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1.
Int J Hyperthermia ; 38(1): 202-212, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33682604

RESUMEN

Increased transmissibility of the pandemic severe acute respiratory coronavirus 2 (SARS-CoV-2) has been noted to occur at lower ambient temperatures. This is seemingly related to a better replication of most respiratory viruses, including SARS-CoV-2, at lower-than-core body temperatures (i.e., 33 °C vs 37 °C). Also, intrinsic characteristics of SARS-CoV-2 make it a heat-susceptible pathogen. Thermotherapy has successfully been used to combat viral infections in plants which could otherwise result in great economic losses; 90% of viruses causing infections in plants are positive-sense single-stranded ribonucleic acid (+ssRNA) viruses, a characteristic shared by SARS-CoV-2. Thus, it is possible to envision the use of heat-based interventions (thermotherapy or mild-temperature hyperthermia) in patients with COVID-19 for which moderate cycles (every 8-12 h) of mild-temperature hyperthermia (1-2 h) have been proposed. However, there are potential safety and mechanistic concerns which could limit the use of thermotherapy only to patients with mild-to-moderate COVID-19 to prevent disease progression rather than to treat patients who have already progressed to severe-to-critical COVID-19. Here, we review the characteristics of SARS-CoV-2 which make it a heat-susceptible virus, potential host mechanisms which could be enhanced at higher temperatures to aid viral clearance, and how thermotherapy could be investigated as a modality of treatment in patients with COVID-19 while taking into consideration potential risks.


Asunto(s)
/terapia , Hipertermia Inducida , Animales , Temperatura Corporal , Genes Virales , Humanos , Plantas/virología , Interferencia de ARN , /aislamiento & purificación
2.
Dokl Biochem Biophys ; 496(1): 27-31, 2021 May.
Artículo en Inglés | MEDLINE | ID: mdl-33689070

RESUMEN

The coronavirus family consists of lipid-containing envelope viruses that have a single-stranded RNA genome that encodes 25-30 proteins in different viruses by the mechanism of positive-polarity strategy. In addition, extended open reading trnslation frames (ORFs, genes) located in a negative-sense orientation were found in the genomes of coronaviruses. The size of negative-sense genes varies in the range of 150-450 nt, which corresponds to polypeptides encoded by negative-polarity genes (negative gene proteins, NGP) with m. m. 5-30 × 103 kDa. Coronaviruses show marked differences from virus to virus in the number of negative genes detected. These negative-sense genes in the coronavirus genome allow this family to be considered as viruses developing an ambisense genome strategy.


Asunto(s)
Genes Virales/genética , Genómica , ARN Viral/genética , /genética , Secuencia de Bases , Sistemas de Lectura Abierta/genética
3.
Sci Rep ; 11(1): 5372, 2021 03 08.
Artículo en Inglés | MEDLINE | ID: mdl-33686189

RESUMEN

Wastewater-based epidemiology (WBE) is a great approach that enables us to comprehensively monitor the community to determine the scale and dynamics of infections in a city, particularly in metropolitan cities with a high population density. Therefore, we monitored the time course of the SARS-CoV-2 RNA concentration in raw sewage in the Frankfurt metropolitan area, the European financial center. To determine the SARS-CoV-2 RNA concentration in sewage, we continuously collected 24 h composite samples twice a week from two wastewater treatment plant (WWTP) influents (Niederrad and Sindlingen) serving the Frankfurt metropolitan area and performed RT-qPCR analysis targeting three genes (N gene, S gene, and ORF1ab gene). In August, a resurgence in the SARS-CoV-2 RNA load was observed, reaching 3 × 1013 copies/day, which represented similar levels compared to April with approx. 2 × 1014 copies/day. This corresponds to a continuous increase again in COVID-19 cases in Frankfurt since August, with an average of 28.6 incidences, compared to 28.7 incidences in April. Different temporal dynamics were observed between different sampling points, indicating local dynamics in COVID-19 cases within the Frankfurt metropolitan area. The SARS-CoV-2 RNA load to the WWTP Niederrad ranged from approx. 4 × 1011 to 1 × 1015 copies/day, the load to the WWTP Sindlingen from approx. 1 × 1011 to 2 × 1014 copies/day, which resulted in a preceding increase in these loading in July ahead of the weekly averaged incidences. The study shows that WBE has the potential as an early warning system for SARS-CoV-2 infections and a monitoring system to identify global hotspots of COVID-19.


