Karl Ocius

J Pharm Biomed Anal. 2022 Apr 1;215:114749. doi: 10.1016/j.jpba.2022.114749. Online ahead of print.

ABSTRACT

Tuberculosis (TB), caused by Mycobacterium tuberculosis (MTB), is one of the 10 leading causes of death worldwide, especially in low-income areas. A rapid, low-cost diagnostic assay for TB with high sensitivity and specificity is not currently available. Bio-functionalized magnetic nanoparticles (MNPs) which are able to efficiently detect and concentrate biomolecules from complex biological samples, allows improving the diagnostic immunoassays. In this way, a proof-of-concept of MNP-based sandwich immunoassay was developed to detect various MTB protein antigens. The superficial and secretory antigenic proteins considered in this research were: CFP10, ESAT6, MTC28, MPT64, 38 kDa protein, Ag85B, and MoeX. The proteins were cloned and expressed in an E. coli system. Polyclonal antibodies (ab) against the recombinant antigens were elicited in rabbits and mice. Antibodies were immobilized on the surface of amine-silanized nanoparticles (MNP@Si). The functionalized MNP@Si@ab were tested in a colorimetric sandwich enzyme-linked immunosorbent assay (sELISA-MNP@Si@ab) to recognize the selected antigens in sputum samples. The selected MTB antigens were successfully detected in sputum from TB patients in a shorter time (~ 4 h) using the sELISA-MNP@Si@ab, compared to the conventional sELISA (~15 h) standardized in home. Moreover, the sELISA-MNP@Si@ab showed the higher sensitivity in the real biological samples from infected patients.

PMID:35447489 | DOI:10.1016/j.jpba.2022.114749

Bioimpacts. 2022;12(2):139-146. doi: 10.34172/bi.2021.23378. Epub 2021 Oct 9.

ABSTRACT

Introduction: With the outbreak of coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the interaction between the host and SARS-CoV-2 was widely studied. However, it is unclear whether and how SARS-CoV-2 infection affects lung microflora, which contribute to COVID-19 complications. Methods: Here, we analyzed the metatranscriptomic data of bronchoalveolar lavage fluid (BALF) of 19 COVID-19 patients and 23 healthy controls from 6 independent projects and detailed the active microbiota landscape in both healthy individuals and COVID-19 patients. Results: The infection of SARS-CoV-2 could deeply change the lung microbiota, evidenced by the α-diversity, β-diversity, and species composition analysis based on bacterial microbiota and virome. Pathogens (e.g., Klebsiella oxytoca causing pneumonia as well), immunomodulatory probiotics (e.g., lactic acid bacteria and Faecalibacterium prausnitzii, a butyrate producer), and Tobacco mosaic virus (TMV) were enriched in the COVID-19 group, suggesting a severe microbiota dysbiosis. The significant correlation between Rothia mucilaginosa, TMV, and SARS-CoV-2 revealed drastic inflammatory battles between the host, SARS-CoV-2, and other microbes in the lungs. Notably, TMV only existed in the COVID-19 group, while human respirovirus 3 (HRV 3) only existed in the healthy group. Our study provides insights into the active microbiota in the lungs of COVID-19 patients and would contribute to the understanding of the infection mechanism of SARS-CoV-2 and the treatment of the disease and complications. Conclusion: SARS-COV-2 infection deeply altered the lung microbiota of COVID-19 patients. The enrichment of several other pathogens, immunomodulatory probiotics (lactic acid or butyrate producers), and TMV in the COVID-19 group suggests a complex and active lung microbiota disorder.

PMID:35411293 | PMC:PMC8905590 | DOI:10.34172/bi.2021.23378

Animal Model Exp Med. 2022 Apr 12. doi: 10.1002/ame2.12227. Online ahead of print.

