RESUMO
The current COVID-19 pandemic has made patent the need for rapid and cost-effective diagnostic tests, crucial for future infectious outbreaks. Loop-mediated isothermal amplification (LAMP) is a promising and decentralized alternative to qPCR. In this work we have developed a sensitive, fast, and simple innovative methodology for quantification of SARS-CoV-2 RNA copies, combining reverse-transcription LAMP with electrochemical detection. This is based on the oxidation of phenol red (PR), a visual and electroactive LAMP indicator, which oxidation peak potential (Ep) changes with the progress of the LAMP reaction. Using that Ep shift as analytical signal, a calibration curve was obtained for fragment N1 copies of SARS-CoV2 (which provided better results than N or S fragments), with a potential shift of 16.2 mV per order of magnitude, and a practical limit of detection of 21 copies·µL-1. Moreover, the precision of Ep is excellent (RSD < 2%): 557 ± 5 mV for negative and 602 ± 7 mV for positive (2148 N fragment RNA copies·µL-1·-1) LAMP controls. This methodology has been applied to the analysis of nasopharyngeal swab samples, resulting in total concordance with clinical RT-qPCR results. Advances towards fully decentralization have been achieved by designing and fabricating a small portable heater for isothermal procedures, obtaining comparable results to those from a commercial benchtop thermal cycler.
Assuntos
COVID-19 , SARS-CoV-2 , Humanos , SARS-CoV-2/genética , COVID-19/diagnóstico , RNA Viral/genética , RNA Viral/análise , Fenolsulfonaftaleína , Pandemias , Técnicas de Laboratório Clínico/métodos , Teste para COVID-19 , Técnicas de Amplificação de Ácido Nucleico/métodos , Sensibilidade e EspecificidadeRESUMO
Honey authenticity guarantee is crucial for consumer health and fair-trading commerce. New visual false-positive-free molecular lateral flow strip (LFS), termed 5'-3' exonuclease activity -directed false positive-eradicating PCR assisted lateral flow strip (FPE-PCR-LFS) was developed. This FPE-PCR-LFS explored the availability of using a signal-probe as the mediator to integrate the efficient amplification module with visual LFS module. With the genomic DNA extracted from target honey, the designed signal probe would be hydrolyzed and exhausted by the 5'-3' exonuclease activity of Taq DNA polymerase in the amplification process. The hydrolyzed signal probe would not be recognized and capture on the T line with only C line of LFS, reflecting the authenticity of the tested honey. And as low as 0.5% authenticity can be accurately identified in commercial honey samples. Significantly, the false-positive-interference was successfully eradicated for the final visual results judgement, which would greatly widen the application of molecular PCR-LFS in various fields.
Assuntos
Mel , Ácidos Nucleicos , Reação em Cadeia da Polimerase/métodos , Técnicas de Amplificação de Ácido Nucleico/métodos , Exonucleases , Sensibilidade e EspecificidadeRESUMO
Nucleic acids amplification is a widely used technique utilized for different manipulations with DNA and RNA. Although, polymerase chain reaction (PCR) remains the most popular amplification method, isothermal approaches are gained more attention last decades. Among these, loop-mediated isothermal amplification (LAMP) became an excellent alternative to PCR. LAMP requires an increased number of primers and, therefore, is considered a highly specific amplification reaction compared to PCR. LAMP primers design is still a non-trivial task, and all niceties should be taken into account during their selection. Here, we report on a new program called LAMPrimers iQ destined for high-quality LAMP primers design. LAMPrimers iQ is based on an original algorithm considering rigorous criteria for primers selection. Unlike alternative programs, LAMPrimers iQ can process long DNA or RNA sequences, and completely avoid primers that can form homo- and heterodimers. The quality of the primers designed was checked using SARS-CoV-2 coronavirus RNA as a model target. It was shown that primers selected with LAMPrimers iQ provide higher specificity and reliable detection of viral RNA compared to those obtained by alternative programs. The program is available at https://github.com/Restily/LAMPrimers-iQ.
