Ecuadorian Acanthocinini: description of two new species and transference and notes on Sympagus cooperi Monn & Monn.

Two new Acanthocinini are described from Ecuador: Nealcidion apunctatum sp. nov.; and Xenocona nubicola sp. nov. Sympagus cooperi Monn & Monn, 2017 is transferred to Stenolis Bates, 1864, and notes on the differences between these two genera are provided.

Dissecting the Heterogeneous Glycan Profiles of Recombinant Coronavirus Spike Proteins with Individual Ion Mass Spectrometry.

Surface-embedded glycoproteins, such as the spike protein trimers of coronaviruses MERS, SARS-CoV, and SARS-CoV-2, play a key role in viral function and are the target antigen for many vaccines. However, their significant glycan heterogeneity poses an analytical challenge. Here, we utilized individual ion mass spectrometry (I2MS), a multiplexed charge detection measurement with similarities to charge detection mass spectrometry (CDMS), in which a commercially available Orbitrap analyzer is used to directly produce mass profiles of these heterogeneous coronavirus spike protein trimers under native-like conditions. Analysis by I2MS shows that glycosylation contributes to the molecular mass of each protein trimer more significantly than expected by bottom-up techniques, highlighting the importance of obtaining complementary intact mass information when characterizing glycosylation of such heterogeneous proteins. Enzymatic dissection to remove sialic acid or N-linked glycans demonstrates that I2MS can be used to better understand the glycan profile from a native viewpoint. Deglycosylation of N-glycans followed by I2MS analysis indicates that the SARS-CoV-2 spike protein trimer contains glycans that are more difficult to remove than its MERS and SARS-CoV counterparts, and these differences are correlated with solvent accessibility. I2MS technology enables characterization of protein mass and intact glycan profile and is orthogonal to traditional mass analysis methods such as size exclusion chromatography-multiangle light scattering (SEC-MALS) and field flow fractionation-multiangle light scattering (FFF-MALS). An added advantage of I2MS is low sample use, requiring 100-fold less than other methodologies. This work highlights how I2MS technology can enable efficient development of vaccines and therapeutics for pharmaceutical development.

Outline, phylogenetic and divergence times analyses of the genus Haploporus (Polyporales, Basidiomycota): two new species are proposed.

Haploporus species have a worldwide distribution and 27 species have been accepted. In this study, two new species, Haploporuscrystallinus and H.dextrinoideus, are proposed from South America, based on the molecular fragments (ITS, LSU and mtSSU) and morphological evidence. Molecular clock analysis was performed and the result suggests that the ancestor of Polyporales originated between the Late Jurassic and Early Cretaceous period, with a mean stem of 159.8 Mya [95% higher posterior density (HPD) of 142.4-184.1 Mya] and the genus Haploporus occurred at a mean stem of 108.3 Mya (95% HPD of 88.5-128.2 Mya). In addition, most species of the genus are diversified between 60.5 Mya and 1.8 Mya, during the Paleogene to Neogene. A key to the accepted species of the genus Haploporus is provided.

Taxonomic and phylogenetic contributions to Fuscoporia (Hymenochaetales, Basidiomycota): two new species from Hawaii with a key to North American species.

Fuscoporia is a cosmopolitan, poroid, wood-decaying genus, belonging to the Hymenochaetales. During a study of wood-inhabiting fungi in the USA, four unknown specimens were collected from Hawaii. Both morphological criteria and molecular genetic analyses based on the ITS+nLSU+EF1-α datasets and the nLSU dataset confirmed that these four specimens represent two new species of Fuscoporia, and they are described as F. hawaiiana and F. minutissima. Fuscoporia hawaiiana is characterized by pileate basidiocarps, the absence of cystidioles, hooked hymenial setae, broadly ellipsoid to subglobose basidiospores measuring 4-6 × 3.5-4.5 µm. Fuscoporia minutissima is distinguished by small pores (10-13 per mm) and basidiospores (3.4-4 × 2.4-3 µm). The taxonomic status of the two new species is briefly discussed. A key to the North American species of Fuscoporia is provided.

Development and Characterization of an In Vitro Cell-Based Assay to Predict Potency of mRNA-LNP-Based Vaccines.

