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1.
AMB Express ; 12(1): 1, 2022 Jan 06.
Artigo em Inglês | MEDLINE | ID: mdl-34989907

RESUMO

Yersiniosis is an infectious zoonotic disease caused by two enteropathogenic species of Gram-negative genus Yersinia: Yersinia enterocolitica and Yersinia pseudotuberculosis. Pigs and other wild and domestic animals are reservoirs for these bacteria. Infection is usually spread to humans by ingestion of contaminated food. Yersiniosis is considered a rare disease, but recent studies indicate that it is overlooked in the diagnostic process therefore the infections with this bacterium are not often identified. Reliable diagnosis of Yersiniosis by culturing is difficult due to the slow growth of the bacteria easily overgrown by other more rapidly growing microbes unless selec-tive growth media is used. Phage adhesins recognizing bacteria in a specific manner can be an excellent diagnostic tool, es-pecially in the diagnosis of pathogens difficult for culturing. In this study, it was shown that Gp17, the tail fiber protein (TFP) of phage φYeO3-12, specifically recognizes only the pathogenic Yersinia enterocolitica serotype O:3 (YeO:3) bacteria. The ELISA test used in this work confirmed the specific interaction of this protein with YeO:3 and demonstrated a promising tool for developing the pathogen recognition method based on phage adhesins.

2.
Expert Rev Mol Diagn ; 21(9): 925-937, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34289786

RESUMO

INTRODUCTION: Examination of disease biomarkers mostly performed on crude materials, such as serum, meets some obstacles, resulting from sample complexity and the wide range of concentrations and sizes of the components. Techniques currently used in clinical diagnostics are usually time-consuming and expensive. The more sensitive and portable devices are needed for early diagnostics. Chemical sensors are devices that convert chemical information into parameters suitable for fast and precise processing and measurement. AREA COVERED: We review the use of biosensors and their possible application in early diagnostics of some diseases like cancer or viral infections. We focus on different types of biorecognition and some technical modifications, lowering the limit of detection potentially attractive to medical practitioners. EXPERT OPINION: Among the new diagnostic strategies, the use of biosensors is of increasing interest. In these techniques, the capture ligand interacts with the analyte of interest. Measuring interactions between partners in real time by surface plasmon resonance yields valuable information about kinetics and affinity in a short time and without labels. Importantly, the tendency in such techniques is to make biosensor devices smaller and the test results apparent with the naked eye, so they can be used in point-of-care medicine.

3.
Molecules ; 25(19)2020 Sep 24.
Artigo em Inglês | MEDLINE | ID: mdl-32987777

RESUMO

For the first time, we are introducing TTPBgp12 and TFPgp17 as new members of the tail tubular proteins B (TTPB) and tail fiber proteins (TFP) family, respectively. These proteins originate from Yersinia enterocolitica phage φYeO3-12. It was originally thought that these were structural proteins. However, our results show that they also inhibit bacterial growth and biofilm formation. According to the bioinformatic analysis, TTPBgp12 is functionally and structurally similar to the TTP of Enterobacteria phage T7 and adopts a ß-structure. TFPgp17 contains an intramolecular chaperone domain at its C-terminal end. The N-terminus of TFPgp17 is similar to other representatives of the TFP family. Interestingly, the predicted 3D structure of TFPgp17 is similar to other bacterial S-layer proteins. Based on the thermal unfolding experiment, TTPBgp12 seems to be a two-domain protein that aggregates in the presence of sugars such as maltose and N-acetylglucosamine (GlcNAc). These sugars cause two unfolding events to transition into one global event. TFPgp17 is a one-domain protein. Maltose and GlcNAc decrease the aggregation temperature of TFPgp17, while the presence of N-acetylgalactosamine (GalNAc) increases the temperature of its aggregation. The thermal unfolding analysis of the concentration gradient of TTPBgp12 and TFPgp17 indicates that with decreasing concentrations, both proteins increase in stability. However, a decrease in the protein concentration also causes an increase in its aggregation, for both TTPBgp12 and TFPgp17.


