Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 10.577
Filtrar
1.
Proc Natl Acad Sci U S A ; 118(39)2021 09 28.
Artigo em Inglês | MEDLINE | ID: mdl-34548403

RESUMO

Many biological functions are leaky, and organisms that perform them contribute some of their products to a community "marketplace" in which nonperforming individuals may compete for them. Leaky functions are partitioned unequally in microbial communities, and the evolutionary forces determining which species perform them and which become beneficiaries are poorly understood. Here, we demonstrate that the market principle of comparative advantage determines the distribution of a leaky antibiotic resistance gene in an environment occupied by two "species"-strains of Escherichia coli growing on mutually exclusive resources and thus occupying separate niches. Communities comprised of antibiotic-resistant cells were rapidly invaded by sensitive cells of both types. While the two phenotypes coexisted stably for 500 generations, in 15/18 replicates, antibiotic sensitivity became fixed in one species. Fixation always occurred in the same species despite both species being genetically identical except for their niche-defining mutation. In the absence of antibiotic, the fitness cost of resistance was identical in both species. However, the intrinsic resistance of the species that ultimately became the sole helper was significantly lower, and thus its reward for expressing the resistance gene was higher. Opportunity cost of resistance, not absolute cost or efficiency of antibiotic removal, determined which species became the helper, consistent with the economic theory of comparative advantage. We present a model that suggests that this market-like dynamic is a general property of Black Queen systems and, in communities dependent on multiple leaky functions, could lead to the spontaneous development of an equitable and efficient division of labor.


Assuntos
Antibacterianos/farmacologia , Evolução Biológica , Resistência Microbiana a Medicamentos , Escherichia coli/fisiologia , Microbiota , Escherichia coli/efeitos dos fármacos , Modelos Teóricos
2.
J Immunol ; 207(7): 1848-1856, 2021 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-34452933

RESUMO

Immune cell responses are strikingly altered in patients with severe coronavirus disease 2019 (COVID-19), but the immunoregulatory process in these individuals is not fully understood. In this study, 23 patients with mild and 22 patients with severe COVID-19 and 6 asymptomatic carriers of COVID-19 were enrolled, along with 44 healthy controls (HC). Peripheral immune cells in HC and patients with COVID-19 were comprehensively profiled using mass cytometry. We found that in patients with severe COVID-19, the number of HLA-DRlow/- monocytes was significantly increased, but that of mucosal-associated invariant T (MAIT) cells was greatly reduced. MAIT cells were highly activated but functionally impaired in response to Escherichia coli and IL-12/IL-18 stimulation in patients with severe COVID-19, especially those with microbial coinfection. Single-cell transcriptome analysis revealed that IFN-stimulated genes were significantly upregulated in peripheral MAIT cells and monocytes from patients with severe COVID-19. IFN-α pretreatment suppressed MAIT cells' response to E. coli by triggering high levels of IL-10 production by HLA-DRlow/--suppressive monocytes. Blocking IFN-α or IL-10 receptors rescued MAIT cell function in patients with severe COVID-19. Moreover, plasma from patients with severe COVID-19 inhibited HLA-DR expression by monocytes through IL-10. These data indicate a unique pattern of immune dysregulation in severe COVID-19, which is characterized by enrichment of suppressive HLA-DRlow/- monocytes associated with functional impairment of MAIT cells through the IFN/IL-10 pathway.


Assuntos
COVID-19/imunologia , Infecções por Escherichia coli/imunologia , Escherichia coli/fisiologia , Interleucina-10/metabolismo , Monócitos/imunologia , Células T Invariantes Associadas à Mucosa/imunologia , SARS-CoV-2/fisiologia , Adolescente , Adulto , Doenças Assintomáticas , Células Cultivadas , Criança , Coinfecção , Progressão da Doença , Feminino , Humanos , Tolerância Imunológica , Ativação Linfocitária , Masculino , Pessoa de Meia-Idade , Índice de Gravidade de Doença , Adulto Jovem
3.
Chem Commun (Camb) ; 57(69): 8648-8651, 2021 Sep 07.
Artigo em Inglês | MEDLINE | ID: mdl-34369943

RESUMO

We report that Thioflavin T (ThT), the reference fluorogenic probe for amyloid detection, displays photodynamic activity against bacterial biofilms. ThT recognizes key structures of the biofilm matrix, disrupting the complex architecture and efficiently inactivating bacterial cells. We also show that ThT phototherapy synergistically boosts the activity of conventional antimicrobials.


