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Virulence Dec 2024β-N-acetylglucosaminidase (NagZ), a cytosolic glucosaminidase, plays a pivotal role in peptidoglycan recycling. Previous research demonstrated that NagZ knockout...
β-N-acetylglucosaminidase (NagZ), a cytosolic glucosaminidase, plays a pivotal role in peptidoglycan recycling. Previous research demonstrated that NagZ knockout significantly eradicated AmpC-dependent β-lactam resistance in . However, NagZ's role in the virulence of remains unclear. Our study, incorporating data on mouse and larval mortality rates, inflammation markers, and histopathological examinations, revealed a substantial reduction in the virulence of following NagZ knockout. Transcriptome sequencing uncovered differential gene expression between NagZ knockout and wild-type strains, particularly in nucleotide metabolism pathways. Further investigation demonstrated that NagZ deletion led to a significant increase in cyclic diguanosine monophosphate (c-di-GMP) levels. Additionally, transcriptome sequencing and RT-qPCR confirmed significant differences in the expression of ECL_03795, a gene with an unknown function but speculated to be involved in c-di-GMP metabolism due to its EAL domain known for phosphodiesterase activity. Interestingly, in ECL_03795 knockout strains, a notable reduction in the virulence was observed, and virulence was rescued upon complementation with ECL_03795. Consequently, our study suggests that NagZ's function on virulence is partially mediated through the ECL_03795→c-di-GMP pathway, providing insight into the development of novel therapies and strongly supporting the interest in creating highly efficient NagZ inhibitors.
Topics: Animals; Virulence; Mice; Enterobacter cloacae; Larva; Moths; Acetylglucosaminidase; Cyclic GMP; Enterobacteriaceae Infections; Virulence Factors; Bacterial Proteins; Female; Gene Expression Regulation, Bacterial; Gene Knockout Techniques
PubMed: 38912723
DOI: 10.1080/21505594.2024.2367652 -
Advances in Nutrition (Bethesda, Md.) Jun 2024Microbiota in early life is closely associated with the health of infants, especially premature ones. Probiotics are important drivers of gut microbiota development in... (Meta-Analysis)
Meta-Analysis Review
Microbiota in early life is closely associated with the health of infants, especially premature ones. Probiotics are important drivers of gut microbiota development in preterm infants; however, there is no consensus regarding the characteristics of specific microbiota in preterm infants receiving probiotics. In this study, we performed a meta-analysis of 5 microbiome data sets (1816 stool samples from 706 preterm infants) to compare the gut microbiota of preterm infants exposed to probiotics with that of preterm infants not exposed to probiotics across populations. Despite study-specific variations, we found consistent differences in gut microbial composition and predicted functional pathways between the control and probiotic groups across different cohorts of preterm infants. The enrichment of Acinetobacter, Bifidobacterium, and Lactobacillus spp and the depletion of the potentially pathogenic bacteria Finegoldia, Veillonella, and Klebsiella spp. were the most consistent changes in the gut microbiota of preterm infants supplemented with probiotics. Probiotics drove microbiome transition into multiple preterm gut community types, and notably, preterm gut community type 3 had the highest α-diversity, with enrichment of Bifidobacterium and Bacteroides spp. At the functional level, the major predicted microbial pathways involved in peptidoglycan biosynthesis consistently increased in preterm infants supplemented with probiotics; in contrast, the crucial pathways associated with heme biosynthesis consistently decreased. Interestingly, Bifidobacterium sp. rather than Lactobacillus sp. gradually became dominant in gut microbiota of preterm infants using mixed probiotics, although both probiotic strains were administered at the same dosage. Taken together, our meta-analysis suggests that probiotics contribute to reshaping the microbial ecosystem of preterm infants at both the taxonomic and functional levels of the bacterial community. More standardized and relevant studies may contribute to better understanding the crosstalk among probiotics, the gut microbiota, and subsequent disease risk, which could help to give timely nutritional feeding guidance to preterm infants. This systematic review and meta-analysis was registered at PROSPERO (https://www.crd.york.ac.uk/PROSPERO/) as CRD42023447901.
Topics: Humans; Gastrointestinal Microbiome; Probiotics; Infant, Premature; Infant, Newborn; Bifidobacterium; Feces; Bacteria; Lactobacillus; Female
PubMed: 38908894
DOI: 10.1016/j.advnut.2024.100233 -
ACS Infectious Diseases Jun 2024Endolysins, peptidoglycan hydrolases derived from bacteriophages (phages), are being developed as a promising alternative to conventional antibiotics. To obtain highly...
