Despite recent advances in prophylactic vaccination, SARS-CoV-2 infections continue to cause significant morbidity. A better understanding of immune response differences between vaccinated individuals with and without later SARS-CoV-2 breakthrough infection is urgently needed. CoV-ADAPT is a prospective long-term study comparing humoral (anti-spike-RBD-IgG, neutralization capacity, avidity) and cellular (spike-induced T-cell interferon-γ release) immune responses in individuals vaccinated against SARS-CoV-2 at four different time points (three before and one after third vaccination). In this cohort study, 62 fully vaccinated individuals presented with SARS‑CoV-2 breakthrough infections vs 151 without infection 3-7 months following third vaccination. Breakthrough infections significantly increased anti-spike-RBD-IgG (p<0.01), but not spike-directed T-cell interferon-γ release (TC), antibody neutralization capacity or avidity. Anti-spike-RBD-IgG and antibody avidity decreased with age (p<0.01) and females showed higher anti-spike-RBD-IgG (p<0.01), and a tendency towards higher antibody avidity (p=0.051). The association between humoral and cellular immune responses previously reported at various time points was lost in subjects after breakthrough infections (p=0.807). Finally, a machine-learning approach based on our large immunological data set (a total of 49 variables) from different time points was unable to predict breakthrough infections (AUC: 0.55). In conclusion, distinct differences in humoral vs cellular immune responses in fully vaccinated individuals with or without breakthrough infection could be demonstrated. Breakthrough infections predominantly drive the humoral response without boosting the cellular component. Breakthrough infections could not be predicted based on immunological data, which indicates a superior role of environmental factors (e.g. virus exposure) in individualized risk assessment.

Rabies is an ancient neuroinvasive viral (genus Lyssavirus, family Rhabdoviridae) disease affecting approximately 59,000 people worldwide. The central nervous system (CNS) is targeted, and rabies has a case fatality rate of almost 100% in humans and animals. Rabies is entirely preventable through proper vaccination, and thus, the highest incidence is typically observed in developing countries, mainly in Africa and Asia. However, there are still cases in European countries and the US. Recently, demographic, increasing income levels, and the coronavirus disease 2019 (COVID-19) pandemic have caused a massive raising in the animal population, enhancing the need for preventive measures (e.g., vaccination, surveillance, animal control programs), post-exposure prophylaxis, and a better understanding of rabies pathophysiology to identify therapeutic targets, since there is no effective treatment after the onset of clinical manifestations. Here we review the neuroimmune biology and mechanisms of rabies. Its pathogenesis involves a complex and poorly understood modulation of immune and brain functions associated with metabolic, synaptic, and neuronal impairments, resulting in fatal outcomes without significant histopathological lesions in the CNS. In this context, the neuroimmunological and neurochemical aspects of excitatory/inhibitory signaling (e.g., GABA/glutamate crosstalk) are likely related to the clinical manifestations of rabies infection. Uncovering new links between immunopathological mechanisms and neurochemical imbalance will be essential to identify novel potential therapeutic targets to reduce rabies morbidity and mortality.

Severe fever with thrombocytopaenia syndrome virus (SFTSV) and Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) can cause the hyperproduction of inflammatory cytokines, which have pathological effects in patient including severe or fatal cytokine storms. To characterize the effect of SFTSV and SARS-CoV-2 infection on the production of cytokines in SFTS and COVID-19 patients, we performed an analysis of cytokines in SFTS and COVID-19 patients and also investigated the role of IL-10 in vitro studies: LPS-induced THP-1-derived macrophages, SFTSV infection of THP-1 cells, and SARS-CoV-2 infection of THP-1 cells. In this study, we found that levels of both IL-10 and IL-6 were significantly elevated, the level of TGF-β was significantly decreased and IL-10 was elevated earlier than IL-6 in severe and critical COVID-19 and fatal SFTS patients, and inhibition of IL-10 signalling decreased the production of IL-6 and elevated that of TGF-β. Therefore, the hyperproduction of IL-10 and IL-6 and the low production of TGF-β have been linked to cytokine storm-induced mortality in fatal SFTS and severe and critically ill COVID-19 patients and that IL-10 can play an important role in the host immune response to severe and critical SARS-CoV-2 and fatal SFTSV infection.