Assuntos
Antibacterianos/farmacologia , Benzotiazóis/farmacologia , Biofilmes/efeitos dos fármacos , Fármacos Fotossensibilizantes/farmacologia , Escherichia coli/efeitos dos fármacos , Escherichia coli/fisiologia , Luz , Testes de Sensibilidade Microbiana , Staphylococcus aureus/fisiologia
4.
Gene ; 803: 145890, 2021 Nov 30.
Artigo em Inglês | MEDLINE | ID: mdl-34375634

RESUMO

Escherichia coli Nissle 1917 (EcN) is an efficient probiotic strain extensively used worldwide because of its several health benefits. Adhesion to the intestinal cells is one of the prerequisites for a probiotic strain. To identify the genes essential for the adhesion of EcN on the intestinal cells, we utilized a quantitative genetic footprinting approach called transposon insertion sequencing (INSeq). A transposon insertion mutant library of EcN comprising of ~17,000 mutants was used to screen the adherence to the intestinal epithelial cells, Caco-2. The transposon insertion sites were identified from the input and output population by employing next-generation sequencing using the Ion torrent platform. Based on the relative abundance of reads in the input and output pools, we identified 113 candidate genes that are essential for the fitness of EcN during the adhesion and colonization on the Caco-2 cells. Functional categorization revealed that these fitness genes are associated with carbohydrate transport and metabolism, cell wall/membrane/envelope biogenesis, post-translational modification, stress response, motility and adhesion, and signal transduction. To further validate the genes identified in our INSeq analysis, we constructed individual knock-out mutants in five genes (cyclic di-GMP phosphodiesterase (gmp), hda, uidC, leuO, and hypothetical protein-coding gene). We investigated their ability to adhere to Caco-2 cells. Evaluation of these mutants showed reduced adhesion on Caco-2 cells, confirming their role in adhesion. Understanding the functions of these genes may provide novel insights into molecular regulation during colonization of probiotic bacteria to the intestinal cells, and useful to develop designer probiotic strains.


Assuntos
Proteínas de Escherichia coli/genética , Escherichia coli/fisiologia , Mutagênese Insercional , Análise de Sequência de DNA/métodos , Aderência Bacteriana , Células CACO-2 , Elementos de DNA Transponíveis , Escherichia coli/genética , Aptidão Genética , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Probióticos
5.
Am J Physiol Endocrinol Metab ; 321(3): E338-E350, 2021 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-34280051

RESUMO

Obesity is associated with metabolic, immunological, and infectious disease comorbidities, including an increased risk of enteric infection and inflammatory bowel disease such as Crohn's disease (CD). Expansion of intestinal pathobionts such as adherent-invasive Escherichia coli (AIEC) is a common dysbiotic feature of CD, which is amplified by prior use of oral antibiotics. Although high-fat, high-sugar diets are associated with dysbiotic expansion of E. coli, it is unknown if the content of fat or another dietary component in obesogenic diets is sufficient to promote AIEC expansion. Here, we found that administration of an antibiotic combined with feeding mice an obesogenic low-fiber, high-sucrose, high-fat diet (HFD) that is typically used in rodent-obesity studies promoted AIEC intestinal expansion. Even a short-term (i.e., 1 day) pulse of HFD feeding before infection was sufficient to promote AIEC expansion, indicating that the magnitude of obesity was not the main driver of AIEC expansion. Controlled-diet experiments demonstrated that neither dietary fat nor sugar were the key determinants of AIEC colonization, but that lowering dietary fiber from approximately 13% to 5%-6% was sufficient to promote the intestinal expansion of AIEC when combined with antibiotics in mice. When combined with antibiotics, lowering fiber promoted AIEC intestinal expansion to a similar extent as widely used HFDs in mice. However, lowering dietary fiber was sufficient to promote AIEC intestinal expansion without affecting body mass. Our results show that low dietary fiber combined with oral antibiotics are environmental factors that promote the expansion of Crohn's disease-associated pathobionts in the gut.NEW & NOTEWORTHY It is commonly thought that obesity or a high-fat diet alters pathogenic bacteria and promotes inflammatory gut diseases. We found that lower dietary fiber is a key factor that expands a gut pathobiont linked to Crohn's disease, independent of obesity status in mice.


Assuntos
Doença de Crohn/microbiologia , Fibras na Dieta/administração & dosagem , Intestinos/microbiologia , Obesidade/microbiologia , Animais , Escherichia coli/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL
6.
Appl Environ Microbiol ; 87(18): e0093821, 2021 08 26.
Artigo em Inglês | MEDLINE | ID: mdl-34260307

