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Tulin G, Méndez AA, Figueroa NR, Smith C, Folmer MP, Serra D, Wade JT, Checa SK, Soncini FC. Integration of BrfS into the biofilm-controlling cascade promotes sessile Salmonella growth at low temperatures. Biofilm 2025; 9:100254. [PMID: 39927094 PMCID: PMC11804604 DOI: 10.1016/j.bioflm.2025.100254] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/16/2024] [Revised: 01/14/2025] [Accepted: 01/15/2025] [Indexed: 02/11/2025] Open
Abstract
Biofilm formation is stimulated by different stress-related physiological and environmental conditions. In Salmonella and Escherichia coli, curli fibers and phosphoethanolamine-cellulose are the major extracellular components of biofilms. The production of both is under the control of CsgD, a transcriptional regulator whose expression is modulated by a number of factors responding to different signals. The atypical MerR-like regulator MlrA is key in the activation of csgD transcription in both Salmonella and E. coli. Recently, MlrB, a SPI-2-encoded MlrA-like regulator that counteracts MlrA by repressing csgD transcription and biofilm formation inside macrophages was identified. Here, we characterize STM1266, a Salmonella-specific MlrA-like regulator, recently renamed BrfS. In contrast to mlrA, brfS transcription increases in minimal growth media and at 20 °C, a temperature not commonly tested in laboratories. Under these conditions, as well as in salt-limited rich medium, deletion or overexpression of brfS affects extracellular matrix production. Using transcriptomics, we uncovered genes under BrfS control relevant for biofilm formation such as csgB and bapA. Transcriptional analysis of these genes in mutants lacking brfS, csgD or both, indicates that BrfS controls curli biosynthesis both in a CsgD-dependent and independent manner. By contrast, at low temperatures, bapA transcription depends only on BrfS, and neither deletion of csgD nor of mlrA modify its expression. Based on these results, we propose that BrfS contributes to Salmonella persistence in the environment, where the pathogen encounters low temperatures and nutrient limitation.
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Affiliation(s)
- Gonzalo Tulin
- Instituto de Biología Molecular y Celular de Rosario, Facultad de Ciencias Bioquímicas y Farmacéuticas, Universidad Nacional de Rosario, Consejo Nacional de Investigaciones Científicas y Técnicas, Rosario, Argentina
| | - Andrea A.E. Méndez
- Instituto de Biología Molecular y Celular de Rosario, Facultad de Ciencias Bioquímicas y Farmacéuticas, Universidad Nacional de Rosario, Consejo Nacional de Investigaciones Científicas y Técnicas, Rosario, Argentina
| | - Nicolás R. Figueroa
- Instituto de Biología Molecular y Celular de Rosario, Facultad de Ciencias Bioquímicas y Farmacéuticas, Universidad Nacional de Rosario, Consejo Nacional de Investigaciones Científicas y Técnicas, Rosario, Argentina
- Current position: Centro de Estudios Fotosintéticos y Bioquímicos, Consejo Nacional de Investigaciones Científicas y Técnicas, Rosario, Argentina
| | - Carol Smith
- Wadsworth Center, New York State Department of Health, Albany, NY, USA
| | - María P. Folmer
- Instituto de Biología Molecular y Celular de Rosario, Facultad de Ciencias Bioquímicas y Farmacéuticas, Universidad Nacional de Rosario, Consejo Nacional de Investigaciones Científicas y Técnicas, Rosario, Argentina
| | - Diego Serra
- Instituto de Biología Molecular y Celular de Rosario, Facultad de Ciencias Bioquímicas y Farmacéuticas, Universidad Nacional de Rosario, Consejo Nacional de Investigaciones Científicas y Técnicas, Rosario, Argentina
| | - Joseph T. Wade
- Wadsworth Center, New York State Department of Health, Albany, NY, USA
- Department of Biomedical Sciences, School of Public Health, University at Albany, Albany, NY, USA
- RNA Institute, University at Albany, SUNY, Albany, NY, USA
| | - Susana K. Checa
- Instituto de Biología Molecular y Celular de Rosario, Facultad de Ciencias Bioquímicas y Farmacéuticas, Universidad Nacional de Rosario, Consejo Nacional de Investigaciones Científicas y Técnicas, Rosario, Argentina
| | - Fernando C. Soncini
- Instituto de Biología Molecular y Celular de Rosario, Facultad de Ciencias Bioquímicas y Farmacéuticas, Universidad Nacional de Rosario, Consejo Nacional de Investigaciones Científicas y Técnicas, Rosario, Argentina
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2
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Fritts RK, Ebmeier CC, Copley SD. Transcriptomic and proteomic ramifications of segmental amplification in Escherichia coli. Proc Natl Acad Sci U S A 2025; 122:e2422424122. [PMID: 40372434 PMCID: PMC12107188 DOI: 10.1073/pnas.2422424122] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/29/2024] [Accepted: 04/17/2025] [Indexed: 05/16/2025] Open
Abstract
Gene amplification can drive adaptation by rapidly increasing the cellular dosage of critical gene products. Segmental amplifications often encompass large genomic regions surrounding the gene(s) under selection for higher dosage. Overexpression of coamplified neighboring genes imposes a substantial metabolic burden. While compensatory mutations can decrease inappropriate overexpression of coamplified genes, it takes time for such mutations to arise. The extent to which intrinsic regulatory mechanisms modulate expression of coamplified genes in the immediate aftermath of segmental amplification is largely unknown. To address the collateral effects of segmental amplification, we evolved replicate cultures of an Escherichia coli mutant under conditions that select for higher dosage of an inefficient enzyme whose weak activity limits growth rate. Segmental amplifications encompassing the gene encoding the weak-link enzyme arose in all populations. Amplified regions ranged in size (33 to 125 kb) and copy number (2 to ≥14 copies). We performed RNA-seq and label-free proteomics to quantify expression of amplified genes present at 2, 6, and 14 copies. mRNA expression generally scales with gene copy number, but protein expression scales less well with both gene copy number and mRNA expression. We characterize the molecular mechanisms underlying discrepancies between gene copy number and expression for several cases. We also show that segmental amplifications can have system-wide consequences by indirectly altering expression of nonamplified genes. Our findings indicate that the fitness benefit derived from segmental amplification depends on the combined effects of amplicon size, gene content, and copy number as well as collateral effects on nonamplified genes.
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Affiliation(s)
- Ryan K. Fritts
- Department of Molecular, Cellular and Developmental Biology, University of Colorado Boulder, Boulder80309, CO
- Cooperative Institute for Research in Environmental Sciences, University of Colorado Boulder, Boulder80309, CO
| | | | - Shelley D. Copley
- Department of Molecular, Cellular and Developmental Biology, University of Colorado Boulder, Boulder80309, CO
- Cooperative Institute for Research in Environmental Sciences, University of Colorado Boulder, Boulder80309, CO
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3
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Sharma A, Tayal S, Bhatnagar S. Analysis of stress response in multiple bacterial pathogens using a network biology approach. Sci Rep 2025; 15:15342. [PMID: 40316612 PMCID: PMC12048639 DOI: 10.1038/s41598-025-91269-5] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/26/2024] [Accepted: 02/19/2025] [Indexed: 05/04/2025] Open
Abstract
Stress response in bacterial pathogens promotes adaptation, virulence and antibiotic resistance. In this study, a network approach is applied to identify the common central mediators of stress response in five emerging opportunistic pathogens; Enterococcus faecium Aus0004, Staphylococcus aureus subsp. aureus USA300, Klebsiella pneumoniae MGH 78,578, Pseudomonas aeruginosa PAO1, and Mycobacterium tuberculosis H37Rv. A Protein-protein interaction network (PPIN) was constructed for each stressor using Cytoscape3.7.1 from the differentially expressed genes obtained from Gene expression omnibus datasets. A merged PPIN was constructed for each bacterium. Hub-bottlenecks in each network were the central stress response proteins and common pathways enriched in stress response were identified using KOBAS3.0. 31 hub-bottlenecks were common to each individual stress response, merged networks in all five pathogens and an independent cross stress (CS) response dataset of Escherichia coli. The 31 central nodes are in the RpoS mediated general stress regulon and also regulated by other stress response systems. Analysis of the 20 common metabolic pathways modulating stress response in all five bacteria showed that carbon metabolism pathway had the highest crosstalk with other pathways like amino acid biosynthesis and purine metabolism pathways. The central proteins identified can serve as targets for novel wide-spectrum antibiotics to overcome multidrug resistance.
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Affiliation(s)
- Anjali Sharma
- Computational and Structural Biology Laboratory, Department of Biological Sciences and Engineering, Netaji Subhas University of Technology, Dwarka, New Delhi, 110078, India
| | - Sonali Tayal
- Computational and Structural Biology Laboratory, Department of Biological Sciences and Engineering, Netaji Subhas University of Technology, Dwarka, New Delhi, 110078, India
| | - Sonika Bhatnagar
- Computational and Structural Biology Laboratory, Department of Biological Sciences and Engineering, Netaji Subhas University of Technology, Dwarka, New Delhi, 110078, India.
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4
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Guha M, Singh A, Butzin NC. Priestia megaterium cells are primed for surviving lethal doses of antibiotics and chemical stress. Commun Biol 2025; 8:206. [PMID: 39922941 PMCID: PMC11807137 DOI: 10.1038/s42003-025-07639-2] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/17/2024] [Accepted: 01/31/2025] [Indexed: 02/10/2025] Open
Abstract
Antibiotic resistant infections kill millions worldwide yearly. However, a key factor in recurrent infections is antibiotic persisters. Persisters are not inherently antibiotic-resistant but can withstand antibiotic exposure by entering a non-dividing state. This tolerance often results in prolonged antibiotic usage, increasing the likelihood of developing resistant strains. Here, we show the existence of "primed cells" in the Gram-positive bacterium Priestia megaterium, formerly known as Bacillus megaterium. These cells are pre-adapted to become persisters prior to lethal antibiotic stress. Remarkably, this prepared state is passed down through multiple generations via epigenetic memory, enhancing survival against antibiotics and other chemical stress. Previously, two distinct types of persisters were proposed: Type I and Type II, formed during stationary and log phases, respectively. However, our findings reveal that primed cells contribute to an increase in persisters during transition and stationary phases, with no evidence supporting distinct phenotypes between Type I and Type II persisters.
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Affiliation(s)
- Manisha Guha
- Department of Biology and Microbiology, South Dakota State University, Brookings, SD, USA
| | - Abhyudai Singh
- Electrical & Computer Engineering, University of Delaware, Newark, DE, USA
| | - Nicholas C Butzin
- Department of Biology and Microbiology, South Dakota State University, Brookings, SD, USA.
- Department of Chemistry and Biochemistry, South Dakota State University, Brookings, SD, USA.
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Akter S, Rahman MA, Ashrafudoulla M, Mahamud AGMSU, Chowdhury MAH, Ha SD. Mechanistic and bibliometric insights into RpoS-mediated biofilm regulation and its strategic role in food safety applications. Crit Rev Food Sci Nutr 2025:1-15. [PMID: 39879107 DOI: 10.1080/10408398.2025.2458755] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/31/2025]
Abstract
Biofilm, complex structures formed by microorganisms within an extracellular polymeric matrix, pose significant challenges in the sector by harboring dangerous pathogens and complicating decontamination, thereby increasing the risk of foodborne illnesses. This article provides a comprehensive review of the sigma factor, rpoS's role in biofilm development, specifically in gram-negative bacteria, and how the genetic, environmental, and regulatory elements influence rpoS activity with its critical role in bacterial stress responses. Our findings reveal that rpoS is a pivotal regulator of biofilm formation, enhancing bacterial survival in adverse conditions. Key factors affecting rpoS activity include oxidative and osmotic stress and nutrient availability. Understanding rpoS-mediated regulatory pathways is essential for developing targeted biofilm management strategies to improve food quality and safety. Furthermore, a bibliometric analysis highlights significant research trends and gaps in the literature, guiding future research directions. Future research should focus on detailed mechanistic studies of rpoS-mediated biofilm regulation, the development of specific rpoS inhibitors, and innovative approaches like biofilm-resistant surface coatings. This knowledge can lead to more effective contamination prevention and overall food safety enhancements.
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Affiliation(s)
- Shirin Akter
- Food Safety and Regulatory Science, Chung-Ang University, Anseong-Si, Republic of Korea
- GreenTech-Based Food Safety Research Group, Chung-Ang University, Anseong, Republic of Korea
- Department of Fisheries and Marine Bioscience, Bangabandhu Sheikh Mujibur Rahman Science and Technology University, Gopalganj, Bangladesh
| | - Md Ashikur Rahman
- Food Safety and Regulatory Science, Chung-Ang University, Anseong-Si, Republic of Korea
- GreenTech-Based Food Safety Research Group, Chung-Ang University, Anseong, Republic of Korea
- Bangladesh Fisheries Research Institute, Mymensingh, Bangladesh
| | - Md Ashrafudoulla
- Food Safety and Regulatory Science, Chung-Ang University, Anseong-Si, Republic of Korea
- National Institutes of Health, Bethesda, MD, USA
- Department of Food Science, Center for Food Safety, University of Arkansas System Division of Agriculture, Fayetteville, AR, USA
| | | | - Md Anamul Hasan Chowdhury
- Food Safety and Regulatory Science, Chung-Ang University, Anseong-Si, Republic of Korea
- GreenTech-Based Food Safety Research Group, Chung-Ang University, Anseong, Republic of Korea
| | - Sang-Do Ha
- Food Safety and Regulatory Science, Chung-Ang University, Anseong-Si, Republic of Korea
- GreenTech-Based Food Safety Research Group, Chung-Ang University, Anseong, Republic of Korea
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Wang M, González L, Saha S, Josić K, Mugler A, Bennett MR. Hyperballistic intercellular signaling through growth assisted positive feedback. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2024:2024.12.04.626899. [PMID: 39677820 PMCID: PMC11643040 DOI: 10.1101/2024.12.04.626899] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/17/2024]
Abstract
Intercellular signaling in bacteria is often mediated by small molecules secreted by cells. These small molecules disperse via diffusion which limits the speed and spatial extent of information transfer in spatially extended systems. Theory shows that a secondary signal and feedback circuits can speed up the flow of information and allow it to travel further. Here, we construct and test several synthetic circuits in Escherichia coli to determine to what extent a secondary signal and feedback can improve signal propagation in bacterial systems. We find that positive feedback-regulated secondary signals propagate further and faster than diffusion-limited signals. Additionally, the speed at which the signal propagates can accelerate in time, provided the density of the cells within the system increases. These findings provide the foundation for creating fast, long-range signal propagation circuits in spatially extended bacterial systems.
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Kaboudari A, Aliakbarlu J, Mehdizadeh T. Simultaneous Effects of Food-related Stresses on the Antibiotic Resistance of Foodborne Salmonella Serotypes. J Food Prot 2024; 87:100350. [PMID: 39168450 DOI: 10.1016/j.jfp.2024.100350] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/27/2023] [Revised: 07/21/2024] [Accepted: 08/14/2024] [Indexed: 08/23/2024]
Abstract
Antibiotic resistance has become one of the most critical issues in the field of public health in recent years. Exposure to food environment stresses may result in the development of antibiotic resistance in Salmonella. The present study aimed to investigate the simultaneous effects of food-related stresses (osmotic pressure, acid, heat, cold, and freezing stresses) on the antibiotic resistance changes in Salmonella Enteritidis and Salmonella Typhimurium. A factorial design with five factors at two levels was used to evaluate the main and interactive effects of stress factors on the antibiotic resistance of Salmonella serotypes. The changes in the antibiotic resistance of Salmonella serotypes were evaluated using the disc diffusion assay. The results showed that the different stresses had different effects on the antibiotic resistance of Salmonella serotypes. The freezing time and osmotic stresses had the most significant effects on the antibiotic resistance (P < 0.05). S. Enteritidis showed the slightest changes after exposure to stresses. The results also showed that a low level (24 h) of freezing time decreased the antibiotic resistance, but at a high level (96 h) increased it. The results emphasized that food processing and storage conditions should be considered as crucial factors in developing antibiotic resistance in Salmonella.