Omicron BF.7 became the predominant SARS-CoV-2 variant in Beijing after the adjustment of COVID-19 response strategies in December 2022. The ability of antibodies elicited by BF.7 infection to cross-react with SARS-CoV-2-like viruses is unknown. This study aimed to investigate the cross-reactive neutralizing antibodies against SARS-CoV-2-related pangolin coronavirus GX_P2V in sera from vaccinated and/or SARS-CoV-2 infected individuals. All vaccinated individuals who recovered from Omicron BF.7 breakthrough infections exhibited substantially higher levels of neutralizing antibodies against GX_P2V among collected subject populations (geometric mean titer [GMT] = 362). Uninfected individuals who received four-mixed-dose vaccines also demonstrated higher levels of neutralizing antibodies (GMT = 44) against GX_P2V than those who received two- or three-dose vaccines and those who recovered from wild type SARS-CoV-2. This finding highlights the significance of prior and hybrid booster vaccinations with wild-type SARS-CoV-2 vaccines in generating potent cross-protective immunity against future spillovers of SARS-CoV-2-like viruses.

Several biomarkers have been evaluated as predictors of severity or in directing the treatment of COVID-19, however there are no conclusive results with prediction of the pathobiology of the infection. In this study, we evaluated serum levels of cytokines, chemokines, and cell growth factors in association with the pathobiology of mild to moderate SARS-CoV-2 infection. Those markers may act as immuno-inflammatory biomarkers in adults with mild to moderate flu syndrome who sought care at health units. Serum levels of SARS-CoV-2 infected patients (n=113) and flu symptoms individuals negative for SARS-CoV-2 (n=58), tested by the RT-qPCR test - nasal swab were compared to healthy controls (n=53). Participants who were symptomatic but negative for SARS-CoV-2 were tested for Influenza A/B and Respiratory Syncytial Virus (RSV). Results showed that the pro-inflammatory cytokines IL-1β, MCP-3, TNF-α and G-CSF were increased in symptomatic patients and the cytokines IL-6 and IL-10 were associated with patients positive for SARS-CoV-2 when compared to healthy controls. Symptoms associated with COVID-19 were fever, anosmia, ageusia and myalgia. For patients without SARS-CoV-2 infection their major symptom was sore throat. Five percent (4/83) of SARS-CoV-2 negative patients were positive for RSV. The pathobiology of mild to moderate SARS-CoV-2 infection was associated with increasing pro-inflammatory cytokines and also a pleiotropic IL-6 and anti-inflammatory IL-10 cytokines compared to healthy controls.

Objectives The aim of this study was to assess the potential of noninvasive and repeated monitoring of rhACE2 distribution and content in organs using the ACE2-specific nuclide probe 68Ga-HZ20. Methods We optimized the labeling conditions of the probe and evaluated its safety. A mouse organ in situ rhACE2 high-aggregation model was constructed for the first time, and in vivo real-time PET imaging of rhACE2 was performed using the ACE2-specific PET agent 68Ga-HZ20. The distribution and uptake of the probe were analyzed, and the model was validated. Results This radiotracer exhibited reliable radiochemical properties in vitro and maintained a high affinity for rhACE2 in vivo. In terms of probe uptake, 68Ga-HZ20 showed a good target-to-nontarget ratio, and the correlation between the uptake value of the probe and the dose of rhACE2 was >90% in both models; the probe was rapidly cleared from the circulatory system and excreted by the kidneys and urinary system. No organs were damaged after the injection of high doses of probe. Conclusions This technology for noninvasively and repeatedly monitoring the content and distribution of rhACE2 in vivo aids in clarifying the resident capacity of rhACE2 in organs and in analyzing the preventive effect of rhACE2 against SARS-CoV-2 and the effectiveness of therapies for COVID-19.

Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) enters the host cell by binding to angiotensin-converting enzyme 2 (ACE2). While evolutionarily conserved, ACE2 glycoproteins differ across various species and differential interactions with Spike (S) glycoproteins of SARS-CoV-2 viruses impact species specificity. Reverse zoonoses led to SARS-CoV-2 outbreaks on multiple American mink ( Mustela vison) farms during the pandemic and gave rise to mink-associated S substitutions known for transmissibility between mink and zoonotic transmission to humans. In this study, we used bio-layer interferometry (BLI) to discern the differences in binding affinity between multiple human and mink-derived S glycoproteins of SARS-CoV-2 and their respective ACE2 glycoproteins. Further, we conducted a structural analysis of a mink variant S glycoprotein and American mink ACE2 (mvACE2) using cryo-electron microscopy (cryo-EM), revealing four distinct conformations. We discovered a novel intermediary conformation where the mvACE2 glycoprotein is bound to the receptor-binding domain (RBD) of the S glycoprotein in a “down” position, approximately 34° lower than previously reported “up” RBD. Finally, we compared residue interactions in the S-ACE2 complex interface of S glycoprotein conformations with varying RBD orientations. These findings provide valuable insights into the molecular mechanisms of SARS-CoV-2 entry.