Assuntos
Caudovirales , Proteínas Estruturais Virais , Yersinia enterocolitica/virologia , Caudovirales/química , Caudovirales/genética , Caudovirales/metabolismo , Domínios Proteicos , Proteínas Estruturais Virais/química , Proteínas Estruturais Virais/genética , Proteínas Estruturais Virais/metabolismo
4.
Front Microbiol ; 11: 1356, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32636826

RESUMO

We report here the complete genome sequence and characterization of Yersinia bacteriophage vB_YenP_ϕ80-18. ϕ80-18 was isolated in 1991 using a Y. enterocolitica serotype O:8 strain 8081 as a host from a sewage sample in Turku, Finland, and based on its morphological and genomic features is classified as a podovirus. The genome is 42 kb in size and has 325 bp direct terminal repeats characteristic for podoviruses. The genome contains 57 predicted genes, all encoded in the forward strand, of which 29 showed no similarity to any known genes. Phage particle proteome analysis identified altogether 24 phage particle-associated proteins (PPAPs) including those identified as structural proteins such as major capsid, scaffolding and tail component proteins. In addition, also the DNA helicase, DNA ligase, DNA polymerase, 5'-exonuclease, and the lytic glycosylase proteins were identified as PPAPs, suggesting that they might be injected together with the phage genome into the host cell to facilitate the take-over of the host metabolism. The phage-encoded RNA-polymerase and DNA-primase were not among the PPAPs. Promoter search predicted the presence of four phage and eleven host RNA polymerase -specific promoters in the genome, suggesting that early transcription of the phage is host RNA-polymerase dependent and that the phage RNA polymerase takes over later. The phage tolerates pH values between 2 and 12, and is stable at 50°C but is inactivated at 60°C. It grows slowly with a 50 min latent period and has apparently a low burst size. Electron microscopy revealed that the phage has a head diameter of about 60 nm, and a short tail of 20 nm. Whole-genome phylogenetic analysis confirmed that ϕ80-18 belongs to the Autographivirinae subfamily of the Podoviridae family, that it is 93.2% identical to Yersinia phage fHe-Yen3-01. Host range analysis showed that ϕ80-18 can infect in addition to Y. enterocolitica serotype O:8 strains also strains of serotypes O:4, O:4,32, O:20 and O:21, the latter ones representing similar to Y. enterocolitica serotype O:8, the American pathogenic biotype 1B strains. In conclusion, the phage ϕ80-18 is a promising candidate for the biocontrol of the American biotype 1B Y. enterocolitica.

5.
Sci Rep ; 10(1): 4196, 2020 03 06.
Artigo em Inglês | MEDLINE | ID: mdl-32144374

RESUMO

Tail Tubular Protein A (TTPA) was long thought to be strictly a structural protein of environmental bacteriophages. However, our recent work has suggested that some TTPAs have additional functional features and thus are dual-function proteins. This study introduces a new TTPA family member, TTPAgp11, which belongs to Yersinia phage phiYeO3-12. We cloned the gene, expressed it and then purified the phage protein. The protein, including its hydrolytic activity, was characterized. Our enzymatic activity tests showed that TTPAgp11 displayed hydrolytic activity towards Red-starch, suggesting that this enzyme could be classified as part as the α - 1, 4-glucosidase family. Protein folding and aggregation tests indicated that TTPAgp11 is a single-domain protein whose aggregation can be induced by maltose or N-acetylglucosamine. The spatial structure of TTPAgp11 seemed to resemble that of the first reported dual-function TTPA, TTPAgp31, which was isolated from Klebsiella pneumoniae phage 32.


