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1.
Talanta ; 236: 122827, 2022 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-34635217

RESUMO

Cryptococcal meningitis (CM) is a global threat with significant attributable morbidity and mortality. Information on microfluidic detection for CM diagnosis is still limited. We developed a multifunctional microfluidic module that integrated the pathogen enrichment and on-chip nucleic acid extraction. The module adopted a simple filtration membrane to effectively capture Cryptococcus cells and simplify the process, and combined lyticase digestion, alkaline lysis and heating methods to optimize the strategy to achieve nucleic acid extraction. The entire process was operated in the module, which reduced the exposure risk of directly processing cryptococcal samples. A portable one-pot lyophilized LAMP reagent bead with no temperature limit was developed, which improved the flexibility of operation. This module did not require any additional instrument, and is promising to develop a simple, rapid, and efficient approach to realize the "sample in and answer out" detection of real CSF samples. This microfluidic module had practical prospects and is expected to replace LFA for efficacy evaluation and follow-up in the middle and late stages of CM treatment, and could be used as an auxiliary method to confirm cases with questionable LFA results in the early diagnosis of CM.


Assuntos
Meningite Criptocócica , Ácidos Nucleicos , Humanos , Meningite Criptocócica/diagnóstico , Microfluídica , Análise de Sequência com Séries de Oligonucleotídeos
2.
Talanta ; 236: 122866, 2022 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-34635248

RESUMO

Small molecular contaminants (such as mycotoxins, antibiotics, pesticide residues, etc.) in food and environment have given rise to many biological and ecological toxicities, which has attracted worldwide attention in recent years. Meanwhile, due to the advantages of aptamers such as high specificity and stability, easy synthesis and modification, as well as low cost and immunogenicity, various aptasensors for the detection of small molecular contaminants have been flourishing. An aptasensor as a whole is composed of an aptamer-based target recognizer and a signal transducer, which are fields of concentrated research. In the practical detection applications, in order to achieve the quantitative detection of small molecular contaminants at low abundance in real samples, a large number of signal enhancing strategies have been utilized in the development of aptasensors. Recent years is a vintage period for efficient signal enhancing strategies of aptasensors by the aid of nanomaterials and nucleic acid amplification that are applied in the elements for target recognition and signal conversion. Therefore, this paper meticulously reviews the signal enhancing strategies based on nanomaterials (including the (quasi-)zero-dimensional, one-dimensional, two-dimensional and three-dimensional nanomaterials) and nucleic acid amplification (including enzyme-assisted nucleic acid amplification and enzyme-free nucleic acid amplification). Furthermore, the challenges and future trends of the abovementioned signal enhancing strategies for application are also discussed in order to inspire the practitioners in the research and development of aptasensors for small molecular contaminants.


Assuntos
Aptâmeros de Nucleotídeos , Técnicas Biossensoriais , Nanoestruturas , Ácidos Nucleicos , Técnicas de Amplificação de Ácido Nucleico
3.
Anal Chim Acta ; 1186: 339134, 2021 Nov 22.
Artigo em Inglês | MEDLINE | ID: mdl-34756259

RESUMO

In recent years, single particle inductively coupled plasma mass spectrometry (SP-ICP-MS) has become a powerful tool for biological quantitative analysis. Homogeneous analysis method requires no separation and washing steps, which is suited for the analysis of highly infectious pathogens, so as to reduce the risk of infection during the operation. SARS-CoV-2 spreads all over the world, and its early infection symptoms are similar to influenza, which brings inconvenience to triage. Therefore, developing novel analytical method for simultaneous detection of multiple viral nucleic acids is essential. Taking the advantages of SP-ICP-MS and homogeneous analysis strategy, a SP-ICP-MS homogeneous nucleic acid assay by using gold nanoparticles (Au NPs) and silver nanoparticles (Ag NPs) probes was established for simultaneous sensitive analysis of SARS-CoV-2 and influenza A (H3N2). In the present of target SARS-CoV-2 or H3N2 nucleic acids, corresponding Au NPs or Ag NPs probes form larger aggregates, resulting in increased pulse signal intensity and reduced pulse signal frequency of the corresponding NPs in SP-ICP-MS measurement. In this assay, the reaction system of Au NPs and Ag NPs probes does not interfere with each other, and there was no separation and washing procedure, which facilitates operation, saves the analysis time, and improves the analysis efficiency. The linear range of this method is 5-1000 pmol L-1, with low-level limits of quantification of target nucleic acid. The developed SP-ICP-MS simultaneous homogeneous detection method has a good potential for detecting nucleic acid, protein, cell and other biological samples by changing different modification sequences on the NPs probes.