Messenger RNA (mRNA) vaccines have emerged as a flexible platform for vaccine development. The evolution of lipid nanoparticles as effective delivery vehicles for modified mRNA encoding vaccine antigens was demonstrated by the response to the COVID-19 pandemic. The ability to rapidly develop effective SARS-CoV-2 vaccines from the spike protein genome, and to then manufacture multibillions of doses per year was an extraordinary achievement and a vaccine milestone. Further development and application of this platform for additional pathogens is clearly of interest. This comes with the associated need for new analytical tools that can accurately predict the performance of these mRNA vaccine candidates and tie them to an immune response expected in humans. Described here is the development and characterization of an imaging based in vitro assay able to quantitate transgene protein expression efficiency, with utility to measure lipid nanoparticles (LNP)-encapsulated mRNA vaccine potency, efficacy, and stability. Multiple biologically relevant adherent cell lines were screened to identify a suitable cell substrate capable of providing a wide dose-response curve and dynamic range. Biologically relevant assay attributes were examined and optimized, including cell monolayer morphology, antigen expression kinetics, and assay sensitivity to LNP properties, such as polyethylene glycol-lipid (or PEG-lipid) composition, mRNA mass, and LNP size. Collectively, this study presents a strategy to quickly optimize and develop a robust cell-based potency assay for the development of future mRNA-based vaccines.

New descriptions, synonymies and records in South American Lamiinae(Coleoptera, Cerambycidae).

Two new species are described from Ecuador: Colobothea androwi sp. nov. and Tybalmia diffusa sp. nov. A provisional key to species of Tybalmia Thomson, 1868 is provided. Tybalmia tetrops Bates, 1872 is synonymized with T. mydas (Lucas, 1859), and a new state record in Brazil is provided. Tybalmia orbis Dillon & Dillon, 1945 is synonymized with T. pupillata (Pascoe, 1859), and a new country record is provided.

Phylogeny and diversity of Rigidoporus (Hymenochaetales, Basidiomycota), including three new species from Asia.

Phylogenetic and morphological analyses on Rigidoporus were carried out. The genus Rigidoporus (Hymenochaetales, Basidiomycota), typified by R. microporus (Fr.) Overeem. (synonym Polyporus micromegas Mont.), was established by Murrill in 1905. The genus is mainly characterized by annual to perennial, resupinate, effused-reflexed to pileate or stipitate basidiomata with azonate or concentrically zonate and sulcate upper surface, a monomitic to pseudo-dimitic hyphal structure, simple-septate generative hyphae, and ellipsoid to globose basidiospores. Phylogeny on species of the genus is reconstructed with two loci DNA sequences including the internal transcribed spacer regions and the large subunit. Three new species in Rigidoporus are described and illustrated from Asia, and one new combination in the genus is proposed. The main morphological characteristics of the currently accepted species of Rigidoporus are provided.

Two new species of Haploporus (Polyporales, Basidiomycota) from China and Ecuador based on morphology and phylogeny.

At present, 25 species are accepted in Haploporus and are distributed in Asia, Europe, North America, South America, Australia, and Africa. In this study, two new species, Haploporus ecuadorensis from Ecuador and H. monomitica from China, are described and illustrated based on morphological examination and phylogenetic analyses. H. ecuadorensis is characterized by annual, resupinate basidiomata with pinkish buff to honey yellow hymenophore when dry, round to angular pores of 2-4 per mm, a dimitic hyphal structure with generative hyphae bearing clamp connections, hyphae at dissepiment edge usually with one or two simple septa, the presence of dendrohyphidia and cystidioles, and oblong to ellipsoid basidiospores measuring 14.9-17.9 × 6.9-8.8 µm. Haploporus monomitica differs from other Haploporus species in that it has a monomitic hyphal system and strongly dextrinoid basidiospores. The differences between the new species and morphologically similar and phylogenetically related species are discussed. In addition, an updated key to 27 species of Haploporus is provided.

Two new species of Fistulina (Agaricales, Basidiomycota) from the Northern Hemisphere.

Phylogenetic and morphological analyses on samples of Fistulina from East Asia and North America were carried out, and two new species were described, namely, Fistulina americana and Fistulina orientalis, both previously known as Fistulina hepatica. The former is characterized by lateral stipitate basidiocarps, relatively small pores (7-8 per mm), a monomitic hyphal system with both clamp connections and simple septa, and ellipsoid basidiospores of 4-4.8 × 3-3.3 µm, and the species has been found on Quercus in North-East USA. F. orientalis is characterized by lateral stipitate basidiocarps, very small pores (11-12 per mm) with pruinose dissepiments, a monomitic hyphal system with both clamp connections and simple septa, and ovoid to subglobose basidiospores of 3-4 × 2.7-3 µm, and the species has been found on Castanopsis in East Asia. Phylogenetically, samples of F. americana and F. orientalis form two new lineages nested in the Fistulina clade.

A new contribution to Megasporoporia sensu lato: Six new species and three new combinations.

