Biology subjects

Nakano, R.

Publications and source records attributed to Nakano, R..

8 recordsLinked to original sources

bioRxiv2024

A Deep Learning-Based Evaluation for Microscopic Agglutination Test

The Microscopic Agglutination Test (MAT) is widely recognized as the gold standard for diagnosing zoonosis leptospirosis. However, its reliance on examiners subjective evaluations often leads to inconsistent results. To address this limitation, we propose a deep neural network replicating the agglutination rate estimation in MAT. By leveraging a pre-trained DenseNet121, the network parameters are optimized efficiently during training. We validated our approach using an in-house dataset, and experimental results show that the proposed network achieves accurate agglutination rate estimates. In addition, we employed a standard visualization technique to elucidate the decision-making process, revealing that the network captures image features indicative of leptospire abundance. Overall, our findings suggest that deep learning effectively estimates agglutination rates and that enhancing interpretability supports medical experts in understanding the underlying functionality of deep learning models.

microbiology↗

bioRxiv2022

Antiviral effect of candies containing persimmon-derived tannin against SARS-CoV-2 delta strain

Inactivation of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in the mouth has the potential to reduce the spread of coronavirus disease 2019 (COVID-19) because the virus is readily transmitted by dispersed saliva. Persimmon-derived tannin has strong antioxidant and antimicrobial activity owing to its strong adhesiveness to proteins, and it also exhibited antiviral effects against non-variant and alpha variant SARS-CoV-2 in our previous study. In this report, we first demonstrated the antiviral effects of persimmon-derived tannin against the delta variant of SARS-CoV-2 in vitro via the plaque assay method. We then examined the effects of candy containing persimmon-derived tannin. Our plaque assay results show that saliva samples provided by healthy volunteers while they were eating tannin-containing candy remarkably suppressed the virus titers of the SARS-CoV-2 delta variant. In addition, we found that the SARS-CoV-2 viral load in saliva from patients with COVID-19 that was collected immediately after they had eaten the tannin-containing candy was below the level of detection by PCR for SARS-CoV-2. These data suggest that adding persimmon-derived tannin to candy and holding such candy in the mouth is an effective method by which to inactivate the SARS-CoV-2 in saliva, and the application of this approach has potential for inhibiting the transmission of COVID-19.

microbiology↗

bioRxiv2022

Degenerate oligonucleotide primer MIG-seq: an effective PCR-based method for high-throughput genotyping

Multiplexed inter-simple sequence repeats genotyping by sequencing (MIG-seq) is an next-generation sequencing library construction method developed for the analysis of DNA in ecology. Although MIG-seq can generate libraries from low-quality DNA, few polymorphisms can be obtained in species with small genomes. In this study, we developed degenerate oligonucleotide primer MIG-seq (dpMIG-seq) as an effective polymorphism discovery method that allows for variation in the number of polymorphisms while retaining the advantages of MIG-seq, including independence from DNA quality. In dpMIG-seq, a proportion of the simple sequence repeats in the primer sequence of the first PCR in MIG-seq was changed to degenerate oligonucleotides to enable annealing to a wider range of sequences. In tests of several crop species other than wheat, the number of loci that could be sequenced using dpMIG-seq with a data volume of 0.3 gigabases (Gb) was increased compared with that sequenced using MIG-seq. In wheat, the number of polymorphisms obtained via dpMIG-seq was higher than that obtained via MIG-seq when a data volume of about [≥]2 Gb was obtained. In dpMIG-seq, different loci could be sequenced by changing the positions of the degenerate oligonucleotides. By applying dpMIG-seq, we constructed a linkage map consisting of 5,142 markers for the rice inter-subspecies F2 population, and we detected quantitative trait loci for heading date in the regions where known heading-related genes were located. Overall, our results show that dpMIG-seq is a useful tool for the genetic analysis of crop species.

