[Transcriptional regulation mechanism of differential accumulation of flavonoids in different varieties of Lonicera macranthoides based on metabonomics and transcriptomics].

This study aims to explore the molecular regulatory mechanism of the differential accumulation of flavonoids between 'Xianglei' and the wild type of Lonicera macranthoides. The flowers, stems, and leaves of the two varieties of L. macranthoides were collected. Ultra-performance liquid chromatography-mass spectrometry(UPLC-MS) and high-throughput sequencing(RNA-seq) were employed to screen out the differential flavonoids, key differentially expressed genes(DEGs) and transcription factors(TFs). Fourteen DEGs were randomly selected for verification by qRT-PCR. The results showed that a total of 17 differential flavonoids were obtained, including naringin chalcone, apigenin, and quercetin. The transcriptomic analysis predicted 19 DEGs associated with flavonoids, including 2 genes encoding chitin synthase(CHS) and 3 genes encoding chalcone isomerase(CHI). The regulatory network analysis and weighted gene co-expression network analysis(WGCNA) screen out the key enzyme genes CHS1, FLS1, and HCT regulating the accumulation of flavonoids. MYB12 and LBD4 may be involved in the biosynthesis of flavonoids by regulating the expression of key enzyme genes CHS1, FLS1, and HCT. The qRT-PCR and RNA-seq results were similar regarding the expression patterns of the 14 randomly selected DEGs. This study preliminarily analyzed the transcriptional regulatory mechanism for the differential accumulation of flavonoids in the two varieties of L. macranthoides and laid a foundation for further elucidating the regulatory effects of key enzyme genes and TFs on the accumulation of flavonoids.

Swin Unet3D: a three-dimensional medical image segmentation network combining vision transformer and convolution.

BACKGROUND: Semantic segmentation of brain tumors plays a critical role in clinical treatment, especially for three-dimensional (3D) magnetic resonance imaging, which is often used in clinical practice. Automatic segmentation of the 3D structure of brain tumors can quickly help physicians understand the properties of tumors, such as the shape and size, thus improving the efficiency of preoperative planning and the odds of successful surgery. In past decades, 3D convolutional neural networks (CNNs) have dominated automatic segmentation methods for 3D medical images, and these network structures have achieved good results. However, to reduce the number of neural network parameters, practitioners ensure that the size of convolutional kernels in 3D convolutional operations generally does not exceed [Formula: see text], which also leads to CNNs showing limitations in learning long-distance dependent information. Vision Transformer (ViT) is very good at learning long-distance dependent information in images, but it suffers from the problems of many parameters. What's worse, the ViT cannot learn local dependency information in the previous layers under the condition of insufficient data. However, in the image segmentation task, being able to learn this local dependency information in the previous layers makes a big impact on the performance of the model. METHODS: This paper proposes the Swin Unet3D model, which represents voxel segmentation on medical images as a sequence-to-sequence prediction. The feature extraction sub-module in the model is designed as a parallel structure of Convolution and ViT so that all layers of the model are able to adequately learn both global and local dependency information in the image. RESULTS: On the validation dataset of Brats2021, our proposed model achieves dice coefficients of 0.840, 0.874, and 0.911 on the ET channel, TC channel, and WT channel, respectively. On the validation dataset of Brats2018, our model achieves dice coefficients of 0.716, 0.761, and 0.874 on the corresponding channels, respectively. CONCLUSION: We propose a new segmentation model that combines the advantages of Vision Transformer and Convolution and achieves a better balance between the number of model parameters and segmentation accuracy. The code can be found at https://github.com/1152545264/SwinUnet3D .

[Mining and identification of members of MYB transcription factor family in Lonicera macranthoides].