ABSTRACT

Zebrafish (Danio rerio) have attracted much attention over the past decade as a reliable model for gut microbiome research. Owing to their low cost, strong genetic and development coherence, efficient preparation of germ-free (GF) larvae, availability in high-throughput chemical screening, and fitness for intravital imaging in vivo, zebrafish have been extensively used to investigate microbiome-host interactions and evaluate the toxicity of environmental pollutants. In this review, the advantages and disadvantages of zebrafish for studying the role of the gut microbiome compared with warm-blooded animal models are first summarized. Then, the roles of zebrafish gut microbiome on host development, metabolic pathways, gut-brain axis, and immune disorders and responses are addressed. Furthermore, their applications for the toxicological assessment of aquatic environmental pollutants and exploration of the molecular mechanism of pathogen infections are reviewed. We highlight the great potential of the zebrafish model for developing probiotics for xenobiotic detoxification, resistance against bacterial infection, and disease prevention and cure. Overall, the zebrafish model promises a brighter future for gut microbiome research.

PMID:35415967 | DOI:10.1002/ame2.12227

RSC Adv. 2022 Jan 5;12(2):1105-1120. doi: 10.1039/d1ra07594b. eCollection 2021 Dec 22.

ABSTRACT

Invasive bacterial and fungal infections have notably increased the burden on the health care system and especially in immune compromised patients. These invasive bacterial and fungal species mimic and interact with the host extracellular matrix and increase the adhesion and internalization into the host system. Further, increased resistance of traditional antibiotics/antifungal drugs led to the demand for other therapeutics and preventive measures. Presently, metallic nanoparticles have wide applications in health care sectors. The present study has been designed to evaluate the advantage of Ag/Sn-SnO₂ composite nanoparticles over the single oxide/metallic nanoparticles. By using in silico molecular docking approaches, herein we have evaluated the effects of Ag/Sn-SnO₂ nanoparticles on adhesion and invasion responsible molecular targets such as LpfD (E. coli), Als3 (C. albicans) and on virulence/resistance causing PqsR (P. aeruginosa), RstA (Bmfr) (A. baumannii), FoxA (K. pneumonia), Hsp90 and Cyp51 (C. albicans). These Ag/Sn-SnO₂ nanoparticles exhibited higher antimicrobial activities, especially against the C. albicans, which are the highest ever reported results. Further, Ag/Sn-SnO₂ NPs exhibited interaction with the heme proionate residues such as Lys143, His468, Tyr132, Arg381, Phe105, Gly465, Gly464, Ile471 and Ile304 by forming hydrogen bonds with the Arg 381 residue of lanosterol 1 4α-demethylase and increased the inhibition of the Candida strains. Additionally, the Ag/Sn-SnO₂ nanoparticles exhibited extraordinary inhibitory properties by targeting different proteins of bacteria and Candida species followed by several molecular pathways which indicated that it can be used to eliminate the resistance to traditional antibiotics.

PMID:35425144 | PMC:PMC8978812 | DOI:10.1039/d1ra07594b

Cell Host Microbe. 2022 Apr 13;30(4):556-569.e5. doi: 10.1016/j.chom.2022.03.014.

ABSTRACT

Abundance and diversity of bacteria and their viral predators, bacteriophages (phages), in the digestive tract are associated with human health. Particularly intriguing is the long-term coexistence of these two antagonistic populations. We performed genome-wide RNA sequencing on a human enteroaggregative Escherichia coli isolate to identify genes differentially expressed between in vitro conditions and in murine intestines. We experimentally demonstrated that four of these differentially expressed genes modified the interactions between E. coli and three virulent phages by either increasing or decreasing its susceptibility/resistance pattern and also by interfering with biofilm formation. Therefore, the regulation of bacterial genes expression during the colonization of the digestive tract influences the coexistence of phages and bacteria, highlighting the intricacy of tripartite relationships between phages, bacteria, and the animal host in intestinal homeostasis.

PMID:35421351 | DOI:10.1016/j.chom.2022.03.014

J Med Microbiol. 2022 Apr;71(4). doi: 10.1099/jmm.0.001512.

