The evaluation of an automated mariPOC SARS-CoV-2 antigen test compared to RT-qPCR SARS-CoV-2 assay and comparison of its sensitivity in Delta- and Omicron-variant samples.

BACKGROUND: The rapid diagnostics tests for SARS-CoV-2 antigen vary in their sensitivities, and moreover, genomic mutations may further affect the performance of the assays. We aimed to evaluate the analytical performance of an automated antigen assay and compare its sensitivity in Delta- and Omicron-variant positive clinical samples. MATERIAL AND METHODS: The analytical performance of an automated mariPOC SARS-CoV-2 antigen test was evaluated on a population of community-dwelling subjects with mild respiratory symptoms or being asymptomatic investigated by the RT-qPCR Allplex™ SARS-CoV-2 assay. The sensitivity and specificity of the antigen test were evaluated on prospective 621 nasopharyngeal swabs along with oropharyngeal swabs. The sensitivity regarding variants determined by the Allplex™ SARS-CoV-2 Variant assays was analysed in additional, retrospective 158 Delta and 59 Omicron samples. RESULTS: The overall sensitivity of the antigen test in prospective samples was 77.9% (113/145; 95% confidence interval [CI] 70.3-84.4) with the specificity of 99.8% (95% CI 98.8-100). Regarding the variant, the sensitivity was higher in Omicron-variant samples, 93.2% (55/59; 95% CI 83.5-98.1), compared to Delta-variant samples, 71.5% (113/158; 95% CI 63.8-78.4; p = .001). CONCLUSION: In community-dwelling subjects with mild respiratory symptoms or being asymptomatic, the automated mariPOC SARS-CoV-2 antigen test showed high sensitivity over 98.0% in subgroup samples with cycle threshold (Ct) values < 25. Regarding the variant, the antigen test sensitivity was higher in the Omicron-variant samples compared to the Delta-variant samples. The analytical performance of the antigen test can differ between the SARS-CoV-2 variants, and a re-evaluation should be performed for new circulating lineages.

Distribution of SARS-CoV-2 Lineages in the Czech Republic, Analysis of Data from the First Year of the Pandemic.

In the Czech Republic, the current pandemic led to over 1.67 million SARS-CoV-2- positive cases since the recording of the first case on 1 March 2020. SARS-CoV-2 genome analysis is an important tool for effective real-time quantitative PCR (RT-qPCR) diagnostics, epidemiology monitoring, as well as vaccination strategy. To date, there is no comprehensive report on the distribution of SARS-CoV-2 genome variants in either the Czech Republic, including Central and Eastern Europe in general, during the first year of pandemic. In this study, we have analysed a representative cohort of SARS-CoV-2 genomes from 229 nasopharyngeal swabs of COVID-19 positive patients collected between March 2020 and February 2021 using validated reference-based sequencing workflow. We document the changing frequency of dominant variants of SARS-CoV-2 (from B.1 -> B.1.1.266 -> B.1.258 -> B.1.1.7) throughout the first year of the pandemic and list specific variants that could impact the diagnostic efficiency RT-qPCR assays. Moreover, our reference-based workflow provided evidence of superinfection in several samples, which may have contributed to one of the highest per capita numbers of COVID-19 cases and deaths during the first year of the pandemic in the Czech Republic.

Evaluation of a Gastrointestinal Pathogen Panel Immunoassay in Stool Testing of Patients with Suspected Clostridioides (Clostridium) difficile Infection.

Clostridioides (Clostridium) difficile infection (CDI) is the most common causative pathogen of health care-associated gastrointestinal infections; however, due to the overlap of clinical symptoms with those of other causes of acute gastroenteritis, the selection of the most appropriate laboratory test is difficult. From April to October 2018, 640 stool samples requested for CDI testing were examined using the mariPOC CDI and Gastro test (ArcDia), which allows the detection of C. difficile glutamate dehydrogenase (GDH) and toxin A/B, norovirus genogroups GI and GII.4, rotavirus, adenovirus, and Campylobacter spp. In parallel, the C. Diff Quik Chek Complete test (Alere) was used as a routine diagnostic assay, and C. difficile toxigenic culture was used as a reference method. The sensitivity of the mariPOC CDI and Gastro test was comparable to that of C. Diff Quik Chek Complete for the detection of GDH (96.40% [95% confidence interval {CI}, 91.81% to 98.82%] versus 95.68% [95% CI, 90.84 to 98.40%]; P = 1.00) and was higher for the detection of toxin A/B (66.67% [95% CI, 57.36 to 75.11%] versus 55.56% [95% CI, 46.08 to 64.74%]; P = 0.00). The specificity of the mariPOC CDI and Gastro test was lower than that of C. Diff Quik Chek Complete for GDH detection (95.21% [95% CI, 92.96% to 96.91%] versus 97.60% [95% CI, 95.85% to 98.76%]; P = 0.04) and comparable to that of C. Diff Quik Chek Complete for toxin A/B detection (99.24 [95% CI, 98.05% to 99.79%] versus 99.81% [95% CI, 98.94% to 100.0%]; P = 0.37). In 29 cases (4.53%), other causative agents of diarrhea were detected (Campylobacter spp. [n = 17], rotavirus [n = 7], and norovirus genogroup GII.4 [n = 5]).

