Journal of Medical Virology

BackgroundCOVID-19 is a disease with global public health emergency that have shook the world since its first detection in China in December, 2019. Severe acute respiratory syndrome Coronavirus 2 (SARS-CoV-2) is the pathogen responsible behind this pandemic. The lethality of different viral strains is found to vary in different geographical locations but the molecular mechanism is yet to be known. MethodsAvailable data of whole genome sequencing of different viral strains published by different countries were retrieved and then analysed using Multiple Sequence Alignment and Pair-wise Sequence Alignment leading to Phylogenetic tree construction. Each location and the corresponding genetic variations were screened in depth. Then the variations are analysed at protein level giving special emphasis on Non Synonymous amino acid substitutions. The fatality rates in different countries were matched against the mutation number, rarity of the nucleotide alterations and functional impact of the Non Synonymous changes at protein level, separately and in combination. FindingsAll the viral strains have been found to evolve from the viral strain of Taiwan (MT192759) which is 100% identical with the ancestor SARS-CoV-2 sequences of Wuhan (NC 045512.2; submitted on 5th Jan, 2020). Transition from C to T (C>T) is the most frequent mutation in this viral genome and mutations A>T, G>A, T>A are the rarest ones, found in countries with maximum fatality rate i.e Italy, Spain and Sweden. 20 Non Synonymous mutations are located in viral genome spanning Orf1ab polyprotein, Surface glycoprotein, Nucleocapsid protein etc. The functional effect on the structure and function of the protein can favourably or unfavourably interact with the host body. InterpretationThe fatality outcome depends on three important factors (a) number of mutation (b) rarity of the allelic variation and (c) functional consequence of the mutation at protein level. The molecular divergence, evolved from the ancestral strain (S) lead to extremely lethal (E), lethal(L) and non lethal (N) strains with the involvement of an Intermediate strain(I).

In December 2019, a cluster of acute respiratory illness, now known as SARS-CoV-2 pneumonia, occurred in Wuhan, China. World Health Organization (WHO) declared the rapidly spreading coronavirus outbreak a pandemic on March 11, 2020, acknowledging what has seemed clear for some time -- the virus will likely spread to all countries on the globe. As of February 11, 2020, the Chinese Center for Disease Control and Prevention (China CDC) has officially reported that there are 2.0% (889) asymptomatic cases, 2.3% (1,023) death cases, and 80.9% mild cases among 44,672 confirmed cases. 51.4% (22,981) were male and 48.6% (21,691) were female. Lymphopenia, in particular T lymphopenia, was common among patients with SARS-COV-2 in the observation. A notable drop in CD4 and CD8 lymphocyte counts occurred early in the course of the syndrome and was associated with adverse outcomes. The appearing a phenomenon of lymphocyte depletion (PLD) suggested severe adverse outcomes. The outcome observed: 60% had discharged and 20% had die.

SARS-CoV-2 variants, along with vaccination, mark the second year of the pandemic. The spike region is a focal point in COVID-19 pathogenesis, with different amino acid changes potentially modulating vaccine response and some being part of variant signatures. NGS is the standard tool to sequence the virus but limitations of different sources hinders expansion of genomic surveillance in many places. To improve surveillance capability we developed a Sanger based sequencing protocol to obtain coverage of most (>95%) spike gene. Eleven nasopharyngeal swabs collections had RNA extracted for real time PCR diagnosis and leftover RNA had up to 3785 bp sequenced at an ABI3500 using dye termination chemistry of nested PCR products of two reactions of one-step RT-PCR. P1 amino acid mutations signatures were present in 18% (2/11), with 82% (9/11) with three or more additional amino acid changes (GISAID CoVsurver list). Most sequences (86%, 6/7) from 2021 have the E484K, whereas the mutation was not present in samples collected in 2020 (0/4, p=0.015).The swiftness that favorable mutations to the virus may prevail and their potential impact in vaccines and other current interventions need broader surveillance and more public health attention.