Asunto(s)
Monitoreo del Ambiente , ARN Viral/análisis , Aguas Residuales/virología , /epidemiología , Ciudades , Monitoreo Epidemiológico , Genes Virales , Alemania , Aguas del Alcantarillado/virología , Factores de Tiempo , Carga Viral , Purificación del Agua
4.
Arch Virol ; 166(5): 1337-1344, 2021 May.
Artículo en Inglés | MEDLINE | ID: mdl-33683473

RESUMEN

A reservoir of antibiotic resistance genes (ARGs) is present in pathogenic, commensal, and environmental bacteria as well as in mobile genetic elements, including bacteriophages. Wastewater treatment plants (WWTPs) are considered hotspots for the spread of ARGs. The aim of this work was to analyze the diversity of the highly prevalent ARGs blaCTX-M and blaTEM in bacterial and bacteriophage fractions associated with human and animal environments through the study of urban waste and animal residues discharged into WWTPs to provide information about the composition and maintenance of the current resistome in Buenos Aires, Argentina. The results showed that a putative extended-spectrum variant of the blaTEM gene was the most frequently detected, with blaTEM-116 being the most prevalent, while a recently described type, blaTEM-229, was also found. In the bacteriophage fraction, we detected blaCTX-M genes from four out of the five clusters described. The detection of blaCTX- M-9-like and blaCTX-M-25-like genes was unexpected based on surveys of the ARGs from clinical pathogens circulating regionally. The finding of divergent blaCTX-M sequences associated with previously reported environmental genes argues in favor of the natural environment as a reservoir of resistance genes. ARGs were detected in bacteriophages as frequently as in bacterial communities, and furthermore, the blaCTX-M genes were more diverse in the bacteriophage fraction. Bacteriophages might therefore play a role in the spread of ARGs in the environment, but they might also be used as "reporters" for monitoring circulating ARGs.


Asunto(s)
Bacteriófagos/genética , Aguas Residuales/virología , beta-Lactamasas/genética , Animales , Argentina , Bacterias/genética , Farmacorresistencia Microbiana/genética , Genes Bacterianos/genética , Genes Virales/genética , Variación Genética , Humanos , Filogenia , Aguas Residuales/microbiología , beta-Lactamasas/clasificación
5.
Biochem Biophys Res Commun ; 550: 8-14, 2021 04 23.
Artículo en Inglés | MEDLINE | ID: mdl-33676232

RESUMEN

The SARS-CoV-2 Variant of Concern 202012/01 (VOC-202012/01) emerged in southeast England and rapidly spread worldwide. This variant is believed to be more transmissible, with all attention being given to its spike mutations. However, VOC-202012/01 has also a mutation (Q27stop) that truncates the ORF8, a likely immune evasion protein. Removal of ORF8 changes the clinical outset of the disease, which may affect the virus transmissibility. Here I provide a detailed analysis of all reported ORF8-deficient lineages found in the background of relevant spike mutations, identified among 231,433 SARS-CoV-2 genomes. I found 19 ORF8 nonsense mutations, most of them occurring in the 5' half of the gene. The ORF8-deficient lineages were rare, representing 0.67% of sequenced genomes. Nevertheless, I identified two clusters of related sequences that emerged recently and spread in different countries. The widespread D614G spike mutation was found in most ORF-deficient lineages. Although less frequent, HV69-70del and L5F spike mutations occurred in the background of six different ORF8 nonsense mutations. I also confirmed that VOC-202012/01 is the ORF8-deficient variant with more spike mutations reported to date, although other variants could have up to six spike mutations, some of putative biological relevance. Overall, these results suggest that monitoring ORF8-deficient lineages is important for the progression of the COVID-19 pandemic, particularly when associated with relevant spike mutations.


