RESUMO
BACKGROUND: This systematic review evaluates pneumolysin (PLY) as a target for new treatments against pneumococcal infections. Pneumolysin is one of the main virulence factors produced by all types of pneumococci. This toxin (53 kDa) is a highly conserved protein that binds to cholesterol in eukaryotic cells, forming pores that lead to cell destruction. METHODS: The databases consulted were MEDLINE, Web of Science, and Scopus. Articles were independently screened by title, abstract, and full text by two researchers, and using consensus to resolve any disagreements that occurred. Articles in other languages different from English, patents, cases report, notes, chapter books and reviews were excluded. Searches were restricted to the years 2000 to 2021. Methodological quality was evaluated using OHAT framework. RESULTS: Forty-one articles describing the effects of different molecules that inhibit PLY were reviewed. Briefly, the inhibitory molecules found were classified into three main groups: those exerting a direct effect by binding and/or blocking PLY, those acting indirectly by preventing its effects on host cells, and those whose mechanisms are unknown. Although many molecules are proposed as toxin blockers, only some of them, such as antibiotics, peptides, sterols, and statins, have the probability of being implemented as clinical treatment. In contrast, for other molecules, there are limited studies that demonstrate efficacy in animal models with sufficient reliability. DISCUSSION: Most of the studies reviewed has a good level of confidence. However, one of the limitations of this systematic review is the lack of homogeneity of the studies, what prevented to carry out a statistical comparison of the results or meta-analysis. CONCLUSION: A panel of molecules blocking PLY activity are associated with the improvement of the inflammatory process triggered by the pneumococcal infection. Some molecules have already been used in humans for other purposes, so they could be safe for use in patients with pneumococcal infections. These patients might benefit from a second line treatment during the initial stages of the infection preventing acute respiratory distress syndrome and invasive pneumococcal diseases. Additional research using the presented set of compounds might further improve the clinical management of these patients.
Assuntos
Infecções Pneumocócicas , Animais , Humanos , Reprodutibilidade dos Testes , Infecções Pneumocócicas/tratamento farmacológico , Infecções Pneumocócicas/complicações , Streptococcus pneumoniae , Estreptolisinas/metabolismo , Proteínas de Bactérias/metabolismoRESUMO
Western blot analysis is widely used for detecting protein expression, analysis of protein-protein interactions, and searching for new biomarkers. Also, it is a diagnostic tool used for detection of human diseases and microorganism infections.Some Streptococcus pneumoniae proteins are important virulence factors and a few of them are diagnostic markers. Here, we describe the detection of two pneumococcal proteins, pneumolysin and PpmA, in human urine by using monoclonal and polyclonal antibodies.
Assuntos
Infecções Pneumocócicas/metabolismo , Infecções Pneumocócicas/urina , Anticorpos Antibacterianos/metabolismo , Anticorpos Monoclonais/metabolismo , Proteínas de Bactérias/urina , Western Blotting , Humanos , Infecções Pneumocócicas/diagnóstico , Streptococcus pneumoniae/imunologia , Streptococcus pneumoniae/metabolismo , Estreptolisinas/urinaRESUMO
Recombinant antibodies such as Fab and scFv are monovalent and small in size, although their functional affinity can be improved through tag-specific immobilization. In order to find the optimum candidate for oriented immobilization, we generated Fab and scFv fragments derived from an anti-pneumolysin monoclonal antibody PLY-7, with histidine and cysteine residues added in diverse arrangements. Tagged antibody fragments scFv-Cys7-His6, His6-scFv-Cys7, and Fab-Cys7 lost considerable affinity for the antigen; however, Fab-His6, Fab-Cys1, and scFv-His6-Cys1 were able to detect immobilized antigen, revealing that the position and number of histidine and cysteine residues are involved differently in the reactivity of antibody fragments. Random and orientated immobilizations were carried out using conventional polystyrene and commercial surface-pretreated ELISA plates. The best orientation performance was obtained with Fab-Cys1-biotin on streptavidin-coated plates with increased signal levels of 62%, while oriented immobilization of Fab-His6 and scFv-His6-Cys1 on nickel- and maleimide-coated plates failed to improve the ELISA sensitivity.
Assuntos
Fragmentos de Imunoglobulinas/química , Estreptolisinas/imunologia , Proteínas de Bactérias/química , Proteínas de Bactérias/imunologia , Fragmentos de Imunoglobulinas/genética , Fragmentos de Imunoglobulinas/imunologia , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologia , Estreptolisinas/químicaRESUMO
A Streptomyces sp. isolated from a patient who had had breast reconstruction after a mastectomy was identified at the species level by comparative sequence analysis of 16S ribosomal DNA (rDNA) and the hypervariable alpha-region of the 16S rDNA.