RESUMO

Biofilm formation is important in both the environmental and intestinal phases of the Vibrio cholerae life cycle. Nevertheless, most studies of V. cholerae biofilm formation focus on monospecies cultures, whereas nearly all biofilm communities found in nature consist of a variety of microorganisms. Multispecies biofilms formed between V. cholerae and other bacteria in the environment and the interactions that exist between these species are still poorly understood. In this study, the influence of Escherichia coli on the biofilm formation of V. cholerae was studied in the context of both in vitro coculture and in vivo coinfection. To understand the underlying synergistic mechanisms between these two species and to investigate the role of E. coli in V. cholerae biofilm formation, different pathotypes of E. coli and corresponding deletion mutants lacking genes that influence flagellar motility, curli fibers, or type I pili were cocultured with V. cholerae. Our findings demonstrate that the presence of commensal E. coli increases biofilm formation at the air-liquid interface in vitro and the generation of biofilm-like multicellular clumps in mouse feces. Examination of laboratory E. coli flagellar-motility ΔfliC and ΔmotA mutants in dual-species biofilm formation suggests that flagellar motility plays an important role in the synergistic interaction and coaggregation formation between V. cholerae and E. coli. This study facilitates a better understanding of how V. cholerae resides in harsh environments and colonizes the intestine. IMPORTANCE Biofilms play an important role in the V. cholerae life cycle. Until now, only monospecies biofilm formation of V. cholerae has been well studied. However, in nature, bacteria live in complex microbial communities, where biofilm is mostly composed of multiple microbial species that interact to cooperate with or compete against each other. Uncovering how V. cholerae forms multispecies biofilms is critical for furthering our understanding of how V. cholerae survives in the environment and transitions to infecting the human host. In this work, the dual-species biofilm containing V. cholerae and Escherichia coli was investigated. We demonstrate that the presence of commensal E. coli increased overall biofilm formation. Furthermore, we demonstrate that the motility of E. coli flagella is important for V. cholerae and E. coli to form coaggregation clumps in a dual-species biofilm. These results shed light on a new mechanism for understanding the survival and pathogenesis of V. cholerae.


Assuntos
Biofilmes , Escherichia coli/fisiologia , Vibrio cholerae/fisiologia , Animais , Cólera/microbiologia , Infecções por Escherichia coli/microbiologia , Fezes/microbiologia , Camundongos , Interações Microbianas
7.
J Immunol ; 207(3): 974-984, 2021 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-34282000

RESUMO

K9CATH is the sole cathelicidin in canines (dogs) and exhibits broad antimicrobial activity against both Gram-positive and Gram-negative bacteria. K9CATH also modulates inflammatory responses and binds to LPS. These activities depend on the secondary structure and a net-positive charge of the peptide. Peptidylarginine deiminases (PAD) convert cationic peptidyl arginine to neutral citrulline. Thus, we hypothesized that citrullination is a biologically relevant modification of the peptide that would reduce the antibacterial and LPS-binding activities of K9CATH. Recombinant PAD2 and PAD4 citrullinated K9CATH to various extents and circular dichroism spectroscopy revealed that both native and citrullinated K9CATH exhibited similar α-helical secondary structures. Notably, citrullination of K9CATH reduced its bactericidal activity, abolished its ability to permeabilize the membrane of Gram-negative bacteria and reduced the hemolytic capacity. Electron microscopy showed that citrullinated K9CATH did not cause any morphological changes of Gram-negative bacteria, whereas the native peptide caused clear alterations of membrane integrity, concordant with a rapid bactericidal effect. Finally, citrullination of K9CATH impaired its capacity to inhibit LPS-mediated release of proinflammatory molecules from mouse and canine macrophages. In conclusion, citrullination attenuates the antibacterial and the LPS-binding properties of K9CATH, demonstrating the importance of a net positive charge for antibacterial lysis of bacteria and LPS-binding effects and suggests that citrullination is a means to regulate cathelicidin activities.


Assuntos
Antibacterianos/metabolismo , Anti-Inflamatórios/metabolismo , Peptídeos Catiônicos Antimicrobianos/metabolismo , Infecções por Escherichia coli/imunologia , Escherichia coli/fisiologia , Macrófagos/imunologia , Infecções por Pasteurella/metabolismo , Pasteurella multocida/fisiologia , Desiminases de Arginina em Proteínas/metabolismo , Animais , Antibacterianos/química , Anti-Inflamatórios/química , Peptídeos Catiônicos Antimicrobianos/química , Citrulinação , Cães , Imunidade Inata , Mediadores da Inflamação/metabolismo , Lipopolissacarídeos/metabolismo , Camundongos , Ligação Proteica , Células RAW 264.7
8.
Inorg Chem ; 60(15): 11058-11069, 2021 Aug 02.
Artigo em Inglês | MEDLINE | ID: mdl-34255500