Asunto(s)
/transmisión , Monitoreo Epidemiológico , Eliminación de Gen , Glicoproteína de la Espiga del Coronavirus/genética , Proteínas Virales/genética , /epidemiología , Codón sin Sentido , Codón de Terminación/genética , Evolución Molecular , Genes Virales/genética , Humanos , Filogenia , Selección Genética , Factores de Tiempo , Reino Unido/epidemiología
6.
Virus Res ; 297: 198398, 2021 05.
Artículo en Inglés | MEDLINE | ID: mdl-33753180

RESUMEN

Commercially available reverse transcription-polymerase chain reaction (RT-PCR) kits are being used as an important tool to diagnose SARS-CoV-2 infection in clinical laboratories worldwide. However, some kits lack sufficient clinical evaluation due to the need for emergency use caused by the current COVID-19 pandemic. Here we found that a novel insertion/deletion mutation in the nucleocapsid (N) gene of SARS-CoV-2 samples is a cause of negative results for the N gene in a widely used assay that received emergency use authorization (EUA) from US FDA and Conformite Europeenne-in vitro diagnostics (CE-IVD) from EU. Although SARS-CoV-2 is diagnosed positive by other target probes in the assay, our findings provide an evidence of the genetic variability and rapid evolution of SARS-CoV-2 as well as a reference in designing commercial RT-PCR assays.


Asunto(s)
/virología , Mutación INDEL , /genética , /diagnóstico , Reacciones Falso Negativas , Genes Virales , Humanos , Tamizaje Masivo , Pandemias , ARN Viral/análisis , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , /aislamiento & purificación
8.
Viruses ; 13(2)2021 02 08.
Artículo en Inglés | MEDLINE | ID: mdl-33567491

RESUMEN

African swine fever (ASF), caused by the African swine fever virus (ASFV), is a major epidemic disease endangering the swine industry. Although a number of vaccine candidates have been reported, none are commercially available yet. To explore the effect of unknown genes on the biological characteristics of ASFV and the possibility of a gene-deleted isolate as a vaccine candidate, the strain SY18ΔL7-11, with deletions of L7L-L11L genes from ASFV SY18, was constructed, and its biological properties were analyzed. The results show that deletion of genes L7L-L11L did not affect replication of the virus in vitro. Virulence of SY18△L7-11 was significantly reduced, as 11 of the 12 pigs survived for 28 days after intramuscular inoculation with a low dose (103 TCID50) or a high dose (106 TCID50) of SY18ΔL7-11. All 11 surviving pigs were completely protected against challenge with the parental ASFV SY18 on 28 days postinoculation (dpi). Transient fever and/or irregularly low levels of genomic DNA in the blood were monitored in some pigs after inoculation. No ASF clinical signs or viremia were monitored after challenge. Antibodies to ASFV were induced in all pigs from 14 to 21 days postinoculation. IFN-γ was detected in most of the inoculated pigs, which is usually inhibited in ASFV-infected pigs. Overall, the results demonstrate that SY18ΔL7-11 is a candidate for further constructing safer vaccine(s), with better joint deletions of other gene(s) related to virulence.


Asunto(s)
Virus de la Fiebre Porcina Africana/inmunología , Fiebre Porcina Africana/prevención & control , Genes Virales/genética , Vacunas Virales/genética , Virus de la Fiebre Porcina Africana/genética , Virus de la Fiebre Porcina Africana/patogenicidad , Animales , Anticuerpos Antivirales/sangre , Células Cultivadas , Eliminación de Gen , Inyecciones Intramusculares , Interferón gamma/sangre , Macrófagos/virología , Porcinos , Vacunas Atenuadas/administración & dosificación , Vacunas Atenuadas/genética , Vacunas Virales/administración & dosificación , Virulencia/genética
9.
Viruses ; 13(2)2021 02 09.
Artículo en Inglés | MEDLINE | ID: mdl-33572446