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Affiliation(s)
- Ata Kaboudari
- Department of Food Hygiene and Quality Control, Faculty of Veterinary Medicine, Urmia University, Urmia 5756151818, Iran
| | - Javad Aliakbarlu
- Department of Food Hygiene and Quality Control, Faculty of Veterinary Medicine, Urmia University, Urmia 5756151818, Iran.
| | - Tooraj Mehdizadeh
- Department of Food Hygiene and Quality Control, Faculty of Veterinary Medicine, Urmia University, Urmia 5756151818, Iran
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8
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Iqbal MZ, He P, He P, Wu Y, Munir S, He Y. The Response of Murine Gut Microbiome in the Presence of Altered rpoS Gene of Klebsiella pneumoniae. Int J Mol Sci 2024; 25:9222. [PMID: 39273171 PMCID: PMC11395600 DOI: 10.3390/ijms25179222] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/27/2024] [Revised: 08/05/2024] [Accepted: 08/08/2024] [Indexed: 09/15/2024] Open
Abstract
The murine model is invaluable for studying intricate interactions among gut microbes; hosts; and diseases. However; the impact of genetic variations in the murine microbiome; especially in disease contexts such as Klebsiella pneumoniae (Kp) infection; still needs to be explored. Kp; an opportunistic global pathogen; is becoming increasingly prevalent in regions like Asia; especially China. This study explored the role of the gut microbiota during Kp infection using mouse model; including wild-type and rpoS mutants of Kp138; KpC4; and KpE4 from human; maize; and ditch water; respectively. Under stress conditions; RpoS reconfigures global gene expression in bacteria; shifting the cells from active growth to survival mode. Our study examined notable differences in microbiome composition; finding that Lactobacillus and Klebsiella (particularly in WKp138) were the most abundant genera in mice guts at the genus level in all wild-type treated mice. In contrast; Firmicutes were predominant in the healthy control mice. Furthermore; Clostridium was the dominant genus in all mutants; mainly in ∆KpC4; and was absent in wild-type treated mice. Differential abundance analysis identified that these candidate taxa potentially influence disease progression and pathogen virulence. Functional prediction analysis showed that most bacterial groups were functionally involved in biosynthesis; precursor metabolites; degradation; energy generation; and metabolic cluster formation. These findings challenge the conventional understanding and highlight the need for nuanced interpretations in murine studies. Additionally; this study sheds light on microbiome-immune interactions in K. pneumoniae infection and proposes new potential therapeutic strategies.
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Affiliation(s)
| | | | | | | | - Shahzad Munir
- State Key Laboratory for Conservation and Utilization of Bioresources in Yunnan, Yunnan Agricultural University, Kunming 650201, China
| | - Yueqiu He
- State Key Laboratory for Conservation and Utilization of Bioresources in Yunnan, Yunnan Agricultural University, Kunming 650201, China
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9
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Maksimova Y, Eliseeva A, Maksimov A. Metabolic and Morphological Aspects of Adaptation of Alkaliphilic Bacillus aequororis 5-DB and Alkali-Tolerant Bacillus subtilis ATCC 6633 to Changes in pH and Mineralization. Int J Microbiol 2024; 2024:3087296. [PMID: 39081933 PMCID: PMC11288695 DOI: 10.1155/2024/3087296] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/10/2024] [Revised: 07/01/2024] [Accepted: 07/10/2024] [Indexed: 08/02/2024] Open
Abstract
The goal of the study is to evaluate metabolic and morphological changes of the facultative alkaliphile Bacillus aequororis 5-DB and the weakly alkali-resistant B. subtilis ATCC 6633 in a wide pH range and at different NaCl concentrations. The alkaliphile B. aequororis 5-DB is shown to have a broader general resistance to adverse factors (wide pH range, 50 g/L NaCl) than a weakly alkali-tolerant strain of the same genus. This alkaliphile is also shown to have a significantly greater resistance not only to high pH but also to low pH in comparison with B. subtilis ATCC 6633. The resistance of B. aequororis 5-DB to low pH was expressed in higher metabolic activity, maintenance of ΔpH, and no significant cell damage. The selected set of methods (reduction of resazurin to resorufin by cell dehydrogenases, bioluminescent method for determining ATP, AFM, and measurement of intracellular pH) allows us to adequately assess the ability of microbial cells to withstand harsh environmental factors. Nonspecific resistance of B. aequororis 5-DB was proven using a complex of selected methods. Tolerance to a wide range of pH and high salt concentrations may be useful for biotechnological applications of the strain.
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Affiliation(s)
- Yuliya Maksimova
- Laboratory of Molecular BiotechnologyInstitute of Ecology and Genetics of Microorganisms Ural Branch Russian Academy of Sciences, Golev Str., 13, Perm 614081, Russia
- Department of Microbiology and ImmunologyPerm State University, Bukirev Str., 15, Perm 614990, Russia
| | - Ann Eliseeva
- Laboratory of Molecular BiotechnologyInstitute of Ecology and Genetics of Microorganisms Ural Branch Russian Academy of Sciences, Golev Str., 13, Perm 614081, Russia
| | - Aleksandr Maksimov
- Laboratory of Molecular BiotechnologyInstitute of Ecology and Genetics of Microorganisms Ural Branch Russian Academy of Sciences, Golev Str., 13, Perm 614081, Russia
- Department of Microbiology and ImmunologyPerm State University, Bukirev Str., 15, Perm 614990, Russia
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Lv F, Zhan Y, Feng H, Sun W, Yin C, Han Y, Shao Y, Xue W, Jiang S, Ma Y, Hu H, Wei J, Yan Y, Lin M. Integrated Hfq-interacting RNAome and transcriptomic analysis reveals complex regulatory networks of nitrogen fixation in root-associated Pseudomonas stutzeri A1501. mSphere 2024; 9:e0076223. [PMID: 38747590 PMCID: PMC11332353 DOI: 10.1128/msphere.00762-23] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/17/2023] [Accepted: 04/10/2024] [Indexed: 06/26/2024] Open
Abstract
The RNA chaperone Hfq acts as a global regulator of numerous biological processes, such as carbon/nitrogen metabolism and environmental adaptation in plant-associated diazotrophs; however, its target RNAs and the mechanisms underlying nitrogen fixation remain largely unknown. Here, we used enhanced UV cross-linking immunoprecipitation coupled with high-throughput sequencing to identify hundreds of Hfq-binding RNAs probably involved in nitrogen fixation, carbon substrate utilization, biofilm formation, and other functions. Collectively, these processes endow strain A1501 with the requisite capabilities to thrive in the highly competitive rhizosphere. Our findings revealed a previously uncharted landscape of Hfq target genes. Notable among these is nifM, encoding an isomerase necessary for nitrogenase reductase solubility; amtB, encoding an ammonium transporter; oprB, encoding a carbohydrate porin; and cheZ, encoding a chemotaxis protein. Furthermore, we identified more than 100 genes of unknown function, which expands the potential direct regulatory targets of Hfq in diazotrophs. Our data showed that Hfq directly interacts with the mRNA of regulatory proteins (RsmA, AlgU, and NifA), regulatory ncRNA RsmY, and other potential targets, thus revealing the mechanistic links in nitrogen fixation and other metabolic pathways. IMPORTANCE Numerous experimental approaches often face challenges in distinguishing between direct and indirect effects of Hfq-mediated regulation. New technologies based on high-throughput sequencing are increasingly providing insight into the global regulation of Hfq in gene expression. Here, enhanced UV cross-linking immunoprecipitation coupled with high-throughput sequencing was employed to identify the Hfq-binding sites and potential targets in the root-associated Pseudomonas stutzeri A1501 and identify hundreds of novel Hfq-binding RNAs that are predicted to be involved in metabolism, environmental adaptation, and nitrogen fixation. In particular, we have shown Hfq interactions with various regulatory proteins' mRNA and their potential targets at the posttranscriptional level. This study not only enhances our understanding of Hfq regulation but, importantly, also provides a framework for addressing integrated regulatory network underlying root-associated nitrogen fixation.
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Affiliation(s)
- Fanyang Lv
- Biotechnology Research Institute/National Key Laboratory of Agricultural Microbiology, Chinese Academy of Agricultural Sciences, Beijing, China
| | - Yuhua Zhan
- Biotechnology Research Institute/National Key Laboratory of Agricultural Microbiology, Chinese Academy of Agricultural Sciences, Beijing, China
| | - Haichao Feng
- College of Agriculture, Henan University, Kaifeng, Henan, China
| | - Wenyue Sun
- Biotechnology Research Institute/National Key Laboratory of Agricultural Microbiology, Chinese Academy of Agricultural Sciences, Beijing, China
| | - Changyan Yin
- Biotechnology Research Institute/National Key Laboratory of Agricultural Microbiology, Chinese Academy of Agricultural Sciences, Beijing, China
| | - Yueyue Han
- Biotechnology Research Institute/National Key Laboratory of Agricultural Microbiology, Chinese Academy of Agricultural Sciences, Beijing, China
| | - Yahui Shao
- Biotechnology Research Institute/National Key Laboratory of Agricultural Microbiology, Chinese Academy of Agricultural Sciences, Beijing, China
| | - Wei Xue
- Biotechnology Research Institute/National Key Laboratory of Agricultural Microbiology, Chinese Academy of Agricultural Sciences, Beijing, China
| | - Shanshan Jiang
- Biotechnology Research Institute/National Key Laboratory of Agricultural Microbiology, Chinese Academy of Agricultural Sciences, Beijing, China
| | - Yiyuan Ma
- Biotechnology Research Institute/National Key Laboratory of Agricultural Microbiology, Chinese Academy of Agricultural Sciences, Beijing, China
| | - Haonan Hu
- Biotechnology Research Institute/National Key Laboratory of Agricultural Microbiology, Chinese Academy of Agricultural Sciences, Beijing, China
| | - Jinfeng Wei
- College of Agriculture, Henan University, Kaifeng, Henan, China
| | - Yongliang Yan
- Biotechnology Research Institute/National Key Laboratory of Agricultural Microbiology, Chinese Academy of Agricultural Sciences, Beijing, China
| | - Min Lin
- College of Agriculture, Henan University, Kaifeng, Henan, China
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11
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Li Z, Zhang M, Lei G, Lu X, Yang X, Kan B. A Single Base Change in the csgD Promoter Resulted in Enhanced Biofilm in Swine-Derived Salmonella Typhimurium. Microorganisms 2024; 12:1258. [PMID: 39065026 PMCID: PMC11278976 DOI: 10.3390/microorganisms12071258] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/08/2024] [Revised: 06/08/2024] [Accepted: 06/19/2024] [Indexed: 07/28/2024] Open
Abstract
Pathogenic Salmonella strains causing gastroenteritis typically can colonize and proliferate in the intestines of multiple host species. They retain the ability to form red dry and rough (rdar) biofilms, as seen in Salmonella enterica serovar Typhimurium. Conversely, Salmonella serovar like Typhi, which can cause systemic infections and exhibit host restriction, are rdar-negative. In this study, duck-derived strains and swine-derived strains of S. Typhimurium locate on independent phylogenetic clades and display relative genomic specificity. The duck isolates appear more closely related to human blood isolates and invasive non-typhoidal Salmonella (iNTS), whereas the swine isolates were more distinct. Phenotypically, compared to duck isolates, swine isolates exhibited enhanced biofilm formation that was unaffected by the temperature. The transcriptomic analysis revealed the upregulation of csgDEFG transcription as the direct cause. This upregulation may be mainly attributed to the enhanced promoter activity caused by the G-to-T substitution at position -44 of the csgD promoter. Swine isolates have created biofilm polymorphisms by altering a conserved base present in Salmonella Typhi, iNTS, and most Salmonella Typhimurium (such as duck isolates). This provides a genomic characteristics perspective for understanding Salmonella transmission cycles and evolution.
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Affiliation(s)
- Zhe Li
- National Key Laboratory of Intelligent Tracking and Forecasting for Infectious Diseases, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing 102206, China; (Z.L.)
| | - Mengke Zhang
- National Key Laboratory of Intelligent Tracking and Forecasting for Infectious Diseases, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing 102206, China; (Z.L.)
- School of Light Industry, Beijing Technology and Business University, Beijing 100048, China
| | - Gaopeng Lei
- Center for Disease Control and Prevention of Sichuan Province, Chengdu 610041, China
| | - Xin Lu
- National Key Laboratory of Intelligent Tracking and Forecasting for Infectious Diseases, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing 102206, China; (Z.L.)
| | - Xiaorong Yang
- Center for Disease Control and Prevention of Sichuan Province, Chengdu 610041, China
| | - Biao Kan
- National Key Laboratory of Intelligent Tracking and Forecasting for Infectious Diseases, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing 102206, China; (Z.L.)
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Guha M, Singh A, Butzin NC. Gram-positive bacteria are primed for surviving lethal doses of antibiotics and chemical stress. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2024:2024.05.28.596288. [PMID: 38895422 PMCID: PMC11185512 DOI: 10.1101/2024.05.28.596288] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 06/21/2024]
Abstract
Antibiotic resistance kills millions worldwide yearly. However, a major contributor to recurrent infections lies in a small fraction of bacterial cells, known as persisters. These cells are not inherently antibiotic-resistant, yet they lead to increased antibiotic usage, raising the risk of developing resistant progenies. In a bacterial population, individual cells exhibit considerable fluctuations in their gene expression levels despite being cultivated under identical, stable conditions. This variability in cell-to-cell characteristics (phenotypic diversity) within an isogenic population enables persister cells to withstand antibiotic exposure by entering a non-dividing state. We recently showed the existence of "primed cells" in E. coli. Primed cells are dividing cells prepared for antibiotic stress before encountering it and are more prone to form persisters. They also pass their "prepared state" down for several generations through epigenetic memory. Here, we show that primed cells are common among distant bacterial lineages, allowing for survival against antibiotics and other chemical stress, and form in different growth phases. They are also responsible for increased persister levels in transition and stationary phases compared to the log phase. We tested and showed that the Gram-positive bacterium Bacillus megaterium, evolutionarily very distant from E. coli, forms primed cells and has a transient epigenetic memory that is maintained for 7 generations or more. We showed this using ciprofloxacin and the non-antibiotic chemical stress fluoride. It is well established that persister levels are higher in the stationary phase than in the log phase, and B. megaterium persisters levels are nearly identical from the early to late-log phase but are ~2-fold and ~4-fold higher in the transition and stationary phase, respectively. It was previously proposed that there are two distinct types of persisters: Type II forms in the log phase, while Type I forms in the stationary phase. However, we show that primed cells lead to increased persisters in the transition and stationary phase and found no evidence of Type I or II persisters with distant phenotypes. Overall, we have provided substantial evidence of the importance of primed cells and their transitory epigenetic memories to surviving stress.
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Affiliation(s)
- Manisha Guha
- Department of Biology and Microbiology; South Dakota State University; Brookings, SD, 57006; USA
| | - Abhyudai Singh
- Electrical & Computer Engineering; University of Delaware; Newark, DE 19716; USA
| | - Nicholas C. Butzin
- Department of Biology and Microbiology; South Dakota State University; Brookings, SD, 57006; USA
- Department of Chemistry and Biochemistry; South Dakota State University; Brookings, SD, 57006; USA
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Cao J, Xue B, Yang S, Yang X, Zhang X, Qiu Z, Shen Z, Wang J. Chlorite and bromate alter the conjugative transfer of antibiotic resistance genes: Co-regulation of oxidative stress and energy supply. JOURNAL OF HAZARDOUS MATERIALS 2024; 471:134257. [PMID: 38636236 DOI: 10.1016/j.jhazmat.2024.134257] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 01/22/2024] [Revised: 04/06/2024] [Accepted: 04/08/2024] [Indexed: 04/20/2024]
Abstract
The widespread use of disinfectants during the global response to the 2019 coronavirus pandemic has increased the co-occurrence of disinfection byproducts (DBPs) and antibiotic resistance genes (ARGs). Although DBPs pose major threats to public health globally, there is limited knowledge regarding their biological effects on ARGs. This study aimed to investigate the effects of two inorganic DBPs (chlorite and bromate) on the conjugative transfer of RP4 plasmid among Escherichia coli strains at environmentally relevant concentrations. Interestingly, the frequency of conjugative transfer was initially inhibited when the exposure time to chlorite or bromate was less than 24 h. However, this inhibition transformed into promotion when the exposure time was extended to 36 h. Short exposures to chlorite or bromate were shown to impede the electron transport chain, resulting in an ATP shortage and subsequently inhibiting conjugative transfer. Consequently, this stimulates the overproduction of reactive oxygen species (ROS) and activation of the SOS response. Upon prolonged exposure, the resurgent energy supply promoted conjugative transfer. These findings offer novel and valuable insights into the effects of environmentally relevant concentrations of inorganic DBPs on the conjugative transfer of ARGs, thereby providing a theoretical basis for the management of DBPs.
