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1.
Med. lab ; 26(3): 237-259, 2022. Tabs, ilus, Grafs
Article in Spanish | WHO COVID, LILACS (Americas) | ID: covidwho-20239968

ABSTRACT

La enfermedad COVID­19 es causada por el virus SARS-CoV-2, descrito por primera vez en diciembre del 2019 en Wuhan, China, y declarada en marzo del 2020 como una pandemia mundial. Actualmente existen diversos métodos diagnósticos para COVID-19, siendo el estándar de oro la detección del material genético mediante la reacción en cadena de la polimerasa (PCR), en su variante, la RT-PCR, que detecta el material genético de tipo ARN presente en el virus. Sin embargo, es necesario disponer de pruebas rápidas con alta sensibilidad y precisión para realizarlas a gran escala y brindar un diagnóstico oportuno. Adicionalmente, se debe disponer de otras herramientas que, si bien no van a establecer un diagnóstico, le van a permitir al profesional brindar un mejor manejo clínico y epidemiológico que ayuden a predecir el agravamiento del paciente y su posible ingreso a UCI, destacando entre estas los niveles de dímero D, linfocitos, ferritina, urea y creatinina, entre otras. En esta revisión se evalúa la utilidad y limitaciones de los diferentes métodos diagnósticos para COVID-19, al igual que las características, fisiopatología y respuesta inmune al SARS-CoV-2, así como algunos aspectos preanalíticos de importancia que ayudan a minimizar errores en el diagnóstico como consecuencia de procedimientos incorrectos en la toma, transporte y conservación de la muestra, y que permiten al profesional emitir resultados veraces y confiables. Lo anterior se realizó basado en artículos originales, revisiones y guías clínicas


COVID­19 is caused by the SARS-CoV-2 virus, first described in December 2019 in Wuhan, China, and declared a global pandemic in March 2020. Currently there are various diagnostic methods for COVID-19, the gold standard is the detection of genetic material through polymerase chain reaction (PCR) in its variant, RT-PCR, which detects RNA-type genetic material present in the virus. However, it is necessary to have rapid tests with high sensitivity and precision to be performed on a large scale and provide timely diagnosis. Furthermore, other tools must be available, and although they will not establish the diagnosis, will allow the professional to provide better clinical and epidemiological management that will help predict the worsening of the patient and possible admission to the ICU. Among these, levels of D-dimer, lymphocytes, ferritin, urea and creatinine. In this review, the usefulness and limitations of the different diagnostic methods for COVID-19 are evaluated, as well as the characteristics, pathophysiology and immune response to SARS-CoV-2, and some important preanalytical aspects that allow minimizing diagnostic errors as a consequence of incorrect procedures in the collection, transport and conservation of the sample, that allow the professional to yield accurate and reliable results. This article was completed based on original articles, reviews and clinical guidelines


Subject(s)
SARS-CoV-2 , Polymerase Chain Reaction , Inflammation Mediators , Containment of Biohazards , Diagnosis , Ferritins , COVID-19 , L-Lactate Dehydrogenase , Methods
2.
Biosensors (Basel) ; 13(5)2023 May 05.
Article in English | MEDLINE | ID: covidwho-20239011

ABSTRACT

We developed a microfluidic chip integrated with nucleic acid purification and droplet-based digital polymerase chain reaction (ddPCR) modules to realize a 'sample-in, result-out' infectious virus diagnosis. The whole process involved pulling magnetic beads through drops in an oil-enclosed environment. The purified nucleic acids were dispensed into microdroplets by a concentric-ring, oil-water-mixing, flow-focusing droplets generator driven under negative pressure conditions. Microdroplets were generated with good uniformity (CV = 5.8%), adjustable diameters (50-200 µm), and controllable flow rates (0-0.3 µL/s). Further verification was provided by quantitative detection of plasmids. We observed a linear correlation of R2 = 0.9998 in the concentration range from 10 to 105 copies/µL. Finally, this chip was applied to quantify the nucleic acid concentrations of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The measured nucleic acid recovery rate of 75 ± 8.8% and detection limit of 10 copies/µL proved its on-chip purification and accurate detection abilities. This chip can potentially be a valuable tool in point-of-care testing.