Assuntos
DNA , Técnicas de Amplificação de Ácido Nucleico , Sensibilidade e Especificidade , Técnicas de Amplificação de Ácido Nucleico/métodos , Software , RNARESUMO
PURPOSE: A new nuclear Overhauser enhancement (NOE)-mediated saturation transfer signal at around -1.6 ppm, termed NOE(-1.6), has been reported at high fields of 7T and 9.4T previously. This study aims to validate the presence of this signal at a relatively low field of 4.7T and evaluate its variations in different brain regions and tumors. METHODS: Rats were injected with monocrystalline iron oxide nanoparticles to reduce the NOE(-1.6) signal. CEST signals were measured using different saturation powers before and after injection to assess the presence of this signal. Multiple-pool Lorentzian fits, with/without inclusion of the NOE(-1.6) pool, were performed on CEST Z-spectra obtained from healthy rat brains and rats with 9L tumors. These fits aimed to further validate the presence of the NOE(-1.6) signal and quantify its amplitude. RESULTS: The NOE(-1.6) signal exhibited a dramatic change following the injection of monocrystalline iron oxide nanoparticles, confirming its presence at 4.7T. The NOE(-1.6) signal reached its peak at a saturation power of â¼0.75 µT, indicating an optimized power level. The multiple-pool Lorentzian fit without the NOE(-1.6) pool showed higher residuals around -1.6 ppm compared to the fit with this pool, further supporting the presence of this signal. The NOE(-1.6) signal did not exhibit significant variation in the corpus callosum and caudate putamen regions, but it showed a significant decrease in tumors, which aligns with previous findings at 9.4T. CONCLUSION: This study successfully demonstrated the presence of the NOE(-1.6) signal at 4.7T, which provides valuable insights into its potential applications at lower field strengths.
Assuntos
Neoplasias Encefálicas , Glioma , Ratos , Animais , Neoplasias Encefálicas/diagnóstico por imagem , Neoplasias Encefálicas/patologia , Glioma/patologia , Imageamento por Ressonância Magnética/métodos , Interpretação de Imagem Assistida por Computador/métodos , Algoritmos , Sensibilidade e Especificidade , Encéfalo/diagnóstico por imagem , Encéfalo/patologiaRESUMO
Accurate and rapid diagnosis of infectious diseases plays a key role in clinical practice, especially in resource-limited countries. In this study, we integrated sunrise-type smart amplification process (s-SmartAmp), a convenient and sensitive isothermal amplification method for nucleic acid, into a portable 3D-printed device equipped with smartphone-assisted image analysis capabilities to develop a novel fluorescence-based sensing system for the on-site diagnosis of tuberculosis (TB). To increase the efficiency of fluorescence (or Förster) resonance energy transfer, two types of sunrise probe systems were compared to detect the IS6110 DNA sequence of TB. Subsequently, linear regression was conducted to compare the performance of s-SmartAmp and loop-mediated isothermal amplification (LAMP). The results indicated that, compared with LAMP, s-SmartAmp yielded more stable and precise results with lower background interference and high linear correlation coefficients (R2 = 0.9994 and 1, respectively) for the FAM-TAMRA and FITC-BHQ-1 probe system. The detection time was 45 min with a detection limit of 10 fg/µL. To evaluate the performance of our proposed on-site sensing system, we used s-SmartAmp 3D-printed ultraviolet light-emitting diode device to test multiple clinical samples of TB. Our findings suggest that the proposed system has the potential to achieve accurate and rapid on-site diagnosis of TB.
Assuntos
Técnicas Biossensoriais , Tuberculose , Humanos , Smartphone , Raios Ultravioleta , Tuberculose/diagnóstico , Técnicas de Amplificação de Ácido Nucleico/métodos , Impressão Tridimensional , Sensibilidade e EspecificidadeRESUMO
Methicillin-resistant Staphylococcus aureus (MRSA), a common zoonotic multidrug-resistant bacterium, puts a great threat to public health and food safety. Rapid and reliable detection of MRSA is crucial to guide effective patient treatment at early stages of infection and control the spread of MRSA infections. Herein, we developed a Simultaneous dual-gene and ulTra-sensitive detection for methicillin-resistant Staphylococcus aureus using Argonaute-DNAzyme tandem Detection (STAND). Simply, loop-mediated isothermal amplification (LAMP) was used for the amplification of the species-specific mecA and nuc gene, followed by STAND enabled by the site-specific cleavage of programable Argonaute. The Argonaute-DNAzyme tandem reaction rendered a conceptually novel signal amplification and transduction module that was more sensitive (1 or 2 order of magnitude higher) than the original Argonaute-based biosensing. With the strategy, the target nucleic acid signals gene were dexterously converted into fluorescent signals. STAND could detect the nuc gene and mecA gene simultaneously in a single reaction with 1 CFU/mL MRSA and a dynamic range from 1 to 108 CFU/mL. This method was confirmed by clinical samples and challenged by identifying contaminated foods and MRSA-infected animals. This work enriches the arsenal of Argonaute-mediated biosensing and presents a novel biosensing strategy to detect pathogenic bacteria with ultra-sensitivity, specificity and on-site capability.
