ScRNA-Seq study of neutrophils reveals vast heterogeneity and breadth of inflammatory responses in severe COVID-19 patients

Severe cases of COVID-19 are characterized by dysregulated immune responses, but specific mechanisms contributing to the most severe outcomes remain unclear. Neutrophils are the most abundant leukocyte population in human hosts and reach markedly high numbers during severe COVID-19. However, a detailed examination of their responses has been largely overlooked in the COVID-19 literature to date. Here, we report for the first time a dedicated study of neutrophil responses using single-cell RNA sequencing (scRNA-Seq) of fresh leukocytes from 11 hospitalized adult patients with mild and severe COVID-19 disease and 5 healthy controls. We observed that neutrophils display a pronounced inflammatory profile, with dramatic disruption of predicted cell-cell interactions as the severity of the disease increases. We also identified unique mature and immature neutrophil subpopulations based on transcriptomic profiling, including an antiviral phenotype, and changes in the proportion of each population linked to the severity of the disease. Finally, pathway analysis revealed increased markers of oxidative phosphorylation and ribosomal genes, along with downregulation of many antiviral and host defense pathway genes during severe disease compared to mild infections. Collectively, our findings indicate that neutrophils are capable of mounting effective antiviral defenses but adopt a form of immune dysregulation characterized by excess cellular stress, thereby contributing to the pathogenesis of severe COVID-19.

Distribution of GC-rich heterochromatin and ribosomal genes in three fungus-farming ants (Myrmicinae, Attini, Attina): insights on chromosomal evolution

Cytogenetic studies on fungus-farming ants have shown remarkable karyotype diversity, suggesting different chromosomal rearrangements involved in karyotype evolution in some genera. A notable cytogenetic characteristic in this ant group is the presence of GC-rich heterochromatin in the karyotypes of some ancient and derivative species. It was hypothesized that this GC-rich heterochromatin may have a common origin in fungus-farming ants, and the increase in species studied is important for understanding this question. In addition, many genera within the subtribe Attina have few or no cytogenetically studied species; therefore, the processes that shaped their chromosomal evolution remain obscure. Thus, in this study, we karyotyped, through classical and molecular cytogenetic techniques, the fungus-farming ants Cyphomyrmex transversus Emery, 1894, Sericomyrmex maravalhas Ješovnik et Schultz, 2017, and Mycetomoellerius relictus (Borgmeier, 1934), to provide insights into the chromosomal evolution in these genera and to investigate the presence the GC-rich heterochromatin in these species. Cyphomyrmex transversus (2n = 18, 10m + 2sm + 6a) and S. maravalhas (2n = 48, 28m + 20sm) showed karyotypes distinct from other species from their genera. Mycetomoellerius relictus (2n = 20, 20m) presented the same karyotype as the colonies previously studied. Notably, C. transversus presented the lowest chromosomal number for the genus and a distinct karyotype from the other two previously observed for this species, showing the existence of a possible species complex and the need for its taxonomic revision. Chromosomal banding data revealed GC-rich heterochromatin in all three species, which increased the number of genera with this characteristic, supporting the hypothesis of a common origin of GC-rich heterochromatin in Attina. Although a single chromosomal pair carries rDNA genes in all studied species, the positions of these rDNA clusters varied. The rDNA genes were located in the intrachromosomal region in C. transversus and M. relictus, and in the terminal region of S. maravalhas. The combination of our molecular cytogenetic data and observations from previous studies corroborates that a single rDNA site located in the intrachromosomal region is a plesiomorphic condition in Attina. In addition, cytogenetic data obtained suggest centric fission events in Sericomyrmex Mayr, 1865, and the occurrence of inversions as the origin of the location of the ribosomal genes in M. relictus and S. maravalhas. This study provides new insights into the chromosomal evolution of fungus-farming ants.

