ABSTRACT
Klebsiella pneumoniae has become one of the most intractable gram-negative pathogens infecting humans and animals due to its severe antibiotic resistance. Bacteriophages and protein products derived from them are receiving increasing amounts of attention as potential alternatives to antibiotics. In this study, we isolated and investigated the characteristics of a new lytic phage, P1011, which lyses K5 K. pneumoniae specifically among 26 serotypes. The K5-specific capsular polysaccharide-degrading depolymerase dep1011 was identified and expressed. By establishing murine infection models using bovine strain B16 (capable of supporting phage proliferation) and human strain KP181 (incapable of sustaining phage expansion), we explored the safety and efficacy of phage and dep1011 treatments against K5 K. pneumoniae. Phage P1011 resulted in a 60% survival rate of the mice challenged with K. pneumoniae supporting phage multiplication, concurrently lowering the bacterial burden in their blood, liver, and lungs. Unexpectedly, even when confronted with bacteria impervious to phage multiplication, phage therapy markedly decreased the number of viable organisms. The protective efficacy of the depolymerase was significantly better than that of the phage. The depolymerase achieved 100% survival in both treatment groups regardless of phage propagation compatibility. These findings indicated that P1011 and dep1011 might be used as potential antibacterial agents to control K5 K. pneumoniae infection.
Subject(s)
Bacteriophages , Klebsiella Infections , Klebsiella pneumoniae , Animals , Klebsiella pneumoniae/virology , Klebsiella pneumoniae/physiology , Mice , Klebsiella Infections/therapy , Klebsiella Infections/veterinary , Klebsiella Infections/microbiology , Bacteriophages/physiology , Disease Models, Animal , Phage Therapy , Female , Glycoside Hydrolases/metabolism , CattleABSTRACT
OBJECTIVES: Hemoperfusion (HP) is used to treat various diseases, including sepsis and acute respiratory distress syndrome. However, few studies have explored the efficiency of HP in dermatomyositis-associated acute exacerbation of interstitial lung disease. METHODS: We conducted a retrospective study. Two hundred and sixteen patients with dermatomyositis-associated acute exacerbation of interstitial lung disease were included. Patients were divided into the HP group (treatment group) and the control group. Changes in oxygenation, hemodynamic parameters, lung ultrasound scores, and inflammatory cytokine levels were evaluated before and after HP in the treatment group. The length of intensive care unit (ICU) stays, duration of ventilator therapy, mortality rate, and incidence of complications were compared between the treatment and control groups. RESULTS: Hemodynamic and oxygenation variables in the treatment group significantly improved after treatment. However, the levels of the inflammatory factors significantly decreased after treatment. The length of ICU stay and the duration of ventilator therapy were significantly shorter in the treatment group than in the control group. The mortality rate of the treatment group was significantly lower than that of the control group. CONCLUSION: This study demonstrated that HP could improve treatment efficacy in patients with dermatomyositis-associated acute exacerbation of interstitial lung disease.
ABSTRACT
Dysfunction of mesangial cells plays a major role in the pathogenesis of diabetic kidney disease (DKD), the leading cause of kidney failure. However, the underlying molecular mechanisms are incompletely understood. By unbiased gene expression analysis of glucose-exposed mesangial cells, we identified the transmembrane receptor CD248 as the most upregulated gene, and the maladaptive unfolded protein response (UPR) as one of the most stimulated pathways. Upregulation of CD248 was further confirmed in glucose-stressed mesangial cells in vitro, in kidney glomeruli isolated from diabetic mice (streptozotocin; STZ and db/db models, representing type 1 and type 2 diabetes mellitus, respectively) in vivo, and in glomerular kidney sections from patients with DKD. Time course analysis revealed that glomerular CD248 induction precedes the onset of albuminuria, mesangial matrix expansion and maladaptive UPR activation (hallmarked by transcription factor C/EBP homologous protein (CHOP) induction) but is paralleled by loss of the adaptive UPR regulator spliced X box binding protein (XBP1). Mechanistically, CD248 promoted maladaptive UPR signaling via inhibition of the inositol requiring enzyme 1α (IRE1α)-mediated transcription factor XBP1 splicing in vivo and in vitro. CD248 induced a multiprotein complex comprising heat shock protein 90, BH3 interacting domain death agonist (BID) and IRE1α, in which BID impedes IRE1α-mediated XBP1 splicing and induced CHOP mediated maladaptive UPR signaling. While CD248 knockout ameliorated DKD-associated glomerular dysfunction and reverses maladaptive unfolded protein response signaling, concomitant XBP1 deficiency abolished the protective effect in diabetic CD248 knockout mice, supporting a functional interaction of CD248 and XBP1 in vivo. Hence, CD248 is a novel mesangial cell receptor inducing maladaptive UPR signaling in DKD.
