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Seasonal human coronaviruses (HCoVs) cause respiratory infections, especially in children. Currently, the knowledge on early childhood seasonal coronavirus infections and the duration of antibody levels following the first infections is limited. Here we analyzed serological follow-up samples to estimate the rate of primary infection and reinfection(s) caused by seasonal coronaviruses in early childhood. Serum specimens were collected from 140 children at ages of 13, 24, and 36 months (1, 2, and 3 years), and IgG antibody levels against recombinant HCoV nucleoproteins (N) were measured by enzyme immunoassay (EIA). Altogether, 84% (118/140) of the children were seropositive for at least one seasonal coronavirus N by the age of 3 years. Cumulative seroprevalences for HCoVs 229E, HKU1, NL63, and OC43 increased by age, and they were 45%, 27%, 70%, and 44%, respectively, at the age of 3 years. Increased antibody levels between yearly samples indicated reinfections by 229E, NL63, and OC43 viruses in 20-48% of previously seropositive children by the age of 3 years. Antibody levels declined 54-73% or 31-77% during the year after seropositivity in children initially seropositive at 1 or 2 years of age, respectively, in case there was no reinfection. The correlation of 229E and NL63, and OC43 and HKU1 EIA results, suggested potential cross-reactivity between the N specific antibodies inside the coronavirus genera. The data shows that seasonal coronavirus infections and reinfections are common in early childhood and the antibody levels decline relatively rapidly. IMPORTANCE The rapid spread of COVID-19 requires better knowledge on the rate of coronavirus infections and coronavirus specific antibody responses in different population groups. In this work we analyzed changes in seasonal human coronavirus specific antibodies in young children participating in a prospective 3-year serological follow-up study. We show that based on seropositivity and changes in serum coronavirus antibody levels, coronavirus infections and reinfections are common in early childhood and the antibodies elicited by the infection decline relatively rapidly. These observations provide further information on the characteristics of humoral immune responses of coronavirus infections in children. The rapid spread of COVID-19 requires better knowledge on the rate of coronavirus infections and coronavirus specific antibody responses in different population groups. In this work we analyzed changes in seasonal human coronavirus specific antibodies in young children participating in a prospective 3-year serological follow-up study.

Funding Information: This work was supported by Business Finland corona-co-creation funding for the project Antiviral Fibers?pilot with extracts from Finnish forests (grant: 40699/31/2020). This study was also funded by the Natural Resources Institute Finland?s strategic research funding to the projects ?More, faster, higher quality: potential of short-rotation aspen and willow biomass for novel products in bioeconomy? (AspenWill) and ?Added value potential of new and under-utilized fibre sources in Finnish value networks of green bioeconomy: prefeasibility, prototyping, and market acceptance? (VALUEPOT). In addition, Academy of Finland has supported this study via the project ?Antivirals from Forest Biomasses: Structure, Function, and Applicability? (grant: 342250). Work was also funded by Jane and Aatos Erkko Foundation: Novel probes for discovering antivirals. Publisher Copyright: Copyright © 2021 Tienaho, Reshamwala, Sarjala, Kilpeläinen, Liimatainen, Dou, Viherä-Aarnio, Linnakoski, Marjomäki and Jyske. Earlier studies have shown that the bark of Salix L. species (Salicaceae family) is rich in extractives, such as diverse bioactive phenolic compounds. However, we lack knowledge on the bioactive properties of the bark of willow species and clones adapted to the harsh climate conditions of the cool temperate zone. Therefore, the present study aimed to obtain information on the functional profiles of northern willow clones for the use of value-added bioactive solutions. Of the 16 willow clones studied here, 12 were examples of widely distributed native Finnish willow species, including dark-leaved willow (S. myrsinifolia Salisb.) and tea-leaved willow (S. phylicifolia L.) (3 + 4 clones, respectively) and their natural and artificial hybrids (3 + 2 clones, respectively). The four remaining clones were commercial willow varieties from the Swedish willow breeding program. Hot water extraction of bark under mild conditions was carried out. Bioactivity assays were used to screen antiviral, antibacterial, antifungal,yeasticidal, and antioxidant activities, as well as the total phenolic content of the extracts. Additionally, we introduce a fast and less labor-intensive steam-debarking method for Salix spp. feedstocks. Clonal variation was observed in the antioxidant properties of the bark extracts of the 16 Salix spp. clones. High antiviral activity against a non-enveloped enterovirus, coxsackievirus A9, was found, with no marked differences in efficacy between the native clones. All the clones also showed antibacterial activity against Staphylococcus aureus and Escherichia coli, whereas no antifungal (Aspergillus brasiliensis) or yeasticidal (Candida albicans) efficacy was detected. When grouping the clone extract results into Salix myrsinifolia, Salix phylicifolia, native hybrid, artificial hybrid, and commercial clones, there was a significant difference in the activities between S. phylicifolia clone extracts and commercial clone extracts in the favor of S. phylicifolia in the antibacterial and antioxidant tests. In some antioxidant tests, S. phylicifolia clone extracts were also significantly more active than artificial clone extracts. Additionally, S. myrsinifolia clone extracts showed significantly higher activities in some antioxidant tests than commercial clone extracts and artificial clone extracts. Nevertheless, the bark extracts of native Finnish willow clones showed high bioactivity. The obtained knowledge paves the way towards developing high value-added biochemicals and other functional solutions based on willow biorefinery approaches.