Assuntos
COVID-19 , Influenza Humana , Nanopartículas Metálicas , Ácidos Nucleicos , Ouro , Humanos , Vírus da Influenza A Subtipo H3N2 , Espectrometria de Massas , SARS-CoV-2 , Prata
4.
Sensors (Basel) ; 21(21)2021 Oct 26.
Artigo em Inglês | MEDLINE | ID: mdl-34770415

RESUMO

In the past few decades, biosensors have been gradually developed for the rapid detection and monitoring of human diseases. Recently, functional nucleic-acid (FNA) biosensors have attracted the attention of scholars due to a series of advantages such as high stability and strong specificity, as well as the significant progress they have made in terms of biomedical applications. However, there are few reports that systematically and comprehensively summarize its working principles, classification and application. In this review, we primarily introduce functional modes of biosensors that combine functional nucleic acids with different signal output modes. In addition, the mechanisms of action of several media of the FNA biosensor are introduced. Finally, the practical application and existing problems of FNA sensors are discussed, and the future development directions and application prospects of functional nucleic acid sensors are prospected.


Assuntos
Técnicas Biossensoriais , Ácidos Nucleicos , Humanos
5.
Zhongguo Xue Xi Chong Bing Fang Zhi Za Zhi ; 33(5): 452-456, 2021 Oct 27.
Artigo em Chinês | MEDLINE | ID: mdl-34791841

RESUMO

OBJECTIVE: To develop a fluorescent recombinase-aided isothermal amplification (RAA)-based nucleic acid assay for detection of Leshimania. METHODS: Specific primers and probes were designed targeting Leishmania internal transcribed spacer 1 (ITS1) gene for RAA assay, and a fluorescent RAA assay was developed for detection of Leishmania following screening of primer pairs and optimization of primer and probe concentrations. The sensitivity of RAA assay for detection of Leishmania was evaluated using recombinant plasmid containing Leishmania ITS1 gene sequences at different copies and Leshimania genomic DNA at different concentrations as templates, and the specificity of RAA assay for detection of Leishmania was evaluated using the genomic DNA of transfusion-transmitted parasites, including Babesia microti, Toxoplasma gondii, Plamodium vivax, P. ovale, P. falciparum, P. malariae, L. donovani and L. infantum. RESULTS: After the optimal primer pair was screened from 9 pairs of primer combinations, the final primer and probe concentrations were optimized as 0.3 µmol/L and 0.08 µmol/L, respectively. Nucleic acid detection of Leishmania was completed by the fluorescent RAA assay at an isothermal temperature of 39 °C within 20 min. Remarkable florescent signals were seen within 5 min following RAA detection of genomic DNA of L. donovani and L. infantum, and no cross-reactions were observed with B. microti, T. gondii, P. vivax, P. ovale, P. falciparum or P. malariae. The lowest limitation of detection of the fluorescent RAA assay was 10 copies/µL recombinant plasmid containing Leishmania ITS1 gene sequences and 1 fg/µL Leishmania genomic DNA. CONCLUSIONS: A rapid, simple, sensitive and specific fluorescent RAA assay is successfully developed for detection of L. donovani and L. infantum, which is effective for field screening of leishmaniasis.