As a large family of transcription factors, the MYB family plays a vital role in regulating flower development. We studied the MYB family members in Lonicera macranthoides for the first time and identified three sequences of 1R-MYB, 47 sequences of R2R3-MYB, two sequences of 3R-MYB, and one sequence of 4R-MYB from the transcriptome data. Further, their physicochemical properties, conserved domains, phylogenetic relationship, protein structure, functional information, and expression were analyzed. The results show that the 53 MYB transcription factors had different conserved motifs, physicochemical properties, structures, and functions in wild type and 'Xianglei' cultivar of L. macranthoides, indicating their conservation and diversity in evolution. The transcript level of LmMYB was significantly different between the wild type and 'Xianglei' cultivar as well as between flowers and leaves, and some genes were specifically expressed. Forty-three out of 53 LmMYB sequences were expressed in both flowers and leaves, and 9 of the LmMYB members showed significantly different transcript levels between the wild type and 'Xianglei' cultivar, which were up-regulated in the wild type. The results provide a theoretical basis for further studying the specific functional mechanism of the MYB family.

MAIT Cells in Respiratory Viral Infections in Mouse and Human.

Mucosal associated invariant T (MAIT) cells were first identified as specific for bacterial, mycobacterial, and fungal organisms, which detect microbially-derived biosynthetic ligands presented by MHC-related protein 1 (MR1). More recently two unexpected, additional roles have been identified for these ancient and abundant cells: a TCR-depen-dent role in tissue repair and a TCR-independent role in antiviral host defence. Data from several classes of viral disease shows their capability for activation by the cytokines interleukin (IL)-12, IL-15, IL-18, and type I interferon. MAIT cells are abundant at mucosal surfaces, particularly in the lung, and it seems likely a primary reason for their striking evolutionary conservation is an important role in early innate defence against respiratory infections, including both bacteria and viruses. Here we review evidence for their TCR-independent activation, observational human data for their activation in influenza A virus, and in vivo murine evidence of their protection against severe influenza A infection, mediated at least partially via IFN-gamma. We then survey evidence emerging from other respiratory viral infections including recent evidence for an important adjuvant role in adenovirus infection, specifically chimpanzee adenoviruses used in recent coronavirus vaccines, and data for strong associations between MAIT cell responses and adverse outcomes from coronavirus-19 (COVID-19) disease. We speculate on potential translational implications of these findings, either using corticosteroids or inhibitory ligands to suppress deleterious MAIT cell responses, or the potential utility of stimulatory MR1 ligands to boost MAIT cell frequencies to enhance innate viral defences.

[Cloning and function analysis of chalcone isomerase gene and chalcone synthase gene in Lonicera macranthoides].

In order to explore the functions of genes of key rate-limiting enzymes chalcone isomerase(CHI) and chalcone synthase(CHS) in the biosynthesis of flavonoids in Lonicera macranthoides, this study screened and cloned the cDNA sequences of CHI and CHS genes from the transcriptome data of conventional variety and 'Xianglei' of L. macranthoides. Online bioinformatics analysis software was used to analyze the characteristics of the encoded proteins, and quantitative reverse-transcription polymerase chain reaction(qRT-PCR) to detect the expression of CHI and CHS in different parts of the varieties at different flowering stages. The content of luteo-loside was determined by high performance liquid chromatography(HPLC) and the correlation with the expression of the two genes was analyzed. The results showed that the CHI and CHS of the two varieties contained a 627 bp and 1170 bp open reading frame(ORF), respectively, and the CHI protein and CHS protein were stable, hydrophilic, and non-secretory. qRT-PCR results demonstrated that CHI and CHS of the two varieties were differentially expressed in stems and leaves at different flowering stages, particularly the key stages. Based on HPLC data, luteoloside content was in negative correlation with the relative expression of the genes. Thus, CHI and CHS might regulate the accumulation of flavonoids in L. macranthoides, and the specific functions should be further studied. This study cloned CHI and CHS in L. macranthoides and analyzed their expression for the first time, which laid a basis for investigating the molecular mechanism of the differences in flavonoids such as luteoloside in L. macranthoides and variety breeding.

Fis Contributes to Resistance of Pseudomonas aeruginosa to Ciprofloxacin by Regulating Pyocin Synthesis.

