Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was first identified in Wuhan, China, in late 2019 and is the causative agent of the coronavirus disease 2019 (COVID-19) pandemic. Quantitative reverse-transcription polymerase chain reaction (qRT-PCR) represents the gold standard for diagnostic assays even if it cannot precisely quantify viral RNA copies. Thus, we decided to compare qRT-PCR with digital polymerase chain reaction (dPCR), which is able to give an accurate number of RNA copies that can be found in a specimen. However, the aforementioned methods are not capable to discriminate if the detected RNA is infectious or not. For this purpose, it is necessary to perform an endpoint titration on cell cultures, which is largely used in the research field and provides a tissue culture infecting dose per mL (TCID50/mL) value. Both research and diagnostics call for a model that allows the comparison between the results obtained employing different analytical methods. The aim of this study is to define a comparison among two qRT-PCR protocols (one with preliminary RNA extraction and purification and an extraction-free qRT-PCR), a dPCR and a titration on cell cultures. The resulting correlations yield a faithful estimation of the total number of RNA copies and of the infectious viral burden from a Ct value obtained with diagnostic routine tests. All these estimations take into consideration methodological errors linked to the qRT-PCR, dPCR and titration assays.

Adenoviral and mRNA vaccines encoding the viral spike (S) protein have been deployed globally to contain severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Older individuals are particularly vulnerable to severe infection, probably reflecting age-related changes in the immune system, which can also compromise vaccine efficacy. It is nonetheless unclear to what extent different vaccine platforms are impacted by immunosenescence. Here, we evaluated S protein-specific immune responses elicited by vaccination with two doses of BNT162b2 or ChAdOx1-S and subsequently boosted with a single dose of BNT162b2 or mRNA-1273, comparing age-stratified participants with no evidence of previous infection with SARS-CoV-2. We found that aging profoundly compromised S protein-specific IgG titers and further limited S protein-specific CD4⁺ and CD8⁺ T cell immunity as a probable function of progressive erosion of the naive lymphocyte pool in individuals vaccinated initially with BNT162b2. Our results demonstrate that primary vaccination with ChAdOx1-S and subsequent boosting with BNT162b2 or mRNA-1273 promotes sustained immunological memory in older adults and potentially confers optimal protection against coronavirus disease 2019.

Orf virus (ORFV) belongs to the genus Parapoxvirus (Poxviridae family). It is the causative agent of contagious ecthyma (CE) that is an economically detrimental disease affecting small ruminants globally. Contagious ecthyma outbreaks are usually reported in intensive breeding of sheep and goats but they have also been reported in wildlife species. Notably, ORFV can infect humans, leading to a zoonotic disease. This study aims to elucidate the global evolutionary history of ORFV genomes in sheep and goats, including the first genomes from Central America in the analyses. In comparison to the last study on ORFV whole genomes, the database now includes 11 more sheep and goat genomes, representing an increase of 42%. The analysis of such a broader database made it possible to obtain a fine molecular dating of the coalescent time for ORFV S and G genomes, further highlighting the genetic structuring between sheep and goat genomes and corroborating their emergence in the latter half of 20th century.

The incidence of total joint arthroplasty is increasing over time since the last decade and expected to be more than 4 million by 2030. As a consequence, the detection of infections associated with surgical interventions is increasing and prosthetic joint infections are representing both a clinically and economically challenging problem. Many pathogens, from bacteria to fungi, elicit the immune system response and produce a polymeric matrix, the biofilm, that serves as their protection. In the last years, the implementation of diagnostic methodologies reduced the error rate and the turn-around time: polymerase chain reaction, targeted or broad-spectrum, and next-generation sequencing have been introduced and they represent a robust approach nowadays that frees laboratories from the unique approach based on culture-based techniques.

Literature offers plenty of cases of immunocompromised patients, who develop chronic and severe SARS-CoV-2 infections. The aim of this study is to provide further insight into SARS-CoV-2 evolutionary dynamic taking into exam a subject suffering from follicular lymphoma, who developed a persistent infection for over 7 months. Eight nasopharyngeal swabs were obtained, and were analyses by qRT-PCR for diagnostic purposes. All of them were considered eligible (Ct < 30) for NGS sequencing. Sequence analysis showed that all sequences matched the B.1.617.2 AY.122 lineage, but they differed by few mutations identifying three genetically similar subpopulations, which evolved during the course of infection, demonstrating that prolonged replication is paralleled with intra-host virus evolution. These evidences support the hypothesis that SARS-CoV-2 adaptive capacities are able to shape a heterogeneous viral population in the context of immunocompromised patients. Spill-over of viral variants with enhanced transmissibility or immune escape capacities from these subjects is plausible.

The recent emergence of a number of new SARS-CoV-2 variants resulting from recombination between two distinct parental lineages or sub-lineages within the same lineage has sparked the debate regarding potential enhanced viral infectivity and immune escape. Among these, XBB, recombinant of BA.2.10 and BA.2.75, has caused major concern in some countries due to its rapid increase in prevalence. In this study, we tested XBB escape capacity from mRNA-vaccine-induced (BNT162b2) neutralising antibodies compared to B.1 ancestral lineage and another co-circulating variant (B.1.1.529 BA.5) by analysing sera collected 30 days after the second dose in 92 healthcare workers. Our data highlighted an enhanced and statistically significant immune escape ability of the XBB recombinant. Although these are preliminary results, this study highlights the importance of immune escape monitoring of new and forthcoming variants and of the reformulation of existing vaccines.