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Affiliation(s)
- Jinrui Cao
- Tianjin Institute of Environmental and Operational Medicine, Tianjin 300050, China
| | - Bin Xue
- Tianjin Institute of Environmental and Operational Medicine, Tianjin 300050, China; Key Laboratory of Risk Assessment and Control for Environment and Food Safety, Tianjin 300050, China
| | - Shuran Yang
- Tianjin Institute of Environmental and Operational Medicine, Tianjin 300050, China
| | - Xiaobo Yang
- Tianjin Institute of Environmental and Operational Medicine, Tianjin 300050, China; Key Laboratory of Risk Assessment and Control for Environment and Food Safety, Tianjin 300050, China
| | - Xi Zhang
- Tianjin Institute of Environmental and Operational Medicine, Tianjin 300050, China; Key Laboratory of Risk Assessment and Control for Environment and Food Safety, Tianjin 300050, China
| | - Zhigang Qiu
- Tianjin Institute of Environmental and Operational Medicine, Tianjin 300050, China; Key Laboratory of Risk Assessment and Control for Environment and Food Safety, Tianjin 300050, China
| | - Zhiqiang Shen
- Tianjin Institute of Environmental and Operational Medicine, Tianjin 300050, China; Key Laboratory of Risk Assessment and Control for Environment and Food Safety, Tianjin 300050, China.
| | - Jingfeng Wang
- Tianjin Institute of Environmental and Operational Medicine, Tianjin 300050, China; Key Laboratory of Risk Assessment and Control for Environment and Food Safety, Tianjin 300050, China.
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14
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Kireina D, Parreira VR, Goodridge L, Farber JM. Survival and Expression of rpoS and grxB of Cronobacter sakazakii in Powdered Infant Formula Under Simulated Gastric Conditions of Newborns. J Food Prot 2024; 87:100269. [PMID: 38519033 DOI: 10.1016/j.jfp.2024.100269] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/31/2023] [Revised: 03/15/2024] [Accepted: 03/18/2024] [Indexed: 03/24/2024]
Abstract
Cronobacter sakazakii can cause severe illnesses in infants, predominantly in preterm newborns, with consumption of contaminated powdered infant formula (PIF) being the major vehicle of infection. Using a dynamic human gastrointestinal simulator called the SHIME, this study examined the effects of gastric acidity and gastric digestion time of newborns on the survival and expression of stress genes of C. sakazakii. Individual strains, inoculated at 7 log CFU/mL into reconstituted PIF, were exposed to gastric pH values of 4.00, 5.00 and 6.00 for 4 h with gradual acidification. The survival results showed that C. sakazakii grew in the stomach portion of the SHIME during a 4-h exposure to pH 4.00, 5.00 and 6.00 by 0.96-1.05, 1.02-1.28 and 1.11-1.73 log CFU/mL, respectively. The expression of two stress genes, rpoS and grxB, throughout gastric digestion was evaluated using reverse transcription qPCR. The upregulation of rpoS and grxB during the 4-h exposure to simulated gastric fluid at pH 4.00 showed that C. sakazakii strains may be experiencing the most stress in the pH 4.00 treatment. The gene expression results also suggest that C. sakazakii strains appeared to develop an acid adaptation response during the 4-h exposure that may facilitate their survival. Altogether, this study highlights that a combination of low gastric acidity, long digestion time in the presence of reconstituted PIF, created a favorable environment for the adaptation and survival of C. sakazakii in the simulation of a newborn's stomach. This study gives directions for future research to further advance our understanding of the behavior of C. sakazakii in the GI tract of newborns.
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Affiliation(s)
- Devita Kireina
- Canadian Research Institute for Food Safety, Department of Food Science, University of Guelph, Guelph, ON N1G 2W1, Canada
| | - Valeria R Parreira
- Canadian Research Institute for Food Safety, Department of Food Science, University of Guelph, Guelph, ON N1G 2W1, Canada.
| | - Lawrence Goodridge
- Canadian Research Institute for Food Safety, Department of Food Science, University of Guelph, Guelph, ON N1G 2W1, Canada
| | - Jeffrey M Farber
- Canadian Research Institute for Food Safety, Department of Food Science, University of Guelph, Guelph, ON N1G 2W1, Canada
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15
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Ibrahim N, Nesbitt D, Guo Q(T, Lin J, Svircev A, Wang Q, Weadge JT, Anany H. Improved Viability of Spray-Dried Pantoea agglomerans for Phage-Carrier Mediated Control of Fire Blight. Viruses 2024; 16:257. [PMID: 38400033 PMCID: PMC10893313 DOI: 10.3390/v16020257] [Citation(s) in RCA: 4] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/25/2023] [Revised: 01/26/2024] [Accepted: 01/30/2024] [Indexed: 02/25/2024] Open
Abstract
Fire blight, caused by Erwinia amylovora, is a devastating bacterial disease that threatens apple and pear production. It is mainly controlled by using antibiotics, such as streptomycin. Due to development of E. amylovora resistant strains and the excessive agricultural use of antibiotics, there is an increased awareness of the possibility of antibiotic resistance gene transfer to other microbes. Urgent development of biocontrol agents (BCAs) is needed that can be incorporated into integrated pest management programs as antibiotic alternatives. A novel phage-carrier system (PCS) that combines an antagonistic bacterium, Pantoea agglomerans, with its ability to act as a phage-carrier bacterium for Erwinia phages has been developed. The low viability of P. agglomerans cells following spray-drying (SD) has been a challenge for the industrial-scale production of this PCS. Here, an SD protocol was developed for P. agglomerans by modifying the growth medium and bacterial cell formulation using D(+)-trehalose and maltodextrin. The developed protocol is amenable to the industrial-scale production of the BCA/PCS. The P. agglomerans viability was greater than 90% after SD and had a shelf life at 4 °C of 4 months, and reconstituted cells showed a 3 log reduction in E. amylovora counts with a pear disc assay.
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Affiliation(s)
- Nassereldin Ibrahim
- Agriculture and Agri-Food Canada (AAFC), Guelph Research and Development Centre (GRDC), 93 Stone Rd W., Guelph, ON N1G 5C9, Canada; (N.I.); (Q.G.); (Q.W.)
- Department of Biology, Wilfrid Laurier University, Waterloo, ON N2L 3C5, Canada;
- Genetic Engineering and Biotechnology Research Institute, University of Sadat City, Sadat City 32897, Egypt
| | - Darlene Nesbitt
- Agriculture and Agri-Food Canada (AAFC), Vineland Station, ON L0R 2E0, Canada; (D.N.); (A.S.)
| | - Qian (Tracy) Guo
- Agriculture and Agri-Food Canada (AAFC), Guelph Research and Development Centre (GRDC), 93 Stone Rd W., Guelph, ON N1G 5C9, Canada; (N.I.); (Q.G.); (Q.W.)
| | - Janet Lin
- National Microbiology Laboratory, National Health Agency, 110 Stone Rd. W., Guelph, ON N1G 3W4, Canada;
| | - Antonet Svircev
- Agriculture and Agri-Food Canada (AAFC), Vineland Station, ON L0R 2E0, Canada; (D.N.); (A.S.)
| | - Qi Wang
- Agriculture and Agri-Food Canada (AAFC), Guelph Research and Development Centre (GRDC), 93 Stone Rd W., Guelph, ON N1G 5C9, Canada; (N.I.); (Q.G.); (Q.W.)
| | - Joel T. Weadge
- Department of Biology, Wilfrid Laurier University, Waterloo, ON N2L 3C5, Canada;
| | - Hany Anany
- Agriculture and Agri-Food Canada (AAFC), Guelph Research and Development Centre (GRDC), 93 Stone Rd W., Guelph, ON N1G 5C9, Canada; (N.I.); (Q.G.); (Q.W.)
- Food Science Department, University of Guelph, Guelph, ON N1G 2W1, Canada
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16
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Jeong GJ, Khan F, Tabassum N, Kim YM. Cellular and physiological roles of sigma factors in Vibrio spp.: A comprehensive review. Int J Biol Macromol 2024; 254:127833. [PMID: 37918595 DOI: 10.1016/j.ijbiomac.2023.127833] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/23/2023] [Revised: 10/30/2023] [Accepted: 10/30/2023] [Indexed: 11/04/2023]
Abstract
Vibrio species are motile gram-negative bacteria commonly found in aquatic environments. Vibrio species include pathogenic as well as non-pathogenic strains. Pathogenic Vibrio species have been reported in invertebrates and humans, whereas non-pathogenic strains are involved in symbiotic relationships with their eukaryotic hosts. These bacteria are also able to adapt to fluctuations in temperature, salinity, and pH, in addition to oxidative stress, and osmotic pressure in aquatic ecosystems. Moreover, they have also developed protective mechanisms against the immune systems of their hosts. Vibrio species accomplish adaptation to changing environments outside or inside the host by altering their gene expression profiles. To this end, several sigma factors specifically regulate gene expression, particularly under stressful environmental conditions. Moreover, other sigma factors are associated with biofilm formation and virulence as well. This review discusses different types of sigma and anti-sigma factors of Vibrio species involved in virulence and regulation of gene expression upon changes in environmental conditions. The evolutionary relationships between sigma factors with various physiological roles in Vibrio species are also discussed extensively.
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Affiliation(s)
- Geum-Jae Jeong
- Department of Food Science and Technology, Pukyong National University, Busan 48513, Republic of Korea
| | - Fazlurrahman Khan
- Institute of Fisheries Sciences, Pukyong National University, Busan 48513, Republic of Korea; Marine Integrated Biomedical Technology Center, The National Key Research Institutes in Universities, Pukyong National University, Busan 48513, Republic of Korea; Research Center for Marine Integrated Bionics Technology, Pukyong National University, Busan 48513, Republic of Korea.
| | - Nazia Tabassum
- Marine Integrated Biomedical Technology Center, The National Key Research Institutes in Universities, Pukyong National University, Busan 48513, Republic of Korea; Research Center for Marine Integrated Bionics Technology, Pukyong National University, Busan 48513, Republic of Korea
| | - Young-Mog Kim
- Department of Food Science and Technology, Pukyong National University, Busan 48513, Republic of Korea; Marine Integrated Biomedical Technology Center, The National Key Research Institutes in Universities, Pukyong National University, Busan 48513, Republic of Korea; Research Center for Marine Integrated Bionics Technology, Pukyong National University, Busan 48513, Republic of Korea.
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17
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Cancino-Diaz ME, Guerrero-Barajas C, Betanzos-Cabrera G, Cancino-Diaz JC. Nucleotides as Bacterial Second Messengers. Molecules 2023; 28:7996. [PMID: 38138485 PMCID: PMC10745434 DOI: 10.3390/molecules28247996] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/26/2023] [Revised: 11/21/2023] [Accepted: 11/23/2023] [Indexed: 12/24/2023] Open
Abstract
In addition to comprising monomers of nucleic acids, nucleotides have signaling functions and act as second messengers in both prokaryotic and eukaryotic cells. The most common example is cyclic AMP (cAMP). Nucleotide signaling is a focus of great interest in bacteria. Cyclic di-AMP (c-di-AMP), cAMP, and cyclic di-GMP (c-di-GMP) participate in biological events such as bacterial growth, biofilm formation, sporulation, cell differentiation, motility, and virulence. Moreover, the cyclic-di-nucleotides (c-di-nucleotides) produced in pathogenic intracellular bacteria can affect eukaryotic host cells to allow for infection. On the other hand, non-cyclic nucleotide molecules pppGpp and ppGpp are alarmones involved in regulating the bacterial response to nutritional stress; they are also considered second messengers. These second messengers can potentially be used as therapeutic agents because of their immunological functions on eukaryotic cells. In this review, the role of c-di-nucleotides and cAMP as second messengers in different bacterial processes is addressed.
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Affiliation(s)
- Mario E. Cancino-Diaz
- Departamentos Microbiología and Inmunología, Escuela Nacional de Ciencias Biológicas, Instituto Politécnico Nacional, Manuel Carpio, Plutarco Elías Calles, Miguel Hidalgo, Ciudad de México 11350, Mexico
| | - Claudia Guerrero-Barajas
- Departamento de Bioprocesos, Unidad Profesional Interdisciplinaria de Biotecnología, Instituto Politécnico Nacional, Av. Acueducto, La Laguna Ticoman, Gustavo A. Madero, Ciudad de México 07340, Mexico;
| | - Gabriel Betanzos-Cabrera
- Área Académica de Nutrición y Medicina, Instituto de Ciencias de la Salud, Universidad Autónoma del Estado de Hidalgo, Carretera Pachuca-Actopan Camino a Tilcuautla s/n, Pueblo San Juan Tilcuautla, Pachuca Hidalgo 42160, Mexico;
| | - Juan C. Cancino-Diaz
- Departamentos Microbiología and Inmunología, Escuela Nacional de Ciencias Biológicas, Instituto Politécnico Nacional, Manuel Carpio, Plutarco Elías Calles, Miguel Hidalgo, Ciudad de México 11350, Mexico
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18
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Thani AB. DNA supercoiling and regulation of intrinsic β-lactamase in pathogenic Escherichia coli. Arch Microbiol 2023; 205:385. [PMID: 37980630 DOI: 10.1007/s00203-023-03716-4] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/03/2023] [Revised: 10/18/2023] [Accepted: 10/19/2023] [Indexed: 11/21/2023]
Abstract
This review addresses the involvement of DNA supercoiling in the development of virulence and antibiotic profiles for uropathogenic Escherichia coli and the emergence of new pathotypes such as strain ST131 (serotype O25:H4). The mechanism suggests a role for topoisomerase enzymes and associated mutations in altering the chromosomal supercoiling state and introducing the required DNA twists for expression of intrinsic β-lactamase by ampC and certain virulence factors. In Escherichia coli, constitutive hyperexpression of intrinsic ampC is associated with specific mutations in the promoter and attenuator regions. However, many reports have documented the involvement of slow growth interventions in the expression of intrinsic resistance determinants. There is evidence that a stationary phase transcriptional switch protein, "BolA," is involved in the expression of the intrinsic ampC gene under starvation conditions. The process involves changes in the activity of the enzyme "gyrase," which leads to a change in the chromosomal DNA topology. Consequently, the DNA is relaxed, and the expression of the bolA gene is upregulated. The evolution of the extraintestinal pathogenic E. coli strain ST131 has demonstrated successful adaptability to various stress conditions and conferred compensatory mutations that endowed the microbe with resistance to fluoroquinolones and β-lactams. The results of this study provided new insights into the evidence for the influence of DNA topology in the expression of virulence genes and various determinants of antibiotic resistance (e.g., the intrinsic ampC gene) in Escherichia coli pathotypes.
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Affiliation(s)
- Ali Bin Thani
- Department of Biology, College of Science, University of Bahrain, Zallaq, Kingdom of Bahrain.
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19
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Zhang J, Zheng M, Tang Z, Zhong S, Bu T, Li Q. The Regulatory Functions of the Multiple Alternative Sigma Factors RpoE, RpoHI, and RpoHII Depend on the Growth Phase in Rhodobacter sphaeroides. Microorganisms 2023; 11:2678. [PMID: 38004690 PMCID: PMC10673084 DOI: 10.3390/microorganisms11112678] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/10/2023] [Revised: 10/26/2023] [Accepted: 10/27/2023] [Indexed: 11/26/2023] Open
Abstract
Bacterial growth, under laboratory conditions or in a natural environment, goes through different growth phases. Some gene expressions are regulated with respect to the growth phase, which allows bacteria to adapt to changing conditions. Among them, many gene transcriptions are controlled by RpoHI or RpoHII in Rhodobacter sphaeroides. In a previous study, it was proven that the alternative sigma factors, RpoE, RpoHI, and RpoHII, are the major regulators of oxidative stress. Moreover, the growth of bacteria reached a stationary phase, and following the outgrowth, rpoE, rpoHI, and rpoHII mRNAs increased with respect to the growth phase. In this study, we demonstrated the regulatory function of alternative sigma factors in the rsp_0557 gene. The gene rsp_0557 is expressed with respect to the growth phase and belongs to the RpoHI/RpoHII regulons. Reporter assays showed that the antisigma factor ChrR turns on or over the RpoE activity to regulate rsp_0557 expression across the growth phase. In the exponential phase, RpoHII and sRNA Pos19 regulate the expression of rsp_0557 to an appropriate level under RpoE control. In the stationary phase, RpoHI and Pos19 stabilize the transcription of rsp_0557 at a high level. During outgrowth, RpoHI negatively regulates the transcription of rsp_0557. Taken together, our data indicate that these regulators are recruited by cells to adapt to or survive under different conditions throughout the growth phase. However, they still did not display all of the regulators involved in growth phase-dependent regulation. More research is still needed to learn more about the interaction between the regulators and the process of adapting to changed growth conditions and environments.
