Microbiological research最新文献

{"title":"Temperate bacteriophage SapYZUs7 alters Staphylococcus aureus fitness balance by regulating expression of phage resistance, virulence and antimicrobial resistance gene.","authors":"Wenyuan Zhou, Yajie Li, Yuhong Wu, Weicheng Hu, Wenjuan Li, Aiping Deng, Yeling Han, Guoqiang Zhu, Zhenquan Yang","doi":"10.1016/j.micres.2024.128040","DOIUrl":"10.1016/j.micres.2024.128040","url":null,"abstract":"<p><p>Temperate bacteriophages are crucial for maintaining the pathogenicity and fitness of S. aureus, which also show promise as a biocontrol agent for S. aureus. However, the fitness benefit and cost of lysogeny by S. aureus temperate phages and their underlying mechanisms remain unexplored. In this study, phage resistance, virulence, antimicrobial resistance (AMR), transcriptome, and metabolome of phage SapYZUs7 lysogenic and non-lysogenic S. aureus strains were compared. Whole-genome analysis revealed that SapYZUs7 harbouring smaII associated with a single-protein MazF-like antiphage system could be integrated into the genome of S. aureus isolates. Notably, lysogenic S. aureus exhibited higher phage resistance, a lower growth rate, and inhibited metabolic activity compared to the parental strains, indicating interference of phage reproduction by smaII. Moreover, prophages carrying smaII are widely distributed across S. aureus and harboured other virulence factor (VF) and AMR genes. Besides, the SapYZUs7-integration increased phagocytosis resistance but decreased adhesion, biofilm formation, and AMR. The combined use of SapYZUs7 and antibiotics exhibited a better bactericidal effect than SapYZUs7 or the antibiotics alone. Consistently, integrated omics analysis suggested that SapYZUs7-lysogeny downregulated multiple VF and AMR genes. Our analysis suggests that SmaII drives mutualistic phage-host interactions through lysogenic conversion. The fitness cost of SapYZUs7-integration is the downregulated expression of VF and AMR genes, serving as an alternative candidate as a biocontrol agent for methicillin-resistant S. aureus and multidrug-resistant S. aureus.</p>","PeriodicalId":18564,"journal":{"name":"Microbiological research","volume":"292 ","pages":"128040"},"PeriodicalIF":6.1,"publicationDate":"2025-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142903380","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A novel Brucella T4SS effector RS15060 acts on bacterial morphology, lipopolysaccharide core synthesis and host proinflammatory responses, which is beneficial for Brucella melitensis virulence.","authors":"Yi Yin, Mingxing Tian, Guangdong Zhang, Chan Ding, Shengqing Yu","doi":"10.1016/j.micres.2024.128015","DOIUrl":"10.1016/j.micres.2024.128015","url":null,"abstract":"<p><p>Brucella relies on the type IV secretion system (T4SS) to establish replication niches within host cells. However, the Brucella T4SS effectors and their functions have not been fully identified. In this study, we investigated the function of Brucella RS15060, a novel T4SS effector discovered in our previous study, on the bacterial biological characteristics and pathogenesis by construction of the gene deletion and complementation strains. We found that deletion of the rs15060 gene weakened abilities of Brucella to replicate within host cells and establish chronic infection in mice but enhanced abilities to adhere/invade HeLa cells and evade lysosomal degradation in the early stage of infection. In addition, the rs15060 deletion Brucella strain showed significant changes in bacterial shape, cell wall thickness, and sensitivity to bactericidal factors. Furthermore, the rs15060 deletion strain showed an increased synthesis of bacterial lipopolysaccharide core and induced a stronger host's inflammatory response. The Brucella rs15060 complementation strain restored the altered biological characteristics. Moreover, BLASTP prediction and 3D structure simulation revealed that the Brucella RS15060 contains NAD(P)-binding and active motifs in structure, which are important for proteins to exert NAD dependent epimerase/dehydratase activity. The complementation strain with mutation on NAD(P)-binding and/or active motifs of RS15060 did not restore the altered characteristics, suggesting that the Brucella RS15060 is a potential NAD dependent epimerase/dehydratase, and the predicted NAD(P)-binding and/or active motifs play an important role on bacterial cell wall and LPS core synthesis, which is crucial for maintaining bacterial morphology and exerting virulence.