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Abstract— Variability in infection rates of trematodes Diplostomum pseudospathaceum in 0+ rainbow trout Oncorhynchus mykiss under different hydraulic regimes was studied in the experimental setup with regulated flow rates and environmental heterogeneity. The average infection rate in the conditions of the current was 1.5 times less than that in the stagnant water. An increase of the flow rate from 3.2 to 11.3 cm/s did not result in lower infection rates, while the interindividual variability in the infection rate (coefficient of variation) tended to increase along with changing the still-water conditions (20%) to the high flow rate environments (40%) within the experiment. A decrease in the average infection rate and an increase in the variability within the heterogenous flow of water could indicate the fact that some fish effectively avoid infection. It may be caused by redistribution of cercariae suspended in the water column across microhabitats with different flow rates together with fish behavior which allows them to choose sites with low parasite concentration.

Enteroviruses are one of the most abundant viruses causing mild to serious acute infections in humans and also contributing to chronic diseases like type 1 diabetes. Presently, there are no approved antiviral drugs against enteroviruses. Here, we studied the potency of vemurafenib, an FDA-approved RAF kinase inhibitor for treating BRAFV600E mutant-related melanoma, as an antiviral against enteroviruses. We showed that vemurafenib prevented enterovirus translation and replication at low micromolar dosage in an RAF/MEK/ERK-independent manner. Vemurafenib was effective against group A, B, and C enteroviruses, as well as rhinovirus, but not parechovirus or more remote viruses such as Semliki Forest virus, adenovirus, and respiratory syncytial virus. The inhibitory effect was related to a cellular phosphatidylinositol 4-kinase type IIIβ (PI4KB), which has been shown to be important in the formation of enteroviral replication organelles. Vemurafenib prevented infection efficiently in acute cell models, eradicated infection in a chronic cell model, and lowered virus amounts in pancreas and heart in an acute mouse model. Altogether, instead of acting through the RAF/MEK/ERK pathway, vemurafenib affects the cellular PI4KB and, hence, enterovirus replication, opening new possibilities to evaluate further the potential of vemurafenib as a repurposed drug in clinical care. peerReviewed

Interactions among symbiotic organisms and their hosts are major drivers of ecological and evolutionary processes. Monitoring the infection patterns among natural populations and identifying factors affecting these interactions are critical for understanding symbiont–host relationships. However, many of these interactions remain understudied since the knowledge about the symbiont species is lacking, which hinders the development of appropriate tools. In this study, we developed a digital droplet PCR (ddPCR) assay based on apicomplexan COX1 gene to detect an undescribed agamococcidian symbiont. We show that the method gives precise and reproducible results and enables detecting cryptic symbionts in low target concentration. We further exemplify the assay's use to survey seasonally sampled natural host (Pygospio elegans) populations for symbiont infection dynamics. We found that symbiont prevalence differs spatially but does not show seasonal changes. Infection load differed between populations and was low in spring and significantly increased towards fall in all populations. We also found that the symbiont prevalence is affected by host length and population density. Larger hosts were more likely to be infected, and high host densities were found to have a lower probability of infection. The observed variations could be due to characteristics of both symbiont and host biology, especially the seasonal variation in encounter rates. Our findings show that the developed ddPCR assay is a robust tool for detecting undescribed symbionts that are otherwise difficult to quantify, enabling further insight into the impact cryptic symbionts have on their hosts., Many symbiont‐host interactions are understudied due to the lack of knowledge about the symbiont species hindering the development of appropriate tools. Here, we developed a ddPCR assay to detect an undescribed agamococcidian symbiont in its host, Pygospio elegans, and show that the method enables detecting cryptic symbionts even in low target concentration. We further exemplify the assay's use to survey seasonally sampled natural host populations for symbiont infection dynamics.

Upon infection, viruses hijack the cell machinery and remodel host cell structures to utilize them for viral proliferation. Since viruses are about a thousand times smaller than their host cells, imaging virus-host interactions at high spatial resolution is like looking for a needle in a haystack. Scouting gross cellular changes with fluorescent microscopy is only possible for well-established viruses, where fluorescent tagging is developed. Soft X-ray tomography (SXT) offers 3D imaging of entire cells without the need for chemical fixation or labeling. Here, we use full-rotation SXT to visualize entire human B cells infected by the herpes simplex virus 1 (HSV-1). We have mapped the temporospatial remodeling of cells during the infection and observed changes in cellular structures, such as the presence of cytoplasmic stress granules and multivesicular structures, formation of nuclear virus-induced dense bodies, and aggregates of capsids. Our results demonstrate the power of SXT imaging for scouting virus-induced changes in infected cells and understanding the orchestration of virus-host remodeling quantitatively.

SARS- CoV- 2 has been detected both in air and on surfaces, but questions remain about the patient-specific and environmental factors affecting virus transmission. Additionally, more detailed information on viral sampling of the air is needed. This prospective cohort study (N= 56) presents results from 258 air and 252 surface samples from the surroundings of 23 hospitalized and eight home- treated COVID-19 index patients between July 2020 and March 2021 and compares the results between the measured environments and patient factors. Additionally, epidemiological and experimental investigations were performed. The proportions of qRT- PCR- positiveair (10.7% hospital/17.6% homes) and surface samples (8.8%/12.9%) showed statisti-cal similarity in hospital and homes. Significant SARS- CoV- 2 air contamination was observed in a large (655.25 m3) mechanically ventilated (1.67 air changes per hour, 32.4– 421 L/s/patient) patient hall even with only two patients present. All positive air samples were obtained in the absence of aerosol- generating procedures. In four cases, positive environmental samples were detected after the patients had devel-oped a neutralizing IgG response. SARS- CoV- 2 RNA was detected in the following particle sizes: 0.65– 4.7 μm , 7. 0– 12.0 μm, >10μm, and

Parvoviruses are small non-enveloped single-stranded DNA viruses, which depend on host cell nuclear transcriptional and replication machinery. After endosomal exposure of nuclear localization sequence and a phospholipase A2 domain on the capsid surface, and escape into the cytosol, parvovirus capsids enter the nucleus. Due to the small capsid diameter of 18–26 nm, intact capsids can potentially pass into the nucleus through nuclear pore complexes (NPCs). This might be facilitated by active nuclear import, but capsids may also follow an alternative entry pathway that includes activation of mitotic factors and local transient disruption of the nuclear envelope. The nuclear entry is followed by currently undefined events of viral genome uncoating. After genome release, viral replication compartments are initiated and infection proceeds. Parvoviral genomes replicate during cellular S phase followed by nuclear capsid assembly during virus-induced S/G2 cell cycle arrest. Nuclear egress of capsids occurs upon nuclear envelope degradation during apoptosis and cell lysis. An alternative pathway for nuclear export has been described using active transport through the NPC mediated by the chromosome region maintenance 1 protein, CRM1, which is enhanced by phosphorylation of the N-terminal domain of VP2. However, other alternative but not yet uncharacterized nuclear export pathways cannot be excluded. peerReviewed

