Multiplexed RT-qPCR Coupled with Whole-Genome Sequencing to Monitor a SARS-CoV-2 Omicron Variant of Concern in a Hospital Laboratory Setting in Latvia.

At the end of 2021, the SARS-CoV-2 Omicron variant of concern (VOC) displaced the previously dominant Delta VOC and enhanced diagnostic and therapeutic challenges worldwide. Respiratory specimens submitted to the Riga East University Hospital Laboratory Service by the central and regional hospitals of Latvia from January to March 2022 that were positive for SARS-CoV-2 RNA were tested by commercial multiplexed RT-qPCR targeting three of the Omicron VOC signature mutations: ΔH69/V70, E484A, and N501Y. Of the specimens tested and analyzed in parallel by whole-genome sequencing (WGS), 964 passed the internal quality criteria (genome coverage ≥90%, read depth ≥400×) and the Nextstrain's quality threshold for "good". We validated the detection accuracy of RT-qPCR for each target individually by using WGS as a control. The results were concordant with both approaches for 938 specimens, with the correct classification rate exceeding 96% for each target (CI 95%); however, the presumptive WHO label was misassigned for 21 specimens. The RT-qPCR genotyping provided an acceptable means to pre-monitor the prevalence of the two presumptive Omicron VOC sublineages, BA.1 and BA.2.

Distinguishable codon usage and amino acid composition patterns among substrates of leaderless secretory pathways from proteobacteria.

The combined set of codon usage frequencies (61 sense codons) from the 111 annotated sequences of leaderless secreted type I, type III, type IV, and type VI proteins from proteobacteria were subjected to the forward and backward selection to obtain a combination of most effective predictor variables for classification/prediction purposes. The group of 24 codon frequencies displayed a strong discriminatory power with an accuracy of 100% for originally grouped and 97.3 +/- 1.6% for cross-validated (LOOCV) cases and an acceptable error rate (0.062 +/- 0.012) in k-fold (k = 6) cross-validation (KCV). The summary frequencies of synonymous codons for ten amino acids as the alternative predictor variables revealed a comparable discriminatory power (92.8 +/- 2.5% for LOOCV), however at somewhat lower levels of prediction accuracy (0.106 +/- 0.015 of KCV). A number of significant (p < 0.001) differences were found among indices of codon usage and amino acid composition depending on a definite secretion type. About 60% of secretion substrates were characterized as apparently originated from horizontal gene transfer events or putative alien genes and found to be unequally allocated in respect of groups. The proposed prediction approaches could be used to specify secretome proteins from genomic sequences as well as to assess the compatibility between bacterial secretion pathways and secretion substrates.

Relationship between Metabolic Fluxes and Sequence-Derived Properties of Enzymes.

Metabolic fluxes are key parameters of metabolic pathways being closely related to the kinetic properties of enzymes, thereby could be dependent on. This study examines possible relationships between the metabolic fluxes and the physical-chemical/structural features of enzymes from the yeast Saccharomyces cerevisiae glycolysis pathway. Metabolic fluxes were quantified by the COPASI tool using the kinetic models of Hynne and Teusink at varied concentrations of external glucose. The enzyme sequences were taken from the UniProtKB and the average amino acid (AA) properties were computed using the set of Georgiev's uncorrelated scales that satisfy the VARIMAX criterion and specific AA indices that show the highest correlations with those. Multiple linear regressions (88.41%

Relationships between kinetic constants and the amino acid composition of enzymes from the yeast Saccharomyces cerevisiae glycolysis pathway.

The kinetic models of metabolic pathways represent a system of biochemical reactions in terms of metabolic fluxes and enzyme kinetics. Therefore, the apparent differences of metabolic fluxes might reflect distinctive kinetic characteristics, as well as sequence-dependent properties of the employed enzymes. This study aims to examine possible linkages between kinetic constants and the amino acid (AA) composition (AAC) for enzymes from the yeast Saccharomyces cerevisiae glycolytic pathway. The values of Michaelis-Menten constant (KM), turnover number (kcat), and specificity constant (ksp = kcat/KM) were taken from BRENDA (15, 17, and 16 values, respectively) and protein sequences of nine enzymes (HXK, GADH, PGK, PGM, ENO, PK, PDC, TIM, and PYC) from UniProtKB. The AAC and sequence properties were computed by ExPASy/ProtParam tool and data processed by conventional methods of multivariate statistics. Multiple linear regressions were found between the log-values of kcat (3 models, 85.74% < Radj.2 <94.11%, p < 0.00001), KM (1 model, Radj.2 = 96.70%, p < 0.00001), ksp (3 models, 96.15% < Radj.2 < 96.50%, p < 0.00001), and the sets of AA frequencies (four to six for each model) selected from enzyme sequences while assessing the potential multicollinearity between variables. It was also found that the selection of independent variables in multiple regression models may reflect certain advantages for definite AA physicochemical and structural propensities, which could affect the properties of sequences. The results support the view on the actual interdependence of catalytic, binding, and structural residues to ensure the efficiency of biocatalysts, since the kinetic constants of the yeast enzymes appear as closely related to the overall AAC of sequences.