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1.
Int J Infect Dis ; 91: 182-187, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31770617

RESUMO

BACKGROUND: Whether T-cell interferon-γ responses to Mycobacterium tuberculosis-specific antigens can be influenced by tuberculosis preventive treatment in a high-endemic country is uncertain. METHODS: In this prospective, open-label, controlled study, 513 individuals with silicosis were randomly selected for TB preventive treatment with rifapentine and isoniazid or for observation. QuantiFERON-TB Gold in-tube (QFT-GIT) assay was used to measure IFN-γ response to M. tuberculosis antigens at baseline (T0) and at 6 (T1) and 33 (T2) months after completion of therapy. RESULTS: A total of 220 subjects were included in the final analysis: 105 and 115 in the prevention and observation arms, respectively. The proportions of QFT-GIT reversion from baseline to T1 were similar in the prevention and observation arms (18.4% vs 12.8%, P=0.566). However, reversion from baseline to T2 was more frequent in the prevention arm than in the observation arm, but the difference was not significant (24.2% vs 6.3%, P=0.881). No significant difference was observed in the quantitative responses of QFT-GIT between the two arms during follow-up at T1 (P=0.648) and T2 (P=0.918). CONCLUSIONS: Preventive tuberculosis treatment has no effect on interferon-γ responses measured by serial QFT-GIT assays in a high tuberculosis-endemic country. CLINICAL TRIALS REGISTRATION: http://www.clinicaltrials.gov NCT02430259.


Assuntos
Antituberculosos/uso terapêutico , Interferon gama/sangue , Tuberculose/prevenção & controle , Antígenos de Bactérias/análise , Antígenos de Bactérias/imunologia , China/epidemiologia , Testes Diagnósticos de Rotina , Doenças Endêmicas/prevenção & controle , Doenças Endêmicas/estatística & dados numéricos , Feminino , Seguimentos , Humanos , Testes de Liberação de Interferon-gama , Isoniazida/uso terapêutico , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Rifampina/análogos & derivados , Rifampina/uso terapêutico , Teste Tuberculínico , Tuberculose/sangue , Tuberculose/epidemiologia , Tuberculose/microbiologia
2.
J Fish Dis ; 43(1): 81-89, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31701546

RESUMO

Mycobacteriosis in cultured fish is a challenge for the aquaculture industry worldwide. Treatment by chemical administration is difficult and no effective vaccine has been developed. Therefore, detection and isolation by early diagnosis are important for prevention of the spread of the disease. In mammals, interferon gamma release assays have been established for detection of tuberculosis; these tests are based on the delayed-type hypersensitivity (DTH) response against culture filtrate protein-10 (CFP-10) and the 6-kDa early secreted antigen target (ESAT-6) of Mycobacterium tuberculosis. On the other hand, little is known about the fish immune response against the ESAT-6 and CFP-10 proteins of mycobacteria, although these responses should find application in the diagnosis of mycobacteriosis in fish. In the present study, we identified ESAT-6 and CFP-10 from Mycobacterium pseudoshottsii and cloned the corresponding genes. Intraperitoneal injection of the corresponding DNA plasmid constructs in ginbuna crucian carp yielded increased expression of the fish interferon-γ1-1-encoding gene (IFN-γ1-1). In contrast, IFN-γ1-1 expression accompanied by DTH response was observed only in the CFP-10-DNA plasmid-injected fish. Furthermore, fish that had been prophylactically injected with CFP-10-DNA plasmid exhibited increased survival of M. pseudoshottsii infection. Taken together, these results suggested that CFP-10 may facilitate diagnosis of mycobacteriosis.


Assuntos
Antígenos de Bactérias/análise , Proteínas de Bactérias/análise , Doenças dos Peixes/diagnóstico , Carpa Dourada , Infecções por Mycobacterium/veterinária , Mycobacterium/fisiologia , Sequência de Aminoácidos , Animais , Antígenos de Bactérias/química , Antígenos de Bactérias/genética , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Doenças dos Peixes/microbiologia , Mycobacterium/química , Mycobacterium/genética , Infecções por Mycobacterium/diagnóstico , Infecções por Mycobacterium/microbiologia , Filogenia , Alinhamento de Sequência
3.
BMC Infect Dis ; 19(1): 1023, 2019 Dec 02.
Artigo em Inglês | MEDLINE | ID: mdl-31791276

RESUMO

BACKGROUND: Staphylococcus aureus carriage is a known risk factor for staphylococcal disease. However, the carriage rates vary by country, demographic group and profession. This study aimed to determine the S. aureus carriage rate in children in Eastern Uganda, and identify S. aureus lineages that cause infection in Uganda. METHODS: Nasopharyngeal samples from 742 healthy children less than 5 years residing in the Iganga/Mayuge Health and Demographic Surveillance Site in Eastern Uganda were processed for isolation of S. aureus. Antibiotic susceptibility testing based on minimum inhibitory concentrations (MICs) was determined by the BD Phoenix™ system. Genotyping was performed by spa and SCCmec typing. RESULTS: The processed samples yielded 144 S. aureus isolates (one per child) therefore, the S. aureus carriage rate in children was 19.4% (144/742). Thirty one percent (45/144) of the isolates were methicillin resistant (MRSA) yielding a carriage rate of 6.1% (45/742). All isolates were susceptible to rifampicin, vancomycin and linezolid. Moreover, all MRSA were susceptible to vancomycin, linezolid and clindamycin. Compared to methicillin susceptible S. aureus (MSSA) isolates (68.8%, 99/144), MRSA isolates were more resistant to non-beta-lactam antimicrobials -trimethoprim/sulfamethoxazole 73.3% (33/45) vs. 27.3% (27/99) [p < 0.0001]; erythromycin 75.6% (34/45) vs. 24.2% (24/99) [p < 0.0001]; chloramphenicol 60% (27/45) vs. 19.2% (19/99) [p < 0.0001]; gentamicin 55.6% (25/45) vs. 25.3% (25/99) [p = 0.0004]; and ciprofloxacin 35.6% (16/45) vs. 2% (2/99) [p < 0.0001]. Furthermore, 42 MRSA (93.3%) were multidrug resistant (MDR) and one exhibited high-level resistance to mupirocin. Overall, 61 MSSA (61.6%) were MDR, including three mupirocin and clindamycin resistant isolates. Seven spa types were detected among MRSA, of which t037 and t064 were predominant and associated with SCCmec types I and IV, respectively. Fourteen spa types were detected in MSSA which consisted mainly of t645 and t4353. CONCLUSIONS: S. aureus carriage rate in healthy children in Eastern Uganda is high and comparable to rates for hospitalized patients in Kampala. The detection of mupirocin resistance is worrying as it could rapidly increase if mupirocin is administered in a low-income setting. S. aureus strains of spa types t064, t037 (MRSA) and t645, t4353 (MSSA) are prevalent and could be responsible for majority of staphylococcal infections in Uganda.