Coinfections of two or more parasites within one host are more of a rule than an exception in nature. Interactions between coinfecting parasites can greatly affect their abundance and prevalence. Characteristics of the host, such as genetic diversity, can also affect the infection dynamics of coinfecting parasites. Here we investigate for the first time the association of coinfection patterns of two marine apicomplexans, Rhytidocystis sp. and Selenidium pygospionis, with the genetic diversity of their host, the polychaete Pygospio elegans, from natural populations. Host genetic diversity was determined with seven microsatellite loci and summarized as allelic richness, inbreeding coefficient, and individual heterozygosity. We detected nonsignificant correlations between infection loads and both individual host heterozygosity and population genetic diversity. Prevalence and infection load of Rhytidocystis sp. were higher than that of S. pygospionis and both varied spatially. Coinfections were common and almost all hosts infected by S. pygospionis were also infected by Rhytidocystis sp.. Rhytidocystis sp. infection load was significantly higher in dual infections. Our results suggest that factors other than host genetic diversity might be more important in marine apicomplexan infection patterns and experimental approaches would be needed to further determine how interactions between the apicomplexans and their host influence infection. peerReviewed

Autonomous parvoviruses encode at least two nonstructural proteins, NS1 and NS2. While NS1 is linked to important nuclear processes required for viral replication, much less is known about the role of NS2. Specifically, the function of canine parvovirus (CPV) NS2 has remained undefined. Here we have used proximity-dependent biotin identification (BioID) to screen for nuclear proteins that associate with CPV NS2. Many of these associations were seen both in noninfected and infected cells, however, the major type of interacting proteins shifted from nuclear envelope proteins to chromatin-associated proteins in infected cells. BioID interactions revealed a potential role for NS2 in DNA remodeling and damage response. Studies of mutant viral genomes with truncated forms of the NS2 protein suggested a change in host chromatin accessibility. Moreover, further studies with NS2 mutants indicated that NS2 performs functions that affect the quantity and distribution of proteins linked to DNA damage response. Notably, mutation in the splice donor site of the NS2 led to a preferred formation of small viral replication center foci instead of the large coalescent centers seen in wild-type infection. Collectively, our results provide insights into potential roles of CPV NS2 in controlling chromatin remodeling and DNA damage response during parvoviral replication. publishedVersion

COVID-19 has highlighted the need for indoor risk-reduction strategies. Our aim is to provide information about the virus dispersion and attempts to reduce the infection risk. Indoor transmission was studied simulating a dining situation in a restaurant. Aerosolized Phi6 viruses were detected with several methods. The aerosol dispersion was modeled by using the Large-Eddy Simulation (LES) technique. Three risk-reduction strategies were studied: (1) augmenting ventilation with air purifiers, (2) spatial partitioning with dividers, and (3) combination of 1 and 2. In all simulations infectious viruses were detected throughout the space proving the existence long-distance aerosol transmission indoors. Experimental cumulative virus numbers and LES dispersion results were qualitatively similar. The LES results were further utilized to derive the evolution of infection probability. Air purifiers augmenting the effective ventilation rate by 65% reduced the spatially averaged infection probability by 30%-32%. This relative reduction manifests with approximately 15 min lag as aerosol dispersion only gradually reaches the purifier units. Both viral findings and LES results confirm that spatial partitioning has a negligible effect on the mean infection-probability indoors, but may affect the local levels adversely. Exploitation of high-resolution LES jointly with microbiological measurements enables an informative interpretation of the experimental results and facilitates a more complete risk assessment.

