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1.
Int J Mol Sci ; 22(14)2021 Jul 09.
Artigo em Inglês | MEDLINE | ID: mdl-34299005

RESUMO

Nanoparticles can interact with the complement system and modulate the inflammatory response. The effect of these interactions on the complement activity strongly depends on physicochemical properties of nanoparticles. The interactions of silver nanoparticles with serum proteins (particularly with the complement system components) have the potential to significantly affect the antibacterial activity of serum, with serious implications for human health. The aim of the study was to assess the influence of graphite oxide (GO) nanocomposites (GO, GO-PcZr(Lys)2-Ag, GO-Ag, GO-PcZr(Lys)2) on the antibacterial activity of normal human serum (NHS), serum activity against bacteria isolated from alveoli treated with nanocomposites, and nanocomposite sensitivity of bacteria exposed to serum in vitro (using normal human serum). Additionally, the in vivo cytotoxic effect of the GO compounds was determined with application of a Galleria mellonella larvae model. GO-PcZr(Lys)2, without IR irradiation enhance the antimicrobial efficacy of the human serum. IR irradiation enhances bactericidal activity of serum in the case of the GO-PcZr(Lys)2-Ag sample. Bacteria exposed to nanocomposites become more sensitive to the action of serum. Bacteria exposed to serum become more sensitive to the GO-Ag sample. None of the tested GO nanocomposites displayed a cytotoxicity towards larvae.


Assuntos
Antibacterianos/farmacologia , Anti-Infecciosos/farmacologia , Grafite/química , Nanopartículas Metálicas/química , Nanocompostos/química , Óxidos/química , Soro/efeitos dos fármacos , Animais , Antibacterianos/química , Anti-Infecciosos/química , Sobrevivência Celular/efeitos dos fármacos , Escherichia coli/efeitos dos fármacos , Escherichia coli/crescimento & desenvolvimento , Escherichia coli/efeitos da radiação , Humanos , Raios Infravermelhos , Larva/efeitos dos fármacos , Larva/efeitos da radiação , Lepidópteros/efeitos dos fármacos , Lepidópteros/efeitos da radiação , Nanopartículas Metálicas/administração & dosagem , Nanocompostos/administração & dosagem , Soro/microbiologia , Prata/química
2.
Lett Appl Microbiol ; 73(2): 132-138, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-33844321

RESUMO

The role of mechanical ventilation and catheters in favouring Acinetobacter baumannii infections needs to be better understood. This study evaluated the adherence of 19 isolates of different hospital clusters of A. baumannii to abiotic surfaces and epithelial cells (HEp-2). Of the hydrophobic isolates, 80% adhered to polystyrene, indicating a close relationship between hydrophobicity and adherence. All isolates adhered to epithelial cells to different degrees, and 73·7% showed an aggregated pattern. Analysis of the serum resistance of catheter-tip isolates showed that all were resistant. These worrisome results showed that the high capacity of A. baumannii to adhere to surfaces and survive in human serum could hinder treatment and control of this pathogen.


Assuntos
Acinetobacter baumannii/fisiologia , Aderência Bacteriana , Células Epiteliais/microbiologia , Infecções por Acinetobacter/microbiologia , Antibacterianos/farmacologia , Linhagem Celular , Farmacorresistência Bacteriana Múltipla , Hospitais , Interações Hospedeiro-Patógeno , Humanos , Interações Hidrofóbicas e Hidrofílicas , Poliestirenos/química , Soro/microbiologia
3.
J Microbiol Methods ; 183: 106182, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-33647359

RESUMO

BACKGROUND: Clinical diagnosis of human brucellosis (HB) is often difficult due to non-specific symptoms. Immunological tests have been the most common method used in HB diagnosis, but molecular methods based on quantitative polymerase chain reaction (qPCR) have largely replaced these diagnostic methods. The aim of this study was to validate a HB diagnostic qPCR method; assessing different target Brucella genes, and the influence of biological matrices (serum vs. whole blood) on analytical parameters. MATERIAL AND METHODS: Two target genes, IS711 and bcsp31, for HB molecular diagnosis were evaluated, together with biological matrix type (whole blood and serum) using samples spiked with Brucella abortus. In addition, diagnostic parameters of this qPCR method were evaluated in paired whole blood and serum samples from patients with suspected HB. RESULTS: Both genes could be potential diagnostic targets, but IS711 showed a lower limit of detection. In spiked matrix experiments, whole blood showed a lower limit of detection than serum after probit regression (224 vs. 3681 CFU/mL) and ANOVA analysis showed a significant (p < 0.001) difference between the Cq of whole blood at all dilutions and that of serum. In 12 paired clinical samples, no serum samples and only one whole blood sample tested positive for Brucella using this qPCR detection method. CONCLUSIONS: This standardized qPCR-based Brucella detection method could improve diagnosis of HB, serving as a rapid, highly sensitive, and specific test. Whole blood is better suited to qPCR-based HB diagnosis due to the presence of higher target DNA loads in this matrix, compared to serum.