Subject(s)
Diabetes Mellitus, Experimental , Diabetes Mellitus, Type 2 , Diabetic Nephropathies , Animals , Mice , Antigens, CD/metabolism , Antigens, Neoplasm , Diabetes Mellitus, Experimental/complications , Diabetes Mellitus, Experimental/genetics , Diabetic Nephropathies/genetics , Endoribonucleases/genetics , Endoribonucleases/metabolism , Protein Serine-Threonine Kinases/genetics , Transcription Factors/metabolism , Unfolded Protein Response , HumansABSTRACT
IMPORTANCE: To the best of our knowledge, this study reveals a strong correlation between mass spectra pattern and virulence phenotype among S. suis for the first time. In order to make the findings applicable and to excavate the intrinsic information in the spectra, the classifiers based on the machine learning algorithms were established, and RF (Random Forest)-based models have achieved an accuracy of over 90%. Overall, this study will pave the way for virulent SS2 (Streptococcus suis serotype 2) rapid detection, and the important findings on the association between genotype and mass spectrum may provide a new idea for the genotype-dependent detection of specific pathogens.
Subject(s)
Streptococcal Infections , Streptococcus suis , Humans , Virulence/genetics , Streptococcus suis/genetics , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization , Serogroup , Machine LearningABSTRACT
Avian pathogenic Escherichia coli, a causative agent of avian colibacillosis, has been causing serious economic losses in the poultry industry. The increase in multidrug-resistant isolates and the complexity of the serotypes of this pathogen, especially the recently reported emergence of a newly predominant serogroup of O145, make the control of this disease difficult. To address this challenge, a high-throughput screening approach, called Pan-RV (Reverse vaccinology based on pangenome analysis), is proposed to search for universal protective antigens against the three traditional serogroups and the newly emerged O145. Using this approach, a total of 61 proteins regarded as probable antigens against the four important serogroups were screened from the core genome of 127 Avian pathogenic Escherichia coli (APEC) genomes, and six were verified by Western blots using antisera. Overall, our research will provide a foundation for the development of an APEC subunit vaccine against avian colibacillosis. Given the exponential growth of whole-genome sequencing (WGS) data, our Pan-RV pipeline will make screening of bacterial vaccine candidates inexpensive, rapid, and efficient. IMPORTANCE With the emergence of drug resistance and the newly predominant serogroup O145, the control of Avian pathogenic Escherichia coli is facing a serious challenge; an efficient immunological method is urgently needed. Here, for the first time, we propose a high-throughput screening approach to search for universal protective antigens against the three traditional serogroups and the newly emerged O145. Importantly, using this approach, a total of 61 proteins regarded as probable antigens against the four important serogroups were screened, and three were shown to be immunoreactive with all antisera (covering the four serogroups), thereby providing a foundation for the development of APEC subunit vaccines against avian colibacillosis. Further, our Pan-RV pipeline will provide immunological control strategies for pathogens with complex and variable genetic backgrounds such as Escherichia coli and will make screening of bacterial vaccine candidates more inexpensive, rapid, and efficient.