ABSTRACT

Introduction. Evidence has linked exogenous and endogenous sex hormones with the human microbiome.Hypothesis/Gap statement. The longitudinal effects of oral contraceptives (OC) on the human gut microbiome have not previously been studied.Aim. We sought to examine the longitudinal impact of OC use on the taxonomic composition and metabolic functions of the gut microbiota and endogenous sex steroid hormones after initiation of OC use.Methodology. We recruited ten healthy women who provided blood and stool samples prior to OC use, 1 month and 6 months after starting OC. We measured serum levels of sex hormones, including estradiol, progesterone, sex hormone-binding globulin (SHBG), and total testosterone. Shotgun metagenomic sequencing was performed on DNA extracted from faecal samples. Species and metabolic pathway abundances were determined using MetaPhlAn2 and HUMAnN2. Multivariate association with linear models was used to identify microbial species and metabolic pathways associated with OC use and endogenous levels of sex hormones.Results. The percentage variance of the microbial community explained by individual factors ranged from 9.9 % for age to 2.7 % for time since initiation of OC use. We observed no changes in the diversity or composition of the gut microbiome following OC initiation. However, the relative abundance of the biosynthesis pathways of peptidoglycan, amino acids (lysine, threonine, methionine, and tryptophan), and the NAD salvage pathway increased after OC initiation. In addition, serum levels of estradiol and SHBG were positively associated with Eubacterium ramulus, a flavonoid-degrading bacterium. Similarly, microbes involving biosynthesis of l-lysine, l-threonine, and l-methionine were significantly associated with lower estradiol, SHBG, and higher levels of total testosterone.Conclusion. Our study provides the first piece of evidence supporting the association between exogenous and endogenous sex hormones and gut microbiome composition and function.

PMID:35452382 | DOI:10.1099/jmm.0.001512

Glycobiology. 2022 Apr 22:cwac027. doi: 10.1093/glycob/cwac027. Online ahead of print.

ABSTRACT

Akkermansia muciniphila is an intestinal symbiont known to improve the gut barrier function in mice and humans. Various cell envelope components have been identified to play a critical role in the immune signalling of A. muciniphila, but the chemical composition and role of peptidoglycan (PG) remained elusive. Here, we isolated PG fragments from A. muciniphila MucT (ATCC BAA-835), analysed their composition and evaluated their immune signalling capacity. Structurally, the PG of A. muciniphila was found to be noteworthy due of the presence of some non-acetylated glucosamine residues, which presumably stems from deacetylation of N-acetylglucosamine. Some of the N-acetylmuramic acid (MurNAc) subunits were O-acetylated. The immunological assays revealed that muropeptides released from the A. muciniphila PG could both activate the intracellular NOD1 and NOD2 receptors to a comparable extent as muropeptides from Escherichia coli BW25113. These data challenge the hypothesis that N-non-acetylation of PG can be used as a NOD-1 evasion mechanism. Our results provide new insights into the diversity of cell envelope structures of key gut microbiota members and their role in steering host-microbiome interactions.

PMID:35452117 | DOI:10.1093/glycob/cwac027

Front Vet Sci. 2022 Mar 29;9:819921. doi: 10.3389/fvets.2022.819921. eCollection 2022.

ABSTRACT

Staphylococcus aureus is a common pathogen that can cause pneumonia and a variety of skin diseases. Skin injuries have a high risk of colonization by S. aureus, which increases morbidity and mortality. Due to the emergence of multidrug-resistant strains, antimicrobial peptides are considered to be among the best alternatives to antibiotics due to their unique mechanism of action and other characteristics. MPX is an antibacterial peptide extracted from wasp venom that has antibacterial activity against a variety of bacteria. This study revealed that MPX has good bactericidal activity against S. aureus and that its minimum inhibitory concentration (MIC) is 0.08 μM. MPX (4×MIC) can kill 99.9% of bacteria within 1 h, and MPX has good stability. The research on the bactericidal mechanism found that MPX could destroy the membrane integrity, increase the membrane permeability, change the membrane electromotive force, and cause cellular content leakage, resulting in bactericidal activity. Results from a mouse scratch model experiment results show that MPX can inhibit colonization by S. aureus, which reduces the wound size, decreases inflammation, and promotes wound healing. This study reports the activity of MPX against S. aureus and its mechanism and reveals the ability of MPX to treat S. aureus infection in mice, laying the foundation for the development of new drugs for bacterial infections.