Assuntos
Antígenos de Bactérias/análise , Portador Sadio/epidemiologia , Farmacorresistência Bacteriana , Nariz/microbiologia , Faringe/microbiologia , Infecções Estafilocócicas/epidemiologia , Staphylococcus aureus/isolamento & purificação , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Antígenos de Bactérias/classificação , Antígenos de Bactérias/genética , Portador Sadio/microbiologia , Pré-Escolar , Estudos Transversais , Farmacorresistência Bacteriana/genética , Feminino , Técnicas de Genotipagem/métodos , Humanos , Lactente , Recém-Nascido , Masculino , Staphylococcus aureus Resistente à Meticilina/genética , Staphylococcus aureus Resistente à Meticilina/isolamento & purificação , Testes de Sensibilidade Microbiana , Tipagem Molecular/métodos , Mupirocina/farmacologia , Mupirocina/uso terapêutico , Mucosa Nasal/microbiologia , Vigilância da População/métodos , Infecções Estafilocócicas/tratamento farmacológico , Infecções Estafilocócicas/microbiologia , Staphylococcus aureus/classificação , Staphylococcus aureus/genética , Uganda/epidemiologia
4.
Int. microbiol ; 22(4): 471-478, dic. 2019. graf, tab
Artigo em Inglês | IBECS | ID: ibc-185065

RESUMO

Chlamydia trachomatis is considered as a public health problem due to its high prevalence and increased rates of gynecological disorders. The major outer membrane protein (MOMP) of this bacterium is the most abundant protein in its membrane and has been evaluated not only as a vaccine development candidate but also is used in many diagnostic tests. The MOMP weighs 69 kDa and contains four variable segments (VS 1-4) separated by constant regions. Several research groups have developed recombinant single-variable segments of MOMP expressed in Escherichia coli cytoplasm. But, all variable segments have been used minimally for the diagnosis of a chlamydial infection. In this experiment, the authors obtained the recombinant MOMP of C. trachomatis (rMOMP) in E. coli rMOMP and extracted, purified, and partially characterized it. This was later used to identify anti-Chlamydia trachomatis antibodies in sera of infertile patients by immunodetection assays, enzyme-linked immunosorbent assay (ELISA), and indirect immunofluorescence tests. The ELISA test showed high sensitivity and low specificity of 100 and 58.3%, respectively. The above results obtained were linked to the cross-reactivity of antibodies against C. pneumoniae or C. psittaci. Hence, an evaluation was performed to obtain an optimized test for the diagnosis of C. trachomatis infection


No disponible


Assuntos
Antígenos de Bactérias/análise , Chlamydia trachomatis/isolamento & purificação , Infecções por Chlamydia/diagnóstico , Ensaio de Imunoadsorção Enzimática/métodos , Proteínas Recombinantes/análise , Proteínas da Membrana Bacteriana Externa/metabolismo , Chlamydia trachomatis/genética , Infecções por Chlamydia/microbiologia , Proteínas Recombinantes/genética
5.
PLoS Negl Trop Dis ; 13(11): e0007825, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31756188

RESUMO

BACKGROUND: Enterotoxigenic Escherichia coli (ETEC) cause significant diarrheal morbidity and mortality in children of resource-limited regions, warranting development of effective vaccine strategies. Genetic diversity of the ETEC pathovar has impeded development of broadly protective vaccines centered on the classical canonical antigens, the colonization factors and heat-labile toxin. Two non-canonical ETEC antigens, the EtpA adhesin, and the EatA mucinase are immunogenic in humans and protective in animal models. To foster rational vaccine design that complements existing strategies, we examined the distribution and molecular conservation of these antigens in a diverse population of ETEC isolates. METHODS: Geographically diverse ETEC isolates (n = 1159) were interrogated by PCR, immunoblotting, and/or whole genome sequencing (n = 46) to examine antigen conservation. The most divergent proteins were purified and their core functions assessed in vitro. RESULTS: EatA and EtpA or their coding sequences were present in 57.0% and 51.5% of the ETEC isolates overall, respectively; and were globally dispersed without significant regional differences in antigen distribution. These antigens also exhibited >93% amino acid sequence identity with even the most divergent proteins retaining the core adhesin and mucinase activity assigned to the prototype molecules. CONCLUSIONS: EtpA and EatA are well-conserved molecules in the ETEC pathovar, suggesting that they serve important roles in virulence and that they could be exploited for rational vaccine design.