Megasporoporia sensu lato has recently been intensively studied in China and South America, and four independent clades representing four genera have been recognized phylogenetically. In this study, more samples, mostly from subtropical and tropical Asia, Oceania, and East Africa, are analyzed. A phylogeny based on a 4-gene dataset of sequences (ITS + nLSU + mtSSU + tef) has confirmed the presence of four genera in Megasporoporia sensu lato: Jorgewrightia, Mariorajchenbergia, Megasporia, and Megasporoporia sensu stricto. Six new species, Jorgewrightia austroasiana, Jorgewrightia irregularis, Jorgewrightia tenuis, Mariorajchenbergia subleucoplaca, Megasporia olivacea, and Megasporia sinuosa, are described based on morphology and phylogenetic analysis. Three new combinations are proposed, viz. Jorgewrightia kirkii, Mariorajchenbergia epitephra, and Mariorajchenbergia leucoplaca. To date, 36 species of Megasporoporia sensu lato are accepted and an identification key to these species is provided. In addition, the identification of Dichomitus amazonicus, Dichomitus cylindrosporus, and Megasporoporia hexagonoides is discussed.

Two new forest pathogens in Phaeolus (Polyporales, Basidiomycota) on Chinese coniferous trees were confirmed by molecular phylogeny.

Phaeolus schweinitzii (Fr.) Pat. was originally described in Europe and is considered a common forest pathogen on conifers in the Northern Hemisphere. Our molecular phylogeny based on samples from China, Europe, and North America confirms that P. schweinitzii is a species complex, including six taxa. P. schweinitzii sensu stricto has a distribution in Eurasia; the samples from Northeast and Southwest China are distantly related to P. schweinitzii sensu stricto, and two new species are described after morphological, phylogenetic, and geographical analyses. The species growing on Larix, Picea, and Pinus in Northeast China is described as Phaeolus asiae-orientalis. Another species mostly occurring on Pinus yunnanensis in Southwest China is Phaeolus yunnanensis. In addition, three taxa distributed in North America differ from P. schweinitzii sensu stricto. Phaeolus tabulaeformis (Berk.) Pat. is in Southeast North America, "P. schweinitzii-1" in Northeast North America, and "P. schweinitzii-2" in western North America.

Development of process analytical tools for rapid monitoring of live virus vaccines in manufacturing.

In the development of end-to-end large-scale live virus vaccine (LVV) manufacturing, process analytical technology (PAT) tools enable timely monitoring of critical process parameters (CPP) and significantly guide process development and characterization. In a commercial setting, these very same tools can enable real time monitoring of CPPs on the shop floor and inform harvest decisions, predict peak potency, and serve as surrogates for release potency assays. Here we introduce the development of four advanced PAT tools for upstream and downstream process monitoring in LVV manufacturing. The first tool explores the application of capacitance probes for real time monitoring of viable cell density in bioreactors. The second tool utilizes high content imaging to determine optimum time of infection in a microcarrier process. The third tool uses flow virometry (or nanoscale flow cytometry) to monitor total virus particle counts across upstream and downstream process steps and establishes a robust correlation to virus potency. The fourth and final tool explores the use of nucleic acid dye staining to discriminate between "good" and "damaged" virus particles and uses this strategy to also monitor virus aggregates generated sometimes during downstream processing. Collectively, these tools provide a comprehensive monitoring toolbox and represent a significantly enhanced control strategy for the manufacturing of LVVs.

Descriptions, notes, and a new record in Oideterus Thomson (Coleoptera, Cerambycidae, Prioninae, Anacolini).

The male of Oideterus elegans (Waterhouse, 1880) is described for the first time, female chromatic variations are reported, and the species is recorded from the Ecuadorian department of Napo. Two new species are described from Ecuador: Oideterus leoninus and O. ecuadorensis. The species-group name of O. nigrum Botero, Galileo Santos-Silva, 2019 is corrected. Identification keys to males and females of Oideterus species are provided.

Functional profiling of Covid 19 vaccine candidate by flow virometry.

Vaccine development is a complex process, starting with selection of a promising immunogen in the discovery phase, followed by process development in the preclinical phase, and later by clinical trials in tandem with process improvements and scale up. A large suite of analytical techniques is required to gain understanding of the vaccine candidate so that a relevant immunogen is selected and subsequently manufactured consistently throughout the lifespan of the product. For viral vaccines, successful immunogen production is contingent on its maintained antigenicity and/or infectivity, as well as the ability to characterize these qualities within the context of the process, formulation, and clinical performance. In this report we show the utility of flow virometry during preclinical development of a Covid 19 vaccine candidate based on SARS-CoV-2 spike (S) protein expressed on vesicular stomatitis virus (VSV). Using a panel of monoclonal antibodies, we were able to detect the S protein on the surface of the recombinant VSV virus, monitor the expression levels, detect differences in the antigen based on S protein sequence and after virus inactivation, and monitor S protein stability. Collectively, flow virometry provided important data that helped to guide preclinical development of this vaccine candidate.

SEC coupled with in-line multiple detectors for the characterization of an oncolytic Coxsackievirus.