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Affiliation(s)
| | | | | | | | | | - Qingfeng Li
- College of Life Sciences, Sichuan Agricultural University, Ya’an 625014, China; (J.Z.); (M.Z.); (Z.T.); (S.Z.); (T.B.)
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20
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Okada K, Roobthaisong A, Hamada S. Flagella-related gene mutations in Vibrio cholerae during extended cultivation in nutrient-limited media impair cell motility and prolong culturability. mSystems 2023; 8:e0010923. [PMID: 37642466 PMCID: PMC10654082 DOI: 10.1128/msystems.00109-23] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/06/2023] [Accepted: 04/19/2023] [Indexed: 08/31/2023] Open
Abstract
IMPORTANCE Vibrio cholerae undergoes a transition to a viable but non-culturable (VNC) state when subjected to various environmental stresses. We showed here that flagellar motility was involved in the development of the VNC state of V. cholerae. In this study, motility-defective isolates with mutations in various flagella-related genes, but not motile isolates, were predominantly obtained under the stress of long-term batch culture. Other genomic regions were highly conserved, suggesting that the mutations were selective. During the stationary phase of long-term culture, V. cholerae isolates with mutations in the acetate kinase and flagella-related genes were predominant. This study suggests that genes involved in specific functions in V. cholerae undergo mutations under certain environmental conditions.
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Affiliation(s)
- Kazuhisa Okada
- Research Institute for Microbial Diseases, Osaka University, Suita, Osaka, Japan
- Thailand-Japan Research Collaboration Center on Emerging and Re-emerging Infections, National Institute of Health, Nonthaburi, Thailand
| | - Amonrattana Roobthaisong
- Thailand-Japan Research Collaboration Center on Emerging and Re-emerging Infections, National Institute of Health, Nonthaburi, Thailand
| | - Shigeyuki Hamada
- Research Institute for Microbial Diseases, Osaka University, Suita, Osaka, Japan
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21
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Chen Y, Quirk NF, Tan S. Shining a light on bacterial environmental cue integration and its relation to metabolism. Mol Microbiol 2023; 120:71-74. [PMID: 37433048 PMCID: PMC10348474 DOI: 10.1111/mmi.15065] [Citation(s) in RCA: 2] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/04/2023] [Revised: 04/05/2023] [Accepted: 04/06/2023] [Indexed: 07/13/2023]
Abstract
The ability of a bacterium to successfully colonize its host is dependent on proper adaptation to its local environment. Environmental cues are diverse in nature, ranging from ions to bacterial-produced signals, and to host immune responses that can also be exploited by the bacteria as cues. Simultaneously, bacterial metabolism must be matched to the carbon and nitrogen sources available at a given time and location. While initial characterization of a bacterium's response to a given environmental cue or its ability to utilize a particular carbon/nitrogen source requires study of the signal in question in isolation, actual infection poses a situation where multiple signals are present concurrently. This perspective focuses on the untapped potential in uncovering and understanding how bacteria integrate their response to multiple concurrent environmental cues, and in elucidating the possible intrinsic coordination of bacterial environmental response with its metabolism.
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Affiliation(s)
- Yue Chen
- Department of Molecular Biology and Microbiology, Tufts University School of Medicine, Boston, Massachusetts, USA
| | - Natalia F. Quirk
- Department of Molecular Biology and Microbiology, Tufts University School of Medicine, Boston, Massachusetts, USA
- Graduate Program in Molecular Microbiology, Graduate School of Biomedical Sciences, Tufts University, Boston, Massachusetts, USA
| | - Shumin Tan
- Department of Molecular Biology and Microbiology, Tufts University School of Medicine, Boston, Massachusetts, USA
- Graduate Program in Molecular Microbiology, Graduate School of Biomedical Sciences, Tufts University, Boston, Massachusetts, USA
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22
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Wilhelm RC, Muñoz-Ucros J, Weikl F, Pritsch K, Goebel M, Buckley DH, Bauerle TL. The effects of mixed-species root zones on the resistance of soil bacteria and fungi to long-term experimental and natural reductions in soil moisture. THE SCIENCE OF THE TOTAL ENVIRONMENT 2023; 873:162266. [PMID: 36822431 DOI: 10.1016/j.scitotenv.2023.162266] [Citation(s) in RCA: 2] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 10/20/2022] [Revised: 02/11/2023] [Accepted: 02/12/2023] [Indexed: 06/18/2023]
Abstract
Mixed forest stands tend to be more resistant to drought than species-specific stands partially due to complementarity in root ecology and physiology. We asked whether complementary differences in the drought resistance of soil microbiomes might contribute to this phenomenon. We experimented on the effects of reduced soil moisture on bacterial and fungal community composition in species-specific (single species) and mixed-species root zones of Norway spruce and European beech forests in a 5-year-old throughfall-exclusion experiment and across seasonal (spring-summer-fall) and latitudinal moisture gradients. Bacteria were most responsive to changes in soil moisture, especially members of Rhizobiales, while fungi were largely unaffected, including ectomycorrhizal fungi (EMF). Community resistance was higher in spruce relative to beech root zones, corresponding with the proportions of drought-favored (more in spruce) and drought-sensitive bacterial taxa (more in beech). The spruce soil microbiome also exhibited greater resistance to seasonal changes between spring (wettest) and fall (driest). Mixed-species root zones contained a hybrid of beech- and spruce-associated microbiomes. Several bacterial populations exhibited either enhanced resistance or greater susceptibility to drought in mixed root zones. Overall, patterns in the relative abundances of soil bacteria closely tracked moisture in seasonal and latitudinal precipitation gradients and were more predictive of soil water content than other environmental variables. We conclude that complementary differences in the drought resistance of soil microbiomes can occur and the likeliest form of complementarity in mixed-root zones coincides with the enrichment of drought-tolerant bacteria associated with spruce and the sustenance of EMF by beech.
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Affiliation(s)
- Roland C Wilhelm
- School of Integrative Plant Science, Cornell University, Ithaca, NY, USA; Agronomy Department, Lilly Hall of Life Sciences, Purdue University, West Lafayette, IN, 47904, USA
| | - Juana Muñoz-Ucros
- School of Integrative Plant Science, Cornell University, Ithaca, NY, USA
| | - Fabian Weikl
- Institute of Biochemical Plant Pathology, Helmholtz Zentrum München, Neuherberg, Germany; Technical University of Munich, Professorship of Land Surface Atmosphere Interactions, Freising, Germany
| | - Karin Pritsch
- Institute of Biochemical Plant Pathology, Helmholtz Zentrum München, Neuherberg, Germany
| | - Marc Goebel
- Department of Natural Resources and the Environment, Cornell University, Ithaca, NY, USA
| | - Daniel H Buckley
- School of Integrative Plant Science, Cornell University, Ithaca, NY, USA
| | - Taryn L Bauerle
- School of Integrative Plant Science, Cornell University, Ithaca, NY, USA.
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Wei Y, Dong B, Wu X, Zhao M, Wang D, Li N, Zhang Q, Zhang L, Zhou H. RpoZ regulates 2,4-DAPG production and quorum sensing system in Pseudomonas fluorescens 2P24. Front Microbiol 2023; 14:1160913. [PMID: 37250031 PMCID: PMC10213339 DOI: 10.3389/fmicb.2023.1160913] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/07/2023] [Accepted: 04/26/2023] [Indexed: 05/31/2023] Open
Abstract
Introduction Pseudomonas fluorescens 2P24 was isolated from soil of natural decay associated with wheat take-all and it can effectively control soil-borne diseases caused by a variety of plant pathogens. 2,4-diacetylphloroglucinol (2,4-DAPG), is produced by P. fluorescens 2P24 and plays an important role in the prevention and control of plant diseases. To understand the resistant mechanism, in this study, we conducted experiments to explore the regulation role of rpoZ in the synthesis of the antibiotic 2,4-DAPG and regulation of QS system. Methods A random mini-Tn5 mutagenesis procedure was used to screen regulators for phlA transcription in stain PM901, which containing a phlA∷lacZ transcriptional fusion reporter plasmid. We identified 12 insertion mutants could significantly change phlA gene expression. By analyzing the amino acid sequences of the interrupted gene, we obtained a mutant strain Aa4-29 destroyed the rpoZ gene, which encodes the omiga subunit. We constructed the plasmid of rpoZ mutant (pBBR-△rpoZ) transformed into competent cells of P. fluorescens 2P24 by electro-transformation assay. The strains of P. fluorescens 2P24/pBBR, 2P24-△rpoZ/pBBR, 2P24-△rpoZ/pBBR-rpoZ were used to evaluate the regulation role of rpoZ in 2,4-DAPG production and quorum sensing system. Results According to β-galactosidase activity, we found that rpoZ positively regulated the expression of phlA (a synthesis gene of 2,4-DAPG) and PcoI (a synthesis gene of PcoI/PcoR QS signal system) at the transcriptional level. The production of 2,4-DAPG antibiotic and signal molecule AHL was influenced by rpoZ. Further, rpoZ was involved in regulating rsmA expression. RpoZ also has a certain regulatory effect on rpoS transcription, but no effect on the transcription of phlF, emhABC and emhR. According to the biocontrol assay, P. fluorescens 2P24 strains with rpoZ showed obvious antagonism ability against the Rhizoctonia solani in cotton, while the mutant strain of rpoZ lost the biocontrol effect. RpoZ had a significant effect on the swimming and biofilm formation in P. fluorescens 2P24. Conclusion Our data showed that rpoZ was an important regulator of QS system, 2,4-DAPG in P. fluorescens 2P24. This may imply that P. fluorescens 2P24 has evolved different regulatory features to adapt to different environmental threats.
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Affiliation(s)
- Yarui Wei
- College of Horticulture and Plant Protection, Inner Mongolia Agricultural University, Hohhot, Inner Mongolia, China
| | - Baozhu Dong
- College of Horticulture and Plant Protection, Inner Mongolia Agricultural University, Hohhot, Inner Mongolia, China
| | - Xiaogang Wu
- College of Agriculture, Guangxi University, Nanning, China
| | - Mingmin Zhao
- College of Horticulture and Plant Protection, Inner Mongolia Agricultural University, Hohhot, Inner Mongolia, China
| | - Dong Wang
- College of Horticulture and Plant Protection, Inner Mongolia Agricultural University, Hohhot, Inner Mongolia, China
| | - Na Li
- Erdos Agricultural and Animal Husbandry Technology Promotion Center, Erdos, Inner Mongolia, China
| | - Qian Zhang
- Bayannaoer Agriculture and Animal Husbandry Technology Promotion Center, Bayannaoer, Inner Mongolia, China
| | - Liqun Zhang
- Department of Plant Pathology, China Agricultural University, Beijing, China
| | - Hongyou Zhou
- College of Horticulture and Plant Protection, Inner Mongolia Agricultural University, Hohhot, Inner Mongolia, China
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Wintenberg M, Manglass L, Martinez NE, Blenner M. Global Transcriptional Response of Escherichia coli Exposed In Situ to Different Low-Dose Ionizing Radiation Sources. mSystems 2023; 8:e0071822. [PMID: 36779725 PMCID: PMC10134817 DOI: 10.1128/msystems.00718-22] [Citation(s) in RCA: 4] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/01/2022] [Accepted: 01/11/2023] [Indexed: 02/14/2023] Open
Abstract
Characterization of biological and chemical responses to ionizing radiation by various organisms is essential for potential applications in bioremediation, alternative modes of detecting nuclear material, and national security. Escherichia coli DH10β is an optimal system to study the microbial response to low-dose ionizing radiation at the transcriptional level because it is a well-characterized model bacterium and its responses to other environmental stressors, including those to higher radiation doses, have been elucidated in prior studies. In this study, RNA sequencing with downstream transcriptomic analysis (RNA-seq) was employed to characterize the global transcriptional response of stationary-phase E. coli subjected to 239Pu, 3H (tritium), and 55Fe, at an approximate absorbed dose rate of 10 mGy day-1 for 1 day and 15 days. Differential expression analysis identified significant changes in gene expression of E. coli for both short- and long-term exposures. Radionuclide source exposure induced differential expression in E. coli of genes involved in biosynthesis pathways of nuclear envelope components, amino acids, and siderophores, transport systems such as ABC transporters and type II secretion proteins, and initiation of stress response and regulatory systems of temperature stress, the RpoS regulon, and oxidative stress. These findings provide a basic understanding of the relationship between low-dose exposure and biological effect of a model bacterium that is critical for applications in alternative nuclear material detection and bioremediation. IMPORTANCE Escherichia coli strain DH10β, a well-characterized model bacterium, was subjected to short-term (1-day) and long-term (15-day) exposures to three different in situ radiation sources comprised of radionuclides relevant to nuclear activities to induce a measurable and identifiable genetic response. We found E. coli had both common and unique responses to the three exposures studied, suggesting both dose rate- and radionuclide-specific effects. This study is the first to provide insights into the transcriptional response of a microorganism in short- and long-term exposure to continuous low-dose ionizing radiation with multiple in situ radionuclide sources and the first to examine microbial transcriptional response in stationary phase. Moreover, this work provides a basis for the development of biosensors and informing more robust dose-response relationships to support ecological risk assessment.
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Affiliation(s)
- Molly Wintenberg
- Department of Chemical and Biomolecular Engineering, Clemson University, Clemson, South Carolina, USA
| | - Lisa Manglass
- Department of Environmental Engineering and Earth Sciences, Clemson University, Clemson, South Carolina, USA
- Department of Physics and Engineering, Francis Marion University, Florence, South Carolina, USA
| | - Nicole E. Martinez
- Department of Environmental Engineering and Earth Sciences, Clemson University, Clemson, South Carolina, USA
| | - Mark Blenner
- Department of Chemical and Biomolecular Engineering, Clemson University, Clemson, South Carolina, USA
- Department of Chemical and Biomolecular Engineering, University of Delaware, Newark, Delaware, USA
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Yang H, Song H, Zhang J, Li W, Han Q, Zhang W. Proteomic analysis reveals the adaptation of Vibrio splendidus to an iron deprivation condition. Appl Microbiol Biotechnol 2023; 107:2533-2546. [PMID: 36922441 DOI: 10.1007/s00253-023-12460-0] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/13/2023] [Revised: 02/22/2023] [Accepted: 02/24/2023] [Indexed: 03/18/2023]
Abstract
Vibrio splendidus is a ubiquitous Gram-negative marine bacterium that causes diseases within a wide range of marine cultured animals. Since iron deprivation is the frequent situation that the bacteria usually encounter, we aimed to explore the effect of iron deprivation on the proteomic profile of V. splendidus in the present study. There were 425 differentially expressed proteins (DEPs) responded to the iron deprivation condition. When the cells were grown under iron deprivation condition, the oxidation‒reduction processes, single-organism metabolic processes, the catalytic activity, and binding activity were downregulated, while the transport process, membrane cell component, and ion binding activity were upregulated, apart from the iron uptake processes. Kyoto Encyclopedia of Genes and Genomes analysis showed that various metabolism pathways, biosynthesis pathways, energy generation pathways of tricarboxylic acid cycle, and oxidative phosphorylation were downregulated, while various degradation pathways and several special metabolism pathways were upregulated. The proteomic profiles of cells at a OD600 ≈ 0.4 grown under iron deprivation condition showed high similarity to that of the cells at a OD600 ≈ 0.8 grown without iron chelator 2,2'-bipyridine. Correspondingly, the protease activity, the activity of autoinducer 2 (AI-2), and indole content separately catalyzed by LuxS and TnaA, were measured to verify the proteomic data. Our present study gives basic information on the global protein profiles of V. splendidus grown under iron deprivation condition and suggests that the iron deprivation condition cause the cell growth enter a state of higher cell density earlier. KEY POINTS: • Adaptation of V. splendidus to iron deprivation was explored by proteomic analysis. • GO and KEGG of DEPs under different iron levels or cell densities were determined. • Iron deprivation caused the cell enter a state of higher cell density earlier.