Assuntos
Antígenos de Bactérias/genética , Escherichia coli Enterotoxigênica/genética , Infecções por Escherichia coli/microbiologia , Proteínas de Escherichia coli/genética , Variação Genética , Glicoproteínas de Membrana/genética , Peptídeo Hidrolases/genética , Antígenos de Bactérias/análise , Escherichia coli Enterotoxigênica/química , Escherichia coli Enterotoxigênica/classificação , Escherichia coli Enterotoxigênica/isolamento & purificação , Infecções por Escherichia coli/imunologia , Proteínas de Escherichia coli/análise , Saúde Global , Humanos , Immunoblotting , Glicoproteínas de Membrana/análise , Peptídeo Hidrolases/análise , Reação em Cadeia da Polimerase , Sequenciamento Completo do Genoma
6.
BMC Infect Dis ; 19(1): 796, 2019 Sep 11.
Artigo em Inglês | MEDLINE | ID: mdl-31510926

RESUMO

BACKGROUND: The number of new rickettsial species are rapidly increasing, and increasing numbers of Rickettsia raoultii (R. raoultii) infection cases have been detected in humans. However, neurological abnormalities caused by R. raoultii are rarely reported, especially in northwestern China. CASE PRESENTATION: A 36-year-old Kazakh shepherd with an attached tick on part temporalis, presented with right eyelid droop, lethargy, fever, headache, fever (38.0-41.0 °C) and erythematous rash. The examination of cerebrospinal fluid (CSF) showed cerebrospinal pressure of 200 mm H2O, leukocyte count of 300.0 × 106/L, adenosine deaminase of 2.15 U/L, and total protein concentration of 0.93 g/L. The diagnosis of R. raoultii infection was confirmed by six genetic markers, and semi-quantified by enzyme-linked immunosorbent assay for rickettsial antigen. The patient gradually recovered after treatment with doxycycline and ceftriaxone. R. raoultii DNA was found both in a tick detached from this patient and in 0.18% (2/1107) of blood samples collected from local shepherds. CONCLUSIONS: This is the first reported case with neurological abnormalities caused by R. raoultii in northwestern China. It is vital to detect rickettsial agents both in blood and CSF for tick bite patients with neurological abnormalities. Public health workers and physicians should pay attention to neurological abnormalities caused by Rickettsia.


Assuntos
Doenças do Sistema Nervoso/diagnóstico , Infecções por Rickettsia/diagnóstico , Rickettsia/metabolismo , Picadas de Carrapatos/diagnóstico , Adenosina Desaminase/líquido cefalorraquidiano , Adulto , Animais , Antígenos de Bactérias/análise , Antígenos de Bactérias/imunologia , Ceftriaxona/uso terapêutico , China , DNA Bacteriano/sangue , Doxiciclina/uso terapêutico , Humanos , Contagem de Leucócitos , Masculino , Doenças do Sistema Nervoso/etiologia , Filogenia , RNA Ribossômico 16S/metabolismo , Rickettsia/classificação , Rickettsia/genética , Infecções por Rickettsia/complicações , Infecções por Rickettsia/tratamento farmacológico , Picadas de Carrapatos/complicações , Carrapatos/genética
7.
PLoS One ; 14(9): e0221903, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31479482

RESUMO

Bacterial leaf scorch, caused by Xylella fastidiosa, is a major threat to blueberry production in the southeastern United States. Management of this devastating disease is challenging and often requires early detection of the pathogen to reduce major loss. There are several different molecular and serological detection methods available to identify the pathogen. Knowing the efficiency and suitability of these detection techniques for application in both field and laboratory conditions is important when selecting the appropriate detection tool. Here, we compared the efficiency and the functionality of four different molecular detection techniques (PCR, real-time PCR, LAMP and AmplifyRP® Acceler8™) and one serological detection technique (DAS-ELISA). The most sensitive method was found to be real-time PCR with the detection limit of 25 fg of DNA molecules per reaction (≈9 genome copies), followed by LAMP at 250 fg per reaction (≈90 copies), AmplifyRP® Acceler8™ at 1 pg per reaction (≈350 copies), conventional PCR with nearly 1.25 pg per reaction (≈ 440 copies) and DAS-ELISA with 1x105 cfu/mL of Xylella fastidiosa. Validation between assays with 10 experimental samples gave consistent results beyond the variation of the detection limit. Considering robustness, portability, and cost, LAMP and AmplifyRP® Acceler8™ were not only the fastest methods but also portable to the field and didn't require any skilled labor to carry out. Among those two, AmplifyRP® Acceler8™ was faster but more expensive and less sensitive than LAMP. On the other hand, real-time PCR was the most sensitive assay and required comparatively lesser time than C-PCR and DAS-ELISA, which were the least sensitive assays in this study, but all three assays are not portable and needed skilled labor to proceed. These findings should enable growers, agents, and diagnosticians to make informed decisions regarding the selection of an appropriate diagnostic tool for X. fastidiosa on blueberry.


Assuntos
Mirtilos Azuis (Planta)/microbiologia , Doenças das Plantas/microbiologia , Xylella/genética , Xylella/imunologia , Anticorpos Antibacterianos , Antígenos de Bactérias/análise , Técnicas Bacteriológicas/métodos , DNA Bacteriano/análise , DNA Bacteriano/genética , Ensaio de Imunoadsorção Enzimática/métodos , Técnicas Genéticas , Limite de Detecção , Técnicas de Amplificação de Ácido Nucleico/métodos , Reação em Cadeia da Polimerase/métodos , Reação em Cadeia da Polimerase em Tempo Real/métodos , Xylella/isolamento & purificação
8.
Int J Biol Macromol ; 140: 842-850, 2019 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-31470050

RESUMO

Bacillus anthracis, the causative agent of anthrax, is a harmful pathogen with potential ability as a biological weapon which persuades scientists to develop novel methods to detect anthrax from infected resources. In this study, a multi-walled carbon nanotube (MWCNTs)-based fluorescence aptasensor was fabricated to detect the recombinant protective antigen domain 4 (rPAD4) of Bacillus anthracis as the most important key factor in development of anthrax. First, PAD4 was recombinant expressed in E. coli and purified by Ni-NTA column. Second, the affinity of aptamer to rPAD4 was confirmed by ELAA assay. In aptasensor design, the aptamer was labeled with Gel Green and immobilized on MWCNTs. Upon the adsorption of labeled aptamer on MWCNTs, fluorescence emission was quenched. In contrast, by adding rPAD4 to hybridization reaction and incubation for 10 min, the fluorescence emission was significantly recovered to 85% compared to the control. Detection limit for the sensitivity and specificity of the aptasensor was determined 20 ng/ml and 62.5 ng/ml purified and unpurified rPAD4 protein, respectively. Also, applicability of aptasensor was showed in mouse serum sample. Finally, results indicated that nanosensor has the potential to be developed as a high-sensitive, cost-effective and fast-acting system for measuring of PA in anthrax diagnostic tests.


