Machine learning assisted analysis on TCR profiling data from COVID-19-convalescent and healthy individuals unveils cross-reactivity between SARS-CoV-2 and a wide spectrum of pathogens and other diseases (preprint)

During the last two years, the emergence of SARS-CoV-2 has led to millions of deaths worldwide, with a devastating socio-economic impact on a global scale. The scientific community’s focus has recently shifted towards the association of the T cell immunological repertoire with COVID-19 progression and severity, by utilising T cell receptor sequencing (TCR-Seq) assays. The Multiplexed Identification of T cell Receptor Antigen (MIRA) dataset, which is a subset of the immunoACCESS © study, provides thousands of TCRs that can specifically recognize SARS-CoV-2 epitopes. Our study proposes a novel Machine Learning (ML) assisted approach for analysing TCR-Seq data from the antigens’ point of view, with the ability to accurately distinguish between COVID-19-convalescent and healthy individuals in the case of MIRA dataset. Most SARS-CoV-2 antigens were found to exhibit equal levels of recognition by MIRA TCRs in both convalescent and healthy cohorts, leading to the assumption of putative cross-reactivity between SARS-CoV-2 and other infectious agents. This hypothesis was validated by combining MIRA with other public TCR profiling repositories that host assays and sequencing data concerning a plethora of pathogens. Our study provides evidence regarding the cross-reactivity between SARS-CoV-2 and a wide spectrum of pathogens and diseases, with M. tuberculosis and Influenza virus exhibiting the highest levels of cross-reactivity. These results can potentially shift the emphasis of immunological studies towards an increased application of TCR profiling assays that have the potential to uncover key mechanisms of cell-mediated immune response against pathogens and diseases.

Wastewater monitoring using a novel, cost-effective PCR-based method that rapidly captures the transition patterns of SARS-CoV-2 variant prevalence (from Delta to Omicron) in the absence of conventional surveillance evidence (preprint)

Background: The magnitude of Omicron SARS-CoV-2 variant of concern (VOC) global spread necessitates reinforcement of surveillance implementation. Conventional VOC surveillance based on genotyping of clinical samples is characterized by certain challenges related to available sequencing capacity, population sampling methodologies, and demands in terms of time, labor, and resources. Wastewater-based SARS-CoV-2 VOC surveillance constitutes a valuable supplementary practice, since it does not require extensive sampling, and provides information on the prevalence of the disease in a timely and cost-effective manner. Methods: A highly sensitive real-time RT-PCR assay was developed, for targeted Omicron VOC detection and quantification in wastewater samples. The assay exclusively amplifies sequences with the S:{Delta}69/70 deletion, thus its performance is not hampered by the presence of the Delta VOC. The method was incorporated in the analysis of composite daily samples taken from the main Wastewater Treatment Plant (WWTP) of Thessaloniki, Greece from 1 December 2021 to 9 January 2022. Results: The Omicron VOC was detected for the first time in samples from the Thessaloniki WWTP on 19 December 2021. In the following 10-day period, a rapid increase in Omicron sewage viral load was observed with an estimated early doubling time of 1.86 days. The proportion of the total SARS-CoV-2 load attributed to Omicron reached 91.09% on 7 January, revealing a fast Delta-to-Omicron VOC transition pattern. The detection of Omicron in wastewater preceded the outburst of reported (presumable) Omicron cases in the city by approximately 7 days. Conclusions: The proposed wastewater-based surveillance approach enables rapid, real-time data acquisition on the Omicron VOC prevalence and transmission dynamics. Timely provision of these results to State authorities can readily influence the decision-making process for targeted public health interventions, including control measures, awareness, and preparedness.

A Thermostable, Flexible RNA Vaccine Delivery Platform for Pandemic Response (preprint)

Current RNA vaccines against SARS-CoV-2 are limited by instability of both the RNA and the lipid nanoparticle delivery system, requiring storage at -20{degrees}C or -70{degrees}C and compromising universally accessible vaccine distribution. This study demonstrates the thermostability and adaptability of a nanostructured lipid carrier (NLC) RNA vaccine delivery system for use in pandemic preparedness and pandemic response. Liquid NLC is stable at refrigerated temperatures for [≥] 1 year, enabling stockpiling and rapid deployment by point-of-care mixing with any vaccine RNA. Alternatively, NLC complexed with RNA may be readily lyophilized and stored at room temperature for [≥] 8 months or refrigerated temperature for [≥] 21 months. This thermostable RNA vaccine platform could significantly improve distribution of current and future pandemic response vaccines, particularly in low-resource settings.