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Affiliation(s)
- Huirong Yang
- Key Laboratory of Aquacultural Biotechnology Ministry of Education, Ningbo University, Zhejiang Province, Ningbo, 315832, People's Republic of China
- Key Laboratory of Marine Biotechnology of Zhejiang Province, School of Marine Sciences, Ningbo University, Beilun District, 169 Qixingnan RoadZhejiang Province, Ningbo, 315832, People's Republic of China
| | - Huimin Song
- Key Laboratory of Aquacultural Biotechnology Ministry of Education, Ningbo University, Zhejiang Province, Ningbo, 315832, People's Republic of China
- Key Laboratory of Marine Biotechnology of Zhejiang Province, School of Marine Sciences, Ningbo University, Beilun District, 169 Qixingnan RoadZhejiang Province, Ningbo, 315832, People's Republic of China
| | - Jinxia Zhang
- Key Laboratory of Aquacultural Biotechnology Ministry of Education, Ningbo University, Zhejiang Province, Ningbo, 315832, People's Republic of China
- Key Laboratory of Marine Biotechnology of Zhejiang Province, School of Marine Sciences, Ningbo University, Beilun District, 169 Qixingnan RoadZhejiang Province, Ningbo, 315832, People's Republic of China
| | - Weisheng Li
- Key Laboratory of Aquacultural Biotechnology Ministry of Education, Ningbo University, Zhejiang Province, Ningbo, 315832, People's Republic of China
- Key Laboratory of Marine Biotechnology of Zhejiang Province, School of Marine Sciences, Ningbo University, Beilun District, 169 Qixingnan RoadZhejiang Province, Ningbo, 315832, People's Republic of China
| | - Qingxi Han
- Key Laboratory of Aquacultural Biotechnology Ministry of Education, Ningbo University, Zhejiang Province, Ningbo, 315832, People's Republic of China
- Key Laboratory of Marine Biotechnology of Zhejiang Province, School of Marine Sciences, Ningbo University, Beilun District, 169 Qixingnan RoadZhejiang Province, Ningbo, 315832, People's Republic of China
| | - Weiwei Zhang
- Key Laboratory of Aquacultural Biotechnology Ministry of Education, Ningbo University, Zhejiang Province, Ningbo, 315832, People's Republic of China.
- Key Laboratory of Marine Biotechnology of Zhejiang Province, School of Marine Sciences, Ningbo University, Beilun District, 169 Qixingnan RoadZhejiang Province, Ningbo, 315832, People's Republic of China.
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Sousa FM, Fernandes B, Pereira MM. The protein family of pyruvate:quinone oxidoreductases: Amino acid sequence conservation and taxonomic distribution. BIOCHIMICA ET BIOPHYSICA ACTA. BIOENERGETICS 2023; 1864:148958. [PMID: 36758662 DOI: 10.1016/j.bbabio.2023.148958] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 07/05/2022] [Revised: 12/24/2022] [Accepted: 02/01/2023] [Indexed: 02/11/2023]
Abstract
Pyruvate:quinone oxidoreductases (PQOs) catalyse the oxidative decarboxylation of pyruvate to acetate and concomitant reduction of quinone to quinol with the release of CO2. They are thiamine pyrophosphate (TPP) and flavin-adenine dinucleotide (FAD) containing enzymes, which interact with the membrane in a monotopic way. PQOs are considered as part of alternatives to most recognized pyruvate catabolizing pathways, and little is known about their taxonomic distribution and structural/functional relationship. In this bioinformatics work we tackled these gaps in PQO knowledge. We used the KEGG database to identify PQO coding genes, performed a multiple sequence analysis which allowed us to study the amino acid conservation on these enzymes, and looked at their possible cellular function. We observed that PQOS are enzymes exclusively present in prokaryotes with most of the sequences identified in bacteria. Regarding the amino acid sequence conservation, we found that 75 amino acid residues (out of 570, on average) have a conservation over 90 %, and that the most conserved regions in the protein are observed around the TPP and FAD binding sites. We systematized the presence of conserved features involved in Mg2+, TPP and FAD binding, as well as residues directly linked to the catalytic mechanism. We also established the presence of a new motif named "HEH lock", possibly involved in the dimerization process. The results here obtained for the PQO protein family contribute to a better understanding of the biochemistry of these respiratory enzymes.
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Affiliation(s)
- Filipe M Sousa
- Instituto de Tecnologia Química e Biológica-António Xavier, Universidade Nova de Lisboa, Av. da República EAN, 2780-157 Oeiras, Portugal; BioISI-Biosystems & Integrative Sciences Institute and Department of Chemistry and Biochemistry, Faculty of Sciences, University of Lisboa, 1749-016 Lisboa, Portugal
| | - Bárbara Fernandes
- Instituto de Tecnologia Química e Biológica-António Xavier, Universidade Nova de Lisboa, Av. da República EAN, 2780-157 Oeiras, Portugal; BioISI-Biosystems & Integrative Sciences Institute and Department of Chemistry and Biochemistry, Faculty of Sciences, University of Lisboa, 1749-016 Lisboa, Portugal
| | - Manuela M Pereira
- Instituto de Tecnologia Química e Biológica-António Xavier, Universidade Nova de Lisboa, Av. da República EAN, 2780-157 Oeiras, Portugal; BioISI-Biosystems & Integrative Sciences Institute and Department of Chemistry and Biochemistry, Faculty of Sciences, University of Lisboa, 1749-016 Lisboa, Portugal.
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Jeje O, Ewunkem AJ, Jeffers-Francis LK, Graves JL. Serving Two Masters: Effect of Escherichia coli Dual Resistance on Antibiotic Susceptibility. Antibiotics (Basel) 2023; 12:antibiotics12030603. [PMID: 36978471 PMCID: PMC10044975 DOI: 10.3390/antibiotics12030603] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/07/2023] [Revised: 03/10/2023] [Accepted: 03/13/2023] [Indexed: 03/30/2023] Open
Abstract
The prevalence of multidrug-resistant bacteria and their increased pathogenicity has led to a growing interest in metallic antimicrobial materials and bacteriophages as potential alternatives to conventional antibiotics. This study examines how resistance to excess iron (III) influences the evolution of bacteriophage resistance in the bacterium Escherichia coli. We utilized experimental evolution in E. coli to test the effect of the evolution of phage T7 resistance on populations resistant to excess iron (III) and populations without excess iron resistance. Phage resistance evolved rapidly in both groups. Dual-resistant (iron (III)/phage) populations were compared to their controls (excess iron (III)-resistant, phage-resistant, no resistance to either) for their performance against each stressor, excess iron (III) and phage; and correlated resistances to excess iron (II), gallium (III), silver (I) and conventional antibiotics. Excess iron (III)/phage-resistant populations demonstrated superior 24 h growth compared to all other populations when exposed to increasing concentrations of iron (II, III), gallium (III), ampicillin, and tetracycline. No differences in 24 h growth were shown between excess iron (III)/phage-resistant and excess iron (III)-resistant populations in chloramphenicol, sulfonamide, and silver (I). The genomic analysis identified selective sweeps in the iron (III) resistant (rpoB, rpoC, yegB, yeaG), phage-resistant (clpX →/→ lon, uvaB, yeaG, fliR, gatT, ypjF, waaC, rpoC, pgi, and yjbH) and iron (III)/phage resistant populations (rcsA, hldE, rpoB, and waaC). E. coli selected for resistance to both excess iron (III) and T7 phage showed some evidence of a synergistic effect on various components of fitness. Dual selection resulted in correlated resistances to ionic metals {iron (II), gallium (III), and silver (I)} and several conventional antibiotics. There is a likelihood that this sort of combination antimicrobial treatment may result in bacterial variants with multiple resistances.
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Affiliation(s)
- Olusola Jeje
- Biology Department, North Carolina Agricultural and Technical State University, 1601 E Market Street, Greensboro, NC 27411, USA
| | - Akamu J Ewunkem
- Department of Biological Sciences, Winston Salem State University, 601 S Martin Luther King Jr Drive, Winston Salem, NC 27110, USA
| | - Liesl K Jeffers-Francis
- Biology Department, North Carolina Agricultural and Technical State University, 1601 E Market Street, Greensboro, NC 27411, USA
| | - Joseph L Graves
- Biology Department, North Carolina Agricultural and Technical State University, 1601 E Market Street, Greensboro, NC 27411, USA
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Paesa M, Remirez de Ganuza C, Alejo T, Yus C, Irusta S, Arruebo M, Sebastian V, Mendoza G. Elucidating the mechanisms of action of antibiotic-like ionic gold and biogenic gold nanoparticles against bacteria. J Colloid Interface Sci 2023; 633:786-799. [PMID: 36493743 DOI: 10.1016/j.jcis.2022.11.138] [Citation(s) in RCA: 6] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/22/2022] [Revised: 11/17/2022] [Accepted: 11/27/2022] [Indexed: 11/30/2022]
Abstract
The antimicrobial action of gold depends on different factors including its oxidation state in the intra- and extracellular medium, the redox potential, its ability to produce reactive oxygen species (ROS), the medium components, the properties of the targeted bacteria wall, its penetration in the bacterial cytosol, the cell membrane potential, and its interaction with intracellular components. We demonstrate that different gold species are able to induce bacterial wall damage as a result of their electrostatic interaction with the cell membrane, the promotion of ROS generation, and the consequent DNA damage. In-depth genomic and proteomic studies on Escherichia coli confirmed the superior toxicity of Au (III) vs Au (I) based on the different molecular mechanisms analyzed including oxidative stress, bacterial energetic metabolism, biosynthetic processes, and cell transport. At equivalent bactericidal doses of Au (III) and Au (I) eukaryotic cells were not as affected as bacteria did, maintaining unaffected cell viability, morphology, and focal adhesions; however, increased ROS generation and disruption in the mitochondrial membrane potential were also observed. Herein, we shed light on the antimicrobial mechanisms of ionic and biogenic gold nanoparticles against bacteria. Under selected conditions antibiotic-like ionic gold can exert a strong antimicrobial activity while being harmless to human cells.
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Affiliation(s)
- Monica Paesa
- Department of Chemical Engineering, University of Zaragoza, Campus Río Ebro-Edificio I+D, C/ Poeta Mariano Esquillor S/N, 50018 Zaragoza, Spain; Instituto de Nanociencia y Materiales de Aragón (INMA), CSIC-Universidad de Zaragoza, Zaragoza 50009, Spain
| | - Cristina Remirez de Ganuza
- Department of Chemical Engineering, University of Zaragoza, Campus Río Ebro-Edificio I+D, C/ Poeta Mariano Esquillor S/N, 50018 Zaragoza, Spain; Instituto de Nanociencia y Materiales de Aragón (INMA), CSIC-Universidad de Zaragoza, Zaragoza 50009, Spain
| | - Teresa Alejo
- Department of Chemical Engineering, University of Zaragoza, Campus Río Ebro-Edificio I+D, C/ Poeta Mariano Esquillor S/N, 50018 Zaragoza, Spain; Instituto de Nanociencia y Materiales de Aragón (INMA), CSIC-Universidad de Zaragoza, Zaragoza 50009, Spain; Aragon Health Research Institute (IIS Aragon), 50009-Zaragoza, Spain; Networking Research Center on Bioengineering, Biomaterials and Nanomedicine, CIBER-BBN, 28029-Madrid, Spain
| | - Cristina Yus
- Department of Chemical Engineering, University of Zaragoza, Campus Río Ebro-Edificio I+D, C/ Poeta Mariano Esquillor S/N, 50018 Zaragoza, Spain
| | - Silvia Irusta
- Department of Chemical Engineering, University of Zaragoza, Campus Río Ebro-Edificio I+D, C/ Poeta Mariano Esquillor S/N, 50018 Zaragoza, Spain; Instituto de Nanociencia y Materiales de Aragón (INMA), CSIC-Universidad de Zaragoza, Zaragoza 50009, Spain; Aragon Health Research Institute (IIS Aragon), 50009-Zaragoza, Spain; Networking Research Center on Bioengineering, Biomaterials and Nanomedicine, CIBER-BBN, 28029-Madrid, Spain
| | - Manuel Arruebo
- Department of Chemical Engineering, University of Zaragoza, Campus Río Ebro-Edificio I+D, C/ Poeta Mariano Esquillor S/N, 50018 Zaragoza, Spain; Instituto de Nanociencia y Materiales de Aragón (INMA), CSIC-Universidad de Zaragoza, Zaragoza 50009, Spain; Aragon Health Research Institute (IIS Aragon), 50009-Zaragoza, Spain; Networking Research Center on Bioengineering, Biomaterials and Nanomedicine, CIBER-BBN, 28029-Madrid, Spain.
| | - Víctor Sebastian
- Department of Chemical Engineering, University of Zaragoza, Campus Río Ebro-Edificio I+D, C/ Poeta Mariano Esquillor S/N, 50018 Zaragoza, Spain; Instituto de Nanociencia y Materiales de Aragón (INMA), CSIC-Universidad de Zaragoza, Zaragoza 50009, Spain; Aragon Health Research Institute (IIS Aragon), 50009-Zaragoza, Spain; Networking Research Center on Bioengineering, Biomaterials and Nanomedicine, CIBER-BBN, 28029-Madrid, Spain.
| | - Gracia Mendoza
- Aragon Health Research Institute (IIS Aragon), 50009-Zaragoza, Spain; Networking Research Center on Bioengineering, Biomaterials and Nanomedicine, CIBER-BBN, 28029-Madrid, Spain
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Jaén-Luchoro D, Karlsson R, Busquets A, Piñeiro-Iglesias B, Karami N, Marathe NP, Moore ERB. Knockout of Targeted Plasmid-Borne β-Lactamase Genes in an Extended-Spectrum-β-Lactamase-Producing Escherichia coli Strain: Impact on Resistance and Proteomic Profile. Microbiol Spectr 2023; 11:e0386722. [PMID: 36622237 PMCID: PMC9927464 DOI: 10.1128/spectrum.03867-22] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/01/2022] [Accepted: 12/09/2022] [Indexed: 01/10/2023] Open
Abstract
Resistance to β-lactams is known to be multifactorial, although the underlying mechanisms are not well established. The aim of our study was to develop a system for assessing the phenotypic and proteomic responses of bacteria to antibiotic stress as a result of the loss of selected antimicrobial resistance genes. We applied homologous recombination to knock out plasmid-borne β-lactamase genes (blaOXA-1, blaTEM-1, and blaCTX-M15) in Escherichia coli CCUG 73778, generating knockout clone variants lacking the respective deleted β-lactamases. Quantitative proteomic analyses were performed on the knockout variants and the wild-type strain, using bottom-up liquid chromatography tandem mass spectrometry (LC-MS/MS), after exposure to different concentrations of cefadroxil. Loss of the blaCTX-M-15 gene had the greatest impact on the resulting protein expression dynamics, while losses of blaOXA-1 and blaTEM-1 affected fewer proteins' expression levels. Proteins involved in antibiotic resistance, cell membrane integrity, stress, and gene expression and unknown function proteins exhibited differential expression. The present study provides a framework for studying protein expression in response to antibiotic exposure and identifying the genomic, proteomic, and phenotypic impacts of resistance gene loss. IMPORTANCE The critical situation regarding antibiotic resistance requires a more in-depth effort for understanding underlying mechanisms involved in antibiotic resistance, beyond just detecting resistance genes. The methodology presented in this work provides a framework for knocking out selected resistance factors, to study the adjustments of the bacterium in response to a particular antibiotic stress, elucidating the genetic response and proteins that are mobilized. The protocol uses MS-based determination of the proteins that are expressed in response to an antibiotic, enabling the selection of strong candidates representing putative resistance factors or mechanisms and providing a basis for future studies to understand their implications in antibiotic resistance. This allows us to better understand how the cell responds to the presence of the antibiotic when a specific gene is lost and, consequently, identify alternative targets for possible future treatment development.