Assuntos
Antígenos de Bactérias/análise , Aptâmeros de Nucleotídeos , Toxinas Bacterianas/análise , Técnicas Biossensoriais , DNA de Cadeia Simples/química , Fluorescência , Nanotubos de Carbono/química , Nanotubos de Carbono/ultraestrutura , Proteínas Recombinantes de Fusão
9.
Int J Infect Dis ; 86: 18-24, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31269455

RESUMO

BACKGROUND: The early identification of Mycobacterium tuberculosis infection can prevent tuberculosis (TB) transmission. A skin test with a tuberculosis recombinant allergen (Diaskintest) is a new method for identification that has been implemented in Russia. This study was performed to compare the performances of Diaskintest and QuantiFERON-TB Gold (QFT) in adults and children with suspected TB in Moscow, Russia. METHODS: Adults (n=85) and children (n=96) were tested using Diaskintest and QFT. Concordance and comparative analyses were performed. RESULTS: Diaskintest and QFT were concordant in 84% of adults and 90% of children (overall concordance 87%, κ>0.6, Kc>0.5). The concordance between QFT, Diaskintest, and the final diagnosis was good in adults (86% and 81%, respectively) and moderate in children (77% and 79%, respectively). In adults, QFT had a higher sensitivity for detecting TB than Diaskintest (82% and 68%, respectively); in children, Diaskintest was more sensitive (73% and 65%, respectively). In patients with a confirmed TB diagnosis, negative Diaskintest/QFT results were associated with low disease activity. Combined Diaskintest/QFT results identified TB patients with higher sensitivity and specificity than each test separately. CONCLUSIONS: Diaskintest is a low-cost diagnostic tool that shows a test positivity rate similar to QFT and can be used in combination with QFT as an adjunctive test for TB diagnosis.


Assuntos
Antígenos de Bactérias/análise , Testes Diagnósticos de Rotina/métodos , Teste Tuberculínico/métodos , Tuberculose Pulmonar/diagnóstico , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Criança , Pré-Escolar , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Federação Russa , Sensibilidade e Especificidade , Tuberculose Pulmonar/microbiologia , Adulto Jovem
10.
Braz J Microbiol ; 50(4): 979-984, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31352634

RESUMO

Control of brucellosis as a worldwide zoonotic disease is based on vaccination of animals and diagnosis of infected cases to be eradicated. Accurate and rapid detection of infected animals is of critical importance for preventing the spread of disease. Current detection of brucellosis is based on whole-cell antigens and investigating serum antibodies against Brucella lipopolysaccharide (LPS). The critical disadvantage is misdiagnosis of vaccinated animals as infected ones and also cross-reactions with other Gram-negative bacteria. Recombinant outer membrane protein 2b (Omp2b) of Brucella abortus was evaluated as a novel serodiagnostic target in comparison to conventional tests which are based on LPS. Recombinant Omp2b (rOmp2b) was expressed in Escherichia coli BL21 and purified by Ni2+-based chromatography. rOmp2b was evaluated in an indirect enzyme-linked immunosorbent assay (ELISA) system for diagnosis of brucellosis, with sera from Brucella-infected mice along with negative sera and sera from mice which were inoculated with other Gram-negative species for assurance of specificity. Thereafter, cattle sera collected from different regions were assessed along with known negative and known positive serum samples. We found that Omp2b can discriminate between Brucella-infected animals and non-infected ones. Results for assessment of two hundred and fifty cattle sera by Omp2b-based indirect ELISA which were compared to Rose Bengal plate agglutination test (RBPT) and serum tube agglutination test (SAT) showed that our proposed procedure has the sensitivity of 88.5%, specificity of 100%, and accuracy of 90.8%. We suggest that recombinant Omp2b could be used as a protein antigen for diagnosis of brucellosis in domestic animals and can be evaluated for detection of human brucellosis.


Assuntos
Antígenos de Bactérias/análise , Proteínas de Bactérias/análise , Brucella abortus/isolamento & purificação , Brucelose/veterinária , Doenças dos Bovinos/diagnóstico , Ensaio de Imunoadsorção Enzimática/métodos , Porinas/análise , Testes Sorológicos/métodos , Animais , Antígenos de Bactérias/genética , Antígenos de Bactérias/imunologia , Proteínas de Bactérias/genética , Proteínas de Bactérias/imunologia , Brucella abortus/genética , Brucella abortus/imunologia , Brucelose/diagnóstico , Brucelose/microbiologia , Bovinos , Doenças dos Bovinos/microbiologia , Feminino , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Porinas/genética , Porinas/imunologia
11.
BMC Fam Pract ; 20(1): 75, 2019 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-31153357

RESUMO

BACKGROUND: The national guideline for sore throat, endorsed by the Danish Society of General Medicine, recommends the use of the modified Centor score and streptococcal rapid antigen detection test to guide diagnosis and treatment of sore throat. The aim was to investigate Danish general practitioners (GPs) routine management of sore throat patients with a focus on the modalities used and adherence to the guideline. METHODS: A cross-sectional study. GPs in the Central Denmark Region answered an online questionnaire in October 2017. The main outcome measure was modalities used in the management of sore throat patients. RESULTS: In total, 266 of 500 (53%) GPs answered the survey. Ten percent of participants were adherent or almost adherent to the guideline, while 82% of GPs added one or more extra modalities (general clinical assessment (67%), biochemical parameters (48%), and throat swabs for bacterial culture (18%)) to differentiate viral and bacterial etiology. Sixty-five percent of participants used the Centor Score or modified Centor Score, 96% of GPs used a streptococcal rapid antigen detection test, and all GPs chose narrow-spectrum penicillin as the first-line antibiotic. The most common reasons for non-adherence to the guideline were greater confidence in the clinical assessment (39%), time pressure (33%), and difficulty recalling the guideline (19%). CONCLUSION: Danish GPs rarely adhere to the recommended sore throat management guideline, but use various combinations of different modalities in the assessment of bacterial infection. This practice may increase antibiotic prescription rates.


