The Tick Microbiota Dysbiosis Promote Tick-Borne Pathogen Transstadial Transmission in a Babesia microti–Infected Mouse Model

Ticks are obligate hematophagous ectoparasites. They are important vectors for many pathogens, of both medical and veterinary importance. Antibiotic residues in animal food are known, but very little is known about the effects of antibiotic residues in animals on the microbiome diversity of ticks and tick-borne pathogen transmission. We used a Haemaphysalis longicornis–infested mouse model to evaluate the effect of antibiotic usage on tick microbiome. Nymphal ticks were fed on an antibiotic cocktail-treated or water control mice. Adult ticks molted from nymphs fed on the antibiotic cocktail-treated mouse had a dysbiosed microbiota. Nymphal ticks were also fed on a B. microti–infected mice that had been treated with antibiotic cocktail or water. We found that the B. microti infection in adult ticks with a dysbiosed microbiota (44.7%) was increased compared with the control adult ticks (24.2%) by using qPCR targeting 18S rRNA gene. This may increase the risk of tick-borne pathogens (TBPs) transmission from adult ticks to a vertebrate host. These results show that an antibiotic-treated mouse can induce tick microbiota dysbiosis. Antibiotic treatment of B. microti-infected mouse poses the possibility of increasing transstadial transmission of B. microti from the nymph to the adult H. longicornis. These findings suggest that B. microti transmission may be exacerbated in high antibiotic usage areas.

O-083 Non-invasive, label-free optical analysis to detect aneuploidy within the inner cell mass of the preimplantation embryo

Study question Can we separate between control and reversine-treated cells within the inner cell mass (ICM) of the mouse preimplantation embryo by using label-free and non-invasive hyperspectral microscopy? Summary answer Hyperspectral microscopy is able to discern between control and reversine-treated cells using cellular autofluorescence in the complete absence of fluorescence tags. What is known already Embryo mosaicism (containing cells that are euploid (46 chromosomes) and aneuploid (deviation from the expected number of chromosomes)) affects up to 17.3% of human blastocyst embryos. Current diagnosis of aneuploidy in the IVF clinic involves a biopsy of trophectoderm (TE) cells or spent media followed by sequencing. In some blastocyst embryos these approaches will fail to diagnose of the proportion of aneuploid cells within the fetal lineage (ICM). Study design, size, duration The impact of aneuploidy on cellular metabolism was assessed by using cellular autofluoresence and hyperspectral microscopy (broad spectral profile). Two models were employed: (i) Primary human fibroblast cells with known karyotypes (4-6 independent replicates, euploid n = 467; aneuploid n = 969) and reversine induced aneuploidy in mouse embryos (5-8 independent replicates, 30-44 cells per group). Both models were subjected to hyperspectral imaging to quantify native cell fluorescence. Participants/materials, setting, methods The human model is comprised of euploid (male and female) and aneuploid (triploid and trisomies: 13, 18, 21, XXX, and XXY) primary human fibroblast cells. For the mouse model, we treated embryos with reversine, a reversible spindle assembly checkpoint inhibitor, during the 4- to 8-cell division. Individual blastomeres were dissociated from control and reversine treated 8-cell embryos. Blastomeres were either imaged directly or used to generate chimeric blastocysts with differing ratios of control:reversine-treated cells. Main results and the role of chance Following unsupervised linear unmixing, the relative abundance of metabolic cofactors was quantified: reduced nicotinamide adenine dinucleotide (NAD(P)H) and flavins with the subsequent calculation of the optical redox ratio (ORR: Flavins/[NAD(P)H + Flavins]). Primary human fibroblast cells displayed an increase in the relative abundance of NAD(P)H with a decrease in flavins, leading to a significant reduction in the ORR for aneuploid cells (P < 0.05). The mouse embryos displayed an identical trend as the human model between control and reversine-treated embryos. Mathematical algorithms were applied and able to distinguish between (i) euploid and aneuploid primary human fibroblast cells, (ii) control and reversine-treated mouse blastomeres and (iii) chimeric blastocysts with differing ratios of control and reversine-treated cells. The accuracy of these separations was supported by receiver operating characteristic curves with areas under the curve. We also showed that hyperspectral imaging of the preimplantation embryo does not impact on embryo developmental competence, pregnancy outcome and offspring health in a mouse model. We believe the role of chance is low as both human somatic cells and mouse embryos showed a consistent shift in cellular metabolism in response to human fibroblast cells that are aneuploid and reversine treated mouse embryos. Limitations, reasons for caution Further validation of our approach could include sequencing of the ICM of individual blastocysts to determine the proportion of aneuploid cells in ICM and correlate this with the metabolic profile obtained through hyperspectral imaging. Wider implications of the findings With hyperspectral imaging able to discriminate between (i) euploid and aneuploid human fibroblast cells and (ii) control and reversine-treated mouse embryos, this could be an accurate, non-invasive and label-free optical imaging approach to assess mosaicism within the ICM of mouse embryos, potentially leading to a new diagnostic tool for embryos. Trial registration number Not applicable

