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Vancomycin-resistant enterococci (VRE) infections and outbreaks are still infrequent in Spain. A six-month outbreak, which took place in a haematology ward, its control and management are described in this study. A total of 22 patients were colonised and two bloodstream infections occurred during this period. Even though there were two waves of new colonised patients, a multidisciplinary approach, quick interventions and enhanced infection control policies were required in order to control this outbreak. [ABSTRACT FROM AUTHOR]

• Assessment of the use of sample pooling in retrospective samples. • A pool size of five samples had the highest sensitivity. • Pooling prospective samples identified positive cases with cycle thresholds ranging from 16.7 to 39.4. Limited testing capacity has characterized the ongoing coronavirus disease 2019 (COVID-19) pandemic in Spain, hampering timely control of outbreaks and opportunities to reduce the escalation of community transmission. This study investigated the potential to use sample pooling, followed by one-step retrotranscription and real-time quantitative reverse transcription polymerase chain reaction (RT-qPCR) to increase testing capacity for severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). Various pool sizes (five, 10 and 15 samples) were evaluated prior to RNA extraction followed by standard RT-qPCR for the diagnosis of COVID-19. The pool size achieving reproducible results with individual sample testing was subsequently used to assess nasopharyngeal samples in a tertiary hospital in August 2020. A pool size of five samples had higher sensitivity compared with pool sizes of 10 and 15 samples, showing a mean cycle threshold (Ct) shift of 3.5 [standard deviation (SD) 2.2] between the pooled test and positive samples in the pool. Next, a pool size of five was used to test a total of 895 pools (4475 prospective samples) using two different RT-qPCR kits. The Real Accurate Quadruplex corona-plus PCR Kit (PathoFinder) reported the lowest mean Ct shift [2.2 (SD 2.4)] between the pool and individual samples. This strategy enables detection of individual positive samples in positive pools with Ct of 16.7–39.4. Grouping samples into pools of five for RT-qPCR resulted in an increase in SARS-CoV-2 testing capacity with minimal loss of sensitivity compared with testing each sample individually. [ABSTRACT FROM AUTHOR]

Worldwide outbreaks of enterovirus D68 (EV-D68) in 2014 and 2016 have caused serious respiratory and neurological disease. We collected samples from several European countries during the 2018 outbreak and determined 53 near full-length genome ('whole genome') sequences. These sequences were combined with 718 whole genome and 1,987 VP1-gene publicly available sequences. In 2018, circulating strains clustered into multiple subgroups in the B3 and A2 subclades, with different phylogenetic origins. Clusters in subclade B3 emerged from strains circulating primarily in the US and Europe in 2016, though some had deeper roots linking to Asian strains, while clusters in A2 traced back to strains detected in East Asia in 2015-2016. In 2018, all sequences from the USA formed a distinct subgroup, containing only three non-US samples. Alongside the varied origins of seasonal strains, we found that diversification of these variants begins up to 18 months prior to the first diagnostic detection during a EV-D68 season. EV-D68 displays strong signs of continuous antigenic evolution and all 2018 A2 strains had novel patterns in the putative neutralizing epitopes in the BC- and DE-loops. The pattern in the BC-loop of the USA B3 subgroup had not been detected on that continent before. Patients with EV-D68 in subclade A2 were significantly older than patients with a B3 subclade virus. In contrast to other subclades, the age distribution of A2 is distinctly bimodal and was found primarily among children and in the elderly. We hypothesize that EV-D68's rapid evolution of surface proteins, extensive diversity, and high rate of geographic mixing could be explained by substantial reinfection of adults. Better understanding of evolution and immunity across diverse viral pathogens, including EV-D68 and SARS-CoV-2, is critical to pandemic preparedness in the future. Author summary: Enterovirus D68 (EV-D68) has caused punctuated, global outbreaks of respiratory illness and neurological disease, including being implicated as the cause of acute flaccid myelitis (AFM). Serology studies and surveillance data suggests almost everyone is infected during early childhood. The majority of sequences collected are from young children, while adults retain high antibody titers against strains that circulated when they were young. However, little is known about how outbreaks are connected and how the virus evolves and spreads around the globe. Despite EV-D68's apparent reliance on young, naive hosts, EV-D68 antibody binding sites are reportedly evolving under antigenic pressure, and EV-D68 seems to spread rapidly during outbreaks. In this multi-center European collaboration, we confirm that subclade specific age differences are present in those infected. Further, we were able to quantify between- and within-country migration and the 'hidden' diversification that indicates unsampled circulation between outbreaks. We conclude that the evolution of EV-D68 may be driven by substantial re-infection of adults, explaining the rapid geographic mixing and continuous antigenic evolution. The presence of largely unsampled circulation prior to outbreaks suggests there are gaps in current surveillance practices which could be addressed by expanding genetic surveillance. [ABSTRACT FROM AUTHOR]