Assuntos
Técnicas Biossensoriais , DNA Catalítico , Staphylococcus aureus Resistente à Meticilina , Infecções Estafilocócicas , Animais , Humanos , Staphylococcus aureus Resistente à Meticilina/genética , Sensibilidade e Especificidade , Inocuidade dos Alimentos , Infecções Estafilocócicas/diagnóstico , Infecções Estafilocócicas/microbiologia , Proteínas de Bactérias/genéticaRESUMO
There are currently several validated HPV tests. However, longitudinal data which spans appropriate age ranges, as well as evaluation of potential screening algorithms are necessary for screening programmes choice of test. The objective of our study was to evaluate the performance of HPV mRNA and HPV DNA testing, including partial genotyping, in routine cervical screening. As part of the CERVIVA HPV Primary Screening Study, ThinPrep samples from 10 150 women were tested for HPV mRNA using the Aptima HPV assay and HPV DNA using the Cobas 4800 HPV test. HPV mRNA-positive women were further assessed with the Aptima genotyping assay for HPV 16/18/45. Baseline cytology and prospective follow-up data were collected. The performance of the two tests was examined over 42 months (to date). HPV mRNA demonstrated equivalent sensitivity to HPV DNA testing for detection of CIN2+ (93.2% [92.4-93.9] vs 92.8% [92.0-93.6], respectively) and CIN3+ (94.6% [93.8-95.3] vs 94.6% [93.8-95.3]). HPV mRNA testing had significantly higher specificity compared to HPV DNA for detection of CIN2+ (84.0% [83.5-84.5] vs 80.8% [80.2-81.4], respectively) and CIN3+ (88.44% [88.2-88.6] vs 85.62 [85.4-85.9]). The proportion of CIN2+ and CIN3+, over 3 years (42 months), in HPV-negative women was comparable for both RNA (0.20% and 0.10%) and DNA (0.22% and 0.11%). Genotyping data was comparable across both assay platforms. In the context of HPV primary screening HPV mRNA testing has potential to reduce triage tests and follow-up tests at 12 months compared to DNA testing, with no significant difference in detection of CIN2+ and CIN3+.
Assuntos
Infecções por Papillomavirus , Displasia do Colo do Útero , Neoplasias do Colo do Útero , Feminino , Humanos , Detecção Precoce de Câncer/métodos , RNA Mensageiro/genética , Infecções por Papillomavirus/diagnóstico , Genótipo , Irlanda/epidemiologia , Papillomavirus Humano 16/genética , Estudos Prospectivos , Papillomavirus Humano 18/genética , DNA , Papillomaviridae/genética , Sensibilidade e Especificidade , DNA Viral/genéticaRESUMO
OBJECTIVE: The aim of the work described here was to evaluate the diagnostic performance of a new integrated strategy using breast ultrasound (US) combined with magnetic resonance imaging (MRI) to differentiate benign and malignant breast non-mass-like lesions (NMLs) detected on US. METHODS: From October 2017 to January 2021, 183 NMLs detected on US that had undergone MRI examinations were included in this respective study. Pathological results were used as the reference standard. The integrated diagnostic strategy of breast US combined with MRI based on a combination of MRI Breast Imaging Reporting and Data System (BI-RADS) with discriminant sonographic indicators highly associated with malignancy was established and validated in a cohort of 61 women. The diagnostic performances of US, MRI and the combined method were calculated and compared. RESULTS: In the training set, the area under the receiver operating characteristic curve (AUC), sensitivity and specificity of US, MRI and the integrated diagnostic strategy using US combined with MRI for NMLs were 0.730, 93.7% and 52.3%; 0.849, 94.7% and 75.0%; and 0.901, 92.6% and 87.5%, respectively. Compared with US or MRI alone, the integrated diagnostic strategy significantly increased the AUC (p < 0.001, p = 0.007) and specificity (p < 0.001, p = 0.034) while maintaining high sensitivity (p = 0.774, p = 0.551). In the validation set, the integrated strategy of US combined with MRI (AUC = 0.899) also had good performance compared with US (AUC = 0.728) or MRI (AUC = 0.838). CONCLUSION: The integrated diagnostic strategy of US combined with MRI exhibited good performance for breast NMLs compared with either modality used alone, which can improve the diagnostic specificity while maintaining high sensitivity.
Assuntos
Neoplasias da Mama , Ultrassonografia Mamária , Feminino , Humanos , Ultrassonografia Mamária/métodos , Ultrassonografia , Mama/diagnóstico por imagem , Sensibilidade e Especificidade , Imageamento por Ressonância Magnética/métodos , Neoplasias da Mama/diagnóstico por imagem , Estudos RetrospectivosRESUMO
This study investigated whether combining International Ovarian Tumor Analysis (IOTA) Simple Rules with tumor biomarkers would improve the diagnostic accuracy for early detection of adnexal malignancies. Receiver operating characteristic curve analysis of suspected adnexal tumors was performed in 226 women admitted for surgery at the University Clinical Center of Kosovo. Primary outcome was the diagnostic accuracy of the combination of adnexal mass biomarkers and IOTA Simple Rules. IOTA Simple Rules combined with biomarker indications increased the diagnostic accuracy of classifying adnexal masses. Data analysis of individual measures showed that ferritin had the lowest rate of sensitivity.