Assuntos
Proteínas Virais/química , Proteínas Virais/metabolismo , Sequência de Aminoácidos , Bacteriófagos/genética , Genoma Viral/genética , Hidrólise , Dados de Sequência Molecular , Proteínas Virais/genética
6.
Sci Rep ; 9(1): 13487, 2019 09 17.
Artigo em Inglês | MEDLINE | ID: mdl-31530875

RESUMO

Pseudomonas aeruginosa is an opportunistic pathogen with a capacity to develop antibiotic resistance, which underlies a larger proportion of hospital-acquired infections and higher morbidity and mortality, compared to other bacterial infections. Effective novel approaches for treatment of infections induced by this pathogen are therefore necessary. Phage therapy represents a promising alternative solution to eradicate antibiotic-resistant pathogens. Here, we investigated phage protein efficacy against multi-drug resistant (MDR) P. aeruginosa PAR21 and PAR50 strains isolated from diabetic foot ulcer patients. The results obtained using spot assay, zymography, spectrophotometry and scanning electron microscopy at low voltage (SEM-LV) indicate that the phage protein, PA-PP, exerts activity against P. aeruginosa PAR50 while having no impact on the PAR21 strain. Using LC-MS-MS/MS and comparative analysis of the peptide molecular mass with the protein sequence database, PA-PP was identified as a member of the serine protease family, a result corroborated by its ability to digest casein. We additionally showed a capacity of PA-PP to digest porin protein on the bacterial outer membrane (OM). Moreover, synergistic activity between PA-PP protein and piperacillin led to higher sensitivity of bacterial cells to this antibiotic. Our collective findings suggest that PA-PP targets porin protein on PAR50 OM, thereby increasing its sensitivity to specific antibiotics. The adverse effects observed on bacterial cells using SEM-LV suggest further roles of this protein that remain to be established.


Assuntos
Antibacterianos/farmacologia , Bacteriófagos/fisiologia , Farmacorresistência Bacteriana Múltipla , Pseudomonas aeruginosa/efeitos dos fármacos , Pseudomonas aeruginosa/virologia , Proteínas Virais/metabolismo , Ativação Enzimática , Humanos , Pseudomonas aeruginosa/ultraestrutura , Proteínas Recombinantes , Análise Espectral
7.
J Mol Graph Model ; 92: 8-16, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31302501

RESUMO

Bacteriophage base tailplate proteins were recently discovered to have hydrolytic activity towards disaccharides. The putative assignment of sugar binding sites was based on known lectin structures and identified residues a.a. 40-120 as the potential binding region for disaccharides [1]. To help verify the prediction, an in silico analysis was performed on the structure of a base tailplate protein gp31 from Klebsiella pneumoniae bacteriophage KP32 (PDB: 5MU4) which shows activity towards maltose but not trehalose [1]. Based on the information, a full surface docking was performed for both sugars which identified 2 regions different than originally predicted. The first region clearly favored maltose during the docking phase while the second one allowed for the energetically-equivalent binding of trehalose. To verify the assignment, a molecular dynamics simulation was performed to assess the stability of the docked substrates. MD simulations suggested that the first site included residues D131, D133, and E134, and was also superior for maltose binding while clearly disfavoring trehalose. Analysis of the putative catalytic mechanism suggested residues D131, D133 and E134 as critical for substrate binding. The residue D133 did participate in a stable substrate binding and was positioned near the scissile bond, potentially making it a catalytic residue. Catalytic residues were most likely D131 and D133, one of the two options proposed by Pyra et al. [1]. A comparison with known hydrolase mechanisms suggested that the enzyme most likely retains configuration during hydrolysis of maltose. The findings are discussed for other bacteriophage proteins regarding their potential specificities and catalytic mechanisms.


Assuntos
Bacteriófagos , Sítios de Ligação , Modelos Moleculares , Açúcares/química , Proteínas Virais/química , Bacteriófagos/metabolismo , Bacteriófagos/ultraestrutura , Catálise , Hidrólise , Maltose/química , Maltose/metabolismo , Conformação Molecular , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , Estrutura Molecular , Ligação Proteica , Especificidade por Substrato , Açúcares/metabolismo , Proteínas Virais/metabolismo
8.
Biosens Bioelectron ; 133: 8-15, 2019 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-30903939