RESUMO

In this work, the synthesis, structural and photophysical characterization of six phosphorescent H2O-soluble Pt(II) complexes are reported while addressing their emission maxima, photoluminescence quantum yields (ΦL), lifetimes (τ), aggregation tendency, and microenvironment sensitivity as a function of the substitution pattern on the main tridentate luminophore. Different ancillary ligands, namely, a trisulfonated phosphane and maltohexaose-conjugated pyridines (with or without amide bridges), were introduced and evaluated for the realization of switch-on-photoluminescent labels reporting on the microenvironment sensed in biofilms of Gram+ and Gram- models, namely, Staphylococcus aureus and Escherichia coli. With the aid of confocal luminescence micro(spectro)scopy, we observed that selected complexes specifically interact with the biofilms while leaving planktonic cells unlabeled. By using photoluminescence lifetime imaging microscopy, excited-state lifetimes within S. aureus biofilms were measured. The photoluminescence intensities were drastically boosted, and the excited state lifetimes were significantly prolonged upon binding to the viscous biofilm matrix, mainly due to the suppression of radiationless deactivation pathways upon shielding from physical quenching processes, such as interactions with solvent molecules and 3O2. The best performances were attained for non-aggregating complexes with maltohexaose targeting units and without amide bridges. Notably, in the absence of the maltodextrin, a hydrophobic adamantyl moiety suffices to attain a sizeable labeling capacity. Moreover, photoluminescence studies showed that selected complexes can also effectively interact with E. coli biofilms, where the bacterial cells are able to partially uptake the maltodextrin-based agents. In summary, the herein introduced concepts enable the development of specific biofilm reporters providing spatial resolution as well as lifetime- and spectrum-based readouts. Considering that most theragnostic agents reported so far mainly address metabolically active bacteria at the surface of biofilms but without reaching cells deeply immersed in the matrix, a new platform with a clear structure-property correlation is provided for the early detection of such bacterial arrays.


Assuntos
Biofilmes , Complexos de Coordenação/química , Escherichia coli/fisiologia , Luminescência , Platina/química , Staphylococcus aureus/fisiologia , Microscopia
9.
Int J Mol Sci ; 22(14)2021 Jul 08.
Artigo em Inglês | MEDLINE | ID: mdl-34298964

RESUMO

The rapid emergence of drug-resistant bacteria is a major global health concern. Antimicrobial peptides (AMPs) and peptidomimetics have arisen as a new class of antibacterial agents in recent years in an attempt to overcome antibiotic resistance. A library of phenylglyoxamide-based small molecular peptidomimetics was synthesised by incorporating an N-alkylsulfonyl hydrophobic group with varying alkyl chain lengths and a hydrophilic cationic group into a glyoxamide core appended to phenyl ring systems. The quaternary ammonium iodide salts 16d and 17c showed excellent minimum inhibitory concentration (MIC) of 4 and 8 µM (2.9 and 5.6 µg/mL) against Staphylococcus aureus, respectively, while the guanidinium hydrochloride salt 34a showed an MIC of 16 µM (8.5 µg/mL) against Escherichia coli. Additionally, the quaternary ammonium iodide salt 17c inhibited 70% S. aureus biofilm formation at 16 µM. It also disrupted 44% of pre-established S. aureus biofilms at 32 µM and 28% of pre-established E. coli biofilms 64 µM, respectively. A cytoplasmic membrane permeability study indicated that the synthesised peptidomimetics acted via disruption and depolarisation of membranes. Moreover, the quaternary ammonium iodide salts 16d and 17c were non-toxic against human cells at their therapeutic dosages against S. aureus.


Assuntos
Antibacterianos , Biofilmes/efeitos dos fármacos , Escherichia coli/fisiologia , Peptidomiméticos , Staphylococcus aureus/fisiologia , Antibacterianos/síntese química , Antibacterianos/química , Antibacterianos/farmacologia , Biofilmes/crescimento & desenvolvimento , Peptidomiméticos/síntese química , Peptidomiméticos/química , Peptidomiméticos/farmacologia , Relação Estrutura-Atividade , Compostos de Sulfonilureia/síntese química , Compostos de Sulfonilureia/química , Compostos de Sulfonilureia/farmacologia
10.
Int J Mol Sci ; 22(11)2021 Jun 07.
Artigo em Inglês | MEDLINE | ID: mdl-34200244

RESUMO

Ribosome biogenesis is a highly coordinated and complex process that requires numerous assembly factors that ensure prompt and flawless maturation of ribosomal subunits. Despite the increasing amount of data collected, the exact role of most assembly factors and mechanistic details of their operation remain unclear, mainly due to the shortage of high-resolution structural information. Here, using cryo-electron microscopy, we characterized 30S ribosomal particles isolated from an Escherichia coli strain with a deleted gene for the RbfA factor. The cryo-EM maps for pre-30S subunits were divided into six classes corresponding to consecutive assembly intermediates: from the particles with a completely unresolved head domain and unfolded central pseudoknot to almost mature 30S subunits with well-resolved body, platform, and head domains and partially distorted helix 44. The structures of two predominant 30S intermediates belonging to most populated classes obtained at 2.7 Å resolutions indicate that RbfA acts at two distinctive 30S assembly stages: early formation of the central pseudoknot including folding of the head, and positioning of helix 44 in the decoding center at a later stage. Additionally, it was shown that the formation of the central pseudoknot may promote stabilization of the head domain, likely through the RbfA-dependent maturation of the neck helix 28. An update to the model of factor-dependent 30S maturation is proposed, suggesting that RfbA is involved in most of the subunit assembly process.