Factor for inversion stimulation (Fis) is a versatile DNA binding protein that plays an important role in coordinating bacterial global gene expression in response to growth phases and environmental stresses. Previously, we demonstrated that Fis regulates the type III secretion system (T3SS) in Pseudomonas aeruginosa In this study, we explored the role of Fis in the antibiotic resistance of P. aeruginosa and found that mutation of the fis gene increases the bacterial susceptibility to ciprofloxacin. We further demonstrated that genes related to pyocin biosynthesis are upregulated in the fis mutant. The pyocins are produced in response to genotoxic agents, including ciprofloxacin, and the release of pyocins results in lysis of the producer cell. Thus, pyocin biosynthesis genes sensitize P. aeruginosa to ciprofloxacin. We found that PrtN, the positive regulator of the pyocin biosynthesis genes, is upregulated in the fis mutant. Genetic experiments and electrophoretic mobility shift assays revealed that Fis directly binds to the promoter region of prtN and represses its expression. Therefore, our results revealed novel Fis-mediated regulation on pyocin production and bacterial resistance to ciprofloxacin in P. aeruginosaIMPORTANCEPseudomonas aeruginosa is an important opportunistic pathogenic bacterium that causes various acute and chronic infections in human, especially in patients with compromised immunity, cystic fibrosis (CF), and/or severe burn wounds. About 60% of cystic fibrosis patients have a chronic respiratory infection caused by P. aeruginosa The bacterium is intrinsically highly resistant to antibiotics, which greatly increases difficulties in clinical treatment. Therefore, it is critical to understand the mechanisms and the regulatory pathways that are involved in antibiotic resistance. In this study, we elucidated a novel regulatory pathway that controls the bacterial resistance to fluoroquinolone antibiotics, which enhances our understanding of how P. aeruginosa responds to ciprofloxacin.

Combination of Azithromycin and Gentamicin for Efficient Treatment of Pseudomonas aeruginosa Infections.

BACKGROUND: Trans-translation is a ribosome rescue system that plays an important role in bacterial tolerance to environmental stresses. It is absent in animals, making it a potential treatment target. However, its role in antibiotic tolerance in Pseudomonas aeruginosa remains unknown. METHODS: The role and activity of trans-translation during antibiotic treatment were examined with a trans-translation-deficient strain and a genetically modified trans-translation component gene, respectively. In vitro assays and murine infection models were used to examine the effects of suppression of trans-translation. RESULTS: We found that the trans-translation system plays an essential role in P. aeruginosa tolerance to azithromycin and multiple aminoglycoside antibiotics. We further demonstrated that gentamicin could suppress the azithromycin-induced activation of trans-translation. Compared with each antibiotic individually, gentamicin and azithromycin combined increased the killing efficacy against planktonic and biofilm-associated P. aeruginosa cells, including a reference strain PA14 and its isogenic carbapenem-resistance oprD mutant, the mucoid strain FRD1, and multiple clinical isolates. Furthermore, the gentamicin-azithromycin resulted in improved bacterial clearance in murine acute pneumonia, biofilm implant, and cutaneous abscess infection models. CONCLUSIONS: Combination treatment with gentamicin and azithromycin is a promising strategy in combating P. aeruginosa infections.

Identification of novel genes that promote persister formation by repressing transcription and cell division in Pseudomonas aeruginosa.

OBJECTIVES: Bacterial persisters are a small subpopulation of cells that are highly tolerant of antibiotics and contribute to chronic and recalcitrant infections. Global gene expression in Pseudomonas aeruginosa persister cells and genes contributing to persister formation remain largely unknown. The objective of this study was to examine the gene expression profiles of the persister cells and those that regained growth in fresh medium, as well as to identify novel genes related to persister formation. METHODS: P. aeruginosa persister cells and those that regrew in fresh medium were collected and subjected to RNA sequencing analysis. Genes up-regulated in the persister cells were overexpressed to evaluate their roles in persister formation. The functions of the persister-contributing genes were assessed with pulse-chase assay, affinity chromatography, fluorescence and electron microscopy, as well as a light-scattering assay. RESULTS: An operon containing PA2282-PA2287 was up-regulated in the persister cells and down-regulated in the regrowing cells. PA2285 and PA2287 play key roles in persister formation. PA2285 and PA2287 were found to bind to RpoC and FtsZ, which are involved in transcription and cell division, respectively. Pulse-chase assays demonstrated inhibitory effects of PA2285 and PA2287 on the overall transcription. Meanwhile, light-scattering and microscopy assays demonstrated that PA2285 and PA2287 interfere with cell division by inhibiting FtsZ aggregation. PA2285 and PA2287 are conserved in pseudomonads and their homologous genes in Pseudomonas putida contribute to persister formation. CONCLUSIONS: PA2285 and PA2287 are novel bifunctional proteins that contribute to persister formation in P. aeruginosa.

