Potential drivers of human tick-borne encephalitis in the Örebro region of Sweden, 2010-2021.

Incidence of tick-borne encephalitis (TBE) has increased during the last years in Scandinavia, but the underlying mechanism is not understood. TBE human case data reported between 2010 and 2021 were aggregated into postal codes within Örebro County, south-central Sweden, along with tick abundance and environmental data to analyse spatial patterns and identify drivers of TBE. We identified a substantial and continuing increase of TBE incidence in Örebro County during the study period. Spatial cluster analyses showed significant hotspots (higher number of cases than expected) in the southern and northern parts of Örebro County, whereas a cold spot (lower number of cases than expected) was found in the central part comprising Örebro municipality. Generalised linear models showed that the risk of acquiring TBE increased by 12.5% and 72.3% for every percent increase in relative humidity and proportion of wetland forest, respectively, whereas the risk decreased by 52.8% for every degree Celsius increase in annual temperature range. However, models had relatively low goodness of fit (R2 < 0.27). Results suggest that TBE in Örebro County is spatially clustered, however variables used in this study, i.e., climatic variables, forest cover, water, tick abundance, sheep as indicator species, alone do not explain this pattern.

Dairy milk from cow and goat as a sentinel for tick-borne encephalitis virus surveillance.

Tick-borne encephalitis (TBE) is one of the most severe human tick-borne diseases in Europe. It is caused by the tick-borne encephalitis virus (TBEV), which is transmitted to humans mainly via bites of Ixodes ricinus or I. persulcatus ticks. The geographical distribution and abundance of I. ricinus is expanding in Sweden as has the number of reported human TBE cases. In addition to tick bites, alimentary TBEV infection has also been reported after consumption of unpasteurized dairy products. So far, no alimentary TBEV infection has been reported in Sweden, but knowledge about its prevalence in Swedish ruminants is scarce. In the present study, a total of 122 bulk tank milk samples and 304 individual milk samples (including 8 colostrum samples) were collected from dairy farms (n = 102) in Sweden. All samples were analysed for the presence of TBEV antibodies by ELISA test and immunoblotting. Participating farmers received a questionnaire about milk production, pasteurization, tick prophylaxis used on animals, tick-borne diseases, and TBE vaccination status. We detected specific anti-TBEV antibodies, i.e., either positive (>126 Vienna Units per ml, VIEU/ml) or borderline (63-126 VIEU/ml) in bulk tank milk from 20 of the 102 farms. Individual milk samples (including colostrum samples) from these 20 farms were therefore collected for further analysis. Our results revealed important information for detection of emerging TBE risk areas. Factors such as consumption of unpasteurized milk, limited use of tick prophylaxis on animals and a moderate coverage of human TBE vaccination, may be risk factors for alimentary TBEV infection in Sweden.

Revealing new tick-borne encephalitis virus foci by screening antibodies in sheep milk.

BACKGROUND: Tick distribution in Sweden has increased in recent years, with the prevalence of ticks predicted to spread towards the northern parts of the country, thus increasing the risk of tick-borne zoonoses in new regions. Tick-borne encephalitis (TBE) is the most significant viral tick-borne zoonotic disease in Europe. The disease is caused by TBE virus (TBEV) infection which often leads to severe encephalitis and myelitis in humans. TBEV is usually transmitted to humans via tick bites; however, the virus can also be excreted in the milk of goats, sheep and cattle and infection may then occur via consumption of unpasteurised dairy products. Virus prevalence in questing ticks is an unreliable indicator of TBE infection risk as viral RNA is rarely detected even in large sample sizes collected at TBE-endemic areas. Hence, there is a need for robust surveillance techniques to identify emerging TBEV risk areas at early stages. METHODS: Milk and colostrum samples were collected from sheep and goats in Örebro County, Sweden. The milk samples were analysed for the presence of TBEV antibodies by ELISA and validated by western blot in which milk samples were used to detect over-expressed TBEV E-protein in crude cell extracts. Neutralising titers were determined by focus reduction neutralisation test (FRNT). The stability of TBEV in milk and colostrum was studied at different temperatures. RESULTS: In this study we have developed a novel strategy to identify new TBEV foci. By monitoring TBEV antibodies in milk, we have identified three previously unknown foci in Örebro County which also overlap with areas of TBE infection reported during 2009-2018. In addition, our data indicates that keeping unpasteurised milk at 4 °C will preserve the infectivity of TBEV for several days. CONCLUSIONS: Altogether, we report a non-invasive surveillance technique for revealing risk areas for TBE in Sweden, by detecting TBEV antibodies in sheep milk. This approach is robust and reliable and can accordingly be used to map TBEV "hotspots". TBEV infectivity in refrigerated milk was preserved, emphasising the importance of pasteurisation (i.e. 72 °C for 15 s) prior to consumption.

