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It is accepted that cancer stem cells (CSCs) are key to the occurrence, progression, drug resistance, and recurrence of bladder cancer (BLCA). Here, we aimed to characterize the landscapes of CSCs and investigate the biological and clinical signatures based on a prognostic model constructed by genes associated with CSCs. The malignant epithelial cells were discovered and sorted into six clusters through single cell analysis. C2 was identified as the CSCs. The signaling involved in the interactions between C2, cancer-associated fibroblasts (CAFs), and immune cells mainly consisted of MK, THBS, ANGPTL, VISFATIN, JAM, and ncWNT pathways. The CSC-like prognostic index (CSCLPI) constructed by the random survival forest was a reliable risk factor for BLCA and had a stable and powerful effect on predicting the overall survival of patients with BLCA. The level of CAFs was higher among patients with higher CSCLPI scores, suggesting that CAFs play a significant role in regulating biological characteristics. The CSCLPI-developed survival prediction nomogram has the potential to be applied clinically to predict the 1-, 2-, 3-, and 5-year overall survival of patients with BLCA. The CSCLPI can be used for prognostic prediction and drug treatment evaluation in the clinic.
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BACKGROUND: Our aim is to evaluate the relationship between prevalence of kidney stones (KS) and novel anthropometric indices (AHIs). METHODS: Participants who participated in the KS questionnaire was extracted from the National Health and Nutrition Examination Survey (NHANES) from 2007 to 2018.A series of covariates were also obtained. The novel AHIs include a body shape index (ABSI) and body roundness index (BRI). Weighted multivariable-adjusted logistic regression was performed to investigate the association of KS with AHIs. RESULTS: After relative covariates were adjusted, a greater risk of KS for each z score increase in ABSI (OR = 1.13, 95%CI 1.05-1.22), and the risk of KS augmented by 19% for every 1 BRI z score added (OR = 1.19, 95%CI 1.11-1.27). The results from subgroup analysis showed that among adults aged 20-39 (OR = 1.31, 95%CI 1.04-1.65), male (OR = 1.14, 95%CI 1.02-1.28), the risk of KS is higher with the increase of each ABSI z score. Raising each BRI z score in those who were male aged 20-39 and 40-59 resulted in a higher risk of KS (aged 20-39: OR = 1.34, 95%CI 1.06-1.69; aged 40-59: OR = 1.29, 95%CI 1.09-1.53). In female aged 40-59, increasing each BRI z score led to a higher risk of KS (OR = 1.23, 95%CI 1.07-1.41). A linear association of ABSI z score with the risk of KS and a non-linear relationship between BRI z score and the risk of KS were discovered. CONCLUSION: This study found that the novel AHIs was related to the risk of kidney stones, and can be used as important indicators to evaluate the risk of KS.
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Cálculos Renales , Obesidad , Adulto , Humanos , Masculino , Femenino , Encuestas Nutricionales , Obesidad/epidemiología , Factores de Riesgo , Índice de Masa Corporal , Prevalencia , Cálculos Renales/epidemiología , Cálculos Renales/complicacionesRESUMEN
Costimulatory molecules were considered to be promising and important targets in immunotherapy for various cancers. The present study was intended for generating a costimulatory molecule signature in kidney renal clear cell carcinoma (KIRC), to investigate prognostic implication, elucidate immune atlas, and predict immunotherapy response. All the KIRC samples from the TCGA were randomly divided into the training dataset and the testing dataset in the ratio of 7:3. The Cox and least absolute shrinkage and selection operator (LASSO) regression analysis were used to identify 7 key costimulatory molecules which were associated with prognosis and construct a costimulatory molecule prognostic index (CMsPI), which was validated by internal and external datasets and an independent cohort. Patients in the high-CMsPI group had high mortality. Mutation analysis showed the most common mutational genes and variant types. Immune analysis demonstrated CD8+ T cells were infiltrated at a high level in the high-CMsPI group. In combination of analysis of the immune relevant gene signature and the biomarkers of immunotherapy, we may infer there were more dysfunctional CD8+ T cells in the high-CMsPI group, and the patients of this group were less sensitive to immunotherapy. A nomogram was constructed, and the concordance index was 0.77 (95% CI: 0.74-0.79). Three key signaling pathways were identified to facilitate tumor progression. The CMsPI can be regarded as a promising biomarker for predicting individual prognosis and assessing immunotherapy response in KIRC patients.