Assuntos
Leishmania , Ácidos Nucleicos , Leishmania/genética , Técnicas de Amplificação de Ácido Nucleico , Recombinases , Sensibilidade e Especificidade
6.
Zhongguo Xue Xi Chong Bing Fang Zhi Za Zhi ; 33(5): 464-469, 2021 Oct 26.
Artigo em Chinês | MEDLINE | ID: mdl-34791843

RESUMO

OBJECTIVE: To establish a nucleic acid assay for detection of Paragonimus skrjabini based on the recombinase-aided isothermal amplification (RAA) technique, and to preliminarily evaluate its detection efficiency. METHODS: The metacercariae of P. skrjabini, P. westermani and Euparagonimus cenocopiosus were isolated from crabs, and genomic DNA was extracted for molecular characterization. The cytochrome coxidase 1 (cox1) gene sequence of P. skrjabini was selected as the target gene fragment, and the primers and probes were designed, screened and synthesized for RAA assay. The genomic DNA of P. skrjabini metacercariae from Jiyuan City and Yiyang County of Luoyang City, Henan Province were used as templates for verification of the fluorescent RAA assay. The fluorescent RAA assay was performed to detect different concentrations of plasmids containing target gene fragment and P. skrjabini metacercariae genomic DNA to determine the sensitivity. Fluorescent RAA assay was performed with recombinant plasmids containing P. skrjabini cox1 gene sequences at different concentrations and P. skrjabini genomic DNA as templates to evaluate its sensitivity, and the genomic DNA of P. westermani, E. cenocopiosus, Clonorchis sinensis and Schistosoma japonicum was detected with fluorescent RAA assay to evaluate its specificity. RESULTS: P. skrjabini, P. westermani and E. cenocopiosus metacercariae were isolated from crabs, respectively. Molecular characterization and phylogenetic analysis confirmed their homology with the genes sequences of standard Paragonimus strains in GenBank. A fluorescent RAA assay was successfully established for nucleic acid detection of P. skrjabini, and the genomic DNA of P. skrjabini metacercariae from Jiyuan City and Yiyang County of Luoyang City, Henan Province was amplified using the fluorescent RAA assay within 5 min, while the negative control was not amplified. If the recombinant plasmid containing P. skrjabini cox1 gene sequences was used as templates, the fluorescent RAA assay showed the lowest detection limit of 10 copies/µL, and positive amplification was observed within 5 min. If genomic DNA was used as templates, the fluorescent RAA assay showed the lowest detection limit of 10 pg/µL, and all positive amplifications were found within 5 to 10 min. In addition, the fluorescent RAA assay was tested negative for P. westermani, E. cenocopiosus, C. sinensis and S. japonicum. CONCLUSIONS: A rapid, sensitive and specific fluorescent RAA assay is successfully established for nucleic acid detection of P. skrjabini, which has potential values in rapid field detection and species identification in freshwater crabs in areas endemic for P. skrjabini.


Assuntos
Ácidos Nucleicos , Recombinases , Animais , Técnicas de Amplificação de Ácido Nucleico , Filogenia , Recombinases/genética , Sensibilidade e Especificidade
7.
Anal Chem ; 93(45): 15216-15223, 2021 11 16.
Artigo em Inglês | MEDLINE | ID: mdl-34736322

RESUMO

The development of a sensitive, facile, and cost-effective colorimetric method is of great significance for the point-of-care testing of viral nucleic acid. Herein, we reported a strand displacement amplification assisted CRISPR-Cas12a (SDACC) method for the colorimetric analysis of viral nucleic acid. The hepatitis B virus (HBV) DNA was chosen as the target to trigger strand displacement amplification (SDA) and generate abundant single-strand DNA (ssDNA) products. The ssDNA amplicon hybridized with template DNA to activate the trans-cleavage activity of CRISPR-Cas12a, leading to the nonspecific cleavage of ssDNA on GOx-ssDNA-modified magnetic beads and the release of GOx. The released GOx was capable of catalyzing the substrate solution to generate a color change, which could be directly observed by naked eyes. The SDACC strategy could identify a single-base mismatch located in the DNA sequence and achieve a sensitive detection for HBV DNA with the limit of detection as low as 41.8 fM. Notably, the sophisticated primer design for target amplification and complicated detection process could be circumvented. The current approach realizes a simple, low-cost, and sensitive colorimetric detection for viral nucleic acid and holds great promise for the practical application of virus infection diagnosis.


