Dynamics of viral DNA shedding and culture viral DNA positivity in different clinical samples collected during the 2022 mpox outbreak in Lombardy, Italy.

BACKGROUND: Mpox virus (MPXV) has recently spread outside of sub-Saharan Africa. This large multicentre study was conducted in Lombardy, the most densely populated Italian region accounting for more than 40% of Italian cases. The present study aims to: i) evaluate the presence and the shedding duration of MPXV DNA in different body compartments correlating the MPXV viability with the time to onset of symptoms; ii) provide evidence of MPXV persistence in different body compartment as a source of infection and iii) characterize the MPXV evolution by whole genome sequencing (WGS) during the outbreak occurred in Italy. MATERIAL AND METHODS: The study included 353 patients with a laboratory-confirmed diagnosis of MPXV infection screened in several clinical specimens in the period May 24th - September 1st, 2022. Viral isolation was attempted from different biological matrices and complete genome sequencing was performed for 61 MPXV strains. RESULTS: MPXV DNA detection was more frequent in the skin (94.4%) with the longest median time of viral clearance (16 days). The actively-replicating virus in cell culture was obtained for 123/377 (32.6%) samples with a significant higher viral quantity on isolation positive samples (20 vs 31, p < 0.001). The phylogenetic analysis highlighted the high genetic identity of the MPXV strains collected, both globally and within the Lombardy region. CONCLUSION: Skin lesion is gold standard material and the high viral load and the actively-replicating virus observed in genital sites confirms that sexual contact plays a key role in the viral transmission.

Fourth dose of microneedle array patch of SARS-CoV-2 S1 protein subunit vaccine elicits robust long-lasting humoral responses in mice.

The COVID-19 pandemic has underscored the pressing need for safe and effective booster vaccines, particularly in considering the emergence of new SARS-CoV-2 variants and addressing vaccine distribution inequalities. Dissolving microneedle array patches (MAP) offer a promising delivery method, enhancing immunogenicity and improving accessibility through the skin's immune potential. In this study, we evaluated a microneedle array patch-based S1 subunit protein COVID-19 vaccine candidate, which comprised a bivalent formulation targeting the Wuhan and Beta variant alongside a monovalent Delta variant spike proteins in a murine model. Notably, the second boost of homologous bivalent MAP-S1(WU + Beta) induced a 15.7-fold increase in IgG endpoint titer, while the third boost of heterologous MAP-S1RS09Delta yielded a more modest 1.6-fold increase. Importantly, this study demonstrated that the administration of four doses of the MAP vaccine induced robust and long-lasting immune responses, persisting for at least 80 weeks. These immune responses encompassed various IgG isotypes and remained statistically significant for one year. Furthermore, neutralizing antibodies against multiple SARS-CoV-2 variants were generated, with comparable responses observed against the Omicron variant. Overall, these findings emphasize the potential of MAP-based vaccines as a promising strategy to combat the evolving landscape of COVID-19 and to deliver a safe and effective booster vaccine worldwide.

Second Boost of Omicron SARS-CoV-2 S1 Subunit Vaccine Induced Broad Humoral Immune Responses in Elderly Mice.