Subject(s)
Escherichia coli Infections , Escherichia coli Vaccines , Poultry Diseases , Animals , Escherichia coli/genetics , Serogroup , Escherichia coli Infections/prevention & control , Escherichia coli Infections/veterinary , Poultry , Bacterial Vaccines , Poultry Diseases/prevention & control , Poultry Diseases/microbiology , ChickensABSTRACT
Mycoplasma hyopneumoniae is the etiological agent underlying porcine enzootic pneumonia, a chronic respiratory disease worldwide. The recruitment of plasminogen to the surface and subsequently promotion of plasmin conversion by the surface-located receptor, have been reported to assist the adhesion and invasion of Mycoplasmas. The surface localization and plasminogen-binding ability of M. hyopneumoniae enolase were previously confirmed; however, the biological functions were not be determined, especially the role as a plasminogen receptor. Here, using ELISA and SPR analyses, we confirmed the stable binding of M. hyopneumoniae enolase to plasminogen in a dose-dependent manner. The facilitation of the activation of plasminogen in the presence of tPA and direct activation of plasminogen at low efficiency without tPA addition by M. hyopneumoniae enolase were also determined using a plasmin-specific chromogenic substrate. Notably, the C-terminal and N-terminal regions located in M. hyopneumoniae enolase play an important role in plasminogen binding and activation. Additionally, we demonstrate that M. hyopneumoniae enolase can competitively inhibit the adherence of M. hyopneumoniae to PK15 cells. These results provide insight into the role of enolase in M. hyopneumoniae infection, a mechanism that manipulates the proteolytic system of the host.
Subject(s)
Mycoplasma hyopneumoniae , Animals , Swine , Mycoplasma hyopneumoniae/metabolism , Plasminogen/metabolism , Fibrinolysin/metabolism , Phosphopyruvate Hydratase , Adhesins, Bacterial/metabolismABSTRACT
Streptococcus parasuis (S. parasuis) is a close relative of Streptococcus suis (S. suis), composed of former members of S. suis serotypes 20, 22 and 26. S. parasuis could infect pigs and cows, and recently, human infection cases have been reported, making S. parasuis a potential opportunistic zoonotic pathogen. In this study, we analysed the genomic characteristics of S. parasuis, using pan-genome analysis, and compare some phenotypic determinants such as capsular polysaccharide, integrative conjugative elements, CRISPR-Cas system and pili, and predicted the potential virulence genes by associated analysis of the clinical condition of isolated source animals and genotypes. Furthermore, to discuss the relationship with S. suis, we compared these characteristics of S. parasuis with those of S. suis. We found that the characteristics of S. parasuis are similar to those of S. suis, both of them have "open" pan-genome, their antimicrobial resistance gene profiles are similar and a srtF pilus cluster of S. suis was identified in S. parasuis genome. But S. parasuis still have its unique characteristics, two novel pilus clusters are and three different type CRISPR-Cas system were found. Therefore, this study provides novel insights into the interspecific and intraspecific genetic characteristics of S. parasuis, which can be useful for further study of this opportunistic pathogen, such as serotyping, diagnostics, vaccine development, and study of the pathogenesis mechanism.
Subject(s)
Streptococcal Infections , Streptococcus suis , Swine Diseases , Animals , Cattle , Genomics , Streptococcal Infections/veterinary , Streptococcus , Streptococcus suis/genetics , Swine , Virulence/geneticsABSTRACT
Mycoplasma hyopneumoniae, the causative agent of swine respiratory disease, demonstrates differences in virulence. However, factors associated with this variation remain unknown. We herein evaluated the association between differences in virulence and genotypes as well as phenotype (i.e., biofilm formation ability). Strains 168 L, RM48, XLW-2, and J show low virulence and strains 232, 7448, 7422, 168, NJ, and LH show high virulence, as determined through animal challenge experiments, complemented with in vitro tracheal mucosa infection tests. These 10 strains with known virulence were then subjected to classification via multilocus sequence typing (MLST) with three housekeeping genes, P146-based genotyping, and multilocus variable-number tandem-repeat analysis (MLVA) of 13 loci. MLST and P146-based genotyping identified 168, 168 L, NJ, and RM48 as the same type and clustered them in a single branch. MLVA assigned a different sequence type to each strain. Simpson's index of diversity indicates a higher discriminatory ability for MLVA. However, no statistically significant correlation was found between genotypes and virulence. Furthermore, we investigated the correlation between virulence and biofilm formation ability. The strains showing high virulence demonstrate strong biofilm formation ability, while attenuated strains show low biofilm formation ability. Pearson correlation analysis revealed a significant positive correlation between biofilm formation ability and virulence. To conclude, there was no association between virulence and our genotyping data, but virulence was found to be significantly associated with the biofilm formation ability of M. hyopneumoniae.