Assuntos
Clínicos Gerais , Fidelidade a Diretrizes , Faringite/diagnóstico , Padrões de Prática Médica , Infecções Estreptocócicas/diagnóstico , Adulto , Idoso , Antibacterianos/uso terapêutico , Antígenos de Bactérias/análise , Estudos Transversais , Dinamarca , Feminino , Medicina Geral , Humanos , Masculino , Pessoa de Meia-Idade , Penicilinas/uso terapêutico , Faringite/terapia , Guias de Prática Clínica como Assunto , Infecções Estreptocócicas/tratamento farmacológico
12.
BMC Infect Dis ; 19(1): 495, 2019 Jun 04.
Artigo em Inglês | MEDLINE | ID: mdl-31164085

RESUMO

BACKGROUND: There is currently no research on the diagnostic value of metagenomic next-generation sequencing (mNGS) for a single pathogens in CSF. The aim of this study was to analyse the value of mNGS for identifying Streptococcus pneumoniae (S. pneumoniae) in paediatric bacterial meningitis. METHODS: Bacterial meningitis (BM) cases from October 23, 2014, to December 31, 2016, and December 1, 2017, to July 31, 2018 at Beijing Children's Hospital were reviewed. Clinical features and pathogens were analysed. RESULTS: We diagnosed 135 patients with BM in this study. A total of 43 S. pneumoniae were identified by combination methods. 26/135 (19.3%) patients had positive results in S. pneumoniae by blood and/or cerebrospinal fluid (CSF) culture. Alere BinaxNow®Streptococcus pneumoniae Antigen test was positive in 35/135(25.9%) cases. 32/135 (23.7%) S. pneumoniae were identified by mNGS. Six CSF samples were identified as S. pneumoniae only by mNGS technology. Taking culture as the gold standard, the sensitivity and specificity of mNGS for diagnosing S. pneumoniae meningitis were 73.1 and 88.1%, respectively. The positive predictive value (PPV) and negative predictive value (NPV) of diagnosing S. pneumoniae meningitis by mNGS were 59.4 and 93.2%, respectively. When comparison between mNGS and combined tests (culture and Alere BinaxNow®Streptococcus pneumoniae Antigen test), the sensitivity and specificity of mNGS for S. pneumoniae identification were 70.3 and 93.9%, the PPV and NPV in the identification of S. pneumoniae by mNGS were 81.4 and 89.3%, respectively. The difference in number of unique reads of S. pneumoniaein from CSF sample (< 14 days onset) and CSF sample (> 14 days from onset) was statistically significant (170.5 VS. 13, P = 0.019). The difference in the collected time of CSF for culture and mNGS was statistically significant (4 days VS. 14 days, P < 0.001). CONCLUSIONS: mNGS has high sensitivity and specificity for S. pneumoniae identification. The pathogen load (number of unique reads) of S. pneumonia is related to the CSF collection time. mNGS was less affected than culture by the use of antibiotics before CSF collection.


Assuntos
Sequenciamento de Nucleotídeos em Larga Escala , Meningites Bacterianas/diagnóstico , Metagenômica/métodos , Streptococcus pneumoniae/genética , Streptococcus pneumoniae/isolamento & purificação , Adolescente , Fatores Etários , Antígenos de Bactérias/análise , Antígenos de Bactérias/sangue , Antígenos de Bactérias/líquido cefalorraquidiano , Antígenos de Bactérias/genética , Criança , Pré-Escolar , Testes Diagnósticos de Rotina , Feminino , Humanos , Lactente , Recém-Nascido , Masculino , Meningites Bacterianas/sangue , Meningites Bacterianas/líquido cefalorraquidiano , Meningites Bacterianas/microbiologia , Pediatria/métodos , Reação em Cadeia da Polimerase/métodos , Valor Preditivo dos Testes , Sensibilidade e Especificidade
13.
Ticks Tick Borne Dis ; 10(5): 981-986, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31109835

RESUMO

Capybaras (Hydrochoerus hydrochaeris Linnaeus, 1766) (Rodentia: Caviidae) are important hosts of Amblyomma ticks (Acari: Ixodidae), which in turn can transmit rickettsiae to humans and animals. However, there is a scarcity of studies about the tick fauna and rickettsial infection in the Amazon region. The present study evaluated rickettsial infection in capybaras and ticks in different areas of the municipality of Rio Branco, state of Acre, in the Western Brazilian Amazon, where rickettsiosis has never been reported. Blood sera from 43 capybaras from four localities in Rio Branco were tested by indirect immunofluorescence assay using Rickettsia rickettsii antigens. Ticks were collected from capybaras and from vegetation as well. Ticks were taxonomically identified to the species level and some of them were tested by PCR, targeting a fragment of the rickettsial gltA gene. Additionally, ticks were tested for bacteria from the genus Borrelia and family Anaplasmatacae. All capybaras submitted to the serological examination were considered non-reactive to R. rickettsii. A total of 410 ticks were collected directly from the capybaras. Amblyomma dubitatum Neumann, 1899 was the most abundant species (82.4%), followed by Amblyomma naponense (Packard, 1869) (14.3%), Amblyomma humerale Koch, 1844 (0.7%), Amblyomma pacae Aragão, 1911 (0.4%), Amblyomma rotundatum Koch 1844 (0.2%) and Amblyomma sp. (1.7%). From the environment 262 ticks were collected: Rhipicephalus microplus (Canestrini, 1888) (88.9%), Dermacentor nitens Neumann, 1897 (9.9%), Amblyomma varium Koch, 1844 (0.7%) and A. rotundatum (0.3%). With the exception of A. humerale, A. rotundatum and R. microplus, all other species are reported here for the first time in the state. Some of the ticks sampled (N = 317) were tested by molecular methods for infection by Rickettsia spp. Rickettsia bellii was identified infecting A. dubitatum and A. rotundatum, while Rickettsia amblyommatis only was found infecting A. humerale and Rickettsia sp. strain Tapirapé was found in A. naponense. This is the first detection of R. bellii and Rickettsia sp. strain Tapirapé in Acre. No Borrelia or Anaplasmataceae were found in the tested ticks. These results add relevant knowledge about the Rickettsia spp. and the acarological fauna in the region of the Western Amazon, and are essential for the maintenance of vigilance about possible pathogens that occur in the state and determination of the risks that they pose to humans and animals that inhabit the region.