RESUMEN

Analysis of pooled genomic short read sequence data revealed the presence of nudivirus-derived sequences from U.S. populations of both southern corn rootworm (SCR, Diabrotica undecimpunctata howardi Barber) and western corn rootworm (WCR, Diabrotica virgifera virgifera LeConte). A near complete nudivirus genome sequence was assembled from sequence data for an SCR population with relatively high viral titers. A total of 147,179 bp was assembled from five contigs that collectively encode 109 putative open reading frames (ORFs) including 20 nudivirus core genes. In contrast, genome sequence recovery was incomplete for a second nudivirus from WCR, although sequences derived from this virus were present in three geographically dispersed populations. Only 48,989 bp were assembled with 48 putative ORFs including 13 core genes, representing about 20% of a typical nudivirus genome. Phylogenetic analysis indicated that both corn rootworm nudiviruses grouped with the third known nudivirus of beetles, Oryctes rhinoceros nudivirus in the genus Alphanudivirus. On the basis of phylogenetic and additional analyses, we propose further taxonomic separation of nudiviruses within Alphanudivirus and Betanudivirus into two subfamilies and five genera. Identification of nudivirus-derived sequences from two species of corn rootworm highlights the diversity of viruses associated with these agricultural insect pests.


Asunto(s)
Escarabajos/virología , Nudiviridae/genética , Animales , Escarabajos/clasificación , ADN Viral/genética , Genes Virales , Genoma Viral/genética , Genómica , Nudiviridae/clasificación , Sistemas de Lectura Abierta , Filogenia , /genética
10.
Sci Rep ; 11(1): 4108, 2021 02 18.
Artículo en Inglés | MEDLINE | ID: mdl-33602998

RESUMEN

In December 2019, rising pneumonia cases caused by a novel ß-coronavirus (SARS-CoV-2) occurred in Wuhan, China, which has rapidly spread worldwide, causing thousands of deaths. The WHO declared the SARS-CoV-2 outbreak as a public health emergency of international concern, since then several scientists are dedicated to its study. It has been observed that many human viruses have codon usage biases that match highly expressed proteins in the tissues they infect and depend on the host cell machinery for the replication and co-evolution. In this work, we analysed 91 molecular features and codon usage patterns for 339 viral genes and 463 human genes that consisted of 677,873 codon positions. Hereby, we selected the highly expressed genes from human lung tissue to perform computational studies that permit to compare their molecular features with those of SARS, SARS-CoV-2 and MERS genes. The integrated analysis of all the features revealed that certain viral genes and overexpressed human genes have similar codon usage patterns. The main pattern was the A/T bias that together with other features could propitiate the viral infection, enhanced by a host dependant specialization of the translation machinery of only some of the overexpressed genes. The envelope protein E, the membrane glycoprotein M and ORF7 could be further benefited. This could be the key for a facilitated translation and viral replication conducting to different comorbidities depending on the genetic variability of population due to the host translation machinery. This is the first codon usage approach that reveals which human genes could be potentially deregulated due to the codon usage similarities between the host and the viral genes when the virus is already inside the human cells of the lung tissues. Our work leaded to the identification of additional highly expressed human genes which are not the usual suspects but might play a role in the viral infection and settle the basis for further research in the field of human genetics associated with new viral infections. To identify the genes that could be deregulated under a viral infection is important to predict the collateral effects and determine which individuals would be more susceptible based on their genetic features and comorbidities associated.


Asunto(s)
Betacoronavirus/genética , Infecciones por Coronavirus/genética , Infecciones por Coronavirus/virología , Codón/genética , Uso de Codones , Biología Computacional/métodos , Coronavirus/genética , Infecciones por Coronavirus/metabolismo , Genes Virales , Genoma Viral , Humanos , Coronavirus del Síndrome Respiratorio de Oriente Medio/genética , Filogenia , Virus del SRAS/genética , /genética
11.
Arch Virol ; 166(5): 1485-1488, 2021 May.
Artículo en Inglés | MEDLINE | ID: mdl-33620554

RESUMEN

Fowlpox virus (FWPV), which is the type member of the genus Avipoxvirus, subfamily Chordopoxvirinae, family Poxviridae, can lead to significant losses to the poultry industry. Although a large number of fowlpox virus genomes have been sequenced and characterised globally, there are no sequences available at the genomic level from Australian isolates. Here, we present the first complete genome sequence of a fowlpox virus vaccine strain (FWPV-S) containing an integrated near-full-length reticuloendotheliosis virus (REV) provirus. The genome of FWPV-S showed the highest sequence similarity to a fowlpox virus from the USA (97.74% identity). The FWPV-S genome contained 16 predicted unique genes, while a further two genes were fragmented compared to previously reported FWPV genome sequences. Subsequent phylogenetic analysis showed that FWPV-S was most closely related to other fowlpox viruses. This is the first reported genome sequence of FWPV from Australia.