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Carcinoma de Células Renales , Neoplasias Renales , Linfocitos T CD8-positivos/metabolismo , Carcinoma de Células Renales/genética , Carcinoma de Células Renales/metabolismo , Carcinoma de Células Renales/terapia , Femenino , Humanos , Inmunoterapia , Riñón , Neoplasias Renales/genética , Neoplasias Renales/metabolismo , Neoplasias Renales/terapia , Masculino , Pronóstico , Factores de Transcripción/genéticaRESUMEN
OBJECTIVE: To assess the association of C677T polymorphism of methylenetetrahydrofolate reductase (MTHFR) gene with autistic behavior and inheritance pattern of children patients. METHODS: Ninety three autism patients were selected as the study group, whilst 93 healthy children were selected as the control group. The C677T genotype of the MTHFR gene was determined, and the correlation between the genotype and the autistic behavior and inheritance pattern were investigated. RESULTS: MTHFR gene C677T locus revealed three genotypes CC, CT and TT. Compared with the control group, the study group had fewer CC genotype but more TT genotype (P<0.05). Individuals with the three genotypes showed a statistically significant difference in the frequencies of four problem behaviors (P<0.05). Regression analysis showed that at least one T allele encoding the degree of 1 and 2 for the 4 problem behaviors that were statistically different. MTHFR gene C677T genotype was associated with autism under the recessive inheritance model and allelic inheritance model (P<0.05). CONCLUSION: The C677T polymorphism of the MTHFR gene is associated with autistic behaviors. Children with the TT genotype or T allele are at higher risk of developing autism, particularly direct gaze, complex limb movements, self-injurious behavior and hyperactivity 1 and 2 related with the degree of coding.
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Trastorno Autístico , Problema de Conducta , Trastorno Autístico/genética , Niño , Predisposición Genética a la Enfermedad , Genotipo , Humanos , Patrón de Herencia , Metilenotetrahidrofolato Reductasa (NADPH2)/genéticaRESUMEN
A colorimetric method is presented for the detection of specific nucleotide sequences in plant pathogens. It is based on the use of CRISPR/Cas9-triggered isothermal amplification and gold nanoparticles (AuNPs) as optical probes. The target DNA was recognized and broken up by a given Cas9/sgRNA complex. After isothermal amplification, the product was hybridized with oligonucleotide-functionalized AuNPs. This resulted in the aggregation of AuNPs and a color change from wine red to purple. The visual detection limit is 2 pM of DNA, while a linear relationship exists between the ratio of absorbance at 650 and 525 nm and the DNA concentration in the range from 0.2 pM to 20 nM. In contrast to the previous CRISPR-based amplification platforms, the method has significantly higher specificity with the single-base mismatch and can be visually read out. It was successfully applied to identify the Phytophthora infestans genomic DNA. Graphical abstract Schematic presentation of a colorimetric method for detection of Phytophthora infestans genomic DNA based on CRISPR/Cas9-triggered isothermal amplification. The Cas9 endonuclease cleaves DNA at the design site and the color changes from red to purple with increasing target DNA concentration.