Lyme borreliosis is a multisystemic disease caused by the pleomorphic bacteria of the Borrelia burgdorferi sensu lato complex. The exact mechanisms for the infection to progress into a prolonged sequelae of the disease are currently unknown, although immune evasion and persistence of the bacteria in the host are thought to be major contributors. The current study investigated B. burgdorferi infection processes in two human cell lines, both non-immune and non-phagocytic, to further understand the mechanisms of infection of this bacterium. By utilizing light, confocal, helium ion, and transmission electron microscopy, borrelial infection of chondrosarcoma (SW1353) and dermal fibroblast (BJ) cells were examined from an early 30-min time point to a late 9-days post-infection. Host cell invasion, viability of both the host and B. burgdorferi, as well as, co-localization with lysosomes and the presence of different borrelial pleomorphic forms were analyzed. The results demonstrated differences of infection between the cell lines starting from early entry as B. burgdorferi invaded BJ cells in coiled forms with less pronounced host cell extensions, whereas in SW1353 cells, micropodial interactions with spirochetes were always seen. Moreover, infection of BJ cells increased in a dose dependent manner throughout the examined 9 days, while the percentage of infection, although dose dependent, decreased in SW1353 cells after reaching a peak at 48 h. Furthermore, blebs, round body and damaged B. burgdorferi forms, were mostly observed from the infected SW1353 cells, while spirochetes dominated in BJ cells. Both infected host cell lines grew and remained viable after 9 day post-infection. Although damaged forms were noticed in both cell lines, co-localization with lysosomes was low in both cell lines, especially in BJ cells. The invasion of non-phagocytic cells and the lack of cytopathic effects onto the host cells by B. burgdorferi indicated one mechanism of immune evasion for the bacteria. The differences in attachment, pleomorphic form expressions, and the lack of lysosomal involvement between the infected host cells likely explain the ability of a bacterium to adapt to different environments, as well as, a strategy for persistence inside a host. peerReviewed

Drosera rotundifolia L. is a carnivorous plant used in traditional medicine for its therapeutic properties. Because of its small size, its collection in nature is laborious and different cultivation methods have been studied to ensure availability. However, only a few studies exist where the lab-grown sundew tissue and field-grown sundew would have been compared in their functionality or metabolic profiles. In this study, the antioxidant and antiviral activities of lab-grown and field-grown sundew extracts and their metabolic profiles are examined. The effect of drying methods on the chromatographic profile of the extracts is also shown. Antioxidant activity was significantly higher (5–6 times) in field-grown sundew but antiviral activity against enterovirus strains coxsackievirus A9 and B3 was similar in higher extract concentrations (cell viability ca. 90%). Metabolic profiles showed that the majority of the identified compounds were the same but field-grown sundew contained higher numbers and amounts of secondary metabolites. Freeze-drying, herbal dryer, and oven or room temperature drying of the extract significantly decreased the metabolite content from −72% up to −100%. Freezing was the best option to preserve the metabolic composition of the sundew extract. In conclusion, when accurately handled, the lab-grown sundew possesses promising antiviral properties, but the secondary metabolite content needs to be higher for it to be considered as a good alternative for the field-grown sundew. peerReviewed