PMID:35425831 | PMC:PMC9002018 | DOI:10.3389/fvets.2022.819921

Science. 2022 Apr 15;376(6590):248-249. doi: 10.1126/science.abo7933. Epub 2022 Apr 14.

ABSTRACT

Peptidoglycans from gut microbiota modulate appetite through hypothalamic circuits.

PMID:35420955 | DOI:10.1126/science.abo7933

Int J Mol Sci. 2022 Mar 24;23(7):3537. doi: 10.3390/ijms23073537.

ABSTRACT

The synthesis of a peptidoglycan septum is a fundamental part of bacterial fission and is driven by a multiprotein dynamic complex called the divisome. FtsW and FtsI are essential proteins that synthesize the peptidoglycan septum and are controlled by the regulatory FtsBLQ subcomplex and the activator FtsN. However, their mode of regulation has not yet been uncovered in detail. Understanding this process in detail may enable the development of new compounds to combat the rise in antibiotic resistance. In this review, recent data on the regulation of septal peptidoglycan synthesis is summarized and discussed. Based on structural models and the collected data, multiple putative interactions within FtsWI and with regulators are uncovered. This elaborates on and supports an earlier proposed model that describes active and inactive conformations of the septal peptidoglycan synthesis complex that are stabilized by these interactions. Furthermore, a new model on the spatial organization of the newly synthesized peptidoglycan and the synthesis complex is presented. Overall, the updated model proposes a balance between several allosteric interactions that determine the state of septal peptidoglycan synthesis.

PMID:35408901 | PMC:PMC8998562 | DOI:10.3390/ijms23073537

Neuropsychiatr Dis Treat. 2022 Apr 5;18:749-760. doi: 10.2147/NDT.S355723. eCollection 2022.

ABSTRACT

BACKGROUND: Gut microbiota is associated with anxiety and depression, while exercise has been proved to alleviate depressive symptoms. However, the interaction of exercise, depression, and gut microbiota remains unclear.

METHODS: Male C57/BL6J mice were exposed to chronic unpredictable mild stress (CUMS) for 6 weeks and then were subjected to a 5-week swimming program. Behavioral tests, including sucrose preference test (SPT), open field test (OFT), elevated plus-maze (EPM) test, and tail suspension test (TST), were conducted to assess the anxiety-like and depressive behaviors. Gut microbiota analysis was carried out after sample collection.

RESULTS: This study showed that CUMS induced depressive behaviors, but swimming exercise increased sucrose preference rate in the SPT, increased time in the center and number of rearing in the OFT, decreased time in the closed arm and increased time in the open arm in EPM, and decreased immobility time in the TST. Firmicutes were the predominant phylum in the gut microbiome, followed by the phyla Bacteroidetes and Proteobacteria. We further found that CUMS and swimming influenced the relative abundance of the genus Desulfovibrio, genus Streptococcus, genus p-75-a5. Among the metabolic pathways, aromatic biogenic amine degradation (PWY-7431), mono-trans and polycis decaprenyl phosphate biosynthesis (PWY-6383), chlorosalicylate degradation (PWY-6107), mycothiol biosynthesis (PWY1G-0), mycolyl-arabinogalactan-peptidoglycan complex biosynthesis (PWY-6397), toluene degradation I (aerobic) (via o-cresol) (PWY-5180), toluene degradation II (aerobic) (via 4-methylcatechol) (PWY-5182), and starch degradation III (PWY-6731) may be related to the mechanism of anti-depression effect.

CONCLUSION: Swimming exercise reverses CUMS-induced depressive behaviors, and the alteration of gut microbiota composition and regulation of microbiota metabolic pathways are involved.