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Proteína 9 Asociada a CRISPR/química , Sistemas CRISPR-Cas , Colorimetría/métodos , ADN/análisis , Secuencia de Bases , Técnicas Biosensibles/métodos , ADN/genética , Oro/química , Límite de Detección , Nanopartículas del Metal/química , Técnicas de Amplificación de Ácido Nucleico/métodos , Hibridación de Ácido Nucleico , Oligodesoxirribonucleótidos/química , Oligodesoxirribonucleótidos/genética , Phytophthora infestans/genéticaRESUMEN
OBJECTIVE: To observe the relationship between the methylation status in promoter region of uncoupling protein 1( UCP1) gene and obesity. METHODS: A casecontrol study based on the hospital consisted 116 people was carried out, according to the body mass index( BMI), the subjects were divided into two groups, the overweight and obesity group with 50 samples( BMI≥24. 0) and the normal weight group with 66 samples( 18. 5≤BMI < 24), DNA samples were extracted from white blood cell and treated by hydrogen sulfite. Then mass spectrometry method was used to quantificationally detect the methylation level UCP1 gene promoter. The difference between the two groups was compared and the relationships between CpG sites and BMI were explored. RESULTS: Statistical analysis showed that the methylation differences between normal weight and overweight or obese group were not statistically significant, however, the CpG site UCP1-2_ Cp G_ 10. 11. 12. 13 had statistical significance in correlation coefficients with BMI according to multiple linear regression method( regression coefficient was 15. 370, P <0. 05). CONCLUSION: The UCP1 gene promoter methylation may be a factor for adult obesity.
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Proteínas Portadoras/genética , Metilación de ADN , Obesidad/genética , Proteína Desacopladora 1/genética , Adulto , Índice de Masa Corporal , Estudios de Casos y Controles , Humanos , Sobrepeso , Reacción en Cadena de la Polimerasa , Proteína Desacopladora 1/metabolismoRESUMEN
Zymoseptoria tritici is a globally distributed plant-pathogenic fungus causing Septoria tritici blotch of wheat. In this study, the in vitro growth rates and aggressiveness of 141 genetically distinct isolates sampled from four wheat fields on three continents were assessed to determine the association of these two ecological parameters. Aggressiveness was assessed on two spring wheat cultivars ('Toronit' and 'Greina') in a greenhouse using percentages of leaf area covered by lesions and pycnidia. We found a positive correlation between aggressiveness of pathogen strains on the two cultivars, consistent with a quantitative and host-nonspecific interaction in this pathosystem. We also found a positive correlation between aggressiveness and average growth rate at two temperatures, suggesting that in vitro pathogen growth rate may make a significant contribution to pathogen aggressiveness.
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Ascomicetos , Enfermedades de las Plantas/microbiología , Triticum/microbiología , Ascomicetos/crecimiento & desarrollo , Ascomicetos/patogenicidad , Hojas de la Planta/microbiología , TemperaturaRESUMEN
OBJECTIVE: To detect the gene-based gene-gene interaction by kernel canonical correlation analysis. METHODS: Based on case-control study, statistical simulation studies were conducted to construct and test the KCCU statistic to evaluate gene-based gene-gene interaction of gene FTO and gene PRDMl6 by kernel canonical correlation analysis. RESULTS: The power of KCCU statistic was related to significant level, sample size, minor allele frequency, and it was higher when the gene-gene interaction increased. The power arrived 0.8 at the significant level of 0.05 when the minor allele frequency was higher than 0.05, the interaction odds ratio was higher than 1.5, and the sample size was greater than 5000. CONCLUSION: KCCU statistic is a valid and powerful statistical inference method for detecting gene-based gene-gene interaction in the large sample analysis with high interaction.