Tämä opinnäytetyö käsittelee tutkimustietoa tyypin 1 diabeteksen (T1D) patogeneesin hypoteesista, jonka mukaan kliiniseen sairauteen johtavan autoimmuunireaktion laukaisee haiman saarekkeiden virusinfektio. T1D on autoimmuunitauti, jossa haiman saarekkeiden beetasolut tuhoutuvat, jolloin niiden insuliinin tuotanto menetetään. Insuliinin tuotannon romahtaessa kehon energia-aineenvaihdunta häiriintyy vakavasti. Hoitamattomana tila johtaa kuolemaan. Menetetty haiman insuliinituotanto voidaan korvata insuliinivalmisteilla, mutta tästä huolimatta potilaat kärsivät merkittävästä elämänlaadun laskusta ja lyhyemmästä elinajanennusteesta. Lisäksi insuliinihoito on kallista. Haiman beetasoluja tuhoavan autoimmuunireaktion laukaisevat tekijät ja solutuhon mekanismit tunnetaan edelleenkin vain osittain. Tiedetään, että tietyt HLA-genotyypit altistavat T1D:n puhkeamiselle. Autoimmuuniprosessi alkaa jo kauan ennen kuin T1D voidaan kliinisesti todeta. Tätä ennen haiman saarekkeisiin hakeutuu lymfosyyttejä, jolloin kehittyy saarekkeiden tulehdusreaktio – insuliitti. Myös humoraalisen immuunijärjestelmän aktivoituminen beetasoluja vastaan voidaan usein havaita ennen T1D:n kliinistä ilmentymistä: Verenkierrossa todetaan autovasta-aineita beetasolujen proteiineja kohtaan. Autovasta-aineista tärkeimpiä ovat insuliini-, GAD65-, IA-2- ja ZnT8-vasta-aineet. Laajoissa epidemiologisissa tutkimuksissa on havaittu enterovirusten sukuun kuuluvien coxsackie B -virusten (CVB) olevan yhteydessä T1D:n puhkeamiseen. CVB:t tartuttavat pääasiassa hengitystie-eritteiden tai ulostekontaminaation välityksellä, ja niiden ensisijainen replikaatiopaikka on suolisto. Ne aiheuttavat etenkin lapsissa yleensä vähäoireisia tauteja, mutta ne voivat aiheuttaa myös hankalia monielininfektioita. CVB:ten on osoitettu infektoivan haiman saarekkeiden beetasoluja in vitro ja in vivo sekä kokeellisissa hiirimalleissa että ihmisissä. CVB pääsee beetasoluihin niiden solukalvollaan ilmentämän coxsackie-adenovirusreseptorin (CAR) välityksellä. Hiiritutkimuksissa on havaittu, että vain tietyt CVB:ten alalajit kykenevät aiheuttamaan beetasolujen infektion ja insuliininpuutosdiabeteksen kaltaisen taudin hiirillä. CVB4 E2 ja CVB4 JVB -viruskannat aiheuttavat insuliininpuutosdiabeteksen hiirille. Tutkimuksissa, joissa on käytetty muita CVB-kantoja, ei ole kyetty aiheuttamaan hiirten beetasolujen infektiota. Persistoivassa infektiossa virus ei infektion akuutin lyyttisen vaiheen jälkeen poistu kudoksesta. Todisteita tämänlaisesta matala-asteisesta enterovirusinfektiosta on havaittu vastikään T1D:een sairastuneiden haiman saarekkeista. Haiman saarekkeiden persistoivan CVB-infektion ajatellaan olevan yksi T1D:een johtavan autoimmuuniprosessin laukaiseva tekijä. Persistoivia CVB-infektioita on pystytty kokeellisesti aikaansaamaan ihmisen haimasolulinjoissa in vitro ja hiirillä myös in vivo. Vaikka selkeää kausaliteettia CVB-infektion ja T1D:n kehittymisen välillä ei ole vielä voitu osoittaa, nämä infektiot ovat tärkeä tutkimuskohde T1D:n patogeneesin ymmärtämisen kannalta. Virusten ja T1D:n yhteyden paremmin ymmärtäminen saattaa tulevaisuudessa mahdollistaa T1D:n ehkäisyn esimerkiksi rokotusten avulla. This thesis aims to highlight current evidence on one of the leading hypothesis of type 1 diabetes (T1D) pathogenesis, where the disease is caused by an autoimmune reaction triggered by viral infection in the pancreatic islets. T1D is an autoimmune disease, in which the beta cells within the pancreatic islets are destroyed and their insulin production is lost. The loss of insulin production leads to a crisis in body energy metabolism that, if left untreated, will lead to death. Even if treated with exogenous insulin therapy, T1D causes major decline in quality of life and life expectancy. Insulin treatment is also expensive. The autoimmune progress and the factors that lead to it are now well understood at the moment. It is known that certain HLA-genotypes predispose to T1D. The autoimmune process leading to T1D begins long before the condition manifests clinically. Before that, the pancreatic islets are infiltrated by lymphocytes, leading to an inflammation of the pancreatic islets – insulitis. Activation of the humoral immune system can also be detected before clinical T1D manifestation. Autoantibodies against beta cell proteins are detected in blood, most important being autoantibodies against insulin, GAD65, IA-2, and ZnT8. Coxsackie B viruses (CVBs) that belong in the genus Enterovirus have associated with T1D onset in large epidemiological studies. CVBs are transmitted mainly via respiratory or faecal-oral route and their primary replication site is the gastrointestinal tract. The symptoms of an infection are usually mild in children, but CVBs can also cause severe septic multiorgan infections. CVBs have been shown to infect pancreatic beta cells in vitro and in vivo in mouse models as well as in humans. CVBs can enter the beta cells via the Coxsackievirus and adenovirus receptor (CAR) on the cell surface. Mouse studies have shown that only select few CVB strains can infect beta cells and cause T1D-like syndrome in mice. CVB4 E2 and CVB4 JVB virus strains can cause T1D-like syndrome in mice. Other studies that have used other CVB strains have not been able to infect mouse beta cells. During persistent infection the virus is not cleared after the acute lytic phase. Evidence of persistent enterovirus infection have been found in the pancreases of recent-onset T1D patients. Persistent CVB infection of pancreatic islets is thought to be the factor triggering the autoimmune process leading to T1D. Persistent CVB infections have been established in human pancreatic cell models in vitro and in mouse models also in vivo. Although no definitive causality between CVB infections and the development of T1D has been proven as of to date, these infections are important focus point in the research of T1D pathogenesis. Better understanding of the association between T1D and viruses may help to prevent T1D using e.g., vaccines.

Herpes simplex virus capsids are assembled and packaged in the nucleus and move by diffusion through the nucleoplasm to the nuclear envelope for egress. Analyzing their motion provides conclusions not only on capsid transport but also on the properties of the nuclear environment during infection. We utilized live-cell imaging and single-particle tracking to characterize capsid motion relative to the host chromatin. The data indicate that as the chromatin was marginalized toward the nuclear envelope it presented a restrictive barrier to the capsids. However, later in infection this barrier became more permissive and the probability of capsids to enter the chromatin increased. Thus, although chromatin marginalization initially restricted capsid transport to the nuclear envelope, a structural reorganization of the chromatin counteracted that to promote capsid transport later. Analyses of capsid motion revealed that it was subdiffusive, and that the diffusion coefficients were lower in the chromatin than in regions lacking chromatin. In addition, the diffusion coefficient in both regions increased during infection. Throughout the infection, the capsids were never enriched at the nuclear envelope, which suggests that instead of nuclear export the transport through the chromatin is the rate-limiting step for the nuclear egress of capsids. This provides motivation for further studies by validating the importance of intranuclear transport to the life cycle of HSV-1., Author summary Herpes simplex virus type 1 is a DNA virus that is studied intensively due to its high prevalence in humans, its ability to cause many diseases upon lytic replication or to evade the immune system in a latent state, and its potential use in oncolytic and immunotherapeutic applications. The nuclear replication of herpes simplex virus type 1 leads to the emergence of viral replication compartments together with relocalization and condensation of the host cell chromatin to the nuclear periphery. The reorganization of the host chromatin potentially hampers the intranuclear transport and nuclear egress of newly assembled progeny capsids. Nuclear capsids move by diffusion prior to their viral nuclear egress complex-mediated exit through the nuclear envelope. However, it is not understood how the capsids travel through the chromatin network. Our results show that infection-induced changes in chromatin architecture enhance nuclear diffusion of capsids at late infection, and that this transport is the rate-limiting step in the nuclear egress of the virus. Understanding the kinetics of capsid translocation in the nucleus is of fundamental interest in the field of DNA virus research because it explains how viruses are able to move in the chromatin before their nuclear egress.