Assuntos
Proteínas de Bactérias/genética , Sangue/microbiologia , Brucella/isolamento & purificação , Brucelose/microbiologia , Reação em Cadeia da Polimerase/métodos , Soro/microbiologia , Brucella/classificação , Brucella/genética , Brucelose/sangue , Brucelose/diagnóstico , DNA Bacteriano/sangue , DNA Bacteriano/genética , Humanos
4.
Prev Vet Med ; 185: 105199, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-33229064

RESUMO

In New Zealand, a new diagnostic approach for the control of paratuberculosis in mixed aged milking cows has been developed using a combination of ELISA and quantitative fecal PCR (f-qPCR). Our analysis was designed to evaluate performance of these individual tests in infected or infectious mixed aged cows across the prevalence of infection typically encountered on NZ dairy farms and calculate test accuracy when used as a screening test of serological ELISAs for four separate antigens read in parallel followed by a confirmatory quantitative f-qPCR test. Data from a cross-sectional study of 20 moderate prevalence herds was combined with existing data from 2 low and 20 high prevalence herds forming a dataset of 3845 paired serum and fecal samples. Incidence of clinical Johne's disease (JD) was used to classify herds into three prevalence categories. High (≥ 3% annual clinical JD for the last three years), moderate (<3 - 1%) and low (<1% incidence for at least the last five years). Positive tests were declared if> 50 ELISA units and f-qPCR at two cut-points (≥1 × 104 genomes/mL or >1 × 103 genomes/mL). Fixed Bayesian latent class models at both f-qPCR cut-points, accounted for conditional independence and paired conditional dependence. Mixed models at both f-qPCR cut-points, using a different mechanism to account for conditional dependencies between tests were also implemented. Models (24 in number) were constructed using OpenBUGS. The aim was to identify Mycobacterium avium subsp. paratuberculosis (MAP) infected cows that met at least one of two criteria: shedding sufficient MAP in feces to be detected by f-qPCR or mounting a detectable MAP antibody response. The best fit to the data was obtained by modelling pairwise dependencies between tests in a fixed model or by accounting for dependencies in a mixed model at a fecal cut-off of ≥1 × 104 genomes/mL. Test performance differed with prevalence, but models were robust to prior assumptions. For the fixed model, at a prevalence of 0.29 (95 % probability interval (PI) = 0.25-0.33), as a screening plus confirmatory f-qPCR, post-test probability for disease in a positive animal was 0.84 (95 %PI = 0.80-0.88) and 0.16 (95 %PI = 0.15-0.18) for disease in a test negative animal. In low prevalence herds (0.01(95 %PI = 0.00-0.04)) the equivalent figures were 0.84 (95 %PI = 0.08-0.92) and 0.00 (95 %PI = 0.00-0.02). These results suggest this is a useful tool to control JD on dairy farms, particularly in herds with higher levels of infection, where the sampling and testing cost per animal is defrayed across more detected animals.


Assuntos
Doenças dos Bovinos/diagnóstico , Ensaio de Imunoadsorção Enzimática/veterinária , Fezes/microbiologia , Paratuberculose/diagnóstico , Reação em Cadeia da Polimerase/veterinária , Soro/microbiologia , Animais , Teorema de Bayes , Bovinos , Doenças dos Bovinos/epidemiologia , Estudos Transversais , Indústria de Laticínios , Ensaio de Imunoadsorção Enzimática/instrumentação , Feminino , Incidência , Análise de Classes Latentes , Nova Zelândia/epidemiologia , Paratuberculose/epidemiologia , Reação em Cadeia da Polimerase/instrumentação , Prevalência , Sensibilidade e Especificidade
5.
J Vet Diagn Invest ; 32(6): 892-897, 2020 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-32814516

RESUMO

Fetal bovine serum (FBS) used in cell culture may be contaminated with adventitious agents, which can affect the production of biologicals and the results of clinical laboratory tests. We carried out a retrospective study to determine the incidence of adventitious agent contamination of Argentinean irradiated FBS dating from 2015 to 2019. We analyzed FBS batches for mycoplasma and adventitious viruses (bovine pestiviruses, bovine adenovirus, bluetongue virus, bovine parainfluenza virus 3, rabies virus, bovine parvovirus, bovine herpesvirus 1, bovine respiratory syncytial virus, and reovirus). Cell passages followed by direct immunofluorescence were carried out to check viability of the mentioned adventitious agents. Also, molecular detection of mycoplasma and pestiviruses was performed on the FBS samples. The presence of neutralizing antibodies against pestiviruses was determined. Molecular analyses indicated that frequencies of mycoplasma and pestiviruses in FBS were 14% and 84%, respectively. All of the batches were seronegative for pestiviral antibodies. After cell passages, all FBS samples were negative for hemadsorbent agents and by immunofluorescence for all of the viral species analyzed; PCR assays were negative for mycoplasma and pestiviruses. Our results demonstrate that, of all adventitious agents tested, local FBS batches only had traces of mycoplasma and pestiviruses; gamma irradiation was effective in inactivating them.