COVID-19 is a viral respiratory disease caused by SARS-CoV-2 infection. Global efforts of genomic surveillance of the virus give chance to track the spread of the pandemic. Global emergence of some viral mutations called attention and various studies have been suggested about increased infectivity of the virus. Herein, we sequenced viral genomes isolated from 184 patients in Istanbul and analyzed clinical metadata for the investigation of any viral mutation which affects the disease course of the host. We did not detect any viral mutations affecting the disease outcome in our cohort. Besides, we observed intra-host mutations in 76% of the isolates. Insertion/deletion and stop-gain mutations are also significantly less common among intra-host variants compared to consensus viral genome mutations. Longitudinal genomic surveillance is essential for timely detection of any lineages that might affect clinical outcome, the performance of diagnostic assays, or even the immunological escape of the virus.

Spike (S) protein is a key protein in coronaviruses life cycle. SARS-CoV-2 Omicron BA.1 variant of concern (VoC) presents an exceptionally high number of 30 substitutions, 6 deletions and 3 insertions in the S protein. Recent works revealed major changes in the SARS-CoV-2 Omicron biological properties compared to earlier variants of concern (VoCs). Here, these major changes could be explained, at least in part, by the mutations N764K and/or N856K in S2 subunit. These mutations were not previously detected in other VoCs. N764K and N856K generate two potential cleavage sites for SKI-1/S1P serine protease, known to cleave viral envelope glycoproteins. The new sites where SKI-1/S1P could cleave S protein might impede the exposition of the internal fusion peptide for membrane fusion and syncytia formation. Based on the human protein atlas, SKI-1/S1P protease is not found in lung tissues (alveolar cells type I/II and endothelial cells), but present in bronchus and nasopharynx. This may explain why Omicron has change of tissue tropism. Viruses have evolved to use several host proteases for cleavage/activation of envelope glycoproteins. Mutations that allow viruses to change of protease may have a strong impact in host range, cell and tissue tropism, and pathogenesis.

Little is known about serological responses to MVA-BN (JYNNEOS) against mpox in elderly individuals with or without HIV. In this study, MVA-BN induced sustained IgG levels regardless of HIV status even up to one year. Birth before 1973 correlated with higher IgG. MVA-BN unvaccinated individuals with HIV had lower IgG than vaccinated.

New SARS-CoV-2 variants are continually emerging and have the potential to present challenges to public health responses worldwide. This report discusses the early detection of the lineage JN.2.5 (alias of B.1.1.529.2.86.1.2.5) in Rio de Janeiro, Brazil. The variant was identified in early 2024 through viral whole-genome sequencing and phylogenetic analysis, revealing that circulating strains present genetic similarities to sequences reported in Canada, Poland, Belgium, and England. While the circulation of diverse Omicron sub lineages has significantly impacted COVID-19 epidemiology in Brazil, the consequences of the circulation of the novel variant are uncertain. Considering its immunological escape capabilities and epidemiological behavior observed elsewhere, close monitoring of this variant epidemiology is recommended.

The rapid spread of the SARS-CoV-2 Variant of Concern (VOC) Gamma during late 2020 and early 2021 in Brazilian settings with high seroprevalence raised some concern about the potential role of reinfections in driving the epidemic. Very few cases of reinfection associated with the VOC Gamma, however, have been reported. Here we describe 25 cases of SARS-CoV-2 reinfection confirmed by real-time RT-PCR twice within months apart in Brazil. SARS-CoV-2 genomic analysis confirmed that individuals were primo-infected between March and December 2020 with distinct viral lineages, including B.1.1, B.1.1.28, B.1.1.33, B.1.195 and P.2, and then reinfected with the VOC Gamma between 3 to 12 months after primo-infection. The overall mean cycle threshold (Ct) value of the first (25.7) and second (24.5) episodes were roughly similar for the whole group and 14 individuals displayed mean Ct values < 25.0 at reinfection. Sera of 14 patients tested by plaque reduction neutralization test after reinfection displayed detectable neutralizing antibodies against Gamma and other SARS-CoV-2 variants (B.1.33, B.1.1.28 and Delta). All individuals have milder or no symptoms after reinfection and none required hospitalization. The present study demonstrates that the VOC Gamma was associated with reinfections during the second Brazilian epidemic wave in 2021 and raised concern about the potential infectiousness of reinfected subjects. Although individuals here analyzed failed to mount a long-term sterilizing immunity, they developed a high anti-Gamma neutralizing antibody response after reinfection that may provide some protection against severe disease.