Assuntos
Técnicas Biossensoriais , Ácidos Nucleicos , Sistemas CRISPR-Cas/genética , Colorimetria , DNA , DNA de Cadeia Simples/genética , Técnicas de Amplificação de Ácido Nucleico
8.
BMC Infect Dis ; 21(1): 1147, 2021 Nov 10.
Artigo em Inglês | MEDLINE | ID: mdl-34758738

RESUMO

BACKGROUND: Dengue, chikungunya and zika infections occur in tropical and subtropical regions of the world. We describe the utilization of an in-house nucleic acid test (NAT) targeting all three viruses for febrile returning travelers in Alberta, Canada. METHODS: NAT was performed until 40 days from symptom onset or exposure due to the prolonged duration of zika virus RNA detection. From Sept 1, 2017 to August 31, 2019, 2552 specimens from 1932 patients were tested. RESULTS: Approximately 2% of patients tested were NAT positive for dengue virus (n = 42), chikungunya virus (n = 4), and zika virus (n = 1). The majority presented with fever, myalgia and rash. Regions with the most frequent travel included SouthEast Asia (68.5%), South America (25%) and the Caribbean (6.5%). Ct values were stronger (~ 1.5 logs) for patients within 1-3 days following onset of clinical symptoms than those presenting later. Nineteen patients had urine and plasma submitted; 5 were positive for both specimens and 2 were positive only for dengue virus in the urine. Also, Ct values were lower for plasma when compared to the corresponding urine. RNA was detected until 10 days and 5 days post-exposure in plasma and urine respectively for dengue virus. CONCLUSIONS: Owing to dengue viremia detected beyond the conventional 7 days and low levels of circulating zika virus globally, a cutoff of 14 days from symptom onset to NAT is sufficient to diagnose acute cases. Inclusion of a zoonotic history form that collects appropriate clinical history results in improved test utilization.


Assuntos
Arbovírus , Febre de Chikungunya , Dengue , Ácidos Nucleicos , Infecção por Zika virus , Zika virus , Alberta , Febre de Chikungunya/diagnóstico , Dengue/diagnóstico , Humanos , Laboratórios , Saúde Pública , Zika virus/genética , Infecção por Zika virus/diagnóstico
9.
Sensors (Basel) ; 21(22)2021 Nov 14.
Artigo em Inglês | MEDLINE | ID: mdl-34833643

RESUMO

In recent years, cerium oxide (CeO2) nanoparticles (NPs) have drawn significant attention owing to their intrinsic enzyme mimetic properties, which make them powerful tools for biomolecular detection. In this work, we evaluated the effect of pyrophosphate (PPi) on the oxidase activity of CeO2 NPs. The presence of PPi was found to enhance the oxidase activity of CeO2 NPs, with enhanced colorimetric signals. This particular effect was then used for the colorimetric detection of target nucleic acids. Overall, the PPi-enhanced colorimetric signals of CeO2 NPs oxidase activity were suppressed by the presence of the target nucleic acids. Compared with previous studies using CeO2 NPs only, our proposed system significantly improved the signal change (ca. 200%), leading to more sensitive and reproducible colorimetric analysis of target nucleic acids. As a proof-of-concept study, the proposed system was successfully applied to the highly selective and sensitive detection of polymerase chain reaction products derived from Klebsiella pneumoniae. Our findings will benefit the rapid detection of nucleic acid biomarkers (e.g., pathogenic bacterial DNA or RNA) in point-of-care settings.


Assuntos
Cério , Nanopartículas , Ácidos Nucleicos , Colorimetria , Difosfatos , Oxirredutases
10.
J Immunol ; 207(11): 2813-2827, 2021 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-34740958

RESUMO

Hypoxia-inducible factor-1α (HIF-1α) is an important regulator of glucose metabolism and inflammatory cytokine production in innate immune responses. Viruses modulate HIF-1α to support viral replication and the survival of infected cells, but it is unclear if this transcription factor also plays an important role in regulating antiviral immune responses. In this study, we found that short and long dsRNA differentially engage TLR3, inducing distinct levels of proinflammatory cytokine production (TNF-α and IL-6) in bone marrow-derived macrophages from C57BL/6 mice. These responses are associated with differential accumulation of HIF-1α, which augments NF-κB activation. Unlike TLR4 responses, increased HIF-1α following TLR3 engagement is not associated with significant alterations in glycolytic activity and was more pronounced in low glucose conditions. We also show that the mechanisms supporting HIF-1α stabilization may differ following stimulation with short versus long dsRNA and that pyruvate kinase M2 and mitochondrial reactive oxygen species play a central role in these processes. Collectively, this work suggests that HIF-1α may fine-tune proinflammatory cytokine production during early antiviral immune responses, particularly when there is limited glucose availability or under other conditions of stress. Our findings also suggest we may be able to regulate the magnitude of proinflammatory cytokine production during antiviral responses by targeting proteins or molecules that contribute to HIF-1α stabilization.