Assuntos
Mycoplasma/isolamento & purificação , Soro/microbiologia , Soro/virologia , Vírus/isolamento & purificação , Animais , Anticorpos Antivirais/isolamento & purificação , Bovinos , Meios de Cultura , Reação em Cadeia da Polimerase/veterinária , Estudos Retrospectivos
6.
Cell Rep ; 32(3): 107927, 2020 07 21.
Artigo em Inglês | MEDLINE | ID: mdl-32698013

RESUMO

Antibiotics halt the growth of bacteria by targeting core, essential physiology that is required for life on standard microbiological media. Many more biochemical and virulence processes, however, are required for bacteria to cause infection in a host. Indeed, chemical inhibitors of the latter processes are overlooked using conventional antibiotic drug discovery approaches. Here, we use human blood serum as an alternative growth medium to explore new targets and compounds. High-throughput screening of genetic and chemical libraries identified compounds targeting biological activities required by Klebsiella pneumoniae to grow in serum, such as nucleobase biosynthesis and iron acquisition, and showed that serum can chemically transform compounds to reveal cryptic antibacterial activity. One of these compounds, ruthenium red, was effective in a rat bloodstream infection model. Our data demonstrate that human serum is an effective tool to find new chemical matter to address the current antibiotic resistance crisis.


Assuntos
Antibacterianos/análise , Antibacterianos/farmacologia , Testes Genéticos , Klebsiella pneumoniae/genética , Soro/microbiologia , Bibliotecas de Moléculas Pequenas/análise , Animais , Antibacterianos/química , Dano ao DNA , Modelos Animais de Doenças , Aprovação de Drogas , Feminino , Humanos , Hidrólise , Indóis/farmacologia , Ferro/metabolismo , Infecções por Klebsiella/sangue , Infecções por Klebsiella/microbiologia , Klebsiella pneumoniae/efeitos dos fármacos , Klebsiella pneumoniae/crescimento & desenvolvimento , Fenótipo , Ratos Wistar , Rutênio Vermelho/farmacologia , Bibliotecas de Moléculas Pequenas/química , Triptofano/biossíntese , Uracila/biossíntese
7.
Infect Immun ; 88(10)2020 09 18.
Artigo em Inglês | MEDLINE | ID: mdl-32719154

RESUMO

Haemophilus influenzae is a Gram-negative bacterium that can be classified into typeable (types a through f) and nontypeable (NTHi) groups. This opportunistic pathogen asymptomatically colonizes the mucosal epithelium of the upper respiratory tract, from where it spreads to other neighboring regions, potentially leading to disease. Infection with NTHi can cause otitis media, sinusitis, conjunctivitis, exacerbations of chronic obstructive pulmonary disease, and pneumonia, but it is increasingly causing invasive disease, including bacteremia and meningitis. Invasive NTHi strains are more resistant to complement-mediated killing. However, the mechanisms of complement resistance have never been studied in large numbers of invasive NTHi strains. In this study, we determined the relationship between binding of IgG or IgM and the bacterial survival in normal human serum for 267 invasive H. influenzae strains from Spain, Portugal, and the Netherlands, of which the majority (200 [75%]) were NTHi. NTHi bacteria opsonized with high levels of IgM had the lowest survival in human serum. IgM binding to the bacterial surface, but not IgG binding, was shown to be associated with complement-mediated killing of NTHi strains. We conclude that evasion of IgM binding by NTHi strains increases survival in blood, thereby potentially contributing to their ability to cause severe invasive diseases.


Assuntos
Proteínas do Sistema Complemento/imunologia , Infecções por Haemophilus/imunologia , Haemophilus influenzae/imunologia , Imunoglobulina M/imunologia , Adulto , Idoso , Ativação do Complemento , Europa (Continente)/epidemiologia , Feminino , Infecções por Haemophilus/epidemiologia , Infecções por Haemophilus/microbiologia , Haemophilus influenzae/isolamento & purificação , Haemophilus influenzae/patogenicidade , Humanos , Evasão da Resposta Imune , Imunoglobulina G/imunologia , Masculino , Viabilidade Microbiana , Pessoa de Meia-Idade , Soro/microbiologia
8.
Crit Care ; 24(1): 312, 2020 06 08.
Artigo em Inglês | MEDLINE | ID: mdl-32513224

RESUMO

BACKGROUND: High serum levels of certain aromatic microbial metabolites (AMM) are associated with severity and mortality in critically ill patients. Omics-based studies suggest gut dysbiosis and reduced microbiome diversity in critical conditions. However, the landscape of gut microbial metabolites is still to be outlined, not to mention the interplay correlation between the metabolome and gut microbiome in critically ill patients. The aim of this study was to analyze the association between serum and fecal levels of AMM and compare them with the composition of gut microbiota in critically ill patients in the acute and chronic stages. METHODS: In this prospective observational pilot study, we analyzed the temporal dynamics of the gut microbiome and the AMM spectrum across two distinct subgroups-acute critical ill (ACI) patients with nosocomial pneumonia and chronically critically ill (CCI) patients (9 subjects each group)-as well as performed comparison with 23 healthy volunteers. The AMM levels for each patient were measured using GC-MS in simultaneously taken serum and fecal samples (SFS). These parameters were compared with 16S rRNA fecal microbiome profiles. RESULTS: The observed proportions of bacterial taxa suggest a significant gut dysbiosis in the ACI and the CCI patients. Stronger imbalance in microbiome composition and dynamics observed in the ACI patients compared to the CCI ones resonates with a higher severity in the former group. The total levels of AMM in serum samples were higher for the ACI patients than for the CCI patients (3.7 (1.4-6.3) and 1.1 (1.0-1.6) µM, respectively; p = 0.0003). The qualitative composition of the SFS was also altered. We discovered significant associations between gut microbial taxa levels and metabolite concentrations in blood serum as well as in feces in each of the ACI and the CCI patients. CONCLUSIONS: Aromatic microbial metabolite profiles in the gut and the serum are interlinked and reflect a disruption of the gut microbial community in critically ill patients.