[Comparing traditional and commercial molecular biology detection of gastrointestinal pathogens with AusDiagnostics panels in Motol University Hospital].

BACKGROUND: The aim was to do an internal audit of gastrointestinal pathogen detection at the Department of Medical Microbiology, Motol University Hospital between the years 2014 and 2018 and to test two commercial multiplex molecular biology assays potentially improving the diagnostic process and reducing costs. MATERIAL AND METHODS: Based on data from a laboratory information system (LIS), a total of 45,888 samples were identified which had been tested for the presence of gastrointestinal pathogens using culture, immunochromatographic, microscopic and molecular biology techniques between 2014-2018. Novel multiplex molecular biology detection was used to test 182 nucleic acid isolates obtained from stool samples with the Enteric Viruses (8-well) assay (Viral Panel, EVP) or Faecal Pathogens M (16-well) assay (Microbial Panel, FPM) manufactured by AusDiagnostics. RESULTS: The LIS data showed 6.2 % of positive pathogens causing diarrhea from all tested samples (detection rates: 4.5 % for bacterial agents, 21.6 % for viral agents and 0.4 % for parasitic agents). Valid samples (98.9 % of all tested samples) tested by the molecular biology technique yielded, in descending order: C. difficile toxin B (19 %), norovirus (9 %), astrovirus (8 %), Campylobacter (7 %), sapovirus (6 %), Yersinia enterocolitica (6 %), rotavirus (4 %), enterovirus (3 %), Aeromonas (3 %), adenovirus (2 %) and Salmonella (1 %). There was found at least 1 additional new positive detection in 27 % of stools tested by the Viral Panel and in 40 % of stools tested by the Microbial Panel in comparison with the traditional approach. Introducing the panels into routine diagnostic practice will not reduce the costs. CONCLUSIONS: The introduction of novel multiplex molecular biology assays for detecting gastrointestinal pathogens will considerably increase pathogen detection rates even though the costs will be higher for the Department of Medical Microbiology.

Intracellular sequestration of zinc, cadmium and silver in Hebeloma mesophaeum and characterization of its metallothionein genes.

Sequestration of intracellular heavy metals in eukaryotes involves compartmentalization and binding with cytosolic, cysteine-rich metallothionein (MT) peptides. We examined the roles of these processes in handling of zinc (Zn), cadmium (Cd) and silver (Ag) in sporocarps and a metal-exposed extraradical mycelium of Hebeloma mesophaeum, the Zn-accumulating ectomycorrhizal (EM) species frequently associated with metal disturbed sites. Size exclusion chromatography revealed that the majority of Zn and Cd in the sporocarps and mycelium was contained in a low molecular mass fraction attributable to compartmentalized metal. The staining of hyphal cells with the Zn-specific Zinquin and Cd-specific Leadmium fluorescent tracers labeled Zn and Cd in small, punctuated vesicles and vacuoles, respectively. By contrast, the sporocarp and mycelium Ag was associated with cysteine-rich, 5-kDa peptides. The peptides of the same size were also identified in minor Zn and Cd complexes from the metal-exposed mycelium. We have further isolated and characterized HmMT1, HmMT2 and HmMT3 genes coding for different 5-kDa MTs of H. mesophaeum collected at a lead smelter site. Heterologous complementation assays in metal-sensitive yeast mutants indicated that HmMTs encode functional, metal-specific peptides: only HmMT1 was able to complement sensitivity to Zn; HmMT1 conferred higher tolerance to Cd and Cu than HmMT2 or HmMT3; and both HmMT2 and HmMT3, but not HmMT1, conferred increased tolerance to Ag. The presence of HmMT1 and HmMT3, but not HmMT2, was also confirmed in a H. mesophaeum isolate from an unpolluted site. Gene expression analysis in the extraradical mycelium of this isolate revealed that the transcription of HmMT1 was preferentially induced in the presence of Zn and Cd, while Ag was a stronger inducer of HmMT3. Altogether, these results improve our understanding of the handling of intracellular Zn, Cd and Ag in Hebeloma and represent the first evidence suggesting involvement of MTs in sequestration of Zn in EM fungi.