Assuntos
Doenças dos Anexos , Neoplasias Ovarianas , Feminino , Humanos , Sensibilidade e Especificidade , Diagnóstico Diferencial , Ultrassonografia , Neoplasias Ovarianas/diagnóstico , Neoplasias Ovarianas/patologia , Doenças dos Anexos/diagnóstico , Doenças dos Anexos/patologia , BiomarcadoresRESUMO
Metabolome analysis has gained widespread application in disease diagnosis owing to its ability to provide comprehensive information, including disease phenotypes. In this study, we utilized 3D superstructures fabricated through evaporation-induced microprinting to analyze the metabolome for glaucoma diagnosis. 3D superstructures offer the following advantages: high hotspot density per unit volume of the structure extending from two to three dimensions, excellent signal repeatability due to the reproducibility and defect tolerance of 3D printing, and high thermal stability due to the PVP-enclosed capsule form. Leveraging the superior optical properties of the 3D superstructure, we aimed to classify patients with glaucoma. The signal obtained from the 3D superstructure was employed in a Deep Neural Network (DNN) classification model to accurately classify glaucoma patients. The sensitivity and specificity of the model were determined as 92.05% and 93.51%, respectively. Additionally, the fabrication of 3D superstructures can be performed at any stage, significantly reducing measurement time. Furthermore, their thermal stability allows for the analysis of smaller samples. One notable advantage of 3D superstructures is their versatility in accommodating different target materials. Consequently, they can be utilized for a wide range of metabolic analyses and disease diagnoses.
Assuntos
Técnicas Biossensoriais , Glaucoma , Humanos , Reprodutibilidade dos Testes , Glaucoma/diagnóstico , Redes Neurais de Computação , Sensibilidade e EspecificidadeRESUMO
Genetic testing has been increasingly used in several fields. In many applications, nucleic acid amplification technology is required. However, current methods to detect nucleic acid amplification require expensive reagents and special equipment or exhibit limited sensitivity, which hinders their use. To address this issue, this study reports an assay method for detecting occurrence of acid amplification in post-amplification samples using pyrophosphate, a highly sensitive byproduct of nucleic acid amplification. The method proposed requires two reagents and an automated analyzer. First, hydrogen peroxide is derived from pyrophosphate, an indicator of nucleic acid amplification, and the oxidizing power of hydrogen peroxide is used to produce Fe (III) from Fe (II). The specific metal chelator 5-Br-PAPS forms a complex with the trivalent iron produced, resulting in a highly sensitive coloration. The within-run reproducibility of our method (n = 20) was less than 3.67% at each concentration tested, and the detection limit was 0.075 µmol/L, sufficient for quantitative analysis. The technique described could detect pyrophosphate in a sample that was amplified using the loop-mediated isothermal amplification method after only 10 min. Therefore, the proposed method has the potential to be a new, rapid, and simple detection technique for amplified nucleic acids.
Assuntos
Difosfatos , Ácidos Nucleicos , Sensibilidade e Especificidade , Peróxido de Hidrogênio , Reprodutibilidade dos Testes , Técnicas de Amplificação de Ácido Nucleico/métodos , Ácidos Nucleicos/genéticaRESUMO
This study compared reverse transcription-loop-mediated isothermal amplification (RT-LAMP) and three reverse transcription-quantitative polymerase chain reaction (RT-qPCR) assays targeting the N and E genes of the SARS-CoV-2 genome for detecting RNA in untreated wastewater samples. RT-qPCR assays exhibited consistent amplification down to 2 × 102 GC/reaction, with greater analytical sensitivity at 2 × 101 GC/reaction by US CDC N1 and US CDC N2 assays. In contrast, RT-LAMP exhibited lower sensitivity, detecting SARS-CoV-2 only at or above 2 × 103 GC/reaction. For SARS-CoV-2 seeded wastewater samples, the US CDC N1 assay exhibited greater analytical sensitivity than the US CDC N2, E_Sarbeco, and RT-LAMP assays. Out of 30 wastewater samples, RT-qPCR detected endogenous SARS-CoV-2 RNA in 29 samples, while RT-LAMP identified 27 positive samples, with 20 displaying consistent amplifications in all three RT-LAMP technical replicates. Agreement analysis revealed a strong concordance between RT-LAMP and the US CDC N1 and E_Sarbeco RT-qPCR assays (κ = 0.474) but lower agreement with the US CDC N2 RT-qPCR assay (κ = 0.359). Quantification of SARS-CoV-2 RNA in positive samples revealed a strong correlation between the US CDC N1 and E_Sarbeco assays, while the US CDC N1 and US CDC N2 assays exhibited weak correlation. Logistic regression analysis indicated that RT-LAMP results correlated with RNA quantified by the US CDC N1 and E_Sarbeco assays, with 95 % limits of detection of 3.99 and 3.47 log10 GC/15 mL, respectively. In conclusion, despite lower sensitivity compared to RT-qPCR assays, RT-LAMP may offer advantages for wastewater surveillance, such as rapid results (estimated as twice as fast), and simplicity, making it a valuable tool in the shifting landscape of COVID-19 wastewater surveillance. Furthermore, LAMP positive wastewater samples might be prioritized for SARS-CoV-2 sequencing due to reduced analytical sensitivity. These findings support the use of RT-LAMP as a specific and efficient method for screening wastewater samples for SARS-CoV-2, particularly in resource-limited settings.