Assuntos
Citocinas/biossíntese , Glucose/imunologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/imunologia , Macrófagos/imunologia , Ácidos Nucleicos/imunologia , Receptor 3 Toll-Like/imunologia , Animais , Células Cultivadas , Camundongos , Camundongos Endogâmicos C57BL , Espécies Reativas de Oxigênio/imunologia
11.
Anal Chem ; 93(46): 15482-15492, 2021 11 23.
Artigo em Inglês | MEDLINE | ID: mdl-34767335

RESUMO

There is an urgent need for reliable biosensors to detect nucleic acid of interest in clinical samples. We propose that the accuracy of the present nucleic acid-sensing method can be advanced by avoiding false-positive identifications derived from nonspecific interactions (e.g., nonspecific binding, probe degradation). The challenge is to exploit biosensors that can distinguish false-positive from true-positive samples in nucleic acid screening. In the present study, by learning from the enzymatic cycle in nature, we raise an allostery tool displaying invertible positive/negative cooperativity for reversible or cyclic activity control of the biosensing probe. We demonstrate that the silencing and regeneration of a positive (or negative) allosteric effector can be carried out through toehold displacement or an enzymatic reaction. We, thus, have developed several dynamic biosensors that can repeatedly measure a single nucleic acid sample. The ability to distinguish a false-positive from a true-positive signal is ascribed to the nonspecific interaction presenting equivalent signal variations, while the specific target binding exhibits diverse signal variations according to repeated measurements. Given its precise identification, such consequent dynamic biosensors offer exciting opportunities in physiological and pathological diagnosis.


Assuntos
Técnicas Biossensoriais , Ácidos Nucleicos
12.
Methods Enzymol ; 661: 407-431, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34776222

RESUMO

We present a Chemistry and Structure Screen Integrated Efficiently (CASSIE) approach (named for Greek prophet Cassandra) to design inhibitors for cancer biology and pathogenesis. CASSIE provides an effective path to target master keys to control the repair-replication interface for cancer cells and SARS CoV-2 pathogenesis as exemplified here by specific targeting of Poly(ADP-ribose) glycohydrolase (PARG) and ADP-ribose glycohydrolase ARH3 macrodomains plus SARS CoV-2 nonstructural protein 3 (Nsp3) Macrodomain 1 (Mac1) and Nsp15 nuclease. As opposed to the classical massive effort employing libraries with large numbers of compounds against single proteins, we make inhibitor design for multiple targets efficient. Our compact, chemically diverse, 5000 compound Goldilocks (GL) library has an intermediate number of compounds sized between fragments and drugs with predicted favorable ADME (absorption, distribution, metabolism, and excretion) and toxicological profiles. Amalgamating our core GL library with an approved drug (AD) library, we employ a combined GLAD library virtual screen, enabling an effective and efficient design cycle of ranked computer docking, top hit biophysical and cell validations, and defined bound structures using human proteins or their avatars. As new drug design is increasingly pathway directed as well as molecular and mechanism based, our CASSIE approach facilitates testing multiple related targets by efficiently turning a set of interacting drug discovery problems into a tractable medicinal chemistry engineering problem of optimizing affinity and ADME properties based upon early co-crystal structures. Optimization efforts are made efficient by a computationally-focused iterative chemistry and structure screen. Thus, we herein describe and apply CASSIE to define prototypic, specific inhibitors for PARG vs distinct inhibitors for the related macrodomains of ARH3 and SARS CoV-2 Nsp3 plus the SARS CoV-2 Nsp15 RNA nuclease.