RESUMO

In this work we discussed a label-free biosensing application of long-period gratings (LPGs) optimized in refractive index (RI) sensitivity by deposition of thin tantalum oxide (TaOx) overlays. Comparing to other thin film and materials already applied for maximizing the RI sensitivity, TaOx offers good chemical and mechanical stability during its surface functionalization and other biosensing experiments. It was shown theoretically and experimentally that when RI of the overlay is as high as 2 in IR spectral range, for obtaining LPGs ultrasensitive to RI, the overlay's thickness must be determined with subnanometer precision. In this experiment the TaOx overlays were deposited using Atomic Layer Deposition method that allowed for achieving overlays with exceptionally well-defined thickness and optical properties. The TaOx nano-coated LPGs show RI sensitivity determined for a single resonance exceeding 11,500 nm/RIU in RI range nD= 1.335-1.345 RIU, as expected for label-free biosensing applications. Capability for detection of various in size biological targets, i.e., proteins (avidin) and bacteria (Escherichia coli), with TaOx-coated LPGs was verified using biotin and bacteriophage adhesin as recognition elements, respectively. It has been shown that functionalization process, as well as type of recognition elements and target analyte must be taken into consideration when the LPG sensitivity is optimized. In this work optimized approach made possible detection of small in size biological targets such as proteins with sensitivity reaching 10.21 nm/log(ng/ml).


Assuntos
Avidina/isolamento & purificação , Técnicas Biossensoriais , Escherichia coli/isolamento & purificação , Tecnologia de Fibra Óptica , Nanopartículas , Óxidos/química , Refratometria , Tantálio/química
9.
Mol Neurobiol ; 56(3): 1841-1851, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-29936690

RESUMO

One of the most important scientific discoveries of recent years was the disclosure that the intestinal microflora takes part in bidirectional communication between the gut and the brain. Scientists suggest that human gut microflora may even act as the "second brain" and be responsible for neurodegenerative disorders like Alzheimer's disease (AD). Although human-associated microbial communities are generally stable, they can be altered by common human actions and experiences. Enteric bacteria, commensal, and pathogenic microorganisms, may have a major impact on immune system, brain development, and behavior, as they are able to produce several neurotransmitters and neuromodulators like serotonin, kynurenine, catecholamine, etc., as well as amyloids. However, brain destructive mechanisms, that can lead to dementia and AD, start with the intestinal microbiome dysbiosis, development of local and systemic inflammation, and dysregulation of the gut-brain axis. Increased permeability of the gut epithelial barrier results in invasion of different bacteria, viruses, and their neuroactive products that support neuroinflammatory reactions in the brain. It seems that, inflammatory-infectious hypothesis of AD, with the great role of the gut microbiome, starts to gently push into the shadow the amyloid cascade hypothesis that has dominated for decades. It is strongly postulated that AD may begin in the gut, and is closely related to the imbalance of gut microbiota. This is promising area for therapeutic intervention. Modulation of gut microbiota through personalized diet or beneficial microbiota intervention, alter microbial partners and their products including amyloid protein, will probably become a new treatment for AD.


Assuntos
Doença de Alzheimer/microbiologia , Microbioma Gastrointestinal , Inflamação/microbiologia , Doença de Alzheimer/metabolismo , Animais , Encéfalo/metabolismo , Humanos , Inflamação/metabolismo
10.
Artigo em Inglês | MEDLINE | ID: mdl-30333963

RESUMO

Three Streptococcus agalactiae (group B streptococci, GBS) immunoreactive proteins: enolase (47.4 kDa), inosine 5'-monophosphate dehydrogenase (IMPDH) (53 kDa) and molecular chaperone GroEL (57 kDa) were subjected to investigation. Enolase protein was described in our previous paper, whereas IMPDH and GroEL were presented for the first time. The aim of our paper was to provide mapping of specific epitopes, highly reactive with umbilical cord blood serum. Bioinformatic analyses allowed to select 32 most likely epitopes for enolase, 36 peptides for IMPDH and 41 immunoreactive peptides for molecular chaperone GroEL, which were synthesized by PEPSCAN. Ten peptides: two in enolase, one in IMPDH and seven in molecular chaperone GroEL have been identified as potentially highly selective epitopes that can be used as markers in rapid immunological diagnostic tests or constitute a component of an innovative vaccine against GBS infections.