Currently approved COVID-19 vaccines prevent symptomatic infection, hospitalization, and death from the disease. However, repeated homologous boosters, while considered a solution for severe forms of the disease caused by new SARS-CoV-2 variants in elderly individuals and immunocompromised patients, cannot provide complete protection against breakthrough infections. This highlights the need for alternative platforms for booster vaccines. In our previous study, we assessed the boost effect of the SARS-CoV-2 Beta S1 recombinant protein subunit vaccine (rS1Beta) in aged mice primed with an adenovirus-based vaccine expressing SARS-CoV-2-S1 (Ad5.S1) via subcutaneous injection or intranasal delivery, which induced robust humoral immune responses (1). In this follow-up study, we demonstrated that a second booster dose of a non-adjuvanted recombinant Omicron (BA.1) S1 subunit vaccine with Toll-like receptor 4 (TLR4) agonist RS09 (rS1RS09OM) was effective in stimulating strong S1-specific immune responses and inducing significantly high neutralizing antibodies against the Wuhan, Delta, and Omicron variants in 100-week-old mice. Importantly, the second booster dose elicits cross-reactive antibody responses, resulting in ACE2 binding inhibition against the spike protein of SARS-CoV-2 variants, including Omicron (BA.1) and its subvariants. Interestingly, the levels of IgG and neutralizing antibodies correlated with the level of ACE2 inhibition in the booster serum samples, although Omicron S1-specific IgG level showed a weaker correlation compared to Wuhan S1-specific IgG level. Furthermore, we compared the immunogenic properties of the rS1 subunit vaccine in young, middle-aged, and elderly mice, resulting in reduced immunogenicity with age, especially an impaired Th1-biased immune response in aged mice. Our findings demonstrate that the new variant of concern (VOC) rS1 subunit vaccine as a second booster has the potential to offer cross-neutralization against a broad range of variants and to improve vaccine effectiveness against newly emerging breakthrough SARS-CoV-2 variants in elderly individuals who were previously primed with the authorized vaccines.

Tetravalent SARS-CoV-2 S1 subunit protein vaccination elicits robust humoral and cellular immune responses in SIV-infected rhesus macaque controllers.

IMPORTANCE: The study provides important insights into the immunogenicity and efficacy of a tetravalent protein subunit vaccine candidate against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The vaccine induced both humoral and cellular immune responses in nonhuman primates with controlled SIVagm infection and was able to generate Omicron variant-specific antibodies without specifically vaccinating with Omicron. These findings suggest that the tetravalent composition of the vaccine candidate could provide broad protection against multiple SARS-CoV-2 variants while minimizing the risk of immune escape and the emergence of new variants. Additionally, the use of rhesus macaques with controlled SIVsab infection may better represent vaccine immunogenicity in humans with chronic viral diseases, highlighting the importance of preclinical animal models in vaccine development. Overall, the study provides valuable information for the development and implementation of coronavirus disease 2019 vaccines, particularly for achieving global vaccine equity and addressing emerging variants.

Hierarchical AI enables global interpretation of culture plates in the era of digital microbiology.

Full Laboratory Automation is revolutionizing work habits in an increasing number of clinical microbiology facilities worldwide, generating huge streams of digital images for interpretation. Contextually, deep learning architectures are leading to paradigm shifts in the way computers can assist with difficult visual interpretation tasks in several domains. At the crossroads of these epochal trends, we present a system able to tackle a core task in clinical microbiology, namely the global interpretation of diagnostic bacterial culture plates, including presumptive pathogen identification. This is achieved by decomposing the problem into a hierarchy of complex subtasks and addressing them with a multi-network architecture we call DeepColony. Working on a large stream of clinical data and a complete set of 32 pathogens, the proposed system is capable of effectively assist plate interpretation with a surprising degree of accuracy in the widespread and demanding framework of Urinary Tract Infections. Moreover, thanks to the rich species-related generated information, DeepColony can be used for developing trustworthy clinical decision support services in laboratory automation ecosystems from local to global scale.

Quantitative SARS-CoV-2 subgenomic RNA as a surrogate marker for viral infectivity: Comparison between culture isolation and direct sgRNA quantification.