Assuntos
COVID-19 , Ácidos Nucleicos , Síndrome Respiratória Aguda Grave , Reparo do DNA , Humanos , Simulação de Acoplamento Molecular , SARS-CoV-2
13.
Zhonghua Liu Xing Bing Xue Za Zhi ; 42(6): 1002-1007, 2021 Jun 10.
Artigo em Chinês | MEDLINE | ID: mdl-34814497

RESUMO

Objective: To analysis effectiveness of the "14 plus 7 day quarantine" and "nucleic acid plus total antibody testing" strategy (combined screening strategy) for screenin the imported patients with COVID-19 in Xiamen. Methods: The study populations were overseas travelers arriving in Xiamen from March 17 to December 31, 2020, and overseas travelers who had quarantine outside Xiamen for less than 21 days from July 18 to December 31, 2020. Data were collected and analyzed on the timing of detection, pathways, and test results of the imported patients with COVID-19 after implementing combined screening strategy. Results: A total of 304 imported patients with COVID-19 were found from 174 628 overseas travelers and 943 overseas travelers from other cities. A total of 163 cases (53.6%) were diagnosed by multitime, multisite intensive nucleic acid testing after positive finding in total antibody testing. Among them, 27 (8.9%) were first positive for nucleic acid in 14 plus 7 day quarantine and 136 were first positive for nucleic acid in 14-day quarantine. Only 8 of these individuals were tested positive for nucleic acid after positive total antibody testing. The other 128 individuals were tested positive for nucleic acid after being negative for average 2.3 times (maximum of 6 times). Aditional 155 cases might be detected by using the combined "14 plus 7 day quarantine" and " nucleic acid plus total antibody testing" strategy compared with "14-day quarantine and nucleic acid testing" strategy, accounting for 51.0% of the total inbound infections. So the combined screening strategy doubled the detection rate for imported patients with COVID-19. No second-generation case caused by overseas travelers had been reported in Xiamen as of February 26, 2021. Conclusions: Xiamen's combined screening strategy can effectively screen the imported patients with COVID-19 who were first positive for nucleic acid after 14 day quarantine. Compared with "14 day quarantine and nucleic acid testing", the combined screening strategy improved detection rate and further reduced the risk of the secondary transmission caused by the imported patients with COVID-19.


Assuntos
COVID-19 , Ácidos Nucleicos , Humanos , Programas de Rastreamento , Quarentena , SARS-CoV-2
14.
Zhonghua Liu Xing Bing Xue Za Zhi ; 42(8): 1347-1352, 2021 Aug 10.
Artigo em Chinês | MEDLINE | ID: mdl-34814552

RESUMO

Objective: To analyze the sensitivity and specificity of SARS-CoV-2 nucleic acid testing in 20 348 close contacts of COVID-19 cases in different prevention and control stages in Guangzhou and to provide scientific evidence for optimizing epidemic response strategies. Methods: A total of 20 348 close contacts of COVID-19 cases in Guangzhou were traced between February 21 and September 22,2020. All the close contacts were tested for the nucleic acid of SARS-CoV-2. The sensitivity and specificity of nucleic acid testing and diagnosis in the different prevention and control stages were compared. Results: In 20 348 close contacts, 12 462 were males (61.24%), the median (P25,P75) of age of them was 31.0 years (23.0,43.0), the median number (P25,P75) of nucleic acid testing for them was 2.0 (1.0,3.0), and the median (P25,P75) of their quarantine days was 12.0 (8.0,13.0) days, respectively. A total of 256 COVID-19 cases were confirmed in the close contacts after seven nucleic acid tests. In the 1st, 2nd, 3rd and 7th nucleic acid testing, the sensitivity and specificity were 69.14% and 99.99% (177 cases confirmed), 89.84% and 99.99% (230 cases confirmed), 97.27% and 99.99% (249 cases confirmed), and 100.00% and 99.98%, respectively. In the three stages of COVID-19 prevention and control in China: domestic case stage, imported case stage, and imported case associated local epidemic stage, the sensitivity of the 1st nucleic acid testing was 70.68%, 68.00% and 67.35%, and the specificity was 99.98%, 100.00% and 100.00%, respectively. Conclusions: The sensitivity of nucleic acid testing in the close contacts at the different stages were consistent with slight decrease, which might be related to the increased proportion of asymptomatic infections in the late stage of epidemic prevention and control with COVID-19 in Guangzhou. It is suggested to give three nucleic acid tests to improve the sensitivity and reduce false negative risk.