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Affiliation(s)
- Daniel Jaén-Luchoro
- Department of Infectious Diseases, Institute for Biomedicine, Sahlgrenska Academy of the University of Gothenburg, Gothenburg, Sweden
- Centre for Antibiotic Resistance Research, University of Gothenburg, Gothenburg, Sweden
- Culture Collection University of Gothenburg, Department of Clinical Microbiology, Sahlgrenska University Hospital, Region Västra Götaland and Sahlgrenska Academy, University of Gothenburg, Gothenburg, Sweden
| | - Roger Karlsson
- Department of Clinical Microbiology, Sahlgrenska University Hospital, Region Västra Götaland, Gothenburg, Sweden
- Nanoxis Consulting AB, Gothenburg, Sweden
| | - Antonio Busquets
- Microbiology, Department of Biology, University of the Balearic Islands, Palma de Mallorca, Spain
| | - Beatriz Piñeiro-Iglesias
- Department of Infectious Diseases, Institute for Biomedicine, Sahlgrenska Academy of the University of Gothenburg, Gothenburg, Sweden
- Culture Collection University of Gothenburg, Department of Clinical Microbiology, Sahlgrenska University Hospital, Region Västra Götaland and Sahlgrenska Academy, University of Gothenburg, Gothenburg, Sweden
| | - Nahid Karami
- Department of Infectious Diseases, Institute for Biomedicine, Sahlgrenska Academy of the University of Gothenburg, Gothenburg, Sweden
- Centre for Antibiotic Resistance Research, University of Gothenburg, Gothenburg, Sweden
- Department of Clinical Microbiology, Sahlgrenska University Hospital, Region Västra Götaland, Gothenburg, Sweden
| | | | - Edward R. B. Moore
- Department of Infectious Diseases, Institute for Biomedicine, Sahlgrenska Academy of the University of Gothenburg, Gothenburg, Sweden
- Centre for Antibiotic Resistance Research, University of Gothenburg, Gothenburg, Sweden
- Culture Collection University of Gothenburg, Department of Clinical Microbiology, Sahlgrenska University Hospital, Region Västra Götaland and Sahlgrenska Academy, University of Gothenburg, Gothenburg, Sweden
- Department of Clinical Microbiology, Sahlgrenska University Hospital, Region Västra Götaland, Gothenburg, Sweden
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Shleeva MO, Kaprelyants AS. Hypobiosis of Mycobacteria: Biochemical Aspects. BIOCHEMISTRY (MOSCOW) 2023; 88:S52-S74. [PMID: 37069114 DOI: 10.1134/s0006297923140043] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 02/22/2023]
Abstract
Under suboptimal growth conditions, bacteria can transit to the dormant forms characterized by a significantly reduced metabolic activity, resistance to various stress factors, and absence of cell proliferation. Traditionally, the dormant state is associated with the formation of highly differentiated cysts and spores. However, non-spore-forming bacteria can transfer to the dormant-like hypobiotic state with the generation of less differentiated cyst-like forms (which are different from spores). This review focuses on morphological and biochemical changes occurred during formation of dormant forms of mycobacteria in particular pathogenic M. tuberculosis (Mtb) caused latent forms of tuberculosis. These forms are characterized by the low metabolic activity, the absence of cell division, resistance to some antibiotics, marked morphological changes, and loss of ability to grow on standard solid media ("non-culturable" state). Being produced in vitro, dormant Mtb retained ability to maintain latent infection in mice. After a long period of dormancy, mycobacteria retain a number of stable proteins with a potential enzymatic activity which could participate in maintaining of low-level metabolic activity in period of dormancy. Indeed, the metabolomic analysis showed significant levels of metabolites in the dormant cells even after a long period of dormancy, which may be indicative of residual metabolism in dormant mycobacteria. Special role may play intracellularly accumulated trehalose in dormant mycobacteria. Trehalose appears to stabilize dormant cells, as evidenced by the direct correlation between the trehalose content and cell viability during the long-term dormancy. In addition, trehalose can be considered as a reserve energy substrate consumed during reactivation of dormant mycobacteria due to the ATP-dependent conversion of trehalase from the latent to the active state. Another feature of dormant mycobacteria is a high representation of proteins participating in the enzymatic defense against stress factors and of low-molecular-weight compounds protecting cells in the absence of replication. Dormant mycobacteria contain a large number of hydrolyzing enzymes, which, on the one hand, ensure inactivation of biomolecules damaged by stress. On the other hand, the products of these enzymatic reactions can be used for the maintenance of energy state and vital activity of bacterial cells during their long-term survival in the dormant state, i.e., for creating a situation that we propose to refer to as the "catabolic survival". In general, dormant non-replicating mycobacterial cells can be described as morphologically altered forms that contain principal macromolecules and are stabilized and protected from the damaging factors by an arsenal of proteins and low-molecular-weight compounds. Because of the presumable occurrence of metabolic reactions in such cells, this form of survival should be referred to as hypobiosis.
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Affiliation(s)
- Margarita O Shleeva
- A.N. Bach Institute of Biochemistry, Federal Research Centre "Fundamentals of Biotechnology" of the Russian Academy of Sciences, Moscow, 119071, Russia.
| | - Arseny S Kaprelyants
- A.N. Bach Institute of Biochemistry, Federal Research Centre "Fundamentals of Biotechnology" of the Russian Academy of Sciences, Moscow, 119071, Russia
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Zou M, Wang K, Zhao J, Lu H, Yang H, Huang M, Wang L, Wang G, Huang J, Min X. DegS protease regulates the motility, chemotaxis, and colonization of Vibrio cholerae. Front Microbiol 2023; 14:1159986. [PMID: 37089576 PMCID: PMC10113495 DOI: 10.3389/fmicb.2023.1159986] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/06/2023] [Accepted: 03/14/2023] [Indexed: 04/25/2023] Open
Abstract
In bacteria, DegS protease functions as an activating factor of the σE envelope stress response system, which ultimately activates the transcription of stress response genes in the cytoplasm. On the basis of high-throughput RNA sequencing, we have previously found that degS knockout inhibits the expression of flagellum synthesis- and chemotaxis-related genes, thereby indicating that DegS may be involved in the regulation of V. cholerae motility. In this study, we examined the relationships between DegS and motility in V. cholerae. Swimming motility and chemotaxis assays revealed that degS or rpoE deletion promotes a substantial reduction in the motility and chemotaxis of V. cholerae, whereas these activities were restored in ΔdegS::degS and ΔdegSΔrseA strains, indicating that DegS is partially dependent on σE to positively regulate V. cholerae activity. Gene-act network analysis revealed that the cAMP-CRP-RpoS signaling pathway, which plays an important role in flagellar synthesis, is significantly inhibited in ΔdegS mutants, whereas in response to the overexpression of cyaA/crp and rpoS in the ΔdegS strain, the motility and chemotaxis of the ΔdegS + cyaA/crp and ΔdegS + rpoS strains were partially restored compared with the ΔdegS strain. We further demonstrated that transcription levels of the flagellar regulatory gene flhF are regulated by DegS via the cAMP-CRP-RpoS signaling pathway. Overexpression of the flhF gene in the ΔdegS strain partially restored motility and chemotaxis. In addition, suckling mouse intestinal colonization experiments indicated that the ΔdegS and ΔrpoE strains were characterized by the poor colonization of mouse intestines, whereas colonization efficacy was restored in the ΔdegSΔrseA, ΔdegS + cyaA/crp, ΔdegS + rpoS, and ΔdegS + flhF strains. Collectively, our findings indicate that DegS regulates the motility and chemotaxis of V. cholerae via the cAMP-CRP-RpoS-FlhF pathway, thereby influencing the colonization of suckling mouse intestines.
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Affiliation(s)
- Mei Zou
- Department of Laboratory Medicine, Affiliated Hospital of Zunyi Medical University, Zunyi, Guizhou, China
- School of Laboratory Medicine, Zunyi Medical University, Zunyi, Guizhou, China
| | - Kaiying Wang
- Department of Laboratory Medicine, Affiliated Hospital of Zunyi Medical University, Zunyi, Guizhou, China
- School of Laboratory Medicine, Zunyi Medical University, Zunyi, Guizhou, China
| | - Jiajun Zhao
- Department of Laboratory Medicine, Affiliated Hospital of Zunyi Medical University, Zunyi, Guizhou, China
- School of Laboratory Medicine, Zunyi Medical University, Zunyi, Guizhou, China
| | - Huifang Lu
- Department of Laboratory Medicine, Affiliated Hospital of Zunyi Medical University, Zunyi, Guizhou, China
- School of Laboratory Medicine, Zunyi Medical University, Zunyi, Guizhou, China
| | - Hui Yang
- Department of Laboratory Medicine, Affiliated Hospital of Zunyi Medical University, Zunyi, Guizhou, China
- School of Laboratory Medicine, Zunyi Medical University, Zunyi, Guizhou, China
| | - Meirong Huang
- School of Laboratory Medicine, Zunyi Medical University, Zunyi, Guizhou, China
- Department of Blood Transfusion, Affiliated Hospital of Zunyi Medical University, Zunyi, Guizhou, China
| | - Lu Wang
- Department of Laboratory Medicine, Affiliated Hospital of Zunyi Medical University, Zunyi, Guizhou, China
- School of Laboratory Medicine, Zunyi Medical University, Zunyi, Guizhou, China
| | - Guangli Wang
- Department of Laboratory Medicine, Affiliated Hospital of Zunyi Medical University, Zunyi, Guizhou, China
- School of Laboratory Medicine, Zunyi Medical University, Zunyi, Guizhou, China
| | - Jian Huang
- Department of Laboratory Medicine, Affiliated Hospital of Zunyi Medical University, Zunyi, Guizhou, China
- School of Laboratory Medicine, Zunyi Medical University, Zunyi, Guizhou, China
| | - Xun Min
- Department of Laboratory Medicine, Affiliated Hospital of Zunyi Medical University, Zunyi, Guizhou, China
- School of Laboratory Medicine, Zunyi Medical University, Zunyi, Guizhou, China
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Zabed HM, Akter S, Rupani PF, Akor J, Zhang Y, Zhao M, Zhang C, Ragauskas AJ, Qi X. Biocatalytic gateway to convert glycerol into 3-hydroxypropionic acid in waste-based biorefineries: Fundamentals, limitations, and potential research strategies. Biotechnol Adv 2023; 62:108075. [PMID: 36502965 DOI: 10.1016/j.biotechadv.2022.108075] [Citation(s) in RCA: 9] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/14/2022] [Revised: 12/02/2022] [Accepted: 12/03/2022] [Indexed: 12/14/2022]
Abstract
Microbial conversion of bioenergy-derived waste glycerol into value-added chemicals has emerged as an important bioprocessing technology due to its eco-friendliness, feasible technoeconomics, and potential to provide sustainability in biodiesel and bioethanol production. Glycerol is an abundant liquid waste from bioenergy plants with a projected volume of 6 million tons by 2025, accounting for about 10% of biodiesel and 2.5% of bioethanol yields. 3-Hydroxypropionic acid (3-HP) is a major product of glycerol bioconversion, which is the third largest biobased platform compound with expected market size and value of 3.6 million tons/year and USD 10 billion/year, respectively. Despite these biorefinery values, 3-HP biosynthesis from glycerol is still at an immature stage of commercial exploitation. The main challenges behind this immaturity are the toxic effects of 3-HPA on cells, the distribution of carbon flux to undesirable pathways, low tolerance of cells to glycerol and 3-HP, co-factor dependence of enzymes, low enzyme activity and stability, and the problems of substrate inhibition and specificity of enzymes. To address these challenges, it is necessary to understand the fundamentals of glycerol bioconversion and 3-HP production in terms of metabolic pathways, related enzymes, cell factories, midstream process configurations, and downstream 3-HP recovery, as discussed in this review critically and comprehensively. It is equally important to know the current challenges and limitations in 3-HP production, which are discussed in detail along with recent research efforts and remaining gaps. Finally, possible research strategies are outlined considering the recent technological advances in microbial biosynthesis, aiming to attract further research efforts to achieve a sustainable and industrially exploitable 3-HP production technology. By discussing the use of advanced tools and strategies to overcome the existing challenges in 3-HP biosynthesis, this review will attract researchers from many other similar biosynthesis technologies and provide a common gateway for their further development.
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Affiliation(s)
- Hossain M Zabed
- School of Food & Biological Engineering, Jiangsu University, 301 Xuefu Road, Zhenjiang 212013, Jiangsu Province, China
| | - Suely Akter
- School of Food & Biological Engineering, Jiangsu University, 301 Xuefu Road, Zhenjiang 212013, Jiangsu Province, China
| | - Parveen Fatemah Rupani
- Department of Chemical Engineering, Ku Luven, Jan De Nayerlaan 5, 2860 Sint-Katelijne-Waver, Belgium
| | - Joseph Akor
- School of Food & Biological Engineering, Jiangsu University, 301 Xuefu Road, Zhenjiang 212013, Jiangsu Province, China
| | - Yufei Zhang
- School of Food & Biological Engineering, Jiangsu University, 301 Xuefu Road, Zhenjiang 212013, Jiangsu Province, China
| | - Mei Zhao
- School of Food & Biological Engineering, Jiangsu University, 301 Xuefu Road, Zhenjiang 212013, Jiangsu Province, China
| | - Cunsheng Zhang
- School of Food & Biological Engineering, Jiangsu University, 301 Xuefu Road, Zhenjiang 212013, Jiangsu Province, China
| | - Arthur J Ragauskas
- Department of Chemical and Biomolecular Engineering, University of Tennessee Knoxville, Knoxville, TN 37996, USA; Department of Forestry, Wildlife, and Fisheries, Center for Renewable Carbon, The University of Tennessee Institute of Agriculture, Knoxville, TN 37996, USA; UTK-ORNL Joint Institute for Biological Science, Biosciences Division, Oak Ridge National Laboratory, Oak Ridge, TN 37831, USA.
| | - Xianghui Qi
- School of Food & Biological Engineering, Jiangsu University, 301 Xuefu Road, Zhenjiang 212013, Jiangsu Province, China; School of Life Sciences, Guangzhou University, Guangzhou 510,006, Guangdong Province, China.
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Chiang CJ, Chang CH, Chao YP. Programmed cell-lysis system based on hybrid sigma factor-dependent promoters. J Taiwan Inst Chem Eng 2022. [DOI: 10.1016/j.jtice.2022.104611] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/27/2022]
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Bian Z, Liu W, Jin J, Hao Y, Jiang L, Xie Y, Zhang H. Rcs phosphorelay affects the sensitivity of Escherichia coli to plantaricin BM-1 by regulating biofilm formation. Front Microbiol 2022; 13:1071351. [PMID: 36504793 PMCID: PMC9729257 DOI: 10.3389/fmicb.2022.1071351] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/23/2022] [Accepted: 11/10/2022] [Indexed: 11/27/2022] Open
Abstract
Introduction: Plantaricin BM-1 is a class IIa bacteriocin produced by Lactobacillus plantarum BM-1 that exerts significant antibacterial activity against many foodborne bacteria. Studies have shown that class IIa bacteriocins inhibit Gram-positive bacteria via the mannose phosphotransferase system; however, their mechanism of action against Gram-negative bacteria remains unknown. In this study, we explored the mechanism through which the Rcs phosphorelay affects the sensitivity of Escherichia coli K12 cells to plantaricin BM-1. Methods and Results: The minimum inhibitory concentrations of plantaricin BM-1 against E. coli K12, E. coli JW5917 (rcsC mutant), E. coli JW2204 (rcsD mutant), and E. coli JW2205 (rcsB mutant) were 1.25, 0.59, 1.31, and 1.22 mg/ml, respectively. Growth curves showed that E. coli JW5917 sensitivity to plantaricin BM-1 increased to the same level as that of E. coli K12 after complementation. Meanwhile, scanning electron microscopy and transmission electron microscopy revealed that, under the action of plantaricin BM-1, the appearance of E. coli JW5917 cells did not significantly differ from that of E. coli K12 cells; however, cell contents were significantly reduced and plasmolysis and shrinkage were observed at both ends. Crystal violet staining and laser scanning confocal microscopy showed that biofilm formation was significantly reduced after rcsC mutation, while proteomic analysis identified 382 upregulated and 260 downregulated proteins in E. coli JW5917. In particular, rcsC mutation was found to affect the expression of proteins related to biofilm formation, with growth curve assays showing that the deletion of these proteins increased E. coli sensitivity to plantaricin BM-1. Discussion: Consequently, we speculated that the Rcs phosphorelay may regulate the sensitivity of E. coli to plantaricin BM-1 by affecting biofilm formation. This finding of class IIa bacteriocin against Gram-negative bacteria mechanism provides new insights.