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

Zoonotic diseases, caused by pathogens transmitted between other vertebrate animals and humans, pose a major risk to human health. Rodents are important reservoir hosts for many zoonotic pathogens, and rodent population dynamics affect the infection dynamics of rodent-borne diseases, such as diseases caused by hantaviruses. However, the role of rodent population dynamics in determining the infection dynamics of rodent-associated tick-borne diseases, such as Lyme borreliosis (LB), caused by Borrelia burgdorferi sensu lato bacteria, have gained limited attention in Northern Europe, despite the multiannual abundance fluctuations, the so-called vole cycles, that characterise rodent population dynamics in the region. Here, we quantify the associations between rodent abundance and LB human cases and Puumala Orthohantavirus (PUUV) infections by using two time series (25-year and 9-year) in Finland. Both bank vole (Myodes glareolus) abundance as well as LB and PUUV infection incidence in humans showed approximately 3-year cycles. Without vector transmitted PUUV infections followed the bank vole host abundance fluctuations with two-month time lag, whereas tick-transmitted LB was associated with bank vole abundance ca. 12 and 24 months earlier. However, the strength of association between LB incidence and bank vole abundance ca. 12 months before varied over the study years. This study highlights that the human risk to acquire rodent-borne pathogens, as well as rodent-associated tick-borne pathogens is associated with the vole cycles in Northern Fennoscandia, yet with complex time lags. peerReviewed

Parvoviruses are small single-stranded (ss) DNA viruses, which replicate in the nucleoplasm and affect both the structure and function of the nucleus. The nuclear stage of the parvovirus life cycle starts at the nuclear entry of incoming capsids and culminates in the successful passage of progeny capsids out of the nucleus. In this review, we will present past, current, and future microscopy and biochemical techniques and demonstrate their potential in revealing the dynamics and molecular interactions in the intranuclear processes of parvovirus infection. In particular, a number of advanced techniques will be presented for the detection of infection-induced changes, such as DNA modification and damage, as well as protein–chromatin interactions. peerReviewed

Koronatestitulos on negatiivinen, mutta infektion yleisoireet eivät hellitä. Lääkäri pohtii ääneen, voisiko vika olla korvien välissä. Miten tällainen ajatuksellinen hyppäys tapahtuu, kun COVID-19-taudin eteneminen ja pitkäaikaisvaikutukset ovat huonosti tunnettuja? Vastausta voi hakea infektioiden ja psykosomaattisen lääketieteen historioiden yhtymäkohdista.

The canonical lytic–lysogenic binary has been challenged in recent years, as more evidence has emerged on alternative bacteriophage infection strategies. These infection modes are little studied, and yet they appear to be more abundant and ubiquitous in nature than previously recognized, and can play a significant role in the ecology and evolution of their bacterial hosts. In this review, we discuss the extent, causes and consequences of alternative phage lifestyles, and clarify conceptual and terminological confusion to facilitate research progress. We propose distinct definitions for the terms ‘pseudolysogeny’ and ‘productive or non-productive chronic infection’, and distinguish them from the carrier state life cycle, which describes a population-level phenomenon. Our review also finds that phages may change their infection modes in response to environmental conditions or the physiological state of the host cell. We outline known molecular mechanisms underlying the alternative phage–host interactions, including specific genetic pathways and their considerable biotechnological potential. Moreover, we discuss potential implications of the alternative phage lifestyles for microbial biology and ecosystem functioning, as well as applied topics such as phage therapy.

The overall CoCID concept is centred on providing virologists with a next-generation imaging device, which, through increased penetration and depth of focus, as well as through high natural contrast and sensitivity to organelle density (including virus-related organelles), will produce higher-fidelity ultrastructural images of whole intact cells. These insights will, in turn, help increase our understanding of the links between the structural reorganisation of cells and the mechanisms of viral entry, replication, assembly, and egress in cells. CoCID will provide this valuable imaging capability in the form of a compact lab-scale device that will greatly improve the accessibility of soft X-ray microscopy for the virologists' consortium within the CoCID project as well as the broader disease research community worldwide.

Lähtökohdat: Clostridium difficile -infektiot ovat lisääntyneet maailmalla. Lasten positiivisen löydöksen tulkinta on haastavaa oireettomien kantajien merkittävän osuuden vuoksi. Menetelmät: Aineistoon kerättiin Tampereen yliopistollisen sairaalan (TAYS) lastentautien yksiköistä alle 18-vuotiaiden potilaiden C. difficile -löydökset vuosina 2008–2017. Tapaus-verrokkitutkimuksessa selvitettiin altistavia tekijöitä sekä löydösten liittymistä kliiniseen taudinkuvaan. Tulokset: Tutkimusajankohtana otettiin yhteensä 644 näytettä ja 94 lapselta löytyi C. difficile -toksiinia ulosteesta. Kaikista näytteistä 32,8 % ja positiivisista näytteistä 38,3 % oli otettu ≤ 12 kuukauden ikäisiltä lapsilta. Heistä 31 % sai infektioon antibioottihoidon ja heiltä ei löytynyt infektiolle selittävää tekijää. Yli vuoden ikäisillä potilailla antibioottihoito, nenä-mahaletku, kirurgia sekä perustauti lisäsivät riskiä C. difficile -infektiolle. Päätelmät: Merkittävä osa lasten C. difficile -näytteistä otetaan imeväisikäisiltä lapsilta ja heillä todetaan myös suuri osa positiivisista löydöksistä. Pienten lasten C. difficile -löydöksiin on suhtauduttava kuitenkin kriittisesti oireettoman kantajuuden yleisyyden vuoksi. Lisäksi kaikissa ikäryhmissä löydösten merkitystä on arvioitava kliiniseen kuvaan suhteutettuna.

The microbial community surrounding fish eyed eggs can harbor pathogenic bacteria. In this study we focused on rainbow trout (Oncorhynchus mykiss) eyed eggs and the potential of bacteriophages against the pathogenic bacteria Flavobacterium psychrophilum and F. columnare. An infection bath method was first established, and the effects of singular phages on fish eggs was assessed (survival of eyed eggs, interaction of phages with eyed eggs). Subsequently, bacteria-challenged eyed eggs were exposed to phages to evaluate their effects in controlling the bacterial population. Culture-based methods were used to enumerate the number of bacteria and/or phages associated with eyed eggs and in the surrounding environment. The results of the study showed that, with our infection model, it was possible to re-isolate F. psychrophilum associated with eyed eggs after the infection procedure, without affecting the survival of the eggs in the short term. However, this was not possible for F. columnare, as this bacterium grows at higher temperatures than the ones recommended for incubation of rainbow trout eyed eggs. Bacteriophages do not appear to negatively affect the survival of rainbow trout eyed eggs and they do not seem to strongly adhere to the surface of eyed eggs either. Finally, the results demonstrated a strong potential for short term (24 h) phage control of F. psychrophilum. However, further studies are needed to explore if phage control can be maintained for a longer period and to further elucidate the mechanisms of interactions between Flavobacteria and their phages in association with fish eggs.