</p>","PeriodicalId":18564,"journal":{"name":"Microbiological research","volume":"292 ","pages":"128015"},"PeriodicalIF":6.1,"publicationDate":"2025-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142846995","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Emergence of two novel tmexCD-toprJ subtypes mediating tigecycline resistance in the megaplasmids from Pseudomonas putida.","authors":"Chengzhen Wang, Xun Gao, Xiaoyu Zhang, Chao Yue, Luchao Lv, Litao Lu, Jian-Hua Liu","doi":"10.1016/j.micres.2025.128051","DOIUrl":"10.1016/j.micres.2025.128051","url":null,"abstract":"<p><p>The widespread antimicrobial resistance (AMR) problem poses a serious health threat, leaving few drug choices, including tigecycline, to treat multidrug resistance pathogens. However, a plasmid-borne tigecycline resistance gene cluster, tmexCD1-toprJ1, emerged and conferred tigecycline resistance. In this study, we identified two novel subtypes, tmexCD2.3-toprJ2.3 and tmexCD2.4-toprJ1b, obtained from three chicken-origin Pseudomonas putida isolates. Two types of megaplasmids were found as the vital vehicle of these tmexCD-toprJ variants. Phylogenetic and genomic analysis indicated the two variants were mainly distributed in Pseudomonas and acted as an evolved intermediated state precursor of tmexCD2-toprJ2. Further gene cloning assay revealed both the expression of tmexCD2.3-toprJ2.3 and tmexCD2.4-toprJ1b could confer multiple antimicrobial resistance, mediating 8- to 16-fold increase of tigecycline MIC. Importantly, two key nucleotide differences in promoter region influence the promoter activity between P_tmexC2.3 and P_tmexC2.4, while the downregulation effect of TNfxB on the transcriptional expression level of tmexCD2.3-toprJ2.3 and tmexCD2.4-toprJ1b were observed. The emergency of two novel tmexCD-toprJ variants necessitates preventive measures to curb their spread and highlights concerns about more emerging tmexCD-toprJ variants.</p>","PeriodicalId":18564,"journal":{"name":"Microbiological research","volume":"292 ","pages":"128051"},"PeriodicalIF":6.1,"publicationDate":"2025-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142965759","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"The Actinobacillus pleuropneumoniae apxIV operon encodes an antibacterial toxin-immunity pair.","authors":"Eva Slivenecka, David Jurnecka, Jana Holubova, Ondrej Stanek, Ludmila Brazdilova, Monika Cizkova, Ladislav Bumba","doi":"10.1016/j.micres.2024.128043","DOIUrl":"10.1016/j.micres.2024.128043","url":null,"abstract":"<p><p>The ApxIVA protein belongs to a distinct class of a \"clip and link\" activity of Repeat-in-ToXin (RTX) exoproteins. Along with the three other pore-forming RTX toxins (ApxI, ApxII and ApxIII), ApxIVA serves as a major virulence factor of Actinobacillus pleuropneumoniae, the causative agent of porcine pneumonia. The gene encoding ApxIVA is located on a bicistronic operon downstream of the orf1 gene and is expressed exclusively under in vivo conditions. Both ApxIVA and ORF1 are essential for full virulence of A. pleuropneumoniae, but the molecular mechanisms by which they contribute to the pathogenicity are not yet understood. Here, we provide a comprehensive structural and functional analysis of ApxIVA and ORF1 proteins. Our findings reveal that the N-terminal segment of ApxIVA shares structural similarity with colicin M (ColM)-like bacteriocins and exhibits an antimicrobial activity. The ORF1 protein resembles the colicin M immunity protein (Cmi) and, like Cmi, is exported to the periplasm through its N-terminal signal peptide. Additionally, ORF1 can protect bacterial cells from the antimicrobial activity of ApxIVA, suggesting that ORF1 and ApxIVA function as an antibacterial toxin-immunity pair. Moreover, we demonstrate that fetal bovine serum could elicit ApxIVA and ORF1 production under in vitro conditions. These findings highlight the coordinated action of various RTX determinants, where the fine-tuned spatiotemporal production of ApxIVA may enhance the fitness of A. pleuropneumoniae, facilitating its invasion to a resident microbial community on the surface of airway mucosa.