Cinnamomum japonicum Anthracnose Caused by Colletotrichum fioriniae in Sichuan, China

The Cinnamomum japonicum Sieb is widely cultivated in urban in China. It’s used to make essential, lubricant, soap, and waterproof timber. In September 2019, This new leaf spot was discovered in Chengdu city (30°05′to 31°26′N, 102°54′to 104°53′E), with approximately 61.20% disease incidence. The symptoms started to occur from May to June, the worst from August to September. Firstly, the typical symptom showed round or oval, brown, and slightly sunken necrotic lesions. Gradually, the necrotic lesions increased in number, and expanded; under humid conditions the central part of the spots became black and ruptured, with orange conidial masses emerged at the margin of lesions. Finally, the leaves turn yellow and fall off. Infected tissues from ten samples were cut into small pieces 2 × 2 mm, surface sterilized for 30 s in 3% sodium hypochlorite, 60 s in 75% ethanol, rinsed three times in sterile water, placed onto potato dextrose agar (PDA) amended with streptomycin sulfate (50 μg/mL), and incubated at 25°C in a dark. Finally, 8 typical isolates exhibited the morphology described as C. fioriniae (Amelie Grammen et al. 2019). After 5 days, the colony diameter reached 28.6 to 41.2 mm and had white to light grey aerial mycelium, but was pink at the base. Orange conidia masses formed after 6-7 days, conidia were oval, slender and fusiform with acute ends (Figure 1e, f), measuring 8.3 to 19.6 × 2.9 to 7.1 μm (average: 13.9 × 4.8 μm) (Tashiro et al. 2018; Chechi et al. 2019). For molecular identification, DNA was extracted from 8 fungal colonies using a plant genomic DNA extraction kit (Solarbio, Beijing). The 5.8S nuclear ribosomal genes with the two flanking internal transcribed spacer (ITS), the glyceraldehyde-3-phosphate dehydrogenase (GAPDH), and partial sequences of the actin (ACT), chitin synthase 1 (CHS-1), beta-tubulin (TUB2), histone3 (HIS3), calmodulin (CAL) and glutamine synthetase (GS) genes were amplified and sequenced using the primer pairs ITS1/ITS4 (White et al. 1990), GDF/GDR (Templeton et al. 1992), ACT-512F/ACT-783R, CAL 228F/CAL 737R (Carbone et al. 1999), CHS-354R/CHS-79F (Weir et al. 2012), TUB1F/Bt2bR, CYLH3F/CYLH3R (Crous et al. 2004), and GSF1/GSR1 (Liu et al. 2015), respectively. Sequences were deposited in GenBank (ITS: MT466533, GAPDH: MT460415, ACT: MT460414, CAL: MT954332, CHS-1: MT954330, TUB2: MT460416, HIS3: MT954331, and GS: MT460417). BLAST analysis showed >99.4% identity with several reference sequences of Colletotrichum fioriniae strain CBS 128517 and strain EHS58 (teleomorph of Glomerella fioriniae) previously deposited in GenBank. The conidial suspension (1 × 107 conidia/mL) collected from PDA cultures with 0.05% Tween 80 buffer was used for inoculation by spraying leaves of 5-year-old C. japonicum plants for pathogenicity test. Ten leaves of each plant (10 pots in total) were inoculated with spore suspensions (Approximately 500 μL per leaf). An equal number of control leaves were sprayed with 0.05% Tween 80 buffer to serve as a control. Twenty days later, the inoculated plants showed the similar symptoms as the original diseased plants but the controls remained asymptomatic. The C. fioriniae was re-isolated from the infected leaves and identified by morphological characteristics and DNA sequence analysis. The pathogenicity test was repeated three times with similar results, confirming Koch’s postulates. To our knowledge, this is the first report of C. japonicum anthracnose caused by C. fioriniae in China.

Qualitative and Quantitative Characteristics of Soil Microbiome of Barents Sea Coast, Kola Peninsula