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Affiliation(s)
- Zheng Bian
- Beijing Laboratory of Food Quality and Safety, Beijing Key Laboratory of Agricultural Product Detection and Control of Spoilage Organisms and Pesticide Residue, Beijing Engineering Technology Research Center of Food Safety Immune Rapid Detection, College of Food Science and Engineering, Beijing University of Agriculture, Beijing, China
| | - Wenbo Liu
- Beijing Laboratory of Food Quality and Safety, Beijing Key Laboratory of Agricultural Product Detection and Control of Spoilage Organisms and Pesticide Residue, Beijing Engineering Technology Research Center of Food Safety Immune Rapid Detection, College of Food Science and Engineering, Beijing University of Agriculture, Beijing, China
| | - Junhua Jin
- Beijing Laboratory of Food Quality and Safety, Beijing Key Laboratory of Agricultural Product Detection and Control of Spoilage Organisms and Pesticide Residue, Beijing Engineering Technology Research Center of Food Safety Immune Rapid Detection, College of Food Science and Engineering, Beijing University of Agriculture, Beijing, China
| | - Yanling Hao
- Department of Nutrition and Health, Key Laboratory of Functional Dairy, Co-constructed by Ministry of Education and Beijing Government, China Agricultural University, Beijing, China
| | - Linshu Jiang
- Animal Science and Technology College, Beijing University of Agriculture, Beijing, China
| | - Yuanhong Xie
- Beijing Laboratory of Food Quality and Safety, Beijing Key Laboratory of Agricultural Product Detection and Control of Spoilage Organisms and Pesticide Residue, Beijing Engineering Technology Research Center of Food Safety Immune Rapid Detection, College of Food Science and Engineering, Beijing University of Agriculture, Beijing, China,*Correspondence: Yuanhong Xie, ; Hongxing Zhang,
| | - Hongxing Zhang
- Beijing Laboratory of Food Quality and Safety, Beijing Key Laboratory of Agricultural Product Detection and Control of Spoilage Organisms and Pesticide Residue, Beijing Engineering Technology Research Center of Food Safety Immune Rapid Detection, College of Food Science and Engineering, Beijing University of Agriculture, Beijing, China,*Correspondence: Yuanhong Xie, ; Hongxing Zhang,
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Raad N, Tandon D, Hapfelmeier S, Polacek N. The stationary phase-specific sRNA FimR2 is a multifunctional regulator of bacterial motility, biofilm formation and virulence. Nucleic Acids Res 2022; 50:11858-11875. [PMID: 36354005 PMCID: PMC9723502 DOI: 10.1093/nar/gkac1025] [Citation(s) in RCA: 6] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/25/2022] [Revised: 10/06/2022] [Accepted: 10/20/2022] [Indexed: 11/11/2022] Open
Abstract
Bacterial pathogens employ a plethora of virulence factors for host invasion, and their use is tightly regulated to maximize infection efficiency and manage resources in a nutrient-limited environment. Here we show that during Escherichia coli stationary phase the 3' UTR-derived small non-coding RNA FimR2 regulates fimbrial and flagellar biosynthesis at the post-transcriptional level, leading to biofilm formation as the dominant mode of survival under conditions of nutrient depletion. FimR2 interacts with the translational regulator CsrA, antagonizing its functions and firmly tightening control over motility and biofilm formation. Generated through RNase E cleavage, FimR2 regulates stationary phase biology by fine-tuning target mRNA levels independently of the chaperones Hfq and ProQ. The Salmonella enterica orthologue of FimR2 induces effector protein secretion by the type III secretion system and stimulates infection, thus linking the sRNA to virulence. This work reveals the importance of bacterial sRNAs in modulating various aspects of bacterial physiology including stationary phase and virulence.
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Affiliation(s)
- Nicole Raad
- Department of Chemistry, Biochemistry, and Pharmaceutical Sciences, University of Bern, Bern, Switzerland,Graduate School for Cellular and Biomedical Sciences, Bern, Switzerland
| | - Disha Tandon
- Graduate School for Cellular and Biomedical Sciences, Bern, Switzerland,Institute for Infectious Diseases, University of Bern, Bern, Switzerland
| | | | - Norbert Polacek
- To whom correspondence should be addressed. Tel: +41 31 684 43 20;
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Jiang T, Li C, Zou Y, Zhang J, Gan Q, Yan Y. Establishing an Autonomous Cascaded Artificial Dynamic (AutoCAD) regulation system for improved pathway performance. Metab Eng 2022; 74:1-10. [PMID: 36041638 PMCID: PMC10947494 DOI: 10.1016/j.ymben.2022.08.009] [Citation(s) in RCA: 16] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/20/2022] [Revised: 07/21/2022] [Accepted: 08/23/2022] [Indexed: 11/17/2022]
Abstract
Endogenous metabolic pathways in microbial cells are usually precisely controlled by sophisticated regulation networks. However, the lack of such regulations when introducing heterologous pathways in microbial hosts often causes unbalanced enzyme expression and carbon flux distribution, hindering the construction of highly efficient microbial biosynthesis systems. Here, using naringenin as the target compound, we developed an Autonomous Cascaded Artificial Dynamic (AutoCAD) regulation system to automatically coordinate the pathway expression and redirect carbon fluxes for enhanced naringenin production. The AutoCAD regulation system, consisting of both intermediate-based feedforward and product-based feedback control genetic circuits, resulted in a 16.5-fold increase in naringenin titer compared with the static control. Fed-batch fermentation using the strain with AutoCAD regulation further enhanced the naringenin titer to 277.2 mg/L. The AutoCAD regulation system, with intermediate-based feedforward control and product-triggered feedback control, provides a new paradigm of developing complicated cascade dynamic control to engineer heterologous pathways.
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Affiliation(s)
- Tian Jiang
- School of Chemical, Materials, and Biomedical Engineering, College of Engineering, The University of Georgia, Athens, GA, 30602, USA
| | - Chenyi Li
- School of Chemical, Materials, and Biomedical Engineering, College of Engineering, The University of Georgia, Athens, GA, 30602, USA
| | - Yusong Zou
- School of Chemical, Materials, and Biomedical Engineering, College of Engineering, The University of Georgia, Athens, GA, 30602, USA
| | - Jianli Zhang
- School of Chemical, Materials, and Biomedical Engineering, College of Engineering, The University of Georgia, Athens, GA, 30602, USA
| | - Qi Gan
- School of Chemical, Materials, and Biomedical Engineering, College of Engineering, The University of Georgia, Athens, GA, 30602, USA
| | - Yajun Yan
- School of Chemical, Materials, and Biomedical Engineering, College of Engineering, The University of Georgia, Athens, GA, 30602, USA.
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Behbahani SB, Kiridena SD, Wijayaratna UN, Taylor C, Anker JN, Tzeng TRJ. pH variation in medical implant biofilms: Causes, measurements, and its implications for antibiotic resistance. Front Microbiol 2022; 13:1028560. [PMID: 36386694 PMCID: PMC9659913 DOI: 10.3389/fmicb.2022.1028560] [Citation(s) in RCA: 10] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/26/2022] [Accepted: 09/22/2022] [Indexed: 01/28/2023] Open
Abstract
The advent of implanted medical devices has greatly improved the quality of life and increased longevity. However, infection remains a significant risk because bacteria can colonize device surfaces and form biofilms that are resistant to antibiotics and the host's immune system. Several factors contribute to this resistance, including heterogeneous biochemical and pH microenvironments that can affect bacterial growth and interfere with antibiotic biochemistry; dormant regions in the biofilm with low oxygen, pH, and metabolites; slow bacterial growth and division; and poor antibody penetration through the biofilm, which may also be regions with poor acid product clearance. Measuring pH in biofilms is thus key to understanding their biochemistry and offers potential routes to detect and treat latent infections. This review covers the causes of biofilm pH changes and simulations, general findings of metabolite-dependent pH gradients, methods for measuring pH in biofilms, effects of pH on biofilms, and pH-targeted antimicrobial-based approaches.
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Affiliation(s)
| | | | | | - Cedric Taylor
- Department of Biological Sciences, Clemson University, Clemson, SC, United States
| | - Jeffrey N. Anker
- Department of Chemistry, Clemson University, Clemson, SC, United States
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Heating Rate during Shell Egg Thermal Treatment Elicits Stress Responses and Alters Virulence of Salmonella enterica Serovar Enteritidis; Implications for Shell Egg Pasteurization. Appl Environ Microbiol 2022; 88:e0114022. [PMID: 36197091 PMCID: PMC9599327 DOI: 10.1128/aem.01140-22] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
Thermal pasteurization of shell eggs, at various time-temperature combinations, has been proposed previously and implemented industrially. This study was conducted to determine if shell egg heating rate, which varies with different pasteurization implementations, alters the Salmonella enterica serovar Enteritidis response to different stresses or expression of virulence. Shell eggs, containing Salmonella Enteritidis in yolk, were subjected to a low (2.4°C/min) or a high (3.5°C/min) heating rate during treatments that mimicked the pasteurization temperature come-up stage. The low heating rate protected Salmonella from the following processes: (i) lethal heat at the holding stage, (ii) loss of viability during 8-h cooling after heating, and (iii) sequential antimicrobial ozone treatment. Transcriptional analysis using Salmonella reporter strains revealed that the heat stress response gene grpE was transcribed at 3-fold-higher levels (P = 0.0009) at the low than at the high heating rate. Slow heating also significantly increased the transcription of the Salmonella virulence-related genes sopB (P = 0.0012) and sseA (P = 0.0006) in comparison to fast heating. Salmonella virulence was determined experimentally as 50% lethal dose (LD50) values in an in vivo model. The slow heat treatment mildly increased Salmonella Enteritidis virulence in mice (LD50 of 3.3 log CFU), compared to that in nontreated yolk (LD50 of 3.9 log CFU). However, when ozone application followed the slow heat treatment, Salmonella virulence decreased (LD50 of 4.2 log CFU) compared to that for heat-treated or nontreated yolk. In conclusion, heating shell eggs at a low rate can trigger hazardous responses that may compromise the safety of the final pasteurized products but following the thermal treatment with ozone application may help alleviate these concerns. IMPORTANCE Pasteurization of shell eggs is an important technology designed to protect consumers against Salmonella Enteritidis that contaminates this commodity. A low heating rate is preferred over a high rate during shell egg thermal pasteurization due to product quality concern. However, it is not known whether raising the temperature at different rates, during pasteurizing, would potentially affect product safety determinants. The current study demonstrated that slow heating during the pasteurization come-up stage increased the following risks: (i) resistance of Salmonella to pasteurization holding stage or to subsequent ozone treatment, (ii) recovery of Salmonella during the cooling that followed pasteurization, and (iii) Salmonella's ability to cause disease (i.e., virulence). Our findings inform food processors about potential safety risks to consumers resulting from improper use of processing parameters during shell egg pasteurization. Additionally, treating shell eggs with ozone after heat treatment could alleviate these hazards and protect consumers from natural Salmonella Enteritidis contaminants in shell eggs.
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Immobilized Stenotrophomonas maltophilia KB2 in Naproxen Degradation. Molecules 2022; 27:molecules27185795. [PMID: 36144528 PMCID: PMC9501314 DOI: 10.3390/molecules27185795] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/22/2022] [Revised: 09/05/2022] [Accepted: 09/06/2022] [Indexed: 11/20/2022] Open
Abstract
Immobilization is a commonly used method in response to the need to increase the resistance of microorganisms to the toxic effects of xenobiotics. In this study, a plant sponge from Luffa cylindrica was used as a carrier for the immobilization of the Stenotrophomonas maltophilia KB2 strain since such a carrier meets the criteria for high-quality carriers, i.e., low price and biodegradability. The optimal immobilization conditions were established as a temperature of 30 °C, pH 7.2, incubation time of 72 h, and an optical density of the culture of 1.4. The strain immobilized in such conditions was used for the biodegradation of naproxen, and an average rate of degradation of 3.8 µg/hour was obtained under cometabolic conditions with glucose. The obtained results indicate that a microbiological preparation based on immobilized cells on a luffa sponge can be used in bioremediation processes where it is necessary to remove the introduced carrier.
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Role of RpoS in Regulating Stationary Phase Salmonella Typhimurium Pathogenesis-Related Stress Responses under Physiological Low Fluid Shear Force Conditions. mSphere 2022; 7:e0021022. [PMID: 35913142 PMCID: PMC9429890 DOI: 10.1128/msphere.00210-22] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/24/2022] Open
Abstract
The discovery that biomechanical forces regulate microbial virulence was established with the finding that physiological low fluid shear (LFS) forces altered gene expression, stress responses, and virulence of the enteric pathogen Salmonella enterica serovar Typhimurium during the log phase. These log phase LFS-induced phenotypes were independent of the master stress response regulator, RpoS (σS). Given the central importance of RpoS in regulating stationary-phase stress responses of S. Typhimurium cultured under conventional shake flask and static conditions, we examined its role in stationary-phase cultures grown under physiological LFS. We constructed an isogenic rpoS mutant derivative of wild-type S. Typhimurium and compared the ability of these strains to survive in vitro pathogenesis-related stresses that mimic those encountered in the infected host and environment. We also compared the ability of these strains to colonize (adhere, invade, and survive within) human intestinal epithelial cell cultures. Unexpectedly, LFS-induced resistance of stationary-phase S. Typhimurium cultures to acid and bile salts stresses did not rely on RpoS. Likewise, RpoS was dispensable for stationary-phase LFS cultures to adhere to and survive within intestinal epithelial cells. In contrast, the resistance of these cultures to challenges of oxidative and thermal stresses, and their invasion into intestinal epithelial cells was influenced by RpoS. These findings expand our mechanistic understanding of how physiological fluid shear forces modulate stationary-phase S. Typhimurium physiology in unexpected ways and provide clues into microbial mechanobiology and nuances of Salmonella responses to microenvironmental niches in the infected host. IMPORTANCE Bacterial pathogens respond dynamically to a variety of stresses in the infected host, including physical forces of fluid flow (fluid shear) across their surfaces. While pathogens experience wide fluctuations in fluid shear during infection, little is known about how these forces regulate microbial pathogenesis. This is especially important for stationary-phase bacterial growth, which is a critical period to understand microbial resistance, survival, and infection potential, and is regulated in many bacteria by the general stationary-phase stress response protein RpoS. Here, we showed that, unlike conventional culture conditions, several stationary-phase Salmonella pathogenic stress responses were not impacted by RpoS when bacteria were cultured under fluid shear conditions relevant to those encountered in the intestine of the infected host. These findings offer new insight into how physiological fluid shear forces encountered by Salmonella during infection might impact pathogenic responses in unexpected ways that are relevant to their disease-causing ability.
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Ye C, Feng M, Chen Y, Zhang Y, Chen Q, Yu X. Dormancy induced by oxidative damage during disinfection facilitates conjugation of ARGs through enhancing efflux and oxidative stress: A lagging response. WATER RESEARCH 2022; 221:118798. [PMID: 35779456 DOI: 10.1016/j.watres.2022.118798] [Citation(s) in RCA: 33] [Impact Index Per Article: 11.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 03/18/2022] [Revised: 06/21/2022] [Accepted: 06/24/2022] [Indexed: 06/15/2023]
Abstract
Disinfection is known to greatly alter bacterial characteristics in water, and high horizontal gene transfer (HGT) frequency occurs in eutrophic conditions. Interestingly, these two seemingly irrelevant phenomena were closely linked by a lagging response of the increased conjugation frequency probably via daily water disinfection in this study. Three disinfection methods (UV, chlorine, and UV/chlorine) were selected to investigate the increased frequency of conjugation of ARGs during the stage of continuing culture after disinfection. The results showed that the conjugative transfer frequency was inhibited for all disinfection treatments after 24 h of co-incubation. Unexpectedly, after 3-7 days of co-cultivation, the HGT frequencies were increased by 2.71-5.61-fold and 5.46-13.96-fold in chlorine (30 min) and UV/chlorine (1 min) groups compared to the control, but not in UV-irradiated groups. A neglected lagging response was found for the first time, i.e., oxidative disinfection-induced dormancy promotes conjugative transfer of ARGs. Furthermore, mechanistic insights were gained from (1) membrane permeability, (2) conjugation-regulated system, (3) efflux pump system, and (4) oxidative stress system, suggesting the critical role of enhancing efflux and oxidative stress in the propagation of ARGs. Finally, the known instantaneous effect of oxidation disinfection was compared to address the controversial debate in this research field, proposing that the dormancy level of donor bacteria is the key to evaluating whether it can promote the HGT process. This study has important environmental implications for elucidating the transmission of ARGs after oxidation disinfection.
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Affiliation(s)
- Chengsong Ye
- College of the Environment and Ecology, Xiamen University, Xiamen 361102, PR China
| | - Mingbao Feng
- College of the Environment and Ecology, Xiamen University, Xiamen 361102, PR China.
| | - Yuqi Chen
- College of the Environment and Ecology, Xiamen University, Xiamen 361102, PR China
| | - Yiting Zhang
- College of the Environment and Ecology, Xiamen University, Xiamen 361102, PR China
| | - Qian Chen
- College of the Environment and Ecology, Xiamen University, Xiamen 361102, PR China
| | - Xin Yu
- College of the Environment and Ecology, Xiamen University, Xiamen 361102, PR China.