PMID:35411144 | PMC:PMC8994653 | DOI:10.2147/NDT.S355723

Methods Enzymol. 2022;665:73-103. doi: 10.1016/bs.mie.2021.12.004. Epub 2022 Jan 13.

ABSTRACT

Glycan-protein interactions facilitate some of the most important biomolecular processes in and between cells. They are involved in different cellular pathways, cell-cell interactions and associated with many diseases, making these interactions of great interest. However, their structural and functional diversity poses great challenges in studying them at the molecular level. Surface plasmon resonance (SPR) technology presents great advantages to study glycan-protein interactions due to its superior sensitivity, ability to monitor real-time interactions, relatively simple data interpretation, and most importantly, direct measurement of binding without a need for fluorescent labeling. Here, another dimensionality of SPR in studying glycan-protein interactions is demonstrated via examples of binding between human innate immune receptors and their bacterial peptidoglycan ligands. In order to best resemble interactions in solution, a novel strategy of tethering the carbohydrate at different positions to the biosensor surface is applied to represent the potential displays of the carbohydrate ligand to the receptor. Subsequent kinetic analysis provides insights into the optimized configuration of peptidoglycan fragments for binding with its receptors. The manuscript contains a "how-to guide" to help with the implementation of these methods in other glycan-protein binding systems.

PMID:35379444 | DOI:10.1016/bs.mie.2021.12.004

PLoS Pathog. 2022 Mar 14;18(3):e1010370. doi: 10.1371/journal.ppat.1010370. eCollection 2022 Mar.

ABSTRACT

Borrelia species are amino acid auxotrophs that utilize di- and tri- peptides obtained through their oligopeptide transport system to supply amino acids for replicative growth during their enzootic cycles. However, Borrelia species from both the Lyme disease (LD) and relapsing fever (RF) groups harbor an amino acid transport and catabolism system, the Arginine Deiminase System (ADI), that could potentially augment intracellular L-arginine required for growth. RF spirochetes contain a "complete", four gene ADI (arcA, B, D, and C) while LD spirochetes harbor arcA, B, and sometimes D but lack arcC (encoding carbamate kinase). In this study, we evaluated the role of the ADI system in bacterial survival and virulence and discovered important differences in RF and LD ADIs. Both in vitro and in a murine model of infection, B. hermsii cells significantly reduced extracellular L-arginine levels and that reduction was dependent on arginine deiminase expression. Conversely, B. burgdorferi did not reduce the concentration of L-arginine during in vitro growth experiments nor during infection of the mammalian host, suggesting a fundamental difference in the ability to directly utilize L-arginine compared to B. hermsii. Further experiments using a panel of mutants generated in both B. burgdorferi and B. hermsii, identified important differences in growth characteristics and ADI transcription and protein expression. We also found that the ADI system plays a key role in blood and spleen colonization in RF spirochetes. In this study we have identified divergent metabolic strategies in two closely related human pathogens, that ultimately impacts the host-pathogen interface during infection.

PMID:35286343 | PMC:PMC8947608 | DOI:10.1371/journal.ppat.1010370

Nutrients. 2022 Jan 30;14(3):608. doi: 10.3390/nu14030608.

ABSTRACT

Autism spectrum disorders (ASD) consist of a range of neurodevelopmental conditions accompanied by dysbiosis of gut microbiota. Therefore, a number of microbiota manipulation strategies were developed to restore their balance. However, a comprehensive comparison of the various methods on gut microbiota is still lacking. Here, we evaluated the effect of Bifidobacterium (BF) treatment and fecal microbiota transplantation (FT) on gut microbiota in a propionic acid (PPA) rat model of autism using 16S rRNA sequencing. Following PPA treatment, gut microbiota showed depletion of Bacteroidia and Akkermansia accompanied by a concomitant increase of Streptococcus, Lachnospiraceae, and Paraeggerthella. The dysbiosis was predicted to cause increased levels of porphyrin metabolism and impairments of acyl-CoA thioesterase and ubiquinone biosynthesis. On the contrary, BF and FT treatments resulted in a distinct increase of Clostridium, Bifidobacterium, Marvinbryantia, Butyricicoccus, and Dorea. The taxa in BF group positively correlated with vitamin B12 and flagella biosynthesis, while FT mainly enriched flagella biosynthesis. In contrast, BF and FT treatments negatively correlated with succinate biosynthesis, pyruvate metabolism, nitrogen metabolism, beta-Lactam resistance, and peptidoglycan biosynthesis. Therefore, the present study demonstrated that BF and FT treatments restored the PPA-induced dysbiosis in a treatment-specific manner.