Subject(s)
Biofilms , Mycoplasma hyopneumoniae , Swine Diseases , Animals , Genotype , Multilocus Sequence Typing/veterinary , Mycoplasma hyopneumoniae/genetics , Swine , Swine Diseases/microbiology , VirulenceABSTRACT
Mycoplasma hyorhinis may cause systemic inflammation of pigs, typically polyserositis and arthritis, and is also associated with several types of human cancer. However, the pathogenesis of M. hyorhinis colonizing and breaching the respiratory barrier to establish systemic infection is poorly understood. Glycolytic enzymes are important moonlighting proteins and virulence-related factors in various bacteria. In this study, we investigated the functions of a glycolytic critical enzyme, enolase in the infection and systemic spread of M. hyorhinis. Bacterial surface localization of enolase was confirmed by flow cytometry and colony hybridization assay. Recombinant M. hyorhinis enolase (rEno) was found to adhere to pig kidney (PK-15) cells, and anti-rEno serum significantly decreased adherence. The enzyme was also found to bind host plasminogen and fibronectin, and interactions were specific and strong, with dissociation constant (KD) values of 1.4 nM and 14.3 nM, respectively, from surface plasmon resonance analysis. Activation of rEno-bound plasminogen was confirmed by its ability to hydrolyze plasmin-specific substrates and to degrade a reconstituted extracellular matrix. To explore key sites during these interactions, C-terminal lysine residues of enolase were replaced with leucine, and the resulting single-site and double-site mutants show significantly reduced interaction with plasminogen in far-Western blotting and surface plasmon resonance tests. The binding affinities of all mutants to fibronectin were reduced as well. Collectively, these results imply that enolase moonlights as an important adhesin of M. hyorhinis, and interacts with plasminogen and fibronectin. The two lysine residues in the C-terminus are important binding sites for its multiple binding activities.
Subject(s)
Mycoplasma hyorhinis , Plasminogen , Adhesins, Bacterial , Animals , Fibronectins , Phosphopyruvate Hydratase/genetics , Phosphopyruvate Hydratase/metabolism , Plasminogen/metabolism , SwineABSTRACT
INTRODUCTION: Gastric cancer (GC) has a high incidence and mortality rate, especially in East Asians, and about 90% of GCs are adenocarcinomas. Histological and etiological heterogeneity and ethnic diversity make molecular subtyping of GC complicated, thus making it difficult to determine molecular division systems and standard treatment modalities. Limited cohorts from South Korea, Singapore, Australia, and Japan have been studied; however, the mutational landscape of gastric adenocarcinomas in Chinese patients is still unknown. METHODS: We performed a targeted sequencing panel focusing on cancer-related genes and tumor-associated microorganisms of 529 gastric adenocarcinoma samples with matched blood controls. We identified 449 clinically relevant gene mutations. RESULTS: Approximately 47.1% of Chinese patients with GC harbored at least one actionable mutation. The top somatic mutations were TP53, ARID1A, LRP1B, PIK3CA, ERBB2, CDH1, KRAS, FAT4, CCNE1, and KMT2D. Truncation mutations of ARID1A, KMT2D, RNF43, TGFBR2, and CIC occurred in patients with high tumor mutational burden. Gene amplifications of ERBB2, CCNE1, CDK12, and CCND1 were detected in patients with low tumor mutational burden. Pathway analysis revealed common gene alterations in the Wnt and PI3K/Akt signaling pathways. The ratio of patients with high microsatellite instability was significantly lower than other cohorts, and high microsatellite instability and Epstein-Barr virus (EBV)-positive features seemed mutually inclusive in Chinese patients with GC. In 44 (8.3%) patients, 45 germline mutations were identified, among which SPINK1 mutations, all SPINK1 c.194 + 2T > C, were present in 15.9% (7/44) of patients. Microorganisms found in Chinese patients with GC included Helicobacter pylori, EBV, hepatitis B virus, and human papillomavirus types 16 and 18. CONCLUSION: Identification of varied molecular features by targeted next-generation sequencing provides more insight into patient stratification and offers more possibilities for both targeted therapies and immunotherapies of Chinese patients with GC. IMPLICATIONS FOR PRACTICE: This study investigated the genomic alteration profile of 529 Chinese patients with gastric adenocarcinoma by deep targeting sequencing, which might be the largest Chinese cohort on the genomic research of gastric adenocarcinoma up to now.
