scholarly journals How Bacteria Change after Exposure to Silver Nanoformulations: Analysis of the Genome and Outer Membrane Proteome

Pathogens ◽  
2021 ◽  
Vol 10 (7) ◽  
pp. 817
Author(s):  
Anna Kędziora ◽  
Mateusz Speruda ◽  
Maciej Wernecki ◽  
Bartłomiej Dudek ◽  
Katarzyna Kapczynska ◽  
...  
Keyword(s):  
Outer Membrane ◽  
Optical Density ◽  
Bacterial Cell ◽  
Protein Identification ◽  
Bacterial Genome ◽  
Antibacterial Properties ◽  
Membrane Proteome ◽  
E Coli ◽  
Phenotypic Changes ◽  
Proteomic Approach

Objective: the main purpose of this work was to compare the genetic and phenotypic changes of E. coli treated with silver nanoformulations (E. coli BW25113 wt, E. coli BW25113 AgR, E. coli J53, E. coli ATCC 11229 wt, E. coli ATCC 11229 var. S2 and E. coli ATCC 11229 var. S7). Silver, as the metal with promising antibacterial properties, is currently widely used in medicine and the biomedical industry, in both ionic and nanoparticles forms. Silver nanoformulations are usually considered as one type of antibacterial agent, but their physical and chemical properties determine the way of interactions with the bacterial cell, the mode of action, and the bacterial cell response to silver. Methods: the changes in the bacterial genome, resulting from the treatment of bacteria with various silver nanoformulations, were verified by analyzing of genes (selected with mutfunc) and their conservative and non-conservative mutations selected with BLOSUM62. The phenotype was verified using an outer membrane proteome analysis (OMP isolation, 2-DE electrophoresis, and MS protein identification). Results: the variety of genetic and phenotypic changes in E. coli strains depends on the type of silver used for bacteria treatment. The most changes were identified in E. coli ATCC 11229 treated with silver nanoformulation signed as S2 (E. coli ATCC 11229 var. S2). We pinpointed 39 genes encoding proteins located in the outer membrane, 40 genes of their regulators, and 22 genes related to other outer membrane structures, such as flagellum, fimbria, lipopolysaccharide (LPS), or exopolysaccharide in this strain. Optical density of OmpC protein in E. coli electropherograms decreased after exposure to silver nanoformulation S7 (noticed in E. coli ATCC 11229 var. S7), and increased after treatment with the other silver nanoformulations (SNF) marked as S2 (noticed in E. coli ATCC 11229 var. S2). Increase of FliC protein optical density was identified in turn after Ag+ treatment (noticed in E.coli AgR). Conclusion: the results show that silver nanoformulations (SNF) exerts a selective pressure on bacteria causing both conservative and non-conservative mutations. The proteomic approach revealed that the levels of some proteins have changed after treatment with appropriate SNF.

scholarly journals The Importance of Porins and β-Lactamase in Outer Membrane Vesicles on the Hydrolysis of β-Lactam Antibiotics

2020 ◽  
Vol 21 (8) ◽  
pp. 2822 ◽  
Author(s):  
Si Won Kim ◽  
Jung Seok Lee ◽  
Seong Bin Park ◽  
Ae Rin Lee ◽  
Jae Wook Jung ◽  
...  
Keyword(s):  
Outer Membrane ◽  
Defense Mechanisms ◽  
Antibacterial Properties ◽  
Membrane Vesicles ◽  
Outer Membrane Vesicles ◽  
E Coli ◽  
Lactam Antibiotic ◽  
Hydrolysis Of ◽  
Degradation Ability

Gram-negative bacteria have an outer membrane inhibiting the entry of antibiotics. Porins, found within the outer membrane, are involved in regulating the permeability of β-lactam antibiotics. β-lactamases are enzymes that are able to inactivate the antibacterial properties of β-lactam antibiotics. Interestingly, porins and β-lactamase are found in outer membrane vesicles (OMVs) of β-lactam-resistant Escherichia coli and may be involved in the survival of susceptible strains of E. coli in the presence of antibiotics, through the hydrolysis of the β-lactam antibiotic. In this study, OMVs isolated from β-lactam-resistant E. coli and from mutants, lacking porin or β-lactamase, were evaluated to establish if the porins or β-lactamase in OMVs were involved in the degradation of β-lactam antibiotics. OMVs isolated from E. coli deficient in β-lactamase did not show any degradation ability against β-lactam antibiotics, while OMVs lacking OmpC or OmpF showed significantly lower levels of hydrolyzing activity than OMVs from parent E. coli. These data reveal an important role of OMVs in bacterial defense mechanisms demonstrating that the OmpC and OmpF proteins allow permeation of β-lactam antibiotics into the lumen of OMVs, and antibiotics that enter the OMVs can be degraded by β-lactamase.