Introduction: In this paper we examine the relationship between vaccination against COVID-19 and both the death rate from COVID-19 and the rate of COVID-19 spread. Our goal is determine if vaccination is associated with reduced death and/or spread of disease at the local level. Methods: This analysis was conducted at the county level in the state of Pennsylvania, United States of America, with data that were collected during the first half of 2022 from the state of Pennsylvania’s Covid Dashboard ([COVID-19 Data for Pennsylvania (pa.gov)](https://www.health.pa.gov/topics/disease/coronavirus/pages/cases.aspx) . Result s: Given that, during this time period, the vaccines being used were not geared specifically toward the common variants at that time, we found no statistically significant relationship between disease spread and vaccination rate at the county level. That said, we did find a highly statistically significant relationship between death rate and vaccination rate (p-value = 0.006). Specifically, a 1% increase in vaccination rate was found to correspond to a 0.751% decrease in death rate (95% confidence interval (0.236%, 1.266%)). Conclusions: These results support previous findings from across the world that Covid vaccination is highly efficacious in preventing death from the disease. Even during a time when vaccine design was not optimally matched with the prevailing strains, vaccination was found to reduce death rate. Hence, improving global vaccine availability is vitally important, in order to achieve necessary outcomes.

COVID-19 bivalent ancestral/Omicron mRNA booster vaccinations became available to boost and expand the immunity against SARS-CoV-2 Omicron infections. In a prospective cohort study including 59 healthcare workers, we assessed SARS-CoV-2 ancestral and Omicron BA.5-specific neutralizing antibody and T cell responses in previously infected and infection-naive individuals. Also, we assessed the effect of an ancestral/Omicron BA.1 bivalent mRNA booster vaccination on these immune responses. 10 months after previous monovalent mRNA vaccinations, ancestral SARS-CoV-2 S1-specific T cell and anti-RBD IgG responses remained detectable in most individuals and a previous SARS-CoV-2 infection was associated with increased T cell responses. T cell responses, anti-RBD IgG, and Omicron BA.5 neutralization activity increased after receiving an ancestral/Omicron BA.1 bivalent booster mRNA vaccination. An Omicron BA.5 infection in addition to bivalent vaccination led to a higher ratio of Omicron BA.5 to ancestral strain neutralization activity compared to bivalent vaccination without a recent SARS-CoV-2 infection. In conclusion, SARS-CoV-2 T cell and antibody responses persist for up to 10 months after a monovalent booster mRNA vaccination. An ancestral/Omicron BA.1 bivalent booster mRNA vaccination increases these immune responses and also induces Omicron BA.5 cross-neutralization antibody activity. Finally, our data indicate that hybrid immunity is associated with improved preservation of T cell immunity.

Clinical and histopathological evidence suggest that the male reproductive system may be negatively impacted in patients with coronavirus disease (COVID-19). The objective of this study is to investigate the effect of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) on semen parameters by comparing semen analyses before and after COVID-19 diagnosis in the same patient. We retrospectively analyzed 342 semen analyses by reviewing medical records. The study included two groups of patients: (i) those who underwent two consecutive semen analyses within 6 months, one before (n=114) and one after (n=114) COVID-19 diagnosis, and (ii) a control group (n=114) that was age-matched and did not receive a diagnosis of COVID-19. The study results indicated a significant decrease in semen volume, total sperm count per ejaculate, progressive motile sperm count, total motile sperm count, and normal sperm morphology after SARS-CoV-2 infection in comparison to their respective values before the infection. Subgroup analyses showed that the duration of COVID-19 diagnosis (short-term vs long-term) did not impact the changes in semen parameters. However, fever during the COVID-19 process had a negative effect on semen parameters, particularly sperm concentration, unlike in patients without fever. In conclusion, our findings suggest that SARS-CoV-2 infection is associated with a decline in semen quality, which may potentially impact male fertility. Furthermore, it’s important to note that the negative effects on semen parameters may persist in the long-term. Our results also indicate that fever during active infection could be a significant risk factor that negatively affects spermatogenesis.

