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International Journal of Molecular... Oct 2020Virus detection in natural and clinical samples is a complicated problem in research and diagnostics. There are different approaches for virus isolation and... (Review)
Review
Virus detection in natural and clinical samples is a complicated problem in research and diagnostics. There are different approaches for virus isolation and identification, including PCR, CRISPR/Cas technology, NGS, immunoassays, and cell-based assays. Following the development of genetic engineering methods, approaches that utilize cell cultures have become useful and informative. Molecular biology methods allow increases in the sensitivity and specificity of cell cultures for certain viruses and can be used to generate reporter cell lines. These cell lines express specific reporter proteins (e.g., GFP, luciferase, and CAT) in response to virus infection that can be detected in a laboratory setting. The development of genome editing and synthetic biology methods has given rise to new perspectives regarding the design of virus reporter systems in cell cultures. This review is aimed at describing both virology methods in general and examples of the development of cell-based methods that exist today.
Topics: Animals; Cell Culture Techniques; Genes, Reporter; Genetic Engineering; Humans; Synthetic Biology; Virology; Viruses
PubMed: 33121109
DOI: 10.3390/ijms21217978 -
Pediatric Pulmonology Nov 2022In adults, permissive hypercapnia reduces mortality and ventilation duration. However, in preterm infants, the findings from past research regarding the efficacy and... (Meta-Analysis)
Meta-Analysis Review
CONTEXT
In adults, permissive hypercapnia reduces mortality and ventilation duration. However, in preterm infants, the findings from past research regarding the efficacy and safety of permissive hypercapnia are controversial.
OBJECTIVE
To evaluate the efficacy and safety of permissive hypercapnia versus normocapnia in preterm infants on mechanical ventilation.
DATA SOURCES
MEDLINE, EMBASE, CENTRAL, and CINAHL STUDY SELECTION: Published randomized controlled trials (RCTs), non-RCTs, interrupted time series, cohort studies, case-control studies, and controlled before-and-after studies were included.
DATA EXTRACTION
Two reviewers independently screened the title, abstract, and full text, extracted data, assessed the risk of bias, and evaluated certainty of evidence (CoE) according to the Grading of Recommendations Assessment and Development and Evaluation approach. A meta-analysis of RCTs was performed using the random-effects model.
RESULTS
Four RCTs (693 infants) and one cohort study (371 infants) were included. No significant differences existed between the permissive hypercapnia and normocapnia groups for bronchopulmonary dysplasia (BPD) (risk ratio [RR], 0.94; 95% confidence interval [CI], 0.74-1.18; very low CoE) and a composite outcome of death or BPD (RR, 1.05; 95% CI, 0.90-1.23; very low CoE). Permissive hypercapnia may increase necrotizing enterocolitis (RR, 1.69; 95% CI, 0.98-2.91; very low CoE), but the null or trivial effect cannot be excluded. No significant differences existed between the two groups for any other outcome assessed (very low-to-low CoE).
LIMITATIONS
The sample sizes were less than the optimal sizes for all outcomes assessed, indicating the need for further trials.
CONCLUSIONS
Permissive hypercapnia did not have any significant benefit or harm in preterm infants.
Topics: Bronchopulmonary Dysplasia; Enterocolitis, Necrotizing; Humans; Hypercapnia; Infant; Infant, Newborn; Infant, Premature; Respiration, Artificial
PubMed: 35945674
DOI: 10.1002/ppul.26108 -
Viruses Jul 2023Diseases induced by infection with pathogenic orthohantaviruses are characterized by a pronounced organ-specific manifestation. Pathogenic Eurasian orthohantaviruses...
Diseases induced by infection with pathogenic orthohantaviruses are characterized by a pronounced organ-specific manifestation. Pathogenic Eurasian orthohantaviruses cause hemorrhagic fever with renal syndrome (HFRS) with often massive proteinuria. Therefore, the use of a relevant kidney cell culture would be favorable to analyze the underlying cellular mechanisms of orthohantavirus-induced acute kidney injury (AKI). We tested different human tubular epithelial cell lines for their suitability as an in vitro infection model. Permissiveness and replication kinetics of highly pathogenic Hantaan virus (HTNV) and non-/low-pathogenic Tula virus (TULV) were analyzed in tubular epithelial cell lines and compared to human primary tubular epithelial cells. Ana-lysis of the cell line HK-2 revealed the same results for viral replication, morphological and functional effects as observed for HTNV in primary cells. In contrast, the cell lines RPTEC/TERT1 and TH1 demonstrated only poor infection rates after inoculation with HTNV and are unusable as an infection model. While pathogenic HNTV infects primary tubular and HK-2 cells, non-/low-pathogenic TULV infects neither primary tubular cells nor the cell line HK-2. Our results show that permissiveness of renal cells varies between orthohantaviruses with differences in pathogenicity and that HK-2 cells demonstrate a suitable in vitro model to study viral tropism and pathogenesis of orthohantavirus-induced AKI.