Development and evaluation of novel recombinant adenovirus-based vaccine candidates for infectious bronchitis virus and Mycoplasma gallisepticum in chickens.

Avian infectious bronchitis caused by the infectious bronchitis virus (IBV), and mycoplasmosis caused by Mycoplasma gallisepticum (MG) are two major respiratory diseases in chickens that have resulted in severe economic losses in the poultry industry. We constructed a recombinant adenovirus that simultaneously expresses the S1 spike glycoprotein of IBV and the TM-1 protein of MG (pBH-S1-TM-1-EGFP). For comparison, we constructed two recombinant adenoviruses (pBH-S1-EGFP and pBH-TM-1-EGFP) that express either the S1 spike glycoprotein or the TM-1 protein alone. The protective efficacy of these three vaccine constructs against challenge with IBV and/or MG was evaluated in specific pathogen free chickens. Groups of seven-day-old specific pathogen free chicks were immunized twice, two weeks apart, via the oculonasal route with the pBH-S1-TM-1-EGFP, pBH-S1-EGFP, or pBH-TM-1-EGFP vaccine candidates or the commercial attenuated infectious bronchitis vaccine strain H52 and MG vaccine strain F-36 (positive controls), and challenged with virulent IBV or MG two weeks later. Interestingly, by days 7 and 14 after the booster immunization, pBH-S1-TM-1-EGFP-induced antibody titre was significantly higher (P < 0.01) compared to attenuated commercial IBV vaccine; however, there was no significant difference between the pBH-S1-TM-1-EGFP and attenuated commercial MG vaccine groups (P > 0.05). The clinical signs, the gross, and histopathological lesions scores of the adenovirus vaccine constructs were not significantly different from that of the attenuated commercial IBV or MG vaccines (positive controls) (P > 0.05). These results demonstrate the potential of the bivalent pBH-S1-TM-1-EGFP adenovirus construct as a combination vaccine against IB and mycoplasmosis.

Adenovirus-mediated co-expression of the TRAIL and HN genes inhibits growth and induces apoptosis in Marek's disease tumor cell line MSB-1.

BACKGROUND: The objective of this study was to determine the in vitro tumor-inhibitory effect of a recombinant adenovirus expressing a fusion protein of tumor necrosis factor (TNF) related apoptosis inducing ligand (TRAIL) and hemagglutinin-neuraminidase (HN) genes on the MSB-1 Marek's disease tumor cell line. METHODS: TRAIL and HN genes were amplified from lymphocytes in the peripheral blood of chickens and the LaSota strain of Newcastle disease virus (NDV), respectively, using RT-PCR. The two genes were connected with a 2A connecting peptide by site-directed mutagenesis and gene splicing by overlap extension (SOE). The target gene TRAIL-2A-HN was cloned into the shuttle vector pShuttle-CMV. Homologous recombination was carried out with the vector pAdeasy-1 in the bacterium BJ5183 to construct the recombinant adenovirus plasmid pAd-TRAIL-2A-HN. After linearization, the plasmid was transfected into AD293 cells and packaged. Real-time quantitative PCR (RT-PCR) and fluorescence microscopy confirmed the introduction of the recombinant adenovirus into AD293 cells. The TCID50 method (50% tissue culture infectious dose) was employed to determine viral titers for the exprimental and control viruses, which met criteria for use. The Marek's disease tumor cell line MSB-1 was transfected with the constructed recombinant adenovirus. The infectivity of the recombinant adenovirus and the expression levels of exogenous genes were detected with RT-PCR and western blotting. The effects of the recombinant adenovirus on the growth of MSB-1 cells and cellular apoptosis were determined using flow cytometry. RESULTS: The recombinant adenovirus infected the cultured cells in vitro, and replicated and expressed exogenous genes in the cells. The recombinant adenovirus Ad-TRAIL-2A-HN inhibited the growth of MSB-1 cells and induced apoptosis by expressing exogenous genes. The rate of induced MSB-1 cell apoptosis reached 11.61%, which indicated that TRAIL and HN produced synergistic tumor-inhibiting effects. CONCLUSION: The constructed TRAIL-2A-HN fusion gene combined the apoptosis-inducing function of TRAIL and the adsorptive capacity of HN from NDV for tumor cells, and the capacity of the recombinant adenovirus expressing this fusion gene to induce tumor cell apoptosis was reported. These results provide a basis for future in vivo tumor suppression studies using recombinant adenoviruses.