Subject(s)
COVID-19 , Nucleic Acids , Humans , SARS-CoV-2 , COVID-19/diagnosis , Polymerase Chain Reaction , Nucleic Acids/analysis , Oligonucleotide Array Sequence Analysis
3.
Virus Genes ; 59(3): 343-350, 2023 Jun.
Article in English | MEDLINE | ID: covidwho-20235973

ABSTRACT

The recent widespread emergence of monkeypox (mpox), a rare and endemic zoonotic disease by monkeypox virus (MPXV), has made global headlines. While transmissibility (R0 ≈ 0.58) and fatality rate (0-3%) are low, as it causes prolonged morbidity, the World Health Organization has declared monkeypox as a public health emergency of international concern. Thus, effective containment and disease management require quick and efficient detection of MPXV. In this bioinformatic overview, we summarize the numerous molecular tests available for MPXV, and discuss the diversity of genes and primers used in the polymerase chain reaction-based detection. Over 90 primer/probe sets are used for the detection of poxviruses. While hemagglutinin and A-type inclusion protein are the most common target genes, tumor necrosis factor receptor and complement binding protein genes are frequently used for distinguishing Clade I and Clade II of MPXV. Problems and possibilities in the detection of MPXV have been discussed.


Subject(s)
Monkeypox , Humans , Monkeypox/diagnosis , Monkeypox/pathology , Monkeypox virus/genetics , Polymerase Chain Reaction , DNA, Viral/genetics , Public Health
4.
PLoS One ; 18(6): e0285083, 2023.
Article in English | MEDLINE | ID: covidwho-20235784

ABSTRACT

BACKGROUND/AIM: During the coronavirus disease (COVID-19) pandemic, Germany and various other countries experienced a shortage of polymerase chain reaction (PCR) laboratory tests due to the highly transmissible SARS-CoV-2 Omicron variant that drove an unprecedented surge of infections. This study developed a mathematical model that optimizes diagnostic capacity with lab-based PCR testing. METHODS: A mathematical model was constructed to determine the value of PCR testing in relation to the pre-test probability of COVID-19. Furthermore, the model derives the lower and upper bounds for the threshold pre-test probability of the designated priority group. The model was applied in a German setting using the PCR test-positivity rate at the beginning of February 2022. RESULTS: The value function of PCR testing is bell-shaped with respect to the pre-test probability, reaching a maximum at a pre-test probability of 0.5. Based on a PCR test-positivity rate of 0.3 and assuming that at least two thirds of the tested population have a pre-test probability below, lower and higher pre-test probability thresholds are ≥ 0.1 and 0.7, respectively. Therefore, individuals who have a 25% likelihood of testing positive because they exhibit symptoms should be a higher priority for PCR testing. Furthermore, a positive rapid antigen test in asymptomatic individuals with no known exposure to COVID-19 should be confirmed using PCR. Yet, symptomatic individuals with a positive RAT should be excluded from PCR testing. CONCLUSION: A mathematical model that allows for the optimal allocation of scarce PCR tests during the COVID-19 pandemic was developed.


Subject(s)
COVID-19 , SARS-CoV-2 , Humans , SARS-CoV-2/genetics , COVID-19/diagnosis , COVID-19/epidemiology , COVID-19 Testing/methods , Pandemics , Sensitivity and Specificity , Polymerase Chain Reaction
5.
Virol J ; 20(1): 119, 2023 Jun 08.
Article in English | MEDLINE | ID: covidwho-20235393