The reduction in host fitness caused by parasite infections (virulence) depends on infection intensity and the degree of damage caused per parasite. Environmental conditions can shape both virulence components, but in contrast to infection intensity, environmental impacts on per-parasite damage are poorly understood. Here, we studied the effect of ambient temperature on per-parasite damage, which is jointly determined by the ability of parasites to induce harm (per-parasite pathogenicity) and the ability of hosts to limit damage (tolerance). We experimentally exposed two salmonid species, Atlantic salmon (Salmo salar) and sea trout (Salmo trutta), to replicated genotypes of the eye fluke Diplostomum pseudospathaceum. After development of health damage (eye cataracts) in warm water (16 °C) during the first 12 weeks post exposure, we maintained the fish at either 5 °C (cold water) or 16 °C for another 8 weeks and quantified changes in cataracts as a function of parasite load. We found that per-parasite damage was reduced in cold compared to warm water, suggesting that cold temperatures improved host health. Per-parasite damage was also affected by parasite genotype and host species, but these effects did not change with temperature. Our findings suggest that cold-water seasons, which are often neglected in host-parasite studies due to low infection risk, could allow hosts to recuperate and thus, may have important implications for the ecology and epidemiology of parasite infections. peerReviewed

Keisarileikkaus on suuri kirurginen toimenpide, joka altistaa synnyttäjän monille komplikaatioille. Tavallisimpia näistä komplikaatioista ovat lapsivuodeaikana (synnytystä seuraavat 6–8 viikkoa) esiintyvät infektiot, joiden esiintyvyys on keisarileikkauksen yhteydessä 5–20-kertainen alatiesynnytykseen verrattuna. Lapsivuodeajan infektioihin kuuluvat esimerkiksi endometriitti eli kohdun sisälimakalvon tulehdus, keisarileikkaushaavan infektio, virtsatieinfektio ja verenmyrkytys eli sepsis. Tämän tutkimuksen tavoitteena oli selvittää lapsivuodeajan infektioiden esiintyvyyttä ja infektioiden riskitekijöitä. Tutkimus toteutettiin keräämällä tietoja Tampereen yliopistollisessa sairaalassa (Tays) vuonna 2018 keisarileikkauksella synnyttäneiltä naisilta. Tutkimusaineisto käsitti kaikki vuonna 2018 Taysissa keisarileikkauksella synnyttäneet naiset, kaikkiaan 655 potilasta. Tiedot kehittyneistä infektioista kerättiin potilaskertomuksista. Tutkimuksessa todettiin, että vuonna 2018 Taysissa 31 potilasta (4,7 % tutkimuspopulaatiosta) sairastui yhteen tai useampaan lapsivuodeinfektioon. Kun otetaan huomioon samoille potilaille kehittyneet useammat infektiot, kehittyi infektioita yhteensä 36 kappaletta. Tutkimuksessa havaittiin, että infektioihin tilastollisesti merkitsevässä yhteydessä olivat pidempi synnytyksen jälkeinen sairaalassaoloaika, raskausajan infektiosairastavuus sekä suolilaman kehittyminen. Lisäksi korioamnioniittia (sikiökalvojen ja lapsiveden infektio) esiintyi lähes kaksinkertaisesti niillä, jotka sairastuivat lapsivuodeinfektioon (16,1 % vs. 8,7 %). Tämä yhteys ei kuitenkaan ollut tilastollisesti merkitsevä. Eräänä tutkimustuloksena havaittiin myös, että kaikki Taysin keisarileikkausten yhteydessä tapahtuvat lapsivuodeinfektiot eivät päädy sairaalainfektiorekisteriin. Loppupäätelmänä pystyttiin toteamaan, että Taysin lapsivuodeinfektioiden esiintyvyys on matalammalla tasolla kuin useissa kansainvälisissä kirjallisuuden lähteissä. Tulevaisuudessa tärkeä kehityskohta voisi olla myös avohoidossa kehittyneiden lapsivuodeinfektioiden ottaminen mukaan tutkimuksiin.

Viruses need to open, i.e., uncoat, in order to release their genomes for efficient replication and translation. Especially for non-enveloped viruses, such as enteroviruses, the cues leading to uncoating are less well known. The status of the virus has previously been observed mainly by transmission electron microscopy using negative staining, cryo electron microscopy, X-ray crystallography or gradient separation (reviewed in Tuthill et al., 2010, Myllynen et al., 2016, Ruokolainen et al., 2019). However, monitoring of uncoating has been limited by the lack of methods detecting dynamic changes of the virions. Here, we present a real-time fluorescence based protocol, which detects the viral genome (RNA) during various stages of uncoating in vitro, while RNA is still inside the particle that has been expanded before the actual RNA release, and when the RNA has been totally released from the viral particle. Our method allows to explore how various molecular factors may promote or inhibit virus uncoating. peerReviewed

Echovirus 30 (E30), a member of the enterovirus B species, is a major cause of viral meningitis, targeting children and adults alike. While it is a frequently isolated enterovirus and the cause of several outbreaks all over the world, surprisingly little is known regarding its entry and replication strategy within cells. In this study, we used E30 strain Bastianni (E30B) generated from an infectious cDNA clone in order to study early entry events during infection in human RD cells. E30B required the newly discovered Fc echovirus receptor (FcRn) for successful infection, but not the coxsackievirus and adenovirus receptor (CAR) or decay-accelerating factor (DAF), although an interaction with DAF was observed. Double-stranded RNA replication intermediate was generated between 2 and 3 h postinfection (p.i.), and viral capsid production was initiated between 4 and 5 h p.i. The drugs affecting Rac1 (NSC 23766) and cholesterol (filipin III) compromised infection, whereas bafilomycin A1, dyngo, U-73122, wortmannin, and nocodazole did not, suggesting the virus follows an enterovirus-triggered macropinocytic pathway rather than the clathrin pathway. Colocalization with early endosomes and increased infection due to constitutively active Rab5 expression suggests some overlap and entry to classical early endosomes. Taken together, these results suggest that E30B induces an enterovirus entry pathway, leading to uncoating in early endosomes. IMPORTANCE Echovirus 30 (E30) is a prevalent enterovirus causing regular outbreaks in both children and adults in different parts of the world. It is therefore surprising that relatively little is known of its infectious entry pathway. We set out to generate a cDNA clone and gradient purified the virus in order to study the early entry events in human cells. We have recently studied other enterovirus B group viruses, like echovirus 1 (EV1) and coxsackievirus A9 (CVA9), and found many similarities between those viruses, allowing us to define a so-called “enterovirus entry pathway.” Here, E30 is reminiscent of these viruses, for example, by not relying on acidification for infectious entry. However, despite not using the clathrin entry pathway, E30 accumulates in classical early endosomes.

