27 research outputs found
Comparison of different sequencing techniques for identification of SARS-CoV-2 variants of concern with multiplex real-time PCR.
No full textAs different SARS-CoV-2 variants emerge and with the continuous evolvement of sub lineages of the delta variant, it is crucial that all countries carry out sequencing of at least >1% of their infections, in order to detect emergence of variants with higher transmissibility and with ability to evade immunity. However, due to limited resources as many resource poor countries are unable to sequence adequate number of viruses, we compared to usefulness of a two-step commercially available multiplex real-time PCR assay to detect important single nucleotide polymorphisms (SNPs) associated with the variants and compared the sensitivity, accuracy and cost effectiveness of the Illumina sequencing platform and the Oxford Nanopore Technologies' (ONT) platform. 138/143 (96.5%) identified as the alpha and 36/39 (92.3%) samples identified as the delta variants due to the presence of lineage defining SNPs by the multiplex real time PCR, were assigned to the same lineage by either of the two sequencing platforms. 34/37 of the samples sequenced by ONT had <5% ambiguous bases, while 21/37 samples sequenced using Illumina generated <5%. However, the mean PHRED scores averaged at 32.35 by Illumina reads but 10.78 in ONT. This difference results in a base error probability of 1 in 10 by the ONT and 1 in 1000 for Illumina sequencing platform. Sub-consensus single nucleotide variations (SNV) are highly correlated between both platforms (R2 = 0.79) while indels appear to have a weaker correlation (R2 = 0.13). Although the ONT had a slightly higher error rate compared to the Illumina technology, it achieved higher coverage with a lower number or reads, generated less ambiguous bases and was significantly less expensive than Illumina sequencing technology
Efficacy of rupatadine in reducing the incidence of dengue haemorrhagic fever in patients with acute dengue: A randomised, double blind, placebo-controlled trial
No full textBACKGROUND: Rupatadine was previously shown to reduce endothelial dysfunction in vitro, reduced vascular leak in dengue mouse models and to reduce the extent of pleural effusions and thrombocytopenia in patients with acute dengue. Therefore, we sought to determine the efficacy of rupatadine in reducing the incidence of dengue haemorrhagic fever (DHF) in patients with acute dengue. METHODS AND FINDINGS: A phase 2, randomised, double blind, placebo controlled clinical trial was carried out in patients with acute dengue in Sri Lanka in an outpatient setting. Patients with ≤3 days since the onset of illness were either recruited to the treatment arm of oral rupatadine 40mg for 5 days (n = 123) or the placebo arm (n = 126). Clinical and laboratory features were measured daily to assess development of DHF and other complications. 12 (9.7%) patients developed DHF in the treatment arm compared to 22 (17.5%) who were on the placebo although this was not significant (p = 0.09, relative risk 0.68, 95% CI 0.41 to 1.08). Rupatadine also significantly reduced (p = 0.01) the proportion of patients with platelet counts <50,000 cells/mm(3) and significantly reduced (p = 0.04) persisting vomiting, headache and hepatic tenderness (p<0.0001) in patients. There was a significant difference in the duration of illness (p = 0.0002) although the proportion of individuals who required hospital admission in both treatment arms. Only 2 patients on rupatadine and 3 patients on the placebo developed shock, while bleeding manifestations were seen in 6 patients on rupatadine and 7 patients on the placebo. CONCLUSIONS: Rupatadine appeared to be safe and well tolerated and showed a trend towards a reducing proportion of patients with acute dengue who developed DHF. Its usefulness when used in combination with other treatment modalities should be explored. TRIAL REGISTRATION: International Clinical Trials Registration Platform: SLCTR/2017/024
Comparison of amino acid changes detected in SARS-CoV-2 genomes by both sequencing technologies.
No full textAnnotated amino acid substitutions and deletions detected in each sample (n = 37). Mutations colored in green indicates they are synonymously detected by both sequencing technologies, whereas yellow and red indicate mutations detected exclusively by only one technology. The X axis indicates each amino acid, which is denoted by the original amino acid, its position in the protein and the substitution/deletion. Amino acid deletions are denoted by “del”.</p
Combined maximum likelihood phylogenetic tree created using sequence pairs of 37 the samples.
No full textThe ML tree was generated using the consensus sequences of each sequencing technology with 1000 bootstrap replicates using TIM2+F+R2 model. Tree is rooted on SARS-CoV-2 reference MN908947.3 and with samples sequences by Illumina coloured red and those sequenced by ONT coloured blue. Bootstrap support values are shown on each branch. 21/37 samples coupled together with 90% genome coverage from both Illumina and ONT datasets while 7/37 samples coupled together with less than 98% bootstrap support. 9/37 of the samples which failed to couple with their counterpart from ONT or Illumina had moderate to high (3% - 31%) ambiguous bases in either sequences.</p
Basic sequencing matrices for Illumina and Oxford Nanopore (ONT) outputs of 37.
No full textBasic sequencing matrices for Illumina and Oxford Nanopore (ONT) outputs of 37.</p
PHRED base call quality score distribution of samples sequenced by Illumina and ONT.