Interactions between an atomically precise gold nanocluster Au102(p-MBA)44 (p-MBA = para mercaptobenzoic acid) and a fluorescent organic dye molecule (KU, azadioxatriangulenium) are studied. In solution, the constituents form spontaneously a weakly bound complex leading to quenching of fluorescence of the KU dye via energy transfer. The KU can be separated from the complex by lowering pH, leading to recovery of fluorescence, which forms a basis for an optical reversible pH sensor. However, the sensor is not a stable entity, which could be delivered inside cells. For this purpose, a covalently bound hybrid is synthesized by linking the KU dye to the ligand layer of the cluster via an ester bond. Covalent linking facilitates entry of the cluster–dye hybrids into cells via endocytosis. Inside cells, the hybrids accumulate in endosomes where Au102 releases its cargo via hydrolysis of the ester bond. Changes of the local pH inside endosomes regulate reversible fluorescence due to variations in the interactions between the Au102 cluster and the dye. This work presents a concept for delivering reporter molecules into cells by using atomically precise gold nanoclusters as carriers and paves the way for future developments of cluster-reporter sensors for in vivo measurements of e.g. absolute pH values or ion concentrations. peerReviewed

We report that several viruses from the human enterovirus group B cause massive vimentin rearrangements during lytic infection. Comprehensive studies suggested that viral protein synthesis was triggering the vimentin rearrangements. Blocking the host cell vimentin dynamics with IDPN did not significantly affect the production of progeny viruses and only moderately lowered the synthesis of structural proteins such as VP1. In contrast, the synthesis of the non-structural proteins 2A, 3C, and 3D was drastically lowered. This led to attenuation of the cleavage of the host cell substrates PABP and G3BP1 and reduced caspase activation, thus leading to prolonged cell survival. Furthermore, the localization of the proteins differed in the infected cells. Capsid protein VP1 was found diffusely around the cytoplasm, whereas 2A and 3D followed vimentin distribution. Based on protein blotting, lower amounts of non-structural proteins did not result from proteasomal degradation, but from lower synthesis without intact vimentin cage structure. In contrast, inhibition of Hsp90 chaperone activity, which regulates P1 maturation, lowered the amount of VP1, but had less effect on 2A. The results suggest that, the vimentin dynamics regulate viral non-structural protein synthesis while having no effect on structural protein synthesis or overall infection efficiency. The results presented here shed new light on differential fate of structural and non-structural proteins of enteroviruses, having consequences on host cell survival. Importance A virus needs the host cell in order to replicate and produce new progeny viruses. For this, the virus takes over the host cell and modifies it to become a factory for viral proteins. Irrespective of the specific virus family, these proteins can be divided into structural and non-structural proteins. Structural proteins are the building blocks for the new progeny virions, whereas the non-structural proteins orchestrate the take-over of the host cell and its functions. Here we have shown a mechanism that viruses exploit in order to regulate the host cell. We show that viral protein synthesis induces vimentin cages, which promote production of specific viral proteins that eventually control apoptosis and the host cell death. This study specifies vimentin as the key regulator of these events and indicates that viral proteins have different fates in the cells depending on their association with vimentin cages. peerReviewed

Fluorescent probes for sensing nucleic acids have found widespread use in the field of cell and molecular biology. However, probes combined with potential for post-synthetic conjugation, e.g. for intra-endosomal measurements of RNA, are unavailable. Herein we developed cyanine dyes that can be conjugated to viral capsid or other targets. First, we solved the crystal structure of SYBR Green II. The structural elucidation of this commonly used RNA probe provided the basis for synthesizing similar molecules with much desired function for post-synthetic conjugation. To address this need, cyanine dyes were prepared using an alternative synthesis protocol. All studied compounds showed considerable brightness upon binding to nucleic acids. However, regardless of the common chromophore on the dyes, the observed fluorescence emission intensities varied significantly, where methyl-substituted dye 1 gave values higher than SYBR Green II, whereas compounds 2–5 containing undecyl spacers had lower values. Studying the structure-activity relationship revealed the longer alkyl chains to induce slight perturbation in dye intercalation, as well as demanding larger binding area on the nucleic acid lattice, explaining these differences. To study the potential biological use of the dyes, the RNA genome of enterovirus echovirus 1 was studied in vitro with the probes. A novel method employing the low binding space requirement of 1 was developed to determine the single-to-double-stranded RNA ratio of a sample, whereas compound 4 was covalently bound to the viral capsid and used successfully to monitor the viral RNA release from within the capsid. The presented results open new possibilities for preparation and use of SYBR Green-based nucleic acid probes to further apply these compounds for increasingly demanding targeting in biological contexts. peerReviewed