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New Biotechnology May 2024Cell wall peptidoglycan binding domains (CBDs) of cell lytic enzymes, including bacteriocins, autolysins and bacteriophage endolysins, enable highly selective bacterial...
Cell wall peptidoglycan binding domains (CBDs) of cell lytic enzymes, including bacteriocins, autolysins and bacteriophage endolysins, enable highly selective bacterial binding, and thus, have potential as biorecognition molecules for nondestructive bacterial detection. Here, a novel design for a self-complementing split fluorescent protein (FP) complex is proposed, where a multimeric FP chain fused with specific CBDs ((FP-CBD)) is assembled inside the cell, to improve sensitivity by enhancing the signal generated upon Staphylococcus aureus or Bacillus anthracis binding. Flow cytometry shows enhanced fluorescence on the cell surface with increasing FP stoichiometry and surface plasmon resonance reveals nanomolar binding affinity to isolated peptidoglycan. The breadth of function of these complexes is demonstrated through the use of CBD modularity and the ability to attach enzymatic detection modalities. Horseradish peroxidase-coupled (FP-CBD) complexes generate a catalytic amplification, with the degree of amplification increasing as a function of FP length, reaching a limit of detection (LOD) of 10 cells/droplet (approximately 0.1 ng S. aureus or B. anthracis) within 15 min on a polystyrene surface. These fusion proteins can be multiplexed for simultaneous detection. Multimeric split FP-CBD fusions enable use as a biorecognition molecule with enhanced signal for use in bacterial biosensing platforms.
PubMed: 38750815
DOI: 10.1016/j.nbt.2024.05.004 -
Environmental Health Perspectives May 2024Respiratory tract infections are major contributors to the global disease burden. Quantitative microbial risk assessment (QMRA) holds potential as a rapidly deployable... (Review)
Review
BACKGROUND
Respiratory tract infections are major contributors to the global disease burden. Quantitative microbial risk assessment (QMRA) holds potential as a rapidly deployable framework to understand respiratory pathogen transmission and inform policy on infection control.
OBJECTIVES
The goal of this paper was to evaluate, motivate, and inform further development of the use of QMRA as a rapid tool to understand the transmission of respiratory pathogens and improve the evidence base for infection control policies.
METHODS
We conducted a literature review to identify peer-reviewed studies of complete QMRA frameworks on aerosol inhalation or contact transmission of respiratory pathogens. From each of the identified studies, we extracted and summarized information on the applied exposure model approaches, dose-response models, and parameter values, including risk characterization. Finally, we reviewed linkages between model outcomes and policy.
RESULTS
We identified 93 studies conducted in 16 different countries with complete QMRA frameworks for diverse respiratory pathogens, including SARS-CoV-2, spp., , influenza, and . Six distinct exposure models were identified across diverse and complex transmission pathways. In 57 studies, exposure model frameworks were informed by their ability to model the efficacy of potential interventions. Among interventions, masking, ventilation, social distancing, and other environmental source controls were commonly assessed. Pathogen concentration, aerosol concentration, and partitioning coefficient were influential exposure parameters as identified by sensitivity analysis. Most (84%, ) studies presented policy-relevant content including ) determining disease burden to call for policy intervention, ) determining risk-based threshold values for regulations, ) informing intervention and control strategies, and ) making recommendations and suggestions for QMRA application in policy.
CONCLUSIONS
We identified needs to further the development of QMRA frameworks for respiratory pathogens that prioritize appropriate aerosol exposure modeling approaches, consider trade-offs between model validity and complexity, and incorporate research that strengthens confidence in QMRA results. https://doi.org/10.1289/EHP12695.
Topics: Risk Assessment; Respiratory Tract Infections; Humans; SARS-CoV-2; COVID-19; Staphylococcus aureus; Infection Control; Legionella; Aerosols
PubMed: 38728217
DOI: 10.1289/EHP12695 -
Vaccines Apr 2024Bacterial surface display platforms have been developed for applications such as vaccine delivery and peptide library screening. The type V secretion system is an...