Assuntos
COVID-19 , SARS-CoV-2 , Humanos , SARS-CoV-2/genética , Águas Residuárias , RNA Viral , Colorimetria , Sensibilidade e Especificidade , Vigilância Epidemiológica Baseada em Águas ResiduáriasRESUMO
OBJECTIVE: Endobronchial ultrasound (EBUS) is commonly used to guide transbronchial needle biopsies for the staging of lymph nodes in non-small cell lung cancer patients. Although contrast-enhanced ultrasound (CEUS) and microbubbles (MBs) can improve the diagnostic accuracy in tumors, the ability of contrast-enhanced EBUS (CE-EBUS) to image MBs has not yet been comprehensively evaluated. In this study, we assessed the ability of a CE-EBUS system (Olympus EU-ME2 PREMIER and BF-UC180F bronchoscope) to detect laboratory-synthesized MBs in comparison to clinical (Toshiba Aplio SSA-790A) and pre-clinical (VisualSonics Vevo 2100) CEUS systems in vitro and in vivo, respectively. METHODS: Agar flow phantoms and reference tissue were used to assess CE-EBUS MB imaging in vitro, and A549 tumor-bearing athymic nude and AE17-OVA tumor-bearing C57BL/6 mice were used to assess MB detectability and perfusion in vivo, respectively. RESULTS: Results revealed that despite the lower sensitivity of CE-EBUS to MB concentration in comparison to clinical CEUS, CE-EBUS yielded a similar contrast-to-tissue ratio (CTR) in vitro of 28.9 ± 4.5 dB for CE-EBUS, compared with 29.7 ± 2.6 dB for clinical CEUS (p < 0.05). In vivo, CE-EBUS generated a perfusion curve highly correlated with that obtained with the pre-clinical CEUS system (Pearson correlation coefficient = 0.927, p < 0.05). Moreover, CE-EBUS yielded a CTR 2.7 times higher than that obtained with the pre-clinical ultrasound system. CONCLUSION: These findings together suggest that CE-EBUS can perform contrast imaging comparable to that produced by commercial pre-clinical and clinical ultrasound systems, with potential for clinical characterization of mediastinal lymph nodes in lung cancer patients.
Assuntos
Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Camundongos , Animais , Humanos , Neoplasias Pulmonares/diagnóstico por imagem , Neoplasias Pulmonares/patologia , Microbolhas , Sensibilidade e Especificidade , Metástase Linfática/patologia , Broncoscopia/métodos , Estadiamento de Neoplasias , Camundongos Endogâmicos C57BL , Linfonodos/patologia , Aspiração por Agulha Fina Guiada por Ultrassom Endoscópico , Estudos RetrospectivosRESUMO
Prostate-specific membrane antigen PET (PSMA-PET) has emerged as a powerful imaging tool for prostate cancer primary staging, biochemical recurrence, and advanced disease assessment. This article offers a concise overview of the benefits and challenges associated with PSMA-PET for prostate cancer evaluation. The article highlights the advantages of PSMA-PET over conventional imaging, such as its higher sensitivity and specificity for detecting metastases, and the potential for guiding personalized treatment decisions. However, it also explores the limitations and potential pitfalls for interpretation. Overall, the article aims to provide valuable insights for clinicians and diagnostic imaging physicians in clinical practice.
Assuntos
Próstata , Neoplasias da Próstata , Masculino , Humanos , Próstata/diagnóstico por imagem , Próstata/patologia , Neoplasias da Próstata/diagnóstico por imagem , Neoplasias da Próstata/patologia , Tomografia por Emissão de Pósitrons combinada à Tomografia Computadorizada/métodos , Tomografia Computadorizada por Raios X , Sensibilidade e Especificidade , Antígeno Prostático EspecíficoRESUMO
Species identification has become a significant concern due to the growing use of food alternatives that may cause allergies and reduce nutritional value. To address the issue of fraudulent adulteration of goat milk products with cow milk, we have developed an affordable, portable, and user-friendly platform called microfluidic-integrated nucleic acid lateral flow strips (LFS). This platform enables simultaneous detection of components derived from both goats and cows in goat milk. In this study, we have introduced an innovative nucleic acid labeling method. The loop primers of loop-mediated isothermal amplification (LAMP) have been modified with amplification terminator spacer C3 and an oligonucleotide sequence, thus eliminating the requirement for costly antibodies in traditional nucleic acid LFS. This modification not only lowers costs but also enables multiple detections. Additionally, we have integrated the LAMP and LFS assay steps into a microfluidic chip, allowing convenient on-site detection while effectively preventing aerosol contamination of LAMP products. The testing process includes rapid DNA extraction, followed by a short nucleic acid addition and incubation for visualized results in about 50 min. This platform is user-friendly, requiring no specialized equipment or extensive training, making it suitable for rapid on-site detection of dairy products by personnel in diverse fields.