scholarly journals Cleavage of Braun lipoprotein Lpp from the bacterial peptidoglycan by a paralog of L,D-transpeptidases, LdtF

2021 ◽  
Author(s):  
Raj Bahadur ◽  
Pavan Kumar Chodisetti ◽  
Manjula Reddy
Keyword(s):  
Outer Membrane ◽  
Bacterial Cell ◽  
Drug Targets ◽  
Structural Integrity ◽  
Cell Envelope ◽  
Hydrolytic Enzyme ◽  
Permeability Barrier ◽  
Gram Negative ◽  
E Coli ◽  
Bacterial Cell Envelope

AbstractGram-negative bacterial cell envelope is made up of an outer membrane (OM), an inner membrane (IM) that surrounds the cytoplasm, and a periplasmic space between the two membranes containing peptidoglycan (PG or murein). PG is an elastic polymer that forms a mesh-like sacculus around the IM protecting cells from turgor and environmental stress conditions. In several bacteria including E. coli, the OM is tethered to PG by an abundant OM lipoprotein, Lpp (or Braun lipoprotein) that functions to maintain the structural and functional integrity of the cell envelope. Since its discovery Lpp has been studied extensively and although L,D-transpeptidases, the enzymes that catalyse the formation of PG–Lpp linkages have been earlier identified, it is not known how these linkages are modulated. Here, using genetic and biochemical approaches, we show that LdtF (formerly yafK), a newly-identified paralog of L,D-transpeptidases in E. coli is a murein hydrolytic enzyme that catalyses cleavage of Lpp from the PG sacculus. LdtF also exhibits glycine-specific carboxypeptidase activity on muropeptides containing a terminal glycine residue. LdtF is earlier presumed to be an L,D-transpeptidase; however, our results show that it is indeed an L,D-endopeptidase that hydrolyses the products generated by the L,D-transpeptidases. To summarize, this study describes the discovery of a murein endopeptidase with a hitherto unknown catalytic specificity that removes the PG–Lpp cross-links suggesting a role for LdtF in regulation of PG-OM linkages to maintain the structural integrity of the bacterial cell envelope.Significance statementBacterial cell walls contain a unique protective exoskeleton, peptidoglycan, which is a target of several clinically important antimicrobials. In Gram-negative bacteria, peptidoglycan is covered by an additional lipid layer, outer membrane that serves as permeability barrier against entry of toxic molecules. In some bacteria, an extremely abundant lipoprotein, Lpp staples outer membrane to peptidoglycan to maintain the structural integrity of the cell envelope. In this study, we identify a previously unknown peptidoglycan hydrolytic enzyme that cleaves Lpp from the peptidoglycan sacculus and show how the outer membrane-peptidoglycan linkages are modulated in Escherichia coli. Overall, this study helps in understanding the fundamental bacterial cell wall biology and in identification of alternate drug targets for development of new antimicrobials.

scholarly journals Comparison of long-read sequencing technologies in interrogating bacteria and fly genomes

2021 ◽  
Author(s):  
Eric S Tvedte ◽  
Mark Gasser ◽  
Benjamin C Sparklin ◽  
Jane Michalski ◽  
Carl E Hjelmen ◽  
...  
Keyword(s):  
Bacterial Genome ◽  
Hybrid Approach ◽  
Cost Effective ◽  
Fruit Fly ◽  
Drosophila Ananassae ◽  
Whole Genome Sequencing Data ◽  
Sequencing Data ◽  
E Coli ◽  
Hybrid Approaches ◽  
Long Read

Abstract The newest generation of DNA sequencing technology is highlighted by the ability to generate sequence reads hundreds of kilobases in length. Pacific Biosciences (PacBio) and Oxford Nanopore Technologies (ONT) have pioneered competitive long read platforms, with more recent work focused on improving sequencing throughput and per-base accuracy. We used whole-genome sequencing data produced by three PacBio protocols (Sequel II CLR, Sequel II HiFi, RS II) and two ONT protocols (Rapid Sequencing and Ligation Sequencing) to compare assemblies of the bacteria Escherichia coli and the fruit fly Drosophila ananassae. In both organisms tested, Sequel II assemblies had the highest consensus accuracy, even after accounting for differences in sequencing throughput. ONT and PacBio CLR had the longest reads sequenced compared to PacBio RS II and HiFi, and genome contiguity was highest when assembling these datasets. ONT Rapid Sequencing libraries had the fewest chimeric reads in addition to superior quantification of E. coli plasmids versus ligation-based libraries. The quality of assemblies can be enhanced by adopting hybrid approaches using Illumina libraries for bacterial genome assembly or polishing eukaryotic genome assemblies, and an ONT-Illumina hybrid approach would be more cost-effective for many users. Genome-wide DNA methylation could be detected using both technologies, however ONT libraries enabled the identification of a broader range of known E. coli methyltransferase recognition motifs in addition to undocumented D. ananassae motifs. The ideal choice of long read technology may depend on several factors including the question or hypothesis under examination. No single technology outperformed others in all metrics examined.