ABSTRACT

BACKGROUND: A variety of open-system real-time reverse transcriptase polymerase chain reaction (RT-PCR) assays for several acute respiratory syndrome coronavirus 2 are currently in use. This study aimed to ensure the quality of omicron nucleic acid testing and to assess the comparability of cycle threshold (Ct) values derived from RT-PCR. METHODS: Five external quality assessment (EQA) rounds using the omicron virus-like particles were organized between February 2022 and June 2022. RESULTS: A total of 1401 qualitative EQA reports have been collected. The overall positive percentage agreement was 99.72%, the negative percentage agreement was 99.75%, and the percent agreement was 99.73%. This study observed a significant variance in Ct values derived from different test systems. There was a wide heterogeneity in PCR efficiency among different RT-PCR kits and inter-laboratories. CONCLUSION: There was strong concordance among laboratories performing qualitative omicron nucleic acid testing. Ct values from qualitative RT-PCR tests should not be used for clinical or epidemiological decision-making to avoid the potential for misinterpretation of the results.


Subject(s)
COVID-19 , Nucleic Acids , Humans , COVID-19/diagnosis , SARS-CoV-2/genetics , COVID-19 Testing , Polymerase Chain Reaction , Reverse Transcriptase Polymerase Chain Reaction , Sensitivity and Specificity
6.
Emerg Infect Dis ; 29(5): 1007-1010, 2023 05.
Article in English | MEDLINE | ID: covidwho-20245370

ABSTRACT

Increasing reports of invasive Streptococcus pyogenes infections mandate surveillance for toxigenic lineage M1UK. An allele-specific PCR was developed to distinguish M1UK from other emm1 strains. The M1UK lineage represented 91% of invasive emm1 isolates in England in 2020. Allele-specific PCR will permit surveillance for M1UK without need for genome sequencing.


Subject(s)
Scarlet Fever , Streptococcal Infections , Humans , Streptococcus pyogenes/genetics , Scarlet Fever/epidemiology , Alleles , England/epidemiology , Streptococcal Infections/diagnosis , Streptococcal Infections/epidemiology , Polymerase Chain Reaction , Antigens, Bacterial/genetics , Bacterial Outer Membrane Proteins/genetics
7.
Int J Mol Sci ; 24(10)2023 May 12.
Article in English | MEDLINE | ID: covidwho-20241326

ABSTRACT

A next-generation sequencing (NGS) study identified a very high viral load of Torquetenovirus (TTV) in KD patients. We aimed to evaluate the feasibility of a newly developed quantitative species-specific TTV-PCR (ssTTV-PCR) method to identify the etiology of KD. We applied ssTTV-PCR to samples collected from 11 KD patients and 22 matched control subjects who participated in our previous prospective study. We used the NGS dataset from the previous study to validate ssTTV-PCR. The TTV loads in whole blood and nasopharyngeal aspirates correlated highly (Spearman's R = 0.8931, p < 0.0001, n = 33), supporting the validity of ssTTV-PCR. The ssTTV-PCR and NGS results were largely consistent. However, inconsistencies occurred when ssTTV-PCR was more sensitive than NGS, when the PCR primer sequences mismatched the viral sequences in the participants, and when the NGS quality score was low. Interpretation of NGS requires complex procedures. ssTTV-PCR is more sensitive than NGS but may fail to detect a fast-evolving TTV species. It would be prudent to update primer sets using NGS data. With this precaution, ssTTV-PCR can be used reliably in a future large-scale etiological study for KD.


Subject(s)
Mucocutaneous Lymph Node Syndrome , Torque teno virus , Humans , Torque teno virus/genetics , Mucocutaneous Lymph Node Syndrome/diagnosis , Mucocutaneous Lymph Node Syndrome/genetics , Polymerase Chain Reaction , Prospective Studies , High-Throughput Nucleotide Sequencing/methods
8.
East. Mediterr. health j ; 27(11): 1036-1044, 2021-11.
Article in English | WHOIRIS | ID: gwh-369361