Assuntos
Leite , Ácidos Nucleicos , Feminino , Bovinos , Animais , Técnicas de Amplificação de Ácido Nucleico/métodos , Primers do DNA/genética , Oligonucleotídeos , Cabras , Sensibilidade e EspecificidadeRESUMO
The unprecedented pandemic has raised the demand for prompt, precise, and large-scale virus detection techniques to control the transmission of contagious illnesses. In this study, a loop-mediated isothermal amplification (LAMP) based on-chip platform was developed to address this challenge using rotational diffusometry and functionalized Janus particles. A recombinant plasmid containing a cDNA sequence of the SARS-CoV-2 non-structural protein 2 (nsp2) gene was employed here as a proof-of-concept for COVID-19 detection. Specifically, designed primers and the functionalized Janus particles were simultaneously loaded on a microfluidic chip to perform the LAMP reaction on a hot plate. The optimal Janus particle concentrations for diffusometric analysis were thoroughly validated, and the performance of the on-chip LAMP reaction was assessed using thermal image analysis. Utilization of the highly sensitive rotational diffusometry achieved a limit of detection of 1 pg/µL in just 10 min with a sample volume of 20 µL. Our method delivered a tenfold higher sensitivity than the conventional method by utilizing only half of its usual required time. Overall, this study proposes a potential nucleic acid (NA) amplification device to aid the rapid diagnosis of various diseases by modifying the primers for different target genes.
Assuntos
COVID-19 , SARS-CoV-2 , Humanos , SARS-CoV-2/genética , COVID-19/diagnóstico , DNA Complementar , Sensibilidade e Especificidade , Técnicas de Amplificação de Ácido Nucleico/métodos , Teste para COVID-19 , Primers do DNA , RNA Viral/genética , RNA Viral/análiseRESUMO
Background: Echinococcosis is a neglected tropical disease that is severely underdiagnosed in resource-limited settings. In developed countries, diagnosing echinococcosis is challenging, and reliable serological assays are urgently needed. In the Central European Alps, EM is more common than EG; however, data on the diagnostic performance of assays for EM cases are scarce. We evaluated the suitability of nine antibody assays for routine diagnostics. Methods: Nine commercially available serological assays for detecting anti-Echinococcus antibodies were compared head-to-head using samples collected from 50 patients with echinococcosis and 50 age- and sex-matched control subjects. The assays are Anti-Echinococcus ELISA (IgG) (Euroimmun), Echinococcus IgG ELISA (DRG), Echinococcus IgG ELISA (IBL International), Echinococcus Western Blot IgG (LDBIO Diagnostics), EUROLINE WB (Euroimmun), Hydatidosis ELISA IgG (VirCell), Hydatidosis VIRCLIA IgG Monotest (VirCell), Ridascreen Echinococcus IgG (R-Biopharm), and Virapid Hydatidosis (VirCell). The cases were ranked according to the WHO-Informal Working Group on Echinococcosis (WHO-IWGE) criteria as confirmed, probable, or possible. Results: The performance of the assays varied greatly, with overall sensitivities ranging between 50% and 88% and specificities between 62% and 100%. We observed a trend toward better performance with cases classified as "confirmed" using the WHO-IWGE criteria. Combined analysis with sequential screening and confirmatory testing resulted in a maximum sensitivity of 84% and specificity of 100%. Differentiation between EG and EM infections is clinically relevant but was found to be unreliable. Conclusions: Echinococcus serological assays are highly variable in terms of sensitivity and specificity. Knowledge of the pre-test probability in the patient cohort is required to choose a suitable assay. A combined approach with screening and confirmatory assays may be the best diagnostic strategy in many situations.
