Different scenarios explaining the emergence of novel variants of concern (VOC) of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have been reported, including their evolution in scarcely monitored populations, in animals as alternative hosts, or in immunocompromised individuals. Here we report SARS-CoV-2 immune escape mutations over a period of seven months in an immunocompromised patient with prolonged viral shedding. Signs of infection, viral shedding and mutation events are periodically analyzed using RT-PCR and next-generation sequencing based on naso-pharyngeal swabs, with the results complemented by immunological diagnostics to determine humoral and T cell immune responses. Throughout the infection course, 17 non-synonymous intra-host mutations are noted, with 15 (88.2%) having been previously described as prominent immune escape mutations (S:E484K, S:D950N, S:P681H, S:N501Y, S:del(9), N:S235F and S:H655Y) in VOCs. The high frequency of these non-synonymous mutations is consistent with multiple events of convergent evolution. Thus, our results suggest that specific mutations in the SARS-CoV-2 genome may represent positions with a fitness advantage, and may serve as targets in future vaccine and therapeutics development for COVID-19.

Seroprevalence rates for immunoglobulin G (IgG) antibodies to Tahyna virus (TAHV) and Inkoo virus (INKV) were determined in sera of 1630 blood donors from North, East, and South Tyrol by immunofluorescence assays (IFAs) and confirmatory serum neutralization tests (SNTs). Ten sera (0.6%) reacted positive by TAHV IFA, five of which (0.3%) were confirmed by SNT. Eleven sera (0.7%) reacted positive in the INKV IFA; only one thereof (0.06%) was verified by subsequent SNT. To identify the source of infections, mosquitoes were trapped at 18 sampling sites in the study area, resulting in the collection of 2571 adult mosquitoes: 1254 individuals of the genus Aedes (48.8% of total) including A. albopictus, 640 Culex (24.9%), 303 Coquillettidia (11.8%), 252 Ochlerotatus (9.8%), 49 Anopheles (1.9%), and 73 mosquitoes of the genus Culiseta (2.8%). The mosquitoes were pooled according to species, trapping site, and time, and were tested by RT-PCR for the presence of California serogroup orthobunyavirus nucleic acids. PCR amplification products were obtained in five of 195 pools (2.6%), and all were identified as TAHVs by subsequent sequencing. This represents the first evidence of TAHV circulation and human exposure in the Tyrols and in the alpine region in general. Interestingly, all TAHV sequences were identified in Culex pipiens/torrentium mosquitoes. Whether other California serogroup orthobunyaviruses such as INKV are also circulating in this area is subject of further investigations on larger numbers of mosquitoes.

This study shows that Babesia spp. are present in the Tyrols, which blood donors come into serologic contact with, and that we have to consider how to sustain blood product safety concerning this new challenge. Additionally, it is the first description of B. venatorum in the Tyrols, found in one Ixodes ricinus at the Italian border.