Assuntos
Proteínas de Escherichia coli/metabolismo , Escherichia coli/fisiologia , Proteínas Ribossômicas/metabolismo , Subunidades Ribossômicas Menores de Bactérias/fisiologia , Ribossomos/metabolismo , Sítios de Ligação , Microscopia Crioeletrônica/métodos , Proteínas de Escherichia coli/genética , Modelos Moleculares , Ligação Proteica , Proteínas Ribossômicas/genética , Subunidades Ribossômicas Menores de Bactérias/ultraestrutura , Ribossomos/ultraestrutura
11.
Int J Mol Sci ; 22(12)2021 Jun 14.
Artigo em Inglês | MEDLINE | ID: mdl-34198485

RESUMO

Brain microvascular endothelial cells (BMECs) constitute the structural and functional basis for the blood-brain barrier (BBB) and play essential roles in bacterial meningitis. Although the BBB integrity regulation has been under extensive investigation, there is little knowledge regarding the roles of long non-coding RNAs (lncRNAs) in this event. The present study aimed to investigate the roles of one potential lncRNA, lncRSPH9-4, in meningitic E. coli infection of BMECs. LncRSPH9-4 was cytoplasm located and significantly up-regulated in meningitic E. coli-infected hBMECs. Electrical cell-substrate impedance sensing (ECIS) measurement and Western blot assay demonstrated lncRSPH9-4 overexpression in hBMECs mediated the BBB integrity disruption. By RNA-sequencing analysis, 639 mRNAs and 299 miRNAs were significantly differentiated in response to lncRSPH9-4 overexpression. We further found lncRSPH9-4 regulated the permeability in hBMECs by competitively sponging miR-17-5p, thereby increasing MMP3 expression, which targeted the intercellular tight junctions. Here we reported the infection-induced lncRSPH9-4 aggravated disruption of the tight junctions in hBMECs, probably through the miR-17-5p/MMP3 axis. This finding provides new insights into the function of lncRNAs in BBB integrity during meningitic E. coli infection and provides the novel nucleic acid targets for future treatment of bacterial meningitis.


Assuntos
Barreira Hematoencefálica/metabolismo , Barreira Hematoencefálica/patologia , Escherichia coli/fisiologia , Metaloproteinase 3 da Matriz/metabolismo , Meningites Bacterianas/genética , Meningites Bacterianas/microbiologia , MicroRNAs/metabolismo , RNA Longo não Codificante/metabolismo , Sequência de Bases , Citoplasma/metabolismo , Células Endoteliais/metabolismo , Células Endoteliais/microbiologia , Redes Reguladoras de Genes , Humanos , MicroRNAs/genética , Microvasos/patologia , Modelos Biológicos , Permeabilidade , RNA Longo não Codificante/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Transdução de Sinais , Junções Íntimas/metabolismo , Transcrição Genética , Regulação para Cima/genética
12.
Vet Res ; 52(1): 92, 2021 Jun 22.
Artigo em Inglês | MEDLINE | ID: mdl-34158121

RESUMO

Unlike in chickens, dynamics of the gut microbiome in turkeys is limitedly understood and no data were yet published in context of pathological changes following experimental infection. Thus, the impact of Histomonas meleagridis-associated inflammatory changes in the caecal microbiome, especially the Escherichia coli population and their caecal wall invasion in turkeys was investigated. Birds experimentally inoculated with attenuated and/or virulent H. meleagridis and non-inoculated negative controls were divided based on the severity of macroscopic caecal lesions. The high throughput amplicon sequencing of 16SrRNA showed that the species richness and diversity of microbial community significantly decreased in severely affected caeca. The relative abundances of operational taxonomic units belonging to Anaerotignum lactatifermentans, E. coli, and Faecalibacterium prausnitzii were higher and paralleled with a decreased abundances of those belonging to Alistipes putredinis, Streptococcus alactolyticus, Lactobacillus salivarius and Lactobacillus reuteri in birds with the highest lesion scores. Although the relative abundance of E. coli was higher, the absolute count was not affected by the severity of pathological lesions. Immunohistochemistry showed that E. coli was only present in the luminal content of caecum and did not penetrate even severely inflamed and necrotized caecal wall. Overall, it was demonstrated that the fundamental shift in caecal microbiota of turkeys infected with H. meleagridis was attributed to the pathology induced by the parasite, which only led to relative but not absolute changes in E. coli population. Furthermore, E. coli cells did not show tendency to penetrate the caecal tissue even when the intestinal mucosal barriers were severely compromised.