V937 is an oncolytic virus immunotherapy clinical drug candidate consisting of a proprietary formulation of Coxsackievirus A21 (CVA21). V937 specifically binds to and lyses cells with over-expressed ICAM-1 receptors in a range of tumor cell types and is currently in phase I and II clinical trials. Infectious V937 particles consist of a ∼30 nm icosahedral capsid assembled from four structural viral proteins that encapsidate a viral RNA genome. Rapid and robust analytical methods to quantify and characterize CVA21 virus particles are important to support the process development, regulatory requirements, and validation of new manufacturing platforms. Herein, we describe a size-exclusion chromatography (SEC) method that was developed to characterize the V937 drug substance and process intermediates. Using a 4-in-1 combination of multi-detectors (UV, refractive index, dynamic and static light scattering), we demonstrate the use of SEC for the quantification of the virus particle count, the determination of virus size (molecular weight and hydrodynamic diameter), and the characterization of virus purity by assessing empty-to-full capsid ratios. Through a SEC analysis of stressed V937 samples, we propose CVA21 thermal degradation pathways that result in genome release and particle aggregation.

Two new species of Hymenochaetaceae from tropical Asia and America.

Two new species in Hymenochaetaceae, Fulvifomes acaciae and Pyrrhoderma nigra, are illustrated and described from tropical Asia and America based on morphology and phylogenetic analyses. F. acaciae is characterized by perennial, pileate, and woody hard basidiomata when fresh; ash gray to dark gray, encrusted, concentrically sulcate, and irregularly cracked pileal surface; circular pores of 7-8 per mm with entire dissepiments; a dimitic hyphal system in trama and context; absence of setal element and presence of cystidioles; and broadly ellipsoid, yellowish brown, thick-walled, and smooth basidiospores measuring 5-6 µm × 4-5 µm. P. nigra is characterized by perennial and resupinate basidiomata with dark gray to almost black pore surface when fresh; small and circular pores of 7-9 per mm, a monomitic hyphal system with generative hyphae simple septate, hyphoid setae dominant in subiculum but not in tube trama, and absence of cystidia; and ellipsoid, hyaline, thin-walled basidiospores measuring 4-5 µm × 3-3.6 µm. The differences between the new species and morphologically similar and phylogenetically related species are discussed. Keys to Fulvifomes and Pyrrhoderma have also been provided.

Two new species of Phylloporia (Hymenochaetales) from the Neotropics.

Two new species of Phylloporia, P.crystallina and P.sumacoensis, are described based on 28S ribosomal RNA phylogeny, morphology, host, and geographic distribution. Phylloporiacrystallina is characterized by pileate, perennial basidiomata with a duplex context, small pores 9-10 per mm, a monomitic hyphal system, absence of cystidia and cystidioles, presence of large rhomboid crystals in tube trama, broadly ellipsoid to subglobose basidiospores measuring 2.8-3 × 2-2.3 µm, and growth on angiosperm stump. Phylloporiasumacoensis is characterized by pileate, perennial basidiomata with a duplex context, very small pores 10-12 per mm, a monomitic hyphal system, hyphae at dissepiment edges bearing fine crystals, presence of cystidioles, broadly ellipsoid to subglobose basidiospores measuring 3-3.7 × 2.1-2.8 µm, and growth on living liana.

New species and new geographical records in Ecuadorian Lamiinae (Coleoptera, Cerambycidae).

Three new species of Lamiinae are described from Ecuador: Pseudosparna pallida sp. nov. (Acanthocinini); Acakyra sinuata sp. nov. and Scythropopsis variegata sp. nov. (Acanthoderini). Piriana pygmaea (Bates, 1881) is recorded from Ecuador for the first time. A chromatic variation in Unaporanga lanceolata Martins & Galileo, 2007 is reported, the male is illustrated for the first time, and the species is newly reported from the Ecuadorian department of Pichincha. The morphological differences between Scythropopsis Thomson, 1864, Acakyra Martins & Galileo, 1996, and Irundisaua Martins & Galileo, 2005 are discussed.

Five new species of Nealcidion Monné, 1997 from Ecuador (Coleoptera, Cerambycidae, Lamiinae).

Five new species of Nealcidion Monné, 1997 are described from Ecuador: Nealcidion obliquum sp. nov., Nealcidion flavoviride sp. nov., Nealcidion cephaliferum sp. nov., Nealcidion sublineatum sp. nov., and Nealcidion calcaratum sp. nov. All new species are illustrated, making it easier to recognize the species and to follow the descriptions.

Description of five new species of American Acanthocinini (Coleoptera, Cerambycidae, Lamiinae).

Five new species of Acanthocinini are described: Onalcidion maculatum, from Costa Rica; Lethes x-notatus, from Costa Rica; Leptostylus indistinctus, from Costa Rica; Leptostylus guianensis, from French Guiana; and Trichotithonus thomasvlasaki, from Ecuador.