Assuntos
Equinococose , Echinococcus , Animais , Humanos , Estudos Retrospectivos , Equinococose/diagnóstico , Ensaio de Imunoadsorção Enzimática/métodos , Anticorpos Anti-Helmínticos/análise , Imunoglobulina G , Sensibilidade e EspecificidadeRESUMO
Objetivo En nuestro estudio el objetivo fue investigar la contribución de los hallazgos de la tomografía por emisión de positrones (PET)/tomografía computarizada (TC) en la diferenciación no invasiva entre el derrame pleural (DP) de origen benigno (DPB) y maligno (DPM) en pacientes diagnosticadas de un carcinoma de ovario (CO). Material y método Se incluyeron en el estudio a 32 pacientes diagnosticadas de CO con DP. Se efectuó análisis comparativo entre el DPB y el DPM para los siguientes parámetros: SUVmáx. del DP, índice objeto/fondo (TBRp: target/background ratio) del DP, dividiendo el SUVmáx. del DP por el SUVmedio del flujo sanguíneo mediastínico (MBP: medistinal blood pool), engrosamiento pleural, adenopatías supradiafragmáticas, DP unilateral o bilateral, diámetro del DP, edad de la paciente y niveles de CA125. Resultados La edad media de las 32 pacientes fue de 57±2,8 años. Se observó mayor frecuencia significativa de un TBRp>1,1, engrosamiento pleural y ganglios linfáticos supradiafragmáticos en el DPM respecto al DPB. Aunque no se detectó ningún nódulo pleural en las pacientes con DPB, estos estuvieron presentes en 7 pacientes con DPM. En la distinción DMP-DBP la sensibilidad del TBRp fue del 95,2% y la especificidad del 72,7%, la sensibilidad del engrosamiento pleural fue del 80,9%, y la especificidad del 81,8%, la sensibilidad de los ganglios linfáticos supradiafragmáticos fue del 38% y la especificidad del 90,9%, y la sensibilidad del nódulo pleural fue del 33,3% y la especificidad del 100%. No hubo diferencias significativas entre los 2 grupos respecto al resto de factores. Conclusión El engrosamiento pleural y el valor de TBRp determinados en la PET/TC pueden contribuir a la diferenciación entre DMP y DBP, especialmente en aquellas pacientes con CO en estadio avanzado y mal estado general, o que no son tributarias de ser sometidas a tratamiento quirúrgico (AU)
Objective The present study investigates the ability of non-invasive contribution of positron emission tomography (PET)/computed tomography (CT) to distinguish between benign pleural effusions (BPE) and malignant pleural effusions (MPE) in patients diagnosed with ovarian carcinoma (OC). Material and method Included in the study were 32 OC patients with a PE diagnosis. The cases with BPE and MPE were compared in terms of the PE maximum standardized uptake value (SUVmax), PE SUVmax/mean standardized uptake (SUVmean) value of the mediastinal blood pool (TBRp), the presence of pleural thickening, the presence of supradiaphragmatic lymph node, unilateral or bilateral PE, pleural effusion diameter, patient age and CA125 value. Results The mean age of the 32 patients was 57±2.8 years. TBRp>1.1, pleural thickening and supradiaphragmatic lymph node were observed significantly more frequently in the MPE than the BPE cases. While no pleural nodules were detected in patients with BPE, they were present in 7 of the patients with MPE. The rates of distinction between the MPE and BPE cases were as follows: the sensitivity of the TBRp value was 95.2% and specificity was 72.7%; the sensitivity of pleural thickness was 80.9% and specificity was 81.8%; the sensitivity of supradiaphragmatic lymph node was 38% and specificity was 90.9%; and the sensitivity of the pleural nodule was 33.3% and specificity was 100%. There were no significant differences between two groups in any other factors. Conclusion Pleural thickening and TBRp values ascertained through PET/CT may aid the distinction between MPE-BPE, especially in patients with advanced stage OC with a poor general condition, or those who cannot undergo surgery (AU)
Assuntos
Humanos , Feminino , Pessoa de Meia-Idade , Neoplasias Ovarianas/diagnóstico por imagem , Derrame Pleural Maligno/diagnóstico por imagem , Tomografia por Emissão de Pósitrons combinada à Tomografia Computadorizada , Sensibilidade e Especificidade , Estudos Retrospectivos , Diagnóstico DiferencialRESUMO