Assuntos
Galinhas , Microbioma Gastrointestinal , Doenças das Aves Domésticas/parasitologia , Infecções Protozoárias em Animais/parasitologia , Trichomonadida/fisiologia , Tiflite/veterinária , Animais , Contagem de Colônia Microbiana/veterinária , Escherichia coli/fisiologia , Tiflite/parasitologia
13.
ACS Appl Mater Interfaces ; 13(23): 26735-26747, 2021 Jun 16.
Artigo em Inglês | MEDLINE | ID: mdl-34081856

RESUMO

Microfluidic gradient generators have been used to study cellular migration, growth, and drug response in numerous biological systems. One type of device combines a hydrogel and polydimethylsiloxane (PDMS) to generate "flow-free" gradients; however, their requirements for either negative flow or external clamps to maintain fluid-tight seals between the two layers have restricted their utility among broader applications. In this work, a two-layer, flow-free microfluidic gradient generator was developed using thiol-ene chemistry. Both rigid thiol-acrylate microfluidic resin (TAMR) and diffusive thiol-acrylate hydrogel (H) layers were synthesized from commercially available monomers at room temperature and pressure using a base-catalyzed Michael addition. The device consisted of three parallel microfluidic channels negatively imprinted in TAMR layered on top of the thiol-acrylate hydrogel to facilitate orthogonal diffusion of chemicals to the direction of flow. Upon contact, these two layers formed fluid-tight channels without any external pressure due to a strong adhesive interaction between the two layers. The diffusion of molecules through the TAMR/H system was confirmed both experimentally (using fluorescent microscopy) and computationally (using COMSOL). The performance of the TAMR/H system was compared to a conventional PDMS/agarose device with a similar geometry by studying the chemorepulsive response of a motile strain of GFP-expressing Escherichia coli. Population-based analysis confirmed a similar migratory response of both wild-type and mutant E. coli in both of the microfluidic devices. This confirmed that the TAMR/H hybrid system is a viable alternative to traditional PDMS-based microfluidic gradient generators and can be used for several different applications.


Assuntos
Acrilatos/química , Quimiotaxia , Escherichia coli/fisiologia , Hidrogéis/química , Microfluídica/instrumentação , Cimentos de Resina/química , Compostos de Sulfidrila/química , Adesivos , Microfluídica/métodos
14.
AAPS PharmSciTech ; 22(5): 170, 2021 Jun 03.
Artigo em Inglês | MEDLINE | ID: mdl-34085150

RESUMO

A novel nanofiber insert was prepared with a modified electrospinning method to enhance the ocular residence time of ofloxacin (OFX) and to provide a sustained release pattern by covering hydrophilic polymers, chitosan/polyvinyl alcohol (CS/PVA) nanofibers, with a hydrophobic polymer, Eudragit RL100 in layers, and by glutaraldehyde (GA) cross-linking of CS-PVA nanofibers for the treatment of infectious conjunctivitis. The morphology of the prepared nanofibers was studied using scanning electron microscopy (SEM). The average fiber diameter was found to be 123 ± 23 nm for the single electrospun nanofiber with no cross-linking (OFX-O). The single nanofibers, cross-linked for 10 h with GA (OFX-OG), had an average fiber diameter of 159 ± 30 nm. The amount of OFX released from the nanofibers was measured in vitro and in vivo using UV spectroscopy and microbial assay methods against Staphylococcus aureus, respectively. The antimicrobial efficiency of OFX formulated in cross-linked and non-cross-linked nanofibers was affirmed by observing the inhibition zones of Staphylococcus aureus and Escherichia coli. In vivo studies using the OFX nanofibrous inserts on a rabbit eye confirmed a sustained release pattern for up to 96 h. It was found that the cross-linking of the nanofibers by GA vapor could reduce the burst release of OFX from OFX-loaded CS/PVA in one layer and multi-layered nanofibers. In vivo results showed that the AUC0-96 for the nanofibers was 9-20-folds higher compared to the OFX solution. This study thus demonstrates the potential of the nanofiber technology is being utilized to sustained drug release in ocular drug delivery systems.


Assuntos
Resinas Acrílicas/química , Administração Oftálmica , Quitosana/química , Nanofibras/química , Ofloxacino/química , Álcool de Polivinil/química , Resinas Acrílicas/administração & dosagem , Resinas Acrílicas/farmacocinética , Animais , Antibacterianos/química , Química Farmacêutica/métodos , Quitosana/administração & dosagem , Quitosana/farmacocinética , Preparações de Ação Retardada/administração & dosagem , Preparações de Ação Retardada/química , Preparações de Ação Retardada/farmacocinética , Sistemas de Liberação de Medicamentos/métodos , Avaliação Pré-Clínica de Medicamentos/métodos , Escherichia coli/efeitos dos fármacos , Escherichia coli/fisiologia , Nanofibras/administração & dosagem , Ofloxacino/administração & dosagem , Ofloxacino/farmacocinética , Álcool de Polivinil/administração & dosagem , Álcool de Polivinil/farmacocinética , Coelhos , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/fisiologia
15.
Commun Biol ; 4(1): 669, 2021 06 03.
Artigo em Inglês | MEDLINE | ID: mdl-34083715