Melanoma extracellular vesicles inhibit tumor growth and metastasis by stimulating CD8 T cells.

Tumor cell-derived extracellular vesicles (EVs) play a crucial role in mediating immune responses by carrying and presenting tumor antigens. Here, we suggested that melanoma EVs triggered cytotoxic CD8 T cell-mediated inhibition of tumor growth and metastasis. Our results indicated that immunization of mice with melanoma EVs inhibited melanoma growth and metastasis while increasing CD8 T cells and serum interferon γ (IFN-γ) in vivo. In vitro experiments showed that melanoma EV stimulates dendritic cells (DCs) maturation, and mature dendritic cells induce T lymphocyte activation. Thus, tumor cell-derived EVs can generate anti-tumor immunity in a prophylactic setting and may be potential candidates for cell-free tumor vaccines.

Transcriptomic and metabolomic profiling reveals molecular regulatory network involved in flower development and phenotypic changes in two Lonicera macranthoides varieties.

Due to the medicinal importance of the flowers of Xianglei type (XL) Lonicera macranthoides, it is important to understand the molecular mechanisms that underlie their development. In this study, we elucidated the transcriptomic and metabolomic mechanisms that underlie the flower development mechanism of two L. macranthoides varieties. In this study, 3435 common differentially expressed unigenes (DEGs) and 1138 metabolites were identified. These common DEGs were mainly enriched in plant hormone signal transduction pathways. Metabolomic analysis showed that amino acids were the main metabolites of differential accumulation in wild-type (WT) L. macranthoides, whereas in XL, they were flavonoids and phenylalanine metabolites. Genes and transcription factors (TFs), such as MYB340, histone deacetylase 1 (HDT1), small auxin-up RNA 32 (SAUR32), auxin response factor 6 (ARF6), PIN-LIKES 7 (PILS7), and WRKY6, likely drive metabolite accumulation. Plant hormone signals, especially auxin signals, and various TFs induce downstream flower organ recognition genes, resulting in a differentiation of the two L. macranthoides varieties in terms of their developmental trajectories. In addition, photoperiodic, autonomous, and plant hormone pathways jointly regulated the L. macranthoides corolla opening. SAUR32, Arabidopsis response regulator 9 (ARR9), Gibberellin receptor (GID1B), and Constans-like 10 (COL10) were closely related to the unfolding of the L. macranthoides corolla. These findings offer valuable understanding of the flower growth process of L. macranthoides and the excellent XL phenotypes at the molecular level. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-024-04019-1.

Unveiling IL6R and MYC as Targeting Biomarkers in Imatinib-Resistant Chronic Myeloid Leukemia through Advanced Non-Invasive Apoptosis Detection Sensor Version 2 Detection.

