses of four histochemical markers, namely H2O2 accumulation, cell death, callose accumulation and TUNEL-positive nuclei, and by four indicators, namely loss of chlorophyll, breakdown of chloroplasts, down-regulation of photosynthesis-related genes, and up-regulation of senescence-associated genes. Defense responses were induced in the spl33 mutant, as shown by enhanced resistance to both the fungal pathogen Magnaporthe oryzae and the bacterial pathogen Xanthomonas oryzae pv. oryzae and by up-regulation of defense response genes. Transcriptome analysis of the spl33 mutant and its wild type provided further evidence for the biological effects of loss of SPL33 function in cell death, leaf senescence and defense responses in rice. Detailed analyses showed that reactive oxygen species accumulation may be the cause of cell death in the spl33 mutant, whereas uncontrolled activation of multiple innate immunity-related receptor genes and signaling molecules may be responsible for the enhanced disease resistance observed in spl33. Thus, we have demonstrated involvement of an eEF1A-like protein in programmed cell death and provided a link to defense responses in rice.

Molecular characterization, expression analysis and RNAi knock-down of elongation factor 1α and 1γ from Nilaparvata lugens and its yeast-like symbiont.

In the present paper, four cDNAs encoding the alpha and gamma subunits of elongation factor 1 (EF-1) were cloned and sequenced from Nilaparvata lugens, named NlEF-1α, NlEF-1γ, and its yeast-like symbiont (YLS), named YsEF-1α and YsEF-1γ, respectively. Comparisons with sequences from other species indicated a greater conservation for EF-1α than for EF-1γ. NlEF-1α has two identical copies. The deduced amino acid sequence homology of NlEF-1α and NlEF-1γ is 96 and 64%, respectively, compared with Homalodisca vitripennis and Locusta migratoria. The deduced amino acid sequence homology of YsEF-1α and YsEF-1γ is 96 and 74%, respectively, compared with Metarhizium anisopliae and Ophiocordyceps sinensis. Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) analysis revealed that the expression level of NlEF-1α and NlEF-1γ mRNA in hemolymph, ovary, fat body and salivary glands were higher than the midgut and leg tissue. YsEF-1α and YsEF-1γ was highly expressed in fat body. The expression level of NlEF-1α was higher than that of NlEF-1γ. Through RNA interference (RNAi) of the two genes, the mortality of nymph reached 92.2% at the 11th day after treatment and the ovarian development was severely hindered. The RT-qPCR analysis verified the correlation between mortality, sterility and the down-regulation of the target genes. The expression and synthesis of vitellogenin (Vg) protein in insects injected with NlEF-1α and NlEF-1γ double-stranded RNA (dsRNA) was significantly lower than control groups. Attempts to knockdown the YsEF-1 genes in the YLS was unsuccessful. However, the phenotype of N. lugens injected with YsEF-1α dsRNA was the same as that injected with NlEF-1α dsRNA, possibly due to the high similarity (up to 71.9%) in the nucleotide sequences between NlEF-1α and YsEF-1α. We demonstrated that partial silencing of NlEF-1α and NlEF-1γ genes caused lethal and sterility effect on N. lugens. NlEF-1γ shares low identity with that of other insects and therefore it could be a potential target for RNAi-based pest management.

Heat-induced accumulation of protein synthesis elongation factor 1A implies an important role in heat tolerance in potato.

MAIN CONCLUSION: Potato eukaryotic elongation factor 1A comprises multiple isoforms, some of which are heat-inducible or heat-upregulated and might be important in alleviating adverse effects of heat stress on plant productivity. Heat stress substantially reduces crop productivity worldwide, and will become more severe due to global warming. Identification of proteins involved in heat stress response may help develop varieties for heat tolerance. Eukaryotic elongation factor 1A (eEF1A) is a cytosolic, multifunctional protein that plays a central role in the elongation phase of translation. Some of the non-canonical eEF1A activities might be important in developing plant heat-stress tolerance. In this study, we investigated effects of heat stress (HS) on eEF1A expression at the protein level in potato, a highly heat vulnerable crop. Our results from both the controlled environment and the field have shown that potato eEF1A is a heat-inducible protein of 49.2-kDa with multiple isoforms (5-8). Increase in eEF1A abundance under HS can be mainly attributed to 2-3 basic polypeptides/isoforms. A significant correlation between eEF1A abundance and the potato productivity in the field was observed in two extremely hot years 2011 and 2012. Genomic Southern blot analysis indicated the existence of multiple genes encoding eEF1A in potato. Identification, isolation and utilization of heat-inducible eEF1A genes might be helpful for the development of the heat-tolerant varieties.

No more time to stay 'single' in the detection of Anisakis pegreffii, A. simplex (s. s.) and hybridization events between them: a multi-marker nuclear genotyping approach.