Bacterial surface display platforms have been developed for applications such as vaccine delivery and peptide library screening. The type V secretion system is an attractive anchoring motif for the surface expression of foreign proteins in gram-negative bacteria. SadA belongs to subtype C of the type V secretion system derived from spp. and promotes biofilm formation and host cell adherence. The inner membrane lipoprotein SadB is important for SadA translocation. In this study, SadA was used as an anchoring motif to expose heterologous proteins in using SadB. The ability of SadA to display heterologous proteins on the surface in the presence of SadB was approximately three-fold higher than that in its absence of SadB. Compared to full-length SadA, truncated SadAs (SadA and SadA) showed similar display capacities when exposing the B-cell epitopes of urease B from (UreB158-172aa and UreB349-363aa). We grafted different protein domains, including mScarlet (red fluorescent protein), the urease B fragment (UreBm) from SS1, and/or protective antigen domain 4 from A16R (PAD4), onto SadA or SadA. Whole-cell dot blotting, immunofluorescence, and flow cytometric analyses confirmed the localization of Flag×3-mScarlet (~30 kDa) and Flag×3-UreBm-mScarlet (~58 kDa) to the surface using truncated SadA or SadA as an anchoring motif. However, Flag×3-UreBm-PAD4-mScarlet (~75 kDa) was displayed on using SadA. The oral administrated pSadBA-FUM/StmΔΔ and pSadBA-FUM/StmΔΔ could elicit a significant mucosal and humoral immunity response. SadA could thus be used as an anchoring motif for the surface expression of large heterologous proteins as a potential strategy for attenuated bacterial vaccine development.
PubMed: 38675781
DOI: 10.3390/vaccines12040399 -
Genes Mar 2024is the bacterium responsible for causing the zoonotic disease called anthrax. The disease presents itself in different forms like gastrointestinal, inhalation, and...
is the bacterium responsible for causing the zoonotic disease called anthrax. The disease presents itself in different forms like gastrointestinal, inhalation, and cutaneous. Bacterial spores are tremendously adaptable, can persist for extended periods and occasionally endanger human health. The Anthrax Toxin Receptor-2 () gene acts as membrane receptor and facilitates the entry of the anthrax toxin into host cells. Additionally, mutations in the gene have been linked to various autoimmune diseases, including Hyaline Fibromatosis Syndrome (HFS), Ankylosing Spondylitis (AS), Juvenile Hyaline Fibromatosis (JHF), and Infantile Systemic Hyalinosis (ISH). This study delves into the genetic landscape of , aiming to comprehend its associations with diverse disorders, elucidate the impacts of its mutations, and pinpoint minimal non-pathogenic mutations capable of reducing the binding affinity of the gene with the protective antigen. Recognizing the pivotal role of single-nucleotide polymorphisms (SNPs) in shaping genetic diversity, we conducted computational analyses to discern highly deleterious and tolerated non-synonymous SNPs (nsSNPs) in the gene. The Mutpred2 server determined that the Arg465Trp alteration in the gene leads to altered DNA binding ( = 0.22) with a probability of a deleterious mutation of 0.808; notably, among the identified deleterious SNPs, rs368288611 (Arg465Trp) stands out due to its significant impact on altering the DNA-binding ability of . We propose these SNPs as potential candidates for hypertension linked to the gene, which is implicated in blood pressure regulation. Noteworthy among the tolerated substitutions is rs200536829 (Ala33Ser), recognized as less pathogenic; this highlights its potential as a valuable biomarker, potentially reducing side effects on the host while also reducing binding with the protective antigen protein. Investigating these SNPs holds the potential to correlate with several autoimmune disorders and mitigate the impact of anthrax disease in humans.
Topics: Antigens, Bacterial; Humans; Anthrax; Receptors, Peptide; Mutation; Polymorphism, Single Nucleotide; Bacterial Toxins; Bacillus anthracis; Hyaline Fibromatosis Syndrome; Spondylitis, Ankylosing; Disease Resistance; Receptors, Cell Surface; Protein Binding
PubMed: 38674361
DOI: 10.3390/genes15040426 -
Microbiology Spectrum Apr 2024The high-consequence pathogen causes human anthrax and often results in lethal infections without the rapid administration of effective antimicrobial treatment....
The high-consequence pathogen causes human anthrax and often results in lethal infections without the rapid administration of effective antimicrobial treatment. Antimicrobial resistance profiling is therefore critical to inform post-exposure prophylaxis and treatment decisions, especially during emergencies such as outbreaks or where intentional release is suspected. Whole-genome sequencing using a rapid long-read sequencer can uncover antimicrobial resistance patterns if genetic markers of resistance are known. To identify genomic markers associated with antimicrobial resistance, we isolated derived from the avirulent Sterne strain with elevated minimal inhibitory concentrations to clarithromycin. Mutants were characterized both phenotypically through broth microdilution susceptibility testing and observations during culturing, as well as genotypically with whole-genome sequencing. We identified two different in-frame insertions in the L22 ribosomal protein-encoding gene , which were subsequently confirmed to be involved in clarithromycin resistance through the reversion of the mutant gene to the parent (drug-susceptible) sequence. Detection of the insertions was possible with rapid long-read sequencing, with a time-to-answer within 3 h. The mutations associated with clarithromycin resistance described here will be used in conjunction with known genetic markers of resistance for other antimicrobials to strengthen the prediction of antimicrobial resistance in .IMPORTANCEThe disease anthrax, caused by the pathogen , is extremely deadly if not treated quickly and appropriately. Clarithromycin is an antibiotic recommended for the treatment and post-exposure prophylaxis of anthrax by the Centers for Disease Control and Prevention; however, little is known about the ability of to develop resistance to clarithromycin or the mechanism of that resistance. The characterization of clarithromycin-resistant isolates presented here provides valuable information for researchers and clinicians in the event of a release of the resistant strain. Additionally, knowledge of the genetic basis of resistance provides a foundation for susceptibility prediction through rapid genome sequencing to inform timely treatment decisions.