Endolysins, peptidoglycan hydrolases derived from bacteriophages (phages), are being developed as a promising alternative to conventional antibiotics. To obtain highly active endolysins, a diverse library of these endolysins is vital. We propose here microbial single-cell genome sequencing as an efficient tool to discover dozens of previously unknown endolysins, owing to its culture-independent sequencing method. As a proof of concept, we analyzed and recovered endolysin genes within prophage regions of single-amplified genomes in human skin microbiome samples. We constructed a library of chimeric endolysins by shuffling domains of the natural endolysins and performed high-throughput screening against . One of the lead endolysins, bbst1027, exhibited desirable antimicrobial properties, such as rapid bactericidal activity, no detectable resistance development, and in vivo efficacy. We foresee that this endolysin discovery pipeline is in principle applicable to any bacterial target and boost the development of novel antimicrobial agents.
PubMed: 38906534
DOI: 10.1021/acsinfecdis.4c00039 -
Journal of Bacteriology Jun 2024During spore development in bacteria, a polar septum separates two transcriptionally distinct cellular compartments, the mother cell and the forespore. The conserved...
During spore development in bacteria, a polar septum separates two transcriptionally distinct cellular compartments, the mother cell and the forespore. The conserved serine phosphatase SpoIIE is known for its critical role in the formation of this septum and activation of compartment-specific transcription in the forespore. Signaling between the mother cell and forespore then leads to activation of mother cell transcription and a phagocytic-like process called engulfment, which involves dramatic remodeling of the septum and requires a balance between peptidoglycan synthesis and hydrolysis to ensure septal stability and compartmentalization. Using , we identify an additional role for SpoIIE in maintaining septal stability and compartmentalization at the onset of engulfment. This role for SpoIIE is mediated by SpoIIQ, which anchors SpoIIE in the engulfing membrane. A SpoIIQ mutant (SpoIIQ Y28A) that fails to anchor SpoIIE, results in septal instability and miscompartmentalization during septal peptidoglycan hydrolysis, when other septal stabilization factors are absent. Our data support a model whereby SpoIIE and its interactions with the peptidoglycan synthetic machinery contribute to the stabilization of the asymmetric septum early in engulfment, thereby ensuring compartmentalization during spore development.IMPORTANCEBacterial sporulation is a complex process involving a vast array of proteins. Some of these proteins are absolutely critical and regulate key points in the developmental process. Once such protein is SpoIIE, known for its role in the formation of the polar septum, a hallmark of the early stages of sporulation, and activation of the first sporulation-specific sigma factor, σF, in the developing spore. Interestingly, SpoIIE has been shown to interact with SpoIIQ, an important σF-regulated protein that functions during the engulfment stage. However, the significance of this interaction has remained unclear. Here, we unveil the importance of the SpoIIQ-SpoIIE interaction and identify a role for SpoIIE in the stabilization of the polar septum and maintenance of compartmentalization at the onset of engulfment. In this way, we demonstrate that key sporulation proteins, like SpoIIQ and SpoIIE, function in multiple processes during spore development.
PubMed: 38904397
DOI: 10.1128/jb.00220-24 -
Frontiers in Microbiology 2024(), a zoonotic pathogen with a broad host range, presents a substantial threat to global public health safety. Vaccination stands as an effective strategy for the...
BACKGROUND
(), a zoonotic pathogen with a broad host range, presents a substantial threat to global public health safety. Vaccination stands as an effective strategy for the prevention and control of infection, highlighting an immediate clinical need for the creation of safe and efficient attenuated live vaccines.
METHODS
In this study, a peptidoglycan-associated lipoprotein () gene deletion strain (Δ), was constructed. To assess its virulence, we conducted experiments on biofilm formation capability, motility, as well as cell and mouse infection. Subsequently, we evaluated the immune-protective effect of Δ.
RESULTS
It was discovered that deletion of the gene reduced the biofilm formation capability and motility of . Cell infection experiments revealed that the Δ strain exhibited significantly decreased abilities in invasion, adhesion, and intracellular survival, with downregulation of virulence gene expression, including , , , , , , , and . Mouse infection experiments showed that the LD of Δ increased by 10 times, and its colonization ability in mouse tissue organs was significantly reduced. The results indicated that the gene severely affected the virulence of . Further, immunogenicity evaluation of Δ showed a significant enhancement in the lymphocyte transformation proliferation capability of immunized mice, producing high titers of specific IgG and IgA, suggesting that Δ possesses good immunogenicity. Challenge protection tests demonstrated that the strain could provide 100% immune protection against wild-type strains in mice.
DISCUSSION
This study proves that the gene influences the virulence of , and Δ could serve as a candidate strain for attenuated live vaccines, laying the foundation for the development of attenuated live vaccines against .