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Genes/genética , Estudio de Asociación del Genoma Completo/métodos , Estudios de Casos y Controles , Frecuencia de los Genes , Humanos , Oportunidad RelativaRESUMEN
BACKGROUND: The aim of the study was to assess the relationship between prevalence of kidney stones (KS) and the oxidative balance score (OBS). METHODS AND MATERIALS: Participants who participated in the KS questionnaire was extracted from the National Health and Nutrition Examination Survey (NHANES) from 2007 to 2018. A series of covariates were also obtained. Weighted adjusted logistic regression was performed to investigate the association of KS with OBS. Dose-response relationship between KS and OBS was assessed by restricted cubic spline. RESULTS: In the fully adjusted model, we discovered that the risk of KS decreased by 3% with each OBS unit raised (OR = 0.97, 95% CI: 0.95-0.99, P = 0.01). In the OBS subgroups, in contrast to the lowest quartile OBS, the higher quartile OBS was correlated to the decreased risk of KS prevalence (Q3 vs Q1: OR = 0.7, 95% CI: 0.49-0.99, P = 0.04; Q4 vs Q1: OR = 0.66, 95% CI: 0.44-0.99, P = 0.04), and the results maintained relative stability across three models. We also found that the risk of population with KS was negatively linked with each unit increase in dietary OBS (OR = 0.97, 95% CI: 0.95-0.99, P = 0.005). Finally, we detected that there was a linear association between OBS and the risk of KS prevalence (P non-linear > 0.05). CONCLUSION: The study discovered that OBS that comprehensively reflects an individual's overall burden of oxidative stress was negatively related to the risk of KS, and can be utilized as an important indicator for assessing the risk of KS.
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Cálculos Renales , Adulto , Humanos , Encuestas Nutricionales , Prevalencia , Cálculos Renales/epidemiología , Estrés OxidativoRESUMEN
BACKGROUND: The aim of the study was to investigate the association between methylmalonic acid (MMA), a biomarker of mitochondrial dysfunction, and the risk of prostate cancer (PCa). METHODS AND MATERIALS: The relevant data were collected from the National Health and Nutrition Examination Survey (NHANES). Weighted univariable and multivariable logistic regression analyses were performed to investigate the association between MMA and risk of PCa. A stratified analysis was also carried out. The dose-response relationship was elucidated by conducting a restricted cubic spline function. RESULTS: A total of 2451 participants were included, of which 95 were PCa participants. The fully-adjusted model 2 constructed by weighted multivariable logistic regression analysis showed that the risk of PCa decreased by 53% when every MMA unit was added [OR: 0.47 (0.22-1.00), P = 0.049]. And a decrease in PCa risk was associated with a higher MMA level in MMA subgroups [OR: 0.34 (0.15-0.82), P = 0.02]. The results from a stratified analysis showed that participants in subgroups of other race, BMI (> 30 kg/m2), smoking (former and now), and hypertension (yes), an increase in every MMA unit was linked to a decrease in PCa risk. MMA and the risk of PCa were negatively correlated in a linear manner. CONCLUSION: It was discovered in the study that an increase in MMA level is connected to a decrease in PCa risk. The serum MMA level may be helpful in assessing PCa risk.
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Biomarcadores , Ácido Metilmalónico , Neoplasias de la Próstata , Humanos , Masculino , Neoplasias de la Próstata/sangre , Ácido Metilmalónico/sangre , Persona de Mediana Edad , Biomarcadores/sangre , Anciano , Medición de Riesgo , Mitocondrias , Estudios TransversalesRESUMEN
Background: Alzheimer's disease (AD) and Parkinson's disease (PD) are the leading causes of death among the elderly. Recent research has demonstrated that mitochondrial dysfunction, which is hallmark of neurodegenerative diseases, is a contributor to the development of these diseases. Methods and materials: Methylmalonic acid (MMA), AD, PD, inflammatory markers and covariates were extracted from the National Health and Nutrition Examination Survey (NHANES). The classification of the inflammatory markers was done through quartile conversion. A restricted cubic spike function was performed to study their dose-response relationship. MMA subgroups from published studies were used to explore the correlation between different subgroups and cause-specific mortality. Multivariable weighted Cox regression was carried out to investigate MMA and cause-specific mortality in patients with AD and PD. Weighted survival analysis was used to study the survival differences among MMA subgroups. Results: A non-linear correlation was observed between MMA and AD-specific death and PD-specific mortality. The presence of MMA Q4 was linked to increased death rates among AD patients (HR = 6.39, 95%CI: 1.19-35.24, P = 0.03) after controlling for potential confounders in a multivariable weighted Cox regression model. In PD patients, the MMA Q4 (Q4: HR: 5.51, 95 % CI: 1.26-24, P = 0.02) was also related to increased mortality. The results of survival analysis indicated that the poorer prognoses were observed in AD and PD patients with MMA Q4. Conclusion: The higher level of mitochondria-derived circulating MMA was associated with a higher mortality rate in AD and PD patients. MMA has the potential to be a valuable indicator for evaluating AD and PD patients' prognosis in the clinic.