PubMed: 38666793
DOI: 10.1128/spectrum.04180-23 -
Frontiers in Microbiology 2024Microorganisms are crucial in our ecosystem, offering diverse functions and adaptability. The UNGA Science Summit has underscored the importance of understanding...
BACKGROUND
Microorganisms are crucial in our ecosystem, offering diverse functions and adaptability. The UNGA Science Summit has underscored the importance of understanding microbes in alignment with the UN Sustainable Development Goals. poses significant challenges among various microorganisms due to its harmful effects on both soil and public health. Our study employed computational techniques to investigate the inhibitory effects of curcumin and mangiferin on , with the aim of presenting a novel bio-based approach to microbial management.
METHODS
Employing high-throughput screening, we identified potential binding sites on . Molecular docking revealed that curcumin and mangiferin, when synergistically combined, exhibited strong binding affinities at different sites on the bacterium. Our findings demonstrated a significant drop in binding free energy, indicating a stronger interaction when these compounds were used together.
FINDINGS
Results of Molecular docking indicated binding energies of -8.45 kcal/mol for mangiferin, -7.68 kcal/mol for curcumin, and a notably higher binding energy of -19.47 kcal/mol for the combination of mangiferin and curcumin with CapD protein. Molecular dynamics simulations further validated these interactions, demonstrating increased stability and structural changes in the bacterium.
CONCLUSION
This study highlights the effectiveness of natural compounds like curcumin and mangiferin in microbial management, especially against challenging pathogens like . It emphasizes the potential of sustainable, nature-based solutions and calls for further empirical research to expand upon these findings.
PubMed: 38646635
DOI: 10.3389/fmicb.2024.1304234 -
Frontiers in Cellular and Infection... 2024The metagenomic next-generation sequencing (mNGS) method is preferred for genotyping useful for the identification of organisms, illumination of metabolic pathways, and...
The metagenomic next-generation sequencing (mNGS) method is preferred for genotyping useful for the identification of organisms, illumination of metabolic pathways, and determination of microbiota. It can accurately obtain all the nucleic acid information in the test sample. Anthrax is one of the most important zoonotic diseases, infecting mainly herbivores and occasionally humans. The disease has four typical clinical forms, cutaneous, gastrointestinal, inhalation, and injection, all of which may result in sepsis or meningitis, with cutaneous being the most common form. Here, we report a case of cutaneous anthrax diagnosed by mNGS in a butcher. Histopathology of a skin biopsy revealed PAS-positive bacilli. Formalin-fixed paraffin-embedded (FFPE) tissue sample was confirmed the diagnosis of anthrax by mNGS. He was cured with intravenous penicillin. To our knowledge, this is the first case of cutaneous anthrax diagnosed by mNGS using FFPE tissue. mNGS is useful for identifying pathogens that are difficult to diagnose with conventional methods, and FFPE samples are simple to manage. Compared with traditional bacterial culture, which is difficult to cultivate and takes a long time, mNGS can quickly and accurately help us diagnose anthrax, so that anthrax can be controlled in a timely manner and prevent the outbreak of epidemic events.
Topics: Male; Humans; Anthrax; Paraffin Embedding; Formaldehyde; High-Throughput Nucleotide Sequencing; Metagenomics; Sensitivity and Specificity; Skin Diseases, Bacterial
PubMed: 38638828
DOI: 10.3389/fcimb.2024.1329235 -
Research Square Mar 2024The antimicrobial resistance crisis (AMR) is associated with millions of deaths and undermines the franchise of medicine. Of particular concern is the threat of...