Assuntos
Estado Terminal , Disbiose/microbiologia , Fezes/microbiologia , Soro/microbiologia , Microbioma Gastrointestinal/imunologia , Microbioma Gastrointestinal/fisiologia , Humanos , Projetos Piloto , Estudos Prospectivos
9.
Infect Immun ; 88(9)2020 08 19.
Artigo em Inglês | MEDLINE | ID: mdl-32571987

RESUMO

Even though both cellular and humoral immunities contribute to host defense, the role played by humoral immunity against the airborne opportunistic fungal pathogen Aspergillus fumigatus has been underexplored. In this study, we aimed at deciphering the role of the complement system, the major humoral immune component, against A. fumigatus Mass spectrometry analysis of the proteins extracted from A. fumigatus conidial (asexual spores and infective propagules) surfaces opsonized with human serum indicated that C3 is the major complement protein involved. Flow cytometry and immunolabeling assays further confirmed C3b (activated C3) deposition on the conidial surfaces. Assays using cell wall components of conidia indicated that the hydrophobin RodAp, ß-(1,3)-glucan (BG) and galactomannan (GM) could efficiently activate C3. Using complement component-depleted sera, we showed that while RodAp activates C3 by the alternative pathway, BG and GM partially follow the classical and lectin pathways, respectively. Opsonization facilitated conidial aggregation and phagocytosis, and complement receptor (CR3 and CR4) blockage on phagocytes significantly inhibited phagocytosis, indicating that the complement system exerts a protective role against conidia by opsonizing them and facilitating their phagocytosis mainly through complement receptors. Conidial opsonization with human bronchoalveolar lavage fluid (BALF) confirmed C3 to be the major complement protein interacting with conidia. Nevertheless, complement C2 and mannose-binding lectin (MBL), the classical and lectin pathway components, respectively, were not identified, indicating that BALF activates the alternative pathway on the conidial surface. Moreover, the cytokine profiles were different upon stimulation of phagocytes with serum- and BALF-opsonized conidia, highlighting the importance of studying interaction of conidia with complement proteins in their biological niche.


Assuntos
Aspergillus fumigatus/imunologia , Líquido da Lavagem Broncoalveolar/imunologia , Complemento C3/imunologia , Polissacarídeos Fúngicos/farmacologia , Macrófagos/efeitos dos fármacos , Soro/imunologia , Esporos Fúngicos/imunologia , Aspergilose/genética , Aspergilose/imunologia , Aspergilose/microbiologia , Aspergillus fumigatus/química , Líquido da Lavagem Broncoalveolar/química , Líquido da Lavagem Broncoalveolar/microbiologia , Parede Celular/química , Parede Celular/imunologia , Ativação do Complemento/efeitos dos fármacos , Complemento C3/genética , Citocinas/biossíntese , Citocinas/imunologia , Polissacarídeos Fúngicos/imunologia , Polissacarídeos Fúngicos/isolamento & purificação , Interações entre Hospedeiro e Microrganismos/imunologia , Humanos , Imunidade Celular , Imunidade Humoral , Integrina alfaXbeta2/genética , Integrina alfaXbeta2/imunologia , Antígeno de Macrófago 1/genética , Antígeno de Macrófago 1/imunologia , Macrófagos/imunologia , Macrófagos/microbiologia , Mananas/imunologia , Mananas/isolamento & purificação , Mananas/farmacologia , Proteínas Opsonizantes/farmacologia , Fagocitose/efeitos dos fármacos , Cultura Primária de Células , Ligação Proteica , Espécies Reativas de Oxigênio , Soro/química , Soro/microbiologia , Esporos Fúngicos/química , beta-Glucanas/imunologia , beta-Glucanas/isolamento & purificação , beta-Glucanas/farmacologia
10.
Sci Rep ; 10(1): 6737, 2020 04 21.
Artigo em Inglês | MEDLINE | ID: mdl-32317653

RESUMO

Bacteriophages are abundant in human biomes and therefore in human clinical samples. Although this is usually not considered, they might interfere with the recovery of bacterial pathogens at two levels: 1) by propagating in the enrichment cultures used to isolate the infectious agent, causing the lysis of the bacterial host and 2) by the detection of bacterial genes inside the phage capsids that mislead the presence of the bacterial pathogen. To unravel these interferences, human samples (n = 271) were analyzed and infectious phages were observed in 11% of blood culture, 28% of serum, 45% of ascitic fluid, 14% of cerebrospinal fluid and 23% of urine samples. The genetic content of phage particles from a pool of urine and ascitic fluid samples corresponded to bacteriophages infecting different bacterial genera. In addition, many bacterial genes packaged in the phage capsids, including antibiotic resistance genes and 16S rRNA genes, were detected in the viromes. Phage interference can be minimized applying a simple procedure that reduced the content of phages up to 3 logs while maintaining the bacterial load. This method reduced the detection of phage genes avoiding the interference with molecular detection of bacteria and reduced the phage propagation in the cultures, enhancing the recovery of bacteria up to 6 logs.