Topics: Humans; Epithelial Cells; Acute Kidney Injury; Kidney; Hantaan virus; Orthohantavirus; RNA Viruses
PubMed: 37632012
DOI: 10.3390/v15081670 -
Microbiology Spectrum Aug 2023The microbial community of the urinary tract (urinary microbiota or urobiota) has been associated with human health. Bacteriophages (phages) and plasmids present in the...
The microbial community of the urinary tract (urinary microbiota or urobiota) has been associated with human health. Bacteriophages (phages) and plasmids present in the urinary tract, like in other niches, may shape urinary bacterial dynamics. While urinary Escherichia coli strains associated with urinary tract infection (UTI) and their phages have been catalogued for the urobiome, bacterium-plasmid-phage interactions have yet to be explored. In this study, we characterized urinary E. coli plasmids and their ability to decrease permissivity to E. coli phage (coliphage) infection. Putative F plasmids were predicted in 47 of 67 urinary E. coli isolates, and most of these plasmids carried genes that encode toxin-antitoxin (TA) modules, antibiotic resistance, and/or virulence. Urinary E. coli plasmids, from urinary microbiota strains UMB0928 and UMB1284, were conjugated into E. coli K-12 strains. These transconjugants included genes for antibiotic resistance and virulence, and they decreased permissivity to coliphage infection by the laboratory phage P1vir and the urinary phages Greed and Lust. Plasmids in one transconjugant were maintained in E. coli K-12 for up to 10 days in the absence of antibiotic resistance selection; this included the maintenance of the antibiotic resistance phenotype and decreased permissivity to phage. Finally, we discuss how F plasmids present in urinary E. coli strains could play a role in coliphage dynamics and the maintenance of antibiotic resistance in urinary E. coli. The urinary tract contains a resident microbial community called the urinary microbiota or urobiota. Evidence exists that it is associated with human health. Bacteriophages (phages) and plasmids present in the urinary tract, like in other niches, may shape urinary bacterial dynamics. Bacterium-plasmid-phage interactions have been studied primarily in laboratory settings and are yet to be thoroughly tested in complex communities. This is especially true of the urinary tract, where the bacterial genetic determinants of phage infection are not well understood. In this study, we characterized urinary E. coli plasmids and their ability to decrease permissivity to E. coli phage (coliphage) infection. Urinary E. coli plasmids, encoding antibiotic resistance and transferred by conjugation into naive laboratory E. coli K-12 strains, decreased permissivity to coliphage infection. We propose a model by which urinary plasmids present in urinary E. coli strains could help to decrease phage infection susceptibility and maintain the antibiotic resistance of urinary E. coli. This has consequences for phage therapy, which could inadvertently select for plasmids that encode antibiotic resistance.
Topics: Humans; Escherichia coli; Plasmids; Coliphages; Bacteriophages; Escherichia coli Infections; Urinary Tract; Bacteria; Anti-Bacterial Agents
PubMed: 37409956
DOI: 10.1128/spectrum.01309-23 -
Virology Oct 2023Hendra virus (HeV) and Nipah virus (NiV) are henipaviruses that can cause fatal encephalitis in humans. Many animal models have been used to study henipavirus...
Hendra virus (HeV) and Nipah virus (NiV) are henipaviruses that can cause fatal encephalitis in humans. Many animal models have been used to study henipavirus pathogenesis. In the mouse, HeV infection has previously shown that intranasal challenge can lead to neurological infection, however mice similarly challenged with NiV show no evidence of virus infecting the brain. We generated recombinant HeV (rHeV) and NiV (rNiV) where selected proteins were switched to examine their role in neuroinvasion in the mouse. These viruses displayed similar growth kinetics when compared to wildtype in vitro. In the mouse, infection outcomes with recombinant virus did not differ to infection outcomes of wildtype viruses. Virus was detected in the brain of 5/30 rHeV-challenged mice, but not rNiV-challenged mice. To confirm the permissiveness of mouse neurons to these viruses, primary mouse neurons were successfully infected in vitro, suggesting that other pathobiological factors contribute to the differences in disease outcomes in mice.
PubMed: 37541184
DOI: 10.1016/j.virol.2023.109856 -
Injury Epidemiology Jan 2021Due to the differences in the way gun law permissiveness scales were created and speculation about the politically motivated underpinnings of the various scales, there...