No full textDistribution plot of PHRED (probability of error per base call in a log scale) quality score (x axis) and error probability (secondary x axis) derived from the PHRED score for the data set sequenced from Illumina (n = 37) and ONT (n = 37). The scores of ONT are shown in blue and Illumina in red. The mean PHRED scores/error probability are shown with the dashed line for each technology. The mean PHRED scores averaged at 32.35 in Illumina reads and 10.78 in ONT.</p
Portraits of Wallachian rulers of the House of Basarab in the 14th-16th century. Formation of Younger. Europe - mature standards of portraits of the Early Middle Ages
No full textWydział Nauk Humanistycznych, Instytut Nauk o Sztuce; promotor: prof. Urszula Małgorzata MazurczakPortrety władców wołoskich z dynastii Basarabów w XIV-XVI w., przedstawione w kontekście kształtowania się Młodszej Europy, zostały opracowane w oparciu o szeroki zakres badań sztuki europejskiej. Praca doktorska podzielona została na dwa główne bloki tematyczne, w pierwszej części autorka zarysowuje kontekst kosmopolityczny, zacierający żelazną granicę pomiędzy sztuką bizantyńską - wschodnią, a łacińską - zachodnią. Część druga skupia się natomiast wokół historii Wołoszczyzny oraz analizy portretów Basarabów. Portrety Basarabów, zawężone stricte do malarstwa ściennego z terenów Wołoszczyzny oraz Transylwanii, skupione zostały wokół postaci czternastu z czterdziestu czterech władców. Najstarsze freski, które przedstawiają Basaraba I (1310-1352) dziś określanego imieniem Mikołaja Aleksandra (1352-1364), Mircze I Starego (1386-1418) oraz Michała I (1418-1420) pochodzą z cerkwi książęcej w Curtea de Arges oraz z cerkwi Trójcy Świętej Cozia. Średniowieczne portrety należące zaledwie do dziewięciu dynastów, przetrwały wszelkie zawirowania historii i stanowią świadectwo zarówno sztuki wołoskiej jak i prestiżu władzy Basarabów. Wizerunki fundatorów trzymających w dłoniach modele świątyń, władcy koronowani przez Aniołów stojący przed obliczem Boga czy dumni dynaści wspierający swych synów, legitymizujący ich prawa do objęcia należnego im tronu - pomimo niewielkiej ilości zachowanych obiektów zakres tematyki portretów jest szeroki i czerpiący wyraźnie ze wzorców oraz zasobów treściowych Wschodu.
Portraits of Wallachian rulers, of the House of Basarab in the 14th-16th century, presented in the context of the formation of Younger Europe, have been developed1 based on a wide range of research on European art. This doctoral thesis has been divided into two main thematic blocks. In the first part, the author outlines the cosmopoli tan context, blurring the line between Byzantine - Eastern - art, and Latin - Western - art. Whereas, the second part focuses on the history of Wallachia and examines portraits of the Basarabs. Portraits of Basarabs, narrowed down to walli paintings from the regions of Wallachia and Transylvania, have been focused on fourteen out of forty rulers. The old-est frescoes depicting Basarab I (1310-1352). Nicholas AIexander (1352-1364), Mircea the Elder (1386-1418) and Michael I (1418-1420) are from the Cathedral of Curtea de Arges and the Holy Trinity Church of the Cozia Monastery. Mediaeval portraits of just nine rulers have survived various historical turmoils and are a witness to the prestige oil the Basarabs’ rule as well as Wallachian art. Benefactors holding models of churches in their hands, monarchs stand ing before God and being coronated by angels, or proud rulers supporting their sons and legitimising their right of succession - despite the small number of remaining por traits, there is a wide range of subject matter, which clearly draws from Eastern standards and themes
Correlation of sub-consensus allele frequencies observed for SNV and Indels between two sequencing technologies.
No full texta) Correlation between sub-consensus single nucleotide substitution frequencies observed for Illumina and ONT. Nucleotide substitutions detected exclusively by one technology are indicated in green and blue whereas the substitutions detected by both technologies are colored red. Even though more nucleotide substitutions exclusive to ONT were observed, there is a clear positive correlation (R2 = 0.79) between two sequencing technologies. b) Correlation between sub-consensus indel frequencies observed for Illumina and ONT. More indels exclusive to ONT can be seen with a weak correlation (R2 = 0.13) between the indel frequencies between two technologies suggesting ONT tend to result in more false-positive indels.</p
Molecular Epidemiology of AY.28 and AY.104 Delta Sub-lineages in Sri Lanka
No full textBACKGROUND: The worst SARS-CoV-2 outbreak in Sri Lanka was due to the two Sri Lankan delta sub-lineages AY.28 and AY.104. We proceeded to further characterize the mutations and clinical disease severity of these two sub-lineages. METHODS: 705 delta SARS-CoV-2 genomes sequenced by our laboratory from mid-May to November 2021 using Illumina and Oxford Nanopore were included in the analysis. The clinical disease severity of 440/705 individuals were further analyzed to determine if infection with either AY.28 or AY.104 was associated with more severe disease. Sub-genomic RNA (sg-RNA) expression was analyzed using periscope. RESULTS: AY.28 was the dominant variant throughout the outbreak, accounting for 67.7% of infections during the peak of the outbreak. AY.28 had three lineage defining mutations in the spike protein: A222V (92.80%), A701S (88.06%), and A1078S (92.04%) and seven in the ORF1a: R24C, K634N, P1640L, A2994V, A3209V, V3718A, and T3750I. AY.104 was characterized by the high prevalence of T95I (90.81%) and T572L (65.01%) mutations in the spike protein and by the absence of P1640L (94.28%) in ORF1a with the presence of A1918V (98.58%) mutation. The mean sgRNA expression levels of ORF6 in AY.28 were significantly higher compared to AY.104 (p < 0.0001) and B.1.617.2 (p < 0.01). Also, ORF3a showed significantly higher sgRNA expression in AY.28 compared to AY.104 (p < 0.0001). There was no difference in the clinical disease severity or duration of hospitalization in individuals infected with these sub lineages. CONCLUSIONS: Therefore, AY.28 and AY.104 appear to have a fitness advantage over the parental delta variant (B.1.617.2), while AY.28 also had a higher expression of sg-RNA compared to other sub-lineages. The clinical implications of these should be further investigated