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Dawan J, Ahn J. Bacterial Stress Responses as Potential Targets in Overcoming Antibiotic Resistance. Microorganisms 2022; 10:microorganisms10071385. [PMID: 35889104 PMCID: PMC9322497 DOI: 10.3390/microorganisms10071385] [Citation(s) in RCA: 69] [Impact Index Per Article: 23.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/07/2022] [Revised: 07/03/2022] [Accepted: 07/08/2022] [Indexed: 12/23/2022] Open
Abstract
Bacteria can be adapted to adverse and detrimental conditions that induce general and specific responses to DNA damage as well as acid, heat, cold, starvation, oxidative, envelope, and osmotic stresses. The stress-triggered regulatory systems are involved in bacterial survival processes, such as adaptation, physiological changes, virulence potential, and antibiotic resistance. Antibiotic susceptibility to several antibiotics is reduced due to the activation of stress responses in cellular physiology by the stimulation of resistance mechanisms, the promotion of a resistant lifestyle (biofilm or persistence), and/or the induction of resistance mutations. Hence, the activation of bacterial stress responses poses a serious threat to the efficacy and clinical success of antibiotic therapy. Bacterial stress responses can be potential targets for therapeutic alternatives to antibiotics. An understanding of the regulation of stress response in association with antibiotic resistance provides useful information for the discovery of novel antimicrobial adjuvants and the development of effective therapeutic strategies to control antibiotic resistance in bacteria. Therefore, this review discusses bacterial stress responses linked to antibiotic resistance in Gram-negative bacteria and also provides information on novel therapies targeting bacterial stress responses that have been identified as potential candidates for the effective control of Gram-negative antibiotic-resistant bacteria.
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Affiliation(s)
- Jirapat Dawan
- Department of Biomedical Science, Kangwon National University, Chuncheon 24341, Gangwon, Korea;
| | - Juhee Ahn
- Department of Biomedical Science, Kangwon National University, Chuncheon 24341, Gangwon, Korea;
- Institute of Bioscience and Biotechnology, Kangwon National University, Chuncheon 24341, Gangwon, Korea
- Correspondence: ; Tel.: +82-33-250-6564
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Evolutionary Instability of Collateral Susceptibility Networks in Ciprofloxacin-Resistant Clinical Escherichia coli Strains. mBio 2022; 13:e0044122. [PMID: 35862779 PMCID: PMC9426462 DOI: 10.1128/mbio.00441-22] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/03/2022] Open
Abstract
Collateral sensitivity and resistance occur when resistance development toward one antimicrobial either potentiates or deteriorates the effect of others. Previous reports on collateral effects on susceptibility focus on newly acquired resistance determinants and propose that novel treatment guidelines informed by collateral networks may reduce the evolution, selection, and spread of antimicrobial resistance. In this study, we investigate the evolutionary stability of collateral networks in five ciprofloxacin-resistant, clinical Escherichia coli strains. After 300 generations of experimental evolution without antimicrobials, we show complete fitness restoration in four of five genetic backgrounds and demonstrate evolutionary instability in collateral networks of newly acquired resistance determinants. We show that compensatory mutations reducing efflux expression are the main drivers destabilizing initial collateral networks and identify rpoS as a putative target for compensatory evolution. Our results add another layer of complexity to future predictions and clinical application of collateral networks.
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Regulatory effect of polyamines and indole on expression of stress adaptation genes in <i> Escherichia coli </i>. ACTA BIOMEDICA SCIENTIFICA 2022. [DOI: 10.29413/abs.2022-7.3.16] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/21/2022] Open
Abstract
Background. Indole and polyamines are involved in the regulation of physiological processes in bacteria associated with adaptation to stress, biofilm formation, antibiotic tolerance, and bacterial persistence. However, the molecular targets and mechanisms of action of these metabolites are still poorly understood. In this work, we studied the effect of polyamines and indole on the expression of such genes as: rpoS, relA, and spoT, encoding regulators of the general stress responses and starvation; hns and stpA, encoding global regulators of gene expression; rmf, yqjD, hpf, raiA, rsfS, sra, ettA, encoding ribosome hibernation factors.The aim. To study the regulatory effects of polyamines and indole on the expression of these genes, which are responsible for the adaptation of Escherichia coli to stress.Materials and methods. We used strains of E. coli in this study. The amount of polyamines was studied by thin layer chromatography. The indole concentration was determined by high performance liquid chromatography. Gene expression was studied using real-time RT-PCR.Results. The addition of polyamines putrescine, cadaverine and spermidine to the medium stimulated the expression of all the studied genes. The maximal stimulation was observed at the stationary phase mostly. Putrescine and spermidine had the most significant effect. At 24 h of cultivation, an equimolar conversion of exogenous tryptophan into indole was showed. At this time, the expression of two genes – rmf and raiA – increased.Conclusions. We have shown that polyamines upregulate the expression of all the studied genes at the transcriptional level. The stimulating effect is specific for the phase of the batch culture and the type of polyamine. Indole has a positive effect on the expression of the rmf and raiA genes.
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45
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Dynamic Mechanism of Phase Variations in Bacteria Based on Multistable Gene Regulatory Networks. J Theor Biol 2022; 549:111212. [DOI: 10.1016/j.jtbi.2022.111212] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/06/2020] [Revised: 06/23/2022] [Accepted: 06/28/2022] [Indexed: 11/20/2022]
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Interrelation between Stress Management and Secretion Systems of Ralstonia solanacearum: An In Silico Assessment. Pathogens 2022; 11:pathogens11070730. [PMID: 35889976 PMCID: PMC9325324 DOI: 10.3390/pathogens11070730] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/19/2022] [Revised: 06/18/2022] [Accepted: 06/24/2022] [Indexed: 11/17/2022] Open
Abstract
Ralstonia solanacearum (Rs), the causative agent of devastating wilt disease in several major and minor economic crops, is considered one of the most destructive bacterial plant pathogens. However, the mechanism(s) by which Rs counteracts host-associated environmental stress is still not clearly elucidated. To investigate possible stress management mechanisms, orthologs of stress-responsive genes in the Rs genome were searched using a reference set of known genes. The genome BLAST approach was used to find the distributions of these orthologs within different Rs strains. BLAST results were first confirmed from the KEGG Genome database and then reconfirmed at the protein level from the UniProt database. The distribution pattern of these stress-responsive factors was explored through multivariate analysis and STRING analysis. STRING analysis of stress-responsive genes in connection with different secretion systems of Rs was also performed. Initially, a total of 28 stress-responsive genes of Rs were confirmed in this study. STRING analysis revealed an additional 7 stress-responsive factors of Rs, leading to the discovery of a total of 35 stress-responsive genes. The segregation pattern of these 35 genes across 110 Rs genomes was found to be almost homogeneous. Increasing interactions of Rs stress factors were observed in six distinct clusters, suggesting six different types of stress responses: membrane stress response (MSR), osmotic stress response (OSR), oxidative stress response (OxSR), nitrosative stress response (NxSR), and DNA damage stress response (DdSR). Moreover, a strong network of these stress responses was observed with type 3 secretion system (T3SS), general secretory proteins (GSPs), and different types of pili (T4P, Tad, and Tat). To the best of our knowledge, this is the first report on overall stress response management by Rs and the potential connection with secretion systems.
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Dong S, Liu X, Chen T, Zhou X, Li S, Fu S, Gong H. Mutation of rpoS is Beneficial for Suppressing Organic Acid Secretion During 1,3-Propandiol Biosynthesis in Klebsiella pneumoniae. Curr Microbiol 2022; 79:218. [PMID: 35704098 DOI: 10.1007/s00284-022-02901-w] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/01/2021] [Accepted: 05/09/2022] [Indexed: 11/25/2022]
Abstract
In this study, to reduce the formation of organic acid during 1,3-propanediol biosynthesis in Klebsiella pneumoniae, a method combining UV mutagenesis and high-throughput screening with pH color plates was employed to obtain K. pneumoniae mutants. When compared with the parent strain, the total organic acid formation by the mutant decreased, whereas 1,3-propanediol biosynthesis increased after 24 h anaerobic shake flask culture. Subsequently, genetic changes in the mutant were analyzed by whole-genome sequencing and verified by signal gene deletion. Mutation of the rpoS gene was confirmed to contribute to the regulation of organic acid synthesis in K. pneumoniae. Besides, rpoS deletion eliminated the formation of 2,3-butanediol, the main byproduct produced during 1,3-propanediol fermentation, indicating the role of rpoS in metabolic regulation in K. pneumoniae. Thus, a K. pneumoniae mutant was developed, which could produce lower organic acid during 1,3-propanediol fermentation due to an rpoS mutation in this study.
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Affiliation(s)
- Shufan Dong
- State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, 130 Meilong Road, Shanghai, 200237, People's Republic of China
| | - Xuxia Liu
- State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, 130 Meilong Road, Shanghai, 200237, People's Republic of China
| | - Tianyu Chen
- State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, 130 Meilong Road, Shanghai, 200237, People's Republic of China
| | - Xiaoqin Zhou
- State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, 130 Meilong Road, Shanghai, 200237, People's Republic of China
| | - Shengming Li
- State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, 130 Meilong Road, Shanghai, 200237, People's Republic of China
| | - Shuilin Fu
- State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, 130 Meilong Road, Shanghai, 200237, People's Republic of China
| | - Heng Gong
- State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, 130 Meilong Road, Shanghai, 200237, People's Republic of China.
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48
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Shotgun Proteomics Revealed Preferential Degradation of Misfolded In Vivo Obligate GroE Substrates by Lon Protease in Escherichia coli. MOLECULES (BASEL, SWITZERLAND) 2022; 27:molecules27123772. [PMID: 35744894 PMCID: PMC9228906 DOI: 10.3390/molecules27123772] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 04/26/2022] [Revised: 06/08/2022] [Accepted: 06/08/2022] [Indexed: 11/25/2022]
Abstract
The Escherichia coli chaperonin GroEL/ES (GroE) is one of the most extensively studied molecular chaperones. So far, ~80 proteins in E. coli are identified as GroE substrates that obligately require GroE for folding in vivo. In GroE-depleted cells, these substrates, when overexpressed, tend to form aggregates, whereas the GroE substrates expressed at low or endogenous levels are degraded, probably due to misfolded states. However, the protease(s) involved in the degradation process has not been identified. We conducted a mass-spectrometry-based proteomics approach to investigate the effects of three ATP-dependent proteases, Lon, ClpXP, and HslUV, on the E. coli proteomes under GroE-depleted conditions. A label-free quantitative proteomic method revealed that Lon protease is the dominant protease that degrades the obligate GroE substrates in the GroE-depleted cells. The deletion of DnaK/DnaJ, the other major E. coli chaperones, in the ∆lon strain did not cause major alterations in the expression or folding of the obligate GroE substrates, supporting the idea that the folding of these substrates is predominantly dependent on GroE.
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49
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Sass TH, Ferrazzoli AE, Lovett ST. DnaA and SspA regulation of the iraD gene of Escherichia coli: an alternative DNA damage response independent of LexA/RecA. Genetics 2022; 221:6571813. [PMID: 35445706 PMCID: PMC9157160 DOI: 10.1093/genetics/iyac062] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/19/2021] [Accepted: 10/05/2021] [Indexed: 11/13/2022] Open
Abstract
The transcription factor RpoS of Escherichia coli controls many genes important for tolerance of a variety of stress conditions. IraD promotes the post-translation stability of RpoS by inhibition of RssB, an adaptor protein for ClpXP degradation. We have previously documented DNA damage induction of iraD expression, independent of the SOS response. Both iraD and rpoS are required for tolerance to DNA damaging treatments such as H2O2 and the replication inhibitor azidothymidine in the log phase of growth. Using luciferase gene fusions to the 672 bp iraD upstream region, we show here that both promoters of iraD are induced by azidothymidine. Genetic analysis suggests that both promoters are repressed by DnaA-ATP, partially dependent on a putative DnaA box at -81 bp and are regulated by regulatory inactivation of DnaA, dependent on the DnaN processivity clamp. By electrophoretic mobility shift assays, we show that purified DnaA protein binds to the iraD upstream region, so DnaA regulation of IraD is likely to be direct. DNA damage induction of iraD during log phase growth is abolished in the dnaA-T174P mutant, suggesting that DNA damage, in some way, relieves DnaA repression, possibly through the accumulation of replication clamps and enhanced regulatory inactivation of DnaA. We also demonstrate that the RNA-polymerase associated factor, stringent starvation protein A, induced by the accumulation of ppGpp, also affects iraD expression, with a positive effect on constitutive expression and a negative effect on azidothymidine-induced expression.
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Affiliation(s)
- Thalia H Sass
- Department of Biology, Rosenstiel Basic Medical Sciences Research Center, Brandeis University, Waltham, MA 02454-9110, USA
| | - Alexander E Ferrazzoli
- Department of Biology, Rosenstiel Basic Medical Sciences Research Center, Brandeis University, Waltham, MA 02454-9110, USA
| | - Susan T Lovett
- Department of Biology, Rosenstiel Basic Medical Sciences Research Center, Brandeis University, Waltham, MA 02454-9110, USA
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50
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Hayashi-Nishino M, Aoki K, Kishimoto A, Takeuchi Y, Fukushima A, Uchida K, Echigo T, Yagi Y, Hirose M, Iwasaki K, Shin'ya E, Washio T, Furusawa C, Nishino K. Identification of Bacterial Drug-Resistant Cells by the Convolutional Neural Network in Transmission Electron Microscope Images. Front Microbiol 2022; 13:839718. [PMID: 35369486 PMCID: PMC8965347 DOI: 10.3389/fmicb.2022.839718] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/20/2021] [Accepted: 02/07/2022] [Indexed: 11/13/2022] Open
Abstract
The emergence of bacteria that are resistant to antibiotics is common in areas where antibiotics are used widely. The current standard procedure for detecting bacterial drug resistance is based on bacterial growth under antibiotic treatments. Here we describe the morphological changes in enoxacin-resistant Escherichia coli cells and the computational method used to identify these resistant cells in transmission electron microscopy (TEM) images without using antibiotics. Our approach was to create patches from TEM images of enoxacin-sensitive and enoxacin-resistant E. coli strains, use a convolutional neural network for patch classification, and identify the strains on the basis of the classification results. The proposed method was highly accurate in classifying cells, achieving an accuracy rate of 0.94. Using a gradient-weighted class activation mapping to visualize the region of interest, enoxacin-resistant and enoxacin-sensitive cells were characterized by comparing differences in the envelope. Moreover, Pearson's correlation coefficients suggested that four genes, including lpp, the gene encoding the major outer membrane lipoprotein, were strongly associated with the image features of enoxacin-resistant cells.
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Affiliation(s)
- Mitsuko Hayashi-Nishino
- SANKEN (Institute of Scientific and Industrial Research), Osaka University, Ibaraki, Japan.,Graduate School of Pharmaceutical Sciences, Osaka University, Suita, Japan.,Artificial Intelligence Research Center (AIRC-ISIR), Osaka University, Ibaraki, Japan
| | - Kota Aoki
- SANKEN (Institute of Scientific and Industrial Research), Osaka University, Ibaraki, Japan
| | - Akihiro Kishimoto
- SANKEN (Institute of Scientific and Industrial Research), Osaka University, Ibaraki, Japan
| | - Yuna Takeuchi
- Graduate School of Pharmaceutical Sciences, Osaka University, Suita, Japan
| | - Aiko Fukushima
- SANKEN (Institute of Scientific and Industrial Research), Osaka University, Ibaraki, Japan
| | - Kazushi Uchida
- Graduate School of Pharmaceutical Sciences, Osaka University, Suita, Japan
| | - Tomio Echigo
- Department of Engineering Informatics, Osaka Electro-Communication University, Neyagawa, Japan
| | - Yasushi Yagi
- SANKEN (Institute of Scientific and Industrial Research), Osaka University, Ibaraki, Japan
| | - Mika Hirose
- Institute for Protein Research, Osaka University, Suita, Japan
| | - Kenji Iwasaki
- Life Science Center for Survival Dynamics, Tsukuba Advanced Research Alliance, University of Tsukuba, Tsukuba, Japan
| | - Eitaro Shin'ya
- SANKEN (Institute of Scientific and Industrial Research), Osaka University, Ibaraki, Japan
| | - Takashi Washio
- SANKEN (Institute of Scientific and Industrial Research), Osaka University, Ibaraki, Japan
| | - Chikara Furusawa
- RIKEN, Center for Biosystems Dynamics Research, Suita, Japan.,Universal Biology Institute, The University of Tokyo, Tokyo, Japan
| | - Kunihiko Nishino
- SANKEN (Institute of Scientific and Industrial Research), Osaka University, Ibaraki, Japan.,Graduate School of Pharmaceutical Sciences, Osaka University, Suita, Japan.,Center for Infectious Disease Education and Research, Osaka University, Suita, Japan
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