BACKGROUND
Due to the differences in the way gun law permissiveness scales were created and speculation about the politically motivated underpinnings of the various scales, there have been questions about their reliability.
METHODS
We compared seven gun law permissiveness scales, varying by type and sources, for an enhanced understanding of the extent to which choice of a gun law permissiveness scale could affect studies related to gun violence outcomes in the United States. Specifically, we evaluated seven different scales: two rankings, two counts, and three scores, arising from a range of sources. We calculated Spearman correlation coefficients for each pair of scales compared. Cronbach's standardized alpha and Guttman's lambda were calculated to evaluate the relative reliability of the scales, and we re-calculated Cronbach's alpha after systematically omitting each scale to assess whether the omitted scale contributed to lower internal consistency between scales. Factor analysis was used to determine single factor loadings and estimates. We also assessed associations between permissiveness of gun laws and total firearm deaths and suicides in multivariable regression analyses.
RESULTS
All pairs of scales were highly correlated (average Spearman's correlation coefficient r = 0.77) and had high relative reliability (Cronbach's alpha = 0.968, Guttman's lambda = 0.975). All scales load onto a single factor. The choice of scale did not meaningfully change the parameter estimates for the associations between permissiveness of gun laws and gun deaths and suicides.
CONCLUSION
Gun law permissiveness scales are highly correlated despite any perceived political agenda, and the choice of gun law permissiveness scale has little effect on study conclusions related to gun violence outcomes.
PubMed: 33455576
DOI: 10.1186/s40621-020-00296-5 -
Gut Microbes 2023Non-caloric artificial sweeteners have been widely permitted as table sugar substitutes with high intensities of sweetness. They can pass through the intestinal tract...
Non-caloric artificial sweeteners have been widely permitted as table sugar substitutes with high intensities of sweetness. They can pass through the intestinal tract without significant metabolization and frequently encounter the gut microbiome, which is composed of diverse bacterial species and is a pool of antibiotic resistance genes (ARGs). However, little is known about whether these sweeteners could accelerate the spread of ARGs in the gut microbiome. Here, we established an conjugation model by using that carries chromosome-inserted Tn7 and plasmid-encoded gene as the donor and murine fecal bacteria as the recipient. We found that four commonly used artificial sweeteners (saccharin, sucralose, aspartame, and acesulfame potassium) can increase reactive oxygen species (ROS) production and promote plasmid-mediated conjugative transfer to the gut microbiome. Cell sorting and 16S rRNA gene amplicon sequencing analysis of fecal samples reveal that the tested sweeteners can promote the broad-host-range plasmid permissiveness to both Gram-negative and Gram-positive gut bacteria. The increased plasmid permissiveness was also validated with a human pathogen . Collectively, our study demonstrates that non-caloric artificial sweeteners can induce oxidative stress and boost the plasmid-mediated conjugative transfer of ARGs among the gut microbiota and a human pathogen. Considering the soaring consumption of these sweeteners and the abundance of mobile ARGs in the human gut, our results highlight the necessity of performing a thorough risk assessment of antibiotic resistance associated with the usage of artificial sweeteners as food additives.
Topics: Mice; Humans; Animals; Gastrointestinal Microbiome; Sweetening Agents; RNA, Ribosomal, 16S; Plasmids; Bacteria; Escherichia coli; Anti-Bacterial Agents; Drug Resistance, Multiple
PubMed: 36524841
DOI: 10.1080/19490976.2022.2157698 -
Heliyon Jul 2023This study compared the experiences of parental abusiveness in childhood and sexual permissiveness in adolescence between adolescents raised by dual parents and those...
OBJECTIVE
This study compared the experiences of parental abusiveness in childhood and sexual permissiveness in adolescence between adolescents raised by dual parents and those raised by single parents.
METHOD
The sampling process involved obtaining the sample size (N = 1037) based on statistical power calculations and the resources available, resulting in representativeness. The sampling frame consisted of students from different backgrounds (single-parent and two-parent families). A stratified random sampling helped to enhance the reliability and validity of the findings while minimizing potential biases. An online survey was used to collect data from the sampled students. An independent sample -test analysis was done by using SPSS software and PROCESS macro of the SPSS to test the hypotheses.
RESULTS
A significant and positive correlation was found between parental abusiveness and sexual permissiveness. There was also a significant mean difference in parental abusiveness experience and sexual permissiveness between adolescents from different family types. In addition, the average sexual permissiveness for males was higher than that of female adolescents from the same type of families.
CONCLUSION
Childhood life significantly influences adolescents' characters throughout their entire life.