scholarly journals The Distribution of Several Genomic Virulence Determinants Does Not Corroborate the Established Serotyping Classification of Bacillus thuringiensis

2021 ◽  
Vol 22 (5) ◽  
pp. 2244
Author(s):  
Anton E. Shikov ◽  
Yury V. Malovichko ◽  
Arseniy A. Lobov ◽  
Maria E. Belousova ◽  
Anton A. Nizhnikov ◽  
...  
Keyword(s):  
Bacillus Thuringiensis ◽  
Protein Identification ◽  
Core Gene ◽  
Comparative Genomic ◽  
Virulence Determinants ◽  
Strain Characterization ◽  
Proteomic Approach ◽  
Liquid Chromatography Tandem Mass ◽  
Identity Threshold ◽  
Genome Assemblies

Bacillus thuringiensis, commonly referred to as Bt, is an object of the lasting interest of microbiologists due to its highly effective insecticidal properties, which make Bt a prominent source of biologicals. To categorize the exuberance of Bt strains discovered, serotyping assays are utilized in which flagellin serves as a primary seroreactive molecule. Despite its convenience, this approach is not indicative of Bt strains’ phenotypes, neither it reflects actual phylogenetic relationships within the species. In this respect, comparative genomic and proteomic techniques appear more informative, but their use in Bt strain classification remains limited. In the present work, we used a bottom-up proteomic approach based on fluorescent two-dimensional difference gel electrophoresis (2D-DIGE) coupled with liquid chromatography/tandem mass spectrometry(LC-MS/MS) protein identification to assess which stage of Bt culture, vegetative or spore, would be more informative for strain characterization. To this end, the proteomic differences for the israelensis-attributed strains were assessed to compare sporulating cultures of the virulent derivative to the avirulent one as well as to the vegetative stage virulent bacteria. Using the same approach, virulent spores of the israelensis strain were also compared to the spores of strains belonging to two other major Bt serovars, namely darmstadiensis and thuringiensis. The identified proteins were analyzed regarding the presence of the respective genes in the 104 Bt genome assemblies available at open access with serovar attributions specified. Of 21 proteins identified, 15 were found to be encoded in all the present assemblies at 67% identity threshold, including several virulence factors. Notable, individual phylogenies of these core genes conferred neither the serotyping nor the flagellin-based phylogeny but corroborated the reconstruction based on phylogenomics approaches in terms of tree topology similarity. In its turn, the distribution of accessory protein genes was not confined to the existing serovars. The obtained results indicate that neither gene presence nor the core gene sequence may serve as distinctive bases for the serovar attribution, undermining the notion that the serotyping system reflects strains’ phenotypic or genetic similarity. We also provide a set of loci, which fit in with the phylogenomics data plausibly and thus may serve for draft phylogeny estimation of the novel strains.

scholarly journals The Location of Substitutions and Bacterial Genome Arrangements

2020 ◽  
Author(s):  
Daniella F Lato ◽  
G Brian Golding
Keyword(s):  
Sinorhizobium Meliloti ◽  
Bacterial Genome ◽  
Evolutionary Analysis ◽  
Origin Of Replication ◽  
Ancestral Reconstruction ◽  
Molecular Change ◽  
E Coli ◽  
A Genome ◽  
The Impact ◽  
Molecular Evolutionary Analysis

Abstract Increasing evidence supports the notion that different regions of a genome have unique rates of molecular change. This variation is particularly evident in bacterial genomes where previous studies have reported gene expression and essentiality tend to decrease, while substitution rates usually increases with increasing distance from the origin of replication. Genomic reorganization such as rearrangements occur frequently in bacteria and allow for the introduction and restructuring of genetic content, creating gradients of molecular traits along genomes. Here, we explore the interplay of these phenomena by mapping substitutions to the genomes of Escherichia coli, Bacillus subtilis, Streptomyces, and Sinorhizobium meliloti, quantifying how many substitutions have occurred at each position in the genome. Preceding work indicates that substitution rate significantly increases with distance from the origin. Using a larger sample size and accounting for genome rearrangements through ancestral reconstruction, our analysis demonstrates that the correlation between the number of substitutions and distance from the origin of replication is often significant but small and inconsistent in direction. Some replicons had a significantly decreasing trend (E. coli and the chromosome of S. meliloti), while others showed the opposite significant trend (B. subtilis, Streptomyces, pSymA and pSymB in S. meliloti). dN, dS and ω were examined across all genes and there was no significant correlation between those values and distance from the origin. This study highlights the impact that genomic rearrangements and location have on molecular trends in some bacteria, illustrating the importance of considering spatial trends in molecular evolutionary analysis. Assuming that molecular trends are exclusively in one direction can be problematic.