Subject(s)
Adenocarcinoma , Epstein-Barr Virus Infections , Stomach Neoplasms , Adenocarcinoma/genetics , Asian People/genetics , Australia , China , Herpesvirus 4, Human , High-Throughput Nucleotide Sequencing , Humans , Japan , Mutation , Phosphatidylinositol 3-Kinases , Republic of Korea , Stomach Neoplasms/genetics , Trypsin Inhibitor, Kazal PancreaticABSTRACT
Streptococcus suis is an important zoonotic pathogen. An increase in multi-drug-resistant strains has led to poor performance of traditional antibiotic therapies. Thus, alternative antibacterial agents are urgently needed. In this study, we identified a recombined and expressed lysin PlyARI derived from the novel serotype S. suis (Chz) prophage PhiARI0460-1. The recombinant PlyARI at a concentration of 10 µg/mL showed high bacteriolytic activity against 30 S. suis isolates. The minimum inhibitory concentration (MIC) of PlyARI against S. suis was found to be as low as 2 µg/mL, and the lytic efficiency could be maintained between the range of pH 4 and 12. Additionally, in a mouse infection model, a dose of 0.5 mg of PlyARI protected 10 out of 10 mice that were challenged with highly virulent S. suis strain HA9801. Furthermore, the binding specificity of PlyARI was evaluated by constructing a green fluorescent protein (GFP-ARIb), where GFP was fused with the PlyARI-SH3b (cell wall-binding domain, CBD), revealing a high affinity to S. suis, Staphylococcus aureus, and Streptococcus equi along with exhibiting a medium affinity to Streptococcus pneumoniae as well as Streptococcus agalactiae. Overall, our findings indicated that PlyARI may be an alternative antibacterial agent that was useful in treating and possibly the prevention of Streptococcal infections.
Subject(s)
Streptococcal Infections , Streptococcus suis , Animals , Mice , Microbial Sensitivity Tests , N-Acetylmuramoyl-L-alanine Amidase , Prophages/genetics , Streptococcal Infections/drug therapy , Streptococcus suis/geneticsABSTRACT
Mycoplasma hyorhinis infects pigs causing polyserositis and polyarthritis, and has also been reported in a variety of human tumor tissues. The occurrence of disease is often linked with the systemic invasion of the pathogen. Glyceraldehyde-3-Phosphate Dehydrogenase (GAPDH), one of the key enzymes of glycolysis, was reported as a surface multifunctional molecule in several bacteria. Here, we investigated whether GAPDH could manifest binary functions; as an adhesin to promote colonization as well as a plasminogen receptor functioning in extracellular matrix (ECM) degradation to promote systemic invasion. The surface localization of GAPDH was observed in M. hyorhinis with flow cytometry and colony blot analysis. Recombinant GAPDH (rGAPDH) was found to be able to bind porcine-derived PK-15 and human-derived NCI-H292 cells. The incubation with anti-GAPDH antibody significantly decreased the adherence of M. hyorhinis to both cell lines. To investigate its function in recruiting plasminogen, firstly, the interaction between rGAPDH and plasminogen was demonstrated by ELISA and Far-Western blot assay. The activation of the rGAPDH-bound plasminogen into plasmin was proved by using a chromogenic substrate, and furtherly confirmed to degrade extracellular matrix by using a reconstituted ECM. Finally, the ability of rGAPDH to bind different ECM components was demonstrated, including fibronectin, laminin, collagen type IV and vitronectin. Collectively, our data imply GAPDH as an important adhesion factor of M. hyrohinis and a receptor for hijacking host plasminogen to degrade ECM. The multifunction of GAPDH to bind both plasminogen and ECM components is believed to increase the targeting of proteolysis and facilitate the dissemination of M. hyorhinis.