PMID:35276971 | PMC:PMC8838423 | DOI:10.3390/nu14030608

Adv Sci (Weinh). 2022 Mar 11:e2105807. doi: 10.1002/advs.202105807. Online ahead of print.

ABSTRACT

Due to the complexity, aggressiveness, and heterogeneity of malignant melanoma, it is difficult to eradicate the whole tumor through conventional treatment. Herein, a strategy of metabolic engineering labeled anaerobic oncolytic bacteria (Clostridium butyricum) is demonstrated to achieve the ablation of melanoma. In this system, the metabolic substrate of C. butyricum d-alanine (d-Ala) is first conjugated with a photosensitizer (TPApy) showing aggregation-induced emission (AIE). The yielded metabolic substrate of d-Ala-TPAPy can be metabolically incorporated into bacterial peptidoglycan to form engineered C. Butyricum. Once the engineered C. butyricum is injected into melanoma, the bacteria can only proliferate in an anaerobic zone, stimulate the tumor immune microenvironment, and ablate the tumor hypoxia region. Following that, the relatively rich oxygen content in the peripheral area can induce the death of C. butyricum. The photosensitizer (PS) on the bacteria can subsequently exert a photodynamic effect in the oxygen-rich region and further remove the melanoma residue under light irradiation. Prominent in vivo melanoma ablation results revealed that the engineering oncolytic bacteria can provide a promising regime for solid tumor eradication.

PMID:35277932 | DOI:10.1002/advs.202105807

Bioorg Chem. 2021 Nov 29;119:105532. doi: 10.1016/j.bioorg.2021.105532. Online ahead of print.

ABSTRACT

Microbial polysaccharides composed of N-acetylglucosamine (GlcNAc), such as chitin, peptidoglycan and poly-β-(1 → 6)-GlcNAc (dPNAG), play a critical role in maintaining cell integrity or in facilitating biofilm formation in numerous fungal and bacterial pathogens. Glycosyl hydrolase enzymes that catalyze the degradation of these β-GlcNAc containing polysaccharides play important roles in normal microbial cell physiology and can also be exploited as biocatalysts with applications as anti-fungal, anti-bacterial, or biofilm dispersal agents. Assays to rapidly detect and characterize the activity of such glycosyl hydrolase enzymes can facilitate their development as biocatalyst, however, currently available probes such as 4-methylumbelliferyl-β-GlcNAc (4MU-GlcNAc) are not universally accepted as substrates, and their fluorescent signal is sensitive to changes in pH. Here, we present the development of a new multifunctional fluorescent substrate analog for the detection and characterization of hexosaminidase enzyme activity containing a 7-amino-4-methyl coumarin (AMC) carbamate aglycone. This probe is widely tolerated as a substrate for exo-acting β-hexosaminidase, family 19 endo-chitinase, and the dPNAG hydrolase enzyme Dispersin B (DspB) and enables detection of hexosaminidase enzyme activity via either single wavelength fluorescent measurements or ratiometric fluorescent detection. We demonstrate the utility of this probe to screen for recombinant DspB activity in Escherichia coli cell lysates, and for the development of a high-throughput assay to screen for DspB inhibitors.

PMID:34883361 | DOI:10.1016/j.bioorg.2021.105532

Microbiol Immunol. 2021 Dec 10. doi: 10.1111/1348-0421.12958. Online ahead of print.