PubMed: 38903796
DOI: 10.3389/fmicb.2024.1422202 -
Inorganic Chemistry Jun 2024Pathogenic bacteria have consistently posed a formidable challenge to human health, creating the critical need for effective antibacterial solutions. In response,...
Pathogenic bacteria have consistently posed a formidable challenge to human health, creating the critical need for effective antibacterial solutions. In response, enzyme-metal-organic framework (MOF) composites have emerged as a promising class of antibacterial agents. This study focuses on the development of an enzyme-MOF composite based on HZIF-8, incorporating the advantages of simple synthesis, ZIF-8 antibacterial properties, lysozyme hydrolysis, and high biological safety. Through a one-pot method, core-shell nanoparticles (HZIF-8) were synthesized. This structure enables efficient immobilization of lysozyme and lactoferrin within the HZIF-8, resulting in the formation of the lysozyme-lactoferrin@HZIF-8 (LYZ-LF@HZIF-8) composite. Upon exposure to light irradiation, HZIF-8 itself possessed antibacterial properties. Lysozyme initiated the degradation of bacterial peptidoglycan and lactoferrin synergistically enhanced the antibacterial effect of lysozyme. All of the above ultimately contributed to comprehensive antibacterial activity. Antibacterial assessments demonstrated the efficacy of the LYZ-LF@HZIF-8 composite, effectively eradicating at a cell density of 1.5 × 10 CFU/mL with a low dosage of 200 μg/mL and completely inactivating at 400 μg/mL with the same cell density. The enzyme-MOF composite exhibited significant and durable antibacterial efficacy, with no apparent cytotoxicity in vitro, thereby unveiling expansive prospects for applications in the medical and food industries.
PubMed: 38902911
DOI: 10.1021/acs.inorgchem.4c02082 -
Infection and Immunity Jun 2024Toll-like receptor 9 (TLR9) is an innate immune receptor that localizes to endosomes in antigen presenting cells and recognizes single stranded unmethylated CpG sites on...
Toll-like receptor 9 (TLR9) is an innate immune receptor that localizes to endosomes in antigen presenting cells and recognizes single stranded unmethylated CpG sites on bacterial genomic DNA (gDNA). Previous bioinformatic studies have demonstrated that the genome of the human pathogen contains TLR9 stimulatory motifs, and correlative studies have implied a link between human TLR9 (hTLR9) genotype variants and susceptibility to infection. Here, we present our evaluation of the stimulatory potential of gDNA and its recognition by hTLR9- and murine TLR9 (mTLR9)-expressing cells. Utilizing reporter cell lines, we demonstrate that purified gDNA from can stimulate hTLR9 signaling, albeit at lower levels than gDNA prepared from other Gram-negative bacteria. Interestingly, we found that while is capable of signaling through hTLR9 and mTLR9 during live infections in HEK293 reporter cell lines, signaling only occurs at later developmental time points. Chlamydia-specific induction of hTLR9 is blocked when protein synthesis is inhibited prior to the RB-to-EB conversion, exacerbated by the inhibition of lipooligosaccharide biosynthesis, and is significantly altered during the induction of aberrance/persistence. Our observations support the hypothesis that chlamydial gDNA is released during the conversion between the pathogen's replicative and infectious forms and during treatment with antibiotics targeting peptidoglycan assembly. Given that inclusions do not co-localize with TLR9-containing vacuoles in the pro-monocytic cell line U937, our findings also hint that chlamydial gDNA is capable of egress from the inclusion, and traffics to TLR9-containing vacuoles via an as yet unknown pathway.
PubMed: 38899879
DOI: 10.1128/iai.00063-24 -
EJNMMI Radiopharmacy and Chemistry Jun 2024Infection remains a major cause of morbidity and mortality, regardless of advances in antimicrobial therapy and improved knowledge of microorganisms. With the major... (Review)
Review
BACKGROUND
Infection remains a major cause of morbidity and mortality, regardless of advances in antimicrobial therapy and improved knowledge of microorganisms. With the major global threat posed by antimicrobial resistance, fast and accurate diagnosis of infections, and the reliable identification of intractable infection, are becoming more crucial for effective treatment and the application of antibiotic stewardship. Molecular imaging with the use of nuclear medicine allows early detection and localisation of infection and inflammatory processes, as well as accurate monitoring of treatment response. There has been a continuous search for more specific radiopharmaceuticals to be utilised for infection imaging. This review summarises the most prominent discoveries in specifically bacterial infection imaging agents over the last five years, since 2019.