Subject(s)
Adhesins, Bacterial/genetics , Bacterial Proteins/genetics , Glyceraldehyde-3-Phosphate Dehydrogenases/genetics , Mycoplasma hyorhinis/physiology , Receptors, Cell Surface/genetics , Adhesins, Bacterial/metabolism , Animals , Bacterial Adhesion/physiology , Bacterial Proteins/metabolism , Cell Line , Epithelial Cells/microbiology , Epithelial Cells/physiology , Extracellular Matrix , Glyceraldehyde-3-Phosphate Dehydrogenases/metabolism , Humans , Mycoplasma hyorhinis/genetics , Plasminogen/metabolism , Receptors, Cell Surface/metabolism , Sus scrofaABSTRACT
Primary porcine bronchial epithelial cells (PBECs) are an ideal model to study the molecular and pathogenic mechanisms of various porcine respiratory pathogens. However, the short lifespan of primary PBECs greatly limit their application. Here, we isolated and cultured primary PBECs and established immortalized PBECs by transfecting primary PBECs with the pEGFP-hTERT recombinant plasmid containing human telomerase reverse transcriptase (hTERT). Immortalized PBECs (hTERT-PBECs) retained the morphological and functional features of primary PBECs as indicated by cytokeratin 18 expression, telomerase activity assay, proliferation assays, karyotype analysis, and quantitative reverse-transcriptase polymerase chain reaction. Compared to primary PBECs, hTERT-PBECs had higher telomerase activity, extended replicative lifespan, and displayed enhanced proliferative activity. Moreover, this cell line is not transformed in vitro and does not exhibit a malignant phenotype in vivo, suggesting that it can be safely used in further studies. Besides, hTERT-PBECs were susceptible to swine influenza virus of H3N2 subtype and porcine circovirus type 2. In conclusion, the immortalized hTERT-PBECs represent a valuable in vitro model, which can be widely used in the study of porcine respiratory pathogenic infections.
Subject(s)
Bronchi/cytology , Epithelial Cells/enzymology , Primary Cell Culture/methods , Telomerase/genetics , Animals , Bronchi/enzymology , Cell Proliferation/genetics , Circovirus/pathogenicity , Humans , Karyotype , Orthomyxoviridae Infections/genetics , Orthomyxoviridae Infections/virology , Swine , Telomerase/biosynthesisABSTRACT
Streptococcus suis serotype 2 (SS2) is an important zoonotic pathogen that infects swine and humans with high mortality and morbidity. Although a number of virulence-associated factors have been reported, the understanding of the molecular mechanism underlying SS2 pathogenicity remains limited. Our previous studies revealed that srtBCD-associated protein 2' (SBP2') contributed to the pathogenesis of SS2, but the function of another member in the srtBCD cluster, srtBCD-associated protein 1 (SBP1) was still unknown. Here, we found that sbp1 was widely distributed among high virulent SS2 strains, suggesting that sbp1 may be involved in the pathogenesis of SS2. To investigate the function of SBP1, we firstly conducted Western blotting analyses to confirm that SBP1 was expressed in the high virulent SS2 strain ZY05719 both in vivo and in vitro, then constructed the deletion mutant of sbp1 by homologous recombination. Bacterial adhesion assay, indirect immunofluorescence assay and protein binding assay all demonstrated that SBP1 was associated with adhesion of SS2 to HEp-2â¯cells. However, SBP1 did not influence the invasion, phagocytosis or intracellular survival of SS2. Furthermore, infection assays in vivo showed that inactivation of sbp1 failed to impair the ability of SS2 to cause zebrafish and mouse mortality. Overall, these results indicate that SBP1 is an adhesion-associated factor without the involvement of virulence in Streptococcus suis serotype 2.
Subject(s)
Adhesins, Bacterial/metabolism , Bacterial Adhesion , Streptococcus suis/physiology , Adhesins, Bacterial/genetics , Animals , Blotting, Western , Cell Line, Tumor , Disease Models, Animal , Endocytosis , Epithelial Cells/microbiology , Gene Deletion , Gene Expression Profiling , Humans , Mice , Microbial Viability , Phagocytosis , Streptococcal Infections/microbiology , Streptococcal Infections/pathology , Virulence , ZebrafishABSTRACT
Mycoplasma hyopneumoniae is an important respiratory pathogen that causes great economic losses to the pig industry worldwide. Although some putative virulence factors have been reported, pathogenesis remains poorly understood. Herein, we evaluated the relative abundance of proteins in virulent 168 (F107) and attenuated 168L (F380) M. hyopneumoniae strains to identify virulence-associated factors by two-dimensional electrophoresis (2-DE). Seven proteins were found to be ≥ 1.5-fold more abundant in 168, and protein-protein interaction network analysis revealed that all seven interact with putative virulence factors. Unexpectedly, six of these virulence-associated proteins are encoded by core rather than accessory genomic elements. The most differentially abundant of the seven, fructose-1,6-bisphosphate aldolase (FBA), was successfully cloned, expressed and purified. Flow cytometry demonstrated the surface localisation of FBA, recombinant FBA (rFBA) mediated adhesion to swine tracheal epithelial cells (STEC), and anti-rFBA sera decreased adherence to STEC. Surface plasmon resonance showed that rFBA bound to fibronectin with a moderately strong KD of 469 nM. The results demonstrate that core gene expression contributes to adhesion and virulence in M. hyopneumoniae, and FBA moonlights as an important adhesin, mediating binding to host cells via fibronectin.