Overexpression of mm9_circ_013935 alleviates renal inflammation and fibrosis in diabetic nephropathy via the miR-153-3p/NFIC axis

Background: Circ-RBM4 (mm9_circ_013935) has been revealed to be low-expressed in the renal tissues of diabetic nephropathy (DN) mice while its underlying regulatory mechanism remains unexplored. Methods: The high glucose (HG)-treated mouse podocytes were used to establish the DN cell models. A CCK-8 assay was used to examine the viability of mouse podocytes. The expression of proteins related to fibrosis (Collagen I, Collagen III, fibronectin) was detected using western blot. The concentration of inflammation cytokines (TNF-α, IL-1β, IL-8) in mouse podocytes was assessed by ELISA. The interaction between genes was explored by luciferase reporter assays. Results: HG treatement decreased the viability and elevated the expression of fibrosis and inflammation factors in mouse podocytes. Circ-RBM4 expression was downregulated in HG-treated mouse podocytes. Circ-RBM4 overexpression reversed HG-induced increase in levels of proteins related to fibrosis and the concentration of inflammation factors. MiR-153-3p was revealed to bind with circ-RBM4 and directly targeted nuclear factor I/C (NFIC) in mouse podocytes. Rescue assays indicated that circ-RBM4 attenuated HG-induced fibrosis and inflammation response in mouse podocytes by inhibiting miR-153-3p expression or upregulating NFIC expression. Conclusion: Circ-RBM4 alleviated the renal inflamation and renal fibrosis in DN by targeting the miR-153-3p/NFIC axis.

Therapeutic Effects of a Neoantigen Vaccine With an Immune Checkpoint Inhibitor in a Cisplatin-Treated Mouse Model

Background: Cancer immunotherapy is widely used as a treatment for cancer that works by improving the immune system with fewer side effects than conventional methods. Neoantigen vaccines are one form of immunotherapy that use cancer-specific neoantigens that are extracted from cancer patients and are not recognized by normal cells in the immune system.Methods: In this study, mutant genes of 4T1 mouse breast cancer cells were identified by direct sequence analysis using tumor-specific MHC I (Major Histocompatibility Complex) or MHC II epitopes through in vivo experiments. Results: The neoantigen vaccine with mutant CD4+ or CD8+ T cell-reactive neoantigen peptides was shown to inhibit tumor growth, increase long-term survival, and induce the secretion of IFN-γ (Interferon gamma) in the cisplatin-treated mouse models. In particular, mutant CD4+ T cell neoantigen peptides induced full potential anti-tumor effects, whereas dual treatment with CD4+ (Cluster of differentiation 4) and CD8+ (Cluster of differentiation 8) T cell neoantigen peptides increased the suppression of tumor growth. Moreover, the combination of neoantigen vaccine with mutant CD4+ T cell neoantigen peptide and anti-PD-L1 (Programmed death-ligand 1) as an immune checkpoint inhibitor (ICI) has been shown to have synergistic therapeutic effects in cisplatin-treated mouse models. Conclusion: This study, therefore, proved that cancer cell-derived neoantigens have great potential to induce immunogenic responses and cancer treatment effects, along with synergistic efficiency when applied to various combinational therapies. Through the methods that were used in our experiments, we could contribute to the development of new adjuvants for evaluating efficacy, discovering unfound neoantigens, and investigating immune checkpoint blockade antibodies for non-clinical studies.

Complete Genome Sequence of Lactobacillus johnsonii NCK2677, Isolated from Mice

ABSTRACT We report the closed genome sequence of a Lactobacillus johnsonii strain (NCK2677) that was isolated from a cefoperazone-treated mouse model designed for the study of Clostridioides difficile infection. Illumina and Nanopore sequencing reads were assembled into a circular 1,951,416-bp chromosome with a G+C content of 34.7%, containing 1,865 genes.