The management of chronic myelogenous leukemia (CML) has seen significant progress with the introduction of tyrosine kinase inhibitors (TKIs), particularly Imatinib. However, a notable proportion of CML patients develop resistance to Imatinib, often due to the persistence of leukemia stem cells and resistance mechanisms independent of BCR::ABL1 This study investigates the roles of IL6R, IL7R, and MYC in Imatinib resistance by employing CRISPR/Cas9 for gene editing and the Non-Invasive Apoptosis Detection Sensor version 2 (NIADS v2) for apoptosis assessment. The results indicate that Imatinib-resistant K562 cells (K562-IR) predominantly express IL6R, IL7R, and MYC, with IL6R and MYC playing crucial roles in cell survival and sensitivity to Imatinib. Conversely, IL7R does not significantly impact cytotoxicity, either alone or in combination with Imatinib. Further genetic editing experiments confirm the protective functions of IL6R and MYC in K562-IR cells, suggesting their potential as therapeutic targets for overcoming Imatinib resistance in CML. This study contributes to understanding the mechanisms of Imatinib resistance in CML, proposing IL6R and MYC as pivotal targets for therapeutic strategies. Moreover, the utilization of NIADS v2 enhances our capability to analyze apoptosis and drug responses, contributing to a deeper understanding of CML pathogenesis and treatment options.

Caerin 1.9-Titanium Plates Aid Implant Healing and Inhibit Bacterial Growth in New Zealand Rabbit Mandibles.

OBJECTIVES: With rising rates of maxillofacial fracture, postoperative infection following rigid internal fixation is an important issue that requires immediate resolution. It is important to explore an alternative antibacterial method apart from conventional antibiotics. A controlled experiment was conducted to evaluate the effectiveness of a caerin 1.9 peptide-coated titanium plate in reducing mandibular infection in New Zealand (NZ) rabbits, aiming to minimise the risk of post-metallic implantation infection. METHODS: Twenty-two NZ rabbits were randomly divided into 3 groups. The experiment group received caerin 1.9 peptide-coated titanium plates and mixed oral bacteria exposure. The control group received normal titanium plates with mixed oral bacteria exposure. The untreated group served as a control to prove that bacteria in the mouth can cause infection. Weight, temperature, hepatic function, and C-reactive protein levels were measured. Wound and bone conditions were evaluated. Further analysis included local infection, anatomic conditions, histology, and bacterial load. RESULTS: No significant differences were found in temperature, weight, blood alanine aminotransferase, and C-reactive protein levels amongst the 3 groups. The experiment group showed the lowest amount of bacterial RNA in wounds. Additionally, the experiment group had higher peripheral lymphocyte counts compared to the control group and lower neutrophil counts on the third and seventh day postoperatively. Histologic analysis revealed lower levels of inflammatory cell infiltration, bleeding, and areas of necrosis in the experimental group compared with the controls. CONCLUSIONS: A caerin 1.9-coated titanium plate is able to inhibit bacterial growth in a NZ rabbit mandibular mixed bacteria infection model and is worth further investigation.

Comparative transcriptome analysis to reveal key ethylene genes involved in a Lonicera macranthoides mutant.

BACKGROUND: Lonicera macranthoides Hand.-Mazz. is an important medicinal plant. Xianglei-type (XL) L. macranthoides was formed after many years of cultivation by researchers on the basis of the natural mutant. The corolla of L. macranthoides XL remains unexpanded and its flowering period is nearly three times longer than that of wild-type (WT) plants. However, the molecular mechanism behind this desirable trait remains a mystery. OBJECTIVE: To understand the floral phenotype differences between L. macranthoides and L. macranthoides XL at the molecular level. METHODS: Transcriptome analysis was performed on L. macranthoides XL and WT. One DEG was cloned by RT-PCR amplification and selected for qRT-PCR analysis. RESULTS: Transcriptome analysis showed that there were 5603 differentially expressed genes (DEGs) in XL vs. WT. Enrichment analysis of DEGs showed that pathways related to plant hormone signal transduction were significantly enriched. We identified 23 key genes in ethylene biosynthesis and signal transduction pathways. The most abundant were the ethylene biosynthesis DEGs. In addition, the open reading frames (ORFs) of WT and XL ETR2 were successfully cloned and named LM-ETR2 (GenBank: MW334978) and LM-XL-ETR2 (GenBank: MW334978), respectively. qRT-PCR at different flowering stages suggesting that ETR2 acts in the whole stage of flower development of WT and XL. CONCLUSIONS: This study provides new insight into the molecular mechanism that regulates the development of special traits in the flowers of L. macranthoides XL. The plant hormone ethylene plays an important role in flower development and flowering duration prolongation in L. macranthoides. The ethylene synthesis gene could be more responsible for the flower phenotype of XL. The genes identified here can be used for breeding and improvement of other flowering plants after functional verification.