ABSTRACT

Streptococcus mutans, a cariogenic pathogen, adheres to the tooth surface and forms a biofilm. Bacterial cell surface proteins are associated with adherence to substrates. Sortase A (SrtA) mediates the localization of proteins with an LPXTG motif-containing proteins to the cell surface by covalent binding to peptidoglycan. In S. mutans UA159, 6 SrtA-dependent proteins, SpaP, WapA, WapE, DexA, FruA, and GbpC, were identified. Although some of these proteins were characterized, a comprehensive analysis of the 6 proteins has not been reported. In this study, we constructed mutants deficient in each of these proteins and the SrtA-deficient mutant. The SrtA-deficient mutant showed drastically decreased binding to salivary components, biofilm formation, bacterial coaggregation activity, hydrophobicity, and cellular matrix binding (collagen type I, fibronectin, and laminin). The SpaP-deficient mutant showed significantly reduced binding to salivary components and partially increased coaggregation with Porphyromonas gingivalis, and decreased hydrophobicity, and collagen binding. The WapA-deficient mutant showed slightly decreased coaggregation with Fusobacterium nucleatum. Although the SrtA-deficient mutant showed drastically altered phenotypes, all SrtA-dependent protein-deficient mutants, except the SpaP-deficient mutant, did not show considerable alterations in binding to salivary components. These results indicate that the 6 proteins may coordinately contribute to these activities. In addition, using genomic data of 125 S. mutans strains, we compared the amino acid sequences of each surface protein and found many variations among strains, which may affect the phenotype of cell surface proteins in S. mutans. This article is protected by copyright. All rights reserved.

PMID:34888908 | DOI:10.1111/1348-0421.12958

Anal Chim Acta. 2022 Jan 2;1189:339213. doi: 10.1016/j.aca.2021.339213. Epub 2021 Oct 27.

ABSTRACT

Staphylococcus aureus is a common foodborne pathogen that can cause a suppurative infection after eating contaminated food. Detection of S. aureus plays an important role in the food industry. In this study, a strategy for the detection of S. aureus using magnetic separation (MS) technology combined with rolling circle amplification (MS-RCA) was proposed. The strategy used antibiotics to capture bacteria and employed RCA products as signal output probes. Vancomycin (Van), as a commonly used antibiotic, can recognize peptidoglycan on the cell wall of Gram-positive bacteria and can effectively identify target bacteria. Therefore, we prepared BSAylated-Van functionalized magnetic beads (Van-MBs) for the pre-enrichment of S. aureus. To ensure the selectivity of this method, we used biotin-pig IgG to bind S. aureus. In addition, to amplify the output signal of the MS-RCA strategy, we introduced streptavidin (SA) and successfully obtained the Van-MBs@S. aureus@biotin-pig IgG@SA@biotin-RCA probe complex and used the biotin-avidin-system (BAS) by combining magnetic separation technology and RCA technology to realize the enrichment and specific detection of S. aureus. Furthermore, by optimizing the experimental conditions such as the magnetic separation time and the amount of Van-MBs, the detection performance of this method was improved. Under the optimal conditions, the detection limit of this method for S. aureus was 3.3 × 10² CFU/mL in fruit juice, and it was less affected by other bacteria.

PMID:34815035 | DOI:10.1016/j.aca.2021.339213

Proc Natl Acad Sci U S A. 2021 Nov 23;118(47):e2108244118. doi: 10.1073/pnas.2108244118.

ABSTRACT

Emerging antibiotic resistance demands identification of novel antibacterial compound classes. A bacterial whole-cell screen based on pneumococcal autolysin-mediated lysis induction was developed to identify potential bacterial cell wall synthesis inhibitors. A hit class comprising a 1-amino substituted tetrahydrocarbazole (THCz) scaffold, containing two essential amine groups, displayed bactericidal activity against a broad range of gram-positive and selected gram-negative pathogens in the low micromolar range. Mode of action studies revealed that THCz inhibit cell envelope synthesis by targeting undecaprenyl pyrophosphate-containing lipid intermediates and thus simultaneously inhibit peptidoglycan, teichoic acid, and polysaccharide capsule biosynthesis. Resistance did not readily develop in vitro, and the ease of synthesizing and modifying these small molecules, as compared to natural lipid II-binding antibiotics, makes THCz promising scaffolds for development of cell wall-targeting antimicrobials.