Asunto(s)
Virus de la Viruela de las Aves de Corral/genética , Provirus/genética , Virus de la Reticuloendoteliosis/genética , Vacunas Virales/genética , Animales , Australia , Secuencia de Bases , Células Cultivadas , Embrión de Pollo , ADN Viral/genética , Virus de la Viruela de las Aves de Corral/clasificación , Virus de la Viruela de las Aves de Corral/aislamiento & purificación , Genes Virales , Genoma Viral/genética , Sistemas de Lectura Abierta , Filogenia , Vacunas Virales/clasificación , Vacunas Virales/aislamiento & purificación , Integración Viral
12.
Brief Bioinform ; 22(2): 1430-1441, 2021 03 22.
Artículo en Inglés | MEDLINE | ID: mdl-33569598

RESUMEN

The COVID-19 disease led to an unprecedented health emergency, still ongoing worldwide. Given the lack of a vaccine or a clear therapeutic strategy to counteract the infection as well as its secondary effects, there is currently a pressing need to generate new insights into the SARS-CoV-2 induced host response. Biomedical data can help to investigate new aspects of the COVID-19 pathogenesis, but source heterogeneity represents a major drawback and limitation. In this work, we applied data integration methods to develop a Unified Knowledge Space (UKS) and used it to identify a new set of genes associated with SARS-CoV-2 host response, both in vitro and in vivo. Functional analysis of these genes reveals possible long-term systemic effects of the infection, such as vascular remodelling and fibrosis. Finally, we identified a set of potentially relevant drugs targeting proteins involved in multiple steps of the host response to the virus.


Asunto(s)
Antivirales/uso terapéutico , /tratamiento farmacológico , /genética , /virología , Genes Virales , Humanos , /aislamiento & purificación , Transcriptoma
13.
Methods Mol Biol ; 2244: 133-158, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33555586

RESUMEN

To fully understand the function of cytomegalovirus (CMV) genes, it is imperative that they are studied in the context of infection. Therefore, the targeted deletion of individual viral genes and the comparison of these loss-of-function viral mutants to the wild-type virus allow for the identification of the relevance and role for a particular gene in the viral replication cycle. Targeted CMV mutagenesis has made huge advances over the past 20 years. The cloning of CMV genomes into Escherichia coli as bacterial artificial chromosomes (BAC) allows for not only quick and efficient deletion of viral genomic regions, individual genes, or single-nucleotide exchanges in the viral genome but also the insertion of heterologous genetic sequences for gain-of-function approaches. The conceptual advantage of this strategy is that it overcomes the restrictions of recombinant technologies in cell culture systems. Namely, recombination in infected cells occurs only in a few clones, and their selection is not possible if the targeted genes are relevant for virus replication and are not able to compete for growth against the unrecombined parental viruses. On the other hand, BAC mutagenesis enables the selection for antibiotic resistance in E. coli, providing selective growth advantage to the recombined genomes and thus clonal selection of viruses with even extremely poor fitness. Here we describe the methods used for the generation of a CMV BAC, targeted mutagenesis of BAC clones, and transfection of human cells with CMV BAC DNA in order to reconstitute the viral infection process.


Asunto(s)
Cromosomas Artificiales Bacterianos/genética , Clonación Molecular/métodos , Citomegalovirus/genética , Células Cultivadas , Escherichia coli/genética , Genes Virales/genética , Genoma Viral/genética , Humanos , Mutagénesis/genética , Transfección/métodos , Replicación Viral/genética
14.
ACS Synth Biol ; 10(2): 379-390, 2021 02 19.
Artículo en Inglés | MEDLINE | ID: mdl-33534552