Multifractal scaling analyses of urban street network structure: The cases of twelve megacities in China.

Traffic networks have been proved to be fractal systems. However, previous studies mainly focused on monofractal networks, while complex systems are of multifractal structure. This paper is devoted to exploring the general regularities of multifractal scaling processes in the street network of 12 Chinese cities. The city clustering algorithm is employed to identify urban boundaries for defining comparable study areas; box-counting method and the direct determination method are utilized to extract spatial data; the least squares calculation is employed to estimate the global and local multifractal parameters. The results showed multifractal structure of urban street networks. The global multifractal dimension spectrums are inverse S-shaped curves, while the local singularity spectrums are asymmetric unimodal curves. If the moment order q approaches negative infinity, the generalized correlation dimension will seriously exceed the embedding space dimension 2, and the local fractal dimension curve displays an abnormal decrease for most cities. The scaling relation of local fractal dimension gradually breaks if the q value is too high, but the different levels of the network always keep the scaling reflecting singularity exponent. The main conclusions are as follows. First, urban street networks follow multifractal scaling law, and scaling precedes local fractal structure. Second, the patterns of traffic networks take on characteristics of spatial concentration, but they also show the implied trend of spatial deconcentration. Third, the development space of central area and network intensive areas is limited, while the fringe zone and network sparse areas show the phenomenon of disordered evolution. This work may be revealing for understanding and further research on complex spatial networks by using multifractal theory.

Gravitational scaling analysis on spatial diffusion of COVID-19 in Hubei Province, China.

The spatial diffusion of epidemic disease follows distance decay law in geography and social physics, but the mathematical models of distance decay depend on concrete spatio-temporal conditions. This paper is devoted to modeling spatial diffusion patterns of COVID-19 stemming from Wuhan city to Hubei province, China. The modeling approach is to integrate analytical method and experimental method. The local gravity model is derived from allometric scaling and global gravity model, and then the parameters of the local gravity model are estimated by observational data and least squares calculation. The main results are as below. The local gravity model based on power law decay can effectively describe the diffusion patterns and process of COVID-19 in Hubei Province, and the goodness of fit of the gravity model based on negative exponential decay to the observational data is not satisfactory. Further, the goodness of fit of the model to data entirely became better and better over time, the size elasticity coefficient increases first and then decreases, and the distance attenuation exponent decreases first and then increases. Moreover, the significance of spatial autoregressive coefficient in the model is low, and the confidence level is less than 80%. The conclusions can be reached as follows. (1) The spatial diffusion of COVID-19 of Hubei bears long range effect, and the size of a city and the distance of the city to Wuhan affect the total number of confirmed cases. (2) Wuhan direct transmission is the main process in the spatial diffusion of COVID-19 in Hubei at the early stage, and the horizontal transmission between regions is not significant. (3) The effect of spatial lockdown and isolation measures taken by Chinese government against the transmission of COVID-19 is obvious. This study suggests that the role of urban gravity (size and distance) should be taken into account to prevent and control epidemic disease.

TpiA is a Key Metabolic Enzyme That Affects Virulence and Resistance to Aminoglycoside Antibiotics through CrcZ in Pseudomonas aeruginosa.