genetics↗

bioRxiv2020

Postharvest properties of ultra-late maturing peach cultivars and their attributions to Melting Flesh (M) locus: Re-evaluation of M locus in association with flesh texture

The postharvest properties of two ultra-late maturing peach cultivars, Tobihaku (TH) and Daijumitsuto (DJ), were investigated. Fruit were harvested at commercial maturity and held at 25{degrees}C. TH exhibited the characteristics of normal melting flesh (MF) peach, including rapid fruit softening associated with an increase in endogenous ethylene production In contrast, DJ did not soften at all during three-week experimental period even though substantial ethylene production was observed. Fruit of TH and DJ were treated with 5000 ppm of propylene, an ethylene analog, continuously for seven days. TH softened rapidly whereas DJ maintained high flesh firmness in spite of an increase in endogenous ethylene production, suggesting that DJ but not TH lacked the ability to be softened in response to endogenous and exogenous ethylene/propylene. DNA-seq analysis showed that tandem endo-polygalacturonase (endoPG) genes located at melting flesh (M) locus, Pp-endoPGM (PGM) and Pp-endoPGF (PGF), were deleted in DJ. The endoPG genes at M locus are known to control flesh texture of peach fruit, and it was suggested that the non-softening property of DJ is due to the lack of endoPG genes. On the other hand, TH possessed an unidentified M haplotype that is involved in determination of MF phenotype. Structural identification of the unknown M haplotype, designated as M0, through comparison with previously reported M haplotypes revealed distinct differences between PGM on M0 haplotype (PGM-M0) and PGM on other haplotypes (PGM-M1). Peach M haplotypes were classified into four main haplotypes: M0 with PGM-M0; M1 with both PGM-M1 and PGF; M2 with PGM-M1; and M3 lacking both PGM and PGF. Re-evaluation of M locus in association with MF/non-melting flesh (NMF) phenotypes in more than 400 accessions by using whole genome shotgun sequencing data on database and/or by PCR genotyping demonstrated that M0 haplotype was the common haplotype in MF accessions, and M0 and M1 haplotypes were dominant over M2 and M3 haplotypes and co-dominantly determined the MF trait. It was also assumed on the basis of structural comparison of M haplotypes among Prunus species that the ancestral haplotype of M0 diverged from those of the other haplotypes before the speciation of Prunus persica.

plant biology↗

bioRxiv2019

Aberrant expression of the COX2/PGE2 axis is induced by activation of the RAF/MEK/ERK pathway in BRAFV595E canine urothelial carcinoma

Cancer-promoting inflammation is an important event in cancer development. The cyclooxygenase 2 (COX2)/prostaglandin E2 (PGE2) axis is a prominent inducer of inflammation. Canine urothelial carcinoma (cUC) uniquely overexpresses PGE2 and is often managed well with COX inhibitor monotherapy. In most cases, cUC tissue harbours homologous human BRAFV600E mutation, which causes aberrant activation of the RAF/MEK/ERK pathway in human cancer. However, mechanisms underlying aberrant PGE2 production and the importance of the BRAF mutation remain unclear. We show that activation of the RAF/MEK/ERK pathway in BRAF mutant cUC cells leads to COX2 overexpression and PGE2 production. Drug screening revealed that treatment with inhibitors of the arachidonic acid cascade (FDR<0.086), RAF/MEK/ERK pathway (FDR<0.067), and p38/JNK pathway (FDR<0.067) significantly reduced PGE2 production in cUC cells. We also validated the association between RAF/MEK/ERK pathway activation and COX2/PGE2 overexpression in BRAF mutant cUC cells using protein detection techniques. In histochemical analysis, BRAF mutant cUC tissue showed higher COX2 expression. Therefore, the driver mutation in the BRAF gene probably promotes tumour-promoting inflammation. These findings would benefit dogs suffering from cUC and can be extrapolated to human cancer. Finally, cUC can serve as a valuable model to elucidate the association between driver mutations and tumour-promoting inflammation.