MAIN BODY
Some promising new radiopharmaceuticals evaluated in patient studies are reported here, including radiolabelled bacterial siderophores like [Ga]Ga-DFO-B, radiolabelled antimicrobial peptide/peptide fragments like [Ga]Ga-NOTA-UBI29-41, and agents that target bacterial synthesis pathways (folic acid and peptidoglycan) like [C]para-aminobenzoic acid and D-methyl-[C]-methionine, with clinical trials underway for [F]fluorodeoxy-sorbitol, as well as for C- and F-labelled trimethoprim.
CONCLUSION
It is evident that a great deal of effort has gone into the development of new radiopharmaceuticals for infection imaging over the last few years, with remarkable progress in preclinical investigations. However, translation to clinical trials, and eventually clinical Nuclear Medicine practice, is apparently slow. It is the authors' opinion that a more structured and harmonised preclinical setting and well-designed clinical investigations are the key to reliably evaluate the true potential of the newly proposed infection imaging agents.
PubMed: 38896373
DOI: 10.1186/s41181-024-00279-7 -
Scientific Reports Jun 2024Penicillin binding proteins (PBPs) are involved in biosynthesis, remodeling and recycling of peptidoglycan (PG) in bacteria. PBP-A from Thermosynechococcus elongatus...
Penicillin binding proteins (PBPs) are involved in biosynthesis, remodeling and recycling of peptidoglycan (PG) in bacteria. PBP-A from Thermosynechococcus elongatus belongs to a cyanobacterial family of enzymes sharing close structural and phylogenetic proximity to class A β-lactamases. With the long-term aim of converting PBP-A into a β-lactamase by directed evolution, we simulated what may happen when an organism like Escherichia coli acquires such a new PBP and observed growth defect associated with the enzyme activity. To further explore the molecular origins of this harmful effect, we decided to characterize deeper the activity of PBP-A both in vitro and in vivo. We found that PBP-A is an enzyme endowed with DD-carboxypeptidase and DD-endopeptidase activities, featuring high specificity towards muropeptides amidated on the D-iso-glutamyl residue. We also show that a low promiscuous activity on non-amidated peptidoglycan deteriorates E. coli's envelope, which is much higher under acidic conditions where substrate discrimination is mitigated. Besides expanding our knowledge of the biochemical activity of PBP-A, this work also highlights that promiscuity may depend on environmental conditions and how it may hinder rather than promote enzyme evolution in nature or in the laboratory.
Topics: Escherichia coli; Hydrogen-Ion Concentration; Penicillin-Binding Proteins; Peptidoglycan; Substrate Specificity; Cyanobacteria; Bacterial Proteins; Synechococcus
PubMed: 38890528
DOI: 10.1038/s41598-024-64806-x -
Archives of Virology Jun 2024The inflammasome is a multimeric protein complex that plays a vital role in the defence against pathogens and is therefore considered an essential component of the...
In vitro analysis of the expression of inflammasome, antiviral, and immune genes in an Oreochromis niloticus liver cell line following stimulation with bacterial ligands and infection with tilapia lake virus.
The inflammasome is a multimeric protein complex that plays a vital role in the defence against pathogens and is therefore considered an essential component of the innate immune system. In this study, the expression patterns of inflammasome genes (NLRC3, ASC, and CAS-1), antiviral genes (IFNγ and MX), and immune genes (IL-1β and IL-18) were analysed in Oreochromis niloticus liver (ONIL) cells following stimulation with the bacterial ligands peptidoglycan (PGN) and lipopolysaccharide (LPS) and infection with TiLV. The cells were stimulated with PGN and LPS at concentrations of 10, 25, and 50 µg/ml. For viral infection, 10 TCID of TiLV per ml was used. After LPS stimulation, all seven genes were found to be expressed at specific time points at each of the three doses tested. However, at even higher doses of LPS, NLRC3 levels decreased. Following TiLV infection, all of the genes showed significant upregulation, especially at early time points. However, the gene expression pattern was found to be unique in PGN-treated cells. For instance, NLRC3 and ASC did not show any response to PGN stimulation, and the expression of IFNγ was downregulated at 25 and 50 µg of PGN per ml. CAS-1 and IL-18 expression was downregulated at 25 µg of PGN per ml. At a higher dose (50 µg/ml), IL-1β showed downregulation. Overall, our results indicate that these genes are involved in the immune response to viral and bacterial infection and that the degree of response is ligand- and dose-dependent.
Topics: Animals; Cichlids; Inflammasomes; Fish Diseases; Cell Line; Peptidoglycan; Liver; Lipopolysaccharides; Immunity, Innate; Fish Proteins; Interleukin-18; Ligands; DNA Virus Infections; Gene Expression Regulation; Interleukin-1beta
PubMed: 38888759
DOI: 10.1007/s00705-024-06077-5