A multi-marker nuclear genotyping approach was performed on larval and adult specimens of Anisakis spp. (N = 689) collected from fish and cetaceans in allopatric and sympatric areas of the two species Anisakis pegreffii and Anisakis simplex (s. s.), in order to: (1) identify specimens belonging to the parental taxa by using nuclear markers (allozymes loci) and sequence analysis of a new diagnostic nuclear DNA locus (i.e. partial sequence of the EF1 α-1 nDNA region) and (2) recognize hybrid categories. According to the Bayesian clustering algorithms, based on those markers, most of the individuals (N = 678) were identified as the parental species [i.e. A. pegreffii or A. simplex (s. s.)], whereas a smaller portion (N = 11) were recognized as F1 hybrids. Discordant results were obtained when using the polymerase chain reaction-restriction fragment length polymorphisms (PCR-RFLPs) of the internal transcribed spacer (ITS) ribosomal DNA (rDNA) on the same specimens, which indicated the occurrence of a large number of 'hybrids' both in sympatry and allopatry. These findings raise the question of possible misidentification of specimens belonging to the two parental Anisakis and their hybrid categories derived from the application of that single marker (i.e. PCR-RFLPs analysis of the ITS of rDNA). Finally, Bayesian clustering, using allozymes and EF1 α-1 nDNA markers, has demonstrated that hybridization between A. pegreffii and A. simplex (s. s.) is a contemporary phenomenon in sympatric areas, while no introgressive hybridization takes place between the two species.

Phylogenetic Relationships of Japanese Auritibicen Species (Hemiptera: Cicadidae: Cryptotympanini) Inferred from Mitochondrial and Nuclear Gene Sequences.

We investigated the phylogenetic relationships and divergence times within the genus Auritibicen(Cicadidae: Cicadinae: Cryptotympanini), analyzing five Japanese species (A. japonicus, A. bihamatus,A. kyushyuensis, A. esakii and A. flammatus) and three species from East Asian mainland and Taiwan (A. atrofasciatus, A. intermedius and A. chujoi) using mitochondrial cytochrome oxidase subunit I (COI) and nuclear elongation factor 1-alpha (EF-1a) gene sequences. Although the EF-1a gene tree did not resolve the relationships among these Auritibicen species, the trees based on COI gene and the combined data set showed that Japanese taxa comprised three distinct lineages: the individual species A. flammatus and A. bihamatus, and the A. japonicus group, comprising A. japonicus, A. esakii and A. kyushyuensis from Japan and A. intermedius from Korea. In A. kyushyuensis, which comprises three populations in Kyushu, western Honshu and Shikoku, the specimens from western Honshu and Shikoku were closely related to each other, but not to the specimen from Kyushu; instead, they were sister to the Korean A. intermedius. The incongruence between the gene tree and species tree necessitates further population genetic and morphological studies to confirm the classification and species status of the western Honshu and Shikoku populations of A. kyushyuensis, which were originally described as two independent species. Divergence time estimation suggested that the most recent common ancestor of Auritibicen species studied dated back to the late Pliocene and that the species of the A. japonicus group diverged during the mid Pleistocene. Thus, the Pleistocene climatic fluctuation may have promoted the divergence of the Auritibicen species.

Taxonomy and phylogeny of three species in Perichaena sensu lato (Myxomycetes = Myxogastria) from China.

After Gulielmina was erected and Ophiotheca was resurrected based on some species originally included in Perichaena (Trichiaceae, Trichiales, Myxomyxetes), some specimens from the Herbarium of Fungi of Nanjing Normal University previously identified as Perichaena species were reexamined from morphological and two-gene (nuclear 18S rDNA and elongation factor-1 alpha) phylogenetic perspectives. In this study, two new myxomycete species, Gulielmina subreticulospora and Ophiotheca dictyospora, are described. Gulielmina subreticulospora shows the following character combination: branched plasmodiocarps, single peridium with circular protrusions in the inner surface, capillitium (2.4-)2.8-3.0(-3.4) µm in diameter, spores (7.4-)8.0-8.5(-9.0) µm in diameter and sub-reticulated. Ophiotheca dictyospora shows the following character combination: sessile sporocarps to short plasmodiocarps, single peridium with a densely irregular network and protrusions in the inner surface, capillitium (2.7-)3.5-5.0(-7.1) µm in diameter, uneven, decorated with spines of uneven size, spores (7.7-)8.2-8.6(-9.4) µm in diameter including obviously complete cristate reticulation with serrated edges, with deep and clear grids. Both new taxa were compared with related species and their genetic isolation was statistically tested. Moreover, a comprehensive morphological description and a detailed figure plate are provided for Perichaena verrucifera, and its phylogenetic position is determined.