Assuntos
Chaperonina 60/imunologia , Mapeamento de Epitopos , Epitopos/imunologia , IMP Desidrogenase/imunologia , Fosfopiruvato Hidratase/imunologia , Streptococcus agalactiae/imunologia , Anticorpos Antibacterianos/sangue , Biologia Computacional , Testes Diagnósticos de Rotina/métodos , Imunoensaio/métodos , Infecções Estreptocócicas/diagnóstico
11.
Sensors (Basel) ; 18(9)2018 Sep 02.
Artigo em Inglês | MEDLINE | ID: mdl-30200522

RESUMO

In designing a bacteria biosensor, various issues must be addressed: the specificity of bacteria recognition, the immobilization of biomolecules that act as the bacteria receptor, and the selectivity of sensor surface. The aim of this paper was to examine how the biofunctionalized surface of Ti, Au, and Ru metals reacts in contact with strains of Escherichia coli (E. coli). The focus on metal surfaces results from their future use as electrodes in high frequency biosensors, e.g., resonant circuits or transmission-line sections. First, the surfaces of different metals were chemically functionalized with 3-aminopropyltriethoxysilane (APTES) and glutaraldehyde or with 3-glycidylooxypropyltrimethoxysilane (GPTMS) followed by N-(5-amino-1-carboxypentyl) iminodiacetic acid (AB-NTA) and NiCl2. Secondly, the lipopolysaccharide binding protein (LBP), polyclonal anti-Escherichia coli antibody and bacteriophage protein gp37 were tested as bacteria receptors. The selectivity and specificity have been confirmed by the Enzyme-Linked Immunosorbent Assay (ELISA) and visualized by scanning electron microscopy at low landing energies. We noticed that LBP, polyclonal antibody, and gp37 were successfully immobilized on all studied metals and recognized the E. coli bacteria selectively. However, for the antibody, the highest reactivity was observed when Ti surface was modified, whereas the bacteria binding was comparable between LBP and gp37 on the functionalized Ru surfaces, independent from modification. Thus, all surfaces were biocompatible within the scope of biosensor functionality, with titanium functionalization showing the best performance.


Assuntos
Técnicas Biossensoriais/instrumentação , Técnicas Biossensoriais/métodos , Escherichia coli/isolamento & purificação , Ouro/química , Rutênio/química , Titânio/química , Eletrodos , Ensaio de Imunoadsorção Enzimática
12.
Sci Rep ; 8(1): 10935, 2018 Jul 19.
Artigo em Inglês | MEDLINE | ID: mdl-30026546

RESUMO

The interaction between the T4 bacteriophage gp37 adhesin and the bacterial lipopolysaccharide (LPS) is a well-studied system, however, the affinity and strength of the interaction haven't been analyzed so far. Here, we use atomic force microscopy to determine the strength of the interaction between the adhesin and its receptor, namely LPS taken from a wild strain of E. coli B. As negative controls we used LPSs of E. coli O111:B and Hafnia alvei. To study the interaction an AFM tip modified with the gp37 adhesin was used to scan surfaces of mica covered with one of the three different LPSs. Using the correlation between the surface topography images and the tip-surface interaction we could verify the binding between the specific LPS and the tip in contrast to the very weak interaction between the tip and the non-binding LPSs. Using force spectroscopy we could then measure the binding strength by pulling on the AFM tip until it lifted off from the surface. The force necessary to break the interaction between gp37 and LPS from E. coli B, LPS from E. coli O111:B and LPS from H. alvei were measured to be 70 ± 29 pN, 46 ± 13 pN and 45 ± 14 pN, respectively. The latter values are likely partially due to non-specific interaction between the gp37 and the solid surface, as LPS from E. coli O111:B and LPS from H. alvei have been shown to not bind to gp37, which is confirmed by the low correlation between binding and topography for these samples.