RESUMEN

Generating and characterizing immunoreagents to enable studies of novel emerging viruses is an area where ensembles of synthetic genes, recombinant antibody pipelines, and modular antibody-reporter fusion proteins can respond rapidly. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continues to spread through the global population causing widespread morbidity, mortality, and socioeconomic chaos. Using SARS-CoV-2 as our model and starting with a gBlocks encoded nucleocapsid (N) gene, we purified recombinant protein from E. coli, to serve as bait for selecting semisynthetic nanobodies from our Nomad single-pot library. Clones were isolated in days and first fused to Gaussia luciferase to determine EC50 in the tens of nM range, and second fused to the ascorbate peroxidase derivative APEX2 for sensitive detection of SARS-CoV-2 infected cells. To generate inherently fluorescent immunoreagents, we introduce novel periplasmic sdAb fusions made with mNeonGreen and mScarlet-I, which were produced at milligram amounts. The fluorescent fusion proteins enabled concise visualization of SARS-CoV-2 N in the cytoplasm but not in the nucleus 24 h post infection, akin to the distribution of SARS-CoV N, thereby validating these useful imaging tools. SdAb reactivity appeared specific to SARS-CoV-2 with very much weaker binding to SARS-CoV, and no noticeable cross-reactivity to a panel of overexpressed human codon optimized N proteins from other CoV. High periplasmic expression levels and in silico immortalization of the nanobody constructs guarantees a cost-effective and reliable source of SARS-CoV-2 immunoreagents. Our proof-of-principle study should be applicable to known and newly emerging CoV to broaden the tools available for their analysis and help safeguard human health in a more proactive than reactive manner.


Asunto(s)
/epidemiología , /genética , Sondas Moleculares/genética , Pandemias , /inmunología , Anticuerpos Antivirales/genética , Especificidad de Anticuerpos/genética , Enfermedades Transmisibles Emergentes/virología , Escherichia coli/genética , Técnica del Anticuerpo Fluorescente , Genes Sintéticos , Genes Virales , Células HEK293 , Humanos , Sondas Moleculares/inmunología , Pandemias/prevención & control , Biblioteca de Péptidos , Fosfoproteínas/genética , Fosfoproteínas/inmunología , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/inmunología , Anticuerpos de Dominio Único/genética , Biología Sintética
15.
J Vet Sci ; 22(1): e12, 2021 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-33522164

RESUMEN

BACKGROUND: Bats have been considered natural reservoirs for several pathogenic human coronaviruses (CoVs) in the last two decades. Recently, a bat CoV was detected in the Republic of Korea; its entire genome was sequenced and reported to be genetically similar to that of the severe acute respiratory syndrome CoV (SARS-CoV). OBJECTIVES: The objective of this study was to compare the genetic sequences of SARS-CoV, SARS-CoV-2, and the two Korean bat CoV strains 16BO133 and B15-21, to estimate the likelihood of an interaction between the Korean bat CoVs and the human angiotensin-converting enzyme 2 (ACE2) receptor. METHODS: The phylogenetic analysis was conducted with the maximum-likelihood (ML) method using MEGA 7 software. The Korean bat CoVs receptor binding domain (RBD) of the spike protein was analyzed by comparative homology modeling using the SWISS-MODEL server. The binding energies of the complexes were calculated using PRODIGY and MM/GBGA. RESULTS: Phylogenetic analyses of the entire RNA-dependent RNA polymerase, spike regions, and the complete genome revealed that the Korean CoVs, along with SARS-CoV and SARS-CoV-2, belong to the subgenus Sarbecovirus, within BetaCoVs. However, the two Korean CoVs were distinct from SARS-CoV-2. Specifically, the spike gene of the Korean CoVs, which is involved in host infection, differed from that of SARS-CoV-2, showing only 66.8%-67.0% nucleotide homology and presented deletions within the RBD, particularly within regions critical for cross-species transmission and that mediate interaction with ACE2. Binding free energy calculation revealed that the binding affinity of Korean bat CoV RBD to hACE2 was drastically lower than that of SARS-CoV and SARS-CoV-2. CONCLUSIONS: These results suggest that Korean bat CoVs are unlikely to bind to the human ACE2 receptor.