Carbon metabolism plays an essential role in bacterial pathogenesis and susceptibility to antibiotics. In Pseudomonas aeruginosa, Crc, Hfq, and a small RNA, CrcZ, are central regulators of carbon metabolism. By screening mutants of genes involved in carbon metabolism, we found that mutation of the tpiA gene reduces the expression of the type III secretion system (T3SS) and bacterial resistance to aminoglycoside antibiotics. TpiA is a triosephosphate isomerase that reversibly converts glyceraldehyde 3-phosphate to dihydroxyacetone phosphate, a key step connecting glucose metabolism with glycerol and phospholipid metabolisms. We found that mutation of the tpiA gene enhances the bacterial carbon metabolism, respiration, and oxidative phosphorylation, which increases the membrane potential and promotes the uptake of aminoglycoside antibiotics. Further studies revealed that the level of CrcZ is increased in the tpiA mutant due to enhanced stability. Mutation of the crcZ gene in the tpiA mutant background restored the expression of the T3SS genes and the bacterial resistance to aminoglycoside antibiotics. Overall, this study reveals an essential role of TpiA in the metabolism, virulence, and antibiotic resistance in P. aeruginosaIMPORTANCE The increase in bacterial resistance against antibiotics imposes a severe threat to public health. It is urgent to identify new drug targets and develop novel antimicrobials. Metabolic homeostasis of bacteria plays an essential role in their virulence and resistance to antibiotics. Recent studies demonstrated that antibiotic efficacies can be improved by modulating the bacterial metabolism. Pseudomonas aeruginosa is an important opportunistic human pathogen that causes various infections. The bacterium is intrinsically resistant to antibiotics. In this study, we provide clear evidence that TpiA (triosephosphate isomerase) plays an essential role in the metabolism of P. aeruginosa and influences bacterial virulence and antibiotic resistance. The significance of this work is in identifying a key enzyme in the metabolic network, which will provide clues as to the development of novel treatment strategies against infections caused by P. aeruginosa.

HigB Reciprocally Controls Biofilm Formation and the Expression of Type III Secretion System Genes through Influencing the Intracellular c-di-GMP Level in Pseudomonas aeruginosa.

Toxin-antitoxin (TA) systems play important roles in bacteria persister formation. Increasing evidence demonstrate the roles of TA systems in regulating virulence factors in pathogenic bacteria. The toxin HigB in Pseudomonas aeruginosa contributes to persister formation and regulates the expression of multiple virulence factors and biofilm formation. However, the regulatory mechanism remains elusive. In this study, we explored the HigB mediated regulatory pathways. We demonstrate that HigB decreases the intracellular level of c-di-GMP, which is responsible for the increased expression of the type III secretion system (T3SS) genes and repression of biofilm formation. By analyzing the expression levels of the known c-di-GMP metabolism genes, we find that three c-di-GMP hydrolysis genes are up regulated by HigB, namely PA2133, PA2200 and PA3825. Deletion of the three genes individually or simultaneously diminishes the HigB mediated regulation on the expression of T3SS genes and biofilm formation. Therefore, our results reveal novel functions of HigB in P. aeruginosa.

HigB of Pseudomonas aeruginosa Enhances Killing of Phagocytes by Up-Regulating the Type III Secretion System in Ciprofloxacin Induced Persister Cells.

Bacterial persister cells are dormant and highly tolerant to lethal antibiotics, which are believed to be the major cause of recurring and chronic infections. Activation of toxins of bacterial toxin-antitoxin systems inhibits bacterial growth and plays an important role in persister formation. However, little is known about the overall gene expression profile upon toxin activation. More importantly, how the dormant bacterial persisters evade host immune clearance remains poorly understood. Here we demonstrate that a Pseudomonas aeruginosa toxin-antitoxin system HigB-HigA is required for the ciprofloxacin induced persister formation. Transcriptome analysis of a higA::Tn mutant revealed up regulation of type III secretion systems (T3SS) genes. Overexpression of HigB increased the expression of T3SS genes as well as bacterial cytotoxicity. We further demonstrate that wild type bacteria that survived ciprofloxacin treatment contain higher levels of T3SS proteins and display increased cytotoxicity to macrophage compared to vegetative bacterial cells. These results suggest that P. aeruginosa accumulates T3SS proteins during persister formation, which can protect the persister cells from host clearance by efficiently killing host immune cells.