Subject(s)
Bacterial Adhesion , Fructose-Bisphosphate Aldolase/physiology , Mycoplasma hyopneumoniae/enzymology , Animals , Bacterial Adhesion/physiology , Blotting, Western/veterinary , Electrophoresis, Gel, Two-Dimensional/veterinary , Flow Cytometry/veterinary , Fructose-Bisphosphate Aldolase/genetics , Genome, Bacterial/genetics , Mycoplasma hyopneumoniae/genetics , Mycoplasma hyopneumoniae/pathogenicity , Pneumonia of Swine, Mycoplasmal/microbiology , Proteomics , Respiratory Mucosa/microbiology , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/veterinary , Swine/microbiology , Trachea/microbiology , VirulenceABSTRACT
BACKGROUND: There are several versions of the Breastfeeding Motivation Scale (BMS), which have been shown to measure maternal breastfeeding motivation, but there is not a Chinese version yet. The study aimed to translate the BMS into Chinese and subsequently assess its psychometric properties among Chinese mothers during the postpartum period. METHODS: The study was composed of two phases. The translation of BMS closely followed the principals of good practices. Phase 1 included a comprehensive translation, back-translation, cross-cultural adaptation, and pretest to develop the Chinese version of the BMS. From 1 December 2021 to 1 July 2022, the Chinese version of the BMS was administered to 206 postnatal mothers in our maternity wards to assess its psychometric properties. Phase 2 involved psychometric property testing, including testing of the internal consistency, test-retest reliability, content validity, construct validity, convergent validity and discriminant validity. RESULTS: Minor modifications in four items were recommended after translations. The Cronbach's α coefficient of the Chinese version of the BMS was .887, and the intraclass correlation coefficient was .897 (P < 0.001). The model fit was acceptable (χ2/df = 2.40, P < 0.001, RMSEA = 0.08, CFI = 0.91, IFI = 0.92 and TLI = 0.90) according to the confirmatory factor analysis. The composite reliability values corresponding to each latent variable were 0.733 ~ 0.926, and the average variance extracted values were 0.476 ~ 0.653. The correlations among the five measured variables were all lower than .85 and the square roots of average variance extracted from the variable were greater than the interconstruct correlations among the five measured variables in the model. CONCLUSIONS: The Chinese version of the BMS has good reliability and validity and provides a reliable assessment tool for measuring maternal breastfeeding motivation. It also provides support to develop culturally sensitive interventions for Chinese mothers' who are breastfeeding.
Subject(s)
Breast Feeding , Motivation , Humans , Female , Pregnancy , Surveys and Questionnaires , Reproducibility of Results , ChinaABSTRACT
In order to study the improvement effect of nano-clay and polypropylene fiber on the mechanical properties of recycled aggregates, unconfined compression tests and triaxial shear tests were conducted. The experimental results show that adding polypropylene fibers to recycled aggregates increases the unconfined compressive strength by 27% and significantly improves ductility. We added 6% nano-clay to fiber-reinforced recycled aggregates, which increased the unconfined compressive strength of the recycled aggregates by 49% and the residual stress by 146%. However, the ductility decreased. Under low confining pressures, with the addition of nano-clay, the peak deviatoric stress strength of the fiber-reinforced recycled aggregates first decreased and then increased. When the nano-clay content was 8%, this reached a maximum value. However, under high confining pressures, the recycled aggregate particles were tightly interlocked, so that the improvement effect of the fiber and nano-clay was not obvious. As more nano-clay was added, the friction angle of the fiber-reinforced recycled aggregates decreased, while the cohesion increased. When the content of nano-clay was 8%, the cohesive force increased by 110%. The results of this research indicate that adding both polypropylene fibers and nano-clay to recycled aggregates has a better improvement effect on their strength characteristics than adding only polypropylene fibers. This study can provide a reference for improving the mechanical properties of recycled aggregates and the use of roadbeds.