Assuntos
Bactérias/metabolismo , Bacteriófago T4/metabolismo , Lipopolissacarídeos/metabolismo , Proteínas do Envelope Viral/metabolismo , Escherichia coli/metabolismo , Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/metabolismo , Hafnia alvei/metabolismo , Cinética , Lipopolissacarídeos/química , Microscopia de Força Atômica , Polissacarídeos Bacterianos/química , Polissacarídeos Bacterianos/metabolismo , Ligação Proteica , Proteínas do Envelope Viral/química
13.
Front Microbiol ; 9: 125, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29467739

RESUMO

The elongation factor Tu has been identified as one of the most immunoreactive proteins that was recognized by human sera of GBS (group B streptococcus) positive patients. In this paper, we present the polypeptide-specific epitopes of the bacterial protein that are recognized by human antibodies: 28LTAAITTVLARRLP41 (peptide no. 3) and 294GQVLAKPGSINPHTKF309 (peptide no. 21). To determine the shortest amino acid sequence recognized by antibodies, truncation peptide libraries were prepared using the PEPSCAN method. The analysis of immunoreactivity of peptides with sera of GBS positive and negative women revealed that the most immunoreactive sequence was 306HTKF309. Moreover, we observed that this sequence also showed the highest specificity which was based on ratio of reactivity with sera of GBS positive relative to sera of GBS negative patients. Epitope was synthetized on Wang resin with the Fmoc strategy. Our results open the possibility to use 306HTKF309 peptide in diagnostic assays to determine Streptococcus agalactiae infection in humans.

14.
Sci Rep ; 7(1): 18048, 2017 12 22.
Artigo em Inglês | MEDLINE | ID: mdl-29273737

RESUMO

In this paper, the enzymatic activity, substrate specificity and antibiofilm feature of bacteriophage dual-function tail proteins are presented. So far, tail tubular proteins A-TTPAgp31 and TTPAgp44-have been considered as structural proteins of Klebsiella pneumoniae bacteriophages KP32 and KP34, respectively. Our results show that TTPAgp31 is able to hydrolyze maltose as well as Red-starch. The activity of 1 µM of the protein was calculated as 47.6 milli-Units/assay relating to the α-amylase activity. It degrades capsular polysaccharides (cPS), slime polysaccharides (sPS) and lipopolysaccharide (LPS) of K. pneumoniae PCM 2713 and shows antibiofilm reactivity towards S. aureus PCM 519 and E. faecalis PCM 2673. TTPAgp44 hydrolyses trehalose and cPS of E. faecium PCM 1859. TTPAgp44's activity was also observed in the antibiofilm test against P. aeruginosa PCM 2710 and B. subtilis PCM 2021. TTPAgp31 has been identified as α-1,4-glucosidase whereas, TTPAgp44 exhibits trehalase-like activity. Both proteins contain aspartate and glutamate residues in the ß-stranded region which are essential for catalytic activity of glycoside hydrolases. The significant novelty of our results is that for the first time the bacteriophage tubular proteins are described as the unique enzymes displaying no similarity to any known phage hydrolases. They can be used as antibacterial agents directed against bacterial strains producing exopolysaccharides and forming a biofilm.


Assuntos
Bacteriófagos/metabolismo , Klebsiella pneumoniae/virologia , Maltose/metabolismo , Polissacarídeos/metabolismo , Proteínas da Cauda Viral/metabolismo , Biofilmes , Hidrólise
15.
Sci Rep ; 7(1): 2223, 2017 05 22.
Artigo em Inglês | MEDLINE | ID: mdl-28533535

RESUMO

Tail tubular protein A (TTPA) is a structural tail protein of Klebsiella pneumoniae bacteriophage KP32, and is responsible for adhering the bacteriophage to host cells. For the first time, we found that TTPA also exhibits lytic activity towards capsular exopolysaccharide (EPS) of the multiresistant clinical strain of Klebsiella pneumoniae, PCM2713, and thus should be regarded as a dual-function macromolecule that exhibits both structural and enzymatic actions. Here, we present our crystallographic and enzymatic studies of TTPA. TTPA was crystallized and X-ray diffraction data were collected to a resolution of 1.9 Å. In the crystal, TTPA molecules were found to adopt a tetrameric structure with α-helical domains on one side and ß-strands and loops on the other. The novel crystal structure of TTPA resembles those of the bacteriophage T7 tail protein gp11 and gp4 of bacteriophage P22, but TTPA contains an additional antiparallel ß-sheet carrying a lectin-like domain that could be responsible for EPS binding. The enzymatic activity of TTPA may reflect the presence of a peptidoglycan hydrolase domain in the α-helical region (amino acid residues 126 to 173). These novel results provide new insights into the enzymatic mechanism through which TTPA acts on polysaccharides.