PubMed: 37519719
DOI: 10.1016/j.heliyon.2023.e18101 -
Journal of Virology May 2023Adeno-associated viruses (AAVs) are small, helper-dependent, single-stranded DNA viruses that exploit a broad spectrum of host factors for cell entry. During the course...
Adeno-associated viruses (AAVs) are small, helper-dependent, single-stranded DNA viruses that exploit a broad spectrum of host factors for cell entry. During the course of infection, several AAV serotypes have been shown to transit through the -Golgi network within the host cell. In the current study, we investigated whether the Golgi-localized, calcium-dependent protease furin influences AAV transduction. While CRISPR/Cas9-mediated knockout (KO) of the gene minimally affected the transduction efficiency of most recombinant AAV serotypes tested, we observed a striking increase in transgene expression (~2 log orders) for the African green monkey isolate AAV4. Interrogation of different steps in the infectious pathway revealed that AAV4 binding, uptake, and transcript levels are increased in furin KO cells, but postentry steps such as uncoating or nuclear entry remain unaffected. Recombinant furin does not cleave AAV4 capsid proteins nor alter cellular expression levels of essential factors such as AAVR or GPR108. Interestingly, fluorescent lectin screening revealed a marked increase in 2,3--linked sialoglycan staining on the surface and perinuclear space of furin KO cells. The essential nature of increased sialoglycan expression in furin KO cells in enhancing AAV4 transduction was further corroborated by (i) increased transduction by the closely related isolates AAVrh.32.33 and sea lion AAV and (ii) selective blockade or removal of cellular 2,3--linked sialoglycans by specific lectins or neuraminidase, respectively. Based on the overall findings, we postulate that furin likely plays a key role in regulating expression of cellular sialoglycans, which in turn can influence permissivity to AAVs and possibly other viruses. Adeno-associated viruses (AAVs) are a proven recombinant vector platform for gene therapy and have demonstrated success in the clinic. Continuing to improve our knowledge of AAV-host cell interactions is critical for improving the safety and efficacy. The current study dissects the interplay between furin, a common intracellular protease, and certain cell surface sialoglycans that serve as viral attachment factors for cell entry. Based on the findings, we postulate that differential expression of furin in host cells and tissues is likely to influence gene expression by certain recombinant AAV serotypes.
Topics: Animals; Chlorocebus aethiops; Dependovirus; Virus Internalization; Furin; Genetic Vectors; Capsid Proteins; Transduction, Genetic
PubMed: 37097176
DOI: 10.1128/jvi.00093-23 -
Substance Abuse 2022Alcohol is the most used substance among adolescents in Nigeria. While risk factors for alcohol use among adolescents are well documented in Europe and the US, they...
Alcohol is the most used substance among adolescents in Nigeria. While risk factors for alcohol use among adolescents are well documented in Europe and the US, they have received less attention in the literature on African countries. This study aimed to investigate the factors associated with alcohol experimentation and drunkenness episodes in a national sample of Nigerian adolescents. A total sample of 4,078 secondary-school students participated in the survey during the school year 2015-2016. The survey involved 32 secondary schools of six geopolitical zones (South-South, South-West, South-East, North-Center, North-West, and North-East) and two metropolitan cities (Abuja and Lagos) of Nigeria. Sociodemographic characteristics, parental alcohol use and permissiveness, friends' alcohol use, risk perceptions and beliefs were investigated as correlates of alcohol experimentation and drunkenness episodes through multilevel, mixed-effect logistic regression models. The prevalence of alcohol experimentation was 34.0%, while the prevalence of drunkenness episodes was 13.4%. Results showed that male gender, family structure different from both parents' families, parental and friends' alcohol use, parental permissiveness to drink, low risk perceptions on drinking alcohol, and positive beliefs on consequences of alcohol use were associated with an increased probability of alcohol experimentation and drunkenness episodes. Family affluence and one-parent family structure were related to an increased probability of alcohol experimentation but not of drunkenness episodes. The majority of risk factors analyzed in this study generalize across drinking-behavior outcomes. Since the young population is dominant in Nigeria, alcohol use could become a big public health problem in the near future. High investment in adolescents' well-being by addressing the factors that contribute to drinking behavior might help to reduce the burden of the problem. Evidence-based prevention curriculum addressing knowledge, risk perceptions, beliefs on consequences of alcohol use, and parental behaviors should be implemented as widely and early as possible.
Topics: Adolescent; Adolescent Behavior; Alcohol Drinking; Alcoholic Intoxication; Humans; Male; Nigeria; Students
PubMed: 34214414
DOI: 10.1080/08897077.2021.1944952