Asunto(s)
Quirópteros/virología , Coronavirus/genética , Virus del SRAS/genética , /genética , Animales , Genes Virales/genética , Genoma Viral/genética , Genómica , Humanos , Funciones de Verosimilitud , Filogenia , Receptor de Angiotensina Tipo 2/genética , Receptor de Angiotensina Tipo 2/metabolismo , República de Corea , Análisis de Secuencia de ADN , Homología de Secuencia , Glicoproteína de la Espiga del Coronavirus/genética , Acoplamiento Viral
16.
Euro Surveill ; 26(5)2021 02.
Artículo en Inglés | MEDLINE | ID: mdl-33541485

RESUMEN

In June-November 2020, SARS-CoV-2-infected mink were detected in 290 of 1,147 Danish mink farms. In North Denmark Region, 30% (324/1,092) of people found connected to mink farms tested SARS-CoV-2-PCR-positive and approximately 27% (95% confidence interval (CI): 25-30) of SARS-CoV-2-strains from humans in the community were mink-associated. Measures proved insufficient to mitigate spread. On 4 November, the government ordered culling of all Danish mink. Farmed mink constitute a potential virus reservoir challenging pandemic control.


Asunto(s)
Animales Salvajes/virología , /veterinaria , Brotes de Enfermedades/veterinaria , Reservorios de Enfermedades/veterinaria , Transmisión de Enfermedad Infecciosa/veterinaria , Visón/virología , Pandemias/veterinaria , /aislamiento & purificación , /transmisión , Animales , /virología , Dinamarca/epidemiología , Brotes de Enfermedades/estadística & datos numéricos , Reservorios de Enfermedades/virología , Granjas , Genes Virales , Humanos , Incidencia , Reacción en Cadena de la Polimerasa , Salud Pública , ARN Viral/análisis , ARN Viral/genética , /virología , Secuenciación Completa del Genoma , Zoonosis/transmisión , Zoonosis/virología
17.
Clin Lab ; 67(2)2021 Feb 01.
Artículo en Inglés | MEDLINE | ID: mdl-33616332

RESUMEN

BACKGROUND AND METHODS: 2019 Corona Virus Disease (COVID-19) caused by SARS-CoV-2 is still pandemic now. RT-qPCR detection was the most common method for the diagnosis of SARS-CoV-2 infection, facilitated by amounts of nucleic acid testing kits. However, the accuracy of nucleic acid detection is affected by various factors such as specimen collection, specimen preparation, reagents deficiency, and personnel quality. RESULTS: In this study, we found that unmatched virus preservation solution will inhibit N gene and OFR-1ab gene (two independent genes of SARS-CoV-2) amplification in one-step detection reagent. CONCLUSIONS: Despite just being a particular phenomenon we found in our work to fight 2019-nCoV, we concluded that unmatched virus preservation solution may have an inhibitory effect on SARS-CoV-2 nucleic acid detection which may lead to incorrect clinical diagnosis.


Asunto(s)
/métodos , Genes Virales/efectos de los fármacos , Soluciones Preservantes de Órganos/farmacología , Manejo de Especímenes , /diagnóstico , Errores Diagnósticos/prevención & control , Humanos , Juego de Reactivos para Diagnóstico/normas , /aislamiento & purificación , Manejo de Especímenes/efectos adversos , Manejo de Especímenes/métodos
18.
J Med Virol ; 93(4): 2538-2542, 2021 04.
Artículo en Inglés | MEDLINE | ID: mdl-33415765

RESUMEN

The occurrence of the COVID-19 second-wave outbreak in Europe has pushed laboratories performing molecular SARS-CoV-2 tests to increase their throughput and decrease the result rendering time. In this evaluation, we tested for the first time a new, extraction-free, protocol with the Allplex SARS-CoV-2 Assay RT-qPCR kit on a Nimbus platform. Ninety-one samples, of which 71 previously tested positive with RT-qPCR with extraction were immediately analyzed without extraction, using only a dilution and thermal shock protocol. The positive and negative percentage agreements were respectively 97.2% (95% confidence interval [CI]: 0.90-0.99) and 95.0% (95% CI: 0.76-0.99). The two false negatives observed were very weakly positive with the comparison method. Moderate variations in Ct of the targeted genes were observed (median ± 95% CI): E gene, +2.49 ± 0.44; N gene, +0.98 ± 0.54; RdRP/S genes, +2.64 ± 0.48. On the other hand, the number of tests performed within 24 h raised from 86.4% to 97.8%, the turn-around time decreased from 19:18 to 09:03 (p < .0001), and the number of tests that can be performed per day doubled since this technique was introduced routinely in our laboratory.