Assuntos
Doenças dos Roedores/epidemiologia , Roedores , Rickettsiose do Grupo da Febre Maculosa/veterinária , Infestações por Carrapato/veterinária , Carrapatos/microbiologia , Carrapatos/fisiologia , Anaplasmataceae/isolamento & purificação , Animais , Antígenos de Bactérias/análise , Borrelia/isolamento & purificação , Brasil/epidemiologia , Feminino , Larva/crescimento & desenvolvimento , Larva/microbiologia , Masculino , Ninfa/crescimento & desenvolvimento , Ninfa/microbiologia , Rickettsia/isolamento & purificação , Rickettsia rickettsii/isolamento & purificação , Doenças dos Roedores/microbiologia , Doenças dos Roedores/parasitologia , Rickettsiose do Grupo da Febre Maculosa/microbiologia , Infestações por Carrapato/epidemiologia , Infestações por Carrapato/parasitologia , Carrapatos/classificação
14.
Artigo em Inglês | MEDLINE | ID: mdl-30937288

RESUMO

The tick-borne rickettsial pathogen, Ehrlichia chaffeensis, causes monocytic ehrlichiosis in people and other vertebrate hosts. Mutational analysis in E. chaffeensis genome aids in better understanding of its infection and persistence in host cells and in the development of attenuated vaccines. Our recent RNA deep sequencing study revealed that three genomic mutations caused global changes in the gene expression patterns, which in turn affect the ability of pathogen's survival in a host and the host's ability to induce protection against the pathogen. In this follow-up study, we document the impact of mutations on the pathogen's global protein expression and the influence of protein abundance on a mutant's attenuation and protection of vertebrate host against infection. iTRAQ labeling and mass spectrometry analysis of E. chaffeensis wildtype and mutants identified 564 proteins covering about 63% of the genome. Mutation in ECH_0379 gene encoding for an antiporter protein, causing attenuated growth in vertebrate hosts, led to overexpression of p28 outer membrane proteins, molecular chaperons, and metabolic enzymes, while a mutation downstream to the ECH_0490 gene that caused minimal impact on the pathogen's in vivo growth resulted in major changes in the expression of outer membrane proteins, transcriptional regulators and T4SS proteins. ECH_0660 gene mutation, causing the pathogen's rapid clearance and offering protection against wild type infection challenge in a vertebrate host, had a minimal impact on proteome similar to our prior observations from transcriptome analysis. While the global proteome data revealed fewer translated proteins compared to the transcripts identified from RNA deep sequencing analysis, there is a great deal of correlation noted between the global proteome and transcriptome analysis. Further, global proteome analysis, including the assessment of 2D resolved total and immunoproteomes revealed greater variations in the highly immunogenic p28-Omp proteins.


Assuntos
Antígenos de Bactérias/análise , Ehrlichia chaffeensis/crescimento & desenvolvimento , Ehrlichia chaffeensis/genética , Mutação , Proteoma/análise , Animais , Linhagem Celular , Cães , Perfilação da Expressão Gênica , Virulência
15.
Zhonghua Jie He He Hu Xi Za Zhi ; 42(4): 262-267, 2019 Apr 12.
Artigo em Chinês | MEDLINE | ID: mdl-30955283

RESUMO

Objective: The aim of this study was to determine the performance of the ratio of tuberculosis-specific antigen (TBAg) to phytohemagglutinin (PHA) (TBAg/PHA ratio) in T-SPOT assay in the diagnosis of active tuberculosis (ATB). Methods: Between January 2014 and January 2017, 378 Mycobacterium tuberculosis (MTB) culture positive patients (268 cases of pulmonary tuberculosis, 110 extra-pulmonary tuberculosis) and 824 healthy individuals were recruited from Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology. T-SPOT assay was performed and TBAg/PHA ratio was calculated in all the participants. To validate the study, another group of 223 MTB culture positive TB patients with positive T-SPOT results were recruited from Guangzhou Chest Hospital between January 2017 and December 2017. This was a retrospective case-control study and differences between groups were analyzed using the Mann-Whitney U-test. Results: Of the 378 culture positive ATB patients, 344 patients had positive T-SPOT results. Of the 824 healthy individuals, 204 individuals had positive T-SPOT results. Using healthy individuals as the control group, the sensitivity and specificity of T-SPOT assay in the diagnosis of ATB were 91.0% (344/378) and 75.2% (620/824). Directly using T-SPOT results had a limited accuracy in distinguishing ATB from latent tuberculosis infection (LTBI). The area under the receiver operating characteristic (ROC) curve was between 0.7 and 0.8. However, a further calculation of the TBAg/PHA ratio showed a better performance than TBAg in distinguishing these two conditions, and the area under the ROC curve was 0.881 (95% CI: 0.853-0.909). If using the threshold value of 0.234, the sensitivity and specificity of the TBAg/PHA ratio in distinguishing ATB from LTBI were 69.5% (239/344) and 94.12% (192/204). The validation data showed that the performance of the TBAg/PHA ratio in distinguishing ATB from LTBI was also satisfactory, and the area under the ROC curve was 0.901 (95% CI: 0.872-0.931). Furthermore, the TBAg/PHA ratio had an important role in the diagnosis of extra-pulmonary tuberculosis. If using the threshold value of 0.234, the sensitivity and specificity of the TBAg/PHA ratio in the diagnosis of extra-pulmonary tuberculosis were 79.2% (76/96) and 94.1% (192/204). The area under the ROC curve was 0.932 (95% CI: 0.897-0.967). Conclusions: The TBAg/PHA ratio in T-SPOT assay was better than directly using T-SPOT results in distinguishing ATB from LTBI. This ratio also showed a potential use in the diagnosis of extra-pulmonary tuberculosis.