Assuntos
Bactérias/virologia , Inoviridae/classificação , Myoviridae/classificação , Podoviridae/classificação , RNA Ribossômico 16S/genética , Siphoviridae/classificação , Líquido Ascítico/microbiologia , Líquido Ascítico/virologia , Bactérias/classificação , Bactérias/genética , Hemocultura/métodos , Capsídeo/química , Líquido Cefalorraquidiano/microbiologia , Líquido Cefalorraquidiano/virologia , Filtração/métodos , Humanos , Inoviridae/genética , Inoviridae/isolamento & purificação , Lisogenia/fisiologia , Tipagem Molecular/métodos , Myoviridae/genética , Myoviridae/isolamento & purificação , Podoviridae/genética , Podoviridae/isolamento & purificação , Soro/microbiologia , Soro/virologia , Siphoviridae/genética , Siphoviridae/isolamento & purificação , Urina/microbiologia , Urina/virologia
11.
Infect Disord Drug Targets ; 20(5): 672-692, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31433763

RESUMO

INTRODUCTION: In the context of the global threat of antimicrobial resistance (AMR) among bacterial pathogens against conventional bactericidal antibiotics, investigation on complementary/ alternative approaches to manage bacterial infections is warranted. The present study aimed at investigating the anti-pathogenic potential of Phyllanthus emblica seed extract (PESE) against four different pathogenic bacteria. METHODS: Hydroalcoholic extract of P. emblica seeds was tested for its possible in vitro quorummodulatory potential against Chromobacterium violaceum, Serratia marcescens, Pseudomonas aeruginosa, and Staphylococcus aureus through broth dilution assay. In vivo efficacy of PESE was assayed employing Caenorhabditis elegans as the model host for these four pathogens. RESULTS: PESE was found to exert in vitro quorum-modulatory effect on C. violaceum, S. marcescens, P. aeruginosa, and S. aureus at ≥50 µg/mL. This extract could curb the haemolytic activity of all the four test bacteria by 23-65%, inhibit biofilm formation, and was also able to modulate their antibiotic susceptibility (AS) and catalase activity. Susceptibility of P. aeruginosa and S. aureus to lysis by human serum was enhanced under the influence of this extract by 23% and 49%, respectively. Repeated exposure of both these notorious pathogens to PESE did not induce resistance in them. In vivo assay confirmed the anti-virulence effect of this extract in the C. elegans host, wherein the nematode host challenged with the PESE-treated pathogenic bacteria scored better survival. PESE also displayed notable prebiotic potential by promoting the growth of three probiotic strains. CONCLUSION: To the best of our awareness, this is the first report on the quorum-modulatory potential of P. emblica seed extract, validating its anti-infective potential and prebiotic property.


Assuntos
Infecções Bacterianas/tratamento farmacológico , Etanol/química , Phyllanthus emblica/química , Extratos Vegetais/farmacologia , Animais , Biofilmes/efeitos dos fármacos , Caenorhabditis elegans , Chromobacterium/efeitos dos fármacos , Chromobacterium/fisiologia , Modelos Animais de Doenças , Farmacorresistência Bacteriana Múltipla/efeitos dos fármacos , Humanos , Testes de Sensibilidade Microbiana , Extratos Vegetais/química , Pseudomonas aeruginosa/efeitos dos fármacos , Pseudomonas aeruginosa/fisiologia , Percepção de Quorum/efeitos dos fármacos , Sementes/química , Serratia marcescens/efeitos dos fármacos , Serratia marcescens/fisiologia , Soro/efeitos dos fármacos , Soro/microbiologia , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/fisiologia
12.
Microb Pathog ; 137: 103737, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31513895

RESUMO

Mucor circinelloides is an opportunistic human pathogen that is used to study mucormycosis, a rare but lethal infection in susceptible immunosuppressed patients. However, the virulence characteristics of this pathogen have not been fully elucidated. In this study, sporangiospores (spores) produced on YPG medium supplemented with native blood serum increased the virulence of M. circinelloides compared with spores produced on YPG supplemented with denatured blood serum or on YPG alone. The spores produced from YPG supplemented with native blood serum increased nematode death and led to significant increases in interleukin (IL)-6, IL-1ß, macrophage inhibitory protein-2, and tumour necrosis factor α mRNA levels in liver and lung tissues from infected diabetic mice compared with those in tissues from animals infected with spores produced in the presence of YPG supplemented with denatured blood serum or of YPG alone. Moreover, spores produced from cultures supplemented with native blood serum showed increased germination rates and longer hyphae compared with other spores. The spores produced in YPG supplemented with native blood serum also enhanced resistance to stress factors and H2O2 and increased thermotolerance compared with spores produced under other conditions. In addition, spores produced in presence of blood serum increased the ability of the pathogen to survive in the presence of macrophages. Taken together, our results showed that these factors were important features for fungal virulence in humans and suggested that thermolabile components in the blood serum may induce M. circinelloides virulence.