Evolution of ACE2 and SARS-CoV-2 Interplay Across 247 Vertebrates (preprint)

Severe acute respiratory syndrome coronavirus 2 (SARS CoV 2) cause the most serious pandemics of Coronavirus Disease 2019 (COVID 19), which threatens human health and public safety. SARS-CoV-2 spike (S) protein uses angiotensin-converting enzyme 2 (ACE2) as recognized receptor for its entry into host cell that contributes to the infection of SARS-CoV-2 to hosts. Using computational modeling approach, this study resolved the evolutionary pattern of bonding affinity of ACE2 in 247 jawed vertebrates to the spike (S) protein of SARS-CoV-2. First, high-or-low binding affinity phenotype divergence of ACE2 to the S protein of SARS-CoV-2 has appeared in two ancient species of jawed vertebrates, Scyliorhinus torazame (low-affinity, Chondrichthyes) and Latimeria chalumnae (high-affinity, Coelacanthimorpha). Second, multiple independent affinity divergence events recur in fishes, amphibians-reptiles, birds, and mammals. Third, high affinity phenotypes go up in mammals, possibly implying the rapid expansion of mammals might accelerate the evolution of coronaviruses. Fourth, we found natural mutations at eight amino acid sites of ACE2 can determine most of phenotype divergences of bonding affinity in 247 vertebrates and resolved their related structural basis. Moreover, we also identified high-affinity or low-affinity-associated concomitant mutation group.The group linked to extremely high affinity may provide novel potentials for the development of human recombinant soluble ACE2 (hrsACE2) in treating patients with COVID-19 or for constructing genetically modified SARS-CoV-2 infection models promoting vaccines studies. These findings would offer potential benefits for the treatment and prevention of SARS-CoV-2. Keywords: Vertebrates, ACE2, SARS-CoV-2, Bonding Affinity

Computational insights into differential interaction of mamalian ACE2 with the SARS-CoV-2 spike receptor binding domain (preprint)

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causing agent of the COVID-19 pandemic, has spread globally. Angiotensin-converting enzyme 2 (ACE2) has been identified as the host cell receptor that binds to receptor-binding domain (RBD) of the SARS-COV-2 spike protein and mediates cell entry. Because the ACE2 proteins are widely available in mammals, it is important to investigate the interactions between the RBD and the ACE2 of other mammals. Here we analyzed the sequences of ACE2 proteins from 16 mammals and predicted the structures of ACE2-RBD complexes. Analyses on sequence, structure, and dynamics synergistically provide valuable insights into the interactions between ACE2 and RBD. The comparison results suggest that the ACE2 of bovine, cat and panda form strong binding with RBD, while in the cases of rat, least horseshoe bat, horse, pig, mouse and civet, the ACE2 proteins interact weakly with RBD.

Molecular epidemiology of SARS-CoV-2 in Greece reveals low rates of onward virus transmission after lifting of travel restrictions based on risk assessment during summer 2020 (preprint)

Molecular epidemiology has provided an additive value to traditional public health tools by identifying SARS-CoV-2 clusters, or providing evidence that clusters based on virus sequences and contact tracing are highly concordant. Our aim was to infer the levels of virus importation and to estimate the impact of public health measures related to travel restrictions to local transmission in Greece. Our phylogenetic and phylogeographic analyses included 389 SARS-CoV-2 sequences collected during the first 7 months of the pandemic in Greece and a random collection in 5 replicates of 3,000 sequences sampled globally, as well as the best hits to our dataset identified by BLAST. Phylogenetic analyses revealed the presence of 70 genetically distinct viruses identified as independent introductions into Greece. The proportion of imported strains was 41%, 11.5%, and 8.8% during the three periods of sampling, namely, March (no travel restrictions), April to June (strict travel restrictions), and July to September (lifting of travel restrictions based on a thorough risk assessment), respectively. These findings reveal low levels of onward transmission from imported cases during summer and underscore the importance of targeted public health measures that can increase the safety of international travel during a pandemic.

Assessing training needs in infectious disease management at major ports, airports and ground-crossings in Europe (preprint)

Background: The implementation of core capacities as stated in the International Health Regulations (IHR) is far from complete, and, as the COVID-19 pandemic shows, the spreading of infectious diseases through points of entry (POEs) is a serious problem. To guide training and exercises, we performed a training needs assessment on infectious disease management among professionals at European POE. Methods: We disseminated a digital questionnaire to representatives of designated airports, ports, and ground-crossings in Europe. Topics were derived from the IHR core capacities for POEs. Based on the importance (4-point Likert scale) and training needs (4-point Likert scale), we identified the topics with the highest priority for training. These results were put in further perspective using prior experience (training <3 year, exercise <5 years, events <5 years). Also, preferences for training methodologies were assessed. Results: Fifty questionnaires were included in the analyses, representing 50 POEs from 19 European countries. Importance is high for 26/30 topics, although scores widely vary among respondents. Topics with a high training need (16/30) are amongst others the handling of ill travelers; using and composing the public health emergency contingency plan, and public health measures. Respondents from ports and airports attribute equal importance to most topics, but respondents from ports showed higher training needs on 75% of the topics. POEs are unevenly and generally little experienced. The most preferred training methods were presentations. Simulation is the preferred methodology for training the handling of ill or exposed travelers. Conclusions: The European workforce at designated ports, airports and ground-crossings has a different level of experience and perceives varying importance of the topics assessed in our study. We identified the topics on which training is required. We call for European collaboration between POEs to agree upon the importance of infectious disease management, and to jointly build a trained and prepared workforce that is ready to face the next crisis.