RESUMO

How motile bacteria navigate environmental chemical gradients has implications ranging from health to climate science, but the underlying behavioral mechanisms are unknown for most species. The well-studied navigation strategy of Escherichia coli forms a powerful paradigm that is widely assumed to translate to other bacterial species. This assumption is rarely tested because of a lack of techniques capable of bridging scales from individual navigation behavior to the resulting population-level chemotactic performance. Here, we present such a multiscale 3D chemotaxis assay by combining high-throughput 3D bacterial tracking with microfluidically created chemical gradients. Large datasets of 3D trajectories yield the statistical power required to assess chemotactic performance at the population level, while simultaneously resolving the underlying 3D navigation behavior for every individual. We demonstrate that surface effects confound typical 2D chemotaxis assays, and reveal that, contrary to previous reports, Caulobacter crescentus breaks with the E. coli paradigm.


Assuntos
Algoritmos , Quimiotaxia/fisiologia , Escherichia coli/fisiologia , Ensaios de Triagem em Larga Escala/métodos , Técnicas Analíticas Microfluídicas/métodos , Modelos Biológicos , Caulobacter crescentus/fisiologia , Especificidade da Espécie
16.
Methods Mol Biol ; 2323: 267-280, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34086287

RESUMO

Spontaneous tumor regression following bacterial infection has been observed for hundreds of years. These observations along with anecdotal medical findings in 1890s led to the development of Coley's "toxins," consisting of killed Streptococcus pyogenes and Serratia marcescens bacteria, as the first cancer immunotherapy. The use of this approach, however, was not widely accepted at the time especially after the introduction of radiation therapy as a treatment for cancer in the early 1900s. Over the last 30-40 years there has been renewed interest in the use of bacteria to treat human solid tumors. This is based on the observation that various nonpathogenic anaerobic bacteria can infiltrate and replicate within solid tumors when given intravenously. Bacteria tested as potential anticancer agents include the Gram-positive obligate anaerobes Bifidobacterium and Clostridium, as well as the gram-negative facultative anaerobe Salmonella. Recent advances in synthetic biology and clinical success in cancer immunotherapy provide renewed momentum for developing bacteria-based cancer immunotherapy for cancer treatment and should allow greater potential for the development of novel therapeutic approaches for this devastating disease.


Assuntos
Terapia Biológica/métodos , Neoplasias/terapia , Interferência de RNA , Biologia Sintética/métodos , Animais , Linhagem Celular Tumoral , Ensaios Clínicos Fase I como Assunto , Neoplasias do Colo/microbiologia , Neoplasias do Colo/terapia , Escherichia coli/genética , Escherichia coli/fisiologia , Feminino , Vetores Genéticos/genética , Vetores Genéticos/uso terapêutico , Humanos , Imunoterapia/métodos , Imunoterapia/tendências , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Nus , Neoplasias/microbiologia , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/uso terapêutico , Reação em Cadeia da Polimerase em Tempo Real/métodos , Indução de Remissão , Salmonella typhimurium/genética , Salmonella typhimurium/fisiologia , Especificidade da Espécie , Organismos Livres de Patógenos Específicos , Biologia Sintética/tendências , Ensaios Antitumorais Modelo de Xenoenxerto
17.
J Mater Chem B ; 9(24): 4906-4914, 2021 06 23.
Artigo em Inglês | MEDLINE | ID: mdl-34100486

RESUMO

We investigated the biomaterial interface between the bacteria Escherichia coli DH5α and silicon nanowire patterned surfaces. We optimised the engineering of silicon nanowire coated surfaces using metal-assisted chemical etching. Using a combination of focussed ion beam scanning electron microscopy, and cell viability and transformation assays, we found that with increasing interfacing force, cell viability decreases, as a result of increasing cell rupture. However, despite this aggressive interfacing regime, a proportion of the bacterial cell population remains viable. We found that the silicon nanowires neither resulted in complete loss of cell viability nor partial membrane disruption and corresponding DNA plasmid transformation. Critically, assay choice was observed to be important, as a reduction-based metabolic reagent was found to yield false-positive results on the silicon nanowire substrate. We discuss the implications of these results for the future design and assessment of bacteria-nanostructure interfacing experiments.