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

A burgeoning body of literature suggests that poor childhood health leads to adverse health outcomes. lower educational attainment and weaker labour market outcomes in adulthood. We focus on an important but under-researched topic, which is the role played by infection-related hospitalization (IRH) in childhood and its links to labour market outcomes later in life. The participants aged 24-30 years in 2001 N =1706 were drawn from the Young Finns Study, which includes comprehensive registry data on IRHs in childhood at ages 0-18 years. These data are linked to longitudinal registry information on labour market outcomes (2001-2012) and parental background (1980). The estimations were performed using ordinary least squares (OLS). The results showed that having an additional IRH is associated with lower log earnings (b = 95% confidence interval (CI): -0.193; -0.026). fewer years of being employed (b = -0.018. 95% CI: -0.031; -0.005). a higher probability of receiving any social income transfers (b = 0.012, 95 % CI: -0.002; 0.026) and larger social income transfers, conditional on receiving any (b = 0.085,95 % CI: 0.025; 0.145). IRHs are negatively linked to human capital accumulation. which explains a considerable part of the observed associations between IRHs and labour market outcomes. We did not find support for the hypothesis that adult health mediates the link. (C) 2020 The Authors. Published by Elsevier B.V.

Background Viral infection is implicated in development of autoimmunity. Parvovirus B19 (B19V) nonstructural protein, NS1, a helicase, covalently modifies self double-stranded deoxyribonucleic acid (dsDNA) and induces apoptosis. This study tested whether resulting apoptotic bodies (ApoBods) containing virally modified dsDNA could induce autoimmunity in an animal model. Methods BALB/c mice were inoculated with (1) pristane-induced, (2) B19V NS1-induced, or (3) staurosporine-induced ApoBods. Serum was tested for dsDNA autoantibodies by Crithidia luciliae staining and enzyme-linked immunosorbent assay. Brain, heart, liver, and kidney pathology was examined. Deposition of self-antigens in glomeruli was examined by staining with antibodies to dsDNA, histones H1 and H4, and TATA-binding protein. Results The B19V NS1-induced ApoBod inoculation induced dsDNA autoantibodies in a dose-dependent fashion. Histopathological features of immune-mediated organ damage were evident in pristane-induced and NS1-induced ApoBod groups; severity scores were higher in these groups than in staurosporine-treated groups. Tissue damage was dependent on NS1-induced ApoBod dose. Nucleosomal antigens were deposited in target tissue from pristane-induced and NS1-induced ApoBod inoculated groups, but not in the staurosporine-induced ApoBod inoculated group. Conclusions This study demonstrated proof of principle in an animal model that virally modified dsDNA in apoptotic bodies could break tolerance to self dsDNA and induce dsDNA autoantibodies and end-organ damage., Inoculation of normal mice with apoptotic bodies, generated by parvovirus B19V helicase NS1 expression, induces dsDNA antibodies and organ damage. This study provides proof of concept that viral modification of host cell DNA can lead to autoimmunity.

There is insufficient evidence to support screening of various tick-borne diseases (TBD) related microbes alongside Borrelia in patients suffering from TBD. To evaluate the involvement of multiple microbial immune responses in patients experiencing TBD we utilized enzyme-linked immunosorbent assay. Four hundred and thirty-two human serum samples organized into seven categories followed Centers for Disease Control and Prevention two-tier Lyme disease (LD) diagnosis guidelines and Infectious Disease Society of America guidelines for post-treatment Lyme disease syndrome. All patient categories were tested for their immunoglobulin M (IgM) and G (IgG) responses against 20 microbes associated with TBD. Our findings recognize that microbial infections in patients suffering from TBDs do not follow the one microbe, one disease Germ Theory as 65% of the TBD patients produce immune responses to various microbes. We have established a causal association between TBD patients and TBD associated co-infections and essential opportunistic microbes following Bradford Hill’s criteria. This study indicated an 85% probability that a randomly selected TBD patient will respond to Borrelia and other related TBD microbes rather than to Borrelia alone. A paradigm shift is required in current healthcare policies to diagnose TBD so that patients can get tested and treated even for opportunistic infections.

Preinfection by one parasitic species may facilitate or by contrast hamper the subsequent penetration and/or establishment of other parasites in a host. The biology of interacting species, timing of preinfection, and dosage of subsequent parasite exposure are likely important variables in this multiparasite dynamic infection process. The increased vulnerability to subsequent infection can be an important and often overlooked factor influencing parasite virulence. We investigated how the preinfection by freshwater pearl mussel Margaritifera margaritifera glochidia could influence the success of subsequent infection by the common trematode Diplostomum pseudospathaceum in brown trout Salmo trutta and vice versa whether preinfection by the trematode made fish more susceptible to glochidia infection. The first experiment was repeated twice with different (low and high) exposure doses to initiate the subsequent trematode infection, while in the second experiment we varied the timing of the preinfection with trematodes. The preinfection with glochidia made fish more vulnerable to subsequent infection with trematodes. Since the trematodes penetrate through the gills, we suggest that increased host vulnerability was most likely the result of increased respiration caused by the freshwater pearl mussel glochidia encysted on gills. In turn, brown trout preinfected with trematodes were more vulnerable to the subsequent glochidial infection, but only if they were preinfected shortly before the subsequent infection (20 hr). Fish preinfected with trematodes earlier (2 weeks before the subsequent infection) did not differ in their vulnerability to glochidia. These effects were observed at moderate intensities of infections similar to those that occur in nature. Our study demonstrates how the timing and sequence of exposure to parasitic species can influence infection success in a host–multiparasite system. It indicates that the negative influence of glochidia on host fitness is likely to be underestimated and that this should be taken into consideration when organizing freshwater pearl mussel restoration procedures. peerReviewed