ABSTRACT

Background:There are no data on the association between clinical course and comorbidity in Iranian patients with COVID-19.Aims:To determine noncommunicable disease (NCD), clinical characteristics and prognosis of patients hospitalized with COVID-19 in Isfahan, Islamic Republic of Iran.Methods:This multicentric retrospective observational study was performed on all patients hospitalized with COVID-19 in Isfahan from 17 February to 6 April 2020. We recruited 5055 patients. Data on clinical course and comorbid NCDs such as hypertension, coronary heart disease (CHD), diabetes mellitus (DM), cancer, chronic kidney disease (CKD) and chronic respiratory disease (CRD) were collected. Statistical analyses were done by Mann–Whitney U, χ2 and logistic regression tests using Stata version 14.Results:DM and hypertension were the most prevalent comorbidities in patients with positive and negative reverse transcription polymerase chain reaction (RT-PCR). Odds ratio (95% confidence interval) of mortality-associated factors was significant for DM [1.35 (1.07–1.70)], CHD [1.58 (1.26–1.96)], CRD [2.18 (1.58–3.0)], and cancer [3.55 (2.42–5.21)]. These results remained significant for cancer after adjustment for age, sex and clinical factors. Among patients with positive RT-PCR, death was significantly associated with CRD and cancer, while this association disappeared after adjustment for all potential confounders. There was a significant association between NCDs and higher occurrence of low oxygen saturation, mechanical ventilation requirement and intensive care unit admission after adjustment for age and sex.Conclusion:The presence of NCDs alone did not increase mortality in patients with COVID-19, after adjustment for all potential confounders including clinical factors.


Subject(s)
COVID-19 , Noncommunicable Diseases , Comorbidity , Logistic Models , Polymerase Chain Reaction , Intensive Care Units , Prognosis
9.
East. Mediterr. health j ; 28(4): 249-257, 2022-04.
Article in English | WHOIRIS | ID: gwh-368770

ABSTRACT

Background: Coronavirus disease 2019 (COVID-19) is a worldwide public health emergency. Aims: This study aimed to investigate the epidemiological and clinical characteristics of patients with COVID-19 in Saveh city, Islamic Republic of Iran in 2020. Methods: In this descriptive analytical research, 3181 patients suspected of having COVID-19 who visited Saveh med- ical centres were investigated. Patients were confirmed with COVID-19 using polymerase chain reaction testing. Data on sociodemographic, epidemiological and clinical characteristics of the patients were collected using a validated form through interviews and medical records. The chi-squared, t and Fisher exact tests were used to assess differences in socio- demographic, epidemiological and clinical characteristics between patients with positive and negative polymerase chain reaction results. Logistic regression analysis was used to examine the association between independent variables and death from COVID-19. Results: About half the patients (48.3%) had a history of chronic disease. Diabetes (16.2%), high blood pressure (14.8%) and cardiovascular disease (12.4%) were the most prevalent chronic diseases among patients who were confirmed positive for COVID-19. Risk factors for death among confirmed COVID-19 patients were: intubation (odds ratio (OR) = 8.97; 95% con- fidence interval (CI): 5.15–15.63), age ≥ 80 years (OR = 5.81; 95% CI: 1.91–17.60), oxygen saturation < 93% (OR = 2.48; 95% CI: 1.51–4.08), diabetes (OR = 1.88; 95% CI: 1.00–3.54) and shortness of breath (OR = 1.70; 95% CI: 1.02–2.82). Conclusion: Given the greater risks of contracting and dying from COVID-19 in certain groups of patients, health educa- tion programmes targeting these groups are recommended.


Subject(s)
COVID-19 , Betacoronavirus , Disease Outbreaks , Logistic Models , Odds Ratio , Cardiovascular Diseases , Polymerase Chain Reaction , Diabetes Mellitus , Hypertension
10.
East. Mediterr. health j ; 28(3): 175-182, 2022-03.
Article in English | WHOIRIS | ID: gwh-368762