cancer biology↗

bioRxiv2019

Auxin-mediated rapid degradation of selective proteins in hippocampal neurons

Genetic manipulation of protein levels is a promising approach to identify the function of a specific protein in living organisms. Previous studies demonstrated that the auxin-inducible degron (AID) strategy provides rapid and reversible degradation of various proteins in fungi and mammalian mitotic cells. In this study, we employed this technology to postmitotic neurons to address whether the AID system could be applied to the nervous system. Using adeno-associated viruses, we simultaneously introduced EGFP fused with an AID tag, and an F-box family protein, TIR1 from Oryza sativa (OsTIR1) into hippocampal neurons. In dissociated hippocampal neurons, EGFP fluorescence signals rapidly decreased when adding a plant hormone, auxin. Further, auxin-induced EGFP degradation was also observed in hippocampal acute slices. Taken together, these results open the door for neuroscientists to manipulate protein expression levels by the AID-system in a temporally-controlled manner.

neuroscience↗

bioRxiv2019

Decoding neuronal diversity by single-cell Convert-seq

The conversion of cell fates is controlled by hierarchical gene regulatory networks (GRNs) that induce remarkable alterations in cellular and transcriptome states. The identification of key regulators within these networks from myriad of candidate genes, however, poses a major research challenge. Here we present Convert-seq, combining single-cell RNA sequencing (scRNA-seq) and pooled ectopic gene expression with a new strategy to discriminate sequencing reads derived from exogenous and endogenous transcripts. We demonstrate Convert-seq by associating hundreds of single cells during reprogramming of human fibroblasts to induced neurons (iN) with exogenous and endogenous transcriptional signatures. Convert-seq identified GRNs modulating the emergence of developmental trajectories and predicted combinatorial activation of exogenous transcription factors controlling iN subtype specification. Functional validation of iN subtypes generated by novel combinations of exogenous transcription factors establish Convert-seq as a broadly applicable workflow to rapidly identify key transcription factors and GRNs orchestrating the reprogramming of virtually any cell type.

neuroscience↗

bioRxiv2019

Inoculum size effects in blaIMP-6 and plasmid-mediated quinolone resistance gene-positive Enterobacteriaceae in Japan

The aim of this study was to examine the resistance genes in clinical isolates which produced IMP-6 type metallo-{beta}-lactamase lactamase (MBL) and had mildly reduced susceptibilities to levofloxacin and/or amikacin. The inoculum size effect was also assessed. A total of 14 Enterobacteriaceae isolates (2 Escherichia coli and 12 Klebsiella pneumoniae) which produced IMP-6 MBL, and had mild increases in their MICs for levofloxacin and amikacin were examined. Thirteen out of 14 isolates harbored CTX-M-2, with the remaining isolate co-harboring CTX-M-2 and CTX-M-1 as ESBLs. All isolates carried one or more PMQRs; aac(6')-Ib-cr was the most prevalent (92.8%), followed by oqxA (64.3%), qnrS (42.9%), oqxAB (21.4%), and qnrB (14.3%). The inoculum size effects were significant in all strains for meropenem, 13 for imipenem, 7 for levofloxacin, and 3 for amikacin. Conjugation was successfully performed with 8 isolates and 11 strains were obtained. Eleven of the experimental strains (100%), and 8 strains (72.7%) showed inoculum size effects for meropenem and imipenem, respectively. No inoculum size effect was seen for levofloxacin. Four strains harbored qnr genes and 2 strains harbored qnr genes and QRDR mutations concurrently. blaIMP-6 positive Enterobacteriaceae with mildly reduced susceptibilities to levofloxacin and/or amikacin also harbored at least one plasmid-mediated drug resistance gene. These represent an unrecognized threat, capable of compromising the in vitro activity of many classes of antimicrobial agents. We conclude that IMP-6 MBL plays an important role in decreasing the MIC for carbapenems, whereas qnr does not for levofloxacin.

microbiology↗