</p>","PeriodicalId":18564,"journal":{"name":"Microbiological research","volume":"292 ","pages":"128043"},"PeriodicalIF":6.1,"publicationDate":"2025-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142910039","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Exploitative stress within Bacillus subtilis biofilm determines the spatial distribution of pleomorphic cells.","authors":"Sumon Dey, Ankit Kumar Nayak, Hema Rajaram, Surajit Das","doi":"10.1016/j.micres.2024.128034","DOIUrl":"10.1016/j.micres.2024.128034","url":null,"abstract":"<p><p>Bacteria commonly live in a spatially organized biofilm assemblage. The metabolic activity inside the biofilm leads to segmented physiological microenvironments. In nature, bacteria possess several pleomorphic forms to withstand certain ecological alterations. We hypothesized that pleomorphism also exists within the biofilm, which can be considered as the fundamental niche for bacteria. We report a distinct pattern of cell size variation throughout the biofilm of Bacillus subtilis. Cell size heterogeneity was observed in biofilm development, wherein the frequency of long cells is higher in outer regions, whereas lower in inner regions. Moreover, compared to planktonic cells, bacteria in the biofilm mode reduce their geometric ratio from 8.34 to 3.69 and 2.65 in the outer and inner regions, respectively. There were no significant differences observed in nutrient diffusion from the outer to the inner region, and more than 73 % of cells in the inner region were viable. However, the inner and middle regions were more acidic than the outer of the biofilm. Conclusively, growth rate-independent cell size reduction at low pH suggests that the resulting phenotype switching within biofilm was observed due to the pH gradient of neutral to acidic from the outer to the core of the biofilm. This gradient of H⁺ ions concentration may create exploitative stress within the biofilm, which could favor specific pleomorphic cells to thrive in their specialized niches. By understanding the cell size variation in response to the local environment, we propose a model of biofilm formation by pleomorphic cells.</p>","PeriodicalId":18564,"journal":{"name":"Microbiological research","volume":"292 ","pages":"128034"},"PeriodicalIF":6.1,"publicationDate":"2025-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142896117","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Phosphorylation of the prokaryotic histone-like protein H-NS modulates bacterial virulence in Salmonella Typhimurium.","authors":"Ying Wang, Jinli Ge, Wei Xian, Zhiheng Tang, Baoshuai Xue, Jingchen Yu, Yu-Feng Yao, Huwei Liu, Jiazhang Qiu, Xiaoyun Liu","doi":"10.1016/j.micres.2024.128041","DOIUrl":"10.1016/j.micres.2024.128041","url":null,"abstract":"<p><p>H-NS is a prokaryotic histone-like protein that binds to bacterial chromosomal DNA with important regulatory roles in gene expression. Unlike histone proteins, hitherto post-translational modifications of H-NS are still largely uncharacterized, especially in bacterial pathogens. Salmonella Typhimurium is a primary enteric pathogen and its virulence is mainly dependent on specialized type III secretion systems (T3SSs), which were evolutionarily acquired via horizontal gene transfer. Previous studies have shown that H-NS plays a critical role in silencing foreign T3SS genes. Here, we found that H-NS is phosphorylated at multiple residues in S. Typhimurium, including S45, Y61, S78, S84, T86, and T106. Notably, we demonstrated that phosphorylation of H-NS S78 promotes its dissociation from DNA via a mechanism dependent on dimer formation, thereby leading to transcriptional activation of target genes. Functionally, phosphoryl-H-NS contributes to the expression of T3SS-associated proteins and hence increases bacterial virulence during infection. Therefore, our study reveals a novel mechanism by which covalent modifications of prokaryotic histone-like proteins regulate bacterial virulence of an important human pathogen.