Fe(iii)-complex mediated bacterial cell surface immobilization of eGFP and enzymes

2021 ◽  
Author(s):  
Lilin Feng ◽  
Liang Gao ◽  
Daniel F. Sauer ◽  
Yu Ji ◽  
Haiyang Cui ◽  
...  
Keyword(s):  
Cell Surface ◽  
Bacterial Cell ◽  
Surface Immobilization ◽  
E Coli ◽  
Bacterial Cell Surface ◽  
Reversible Method

A facile and reversible method to immobilize His6-tagged proteins on the E. coli cell surface through the formation of an Fe(iii)-complex.

scholarly journals Green Synthesis and Incorporation of Sericin Silver Nanoclusters into Electrospun Ultrafine Cellulose Acetate Fibers for Anti-Bacterial Applications

Polymers ◽  
2021 ◽  
Vol 13 (9) ◽  
pp. 1411
Author(s):  
Mujahid Mehdi ◽  
Huihui Qiu ◽  
Bing Dai ◽  
Raja Fahad Qureshi ◽  
Sadam Hussain ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Green Synthesis ◽  
Cellulose Acetate ◽  
Antibacterial Properties ◽  
Vital Role ◽  
Silver Nanoclusters ◽  
Composite Fibers ◽  
E Coli ◽  
Antibacterial Performance ◽  
Field Emission Electron Microscopy

Fiber based antibacterial materials have gained an enormous attraction for the researchers in these days. In this study, a novel Sericin Encapsulated Silver Nanoclusters (sericin-AgNCs) were synthesized through single pot and green synthesis route. Subsequently these sericin-AgNCs were incorporated into ultrafine electrospun cellulose acetate (CA) fibers for assessing the antibacterial performance. The physicochemical properties of sericin-AgNCs/CA composite fibers were investigated by transmission electron microscopy (TEM), field emission electron microscopy (FE-SEM), Fourier transform infrared spectroscopy (FTIR) and wide X-ray diffraction (XRD). The antibacterial properties of sericin-AgNCs/CA composite fibers against Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus) were systematically evaluated. The results showed that sericin-AgNCs incorporated in ultrafine CA fibers have played a vital role for antibacterial activity. An amount of 0.17 mg/mL sericin-AgNCs to CA fibers showed more than 90% results and elevated upto >99.9% with 1.7 mg/mL of sericin-AgNCs against E. coli. The study indicated that sericin-AgNCs/CA composite confirms an enhanced antibacterial efficiency, which could be used as a promising antibacterial product.

scholarly journals Interaction of Temporin-L Analogues with the E. coli FtsZ Protein

Antibiotics ◽  
2021 ◽  
Vol 10 (6) ◽  
pp. 704
Author(s):  
Angela Di Somma ◽  
Carolina Canè ◽  
Antonio Moretta ◽  
Angela Duilio
Keyword(s):  
Protein Interactions ◽  
Bacterial Infections ◽  
Antibacterial Properties ◽  
Bacterial Cell Division ◽  
Structural Determinants ◽  
E Coli ◽  
Functional Studies ◽  
Division Process ◽  
Z Ring ◽  
Peptide Complexes

The research of new therapeutic agents to fight bacterial infections has recently focused on the investigation of antimicrobial peptides (AMPs), the most common weapon that all organisms produce to prevent invasion by external pathogens. Among AMPs, the amphibian Temporins constitute a well-known family with high antibacterial properties against Gram-positive and Gram-negative bacteria. In particular, Temporin-L was shown to affect bacterial cell division by inhibiting FtsZ, a tubulin-like protein involved in the crucial step of Z-ring formation at the beginning of the division process. As FtsZ represents a leading target for new antibacterial compounds, in this paper we investigated in detail the interaction of Temporin L with Escherichia coli FtsZ and designed two TL analogues in an attempt to increase peptide-protein interactions and to better understand the structural determinants leading to FtsZ inhibition. The results demonstrated that the TL analogues improved their binding to FtsZ, originating stable protein-peptide complexes. Functional studies showed that both peptides were endowed with a high capability of inhibiting both the enzymatic and polymerization activities of the protein. Moreover, the TL analogues were able to inhibit bacterial growth at low micromolar concentrations. These observations may open up the way to the development of novel peptide or peptidomimetic drugs tailored to bind FtsZ, hampering a crucial process of bacterial life that might be proposed for future pharmaceutical applications.

Sign in / Sign up

Export Citation Format

Share Document