Asunto(s)
/métodos , Reacción en Cadena en Tiempo Real de la Polimerasa/métodos , /aislamiento & purificación , /diagnóstico , /genética , Brotes de Enfermedades , Europa (Continente) , Genes Virales , Humanos , Fosfoproteínas/genética , ARN Viral/análisis , ARN Viral/genética , Sensibilidad y Especificidad
19.
Biosens Bioelectron ; 178: 113015, 2021 Apr 15.
Artículo en Inglés | MEDLINE | ID: mdl-33493896

RESUMEN

Dependable, specific and rapid diagnostic methods for severe acute respiratory syndrome ß-coronavirus (SARS-CoV-2) detection are needed to promote public health interventions for coronavirus disease 2019 (COVID-19). Herein, we have established an entropy-driven amplified electrochemiluminescence (ECL) strategy to detect the RNA-dependent RNA polymerase (RdRp) gene of SARS-CoV-2 known as RdRp-COVID which as the target for SARS-CoV-2 plays an essential role in the diagnosis of COVID-19. For the construction of the sensors, DNA tetrahedron (DT) is modified on the surface of the electrode to furnish robust and programmable scaffolds materials, upon which target DNA-participated entropy-driven amplified reaction is efficiently conducted to link the Ru (bpy)32+ modified S3 to the linear ssDNA at the vertex of the tetrahedron and eventually present an "ECL on" state. The rigid tetrahedral structure of the DT probe enhances the ECL intensity and avoids the cross-reactivity between single-stranded DNA, thus increasing the sensitivity of the assays. The enzyme-free entropy-driven reaction prevents the use of expensive enzyme reagents and facilitates the realization of large-scale screening of SARS-CoV-2 patients. Our DT-based ECL sensor has demonstrated significant specificity and high sensitivity for SARS-CoV-2 with a limit of detection (LOD) down to 2.67 fM. Additionally, our operational method has achieved the detection of RdRp-COVID in human serum samples, which supplies a reliable and feasible sensing platform for the clinical bioanalysis.


Asunto(s)
Técnicas Biosensibles/instrumentación , /diagnóstico , /genética , /aislamiento & purificación , Técnicas Biosensibles/estadística & datos numéricos , /sangre , ADN/química , Técnicas Electroquímicas , Entropía , Genes Virales , Humanos , Límite de Detección , Luminiscencia , Conformación de Ácido Nucleico , Pandemias , Sensibilidad y Especificidad
20.
Sci Rep ; 11(1): 2261, 2021 01 26.
Artículo en Inglés | MEDLINE | ID: mdl-33500503

RESUMEN

The diagnosis of COVID-19 relies on the direct detection of SARS-CoV-2 RNA in respiratory specimens by RT-PCR. The pandemic spread of the disease caused an imbalance between demand and supply of materials and reagents needed for diagnostic purposes including swab sets. In a comparative effectiveness study, we conducted serial follow-up swabs in hospitalized laboratory-confirmed COVID-19 patients. We assessed the diagnostic performance of an in-house system developed according to recommendations by the US CDC. In a total of 96 serial swabs, we found significant differences in the accuracy of the different swab systems to generate a positive result in SARS-CoV-2 RT-PCR, ranging from around 50 to 80%. Of note, an in-house swab system was superior to most commercially available sets as reflected by significantly lower Ct values of viral genes. Thus, a simple combination of broadly available materials may enable diagnostic laboratories to bypass global limitations in the supply of swab sets.


Asunto(s)
/instrumentación , Equipos Desechables/provisión & distribución , Técnicas de Diagnóstico Molecular/instrumentación , /aislamiento & purificación , /métodos , Técnicas de Laboratorio Clínico , Pruebas Diagnósticas de Rutina , Genes Virales , Humanos , Técnicas de Diagnóstico Molecular/métodos , Control de Calidad , ARN Viral/análisis , Reproducibilidad de los Resultados , Asignación de Recursos , Manejo de Especímenes
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