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Background: The oxidative balance score (OBS) can be used to represent the overall burden of oxidative stress in an individual. This study aimed to explore the association between the risk of stroke and OBS. Methods and materials: The National Health and Nutrition Examination Survey (NHANES) from 1999 to 2018 was used to extract a series of variables for participants who took the stroke questionnaire. The construction of OBS relied on diet and lifestyle components, which included 16 nutrients and 4 lifestyle factors. Weighted multivariable-adjusted logistic regression was performed to investigate the association between stroke risk and OBS. A stratified analysis was also conducted. The dose-response relationship between stroke risk and OBS was elucidated by performing a restricted cubic spline function. Results: A total of 20,680 participants were included for analysis, 768 of whom suffered from stroke. Based on weighted multivariable logistic regression analysis, we discovered that the stroke prevalence decreased by 2% for each OBS unit added [OR: 0.98 (0.97-1.00), P < 0.01]. For the OBS subgroup, we also discovered that higher OBS was related to a reduction in the risk of stroke [Q4 vs. Q1: OR:0.65 (0.46-0.90), P < 0.01]. The prevalence of stroke declined by 3% with every OBS unit added to the diet component [OR: 0.97 (0.96-0.99), P < 0.01]. For the dietary OBS subgroup, higher OBS in diet components was associated with a decrease in the prevalence of stroke [Q4 vs. Q1: OR: 0.65, (0.47-0.91), P < 0.05]. Further stratified analysis showed that every OBS unit raised was associated with a decline in stroke prevalence, which was statistically significant in participants in subgroups of ≥60 years, female, no-diabetes mellitus and no-hypertension. OBS and stroke prevalence were correlated in a linear manner. Conclusion: The study found that a higher OBS was associated with a decrease in stroke prevalence, which could be a significant indicator for evaluating stroke risk.
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BACKGROUND: Patients with mild cognitive impairment (MCI) suffer from a high risk of developing Alzheimer's disease (AD). Cumulative evidence has demonstrated that the development of AD is a complex process that could be modulated by miRNAs. Here, we aimed to identify miRNAs involved in the pathway, and interrogate their ability to predict prognosis in patients with MCI. METHODS: We obtained the miRNA-seq profiles and the clinical characteristics of patients with MCI from the Gene Expression Omnibus (GEO). Cox regression analysis was used to construct a risk level model. The receiver operating characteristic (ROC) curve was used to assess the performance of the model for predicting prognosis. Combined with clinical characteristics, factors associated with prognosis were identified and a predictive prognosis nomogram was developed and validated. Through Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis, we evaluated molecular signatures for the candidate miRNAs. RESULTS: Our analysis identified 120 DEmiRNAs. The Cox regression analysis showed that two miRNAs could serve as risk factors for disease development. A risk level model was constructed. Age, apoe4, and risk level were associated with the prognosis. We developed a nomogram to predict disease progression. The calibration curve and concordance index (C-index) demonstrated the reliability of the nomogram. Functional enrichment analysis showed that these miRNAs were involved in regulating both cGMP-PKG and Sphingolipid signaling pathways. CONCLUSION: We have identified miRNAs associated with the development of MCI. These miRNAs could be used for early diagnosis and surveillance in patients with MCI, enabling prediction of the development of AD.