Assuntos
Implante Mamário/efeitos adversos , Infecções por Bactérias Gram-Positivas/diagnóstico , Streptomyces/classificação , Streptomyces/genética , Infecção da Ferida Cirúrgica/microbiologia , DNA Bacteriano/química , DNA Bacteriano/genética , DNA Ribossômico/química , DNA Ribossômico/genética , Feminino , Genes de RNAr , Infecções por Bactérias Gram-Positivas/microbiologia , Humanos , Pessoa de Meia-Idade , Dados de Sequência Molecular , Filogenia , RNA Bacteriano/genética , RNA Ribossômico 16S/genética , Análise de Sequência de DNA , Homologia de Sequência do Ácido Nucleico , Silicones , Streptomyces/isolamento & purificaçãoRESUMO
Streptomyces cinereoruber isolated from a patient with severe respiratory failure that could not be identified by culture was correctly identified at the species level by comparative sequence analysis of 16S rDNA and the hypervariable alpha-region of 16S rDNA. Restriction fragment length polymorphism, morphologic, and biochemical characteristics have confirmed their identity.
Assuntos
Infecções por Haemophilus/complicações , Haemophilus influenzae/isolamento & purificação , Doença Pulmonar Obstrutiva Crônica/microbiologia , Streptomyces/isolamento & purificação , Idoso , Antibacterianos/uso terapêutico , Anti-Inflamatórios/uso terapêutico , Técnicas de Tipagem Bacteriana , DNA Bacteriano/química , DNA Bacteriano/genética , DNA Ribossômico/química , DNA Ribossômico/genética , Infecções por Haemophilus/tratamento farmacológico , Haemophilus influenzae/genética , Humanos , Masculino , Polimorfismo de Fragmento de Restrição , Prednisona/uso terapêutico , Doença Pulmonar Obstrutiva Crônica/tratamento farmacológico , RNA Ribossômico 16S/genética , Radiografia Torácica , Streptomyces/genética , Sulfametoxazol/uso terapêutico , Tomografia Computadorizada por Raios X , Trimetoprima/uso terapêuticoRESUMO
A pneumolysin-specific enzyme-linked immunosorbent assay (PLY-ELISA) for the detection of pneumolysin in urine was developed and evaluated in comparison with the commercially available Binax Now Streptococcus pneumoniae test (Binax, Portland, ME) for the diagnosis of pneumococcal infections. Assay sensitivity was evaluated using urine from 108 patients with culture-confirmed pneumococcal infections. In adults, the sensitivity and specificity of the PLY-ELISA were 56.6% and 92.2%, respectively. In children with nasopharyngeal pneumococcal carriage, PLY-ELISA and Binax Now S. pneumoniae test sensitivities were 62.5% and 87.5%, respectively, while specificities were 94.4% and 27.8%, respectively. In children with nonnasopharyngeal pneumococcal carriage, PLY-ELISA and Binax Now S. pneumoniae test sensitivities were 68.7% and 93.7%, respectively, and test specificities were 94.1% and 41.2%, respectively. The persistence of pneumolysin in urine of pneumococcal pneumonia patients decreased significantly after 4 to 6 days of treatment. Our data suggest that combining the high specificity of the PLY-ELISA with the high sensitivity of the Binax Now S. pneumoniae test would enable pneumococcal infections to be accurately diagnosed in children.
Assuntos
Ensaio de Imunoadsorção Enzimática/métodos , Infecções Pneumocócicas/diagnóstico , Estreptolisinas/urina , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Proteínas de Bactérias/urina , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Infecções Pneumocócicas/urina , Sensibilidade e EspecificidadeRESUMO
Orientation of reagents is a key step in the construction of immunosensors. When the immunoreagent is a recombinant protein, this can be achieved by employing hexahistidine tags. The orientation of recombinant histidine-tagged Fab fragments of monoclonal anti-pneumolysin antibodies on gold films is evaluated. Using histidine as a chelator of Ni or employing an anti-polyhistidine antibody for capturing the His6 residue is considered. Measurements are based in the signal of indigo, which comes from the hydrolysis of 3-indoxylphosphate by alkaline phosphatase (AP). The attachment of the enzyme occurs through the interaction of biotin with AP-labelled streptavidin or employing AP-conjugated immunoreagents. In the case of the interaction Ni-histidine, for the study of the self-assembling process a His-tagged and biotinylated protein (His6-GST-B) was employed. General conditions were studied and non-specific adsorption was avoided with the use of 1-hexanethiol. Improvements of the signal compared with the direct adsorption were only achieved by the use of histidine capturing antibodies. With an optimised ratio anti-polyhis:His6-Fab the signal increases approximately a 100%. Precision is adequate and the response is linear with the concentration of pneumolysin between 0.1 and 10 ng/mL.