ABSTRACT

Background: Clinical features of confirmed COVID-19 cases cover a wide spectrum. Aims: To study the clinical, radiological and virological features of the first 150 patients with COVID-19 in Lebanon. Methods: Our university hospital was designated as the primary COVID-19 care centre in Lebanon. Between 21 February 2020, the date of the first confirmed case of COVID-19 in Lebanon, and 3 April 2020, our team treated 150 patients diagnosed with COVID-19. In this prospective descriptive study, we present our experience in treating these patients, specifically the diagnostic criteria, outcome, and demographic, clinical, radiological and biological characteristics. Results: Ninety-five (63.33%) of the patients were male and 55 (36.67%) were female. Most patients (58%) were aged > 50 years, and 8 (5.33%) were healthcare workers. Diagnosis was based on reverse transcription polymerase chain reaction, and patients were classified as mild, moderate or critical. Fifteen (10%) patients had a critical presentation and fever was the most prominent symptom at presentation. One hundred and thirty-eight (92%) patients underwent radiological evaluation. The most common laboratory findings were lymphocytopenia (34.38%), followed by neutropenia (28.13%), but leukocytosis was not prevalent (1.56%). Old age and comorbidity were significant indicators in patient risk stratification. Chest computed tomography was an invaluable method of diagnosis and management. Our radiological findings were consistent with the published literature. Conclusion: Our study underlines the variable presentation of COVID-19, the difference in severity, and the diverse methods of diagnosis. This suggests the need for a tailored approach, taking into consideration the wide spectrum of presentation.


Subject(s)
COVID-19 , Betacoronavirus , Disease Outbreaks , Risk Assessment , Polymerase Chain Reaction , Health Personnel , Critical Care , Comorbidity , Demography
11.
Analyst ; 148(12): 2758-2766, 2023 Jun 12.
Article in English | MEDLINE | ID: covidwho-2323689

ABSTRACT

This paper introduces an enclosed microfluidic chip that integrates sample preparation and the chamber-based digital polymerase chain reaction (cdPCR). The sample preparation of the chip includes nucleic acid extraction and purification based on magnetic beads, which adsorb nucleic acids by moving around the reaction chambers to complete the reactions including lysis, washing, and elution. The cdPCR area of the chip consists of tens of thousands of regularly arranged microchambers. After the sample preparation processes are completed, the purified nucleic acid can be directly introduced into the microchambers for amplification and detection on the chip. The nucleic acid extraction performance and digital quantification performance of the system were examined using synthetic SARS-CoV-2 plasmid templates at concentrations ranging from 101-105 copies per µL. Further on, a simulated clinical sample was used to test the system, and the integrated chip was able to accurately detect SARS-CoV-2 virus particle samples doped with interference (saliva) with a detection limit of 10 copies per µL. This integrated system could provide a promising tool for point-of-care testing of pathogenic infections.


Subject(s)
Microfluidics , Microfluidics/methods , Polymerase Chain Reaction , Nucleic Acids/analysis , SARS-CoV-2/genetics , SARS-CoV-2/isolation & purification
12.
Nucleic Acids Res ; 51(11): e65, 2023 Jun 23.
Article in English | MEDLINE | ID: covidwho-2322793

ABSTRACT

Despite the need in various applications, accurate quantification of nucleic acids still remains a challenge. The widely-used qPCR has reduced accuracy at ultralow template concentration and is susceptible to nonspecific amplifications. The more recently developed dPCR is costly and cannot handle high-concentration samples. We combine the strengths of qPCR and dPCR by performing PCR in silicon-based microfluidic chips and demonstrate high quantification accuracy in a large concentration range. Importantly, at low template concentration, we observe on-site PCR (osPCR), where only certain sites of the channel show amplification. The sites have almost identical ct values, showing osPCR is a quasi-single molecule phenomenon. Using osPCR, we can measure both the ct values and the absolute concentration of templates in the same reaction. Additionally, osPCR enables identification of each template molecule, allowing removal of nonspecific amplification during quantification and greatly improving quantification accuracy. We develop sectioning algorithm that improves the signal amplitude and demonstrate improved detection of COVID in patient samples.