Detection of subgenomic (sg) SARS-CoV-2 RNAs are frequently used as a correlate of viral infectiousness, but few data about correlation between sg load and viable virus are available. Here, we defined concordance between culture isolation and E and N sgRNA quantification by ddPCR assays in 51 nasopharyngeal swabs collected from SARS-CoV-2 positive hospitalized patients. Among the 51 samples, 14 were SARS-CoV-2 culture-positive and 37 were negative. According to culture results, the sensitivity and specificity of E and N sgRNA assays were 100% and 100%, and 84% and 86%, respectively. ROC analysis showed that the best E and N cut-offs to predict positive culture isolation were 32 and 161 copies/mL respectively, with an AUC (95% CI) of 0.96 (0.91-1.00) and 0.96 (0.92-1.00), and a diagnostic accuracy of 88% and 92%, respectively. Even if no significant correlations were observed between sgRNA amount and clinical presentation, a higher number of moderate/severe cases and lower number of days from symptoms onset characterized patients with sgRNA equal to or higher than sgRNA cut-offs. Overall, this study suggests that SARS-CoV-2 sgRNA quantification could be helpful to estimate the replicative activity of SARS-CoV-2 and can represent a valid surrogate marker to efficiently recognize patients with active infection. The inclusion of this assay in available SARS-CoV-2 diagnostics procedure might help in optimizing fragile patients monitoring and management.

Autoantibodies neutralizing type I IFNs underlie West Nile virus encephalitis in ∼40% of patients.

Mosquito-borne West Nile virus (WNV) infection is benign in most individuals but can cause encephalitis in <1% of infected individuals. We show that ∼35% of patients hospitalized for WNV disease (WNVD) in six independent cohorts from the EU and USA carry auto-Abs neutralizing IFN-α and/or -ω. The prevalence of these antibodies is highest in patients with encephalitis (∼40%), and that in individuals with silent WNV infection is as low as that in the general population. The odds ratios for WNVD in individuals with these auto-Abs relative to those without them in the general population range from 19.0 (95% CI 15.0-24.0, P value <10-15) for auto-Abs neutralizing only 100 pg/ml IFN-α and/or IFN-ω to 127.4 (CI 87.1-186.4, P value <10-15) for auto-Abs neutralizing both IFN-α and IFN-ω at a concentration of 10 ng/ml. These antibodies block the protective effect of IFN-α in Vero cells infected with WNV in vitro. Auto-Abs neutralizing IFN-α and/or IFN-ω underlie ∼40% of cases of WNV encephalitis.

SARS-CoV-2 S1 Subunit Booster Vaccination Elicits Robust Humoral Immune Responses in Aged Mice.

The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants has raised concerns about reduced vaccine effectiveness and the increased risk of infection, and while repeated homologous booster shots are recommended for elderly and immunocompromised individuals, they cannot completely protect against breakthrough infections. In our previous study, we assessed the immunogenicity of an adenovirus-based vaccine expressing SARS-CoV-2 S1 (Ad5.S1) in mice, which induced robust humoral and cellular immune responses (E. Kim, F. J. Weisel, S. C. Balmert, M. S. Khan, et al., Eur J Immunol 51:1774-1784, 2021, https://doi.org/10.1002/eji.202149167). In this follow-up study, we found that the mice had high titers of anti-S1 antibodies 1 year after vaccination, and one booster dose of the nonadjuvanted rS1Beta (recombinant S1 protein of SARS-CoV-2 Beta [B.1.351]) subunit vaccine was effective at stimulating strong long-lived S1-specific immune responses and inducing significantly high neutralizing antibodies against Wuhan, Beta, and Delta strains, with 3.6- to 19.5-fold increases. Importantly, the booster dose also elicited cross-reactive antibodies, resulting in angiotensin-converting enzyme 2 (ACE2) binding inhibition against spikes of SARS-CoV-2, including Omicron variants, persisting for >28 weeks after booster vaccination. Interestingly, the levels of neutralizing antibodies were correlated not only with the level of S1 binding IgG but also with ACE2 inhibition. Our findings suggest that the rS1Beta subunit vaccine candidate as a booster has the potential to offer cross-neutralization against broad variants and has important implications for the vaccine control of newly emerging breakthrough SARS-CoV-2 variants in elderly individuals primed with adenovirus-based vaccines like AZD1222 and Ad26.COV2.S. IMPORTANCE Vaccines have significantly reduced the incidences of severe coronavirus disease 2019 (COVID-19) cases and deaths. However, the emergence of SARS-CoV-2 variants has raised concerns about their increased transmissibility and ability to evade neutralizing antibodies, especially among elderly individuals who are at higher risks of mortality and reductions of vaccine effectiveness. To address this, a heterologous booster vaccination strategy has been considered as a solution to protect the elderly population against breakthrough infections caused by emerging variants. This study evaluated the booster effect of an S1 subunit vaccine in aged mice that had been previously primed with adenoviral vaccines, providing valuable preclinical evidence for elderly people vaccinated with the currently approved COVID-19 vaccines. This study confirms the potential for using the S1 subunit vaccine as a booster to enhance cross-neutralizing antibodies against emerging variants of concern.