Assuntos
Mucor/patogenicidade , Mucormicose/sangue , Soro/microbiologia , Esporos Fúngicos/metabolismo , Animais , Citocinas/metabolismo , Diabetes Mellitus Experimental , Humanos , Peróxido de Hidrogênio , Hifas/crescimento & desenvolvimento , Interleucina-1beta/metabolismo , Interleucina-6/metabolismo , Pulmão , Macrófagos/microbiologia , Masculino , Camundongos , RNA Mensageiro/metabolismo , Esporos Fúngicos/crescimento & desenvolvimento , Virulência
13.
Microb Pathog ; 130: 100-103, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30844472

RESUMO

The epidemiology and prevalence of Q fever in India is largely unknown. There are very few serologic and molecular reports of Q fever in India and these are old reports. The objective of this study was to investigate, for the first time, the presence of Coxiella burnetii infection in sheep and goat flocks of Jammu province of Jammu and Kashmir, India. A total of 148 milk (110 sheep and 38 goats) samples, 282 sera (170 sheep and 112 goats), and 152 vaginal swabs (123 sheep and 29 goats) were collected from farms with incidences of repeated abortion. The LSI Q fever ruminant serum/milk ELISA kit was used to identify anti-C. burnetii antibodies and nested PCR was employed to detect DNA in vaginal swabs. Overall, 42 (38.2%; 95% CI: 29.2-47.9) sheep and 9 (23.7%; 95% CI: 12.0-40.6) goat milk samples, and 21 (12.4%; 95% CI: 8.0-18.5) sheep and 11 (9.8%; 95% CI: 5.2-17.3) goat sera were ELISA positive. In addition, nine (7.3%; 95% CI: 3.6-13.8) vaginal swabs from sheep tested positive by nested PCR; however, C. burnetii could not be found in any of the vaginal swabs from goat. These results indicate that sheep seem to be a more important reservoir of C. burnetii than goats posing a risk for human infection in this area.


Assuntos
Técnicas Bacteriológicas , Coxiella burnetii/isolamento & purificação , Doenças das Cabras/diagnóstico , Febre Q/veterinária , Doenças dos Ovinos/diagnóstico , Animais , Anticorpos Antibacterianos/sangue , Coxiella burnetii/genética , Coxiella burnetii/imunologia , DNA Bacteriano/análise , Ensaio de Imunoadsorção Enzimática , Feminino , Doenças das Cabras/epidemiologia , Doenças das Cabras/microbiologia , Cabras , Índia/epidemiologia , Leite/microbiologia , Técnicas de Diagnóstico Molecular , Reação em Cadeia da Polimerase , Prevalência , Febre Q/diagnóstico , Febre Q/epidemiologia , Testes Sorológicos , Soro/microbiologia , Ovinos , Doenças dos Ovinos/epidemiologia , Doenças dos Ovinos/microbiologia , Vagina/microbiologia
15.
J Clin Microbiol ; 57(5)2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30842231

RESUMO

The Aspergillus-specific lateral-flow device (AspLFD) test is a newly developed point-of-care diagnostic method for invasive pulmonary aspergillosis. However, evidence of the diagnostic performance of the AspLFD for chronic pulmonary aspergillosis (CPA) is limited. Therefore, we conducted a retrospective study to investigate this in comparison with the galactomannan (GM) ß-d-glucan (BDG) test. Fifty patients with chronic pulmonary aspergillosis and 65 patients with respiratory disease, as a control, were enrolled in this study. The majority of the CPA disease entities were chronic pulmonary aspergillosis (64.0%, n = 32), followed by subacute invasive pulmonary aspergillosis (IPA) (20.0%, n = 10) and simple pulmonary aspergilloma (SPA) (16.0%, n = 8). The sensitivity and specificity of the AspLFD test in serum samples were 62.0% and 67.7%, respectively. The GM test (cutoff index, 1.54) showed a sensitivity of 22% and a specificity of 92.3%, while the sensitivity and specificity of the BDG test (cutoff, 19.3 pg/ml) were 48% and 90.8%, respectively. In bronchoalveolar lavage fluid samples, the AspLFD test showed a sensitivity of 66.7% and a specificity of 69.2%, while those of the GM test (cutoff index, 0.6) were 72.7% and 83.1%, respectively. The Aspergillus precipitating antibody test had 70% sensitivity. Unlike the Aspergillus precipitating antibody test, the AspLFD on serum samples showed similar sensitivity to non-fumigatus Aspergillus species. Patients with false-positive results for the AspLFD on serum samples were of a significantly higher age and had a higher prevalence of cavitary lesions in chest computed tomography than patients with negative results in the control group. Given the results in this study, the performance of the AspLFD using serum was acceptable as a point-of-care test for the diagnosis of CPA.