</p>","PeriodicalId":18564,"journal":{"name":"Microbiological research","volume":"292 ","pages":"128041"},"PeriodicalIF":6.1,"publicationDate":"2025-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142907141","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Salmonella: Infection mechanism and control strategies.","authors":"Juane Lu, Hao Wu, Shengbo Wu, Shengli Wang, Hongfei Fan, Haihua Ruan, Jianjun Qiao, Qinggele Caiyin, Mingzhang Wen","doi":"10.1016/j.micres.2024.128013","DOIUrl":"10.1016/j.micres.2024.128013","url":null,"abstract":"<p><p>Salmonella is a foodborne pathogen that predominantly resides in the intestinal tract of humans and animals. Infections caused by Salmonella can lead to various illnesses, including gastroenteritis, bacteremia, septicemia, and focal infections, with severe cases potentially resulting in host mortality. The mechanisms by which Salmonella invades host cells and disseminates throughout the body are partly understood, but there are still many scientific questions to be solved. This review aims to synthesize existing research on the interactions between Salmonella and hosts, detailing a comprehensive infection mechanism from adhesion and invasion to intracellular propagation and systemic spread. Overuse of antibiotics contributes to the emergence of drug-resistant Salmonella strains. An in-depth analysis of the mechanism of Salmonella infection will provide a theoretical basis for the development of novel Salmonella control strategies. These innovative control strategies include antibiotic adjuvants, small molecules, phages, attenuated vaccines, and probiotic therapies, which show huge potential in controlling Salmonella infection.</p>","PeriodicalId":18564,"journal":{"name":"Microbiological research","volume":"292 ","pages":"128013"},"PeriodicalIF":6.1,"publicationDate":"2025-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142829323","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Phototactic signaling network in rod-shaped cyanobacteria: A study on Synechococcus elongatus UTEX 3055.","authors":"Shang-Yu Li, Chenliu He, Cesar Augusto Valades-Cruz, Cheng-Cai Zhang, Yiling Yang","doi":"10.1016/j.micres.2024.127967","DOIUrl":"10.1016/j.micres.2024.127967","url":null,"abstract":"<p><p>Light-controlled motility is advantageous for photosynthetic prokaryotes to better survive in environment with constantly changing light conditions. For cyanobacteria, light is both an energy source for photosynthesis and a stress factor. Consequently, some cyanobacteria evolved the ability to control type-IV pili (T4P)-mediated surface motility using a chemotaxis-like system in response to light signals. Extensive studies on the mechanism of phototaxis has been conducted in the spherical Synechocystis sp. PCC 6803 and the filamentous strain Nostoc punctiforme, while less is explored in rod-shaped cyanobacteria such as Synechococcus species. In this study, we investigated the phototaxis pathway in the unicellular rod-shaped cyanobacterium Synechococcus elongatus UTEX 3055, which exhibits bidirectional phototaxis using a single tax1 operon, in contrast to more complex and multiple gene clusters revealed in Synechocystis sp. PCC 6803. Results obtained by protein-protein interaction assays and protein subcellular localization experiments indicated that proteins encoded by the tax1 operon form large clusters that asymmetrically distributed both between the two poles and within the same pole. In vitro phosphorylation assays and site-directed mutations of conserved phosphorylation sites in PixL_Se, PixG_Se and PixH_Se demonstrate that PixL_Se acts as a histidine kinase, and PixG_Se and PixH_Se as response regulators for signal transduction. We further show that PixG_Se and PixH_Se are recruited to cell poles via interactions with the N-terminal region of PixL_Se. While phosphotransfer reactions in this signaling pathway are critical for phototactic signaling, the two response regulators appear to play different roles in the control of phototaxis. This study provides a framework for further investigation into the complex phototactic signaling network in rod-shaped cyanobacteria with clearly defined cell poles in contrast to round shaped Synechocystis species with virtual cells poles through light-lensing effect.