The soil microbiome of the Barents Sea coast of the Kola Peninsula is here characterized for the first time. The content of copies of ribosomal genes of archaea, bacteria, and fungi was determined by real-time PCR. Reserves and structure of biomass of soil microorganisms such as total biomass of fungi and prokaryotes, length and diameter of mycelium of fungi and actinomycetes, proportion of mycelium in biomass, number of spores and prokaryotic cells, proportion of small and large fungal propagules, and morphology of mycobiota spores were determined. The largest number of ribosomal gene copies was found for bacteria (from 6.47 × 109 to 3.02 × 1011 per g soil). The number of copies of ribosomal genes of fungi and archaea varied within 107–109 copies of genes/g soil. The biomass of microorganisms (prokaryotes and fungi in total) varied from 0.023 to 0.840 mg/g soil. The share of mycobiota in the microbial biomass ranged from 90% to 97%. The number of prokaryotes was not large and varied from 1.87 × 108 to 1.40 × 109 cells/g of soil, while the biomass of fungi was very significant and varied from 0.021 to 0.715 mg/g of soil. The length of actinomycete mycelium was small—from 0.77 to 88.18 m/g of soil, as was the length of fungal hyphae—an order of magnitude higher (up to 504.22 m/g of soil). The proportion of fungal mycelium, an active component of fungal biomass, varied from 25% to 89%. Most (from 65% to 100%) of mycobiota propagules were represented by specimens of small sizes, 2–3 microns. Thus, it is shown that, despite the extreme position on the mainland land of Fennoscandia, local soils had a significant number of microorganisms, on which the productivity of ecosystems largely depends.

A new species of subgenus Asonicogryllus He, 2019 from Yunnan (Orthoptera: Gryllidae: Gryllinae)

We studied the relationship of the genera Loxoblemmus Saussure, 1877 and Asonicogryllus He, 2019 by reconstructing the phylogenetic tree based on 2 mitochondrial genes and 2 ribosomal genes. The result shows that Loxoblemmus is paraphyletic. Thus, we place Asonicogryllus as a subgenus of Loxoblemmus. Meanwhile, we report a new species of Asonicogryllus, L. (A.) bronzus He sp. nov. The new species is similar to L. (A.) kwanghua, but it has well developed forewings and hindwings. The type specimens are deposited in the Museum of Biology, East China Normal University (ECNU).

OTEH-1. Alternative RNA splicing modulates composition of ribosomes and determines spatial phenotype of glioblastoma cells

Glioblastoma (GBM) is an extremely heterogeneous tumor and its different regions are populated with phenotypically distinct types of cancer cells. However, it is still unclear how multiple GBM populations arise from the originally homogenous group of tumor precursor cells. Here we showed that GBM cells from the core and edge of the tumor have different composition of ribosomes due to the alternative RNA splicing of multiple ribosomal genes with highest differences observed for RPL22L1. We found that cells at the edge of the tumor express classical isoform of RPL22L1 (RPL22L1a) while core cells have a novel RPL22L1b isoform. RPL22L1b appears due to low pH condition at the core of the tumor. It allows cells to survive during acidosis, promotes more aggressive phenotype in vivo and correlate with worse patient outcome. Mechanistically, RPL22L1b binds to lncRNA MALAT1 in the nucleus and induces its degradation enhancing stemness of GBM cells. On the other hand, RPL22L1a interacts with ribosomes in cytoplasm and upregulates p53 translation favoring less aggressive edge phenotype of GBM. The splicing switch between RPL22L1 isoforms is regulated by SRSF4 proteins. We identified a small molecule compound that inhibits SRSF4 and impairs splicing of RPL22L1, inducing apoptosis of GBM cells and decreasing tumor growth in vivo. Altogether, our data unraveled the mechanism by which less aggressive edge-like GBM cells acquire more malignant core-like phenotype during tumor growth. It may also explain discrepancies between proteome and transcriptome of GBM cell populations. Targeting this pathway may help to decrease tumor heterogeneity and eliminate therapy resistant cells at the tumor core. Work was supported by the Russian Science Foundation grant 19-44-02027.

Prevalence and molecular identification of Nematodirus helvetianus in camels in Iraq

Background and Aim: Camels from the central part of Iraq are infected with multiple parasitic diseases that have an economic impact by decreasing meat and milk production. This study aimed to evaluate Nematodirus spp. in camels (Camelus dromedarius). Materials and Methods: The study animals consisted of camels slaughtered in the central area of Iraq at the Al-Najaf slaughterhouse. All ages and sexes of camels were examined. Worms were recovered and identified microscopically. For molecular characterization, two Iraqi Nematodirus spp. partial ribosomal genes (ITS1 and ITS2) were sequenced and submitted to the NCBI database. Results: Of 160 camels tested, 29 were infected with Nematodirus spp. (18.13%). Twenty-one nematodes containing the Nematodirus genes were identified in the small intestines of naturally infected camels. BLAST analysis revealed 88.1% sequence similarity with that of Nematodirus helvetianus isolated in China and 87.2% similarity with N. helvetianus isolated in the United States. Conclusion: The prevalence of N. helvetianus warrants the use of anti-helminthic drugs for these animals and a rationale for future control strategies to prevent the transmission of this infection to other livestock.