Purpose Although monoclonal antibodies specific to the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are known, information about the B-cell receptor (BCR) repertoire and its change in patients during COVID-19 disease progression is underreported. Methods We used immunoglobulin heavy chain (IGH) variable region (IGHV) spectratyping and next-generation sequencing of peripheral blood B-cell genomic DNA collected at multiple time points during disease evolution to study B-cell response to SARS-CoV-2 infection in 14 individuals with acute COVID-19. Results We found a broad distribution of responding B-cell clones. The IGH gene usage was not significantly skewed but frequencies of individual IGH genes changed repeatedly. We found predominant usage of unmutated and low mutation-loaded IGHV rearrangements characterizing naïve and extrafollicular B-cells among the majority of expanded peripheral B-cell clonal lineages at most tested time points in most patients. IGH rearrangement usage showed no apparent relation to anti-SARS-CoV-2 antibody titers. Some patients demonstrated mono/oligoclonal populations carrying highly mutated IGHV rearrangements indicating antigen experience at some of the time points tested, including even before anti-SARS-CoV-2 antibodies were detected. Conclusion We present evidence demonstrating that the B-cell response to SARS-CoV-2 is individual and includes different lineages of B-cells at various time points during COVID-19 progression.

The main proteases (M pro) are highly conserved cysteine-rich proteins that can be covalently modified by numerous natural and synthetic compounds. Herein, we constructed an integrative approach to efficiently discover covalent inhibitors of M pro from complex herbal matrices. This work begins with biological screening of sixty clinically used antiviral herbal medicines, among which Lonicera japonica (LJ) demonstrated the strongest anti-M pro effect (IC 50 = 37.82 μg/mL). Mass spectrometry-based chemical analysis and chemoproteomic profiling revealed that LJ extract contains at least 50 constituents, of which 22 exhibited the capability to covalently modify M pro. We subsequently verified the anti-M pro effects of these covalent binders. Gallic acid and quercetin were found to potently inhibit SARS-CoV-2 M pro in dose- and time- dependent manners, with the IC 50 values below 10 µM. The inactivation kinetics, binding affinity and binding mode of gallic acid and quercetin were further characterized by fluorescence resonance energy transfer, surface plasmon resonance, and covalent docking simulations. Overall, this study established a practical approach for efficiently discovering the covalent inhibitors of M pro from herbal medicines by integrating target-based high-throughput screening and mass spectrometry-based assays, which would greatly facilitate the discovery of key anti-viral constituents from medicinal plants.

Objectives: Circulating nucleocapsid (NCP) antigen of SARS-CoV-2 is increased in severely ill COVID-19 patients. However, clinical deterioration of COVID-19 often happens about one week after benign initial presentation. The role of NCP antigenemia as a biomarker in those cases remains unclear. We investigated NCP clearance kinetics in hospitalized patients as a risk assessment tool for predicting necessity of intensive care treatment of COVID-19 patients. Methods: Serum NCP was quantified using a commercial NCP-specific ELISA in hospitalized COVID-19 patients (n=63) during their hospital stay. Results were correlated to COVID-19 disease severity, inflammation parameters, antibody response and results of SARS-CoV-2 PCR from nasopharyngeal swabs.   Results: We demonstrate that NCP antigen levels in serum remained elevated in 45.6% of patients requiring treatment on intensive care units (ICU) after >8 days post positive SARS-CoV-2 PCR, compared to complete clearance in all non-ICU patients. This was in contrast to mucosal clearance of virus as measured by PCR. Antigen clearance was associated with higher IgG against S1 but not NCP.   Conclusions: Detection of NCP antigenemia after 8 days post COVID-19 diagnosis identifies patients who will require intensive care. Lack of NCP clearance after one week can thus help to assess the risk to develop severe COVID-19.

Postnatal cytomegalovirus (HCMV) infection is well characterized in preterm infants, where it can lead to severe symptomatic infection. We analyzed the rate and route of transmission of postnatal HCMV infections in full-term babies during the first year of life. A cohort of 120 HCMV seropositive mothers and their 122 newborns were tested after delivery for HCMV DNA shedding in different bodily fluids. Postnatal HCMV infection was defined as the detection of >2.5×10 2 HCMV-DNA copies/mL in infants’ saliva swabs. Maternal neutralizing antibody serum titer, HCMV specific T-cell response, and HCMV glycoprotein B (gB) IgG on breastmilk were analyzed. HCMV shedding was detected in 67 of 120 mothers (55.8%), and 20 of 122 infants (16.4%) developed HCMV infection within the first three months of life. Six additional infants were infected during the first year, for a postnatal infection rate of 21.3%. Viral shedding was more frequent in breastmilk than saliva, urine and vaginal secretions, and the mothers of infected infants showed higher levels of HCMV-DNA in milk. No association was found between the antibody levels in serum or milk and maternal viral shedding, whereas a slightly lower frequency of HCMV-specific CD4 + T-cells with long-term memory phenotype was observed in women with HCM-DNA-positive milk. About one out of five infants develop HCMV infection within the first year of life. Breastmilk appears the major route of transmission of the infection, maternal saliva have a minor role whereas the role of vaginal secretions is negligible.