Multiple direct interactions of TBP with the MYC oncoprotein.

Transcription factor c-MYC is a potent oncoprotein; however, the mechanism of transcriptional regulation via MYC-protein interactions remains poorly understood. The TATA-binding protein (TBP) is an essential component of the transcription initiation complex TFIID and is required for gene expression. We identify two discrete regions mediating MYC-TBP interactions using structural, biochemical and cellular approaches. A 2.4 -Å resolution crystal structure reveals that human MYC amino acids 98-111 interact with TBP in the presence of the amino-terminal domain 1 of TBP-associated factor 1 (TAF1TAND1). Using biochemical approaches, we have shown that MYC amino acids 115-124 also interact with TBP independently of TAF1TAND1. Modeling reveals that this region of MYC resembles a TBP anchor motif found in factors that regulate TBP promoter loading. Site-specific MYC mutants that abrogate MYC-TBP interaction compromise MYC activity. We propose that MYC-TBP interactions propagate transcription by modulating the energetic landscape of transcription initiation complex assembly.

E3 ubiquitin-protein ligase TRIM21-mediated lysine capture by UBE2E1 reveals substrate-targeting mode of a ubiquitin-conjugating E2.

The E3 ubiquitin-protein ligase TRIM21, of the RING-containing tripartite motif (TRIM) protein family, is a major autoantigen in autoimmune diseases and a modulator of innate immune signaling. Together with ubiquitin-conjugating enzyme E2 E1 (UBE2E1), TRIM21 acts both as an E3 ligase and as a substrate in autoubiquitination. We here report a 2.82-Å crystal structure of the human TRIM21 RING domain in complex with the human E2-conjugating UBE2E1 enzyme, in which a ubiquitin-targeted TRIM21 substrate lysine was captured in the UBE2E1 active site. The structure revealed that the direction of lysine entry is similar to that described for human proliferating cell nuclear antigen (PCNA), a small ubiquitin-like modifier (SUMO)-targeted substrate, and thus differs from the canonical SUMO-targeted substrate entry. In agreement, we found that critical UBE2E1 residues involved in the capture of the TRIM21 substrate lysine are conserved in ubiquitin-conjugating E2s, whereas residues critical for SUMOylation are not conserved. We noted that coordination of the acceptor lysine leads to remodeling of amino acid side-chain interactions between the UBE2E1 active site and the E2-E3 direct interface, including the so-called "linchpin" residue conserved in RING E3s and required for ubiquitination. The findings of our work support the notion that substrate lysine activation of an E2-E3-connecting allosteric path may trigger catalytic activity and contribute to the understanding of specific lysine targeting by ubiquitin-conjugating E2s.

Solution NMR structure of the TRIM21 B-box2 and identification of residues involved in its interaction with the RING domain.

Tripartite motif-containing (TRIM) proteins are defined by the sequential arrangement of RING, B-box and coiled-coil domains (RBCC), where the B-box domain is a unique feature of the TRIM protein family. TRIM21 is an E3 ubiquitin-protein ligase implicated in innate immune signaling by acting as an autoantigen and by modifying interferon regulatory factors. Here we report the three-dimensional solution structure of the TRIM21 B-box2 domain by nuclear magnetic resonance (NMR) spectroscopy. The structure of the B-box2 domain, comprising TRIM21 residues 86-130, consists of a short α-helical segment with an N-terminal short ß-strand and two anti-parallel ß-strands jointly found the core, and adopts a RING-like fold. This ßßαß core largely defines the overall fold of the TRIM21 B-box2 and the coordination of one Zn2+ ion stabilizes the tertiary structure of the protein. Using NMR titration experiments, we have identified an exposed interaction surface, a novel interaction patch where the B-box2 is likely to bind the N-terminal RING domain. Our structure together with comparisons with other TRIM B-box domains jointly reveal how its different surfaces are employed for various modular interactions, and provides extended understanding of how this domain relates to flanking domains in TRIM proteins.