Hosts are typically infected with multiple strains or genotypes of one or several parasite species. These infections can take place simultaneously, but also at different times, i.e. sequentially, when one of the parasites establishes first. Sequential parasite dynamics are common in nature, but also in intensive farming units such as aquaculture. However, knowledge of effects of previous exposures on virulence of current infections in intensive farming is very limited. This is critical as consecutive epidemics and infection history of a host could underlie failures in management practises and medical intervention of diseases. Here, we explored effects of timing of multiple infection on virulence in two common aquaculture parasites, the bacterium Flavobacterium columnare and the fluke Diplostomum pseudospathaceum. We exposed fish hosts first to flukes and then to bacteria in two separate experiments, altering timing between the infections from few hours to several weeks. We found that both short‐term and long‐term difference in timing of the two infections resulted in significant, genotype‐specific decrease in bacterial virulence. Second, we developed a mathematical model, parameterized from our experimental results, to predict the implications of sequential infections for epidemiological progression of the disease, and levels of fish population suppression, in an aquaculture setting. Predictions of the model showed that sequential exposure of hosts can decrease the population‐level impact of the bacterial epidemic, primarily through the increased recovery rate of sequentially infected hosts, thereby substantially protecting the population from the detrimental impact of infection. However, these effects depended on bacterial strain–fluke genotype combinations, suggesting the genetic composition of the parasite populations can greatly influence the degree of host suppression. Overall, these results suggest that host infection history can have significant consequences for the impact of infection at host population level, potentially shaping parasite epidemiology, disease dynamics and evolution of virulence in farming environments. peerReviewed

Viruses depend on cells to replicate and can cause considerable damage to their hosts. However, hosts have developed a plethora of antiviral mechanisms to counterattack or prevent viral replication and to maintain homeostasis. Advantageous features are constantly being selected, affecting host-virus interactions and constituting a harsh race for supremacy in nature. Here, we describe a new antiviral mechanism unveiled by the interaction between a giant virus and its amoebal host. Faustovirus mariensis infects Vermamoeba vermiformis, a free-living amoeba, and induces cell lysis to disseminate into the environment. Once infected, the cells release a soluble factor that triggers the encystment of neighbor cells, preventing their infection. Remarkably, infected cells stimulated by the factor encyst and trap the viruses and viral factories inside cyst walls, which are no longer viable and cannot excyst. This unprecedented mechanism illustrates that a plethora of antiviral strategies remains to be discovered in nature.IMPORTANCE Understanding how viruses of microbes interact with its hosts is not only important from a basic scientific point of view but also for a better comprehension of the evolution of life. Studies involving large and giant viruses have revealed original and outstanding mechanisms concerning virus-host relationships. Here, we report a mechanism developed by Vermamoeba vermiformis, a free-living amoeba, to reduce Faustovirus mariensis dissemination. Once infected, V. vermiformis cells release a factor that induces the encystment of neighbor cells, preventing infection of further cells and/or trapping the viruses and viral factories inside the cyst walls. This phenomenon reinforces the need for more studies regarding large/giant viruses and their hosts.

Leakage of medical residues into the environment can significantly impact natural communities. For example, antibiotic contamination from agriculture and aquaculture can directly influence targeted pathogens, but also other non-targeted taxa of commensals and parasites that regularly co-occur and co-infect the same host. Consequently, antibiotics could significantly alter interspecific interactions and epidemiology of the co-infecting parasite community. We studied how minor environmental concentrations of antibiotic affects the co-infection of two parasites, the bacterium Flavobacterium columnare and the fluke Diplostomum pseudospathaceum , in their fish host. We found that antibiotic in feed, and particularly the minute concentration in water, significantly decreased bacterial virulence and changed the infection success of the flukes. These effects depended on the level of antibiotic resistance of the bacterial strains. Antibiotic, however, did not compensate for the higher virulence of co-infections. Our results demonstrate that even very low environmental concentrations of antibiotic can influence ecology and epidemiology of diseases in co-infection with non-targeted parasites. Leakage of antibiotics into the environment may thus have more complex effects on disease ecology than previously anticipated.

Echoviruses are amongst the most common causative agents of aseptic meningitis worldwide and are particularly devastating in the neonatal population, where they are associated with severe hepatitis, neurological disease, including meningitis and encephalitis, and even death. Here, we identify the neonatal Fc receptor (FcRn) as a pan-echovirus receptor. We show that loss of expression of FcRn or its binding partner beta 2 microglobulin (β2M) renders cells resistant to infection by a panel of echoviruses at the stage of virus attachment, and that a blocking antibody to β2M inhibits echovirus infection in cell lines and in primary human intestinal epithelial cells. We also show that expression of human, but not mouse, FcRn renders nonpermissive human and mouse cells sensitive to echovirus infection and that the extracellular domain of human FcRn directly binds echovirus particles and neutralizes infection. Lastly, we show that neonatal mice expressing human FcRn are more susceptible to echovirus infection by the enteral route. Our findings thus identify FcRn as a pan-echovirus receptor, which may explain the enhanced susceptibility of neonates to echovirus infections.

Coinfections by multiple parasites predominate in the wild. Interactionsbetween parasites can be antagonistic, neutral, or facilitative, and they canhave significant implications for epidemiology, disease dynamics, and evolu-tion of virulence. Coinfections commonly result from sequential exposure ofhosts to different parasites. We argue that the sequential nature of coinfectionsis important for the consequences of infection in both natural and man-madeenvironments. Coinfections accumulate during host lifespan, determining thestructure of the parasite infracommunity. Interactions within the parasite com-munity and their joint effect on the host individual potentially shape evolution ofparasite life-history traits and transmission biology. Overall, sequential coin-fections have the potential to change evolutionary and epidemiological out-comes of host–parasite interactions widely across plant and animal systems. peerReviewed

During lytic herpes simplex virus 1 (HSV-1) infection, the expansion of the viral replication compartments leads to an enrichment of the host chromatin in the peripheral nucleoplasm. We have shown previously that HSV-1 infection induces the formation of channels through the compacted peripheral chromatin. Here, we used three-dimensional confocal and expansion microscopy, soft X-ray tomography, electron microscopy, and random walk simulations to analyze the kinetics of host chromatin redistribution and capsid localization relative to their egress site at the nuclear envelope. Our data demonstrated a gradual increase in chromatin marginalization, and the kinetics of chromatin smoothening around the viral replication compartments correlated with their expansion. We also observed a gradual transfer of capsids to the nuclear envelope. Later in the infection, random walk modeling indicated a gradually faster transport of capsids to the nuclear envelope that correlated with an increase in the interchromatin channels in the nuclear periphery. Our study reveals a stepwise and time-dependent mechanism of herpesvirus nuclear egress, in which progeny viral capsids approach the egress sites at the nuclear envelope via interchromatin spaces. peerReviewed

The study of polymorphic immune genes in host populations is critical for understanding genetic variation in susceptibility to pathogens. Controlled infection experiments are necessary to separate variation in the probability of exposure from genetic variation in susceptibility to infection, but such experiments are rare for wild vertebrate reservoir hosts and their zoonotic pathogens. The bank vole (Myodes glareolus) is an important reservoir host of Borrelia afzelii, a tick-borne spirochete that causes Lyme disease. Bank vole populations are polymorphic for Toll-like receptor 2 (TLR2), an innate immune receptor that recognizes bacterial lipoproteins. To test whether the TLR2 polymorphism influences variation in the susceptibility to infection with B. afzelii, we challenged pathogen-free, lab-born individuals of known TLR2 genotype with B. afzelii-infected ticks. We measured the spirochete load in tissues of the bank voles. The susceptibility to infection with B. afzelii following an infected tick bite was very high (95%) and did not differ between TLR2 genotypes. The TLR2 polymorphism also had no effect on the spirochete abundance in the tissues of the bank voles. Under the laboratory conditions of our study, we did not find that the TLR2 polymorphism in bank voles influenced variation in the susceptibility to B. afzelii infection.