Assuntos
Antígenos de Bactérias/análise , Tuberculose Latente/diagnóstico , Mycobacterium tuberculosis/isolamento & purificação , Mycobacterium/imunologia , Fito-Hemaglutininas/análise , Tuberculose/diagnóstico , Estudos de Casos e Controles , Feminino , Humanos , Interferon gama , Tuberculose Latente/microbiologia , Masculino , Curva ROC , Estudos Retrospectivos , Sensibilidade e Especificidade , Tuberculose/microbiologia
16.
Curr Microbiol ; 76(6): 698-705, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30955044

RESUMO

Group A streptococcus (GAS) is an important human pathogen whose clinical isolates differ in their ability to produce hydrogen peroxide (H2O2). H2O2 is primarily produced by the enzyme lactate oxidase (LctO), an in depth in silico research revealed that all genome-sequenced GAS possess the required gene lctO. The importance of lctO for GAS is underlined by its highly conserved catabolite control element (cre box) as well as its perfect promotor sequence in comparison to the known consensus sequences of the Gram-positive model organism Bacillus subtilis. In this study, we provide further insight in the function and regulation of lactate oxidase by analyzing a large group of clinical GAS isolates. We found that H2O2 production increased over time in the late stationary phase; after 4 days of incubation, 5.4% of the isolates showed a positive result at 37 °C, while the rate increased to 16.4% at 20 °C. This correlation between H2O2 production and low temperatures suggests additional regulatory mechanisms for lctO besides catabolite control protein A (CcpA) and indicates that lctO might play a role for GAS energy metabolism at sub-body temperatures. Furthermore, we could identify that H2O2 production was different among clinical isolates; we could correlate H2O2 production to emm-types, indicating that emm-types 6 and 75 had the highest rate of H2O2 production. The emm-type- and temperature-dependent H2O2 production of clinical GAS isolates might contribute to their different survival strategies.


Assuntos
Antígenos de Bactérias/análise , Proteínas da Membrana Bacteriana Externa/análise , Proteínas de Transporte/análise , Peróxido de Hidrogênio/metabolismo , Oxigenases de Função Mista/genética , Oxigenases de Função Mista/metabolismo , Oxidantes/metabolismo , Streptococcus pyogenes/metabolismo , Streptococcus pyogenes/efeitos da radiação , Proteínas de Bactérias/metabolismo , Metabolismo Energético , Regulação Bacteriana da Expressão Gênica , Humanos , Proteínas Repressoras/metabolismo , Infecções Estreptocócicas/microbiologia , Streptococcus pyogenes/classificação , Streptococcus pyogenes/genética , Temperatura
17.
Methods Mol Biol ; 1969: 205-215, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30877679

RESUMO

Meningococcal Antigen Typing System (MATS) is the combination of a sandwich ELISA (Enzyme Linked Immunosorbent Assay) developed to estimate the level of expression and immunoreactivity of the antigen components (fHbp, NHBA, and NadA) of the 4CMenB vaccine (Bexsero, GSK Vaccines) in circulating, serogroup B meningococcal (MenB) strains, with the molecular typing of PorA, the main antigenic component in the outer membrane vesicles (OMV). MATS has been proven to be a good surrogate of the accepted correlate of protection for meningococcus (hSBA), thus providing a quick, conservative and reproducible method to assess vaccine coverage. The method has been successfully transferred and standardized in several public health laboratories across Europe, North America, and Australia and used to screen thousands of isolates all over the world. Here we describe the sandwich ELISA method applied to assess the expression and cross-reactivity of fHbp, NHBA, and NadA in MenB isolates.


Assuntos
Antígenos de Bactérias/análise , Ensaio de Imunoadsorção Enzimática/métodos , Infecções Meningocócicas/prevenção & controle , Vacinas Meningocócicas/imunologia , Neisseria meningitidis Sorogrupo B/imunologia , Antígenos de Bactérias/imunologia , Austrália/epidemiologia , Reações Cruzadas , Europa (Continente)/epidemiologia , Humanos , Infecções Meningocócicas/epidemiologia , Infecções Meningocócicas/imunologia , Infecções Meningocócicas/microbiologia , Vacinas Meningocócicas/administração & dosagem , América do Norte/epidemiologia , Vacinação
18.
Clin Lab ; 65(3)2019 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-30868844

RESUMO

BACKGROUND: Primary hepatic tuberculosis is a very rare clinical form of tuberculosis, with atypical clinical presentations and nonspecific imaging features. This results in great difficulty to reach a correct and timely clinical diagnosis. Traditionally, liver biopsy is the gold standard for its diagnosis. Here we assessed the effectiveness of using a T-SPOT.TB test in the early diagnosis of primary hepatic tuberculosis. METHODS: We report a case of primary hepatic tuberculosis whose location of hepatic lesion renders it hard to perform a biopsy. Instead, a T-SPOT.TB test was utilized to help in the early diagnosis of this rare form of tuberculosis. A conventional fourdrug regimen for anti-tubercular therapy together with vitamin B6 was initiated and maintained for 6 months. RESULTS: The T-SPOT.TB test was highly positive for ESAT-6 (87 > 20) and CFP-10 (89 > 20). Dull pain in the upper right abdomen was gone 2 months post treatment. The abnormal lesions shown in an MRI reduced significantly 4 months post treatment. Spot count for ESAT-6 and CFP-10 decreased 6 months post treatment. CONCLUSIONS: The results of this study suggest the critical role of T-SPOT.TB test in the earlier diagnosis of prima¬ry hepatic tuberculosis for those patients who have difficulties having a hepatic biopsy.