Assuntos
Líquido da Lavagem Broncoalveolar/microbiologia , Testes Imediatos , Aspergilose Pulmonar/diagnóstico , Soro/microbiologia , Idoso , Idoso de 80 Anos ou mais , Antígenos de Fungos/imunologia , Aspergillus/genética , Ação Capilar , Feminino , Humanos , Aspergilose Pulmonar Invasiva/diagnóstico , Masculino , Mananas/sangue , Mananas/imunologia , Pessoa de Meia-Idade , Aspergilose Pulmonar/sangue , Estudos Retrospectivos , Sensibilidade e Especificidade , Testes Sorológicos/instrumentação , Testes Sorológicos/métodos
16.
Infect Immun ; 87(1)2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30323028

RESUMO

Pseudomonas aeruginosa is an opportunistic pathogen that causes nosocomial pneumonia and infects patients with cystic fibrosis. P. aeruginosa lung infections are difficult to treat due to bacterial resistance to antibiotics, and strains with multidrug resistance are becoming more prevalent. Here, we examined the use of a small host defense peptide, innate defense regulator 1002 (IDR-1002), in an acute P. aeruginosa lung infection in vivo IDR-1002 significantly reduced the bacterial burden in bronchoalveolar lavage fluid (BALF), as well as MCP-1 in BALF and serum, KC in serum, and interleukin 6 (IL-6) in BALF. Transcriptome sequencing (RNA-Seq) was conducted on lungs and whole blood, and the effects of P. aeruginosa, IDR-1002, and the combination of P. aeruginosa and IDR-1002 were evaluated. Differential gene expression analysis showed that P. aeruginosa increased multiple inflammatory and innate immune pathways, as well as affected hemostasis, matrix metalloproteinases, collagen biosynthesis, and various metabolism pathways in the lungs and/or blood. Infected mice treated with IDR-1002 had significant changes in gene expression compared to untreated infected mice, with fewer differentially expressed genes associated with the inflammatory and innate immune responses to microbial infection, and treatment also affected morphogenesis, certain metabolic pathways, and lymphocyte activation. Overall, these results showed that IDR-1002 was effective in treating P. aeruginosa acute lung infections and associated inflammation.


Assuntos
Peptídeos Catiônicos Antimicrobianos/administração & dosagem , Bacteriemia/patologia , Pneumonia/patologia , Infecções por Pseudomonas/patologia , Animais , Bacteriemia/tratamento farmacológico , Carga Bacteriana , Líquido da Lavagem Broncoalveolar/química , Líquido da Lavagem Broncoalveolar/microbiologia , Quimiocina CCL2/análise , Modelos Animais de Doenças , Feminino , Perfilação da Expressão Gênica , Camundongos Endogâmicos C57BL , Pneumonia/tratamento farmacológico , Infecções por Pseudomonas/tratamento farmacológico , Pseudomonas aeruginosa/isolamento & purificação , Soro/química , Soro/microbiologia , Resultado do Tratamento
17.
Small ; 15(3): e1803051, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30358085

RESUMO

Detection and inhibition of bacteria are universally required in clinics and daily life for health care. Developing a dual-functional material is challenging and in demand, engaging advanced applications for both defined bioanalysis and targeted biotoxicity. Herein, magnetic silver nanoshells are designed as a multifunctional platform for the detection and inhibition of bacteria. The optimized magnetic silver nanoshells enable direct laser desorption/ionization mass spectrometry based metabolic analysis of bacteria (≈10 µL-1 ), in complex biofluids. The serum infection process (0-10 h) is monitored by statistics toward clinical classification. Moreover, magnetic silver nanoshells facilitate surface adhesion on bacteria due to nanoscale surface roughness and thus display long-term antibacterial effects. Bacteria metabolism is studied with metabolic biomarkers (e.g., malate and lysine) identified during inhibition, showing cell membrane destruction and dysfunctional protein synthesis mechanisms. This work not only guides the design of material-based approaches for bioanalysis and biotoxicity, but contributes to bacteria-related diagnosis by using specific metabolic biomarkers for sensitive detection and new insights by monitoring metabolomic change of bacteria for antibacterial applications.


Assuntos
Antibacterianos/química , Bactérias , Carga Bacteriana/métodos , Testes de Sensibilidade Microbiana/métodos , Nanoconchas/química , Prata/química , Antibacterianos/síntese química , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Bactérias/citologia , Bactérias/efeitos dos fármacos , Bactérias/isolamento & purificação , Bactérias/metabolismo , Infecções Bacterianas/sangue , Infecções Bacterianas/diagnóstico , Infecções Bacterianas/metabolismo , Escherichia coli/citologia , Escherichia coli/isolamento & purificação , Escherichia coli/metabolismo , Humanos , Metabolômica/métodos , Técnicas Microbiológicas/métodos , Nanoconchas/uso terapêutico , Soro/metabolismo , Soro/microbiologia , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Espectrofotometria/métodos
18.
Anaerobe ; 55: 54-60, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30359695