The number of SARS-CoV-2 recombinants identified during the pandemic has increased since the era of Omicron variants, but XBB.1.5 (or Omicron 23A) is the first lineage comprised of hybrid genomes to predominate at the country and global scales. Very interestingly, the XBB.1.5 recombinant, like the Marseille-4B subvariant (B.1.160/20A.EU2) and the pandemic variant B.1.1.7 (20I/Alpha) previously, has its ORF8 gene inactivated by a stop codon. XBB.1.5 was generated through two successive main events: a recombination between SARS-CoV-2 of lineages BA.2.10.1.1 (BJ.1) and BA.2 75.3.1.1.1 (BM.1.1.1) that generated the XBB (22F) lineage; then ORF8 gene inactivation by a stop codon. We further identified that a stop codon was present at 89 (74%) codons of the ORF8 gene in ≥1 of 15,222,404 genomes available in GISAID, and at 15 codons (12%) in ≥1,000 genomes. Thus, it is very likely that stop codons in ORF8 gene contributed on at least 3 occasions and independently during the SARS-CoV-2 pandemic to the evolutionary success of a lineage that became transiently predominant, most recently XBB.1.5. Such association of gene loss with evolutionary success, which suits the recently described Mistigri rule, is an important biological phenomenon very unknown in virology while largely described in cellular organisms.

1. A survey on the recent COVID-19 epidemic in China was conducted. 2. The epidemic likely made over 85% of people in China sick in December 2022. 3. The epidemic likely ended in early January 2023. 4. The epidemic had the features of spreading more rapidly and widely and having a higher symptomatic rate than previously thought. 5. The above features could lead to many severe cases and deaths due to the shortages of medical resources. 6. The above features greatly enhanced the population immunity in China, which will accelerate the end of the COVID-19 pandemic and reduce the emergence of risky variants of SARS-CoV-2 in China.

SARS-CoV2 infection has caused an increase in mortality and morbidity but with vaccination, the disease severity has significantly reduced. But with emergence of various variants of concerns (VOCs), the vaccine breakthrough infection has also increased. Here, we studied circulating spike specific T follicular response (cTfh) in infection naïve vaccinees and convalescent vaccinees (individuals who got delta breakthrough infection after 2 doses of BBV152 vaccine) to understand their response as they are the most crucial cells that are involve in vaccine mediated protection by helping in B-cell maturation. Our results indicated that cTfh cells in both the groups recognized wild type and delta spike protein but memory response to wild type spike was superior in infection naïve than in convalescent group. The cytokine response particularly IL-21 from cTfh cells was also higher in infection naïve than convalescent vaccines indicating a dampened cTfh response in convalescent vaccines after breakthrough infection. Also, there was positive correlation between IL-21 from cTfh cells and neutralizing antibodies of infection naïve vaccinees. Multiple cytokine analysis also revealed higher inflammation in convalescent vaccinees. Our data indicated necessity of third booster dose may be individual specific depending in the steady state functional phenotype of immune cells.

Objectives In a COVID-19 sero-surveillance cohort study with predominantly healthy and vaccinated individuals, the objectives were (i) to investigate longitudinally the factors associated with the quantitative dynamics of anti-spike IgG antibody levels, (ii) to evaluate whether the antibody levels were associated with protection from SARS-CoV-2 infection and (iii) to assess whether the association was different in the pre-Omicron compared with the Omicron period. Methods The QuantiVac Euroimmun ELISA test was used to quantify anti-S1 IgG levels. The entire study period (16 months), the 11-month pre-Omicron period and the cross-sectional analysis prior to the Omicron surge included 3219, 2310 and 895 reactive serum samples from 949, 919 and 895 study participants, respectively. Mixed-effect linear, mixed-effect time-to-event and logistic regression models were used to achieve the objectives. Results Age and time since infection or vaccination were the only factors associated with a decline of anti-S1 IgG levels. Antibody levels were significantly associated with protection from SARS-CoV-2 infection, and the association was higher for the Omicron than for the Alpha and Delta variants. In a prediction model, it was estimated that >8000 BAU/mL anti-S1 IgG was required to reduce the risk of infection with Omicron variants by 20% to 30% for 90 days. Conclusions Anti-S1 IgG antibody levels are associated with protection from infection. The levels in the pre-Omicron periods were less significant than during the Omicron surge, which in turn required very high levels for protection in a statistical model.