ABSTRACT
Streptococcus suis serotype 2 (SS2) has been reported to be a highly invasive pathogen in swine and a zoonotic agent for humans. Although many bacterial virulence factors have been identified, our an insightful understanding of SS2 pathogenicity is lacking. The gene nadR, encoding nicotinamide-nucleotide adenylyltransferase, was first reported as a regulator and transporter of the intracellular NAD synthesis pathway in Salmonella typhimurium. In this study, we constructed a mutant strain of nadR (ΔnadR) to test the phenotypic and virulence variations between the deletion mutant and the wild-type strain ZY05719. The phenotypic experimental results showed that ΔnadR obviously inhibited cell growth and exhibited shorter chains than WT. The growth defect of ΔnadR was caused by the loss of the function of nadR for transporting the substrates nicotinamide mononucleotide and nicotinamide riboside in the intracellular NAD synthesis pathway. In the process of interaction with the host, ΔnadR participated in adherence and invasion to the host cells, and it was more easily cleared by RAW264.7 macrophages. More importantly, both zebrafish and BALB/c mice in vivo virulence experimental results showed that ΔnadR dramatically attenuated the virulence of SS2, and the ability of ΔnadR to colonize tissues was notably reduced in comparison with that of WT in the BALB/c mice infection model. To the best of our knowledge, this is the first time to demonstrate that nadR not only plays an important role in bacterial growth, but also in connection with the virulence of SS2 as a global transcriptional regulator.
Subject(s)
Streptococcal Infections , Streptococcus suis , Swine Diseases , Animals , Mice , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , NAD/metabolism , Serogroup , Streptococcal Infections/veterinary , Streptococcal Infections/microbiology , Streptococcus suis/pathogenicity , Swine , Virulence/genetics , ZebrafishABSTRACT
Integrating genomics and histology for cancer prognosis demonstrates promise. Here, we develop a multi-classifier system integrating a lncRNA-based classifier, a deep learning whole-slide-image-based classifier, and a clinicopathological classifier to accurately predict post-surgery localized (stage I-III) papillary renal cell carcinoma (pRCC) recurrence. The multi-classifier system demonstrates significantly higher predictive accuracy for recurrence-free survival (RFS) compared to the three single classifiers alone in the training set and in both validation sets (C-index 0.831-0.858 vs. 0.642-0.777, p < 0.05). The RFS in our multi-classifier-defined high-risk stage I/II and grade 1/2 groups is significantly worse than in the low-risk stage III and grade 3/4 groups (p < 0.05). Our multi-classifier system is a practical and reliable predictor for recurrence of localized pRCC after surgery that can be used with the current staging system to more accurately predict disease course and inform strategies for individualized adjuvant therapy.
Subject(s)
Carcinoma, Renal Cell , Kidney Neoplasms , Neoplasm Recurrence, Local , Humans , Carcinoma, Renal Cell/genetics , Carcinoma, Renal Cell/pathology , Kidney Neoplasms/genetics , Kidney Neoplasms/pathology , Kidney Neoplasms/surgery , Male , Female , Neoplasm Recurrence, Local/genetics , Middle Aged , Aged , Prognosis , Genomics/methods , Adult , Neoplasm Staging , Deep Learning , Disease-Free SurvivalABSTRACT
The widespread production and application of graphene oxide (GO) may lead to its dispersion throughout natural water systems, with potential negative effects on living organisms and the ecological environment. This study used gypsum (G) as an adsorbent and examined different conditions (pH, adsorbent dosage, GO initial concentration) for the removal effect of GO by G. The results showed the best adsorption effect for a solution pH of 8.0, gypsum dosage of 60 mg, initial GO concentration of 80 mg·L-1, and temperature of 303 K; at this time, the maximum removal rate of graphene oxide by gypsum was 93.3%. It could be obtained by isotherm and thermodynamic analysis that the GO adsorption by gypsum conforms to the Langmuir isotherm model, it does not easily occur in high-temperature environments, and is a spontaneous exothermic process. In addition, experiments such as SEM, AFM, TGA, XRD, XPS, FTIR, Raman, and Zeta were used to adsorb graphene oxide by gypsum composites (G/GO), through which the mineral interactions with graphene oxides were microscopically characterized. The impact on the adsorption properties of contaminants provides new insights into contaminant removal by gypsum.