PMID:34785593 | DOI:10.1073/pnas.2108244118

PLoS One. 2021 Nov 16;16(11):e0259181. doi: 10.1371/journal.pone.0259181. eCollection 2021.

ABSTRACT

Mycobacterium tuberculosis, the causative agent of tuberculosis remains a global health concern, further compounded by the high rates of HIV-TB co-infection and emergence of multi- and extensive drug resistant TB, all of which have hampered efforts to eradicate this disease. As a result, novel anti-tubercular interventions are urgently required, with the peptidoglycan component of the M. tuberculosis cell wall emerging as an attractive drug target. Peptidoglycan M23 endopeptidases can function as active cell wall hydrolases or degenerate activators of hydrolases in a variety of bacteria, contributing to important processes such as bacterial growth, division and virulence. Herein, we investigate the function of the Rv0950-encoded putative M23 endopeptidase in M. tuberculosis. In silico analysis revealed that this protein is conserved in mycobacteria, with a zinc-binding catalytic site predictive of hydrolytic activity. Transcript analysis indicated that expression of Rv0950c was elevated during lag and log phases of growth and reduced in stationary phase. Deletion of Rv0950c yielded no defects in growth, colony morphology, antibiotic susceptibility or intracellular survival but caused a reduction in cell length. Staining with a monopeptide-derived fluorescent D-amino acid, which spatially reports on sites of active PG biosynthesis or repair, revealed an overall reduction in uptake of the probe in ΔRv0950c. When stained with a dipeptide probe in the presence of cell wall damaging agents, the ΔRv0950c mutant displayed reduced sidewall labelling. As bacterial peptidoglycan metabolism is important for survival and pathogenesis, the role of Rv0950c and other putative M23 endopeptidases in M. tuberculosis should be explored further.

PMID:34784363 | PMC:PMC8594824 | DOI:10.1371/journal.pone.0259181

RSC Med Chem. 2021 Aug 4;12(10):1623-1639. doi: 10.1039/d1md00200g. eCollection 2021 Oct 20.

ABSTRACT

The β-lactams are the most widely used antibacterial agents worldwide. These antibiotics, a group that includes the penicillins and cephalosporins, are covalent inhibitors that target bacterial penicillin-binding proteins and disrupt peptidoglycan synthesis. Bacteria can achieve resistance to β-lactams in several ways, including the production of serine β-lactamase enzymes. While β-lactams also covalently interact with serine β-lactamases, these enzymes are capable of deacylating this complex, treating the antibiotic as a substrate. In this tutorial-style review, we provide an overview of the β-lactam antibiotics, focusing on their covalent interactions with their target proteins and resistance mechanisms. We begin by describing the structurally diverse range of β-lactam antibiotics and β-lactamase inhibitors that are currently used as therapeutics. Then, we introduce the penicillin-binding proteins, describing their functions and structures, and highlighting their interactions with β-lactam antibiotics. We next describe the classes of serine β-lactamases, exploring some of the mechanisms by which they achieve the ability to degrade β-lactams. Finally, we introduce the l,d-transpeptidases, a group of bacterial enzymes involved in peptidoglycan synthesis which are also targeted by β-lactam antibiotics. Although resistance mechanisms are now prevalent for all antibiotics in this class, past successes in antibiotic development have at least delayed this onset of resistance. The β-lactams continue to be an essential tool for the treatment of infectious disease, and recent advances (e.g., β-lactamase inhibitor development) will continue to support their future use.

PMID:34778765 | PMC:PMC8528271 | DOI:10.1039/d1md00200g