Characterization of immune response against monkeypox virus in cohorts of infected patients, historic and newly vaccinated subjects.

Monkeypox virus (MPXV) is a zoonotic disease endemic in the rainforest countries of Central and West Africa. Understanding the immune response in zoonosis is fundamental to prevent and contrast viral spreading. MPXV is a close relative of Variola (smallpox) virus and vaccination with vaccinia virus gives approximatively 85% of protection against MPXV. With the emergence of the recent MPXV outbreak, JYNNEOS vaccine has been proposed to individuals at high-risk of exposure. Comparative data on MPXV immune response in vaccinated or infected subjects are still limited. Here we set-up an immunofluorescence method for the evaluation of humoral response elicited by natural infection and healthy vaccinated subjects, including historically smallpox-vaccinated individuals and newly vaccinated subjects. Neutralization assay was also included, and in vaccinated subjects, cell-mediated response was evaluated. We observed that the natural infection produces a strong immune response that can control the disease. In naïve subjects, a second dose boosts the serological response to levels similar to those of the MPXV patients. Last, smallpox-vaccinated controls retain a degree of protection, even after years from vaccination, most visible in the t-cellular response.

Persistence of Immune Response Elicited by Three Doses of mRNA Vaccine against SARS-CoV-2 in a Cohort of Patients with Solid Tumors: A One-Year Follow-Up.

The role and durability of the immunogenicity of the BNT162b2 mRNA vaccine against severe acute respiratory virus 2 (SARS-CoV-2), in cancer patients one year after receiving the third dose have to be elucidated. We have prospectively evaluated the long-term immunogenicity of the third dose of the SARS-CoV-2 BNT162b2 mRNA vaccine in 55 patients undergoing active treatment. Neutralizing antibody (NT Ab) titers against Omicron variants and total anti-trimeric S IgG levels were measured one year after the third dose. Heparinized whole-blood samples were used for the assessment of the SARS-CoV-2 interferon-γ release assay (IGRA). Thirty-seven patients (67.3%) showed positive total anti-trimeric S IgG one year after the third dose. Looking at the T-cell response against the spike protein, the frequency of responder patients did not decrease significantly between six and twelve months after the third dose. Finally, less than 20% of cancer patients showed an undetectable NT Ab titer against BA.1 and BA.5 variants of concern (VOCs). Underlying therapies seem to not affect the magnitude or frequency of the immune response. Our work underlines the persistence of humoral and cellular immune responses against BNT162b2 in a cohort of cancer patients one year after receiving the third dose, regardless of the type of underlying therapy.

19n01, a broadly neutralizing antibody against omicron BA.1, BA.2, BA.4/5, and other SARS-CoV-2 variants of concern.