Subject(s)
COVID-19 Testing , Polymerase Chain Reaction , Humans , COVID-19 , DNA/genetics , Microfluidics
13.
BMC Microbiol ; 23(1): 135, 2023 05 17.
Article in English | MEDLINE | ID: covidwho-2321595

ABSTRACT

Mycophenolic acid (MPA) is the active ingredient in the most important immunosuppressive pharmaceuticals. It has antifungal, antibacterial, antiviral, anti-psoriasis, and antitumor activities. Therefore, its overproduction in addition to gene expression analysis was our main target. Through this study, we isolated a novel potent mycophenolic acid (MPA) producer strain of the genus Penicillium from the refrigerated Mozzarella cheese and it was identified with the molecular marker ITS and benA genes as P. arizonenseHEWt1. Three MPA overproducer mutants were isolated by exposing the wild type to different doses of gamma-rays, and the fermentation conditions for the highest production of MPA were optimized. The results indicated that MPA amounts produced by the mutants MT1, MT2, and MT3 were increased by 2.1, 1.7, and 1.6-fold, respectively, compared with the wild-type. The growth of both mutant and wild-type strains on PD broth, adjusted to pH 6 and incubated at 25 °C for 15 d, were the best conditions for maximum production of MPA. In a silico study, five orthologs genes of MPA biosynthesizing gene clusters in P. brevicompactum were predicted from the genome of P. arizonense. Sequencing and bioinformatic analyses proved the presence of five putative genes namely mpaA, mpaC, mpaF, mpaG, and mpaH in the P. arizonense HEWt1 genome. Gene expression analysis by qRT-PCR indicated an increase in the transcription value of all annotated genes in the three mutants over the wild type. A highly significant increase in the gene expression of mpaC, mpaF, and mpaH was observed in P. arizonense-MT1 compared with wild-type. These results confirmed the positive correlation of these genes in MPA biosynthesis and are the first report regarding the production of MPA by P. arizonense.Kew word.Mycophenolic acid, Penicillium arizonense, mutagenesis, gene expression.


Subject(s)
Mycophenolic Acid , Penicillium , Mycophenolic Acid/pharmacology , Mycophenolic Acid/metabolism , Immunosuppressive Agents , Penicillium/genetics , Polymerase Chain Reaction
14.
J Infect Chemother ; 29(8): 778-782, 2023 Aug.
Article in English | MEDLINE | ID: covidwho-2325110

ABSTRACT

BACKGROUND: For patients with coronavirus disease 2019 (COVID-19) requiring hospitalization, extending isolation is warranted. As a cautious protocol, ending isolation based on polymerase chain reaction cycle threshold (Ct) value was introduced for patients requiring therapy for >20 days after symptom onset. METHOD: We compared a Ct-based strategy using Smart Gene® between March 2022 and January 2023 with a preceding control period (March 2021 to February 2022) when two consecutive negative reverse transcription-polymerase chain reaction tests using FilmArray® were required for ending isolation. Ct was evaluated on day 21, and ending isolation was permitted in patients with Ct ≥ 38. Although patients with Ct 35-37 were transferred to a non-COVID-19 ward, isolation was continued. RESULTS: The duration of stay on a COVID-19 ward in the Ct group was 9.7 days shorter than that in controls. The cumulative number of tests was 3.7 in controls and 1.2 in the Ct group. There was no nosocomial transmission after ending isolation in either group. The number of days from symptom onset to testing was 20.7 ± 2.1 in Ct group, and five patients had Ct < 35, nine Ct 35-37, and 71 Ct ≥ 38. No patients were moderately or severely immunocompromised. Steroid use was an independent risk factor for prolonged low Ct (odds ratio 9.40, 95% confidence interval 2.31-38.15, p = 0.002) CONCLUSIONS: The efficacy of ending isolation based on Ct values could improve bed utilization without the risk of transmission among patients with COVID-19 requiring therapy for >20 days after symptom onset.


