ABSTRACT
Reliable, robust, large-scale molecular testing for SARS-CoV-2 is essential for monitoring the ongoing Covid-19 pandemic. We have developed a scalable analytical approach to detect viral proteins based on peptide immunoaffinity enrichment combined with liquid chromatography - mass spectrometry (LC-MS). This is a multiplexed strategy, based on targeted proteomics analysis and read-out by LC-MS, capable of precisely quantifying and confirming the presence of SARS-CoV-2 in PBS swab media from combined throat/nasopharynx/saliva samples. The results reveal that the levels of SARS-CoV-2 measured by LC-MS correlate well with their corresponding RT-PCR readout (r=0.79). The analytical workflow shows similar turnaround times as regular RT-PCR instrumentation with a quantitative readout of viral proteins corresponding to cycle thresholds (Ct) equivalents ranging from 21 to 34. Using RT-PCR as a reference, we demonstrate that the LC-MS-based method has 100% negative percent agreement (estimated specificity) and 95% positive percent agreement (estimated sensitivity) when analyzing clinical samples collected from asymptomatic individuals with a Ct within the limit of detection of the mass spectrometer (Ct [≤]30). These results suggest that a scalable analytical method based on LC-MS has a place in future pandemic preparedness centers to complement current virus detection technologies.
ABSTRACT
BackgroundIn March 2020, Stockholm, Sweden was hit by a severe outbreak of SARS-CoV-2. Four weeks later, a systematic study of testing for past or present infections among healthcare workers in the region was launched. Only a minority of COVID-19 related deaths occurred at hospitals and the study was therefore extended to employees in companies providing home care services for the elderly. MethodsFive companies offered participation to 438 employees at work and 405 employees (92.5%) were enrolled. Serum samples were analyzed for IgG to SARS-CoV-2 and throat swabs were tested by for the SARS-CoV-2 virus by PCR. ResultsAmong home care employees, 20.1% (81/403) were seropositive, about twice as many as in a simultaneously enrolled reference population (healthcare workers entirely without patient contact, n=3,671; 9.7% seropositivity). Only 13/379 employees (3.4%) had evidence of a current infection (PCR positivity). Among these, 5 were also seropositive (a sign of past infection or lingering infection after symptoms have resolved) and 3 were positive with only low amounts of virus. The combination of high amounts of virus and no antibodies, a characteristic for pre-symptomatic COVID-19, was thus present only in 5 employees (1.3%). ConclusionsPersonnel providing home service for the elderly appear to be a risk group for SARS-CoV-2 infection. Employees likely to be pre-symptomatic for COVID-19 can be readily identified by screening. Increased attention for protection of employees as well as of the elderly they serve is warranted.
ABSTRACT
BackgroundPre-symptomatic subjects are spreaders of SARS-CoV-2 infection, and strategies that could identify these subjects, particularly in hospital settings, are needed. MethodsWe tested a cohort of 9449 employees at work at the Karolinska University Hospital, Stockholm, Sweden for SARS-CoV-2 RNA and antibodies, linked the screening results to sick leave records and examined the association between screening results and past or future sick leave using multinomial logistic regression. ResultsWe found that healthcare workers with high amounts of SARS-CoV-2 virus, as indicated by the Cycle threshold (Ct) value in the PCR, had the highest risk for sick leave in the two weeks after testing (OR 11{middle dot}97 (CI 95% 6{middle dot}29-22{middle dot}80)) whereas subjects with low amounts of virus had the highest risk for sick leave in the past three weeks before testing (OR 6{middle dot}31 (4{middle dot}38-9{middle dot}08)). Only 2{middle dot}5% of employees were SARS-CoV-2 positive while 10{middle dot}5% were positive by serology and 1{middle dot}2% were positive in both tests. Serology-positive subjects were not at excess risk for future sick leave (OR 1{middle dot}06 (95% CI, 0{middle dot}71-1{middle dot}57)), but virus-positive subjects had a 7{middle dot}23 fold (95% CI, 4{middle dot}52-11{middle dot}57)) increased risk for sick leave within two weeks post testing. ConclusionsScreening of asymptomatic healthcare workers for high amounts of SARS-CoV-2 virus using Ct values will identify pre-symptomatic subjects who will develop disease in the next few weeks. Identification of potentially contagious, pre-symptomatic subjects is likely critical for protecting patients and healthcare workers. Main pointHealthy healthcare workers with low amounts of SARS-CoV-2 nucleic acids will previously have had the disease. Presence of a high amount of SARS-CoV-2 nucleic acids predicts future symptomatic disease.
ABSTRACT
Opportunistic bacteria in apical periodontitis (AP) may pose a risk for systemic dissemination. Mucosal-associated invariant T (MAIT) cells are innate-like T cells with a broad and potent antimicrobial activity important for gut mucosal integrity. It was recently shown that MAIT cells are present in the oral mucosal tissue, but the involvement of MAIT cells in AP is unknown. Here, comparison of surgically resected AP and gingival tissues demonstrated that AP tissues express significantly higher levels of Vα7.2-Jα33, Vα7.2-Jα20, Vα7.2-Jα12, Cα and tumour necrosis factor (TNF), interferon (IFN)-γ and interleukin (IL)-17A transcripts, resembling a MAIT cell signature. Moreover, in AP tissues the MR1-restricted MAIT cells positive for MR1-5-OP-RU tetramer staining appeared to be of similar levels as in peripheral blood but consisted mainly of CD4 subset. Unlike gingival tissues, the AP microbiome was quantitatively impacted by factors like fistula and high patient age and had a prominent riboflavin-expressing bacterial feature. When merged in an integrated view, the examined immune and microbiome data in the sparse partial least squares discriminant analysis could identify bacterial relative abundances that negatively correlated with Vα7.2-Jα33, Cα, and IL-17A transcript expressions in AP, implying that MAIT cells could play a role in the local defence at the oral tissue barrier. In conclusion, we describe the presence of MAIT cells at the oral site where translocation of oral microbiota could take place. These findings have implications for understanding the immune sensing of polymicrobial-related oral diseases.
