The direct approach for determining reference intervals (RIs) is not always practical. This study aimed to generate evidence that a real-world data (RWD) approach could be applied to transfer free ...thyroxine RIs determined in one population to a second population, presenting an alternative to performing multiple RI determinations.
Two datasets (US, n = 10,000; Europe, n = 10,000) were created from existing RWD. Descriptive statistics, density plots and cumulative distributions were produced for each data set and comparisons made. Cumulative probabilities at the lower and upper limits of the RIs were identified using an empirical cumulative distribution function. According to these probabilities, estimated percentiles for each dataset and estimated differences between the two sets of percentiles were obtained by case resampling bootstrapping. The estimated differences were then evaluated against a pre-determined acceptance criterion of ≤7.8% (inter-individual biological variability). The direct approach was used to validate the RWD approach.
The RWD approach provided similar descriptive statistics for both populations (mean: US = 16.1 pmol/L, Europe = 16.4 pmol/L; median: US = 15.4 pmol/L, Europe = 15.8 pmol/L). Differences between the estimated percentiles at the upper and lower limits of the RIs fulfilled the pre-determined acceptance criterion and the density plots and cumulative distributions demonstrated population homogeneity. Similar RI distributions were observed using the direct approach.
This study provides evidence that a RWD approach can be used to transfer RIs determined in one population to another.
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•Real-world data (RWD) approach to determine reference intervals (RIs) was assessed.•Population homogeneity between the US and Europe was seen using the RWD approach.•The RWD approach was validated using the direct approach.•The RWD approach allows for transfer of RIs from one population to another.
Replication of SARS-CoV-2 in the human population is defined by distributions of mutants that are present at different frequencies within the infected host and can be detected by ultra-deep ...sequencing techniques. In this study, we examined the SARS-CoV-2 mutant spectra of amplicons from the spike-coding (S-coding) region of 5 nasopharyngeal isolates derived from patients with vaccine breakthrough. Interestingly, all patients became infected with the Alpha variant, but amino acid substitutions that correspond to the Delta Plus, Iota, and Omicron variants were present in the mutant spectra of the resident virus. Deep sequencing analysis of SARS-CoV-2 from patients with vaccine breakthrough revealed a rich reservoir of mutant types and may also identify tolerated substitutions that can be represented in epidemiologically dominant variants.
SARS-CoV-2 isolates of a given clade may contain low frequency genomes that encode amino acids or deletions which are typical of a different clade.
Here we use high resolution ultra-deep sequencing ...to analyze SARS-CoV-2 mutant spectra.
In 6 out of 11 SARS-CoV-2 isolates from COVID-19 patients, the mutant spectrum of the spike (S)-coding region included two or more amino acids or deletions, that correspond to discordant viral clades. A similar observation is reported for laboratory populations of SARS-CoV-2 USA-WA1/2020, following a cell culture infection in the presence of remdesivir, ribavirin or their combinations. Moreover, some of the clade-discordant genome residues are found in the same haplotype within an amplicon.
We evaluate possible interpretations of these findings, and reviewed precedents for rapid selection of genomes with multiple mutations in RNA viruses. These considerations suggest that intra-host evolution may be sufficient to generate minority sequences which are closely related to sequences typical of other clades. The results provide a model for the origin of variants of concern during epidemic spread─in particular Omicron lineages─that does not require prolonged infection, involvement of immunocompromised individuals, or participation of intermediate, non-human hosts.
Mutant spectra of RNA viruses are important to understand viral pathogenesis and response to selective pressures. There is a need to characterize the complexity of mutant spectra in coronaviruses ...sampled from infected patients. In particular, the possible relationship between SARS-CoV-2 mutant spectrum complexity and disease associations has not been established. In the present study, we report an ultradeep sequencing (UDS) analysis of the mutant spectrum of amplicons from the nsp12 (polymerase)- and spike (S)-coding regions of 30 nasopharyngeal isolates (diagnostic samples) of SARS-CoV-2 of the first COVID-19 pandemic wave (Madrid, Spain, April 2020) classified according to the severity of ensuing COVID-19. Low-frequency mutations and deletions, counted relative to the consensus sequence of the corresponding isolate, were overwhelmingly abundant. We show that the average number of different point mutations, mutations per haplotype, and several diversity indices was significantly higher in SARS-CoV-2 isolated from patients who developed mild disease than in those associated with moderate or severe disease (exitus). No such bias was observed with RNA deletions. Location of amino acid substitutions in the three-dimensional structures of nsp12 (polymerase) and S suggest significant structural or functional effects. Thus, patients who develop mild symptoms may be a richer source of genetic variants of SARS-CoV-2 than patients with moderate or severe COVID-19.