</p>","PeriodicalId":18564,"journal":{"name":"Microbiological research","volume":"292 ","pages":"127967"},"PeriodicalIF":6.1,"publicationDate":"2025-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142786085","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"The 1-acylglycerol-3-phosphate acyltransferase Slc1 is required to regulate mitochondria and lipid droplets.","authors":"Chenhui Zhao, Ke Liu, Yifan Wu, Shuaijie Yan, Jiajia He, Chuanhai Fu","doi":"10.1016/j.micres.2025.128080","DOIUrl":"https://doi.org/10.1016/j.micres.2025.128080","url":null,"abstract":"<p><p>Mitochondria are organelles involved in energy metabolism and biosynthesis. As the metabolites released from mitochondria are raw materials used for lipid synthesis, mitochondria also play important roles in lipid metabolism. Here we report that Slc1, a 1-acylglycerol-3-phosphate O-acyltransferase in the fission yeast Schizosaccharomyces pombe, is required to maintain tubular mitochondrial morphology and normal mitochondrial functions. The absence of Slc1 causes mitochondrial fragmentation, increases mitochondrial fission frequency, reduces mitochondrial respiration, and slows down nitrogen starvation-induced mitophagy. In addition, the absence of Slc1 significantly increases the protein level of Ptl2, which is the triacylglycerol lipase localized on lipid droplets. The phenotypes caused by the absence of Slc1 depend on its acyltransferase enzymatic activity. Therefore, our study uncovers new roles of a lipid synthesis enzyme Slc1 in regulating mitochondria and lipid droplets.</p>","PeriodicalId":18564,"journal":{"name":"Microbiological research","volume":"293 ","pages":"128080"},"PeriodicalIF":6.1,"publicationDate":"2025-01-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143075096","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Characterization of novel Erwinia amylovora-specific phiEaSP1 phage and its application as phage cocktail for managing fire blight in apples.","authors":"Hyeongsoon Kim, Nguyen Trung Vu, Roniya Thapa Magar, Eom-Ji Oh, Ki-Hoon Oh, Jiwon Lee, LeeSeul Kim, In Sun Hwang, Chang-Sik Oh","doi":"10.1016/j.micres.2025.128087","DOIUrl":"https://doi.org/10.1016/j.micres.2025.128087","url":null,"abstract":"<p><p>Erwinia amylovora (Ea) is a devastating bacterial pathogen that causes fire blight disease in Rosaceae family plants, including apples and pears. The use of bacteriophages is an alternative strategy to antibiotics for managing bacterial pathogens. In this study, the Ea-specific virulent phiEaSP1 phage was characterized, and its biocontrol efficacy against Ea was evaluated in apple seedlings. Genomic analyses revealed that phiEaSP1 belongs to the family Chaseviridae, subfamily Cleopatravirinae, and genus Loessnervirus. Most phiEaSP1 particles bound to the host cell surface within 5 min, and one virion made 68 progenies within 20 min of infection. The phage rapidly lysed Ea cells in vitro and maintained its lytic activity after incubation under different environmental conditions, including temperature, pH, and UV-A, as well as in the soil, with surfactants, and on apple seedlings. Receptor analysis using the Tn5 random mutant library of Ea TS3128 demonstrated that phiEaSP1 recognizes lipopolysaccharide as a receptor, whereas phiEaP-8 and phiEaP-21 recognize cellulose as a receptor. Protective efficacy against fire blight was tested on apple seedlings pretreated with the single phiEaSP1 or a phage cocktail containing phiEaSP1, phiEaP-8, and phiEaP-21. No or only weak symptoms were observed in the phage-treated seedlings. The application of a phage cocktail showed better control efficacy, indicating the potential of the phage cocktail, including phiEaSP1, as a preventive agent. Taken together, these results suggest that the use of a phage cocktail containing phiEaSP1 could be a potential strategy for the biocontrol of fire blight disease in apples.</p>","PeriodicalId":18564,"journal":{"name":"Microbiological research","volume":"293 ","pages":"128087"},"PeriodicalIF":6.1,"publicationDate":"2025-01-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143075081","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}