Comparative Cytogenetic Analysis of Three Eumeninae Species (Hymenoptera, Vespidae)

Eumeninae represents the largest subfamily within Vespidae, with 3,600 species described. Of these, only 18 have been cytogenetically analysed. In the present study, we used both classical and molecular techniques to characterise and compare the karyotypes of 3 Eumeninae species, namely, <i>Ancistrocerus</i> sp., <i>Pachodynerus grandis,</i> and <i>Pachodynerus nasidens</i>. <i>Ancistrocerus</i> sp. presented a haploid chromosome number of n = 12, with the first 2 chromosomes of the karyotype being almost entirely heterochromatic and much larger than the remaining chromosomes. The 2 <i>Pachodynerus</i> species presented the same chromosome number (n = 11 and 2n = 22) but displayed different karyotypic formulae<i>.</i> Additionally, chromosomal polymorphisms were observed in the analysed <i>P. nasidens</i> female. In the 3 species, heterochromatin was located in one of the chromosome arms. Fluorochrome staining revealed a balanced composition of AT and GC bases within the chromatin for each of the 3 species, except for few regions that were visibly GC-rich. All species had a single 18S rDNA site that co-localised with GC-rich regions; however, this localisation varied from species to species and not all GC-rich regions corresponded to ribosomal genes. Based on the cytogenetic data obtained here, we discuss the possible numerical/structural rearrangements that may be involved in the karyotypic evolution of the 3 studied species. In addition to the first description of the molecular cytogenetic characteristics of the Eumeninae subfamily and the genus <i>Pachodynerus</i>, this study also provides a relevant contribution towards the discussion of chromosomal evolution in Eumeninae wasps.

A new genus and species of tachinid flies from Iran (Diptera, Tachinidae, Goniini)

Susamyia Ziegler & Gilasian gen. nov. and its type species Susamyia mira Ziegler & Gilasian gen. et sp. nov. are described from southwestern Karkheh National Park and southeastern Jazmourian Wetland of Iran. Drawings of terminalia and head of the male as well as images of habitus, head and egg are provided. Susamyia gen. nov. is incorporated into the identification key to the Palearctic tachinid genera of Tschorsnig & Richter (1998). The systematic position of Susamyia gen. nov. in the tribe Goniini (Exoristinae) is discussed in light of molecular analyses based on the ribosomal genes 16S, 18S and 28S, and the egg type of females. The morphology of the microtype eggs was studied and compared with that of other Goniini, but proved too unreliable to be included in the phylogenetic analysis. Susamyia gen. nov. is believed to be most closely related to the goniine genera Gymnophryxe Villeneuve, 1922, Palesisa Villeneuve, 1929 and Prosopodopsis Townsend, 1926.

rDNA nascent transcripts promote a unique spatial organization during mouse early development

During the first cell cycles of the early development, the chromatin of the embryo is highly reprogrammed alongside that embryonic genome starts its own transcription. The spatial organization of the genome is a major process that contributes to regulating gene transcription in time and space, however, it is poorly studied in the context of early embryos. To study the cause and effect link between transcription and spatial organization in embryos, we focused on the ribosomal genes, that are first silent and begin to transcribe during the 2-cell stage in mouse. We demonstrated that ribosomal sequences are spatially organized in a very peculiar manner from the 2-cell to the 16-cell stage with transcription and processing of ribosomal RNAs excluding mutually. Using drugs inhibiting the RNA polymerase I, we show that this organization, totally different from somatic cells, depends on an active transcription of ribosomal genes and induces a unique chromatin environment that favors major satellite sequences transcription after the 4-cell stage.