Assuntos
COVID-19 , Ácidos Nucleicos , Adulto , Infecções Assintomáticas , Humanos , Masculino , SARS-CoV-2 , Sensibilidade e Especificidade
15.
Zhonghua Liu Xing Bing Xue Za Zhi ; 42(8): 1353-1359, 2021 Aug 10.
Artigo em Chinês | MEDLINE | ID: mdl-34814553

RESUMO

Objective: To establish an index system of population based SARS-CoV-2 nucleic acid screening, and provide reference to determine the screening coverage appropriately. Methods: The literature review and brain storming sessions were used to develop the basic frame and index system of population based SARS-CoV-2 nucleic acid screening. Based on Delphi method and Analytic Hierarchy Process, 21 domestic experts were selected for two rounds of consultation to determine the index system of population based SARS-CoV-2 nucleic acid screening and its weight. Results: The positive indexes of experts in two rounds of consultations were both 100%. The experts' authority coefficients (Cr) were 0.88±0.08 and 0.89±0.07, respectively. And the range of coefficient of variation (CV) were (0.08, 0.24), (0.09, 0.25). The Kendall's W coordination coefficients were 0.34 and 0.22 respectively, which were statistically significant. The index system of population based SARS-CoV-2 nucleic acid screening was established, which had 4 first-level indexes, 11 second-level indexes and 58 third-level indexes. Besides, the weight of each index was determined. Conclusion: The index system of population based SARS-CoV-2 nucleic acid screening has been established, which can provide scientific reference for the health administration to determine the coverage of population based SARS-CoV-2 nucleic acid screening when local COVID-19 epidemic occurs.


Assuntos
COVID-19 , Ácidos Nucleicos , Humanos , Programas de Rastreamento , SARS-CoV-2
16.
Front Cell Infect Microbiol ; 11: 755508, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34722341

RESUMO

COVID-19 continues to circulate globally in 2021, while under the precise policy implementation of China's public health system, the epidemic was quickly controlled, and society and the economy have recovered. During the pandemic response, nucleic acid detection of SARS-CoV-2 has played an indispensable role in the first line of defence. In the cases of emergency operations or patients presenting at fever clinics, nucleic acid detection is required to be performed and reported quickly. Therefore, nucleic acid point-of-care testing (POCT) technology for SARS-CoV-2 identification has emerged, and has been widely carried out at all levels of medical institutions. SARS-CoV-2 POCT has served as a complementary test to conventional polymerase chain reaction (PCR) batch tests, thus forming an experimental diagnosis platform that not only guarantees medical safety but also improves quality services. However, in view of the complexity of molecular diagnosis and the biosafety requirements involved, pathogen nucleic acid POCT is different from traditional blood-based physical and chemical index detection. No guidelines currently exist for POCT quality management, and there have been inconsistencies documented in practical operation. Therefore, Shanghai Society of Molecular Diagnostics, Shanghai Society of Laboratory Medicine, Clinical Microbiology Division of Shanghai Society of Microbiology and Shanghai Center for Clinical Laboratory have cooperated with experts in laboratory medicine to generate the present expert consensus. Based on the current spectrum of major infectious diseases in China, the whole-process operation management of pathogen POCT, including its application scenarios, biosafety management, personnel qualification, performance verification, quality control, and result reporting, are described here. This expert consensus will aid in promoting the rational application and robust development of this technology in public health defence and hospital infection management.


Assuntos
COVID-19 , Ácidos Nucleicos , China , Consenso , Humanos , Testes Imediatos , SARS-CoV-2
17.
Euro Surveill ; 26(45)2021 11.
Artigo em Inglês | MEDLINE | ID: mdl-34763752