Numerous factors have been identified to influence susceptibility to SARS-CoV-2 infection and disease severity. Cancer patients are more prone to clinically evolve to more severe COVID-19 conditions, but the determinants of such a more severe outcome remain largely unknown. We have determined the full-length SARS-CoV-2 genomic sequences of cancer patients and healthcare workers (HCW; non-cancer controls) by deep sequencing and investigated the within-host viral quasispecies of each infection, quantifying intrahost genetic diversity. Naso- and oropharyngeal SARS-CoV-2+ swabs from 57 cancer patients and 14 healthcare workers (HCW) from the Brazilian Cancer Institute were collected in April-May 2020. Complete genome amplification using ARTIC network V3 multiplex primers was performed followed by next-generation sequencing. Assemblies were conducted in Geneious R11, where consensus sequences were extracted and intrahost single nucleotide variants (iSNVs) were identified. Maximum likelihood phylogenetic analysis was performed using PhyMLv.3.0 and lineages were classified using Pangolin and CoV-GLUE. Phylogenetic analysis showed that all but one strain belonged to clade B1.1. Four genetically linked mutations known as the globally dominant SARS-CoV-2 haplotype (C241T, C3037T, C14408T and A23403G) were found in the majority of consensus sequences. SNV signatures of previously characterized Brazilian genomes were also observed in most samples. Another 85 SNVs were found at a lower frequency (1.4-19.7%). Cancer patients displayed a significantly higher intrahost viral genetic diversity compared to HCW (p = 0.009). Intrahost genetic diversity in cancer patients was independent of SARS-CoV-2 Ct values, and was not associated with disease severity, use of corticosteroids, or use of antivirals, characteristics that could influence viral diversity. Such a feature may explain, at least in part, the more adverse outcomes to which cancer/COVID-19 patients experience. Author SummaryCancer patients are more prone to clinically evolve to more severe COVID-19 conditions, but the determinants of such a more severe outcome remain largely unknown. In this study, phylogenetic and variation analysis of SARS-CoV-2 genomes from cancer patients and non-cancer healthcare workers at the Brazilian National Cancer Institute were characterized by deep sequencing. Viral genomes showed signatures characteristic of Brazilian viruses, consistent with the hypothesis of local, community transmission rather than virus importation from abroad. Despite most genomes in patients and healthcare workers belonging to the same lineage, intrahost variability was higher in cancer patients when compared to non-cancer counterparts. The intrahost genomic diversity analysis presented in our study highlights the relaxed evolution of SARS-CoV-2 in a vulnerable population of cancer patients. The high number of minor variations can result in the selection of immune escape variants, resistance to potential drugs, and/or increased pathogenicity. The impact of this higher intrahost variability over time warrants further investigation.

Here we report the clinical features of 25 discharged patients with COVID-19 recovery. Our analysis indicated that there was a significant inverse correlation existed between serum D-Dimer level and the duration of antiviral treatment, while lymphocyte concentration significantly positively correlated with the duration of virus reversal.

Cell-mediated immunity (CMI), which includes T-cells (both T helper and cytotoxic), is critical for effective antiviral defenses against coronavirus disease-2019 (COVID-19). To better understand the immunological characteristics of CD markers on T-cells in post-COVID-19 patients, we investigated the expression of differential CD markers in the patient groups in this study. Flow cytometry was used to quantify total lymphocyte count and assess the levels of expression of CD markers in the samples. The percentage of Lymphocytes decreased significantly in the post-SARS-COV-2 patients in comparison to normal subjects, which is usually happening in any viral infection. In contrast to that, expression of CD8 was increased in the patient group having long SARS-COV-2 infection with comorbid complications with respect to the normal individuals and long SARS-COV-2 infection without comorbid complications. This data revealed that the cellular immunological responses corroborated with an earlier report of COVID-19 infection were mediated by CD8 upregulation and cytotoxic T lymphocyte hyperactivation.

In this work, 28 sequences with 57,570 sites of the B.1.1 variant of SARS-CoV-2, from Brazilian states, were used. All sequences (publicly available on the National Center for Biotechnology Information platform (NCBI)) were aligned with Mega X software and all gaps, ambiguous sites and lost data were extracted, resulting in a region in a segment with 8,799 polymorphic (15.2% of the total) that were analyzed for their molecular diversity, FST, demographic and spatial expansion. Phylogenetic relationships of ancestry revealed the absence of genetically distinct subgroups, which was corroborated by the low value of FST found (15.38%). The low degree of polymorphism found among these samples, corroborated by the almost non-existent genetic distance, helped or established the absence of a genetic structuring pattern, demonstrating a satisfactory pattern of response to vaccines, since all the sequences analyzed were part of the Brazilian strains of variant B.1.1 of SARS-CoV-2 used in vaccines.