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Enfermedad de Alzheimer , MicroARNs , Enfermedad de Alzheimer/diagnóstico , Enfermedad de Alzheimer/genética , Apolipoproteína E4 , Humanos , MicroARNs/genética , MicroARNs/metabolismo , Reproducibilidad de los Resultados , EsfingolípidosRESUMEN
Background: Pyroptosis is a programmed cell death caused by inflammasomes, which is closely related to immune responses and tumor progression. The present study aimed to construct dual prognostic indices based on pyroptosis-associated and immune-associated genes and to investigate the impact of the biological signatures of these genes on Kidney Renal Clear Cell Carcinoma (KIRC). Materials and Methods: All the KIRC samples from the Cancer Genome Atlas (TCGA) were randomly and equally divided into the training and testing datasets. Cox and Least Absolute Shrinkage and Selection Operator (LASSO) regression analysis were used to screen crucial pyroptosis-associated genes (PAGs), and a pyroptosis-associated genes prognostic index (PAGsPI) was constructed. Immune-associated genes (IAGs) related to PAGs were identified, and then screened through Cox and LASSO regression analyses, and an immune-associated genes prognostic index (IAGsPI) was developed. These two prognostic indices were verified by using the testing and the Gene Expression Omnibus (GEO) datasets and an independent cohort. The patients' response to immunotherapy was analyzed. A nomogram was constructed and calibrated. qRT-PCR was used to detect the expression of PAGs and IAGs in the tumor tissues and normal tissues. Functional experiment was carried out. Results: 86 PAGs and 1,774 differentially expressed genes (DEGs) were obtained. After intersecting PAGs with DEGs, 22 differentially expressed PAGs (DEPAGs) were included in Cox and LASSO regression analyses, identifying 5 crucial PAGs. The PAGsPI was generated. Patients in the high-PAGsPI group had a poor prognosis. 82 differentially expressed IAGs (DEIAGs) were highly correlated with DEPAGs. 7 key IAGs were screened out, and an IAGsPI was generated. Patients in the high-IAGsPI group had a poor prognosis. PAGsPI and IAGsPI were verified to be robust and reliable. The results revealed patients in low-PAGsPI group and high-IAGsPI group may be more sensitive to immunotherapy. The calibrated nomogram was proved to be reliable. An independent cohort study also proved that PAGsPI and IAGsPI performed well in prognosis prediction. We found that the expression of AIM2 may affect proliferation of KIRC cells. Conclusion: PAGsPI and IAGsPI could be regarded as potential biomarkers for predicting the prognosis of patients with KIRC.
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Phytophthora infestans, the causal agent of late blight in potatoes and tomatoes, is the most important and ongoing pathogenic threat to agricultural production worldwide. Rapid and early identification of P. infestans is an essential prerequisite for countering the further spread of infection. In this study, a novel method for visual detection of P. infestans has been developed by integrating universal primer mediated asymmetric PCR with gold nanoparticle (AuNP)-based lateral flow biosensor. We employed asymmetric PCR to generate large amounts of single-stranded DNA (ssDNA) by amplifying a region of P. infestans-specific repetitive DNA sequence. The ssDNA products were then applied to the lateral flow biosensor to perform a visual detection using sandwich-type hybridization assays. In the presence of target DNA, sandwich-type hybridization reactions among the AuNP-probe, target DNA and capture probe were performed on the test line of the biosensor, and then a characteristic red band was produced for the accumulation of AuNPs. Quantitative analysis obtained by recording the optical intensity of the red band demonstrated that this biosensor could detect as little as 0.1â¯pg⯵L-1 genomic DNA. Furthermore, the specificity of the biosensor was confirmed by detecting three other Phytophthora species and two pathogenic fungi. We believe this method has potential application in early prediction of potato late blight disease and instigation of management actions to reduce the risk of epidemic development.