Assuntos
Bacteriófagos/metabolismo , Klebsiella pneumoniae/virologia , Proteínas Virais/metabolismo , Sequência de Aminoácidos , Bacteriófagos/genética , Hidrólise , Modelos Moleculares , Ligação Proteica , Conformação Proteica , Multimerização Proteica , Proteínas Virais/química , Proteínas Virais/genética
16.
Postepy Hig Med Dosw (Online) ; 71(0): 78-91, 2017 Feb 14.
Artigo em Inglês | MEDLINE | ID: mdl-28258668

RESUMO

Pseudomonas aeruginosa is an opportunistic pathogen that can cause several acute and chronic infections in humans, and it has become an important cause of nosocomial infections and antibiotic resistance. Biofilm represents an important virulence factor for these bacteria, plays a role in P. aeruginosa infections and avoidance of immune defence mechanisms, and has the ability to protect the bacteria from antibiotics. Alginate, Psl and Pel, three exopolysaccharides, are the main components in biofilm matrix, with many biological functions attributed to them, especially with respect to the protection of the bacterial cell from antibiotics and the immune system. Pseudomonas infections, biofilm formation and development of resistance to antibiotics all require better understanding to achieve the best results using alternative treatment with phage therapy. This review describes the P. aeruginosa pathogenicity and virulence factors with a special focus on the biofilm and its role in infection and resistance to antibiotics and summarizes phage therapy as an alternative approach in treatment of P. aeruginosa infections.


Assuntos
Antibacterianos/farmacologia , Terapia por Fagos/métodos , Pseudomonas aeruginosa/efeitos dos fármacos , Biofilmes/crescimento & desenvolvimento , Infecção Hospitalar/prevenção & controle , Resistência Microbiana a Medicamentos , Humanos , Infecções por Pseudomonas/microbiologia , Pseudomonas aeruginosa/crescimento & desenvolvimento , Fatores de Virulência
17.
Microb Biotechnol ; 10(3): 586-593, 2017 05.
Artigo em Inglês | MEDLINE | ID: mdl-28165193

RESUMO

Here, we compared the abilities of polysaccharides L900/2 and L900/3, which were previously isolated from Lactobacillus rhamnosus LOCK 0900, to modulate the immune response to bystander antigens in a mouse model of ovalbumin (OVA) sensitization. In vivo, both polysaccharides reduced the levels of OVA-specific IgE, IgE-dependent basophil degranulation and IgG2a antibodies, but had no effect on the levels of OVA-specific IgA or IgG1. Interestingly, both polysaccharides triggered recall cellular responses with distinct properties. L900/3 significantly suppressed the OVA-induced upregulations of IL-4, IL-5, IL-10 and IL-13 in re-stimulated spleen cells and mesenteric lymph nodes. Our findings support and expand on our previous in vitro studies by demonstrating that polymer L900/3 might modulate the Th1/Th2 balance and could be a promising candidate molecule for preventing allergic sensitization.


Assuntos
Alérgenos/imunologia , Imunização , Fatores Imunológicos/administração & dosagem , Lactobacillus rhamnosus/química , Polissacarídeos Bacterianos/administração & dosagem , Serpinas/imunologia , Animais , Basófilos/imunologia , Degranulação Celular , Citocinas/biossíntese , Imunoglobulina E/sangue , Imunoglobulina G/sangue , Fatores Imunológicos/isolamento & purificação , Camundongos , Polissacarídeos Bacterianos/isolamento & purificação
18.
Sci Rep ; 6: 37613, 2016 11 21.
Artigo em Inglês | MEDLINE | ID: mdl-27869188