Assuntos
Escherichia coli/efeitos dos fármacos , Escherichia coli/fisiologia , Viabilidade Microbiana/efeitos dos fármacos , Nanofios , Silício/química , Silício/farmacologia , Biotransformação/efeitos dos fármacos , Escherichia coli/metabolismo , Propriedades de Superfície
18.
Molecules ; 26(10)2021 May 18.
Artigo em Inglês | MEDLINE | ID: mdl-34069848

RESUMO

Escherichia coli colonies were grown on different supports for the removal of nitrates from water. A carbon material and different commercial metal oxides, such as SiO2, TiO2 and Al2O3, and their corresponding carbon-metal oxide composites were studied. The physicochemical properties were analyzed by different techniques and the results were correlated with their performance in the denitrification process. Developed biofilms effectively adhere to the supports and always reach the complete reduction of nitrates to gaseous products. Nevertheless, faster processes occur when the biofilm is supported on mesoporous and non-acid materials (carbon and silica).


Assuntos
Agricultura , Biofilmes , Carbono/farmacologia , Nitratos/isolamento & purificação , Óxidos/farmacologia , Águas Residuárias/química , Purificação da Água , Biofilmes/efeitos dos fármacos , Carbono/toxicidade , Desnitrificação , Escherichia coli/crescimento & desenvolvimento , Escherichia coli/fisiologia , Escherichia coli/ultraestrutura , Concentração de Íons de Hidrogênio , Óxidos/toxicidade , Tamanho da Partícula , Termogravimetria , Difração de Raios X
19.
ACS Appl Mater Interfaces ; 13(27): 31406-31417, 2021 Jul 14.
Artigo em Inglês | MEDLINE | ID: mdl-34185501

RESUMO

The use of ultraviolet (UV) and blue irradiation to sterilize surfaces is well established, but commercial applications would be enhanced if the light source is replaced with ambient light. In this paper, it is shown that nanofibers can be explored as an alternative methodology to UV and blue irradiation for bacterial inactivation. It is demonstrated that this is indeed possible using spun nanofibers of poly[lactic-co-(glycolic acid)] (PLGA). This work shows that PLGA spun scaffolds can promote photoinactivation of Staphylococcus aureus and Escherichia coli bacteria with ambient light or with laser irradiation at 630 nm. With the optimized scaffold composition of PLGA85:15 nanofibers, the minimum intensity required to kill the bacteria is much lower than in antimicrobial blue light applications. The enhanced effect introduced by PLGA scaffolds is due to their nanofiber structures since PLGA spun nanofibers were able to inactivate both S. aureus and E. coli bacteria, but cast films had no effect. These findings pave the way for an entirely different method to sterilize surfaces, which is less costly and environmentally friendly than current procedures. In addition, the scaffolds could also be used in cancer treatment with fewer side effects since photosensitizers are not required.


Assuntos
Eletricidade , Escherichia coli/fisiologia , Viabilidade Microbiana/efeitos dos fármacos , Copolímero de Ácido Poliláctico e Ácido Poliglicólico/química , Copolímero de Ácido Poliláctico e Ácido Poliglicólico/farmacologia , Staphylococcus aureus/fisiologia , Raios Ultravioleta , Escherichia coli/efeitos dos fármacos , Escherichia coli/efeitos da radiação , Viabilidade Microbiana/efeitos da radiação , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/efeitos da radiação
20.
Int J Mol Sci ; 22(9)2021 May 06.
Artigo em Inglês | MEDLINE | ID: mdl-34066555

RESUMO

Traditional antimicrobial treatments consist of drugs which target different essential functions in pathogens. Nevertheless, bacteria continue to evolve new mechanisms to evade this drug-mediated killing with surprising speed on the deployment of each new drug and antibiotic worldwide, a phenomenon called antimicrobial resistance (AMR). Nowadays, AMR represents a critical health threat, for which new medical interventions are urgently needed. By 2050, it is estimated that the leading cause of death will be through untreatable AMR pathogens. Although antibiotics remain a first-line treatment, non-antibiotic therapies such as prophylactic vaccines and therapeutic monoclonal antibodies (mAbs) are increasingly interesting alternatives to limit the spread of such antibiotic resistant microorganisms. For the discovery of new vaccines and mAbs, the search for effective antigens that are able to raise protective immune responses is a challenging undertaking. In this context, outer membrane vesicles (OMV) represent a promising approach, as they recapitulate the complete antigen repertoire that occurs on the surface of Gram-negative bacteria. In this review, we present Escherichia coli and Pseudomonas aeruginosa as specific examples of key AMR threats caused by Gram-negative bacteria and we discuss the current status of mAbs and vaccine approaches under development as well as how knowledge on OMV could benefit antigen discovery strategies.


Assuntos
Farmacorresistência Bacteriana , Escherichia coli/fisiologia , Pseudomonas aeruginosa/fisiologia , Animais , Vacinas Bacterianas/imunologia , Infecções por Escherichia coli/epidemiologia , Infecções por Escherichia coli/imunologia , Infecções por Escherichia coli/microbiologia , Humanos , Infecções por Pseudomonas/epidemiologia , Infecções por Pseudomonas/imunologia , Infecções por Pseudomonas/microbiologia
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...