Proteomics and microscopy tools for the study of antimicrobial resistance and germination mechanisms of bacterial spores

Abhyankar, Wishwas; Wen, Jiaming; Swarge, Bhagyashree N.; Tu, Zhiwei; Boer, Richard de; Smelt, J.P.P.M.; Koning, Leo J. de; Manders, Erik M. M.; Koster, Chris G. de; Brul, Stanley

doi:10.1016/j.fm.2018.03.006

Cited by 6 publications

(4 citation statements)

References 52 publications

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“…To this end, our research group previously combined proteomics with metabolomics analysis of clinical specimens, and further revealed some differently expressed genes, proteins, pathways as well as metabolites in the antibiotic resistance of ESBLs-EC, providing solid evidence for mechanism study. 16 In addition to proteomics and metabolomics as potent tools in the research of bacterial drug-resistance, 26–29 we applied whole-genome sequencing (WGS) on same clinical samples in this study to firstly identify DEGs. Based on this, we further performed RT-PCR to determine their relative expression levels.…”

Section: Discussionmentioning

confidence: 99%

GLO1 Contributes to the Drug Resistance of Escherichia coli Through Inducing PER Type of Extended-Spectrum β-Lactamases

Ma¹,

Lai²,

Zan³

et al. 2022

IDR

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Background Escherichia coli -associated antimicrobial resistance (AMR) issue so far needs urgent considerations. This study aims to screen the potent genes associated with extended-spectrum β-lactamases (ESBLs) in drug-resistant Escherichia coli and elucidate the specific drug-resistant mechanism. Methods Clinical ESBLs-EC samples were obtained based on the microbial identification, and the whole genome was sequenced. In combination with the significantly enriched pathways, several differently expressed genes were screened and verified by RT-PCR. Furthermore, through knocking out glyoxalase 1 (GLO1) gene and transfecting overexpressed plasmids, the potential relationship between GLO1 and ESBLs was then investigated. Lastly, the concentrations of β-lactamases in bacteria and supernatant from different groups were examined by enzyme-linked immunosorbent assay (ELISA). Results After successful isolation and identification of ESBLs-EC, the whole genome and eighteen differential metabolic pathways were analyzed to select differently expressed genes, including add, deoD, guaD, speG, GLO1, VNN1, etc. RT-PCR results showed that there were no differences in these genes between the standard bacteria and susceptible Escherichia coli . Remarkably, the relative levels of four genes including speG, Hdac10, GLO1 and Ppcdc were significantly increased in ESBLs-EC in comparison with susceptible strains, whereas other gene expression was decreased. Further experiments utilizing gene knockout and overexpression strains confirmed the role of GLO1. At last, a total of 10 subtypes of β-lactamases were studied using ELISA, including BES-, CTX-M1-, CTX-M2-, OXA1-, OXA2-, OXA10-, PER-, SHV-, TEM-, and VEB-ESBLs, and results demonstrated that GLO1 gene expression only affected PER-β-lactamases but had no effects on other β-lactamases. Conclusion SpeG, Hdac10, GLO1 and Ppcdc might be associated with the drug-resistant mechanism of Escherichia coli . Of note, this study firstly addressed the role of GLO1 in the drug resistance of ESBLs-EC, and this effect may be mediated by increasing PER-β-lactamases.

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Section: Discussionmentioning

confidence: 99%

GLO1 Contributes to the Drug Resistance of Escherichia coli Through Inducing PER Type of Extended-Spectrum β-Lactamases

Ma¹,

Lai²,

Zan³

et al. 2022

IDR

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“…Both traditional biochemical assays and omics require additional approaches for validation. For example, (Abhyankar et al, 2019) used proteomics to study bacterial spore integrity and germination physiology, showing a large variation in sporulation and germination kinetics which necessitated fluorescence imaging of individual spore germination, outgrowth and intracellular pH dynamics. Furthermore, pathogen mutational studies are incomplete without phenotypic characterization of individual pathogens in situ (Conrad et al, 2018).…”

Section: Limitations Of Traditional Methods For Studying Pathogensmentioning

confidence: 99%

AFM-Based Correlative Microscopy Illuminates Human Pathogens

Bhat

Price

Dahms

2021

Front. Cell. Infect. Microbiol.

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Microbes have an arsenal of virulence factors that contribute to their pathogenicity. A number of challenges remain to fully understand disease transmission, fitness landscape, antimicrobial resistance and host heterogeneity. A variety of tools have been used to address diverse aspects of pathogenicity, from molecular host-pathogen interactions to the mechanisms of disease acquisition and transmission. Current gaps in our knowledge include a more direct understanding of host-pathogen interactions, including signaling at interfaces, and direct phenotypic confirmation of pathogenicity. Correlative microscopy has been gaining traction to address the many challenges currently faced in biomedicine, in particular the combination of optical and atomic force microscopy (AFM). AFM, generates high-resolution surface topographical images, and quantifies mechanical properties at the pN scale under physiologically relevant conditions. When combined with optical microscopy, AFM probes pathogen surfaces and their physical and molecular interaction with host cells, while the various modes of optical microscopy view internal cellular responses of the pathogen and host. Here we review the most recent advances in our understanding of pathogens, recent applications of AFM to the field, how correlative AFM-optical microspectroscopy and microscopy have been used to illuminate pathogenicity and how these methods can reach their full potential for studying host-pathogen interactions.

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“…In view of B. subtilis having nearly 75% spore dry weight content, proteins play a very important role in spore resistance [28]. It is, therefore, meaningful to identify and characterize spore proteins in response to the synergistic effects of PT-LK treatment.…”

Section: Tmt Proteomic Analysis Of B Subtilis Spores In Response To P...mentioning

confidence: 99%

Proteomic Response of Bacillus subtilis Spores under High Pressure Combined with Moderate Temperature and Random Peptide Mixture LK Treatment

Pang

Cui

et al. 2022

Foods

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In this study, a method of Bacillus subtilis spore inactivation under high pressure (P, 200 MPa) combined with moderate temperature (T, 80 °C) and the addition of antimicrobial peptide LK (102 μg/mL) was investigated. Spores presented cortex hydrolysis and inner membrane (IM) damage with an 8.16 log reduction in response to treatment with PT-LK, as observed by phase-contrast and inverted fluorescence microscopy and flow cytometry (FCM) analysis. Furthermore, a tandem mass tag (TMT) quantitative proteomics approach was utilized because Liquid chromatography-tandem mass spectrometry (LC–MS/MS) analysis data were used. After treatment with PT-LK, 17,017 polypeptides and 3166 proteins were detected from B. subtilis spores. Among them, 78 proteins showed significant differences in abundance between the PT-LK-treated and control groups, with 49 proteins being upregulated and 29 proteins being downregulated in the PT-LK-treated group. Genetic information processing, metabolism, cellular process, and environmental information processing were the main mechanisms of PT-LK-mediated spore inactivation.

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Proteomics and microscopy tools for the study of antimicrobial resistance and germination mechanisms of bacterial spores

Cited by 6 publications

References 52 publications

GLO1 Contributes to the Drug Resistance of Escherichia coli Through Inducing PER Type of Extended-Spectrum β-Lactamases

GLO1 Contributes to the Drug Resistance of Escherichia coli Through Inducing PER Type of Extended-Spectrum β-Lactamases

AFM-Based Correlative Microscopy Illuminates Human Pathogens

Proteomic Response of Bacillus subtilis Spores under High Pressure Combined with Moderate Temperature and Random Peptide Mixture LK Treatment

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