Objetivos Comparar la validez discriminante y la fiabilidad interobservador de los 2 métodos de corrección del test del reloj más usados en España. Metodología Se han evaluado 2 colecciones de dibujos del reloj obtenidos en un contexto clínico (116 casos; 56,8% mujeres, edad media 73,1±7,7 años) y en una cohorte de voluntarios (2.039 dibujos de 579 sujetos; 59,5% mujeres, edad media 78,3±3,8 años). Todos los sujetos fueron clasificados como sin deterioro cognitivo (DC−) o con deterioro cognitivo (DC+) tras una extensa evaluación clínica y neuropsicológica. Evaluadores expertos han valorado estos dibujos de forma independiente y sin conocimiento del diagnóstico con los métodos de Sunderland y Solomon estandarizados en español por Cacho (rango: 0 a 10) y del Ser (rango: 0 a 7), respectivamente. Se ha calculado la validez discriminante de cada método mediante el área bajo la curva ROC (aROC) en las 2 muestras, y la fiabilidad interobservador mediante el coeficiente de correlación intraclase (CCI) y el coeficiente kappa en la muestra clínica que fue valorada por los 2 evaluadores. Resultados No hay diferencias significativas en la validez discriminante de los métodos de Sunderland y Solomon en ninguna de las muestras (clínica: aROC: 0,73 [IC 95%: 0,64-0,81] y 0,77 [IC 95%: 0,69-0,85], respectivamente, p=0,19; voluntarios: aROC: 0,69 [IC 95%: 0,67-0,71] y 0,72 [IC 95%: 0,69-0,73], respectivamente, p=0,08). Los puntos de corte ≤8 y ≤5 clasifican correctamente al 71 y 73% de la muestra clínica y al 82 y 84% de la muestra de voluntarios, respectivamente. Los 2 métodos tienen una buena concordancia en la muestra clínica (AU)
Objective To compare the discriminant validity and inter-rater reliability of the two scoring systems for the Clock test that are most used in Spain. Methodology Two collections of clock drawings obtained in a clinical context (116 cases; 56.8% women, mean age 73.1±7.7 years) and in a cohort of volunteers (2039 drawings of 579 subjects; 59.5% women, mean age 78.3±3.8 years) have been assessed. All subjects were classified as cognitively normal (CN) or cognitively impaired (CI) after extensive clinical and neuropsychological evaluation. Expert raters have evaluated these drawings independently and without knowledge of the diagnosis using the Sunderland and Solomon systems standardized in Spanish by Cacho (range 0 to 10) and del Ser (range 0 to 7) respectively. The discriminant validity of each method was calculated in the two samples using the area under the ROC curve (aROC), and the inter-rater reliability was calculated in the clinical sample, that was assessed by the two evaluators, using the intraclass correlation coefficient (ICC) and the kappa coefficient. Results There are no significant differences in the discriminant validity of the Sunderland and Solomon systems in any of the samples (clinical: aROC 0.73 [CI95%: 0.64-0.81] and 0.77 [CI95%: 0.69-0.85] respectively, P=.19; volunteers: aROC 0.69 [CI95%: 0.67-0.71] and 0.72 [CI95%: 0.69-0.73] respectively, P=.08). The cut-off points ≤8 and ≤5 correctly classify 71% and 73% of the clinical sample and 82% and 84% of the volunteer sample, respectively. Both systems have good agreement in the clinical sample (Sunderland: ICC 0.90 [CI95%: 0.81-0.93], kappa 0.76 [CI95%: 0.70-0.83]; Solomon: 0.92 [CI95%: 0.88-0.95] and 0.77 [CI95%: 0.71-0.83] respectively), somewhat higher in the second, although the differences are not significant (AU)
Assuntos
Humanos , Masculino , Feminino , Idoso , Idoso de 80 Anos ou mais , Variações Dependentes do Observador , Avaliação Geriátrica/métodos , Disfunção Cognitiva/diagnóstico , Testes Neuropsicológicos , Reprodutibilidade dos Testes , Sensibilidade e EspecificidadeRESUMO
BACKGROUND: The clinical application of coronary MR angiography (MRA) combining diastole and systole imaging has never been described comprehensively in coronary artery disease (CAD) patients. We aimed to design an optimal non-contrast coronary MRA scan protocol combining diastolic and systolic imaging and to (1) evaluate its diagnostic performance for detecting significant coronary stenosis; (2) evaluate the feasibility of this protocol to noninvasively measure the coronary distensibility index (CDI). METHODS: From June 2021 to May 2022, 33 healthy volunteers and 91 suspected CAD patients scheduled for X-ray coronary angiography (CAG) were prospectively enrolled. 3T non-contrast water-fat coronary MRA was carried out twice at diastole and systole. Significant coronary stenosis was defined as a luminal diameter reduction of ≥ 50% using CAG as the reference and was evaluated as follows: (1) by coronary MRA in diastole alone; (2) by coronary MRA in systole alone; (3) by combined coronary MRA in diastole and systole. According to CAG, the patients were divided into significant CAD patients and non-significant CAD patients. The difference in CDI among participants was evaluated. RESULTS: Combined coronary MRA was completed in 31 volunteers and 76 patients. The per-patient sensitivity, specificity, and accuracy of combined coronary MRA were 97.5%, 83.3%, and 90.8%, respectively. Compared with single diastolic mode, combined coronary MRA showed equally high sensitivity but improved specificity on a per-patient basis (83.3% vs. 63.9%, adjusted P = 0.013). The CDI tested by coronary MRA decreased incrementally from healthy volunteers to non-significant and significant CAD patients. CONCLUSION: Compared with single-phase mode, 3 T non-contrast combined coronary MRA significantly improved specificity and may have potential to be a simple noninvasive method to measure CDI.