Subject(s)
COVID-19 , Humans , SARS-CoV-2/genetics , Reverse Transcriptase Polymerase Chain Reaction , Reverse Transcription , Hospitals , Polymerase Chain Reaction , COVID-19 Testing
15.
Sci Rep ; 13(1): 7612, 2023 05 10.
Article in English | MEDLINE | ID: covidwho-2318137

ABSTRACT

Epidemiologic surveillance of circulating SARS-CoV-2 variants is essential to assess impact on clinical outcomes and vaccine efficacy. Whole genome sequencing (WGS), the gold-standard to identify variants, requires significant infrastructure and expertise. We developed a digital droplet polymerase chain reaction (ddPCR) assay that can rapidly identify circulating variants of concern/interest (VOC/VOI) using variant-specific mutation combinations in the Spike gene. To validate the assay, 800 saliva samples known to be SARS-CoV-2 positive by RT-PCR were used. During the study (July 2020-March 2022) the assay was easily adaptable to identify not only existing circulating VAC/VOI, but all new variants as they evolved. The assay can discriminate nine variants (Alpha, Beta, Gamma, Delta, Eta, Epsilon, Lambda, Mu, and Omicron) and sub-lineages (Delta 417N, Omicron BA.1, BA.2). Sequence analyses confirmed variant type for 124/124 samples tested. This ddPCR assay is an inexpensive, sensitive, high-throughput assay that can easily be adapted as new variants are identified.


Subject(s)
COVID-19 , SARS-CoV-2 , Humans , SARS-CoV-2/genetics , COVID-19/diagnosis , COVID-19/epidemiology , Polymerase Chain Reaction , Clinical Decision-Making , Population Surveillance , COVID-19 Testing
16.
Biotechniques ; 74(4): 158-171, 2023 04.
Article in English | MEDLINE | ID: covidwho-2316281

ABSTRACT

The recent cases of COVID-19 have brought the prospect of and requirement for point-of-care diagnostic devices into the limelight. Despite all the advances in point-of-care devices, there is still a huge requirement for a rapid, accurate, easy-to-use, low-cost, field-deployable and miniaturized PCR assay device to amplify and detect genetic material. This work aims to develop an Internet-of-Things automated, integrated, miniaturized and cost-effective microfluidic continuous flow-based PCR device capable of on-site detection. As a proof of application, the 594-bp GAPDH gene was successfully amplified and detected on a single system. The presented mini thermal platform with an integrated microfluidic device has the potential to be used for the detection of several infectious diseases.


Subject(s)
COVID-19 , Humans , COVID-19/diagnosis , Nucleic Acid Amplification Techniques , Polymerase Chain Reaction , Lab-On-A-Chip Devices , DNA
17.
Pediatr Pulmonol ; 58(7): 2111-2123, 2023 Jul.
Article in English | MEDLINE | ID: covidwho-2314963

ABSTRACT

The reported prevalence of chronic lung disease (CLD) due to coronavirus 2 (severe acute respiratory syndrome coronavirus 2 [SARS-CoV-2)]) pneumonia with the severe acute respiratory syndrome in children is unknown and rarely reported in English literature. In contrast to most other respiratory viruses, children generally have less severe symptoms when infected with SARS-CoV-2. Although only a minority of children with SARS-CoV-2 infection require hospitalization, severe cases have been reported. More severe SARS-CoV-2 respiratory disease in infants has been reported in low- and middle-income countries (LMICs) compared to high-income countries (HICs). We describe our experience of five cases of CLD in children due to SARS-CoV-2 collected between April 2020 and August 2022. We included children who had a history of a positive SARS-CoV-2 polymerase chain reaction (PCR) or antigen test or a positive antibody test in the serum. Three patterns of CLD related to SARS-CoV-2 were identified: (1) CLD in infants postventilation for severe pneumonia (n = 3); (2) small airway disease with bronchiolitis obliterans picture (n = 1) and (3) adolescent with adult-like post-SARS-CoV-2 disease (n = 1). Chest computerized tomography scans showed airspace disease and ground-glass opacities involving both lungs with the development of coarse interstitial markings seen in four patients, reflecting the long-term fibrotic consequences of diffuse alveolar damage that occur in children post-SARS-CoV-2 infection. Children with SARS-CoV-2 infection mostly have mild symptoms with little to no long-term sequelae, but the severe long-term respiratory disease can develop.