The antimicrobial resistance crisis (AMR) is associated with millions of deaths and undermines the franchise of medicine. Of particular concern is the threat of bioweapons, exemplified by anthrax. Introduction of novel antibiotics helps mitigate AMR, but does not address the threat of bioweapons with engineered resistance. We reasoned that teixobactin, an antibiotic with no detectable resistance, is uniquely suited to address the challenge of weaponized anthrax. Teixobactinbinds to immutable targets, precursors of cell wall polymers. Here we show that teixobactinis highly efficacious in a rabbit model of inhalation anthrax. Inhaling spores of causes overwhelming morbidity and mortality. Treating rabbits with teixobactinafter the onset of disease rapidly eliminates the pathogen from blood and tissues, normalizes body temperature, and prevents tissue damage. Teixobactinassembles into an irreversible supramolecular structure of the surface of membrane, likely contributing to its unusually high potency against anthrax. Antibiotics evading resistance provide a rational solution to both AMR and engineered bioweapons.
PubMed: 38585816
DOI: 10.21203/rs.3.rs-3991430/v1 -
NPJ Systems Biology and Applications Mar 2024Protective antigen (PA) is a protein produced by Bacillus anthracis. It forms part of the anthrax toxin and is a key immunogen in US and UK anthrax vaccines. In this...
Protective antigen (PA) is a protein produced by Bacillus anthracis. It forms part of the anthrax toxin and is a key immunogen in US and UK anthrax vaccines. In this study, we have conducted experiments to quantify PA in the supernatants of cultures of B. anthracis Sterne strain, which is the strain used in the manufacture of the UK anthrax vaccine. Then, for the first time, we quantify PA production and degradation via mathematical modelling and Bayesian statistical techniques, making use of this new experimental data as well as two other independent published data sets. We propose a single mathematical model, in terms of delay differential equations (DDEs), which can explain the in vitro dynamics of all three data sets. Since we did not heat activate the B. anthracis spores prior to inoculation, germination occurred much slower in our experiments, allowing us to calibrate two additional parameters with respect to the other data sets. Our model is able to distinguish between natural PA decay and that triggered by bacteria via proteases. There is promising consistency between the different independent data sets for most of the parameter estimates. The quantitative characterisation of B. anthracis PA production and degradation obtained here will contribute towards the ambition to include a realistic description of toxin dynamics, the host immune response, and anti-toxin treatments in future mechanistic models of anthrax infection.
Topics: Humans; Bayes Theorem; Bacillus anthracis; Anthrax Vaccines; Anthrax
PubMed: 38553532
DOI: 10.1038/s41540-024-00357-1 -
International Journal of Molecular... Mar 2024Lethal toxin (LT) is the critical virulence factor of , the causative agent of anthrax. One common symptom observed in patients with anthrax is thrombocytopenia, which...
Lethal toxin (LT) is the critical virulence factor of , the causative agent of anthrax. One common symptom observed in patients with anthrax is thrombocytopenia, which has also been observed in mice injected with LT. Our previous study demonstrated that LT induces thrombocytopenia by suppressing megakaryopoiesis, but the precise molecular mechanisms behind this phenomenon remain unknown. In this study, we utilized 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced megakaryocytic differentiation in human erythroleukemia (HEL) cells to identify genes involved in LT-induced megakaryocytic suppression. Through cDNA microarray analysis, we identified Dachshund homolog 1 () as a gene that was upregulated upon TPA treatment but downregulated in the presence of TPA and LT, purified from the culture supernatants of . To investigate the function of DACH1 in megakaryocytic differentiation, we employed short hairpin RNA technology to knock down DACH1 expression in HEL cells and assessed its effect on differentiation. Our data revealed that the knockdown of DACH1 expression suppressed megakaryocytic differentiation, particularly in polyploidization. We demonstrated that one mechanism by which LT induces suppression of polyploidization in HEL cells is through the cleavage of MEK1/2. This cleavage results in the downregulation of the ERK signaling pathway, thereby suppressing gene expression and inhibiting polyploidization. Additionally, we found that known megakaryopoiesis-related genes, such as , , , , , and genes may be positively regulated by . Furthermore, we observed an upregulation of DACH1 during in vitro differentiation of CD34-megakaryocytes and downregulation of DACH1 in patients with thrombocytopenia. In summary, our findings shed light on one of the molecular mechanisms behind LT-induced thrombocytopenia and unveil a previously unknown role for DACH1 in megakaryopoiesis.
Topics: Animals; Humans; Mice; Anthrax; Antigens, Bacterial; Bacillus anthracis; Butyrate Response Factor 1; Cell Differentiation; Leukemia, Erythroblastic, Acute; Thrombocytopenia
PubMed: 38542074
DOI: 10.3390/ijms25063102