RESUMO

Characteristic changes in the microbiota biostructure and a decreased tolerance to intestinal bacteria have been associated with inflammatory bowel disease (IBD). However, few studies have examined the constituents of the intestinal microbiota, including the surface molecules of the bacteria, in healthy and IBD subsets. Here, we compare the chemical structures and immunomodulatory properties of the exopolysaccharides (EPS) of lactobacilli isolated from mice with induced IBD (IBD "+") versus those of healthy mice (IBD "-"). Classical structural analyses were performed using nuclear magnetic resonance spectroscopy and mass spectrometry. Immunomodulatory properties were assessed by stimulation of dendritic cells derived from mouse bone marrow or human peripheral mononuclear blood cells. Our results revealed that EPS produced by IBD "+" species are structurally different from those isolated from IBD "-". Moreover, the structurally different EPS generate different immune responses by dendritic cells. We speculate that resident strains could, upon gut inflammation, switch to producing EPS with specific motifs that are absent from lactobacilli IBD "-", and/or that bacteria with a particular EPS structure might inhabit the inflamed intestinal mucosa. This study may shed light on the role of EPS in IBD and help the development of a specific probiotic therapy for this disease.


Assuntos
Imunomodulação , Doenças Inflamatórias Intestinais/microbiologia , Intestinos/microbiologia , Lactobacillus/metabolismo , Polissacarídeos Bacterianos/química , Polissacarídeos Bacterianos/isolamento & purificação , Animais , Biomarcadores/metabolismo , Células da Medula Óssea/metabolismo , Diferenciação Celular , Citocinas/biossíntese , Células Dendríticas/metabolismo , Citometria de Fluxo , Humanos , Doenças Inflamatórias Intestinais/induzido quimicamente , Doenças Inflamatórias Intestinais/patologia , Espectroscopia de Ressonância Magnética , Camundongos Endogâmicos C57BL
19.
Front Microbiol ; 7: 1537, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-27746766

RESUMO

The Bifidobacteria show great diversity in the cell surface architecture which may influence the physicochemical properties of the bacterial cell and strain specific properties. The immunomodulatory role of bifidobacteria has been extensively studied, however studies on the immunoreactivity of their protein molecules are very limited. Here, we compared six different methods of protein isolation and purification and we report identification of immunogenic and immunoreactive protein of two human Bifidobacterium longum ssp. longum strains. We evaluated potential immunoreactive properties of proteins employing polyclonal sera obtained from germ free mouse, rabbit and human. The protein yield was isolation method-dependent and the reactivity of proteins detected by SDS-PAGE and Western blotting was heterogeneous and varied between different serum samples. The proteins with the highest immunoreactivity were isolated, purified and have them sequenced. Among the immunoreactive proteins we identified enolase, aspartokinase, pyruvate kinase, DnaK (B. longum ssp. longum CCM 7952) and sugar ABC transporter ATP-binding protein, phosphoglycerate kinase, peptidoglycan synthethase penicillin-binding protein 3, transaldolase, ribosomal proteins and glyceraldehyde 3-phosphate dehydrogenase (B. longum ssp. longum CCDM 372).

20.
Biomed Opt Express ; 7(3): 829-40, 2016 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-27231592

RESUMO

This paper presents a novel application of a highly sensitive sensor based on long-period gratings (LPGs) coated with T4 bacteriophage adhesin for Gram-negative bacteria detection. We show here, that the sensor evidently recognizes Escherichia coli K-12 (PCM2560), whereas in the reference tests - ELISA and BIAcore - the results are questionable. For LPGs sensor the resonant wavelength shift observed for E. coli K-12 was approximately half of that measured for E.coli B (positive control). The BIAcore readings (RU) for E. coli K-12 were at 10% level of the signal obtained for E .coli B. These results confirm the improved sensitivity of the LPGs sensor. Moreover, we also show that application of adhesin may allow for efficient detection of E. coli O111 (PCM418), Klebsiella pneumoniae O1 (PCM1) and Yersinia enterocolitica O1 (PCM1879). The specificity of binding bacteria by the adhesin is discussed and it is determined by a distinct region of lipopolysaccharide receptors and/or by the presence of outer-membrane protein C in an outer membrane of Gram-negative bacteria.

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