IntroductionThe objective of this study is to analyse the specific immune response against SARS-CoV-2 in those affected by Long Covid (LC), attributable to T cells (cell-mediated immunity) and to carry out a parallel analysis of the humoral response and lymphocyte typing. MethodologyDescriptive cross-sectional study of 74 patients with LC for at least 4 months since diagnosis. The collected data were: information on the COVID-19 episode and the persistent symptoms, medical history and a specific cell-mediated immunity to SARS-CoV-2 through flow cytometry, assessing the release of interferon-gamma (IFN-{gamma}) by T4 lymphocytes, T8 lymphocytes and NK cells. Descriptive and comparative analyses were carried out. ResultsPatients with LC had negative serology for Covid-19 in 89% of cases but 96% showed specific cellular immunity to SARS-CoV-2 an average of 9.5 months after infection: 89% of this response corresponded to T8 lymphocytes, 58% to NK cells, and 51% to T4 lymphocyte (20% negligibly positive). Most of them had altered immune cell typing and we found that T4 lymphocyte counts were low in 34% of cases and NK cell high in 64%. Macrophage populations were detected in the peripheral blood of 7% of them. Patients displayed a higher percentage of illnesses related to &[Prime]abnormal&[Prime] immune responses, either preceding SARS-CoV-2 infection (43%) or following it in 23% of cases. ConclusionThe immune system appears to have an important involvement in the development of LC and viral persistence could be the cause or consequence of it. Further analysis with a control group should be performed.

Paediatric immunisation had been relevant in reducing the widespread of Hepatitis B virus, as an outcome of the induction of hepatitis B surface antigen specific-IgG antibodies (anti-HBs). Studies revealed alteration effects of memory B cells during antiretroviral therapy (ART). We aimed at assessing anti-HBs response profile with respect to the most prominently used ART regimens in children. a cross-sectional study was conducted in 116 participants made up of 72 HIV-exposed and infected children, subdivided into 20 antiretroviral-naive on one hand and on another hand 52 ARV treated children made up of regimen subgroups, including 8 ABC-3TC-EFV/NVP (ART-R1), 19 ABC-3TC-LPV/r (ART-R2), 21 AZT-3TC-NVP (ART-R3) and 4 AZT-3TC-LPV/r (ART-R4), and 44 HIV-uninfected and unexposed (HUx or control group) children. Participants included in this study were regularly vaccinated children aged between 4 months and 5 years old, born to HIV-infected mothers. An optimized and adapted home-made ELISA and BioELISA(R) Biohit kit were used to measure specific IgM, IgG and IgG subclasses to HBs in children. As result, this study showed that the rates of vaccine protective response in children treated with ART under regimens R1, R2, R3 and R4 were 25%, 38%, 51% and 75%, respectively. These protective response rates were significantly lower (p<0.0001) in children under R1, R2 and R3 than the control group (92%). When comparing anti-HBs specific IgM and IgG response medians; IgM response levels were similar in both control and ARV treated children, whereas R1 (p=0.0045), R2 (p=0.0016), and R4 (p<0.0001) showed significantly lower IgG level compared to the control group. Anti-HBs IgG subclass profile pattern in the control was IgG3{approx}IgG1{approx}IgG4>IgG2. However, IgG3{approx}IgG1{approx}IgG4>IgG4 profile pattern was estimated for children submitted to R1, R2 and R4, and the profile pattern of IgG3>IgG1{approx}IgG4{approx}IgG2 in those treated with R3 which also showed the most prominent anti-HBs IgG response mean rank level.