This study reports the isolation and characterization of a human monoclonal antibody (mAb) called 19n01. This mAb was isolated by using single-cell RNAseq of B cells from donors infected with the ancestral strain. This mAb possesses a potent and broad capacity to bind and neutralize all previously circulating variants of concern (VOCs), including Omicron sublineages BA.1, BA.2, and BA.4/5. The pseudovirus neutralization assay revealed robust neutralization capacity against the G614 strain, BA.1, BA.2, and BA.4/5, with inhibitory concentration (IC50) values ranging from 0.0035 to 0.0164 µg/mL. The microneutralization assay using the G614 strain and VOCs demonstrated IC50 values of 0.013-0.267 µg/mL. Biophysical and structural analysis showed that 19n01 cross-competes with ACE2 binding to the receptor-binding domain (RBD) and the kinetic parameters confirmed the high affinity against the Omicron sublineages (KD of 61 and 30 nM for BA.2 and BA.4/5, respectively). These results suggest that the 19n01 is a remarkably potent and broadly reactive mAb.

Real-life lack of evidence of viable SARS-CoV-2 transmission via inanimate surfaces: The SURFACE study.

INTRODUCTION: Although the potential role of inanimate surfaces in SARS-CoV-2 transmission has yet to be adequately assessed, it is still routine practice to apply deep and expensive environmental disinfection protocols. The aim of this study was to verify the presence of viable virus on different surfaces exposed to droplets released by coughing in SARS-CoV-2 RNA positive patients. METHODS: Patients admitted to hospital with a positive SARS-CoV-2 real-time (RT)-PCR swab were asked to cough on steel, cardboard, plastic and their hands. Surfaces were tested at baseline (T0) and at different timepoints thereafter using swabs dipped in medium, and quickly seeded on VERO E6 cells that were checked every other day for cytopathic effect (CPE). Laboratory-propagated SARS-CoV-2 strains were examined at the same time points and on identical materials. RESULTS: Ten RNA-positive patients were enrolled into the study. The median cycle threshold value was 20.7 (range 13-28.3). Nasopharyngeal swabs from 3 of the patients yielded viable virus 2-10 days post-inoculation. However, in none of the patients was it possible to isolate viable SARS-CoV-2 from sputum under identical experimental conditions. A CPE was instead already visible using laboratory-propagated SARS-CoV-2 strains at 20', 60', 180' while an effect at 24 h required a 6-day incubation. CONCLUSION: The evidence emerging from this real-life study suggests that droplets delivered by SARS-CoV-2 infected patients on common inanimate surfaces did not contain viable virus. In contrast, and in line with several laboratory-based experiments, in vitro adapted viruses could survive and grow on the same fomites.

Tetravalent SARS-CoV-2 S1 Subunit Protein Vaccination Elicits Robust Humoral and Cellular Immune Responses in SIV-Infected Rhesus Macaque Controllers.

The COVID-19 pandemic has highlighted the need for safe and effective vaccines to be rapidly developed and distributed worldwide, especially considering the emergence of new SARS-CoV-2 variants. Protein subunit vaccines have emerged as a promising approach due to their proven safety record and ability to elicit robust immune responses. In this study, we evaluated the immunogenicity and efficacy of an adjuvanted tetravalent S1 subunit protein COVID-19 vaccine candidate composed of the Wuhan, B.1.1.7 variant, B.1.351 variant, and P.1 variant spike proteins in a nonhuman primate model with controlled SIVsab infection. The vaccine candidate induced both humoral and cellular immune responses, with T- and B cell responses mainly peaking post-boost immunization. The vaccine also elicited neutralizing and cross-reactive antibodies, ACE2 blocking antibodies, and T-cell responses, including spike specific CD4+ T cells. Importantly, the vaccine candidate was able to generate Omicron variant spike binding and ACE2 blocking antibodies without specifically vaccinating with Omicron, suggesting potential broad protection against emerging variants. The tetravalent composition of the vaccine candidate has significant implications for COVID-19 vaccine development and implementation, providing broad antibody responses against numerous SARS-CoV-2 variants.

Differential Kinetics of Effector and Memory Responses Induced by Three Doses of SARS-CoV-2 mRNA Vaccine in a Cohort of Healthcare Workers.