RESUMO

Dental biofilms are complex ecosystems containing many bacterial species that live in mutualistic relationships. These interactions can profoundly affect the virulence properties of the community. In this study we investigated how the production of gingipains, virulence factors from Porphyromonas gingivalis important in periodontal disease, was affected by other commonly found members of the sub-gingival microbiome. To mimic the subgingival microbiome, multispecies consortia (P. gingivalis, Fusobacterium nucleatum, Actinomyces naeslundii, Streptococus oralis, Streptococcus mitis, Streptococcus gordonii and Streptococcus cristatus, with or without Parvimonas micra) as well as dual species consortia (P. gingivalis with P. micra, S. oralis or F. nucleatum) were constructed and maintained anaerobically in 10% serum for up to seven days. The number of P. gingivalis was determined by plating on Brucella agar and the gingipain specific fluorogenic substrate BikKam-10 was used to investigate gingipain activity. The effect of secreted products from P. micra on gingipain activity was investigated by adding supernatants from P. micra to P. gingivalis cultures. The most prominent secreted proteins in the supernatant were identified using mass spectrometry. P. gingivalis was unable to grow in serum, either alone or in the presence of S. oralis or F. nucleatum. In contrast, with P. micra growth was significantly enhanced and this was associated with an increase in gingipain activity. In the multi-species consortia, the presence of P. micra caused a 13-fold increase in gingipain activity. Exposure of P. gingivalis to supernatants from P. micra for 24 h caused a 3-fold increase in gingipain activity. This effect was reduced by 43% after heat-treatment of the supernatant. Two dimensional gel electrophoresis revealed that several of the most prominent proteins in the P. micra supernatant were glycolytic enzymes. The results from this study suggests that gingipains are produced in response to a P. micra derived signalling molecule that is most likely a protein. This is the first time it has been shown that P. micra can affect P. gingivalis virulence properties. This is likely to be of significance for the development of be of periodontitis since these two microorganisms are often found together in the subgingival biofilm.


Assuntos
Adesinas Bacterianas/metabolismo , Cisteína Endopeptidases/metabolismo , Firmicutes/crescimento & desenvolvimento , Consórcios Microbianos , Interações Microbianas , Porphyromonas gingivalis/metabolismo , Anaerobiose , Carga Bacteriana , Contagem de Colônia Microbiana , Meios de Cultura , Cisteína Endopeptidases Gingipaínas , Humanos , Porphyromonas gingivalis/crescimento & desenvolvimento , Soro/microbiologia , Fatores de Virulência/metabolismo
19.
Future Microbiol ; 14: 33-45, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30539665

RESUMO

The multidrug-resistant opportunistic yeast species of Candida auris, Candida haemulonii, Candida duobushaemulonii and Candida pseudohaemulonii continue to endanger the healthcare settings around the globe. Due to the lack of a specific qPCR assay for detection of these species from clinical samples, we developed a multiplex qPCR assay. Analytical specificity and sensitivity showed 100% specificity and the sensitivity of up to ten genomes of target species with a high value of reproducibility (R2 >0.99). Subsequently, from spiked serum samples, our qPCR specifically could detect up to ten genomes of C. auris and one genome of C. haemulonii, C. duobushaemulonii and C. pseudohaemulonii (R2 >0.98). Lack of cross reaction with the human DNA, a high degree of specificity and sensitivity, showed the potential of our multiplex PCR for direct detection of C. auris and closely related species from serum samples of suspected patients. Future studies are warranted to assure its applicability in clinical settings.


Assuntos
Candida/isolamento & purificação , Candidemia/diagnóstico , Técnicas de Diagnóstico Molecular , Reação em Cadeia da Polimerase Multiplex/métodos , Soro/microbiologia , Candida/genética , Candidemia/sangue , Primers do DNA/genética , DNA Fúngico/sangue , DNA Fúngico/genética , DNA Fúngico/isolamento & purificação , DNA Ribossômico/genética , Humanos , Limite de Detecção , Reprodutibilidade dos Testes , Sensibilidade e Especificidade
20.
Infect Immun ; 87(2)2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30455196

RESUMO

Nontypeable Haemophilus influenzae (NTHi) bacteria express various molecules that contribute to their virulence. The presence of phosphocholine (PCho) on NTHi lipooligosaccharide increases adhesion to epithelial cells and is an advantage for the bacterium, enabling nasopharyngeal colonization, as measured in humans and animal models. However, when PCho is expressed on the lipooligosaccharide, it is also recognized by the acute-phase protein C-reactive protein (CRP) and PCho-specific antibodies, both of which are potent initiators of the classical pathway of complement activation. In this study, we show that blood isolates, which are exposed to CRP and PCho-specific antibodies in the bloodstream, have a higher survival in serum than oropharyngeal isolates, which was associated with a decreased presence of PCho. PCholow strains showed decreased IgM, CRP, and complement C3 deposition, which was associated with increased survival in human serum. Consistent with the case for the PCholow strains, removal of PCho expression by licA gene deletion decreased IgM, CRP, and complement C3 deposition, which increased survival in human serum. Complement-mediated killing of PChohigh strains was mainly dependent on binding of IgM to the bacterial surface. These data support the hypothesis that a PCholow phenotype was selected in blood during invasive disease, which increased resistance to serum killing, mainly due to lowered IgM and CRP binding to the bacterial surface.


Assuntos
Proteína C-Reativa/metabolismo , Adesão Celular/imunologia , Haemophilus influenzae , Imunoglobulina M/metabolismo , Orofaringe/microbiologia , Fosforilcolina/metabolismo , Soro/microbiologia , Idoso , Feminino , Haemophilus influenzae/imunologia , Haemophilus influenzae/metabolismo , Haemophilus influenzae/patogenicidade , Humanos , Masculino , Pessoa de Meia-Idade
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