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Técnicas Biosensibles , Oro/química , Nanopartículas del Metal/química , Phytophthora infestans/aislamiento & purificación , Solanum tuberosum/parasitología , ADN Protozoario/aislamiento & purificación , Phytophthora infestans/patogenicidadRESUMEN
Potato virus S (PVS) is a major plant pathogen that causes considerable losses in global potato production. Knowledge of the evolutionary history and spatio-temporal dynamics of PVS is vital for developing sustainable management schemes. In this study, we investigated the phylodynamics of the virus by analysing 103 nucleotide sequences of the coat protein gene, sampled between 1985 and 2014. Our Bayesian phylogenetic analyses showed that PVS has been evolving at a rate of 3.32â¯×â¯10-4 substitutions/site/year (95% credibility interval 1.33â¯×â¯10-4-5.58â¯×â¯10-4). We dated the crown group to the year 1325 CE (95% credibility interval 762-1743 CE). Our phylogeographic analyses pointed to viral origins in South America and identified multiple migration pathways between Europe and other regions, suggesting that Europe has been a major hub for PVS transmission. The results of our study have potential implications for developing effective strategies for the control of this pathogen.
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Carlavirus/genética , Enfermedades de las Plantas/virología , Solanum tuberosum/virología , Europa (Continente) , Evolución Molecular , FilogeografíaRESUMEN
Food-borne pathogens have been recognized as a major cause of human infections worldwide. Their identification needs to be simpler, cheaper and more reliable than the traditional methods. Here, we constructed a low-cost paper platform for viable pathogenic bacteria detection with the naked eye. In this study, an effective isothermal amplification method was used to amplify the hlyA mRNA gene, a specific RNA marker in Listeria monocytogenes. The amplification products were applied to the paper-based platform to perform a visual test using sandwich hybridization assays. When the RNA products migrated along the platform by capillary action, the gold nanoparticles accumulated at the designated area. Under optimized experimental conditions, as little as 0.5 pg/µL genomic RNA from L. monocytogenes could be detected. It could also be used to specifically detect 20 CFU/mL L. monocytogenes from actual samples. The whole assay process, including RNA extraction, amplification, and visualization, can be completed within several hours. This method is suitable for point-of-care applications to detect food-borne pathogens, as it can overcome the false-positive results caused by amplifying nonviable L. monocytogenes. Furthermore, the results can be imaged and transformed into a two-dimensional bar code through an Android-based smart phone for further analysis or in-field food safety tracking.
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Técnicas Biosensibles/instrumentación , Oro/química , Listeria monocytogenes/genética , Listeria monocytogenes/aislamiento & purificación , Listeriosis/microbiología , Nanopartículas del Metal/química , ARN Bacteriano/genética , Diseño de Equipo , Microbiología de Alimentos , Análisis de Peligros y Puntos de Control Críticos , Humanos , Listeriosis/diagnóstico , Hibridación de Ácido Nucleico , Papel , Sistemas de Atención de Punto , ARN Bacteriano/aislamiento & purificación , Sensibilidad y EspecificidadRESUMEN
A novel method for detection of rotavirus has been developed by integrating magnetic primer based reverse transcription-polymerase chain reaction (RT-PCR) with electrochemiluminescence (ECL) detection. This is realized by accomplishing RT of rotavirus RNA in traditional way and performing PCR of the resulting cDNA fragment on the surface of magnetic particles (MPs). In order to implement PCR on MPs and achieve rapid ECL detection, forward and reverse primers are bounded to MPs and tris-(2,2'-bipyridyl) ruthenium (TBR), respectively. After RT-PCR amplification, the TBR labels are directly enriched onto the surface of MPs. Then the MPs-TBR complexes can be loaded on the electrode surface and analyzed by magnetic ECL platform without any post-modification or post-incubation process. So some laborious manual operations can be avoided to achieve rapid yet sensitive detection. In this study, rotavirus in fecal specimens was successfully detected within 1.5 h. Experimental results showed that the detection limit of the assay was 0.2 pg µL(-1) of rotavirus. The ECL intensity was linearly with the concentration from 0.2 pg µL(-1) to 400 pg µL(-1). What's more, the specificity of this method was confirmed by detecting other fecal specimens of patients with nonrotavirus-associated gastroenteritis. We anticipate that the proposed magnetic primer based RT-PCR with ECL detection strategy will find numerous applications in food safety field and clinical diagnosis.