We reported the long-term kinetics of immune response after vaccination and evaluated the immunogenicity after a third dose of mRNA vaccine in 86 healthcare workers. Humoral response was analyzed by measuring anti-spike IgG and SARS-CoV-2 NTAbs titer; cell-mediated response was measured as frequency of IFN-γ producing T-cells and cell proliferation. Memory B cells secreting SARS-CoV-2 RBD-IgG were measured by B-spot assay. At three weeks after the third dose (T4), the frequency of subjects showing NT-Abs titer at the upper detection limit (≥640) was significantly higher than that observed at three weeks after the second dose (26/77; 33.7% vs. 9/77; 11.6%; p = 0.0018). Additionally, at T4, all the subjects reached positive levels of T-cell mediated response (median 110 SFU/106 PBMC, IQR 73-231). While the number of IFNγ-producing T-cells decreased between second and third dose administration, the T-cell proliferative response did not decrease but was sustained during the follow-up. Among T-cell subsets, a higher proliferative response was observed in CD4+ than in CD8+ population. Moreover, even if a decline in antibody response was observed between the second and third dose, a sustained persistence of memory B cells was observed. Subsequently, the third dose did not affect the frequency of memory B cells, while it restored or increased the peak antibody levels detected after the second dose.

Variant of Concern-Matched COVID-19 Convalescent Plasma Usage in Seronegative Hospitalized Patients.

COVID-19 convalescent plasma (CCP) has been the only specific anti-viral therapy against SARS-CoV-2 available for more than one year. Following the negative results from most randomized controlled trials on its efficacy in COVID-19 hospitalized patients and the availability of anti-spike monoclonal antibodies (mAbs), the use of CCP has subsequently rapidly faded. However, the continuous appearance of new variants of concern (VOCs), most of which escape mAbs and vaccine-elicited neutralizing antibodies (nAbs), has renewed the interest towards CCP, at least in seronegative immunocompetent patients, and in immunocompromised patients not able to mount a protective immune response. We report here the experience of a single Italian hospital in collecting and transfusing CCP in immunocompromised patients hospitalized for severe COVID-19 between October 2021 and March 2022. During this 6-month period, we collected CCP from 32 vaccinated and convalescent regular blood donors, and infused high nAb-titer CCP units (titered against the specific VOC affecting the recipient) to 21 hospitalized patients with severe COVID-19, all of them seronegative at the time of CCP transfusion. Patients' median age was 66 years (IQR 50-74 years) and approximately half of them (47.6%, 10/21) were immunocompromised. Two patients were rescued after previous failure of mAbs. No adverse reactions following CCP transfusion were recorded. A 28-day mortality rate of 14.3 percent (3/21) was reported, with age, advanced disease stage and late CCP transfusion associated with a worse outcome. This real-life experience also supports the use of CCP in seronegative hospitalized COVID-19 patients during the Delta and Omicron waves.

Optimizing the quality of research in Veterinary Parasitology.

Countless research is carried out until new discoveries are transformed into products or services available to the population. This trajectory can be slower and more costly or even impossible when irreproducible data are obtained in the most diverse fields of science. Thus, quality management appears as an essential tool to guarantee the reliability of academic research results. In this work, we demonstrate the applied strategy to implement a Quality Management System (QMS) in a research laboratory in Veterinary Parasitology and we highlight the adaptable quality requirements in this scientific research environment. For this, the Plan-Do-Check-Act (PDCA) quality tool was used, and two internal audits were performed, one before and one after implementation. The audits reached 67 (41.36%) and 157 (96.91%) points before and after implementation, respectively, with a significant difference between the moments studied. Thus, we demonstrate that the adoption of QMS principles in research is feasible. The methodology applied in this work can be adopted by managers from other laboratories interested in the implementation of quality standards as a support in the reproducibility of research.