Subject(s)
COVID-19 , SARS-CoV-2 , Infant , Adult , Adolescent , Humans , Child , COVID-19/complications , Lung/diagnostic imaging , Polymerase Chain Reaction , Hospitalization
18.
Vet Clin North Am Food Anim Pract ; 39(1): 129-140, 2023 Mar.
Article in English | MEDLINE | ID: covidwho-2320302

ABSTRACT

Laboratory testing is one part of clinical diagnosis, and quick and reliable testing results provide important data to support treatment decision and develop control strategies. Clinical viral testing has been shifting from traditional virus isolation and electron microscopy to molecular polymerase chain reaction and point-of-care antigen tests. This shift in diagnostic methodology also means change from looking for infectious virions or viral particles to hunting viral antigens and genomes. With technological development, it is predicted that metagenomic sequencing will be commonly used in veterinary clinical diagnosis for unveiling the whole picture of microbes involved in diseases in the future.


Subject(s)
Laboratories , Animals , Polymerase Chain Reaction/veterinary
19.
Microbiol Spectr ; 11(3): e0005523, 2023 Jun 15.
Article in English | MEDLINE | ID: covidwho-2319646

ABSTRACT

Coronavirus disease 2019, caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), poses a considerable threat to global public health. This study developed and evaluated a rapid, low-cost, expandable, and sequencing-free high-resolution melting (HRM) assay for the direct detection of SARS-CoV-2 variants. A panel of 64 common bacterial and viral pathogens that can cause respiratory tract infections was employed to evaluate our method's specificity. Serial dilutions of viral isolates determined the sensitivity of the method. Finally, the assay's clinical performance was assessed using 324 clinical samples with potential SARS-CoV-2 infection. Multiplex HRM analysis accurately identified SARS-CoV-2 (as confirmed with parallel reverse transcription-quantitative PCR [qRT-PCR] tests), differentiating between mutations at each marker site within approximately 2 h. For each target, the limit of detection (LOD) was lower than 10 copies/reaction (the LOD of N, G142D, R158G, Y505H, V213G, G446S, S413R, F486V, and S704L was 7.38, 9.72, 9.96, 9.96, 9.50, 7.80, 9.33, 8.25, and 8.25 copies/reaction, respectively). No cross-reactivity occurred with organisms of the specificity testing panel. In terms of variant detection, our results had a 97.9% (47/48) rate of agreement with standard Sanger sequencing. The multiplex HRM assay therefore offers a rapid and simple procedure for detecting SARS-CoV-2 variants. IMPORTANCE In the face of the current severe situation of increasing SARS-CoV-2 variants, we developed an upgraded multiplex HRM method for the predominant SARS-CoV-2 variants based on our original research. This method not only could identify the variants but also could be utilized in subsequent detection of novel variants since the assay has great performance in terms of flexibility. In summary, the upgraded multiplex HRM assay is a rapid, reliable, and economical detection method, which could better screen prevalent virus strains, monitor the epidemic situation, and help to develop measures for the prevention and control of SARS-CoV-2.


Subject(s)
COVID-19 , SARS-CoV-2 , Humans , SARS-CoV-2/genetics , COVID-19/diagnosis , Sensitivity and Specificity , Polymerase Chain Reaction
20.
Int J Mol Sci ; 24(9)2023 May 02.
Article in English | MEDLINE | ID: covidwho-2319617

ABSTRACT

Infectious uveitis is a vision-threatening condition that requires prompt clinical diagnosis and proper treatment. However, rapid and proper diagnosis in infectious uveitis remains challenging. Several examination tests, including polymerase chain reaction (PCR) tests, are transitioning from laboratory-based basic research-level tests to bedside clinical tests, and recently tests have changed to where they can be performed right next to clinicians. In this review, we introduce an updated overview of recent studies that are representative of the current trends in clinical microbiological techniques including PCR tests for infectious uveitis.


Subject(s)
Communicable Diseases , Eye Infections, Bacterial , Uveitis , Humans , Eye , Polymerase Chain Reaction/methods , Uveitis/diagnosis , Uveitis/microbiology , Communicable Diseases/diagnosis , Vision Disorders
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