RESUMO

BackgroundReliable testing for SARS-CoV-2 is key for the management of the COVID-19 pandemic.AimWe estimate diagnostic accuracy for nucleic acid and antibody tests 5 months into the COVID-19 pandemic, and compare with manufacturer-reported accuracy.MethodsWe reviewed the clinical performance of SARS-CoV-2 nucleic acid and antibody tests based on 93,757 test results from 151 published studies and 20,205 new test results from 12 countries in the European Union and European Economic Area (EU/EEA).ResultsPooling the results and considering only results with 95% confidence interval width ≤ 5%, we found four nucleic acid tests, including one point-of-care test and three antibody tests, with a clinical sensitivity ≥ 95% for at least one target population (hospitalised, mild or asymptomatic, or unknown). Nine nucleic acid tests and 25 antibody tests, 12 of them point-of-care tests, had a clinical specificity of ≥ 98%. Three antibody tests achieved both thresholds. Evidence for nucleic acid point-of-care tests remains scarce at present, and sensitivity varied substantially. Study heterogeneity was low for eight of 14 sensitivity and 68 of 84 specificity results with confidence interval width ≤ 5%, and lower for nucleic acid tests than antibody tests. Manufacturer-reported clinical performance was significantly higher than independently assessed in 11 of 32 and four of 34 cases, respectively, for sensitivity and specificity, indicating a need for improvement in this area.ConclusionContinuous monitoring of clinical performance within more clearly defined target populations is needed.


Assuntos
COVID-19 , Ácidos Nucleicos , Humanos , Pandemias , SARS-CoV-2 , Sensibilidade e Especificidade
18.
Nanoscale ; 13(43): 18084-18088, 2021 Nov 11.
Artigo em Inglês | MEDLINE | ID: mdl-34730160

RESUMO

This communication describes a novel water-soluble membrane prepared from chitosan intended for SARS-CoV-2 viral nucleic acid collection and detection. The CSH membrane formed from nanofibers shows promising potential in the quantitative determination of the SARS-CoV-2 viral nucleic acids at a concentration of 102 copies per L in air. The sponge-like structure which allows gas to pass through for collection of viral nucleic acids potentially provides simple, fast, and reliable sampling as well as detection of various types of airborne viruses.


Assuntos
COVID-19 , Ácidos Nucleicos , Humanos , RNA Viral , SARS-CoV-2 , Manejo de Espécimes , Água
19.
Sheng Wu Gong Cheng Xue Bao ; 37(11): 3890-3904, 2021 Nov 25.
Artigo em Chinês | MEDLINE | ID: mdl-34841793

RESUMO

Clustered regularly interspaced short palindromic repeats/clustered regularly interspaced short palindromic repeats -associated protein (CRISPR/Cas) has been developed as a precise, efficient, affordable and sensitive nucleic acid detection tool due to its efficient targeted binding ability and programmability. At present, biosensors based on CRISPR-Cas system have shown excellent performance in the detection of nucleic acid of pathogens, which has attracted widespread attention, and is expected to replace the conventional detection methods. This review summarizes the latest research progress of biosensors based on CRISPR/Cas system for detecting nucleic acid of pathogens.


Assuntos
Técnicas Biossensoriais , Ácidos Nucleicos , Sistemas CRISPR-Cas/genética , Ácidos Nucleicos/genética
20.
Anal Chem ; 93(46): 15288-15294, 2021 11 23.
Artigo em Inglês | MEDLINE | ID: mdl-34735121

RESUMO

Herein, a pipette-tip-enabled digital nucleic acid analyzer for high-performance COVID-19 testing is demonstrated. This is achieved by digital loop-mediated isothermal amplification (digital LAMP or dLAMP) using common laboratory equipment and materials. It is shown that simply fixing a glass capillary inside conventional pipette tips enables the generation of monodisperse, water-in-oil microdroplets with benchtop centrifugation. It is shown that using LAMP, the ORF1a/b gene, a standard test region for COVID-19 screening, can be amplified without a thermal cycler. The amplification allows counting of fluorescent microdroplets so that Poisson analysis can be performed to allow quantification with a limit of detection that is 1 order of magnitude better than those of nondigital techniques and comparable to those of commercial dLAMP platforms. It is envisioned that this work will inspire studies on ultrasensitive digital nucleic acid analyzers demanding both sensitivity and accessibility, which is pivotal to their large-scale applications.


Assuntos
COVID-19 , Ácidos Nucleicos , Teste para COVID-19 , Humanos , Técnicas de Diagnóstico Molecular , Técnicas de Amplificação de Ácido Nucleico , SARS-CoV-2
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