Evaluation of the Neutralizing Antibodies Response against 14 SARS-CoV-2 Variants in BNT162b2 Vaccinated Naïve and COVID-19 Positive Healthcare Workers from a Northern Italian Hospital.

SARS-CoV-2 still represents a global health burden, causing more than six million deaths worldwide. Moreover, the emergence of new variants has posed new issues in terms of vaccine efficacy and immunogenicity. In this study, we aimed to evaluate the neutralizing antibody response against SARS-CoV-2 variants in different cohorts of vaccinated and unvaccinated subjects. Four-fold diluted sera from SARS-CoV-2 naïve and recovered subjects vaccinated with two or three doses of the BNT162b2 vaccine were challenged against 14 SARS-CoV-2 variants, and the SARS-CoV-2 neutralizing antibody titer was measured. Results were compared with those obtained from unvaccinated COVID-19 recovered patients. Overall, a better SARS-CoV-2 NT Abs response was observed in recovered vaccinated subjects after three doses of the vaccine when compared to unvaccinated patients and vaccinated subjects with only two doses. Additionally, the lowest level of response was observed against the Omicron variant. In conclusion, third doses of BNT162b2 vaccine seems to elicit a sustained response against the large majority of variants.

Neutralizing antibody levels against SARS-CoV-2 variants of concern Delta and Omicron in vaccine breakthrough-infected blood donors.

BACKGROUND: Novel SARS-CoV-2 variants of concern (VOC) Delta and Omicron are able to escape some monoclonal antibody therapies, making again COVID-19 convalescent plasma (CCP) a potential frontline treatment. STUDY DESIGN/METHODS: In this study, we investigated the kinetics of anti-SARS-CoV-2 neutralizing antibodies (nAbs) against VOCs Delta and Omicron in vaccine breakthrough infected plasma donors. Serum samples from 19 donors were collected at the time of plasma donation and tested for anti-SARS-CoV-2 nAbs (using live authentic VOC viral neutralization test) and IgG (Liaison® SARS-CoV-2 S1/S2 and Liaison® SARS-CoV-2 TrimericS IgG assays, DiaSorin). Measures were correlated with different variables, including the time between last vaccine dose and CCP donation, and time between SARS-COV-2 infection and CCP donation. RESULTS: nAb titers against VOC Delta and Omicron were directly related to the time interval since last vaccine dose to CCP donation, but inversely related to time since COVID19 breakthrough infection. DISCUSSION: SARS-CoV-2 breakthrough infection in vaccinated in donors boosts nAb titers against VOCs Delta and Omicron, but such titers decay shortly after infection. Therefore, CCP must be collected early after vaccine breakthrough infection.

SARS-CoV-2 variants inactivation of plasma units using a riboflavin and ultraviolet light-based photochemical treatment.

BACKGROUND: Test the ability of Mirasol Pathogen Reduction Technology (PRT, Terumo BCT, Lakewood Co, USA) treatment with riboflavin and ultraviolet light (R + UV) in reducing SARS-CoV-2 infectivity while maintaining blood product quality. MATERIAL AND METHODS: SARS-CoV-2 strains were isolated and titrated to prepare cell free virus for plasma units infection. The units were then under treatment with Mirasol PRT. The infectious titers were determined before and after treatment with an in house microtitration assay on Vero E6 cells. Thirty-six plasma pool bags underwent PRT treatment. RESULTS: In all the experiments, the measured titer following riboflavin and UV treatment was below the limit of detection of microtitration assay for all the different SARS-CoV-2 strains. Despite the high copies number detected by RT-PCR for each viral strain after treatment, viruses were completely inactivated and not able to infect VERO E6 cells. CONCLUSION: Riboflavin and UV light treatment effectively reduced the virus titers of human plasma to the limit of detection in tissue culture, regardless of the strain. These data suggest that pathogen reduction in blood products highlight the safety of CP therapy procedures for critically ill COVID-19 patients, while maintaining blood product quality.