The study shows that mutant spectra of SARS-CoV-2 from diagnostic samples differ in point mutation abundance and complexity and that significantly larger values were observed in virus from patients who developed mild COVID-19 symptoms. Mutant spectrum complexity is not a uniform trait among isolates. The nature and location of low-frequency amino acid substitutions present in mutant spectra anticipate great potential for phenotypic diversification of SARS-CoV-2.
Populations of RNA viruses are composed of complex and dynamic mixtures of variant genomes that are termed mutant spectra or mutant clouds. This applies also to SARS-CoV-2, and mutations that are ...detected at low frequency in an infected individual can be dominant (represented in the consensus sequence) in subsequent variants of interest or variants of concern. Here we briefly review the main conclusions of our work on mutant spectrum characterization of hepatitis C virus (HCV) and SARS-CoV-2 at the nucleotide and amino acid levels and address the following two new questions derived from previous results: (i) how is the SARS-CoV-2 mutant and deletion spectrum composition in diagnostic samples, when examined at progressively lower cut-off mutant frequency values in ultra-deep sequencing; (ii) how the frequency distribution of minority amino acid substitutions in SARS-CoV-2 compares with that of HCV sampled also from infected patients. The main conclusions are the following: (i) the number of different mutations found at low frequency in SARS-CoV-2 mutant spectra increases dramatically (50- to 100-fold) as the cut-off frequency for mutation detection is lowered from 0.5% to 0.1%, and (ii) that, contrary to HCV, SARS-CoV-2 mutant spectra exhibit a deficit of intermediate frequency amino acid substitutions. The possible origin and implications of mutant spectrum differences among RNA viruses are discussed.
Twenty-six cases of Blastoschizomyces capitatus infection were diagnosed in 25 patients at 7 tertiary care hematology units in Spain over a 10-year period. Most patients (92%) had acute leukemia and ...developed infection during a period of severe and prolonged neutropenia. Two patients had esophagitis, and the rest had invasive infection. Fungemia (20 cases) was a common finding, with frequent visceral dissemination. The 30-day mortality associated with this infection was 52%, compared with 57% among patients with systemic infection. In a univariate analysis, the following 3 variables had a positive impact on 30-day survival: removal of the central venous catheter within 5 days after the onset of infection (P = .02), a good performance status (P = .003), and receipt of systemic prophylactic or empirical antifungal therapy before infection onset (P = .006). Outcome for neutropenic patients with B. capitatus infection is still poor. Rapid removal of the central venous catheter and novel antifungal therapies are recommended for treatment of this rare infection.
A study to evaluate the biofilm-development ability in three different media (Middlebrook 7H9, sterile tap water and PBS-5% glucose) was performed with 19 collection strains from 15 different species ...on non-pigmented rapidly growing mycobacteria (NPRGM). A microtiter plate assay was developed to evaluate the percentage of covered surface of the microtiter plate wells in different days from day 1 to day 69.
All strains were able to develop biofilm in all the tested media. Middlebrook 7H9 showed the fastest growth, followed by sterile tap water and PBS-5% glucose. A sigmoid growth curve was detected in all the strains both in Middlebrook 7H9 and in sterile tap water. A difference could be detected for Mycobacterium abscessus in tap water, where it showed faster growth than all the other strains.
Biofilm development seems to be a property of all the species of NPRGM and it depends on the nutrients present in the medium. The microtiter plate assay described here is a useful tool to evaluate differences in biofilm development among the different species of rapidly growing mycobacteria.
Serology in the xxi century: is it still of interest? García Coca, Marta; Fernández Roblas, Ricardo; Gadea Gironés, Ignacio
Enfermedades infecciosas y microbiologia clinica (English ed.),
05/2019, Volume:
37 Suppl 1
Journal Article
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