Assuntos
Tuberculose Hepática/diagnóstico , Antígenos de Bactérias/análise , Proteínas de Bactérias/análise , Feminino , Humanos , Adulto Jovem
19.
J Nanobiotechnology ; 17(1): 37, 2019 Mar 06.
Artigo em Inglês | MEDLINE | ID: mdl-30841927

RESUMO

BACKGROUND: Magnetosomes (also called bacterial magnetic nanoparticles; BMPs) are biomembrane-coated nanoparticles synthesized by magnetotactic bacteria (MTB). Engineered BMPs fused to protein A (termed ∆F-BMP-FA) bind antibodies (Abs) automatically, and thus provide a series of potential advantages. However, no report so far has systematically evaluated functional applicability of genetically engineered BMPs. RESULTS: We evaluated properties of ∆F-BMP-FA, and developed/optimized culture methods for host strain Magnetospirillum gryphiswaldense ΔF-FA, ∆F-BMP-FA extraction conditions, conditions for Ab conjugation to ∆F-BMP-FA surface, and procedures for antigen detection using ∆F-BMP-FA/Ab complexes (termed BMP-A-Ab). Fed-batch culture for 36 h in a 42-L fermentor resulted in yields (dry weight) of 2.26 g/L for strain ΔF-FA and 62 mg/L for ∆F-BMP-FA. Optimal wash cycle number for ∆F-BMP-FA purification was seven, with magnetic separation following each ultrasonication step. Fusion of protein A to BMPs resulted in ordered arrangement of Abs on BMP surface. Linkage rate 962 µg Ab per mg ∆F-BMP-FA was achieved. BMP-A-Ab were tested for detection of pathogen (Vibrio parahaemolyticus; Vp) surface antigen and hapten (gentamicin sulfate). Maximal Vp capture rate for BMP-A-Ab was 90% (higher than rate for commercial immunomagnetic beads), and detection sensitivity was 5 CFU/mL. ∆F-BMP-FA also bound Abs from crude mouse ascites to form complex. Lowest gentamicin sulfate detection line for BMP-A-Ab was 0.01 ng/mL, 400-fold lower than that for double Ab sandwich ELISA, and gentamicin sulfate recovery rate for BMP-A-Ab was 93.2%. CONCLUSION: Our findings indicate that engineered BMPs such as ∆F-BMP-FA are inexpensive, eco-friendly alternatives to commercial immunomagnetic beads for detection or diagnostic immunoassays, and have high Ab-conjugation and antigen-adsorption capacity.


Assuntos
Nanopartículas de Magnetita/química , Magnetossomos/química , Magnetospirillum/química , Proteína Estafilocócica A/química , Animais , Anticorpos/química , Antígenos de Bactérias/análise , Reatores Biológicos , Ensaio de Imunoadsorção Enzimática , Gentamicinas/análise , Haptenos/análise , Limite de Detecção , Camundongos , Engenharia de Proteínas , Propriedades de Superfície , Vibrio parahaemolyticus/isolamento & purificação
20.
Anticancer Res ; 39(3): 1091-1104, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30842138

RESUMO

BACKGROUND/AIM: Several clinical conditions seriously hamper the diagnostic accuracy of the commonly used tests for Helicobacter pylori (Hp), 13C-urea breath test (UBT) and stool antigen test (SAT). The present communication is a critical review of the potential limitations of UBT and SAT, and describes the approach on how these can be avoided. Drawbacks of the Hp tests: False-negative results are most often due to low bacterial load in the stomach due to: i) use of proton pump inhibitor medication; ii) use of antibiotics; iii) presence of atrophic gastritis and hypoacid stomach; iv); bleeding peptic ulcer; v) gastric cancer (GC) and vi) mucosal-associated lymphatic tissue lymphoma. The UBT also gives false-positive results when urease-producing bacterial species, other than Hp colonize an acid-free stomach. Importantly, neither UBT nor SAT are capable of diagnosing atrophic gastritis, thus missing the patients at highest risk for GC. GastroPanel® (Biohit Oyj, Finland) circumvents these shortcomings with a serological test consisting of a panel of stomach-specific biomarkers: pepsinogen I, pepsinogen II, gastrin-17 and Hp antibodies. GastroPanel® is a tool for non-invasive examination of i) dyspeptic patients for exclusion or diagnosis of Hp or atrophic gastritis, also disclosing the status of gastric acid output; ii) for screening of asymptomatic individuals at risk of GC; and iii) for comprehensive diagnosis of Hp infection. GastroSoft® application integrates the biomarker profile with the patient's medical information, accurately classifying the biomarker profiles into eight diagnostic categories. CONCLUSION: Given that Hp is the single most important risk factor of GC, the non-invasive diagnosis and screening of Hp should be based on more accurate and more comprehensive testing than UBT or SAT alone. The GastroPanel® is such test, being completely devoid of the known serious shortcomings of UBT and SAT.


Assuntos
Infecções por Helicobacter/diagnóstico , Antígenos de Bactérias/análise , Bioensaio , Biomarcadores/sangue , Testes Respiratórios , Técnicas de Diagnóstico do Sistema Digestório , Fezes/química , Infecções por Helicobacter/sangue , Humanos , Ureia/metabolismo
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