The spread of the corona virus around the world has spurred travel restrictions and community lockdowns to manage the transmission of infection. In the Philippines, with a large population of overseas Filipino contract workers (OFWs), as well as foreign workers in the local online gaming industry and visitors from nearby countries, the first reported cases were from a Chinese couple visiting the country in mid-January 2020. Three months on, by mid-March, the COVID-19 cases in the Philippines had reached its first 100, before it exploded to the present 178,022 cases (as of August 20, 2020). Here, we report a genomic survey of six (6) whole genomes of the SARS-CoV-2 virus collected from COVID-19 patients seen at the Philippine General Hospital, the major referral hospital for COVID-19 cases in Metro Manila at about the time the Philippines had over a hundred cases. Analysis of commonly observed variants did not reveal a clear pattern of the virus evolving towards a more infectious and severe strain. When combined with other available viral sequences from the Philippines and from GISAID, phylogenomic analysis reveal that the sequenced Philippine isolates can be classified into three primary groups based on collection dates and possible infection sources: (1) January samples collected in the early phases of the pandemic that are closely associated with isolates from Wuhan, China; (2) March samples that are mainly linked to the M/V Diamond Princess Cruise Ship outbreak; and (3) June samples that clustered with European isolates, one of which already harbor the globally prevalent D614G mutation which initially circulated in Europe. The presence of community-acquired viral transmission amidst compulsory and strict quarantine protocols, particularly for repatriated Filipino workers, highlights the need for a refinement of the quarantine, testing, and tracing strategies currently being implemented to adapt to the current pandemic situation.

In 2021, 273 Rocky Mountain Spotted Fever cases have been reported nationwide. In Chihuahua City, fourteen samples were obtained from children suspected of rickettsial infection. The analysis of samples collected from January to December 2021 showed a prevalence of 28.5%, 43% and 28.5% for Rickettsia rickettsii, Ehrlichia canis, and both pathogens in coinfection, respectively. The analysis of clinical hematological and biochemistry analytes showed alterations such as 100% of the children coursed with elevated liver enzymes and coagulation times, 64% showed leukocytosis due to neutrophilia, 55% of them had thrombocytopenia, lymphopenia and hypoalbuminemia, and 45% showed normocytic normochromic anemia. Statistically significant differences were obtained in the chemokines IL-8, RANTES, CXCL9/MIG, and CXCL10/IP-10 across the coinfected and control groups; the differences in IP-10 were significant for patients infected by R. rickettsii compared to the control group. Also, significant differences were observed for IL-1{beta}, IL-6, IL-17, IFN{gamma}, and TNF among the R. rickettsii positive group compared to the control group; on the other hand; the coinfected group exhibited modified levels of IL-6, IL-8, and IL-10 compared with the control group. Finally, significant differences were obtained for CD8 + T lymphocytes subpopulations between positive individuals for R. rickettsii and E. canis.

The authors have withdrawn their manuscript, which means that the paper will no longer be published. This decision was made because the authors used new approaches in analyzing a different sample size, which could result in different findings when compared to the previous version. The authors believe that the new data could be controversial, and therefore, they do not wish for their work to be cited as a reference for any project. Therefore, the authors do not wish this work to be cited as reference for the project. If you have any questions, please contact the corresponding author.

Concomitant infection of multiple SARS-CoV-2 variants has become an increasing concern, as this scenario increases the likelihood of recombinant variants. Detecting co-infection of SARS-CoV-2 variants is difficult to detect by whole genome sequencing approaches, but genotyping methods facilitate detection. We describe 2 cases of Delta/Omicron and 2 cases of Omicron sublineage BA.1/ BA.2 co-infection as detected by a multiplex genotyping fragment analysis method. Findings were confirmed by whole genome sequencing. Review of the patient characteristics revealed co-morbidities and conditions which weaken the immune system and may make them more susceptible to harboring SARS-CoV-2 variant co-infections.

While establishing a regional SARS-Cov-2 variant surveillance by genome sequencing, we have identified three infected individuals in a clinical setting (two long-term hospitalized patients and a nurse) that shared the spike N501Y mutation within a genotype background distinct from the current viral variants of concern. We suggest that the adaptive N501Y mutation, known to increase SARS-CoV-2 transmissibility, arose by convergent evolution around December in Mainz, Germany. Hospitalized patients with a compromised immune system may be a potential source of novel viral variants, which calls for monitoring viral evolution by genome sequencing in clinical settings.

Analysing 92,598 variant screening tests performed on SARS-CoV-2 positive samples collected in France between 1 July and 31 August 2021 shows an increase of Kappa-like infections. Full genome sequencing reveals that these correspond to Delta variants bearing the S:E484Q mutation. Most of these sequences belong to a phylogenetic cluster and also bear the S:T95I mutation. Further monitoring is needed to determine if this trend is driven by undocumented superspreading events or an early signal of future viral evolutionary dynamics.