1.Parasitic diseases represent one of the greatest challenges for aquaculture worldwide and there is an increasing emphasis on ecological solutions to prevent infections. One proposed solution is enriched rearing, where traditional stimulus‐poor rearing tanks are equipped with different types of structures to increase habitat complexity. Such spatial enrichment is known to increase survival of fish during parasite epidemics, but the underlying mechanisms are still unclear. 2.We studied whether enriched rearing affected infection of an important fish pathogen Flavobacterium columnare in young Atlantic salmon (Salmo salar) and sea‐migrating brown trout (Salmo trutta). First, we used natural bacterial exposures and multiple fish populations in a common garden experiment to address the role of host genetic background in effects of enriched rearing. Second, fish from standard and enriched rearing were experimentally exposed to controlled bacterial doses in standard and enriched environments in a full factorial design to explore the relative roles of rearing background and environment of exposure on survival of fish. 3.Enriched rearing significantly increased survival of fish during the natural bacterial outbreak. This effect was also fairly consistent and observed in eight of the ten fish populations. In the controlled exposure, fish exposed in enriched environment had higher survival regardless of their rearing background, suggesting a stronger impact of the environment on the disease progression. Additionally, the survival in the enriched environment was the highest among the fish of enriched rearing background, supporting the idea of their higher resistance. 4.Synthesis and applications. Our study suggests that the enhanced survival of fish in enriched rearing is a result of a combined effect of the environment and improved fish condition and, to a lesser degree, from host genetic background. This has important implications for when and how environmental enrichment should be applied. Overall, these results indicate that environmental enrichment has the potential to improve survival of fish during parasitic epidemics and thus reduce use of antibiotics in aquaculture. peerReviewed

Enteroviruses are small RNA viruses that cause diseases with various symptoms ranging from mild to severe. Enterovirus proteins are translated as a single polyprotein, which is cleaved by viral proteases to release capsid and nonstructural proteins. Here, we show that also cellular calpains have a potential role in the processing of the enteroviral polyprotein. Using purified calpains 1 and 2 in an in vitro assay, we show that addition of calpains leads to an increase in the release of VP1 and VP3 capsid proteins from P1 of enterovirus B species, detected by western blotting. This was prevented with a calpain inhibitor and was dependent on optimal calcium concentration, especially for calpain 2. In addition, calpain cleavage at the VP3-VP1 interface was supported by a competition assay using a peptide containing the VP3-VP1 cleavage site. Moreover, a mass spectrometry analysis showed that calpains can cleave this same peptide at the VP3-VP1 interface, the cutting site being two amino acids aside from 3C&rsquo, s cutting site. Furthermore, we show that calpains cannot cleave between P1 and 2A. In conclusion, we show that cellular proteases, calpains, can cleave structural proteins from enterovirus polyprotein in vitro. Whether they assist polyprotein processing in infected cells remains to be shown.

Influence of fish personality on infection rate is almost not studied. In the experiments on the young-of-the-year Oncorhynchus mykiss and cercariae of a trematode Diplostomum pseudospathaceum we tested the hypothesis that infection rate differs between more and less active (“bold” and “shy”) fish. Will individual differences in infection persist upon re-infection? Fish serve as a second intermediate host for this trematode. A positive correlation was found between the results of consecutive infections. Accumulation of parasites with successive infections leads to an aggregated distribution of D. pseudospathaceum among the hosts, affecting individual fitness and polymorphism in fish populations. Persistent individual differences in parasite burden among fish and, as a result, vulnerability for predators confirms the role of parasites as an important factor of natural selection. peerReviewed

Enterovirus B species typically cause a rapid cytolytic infection leading to efficient release of progeny viruses. However, they are also capable of persistent infections in tissues, which are suggested to contribute to severe chronic states such as myocardial inflammation and type 1 diabetes. In order to understand the factors contributing to differential infection strategies, we constructed a chimera by combining the capsid proteins from a fast cytolysis causing echovirus 1 (EV1) with non-structural proteins from Coxsackievirus B5 (CVB5) showing persistent infection in RD cells. The results showed that the chimera behaved similar to the parental EV1 leading to efficient cytolysis in both permissive A549 and semi-permissive RD cells. In contrast to EV1 and chimera, CVB5 replicated slower in permissive cells and showed persistent infection in semi-permissive cells. However, there was no difference in the efficiency of uptake of CVB5 in A549 or RD cells in comparison to the chimera or EV1. CVB5 virus batches constantly contained significant amounts of empty capsids, also in comparison to its close relative CVB3. During successive passaging of batch containing only intact CVB5, increasing amounts of empty and decreasing amounts of infective capsids were produced. Our results demonstrate that the increased amounts of empty particles and lowering amounts of infective particles is dictated by the CVB5 structural proteins leading to slowing down the infection between passages. Furthermore, the key factor for persistent infection is the low amount of infective particles produced, not the high number of empty particles accumulating. peerReviewed

The current methods to study the distribution and dynamics of viral RNA molecules inside infected cells are not ideal, as electron microscopy and immunohistochemistry can only detect mature virions, and quantitative real-time PCR does not reveal localized distribution of RNAs. We demonstrated here the branched DNA in situ hybridization (bDNA ISH) technology to study both the amount and location of the emerging &minus, RNA and +RNA during acute and persistent enterovirus infections. According to our results, the replication of the viral RNA started 2&ndash, 3 h after infection and the translation shortly after at 3&ndash, 4 h post-infection. The replication hotspots with newly emerging &minus, RNA were located quite centrally in the cell, while the +RNA production and most likely virion assembly took place in the periphery of the cell. We also discovered that the pace of replication of &minus, RNA and +RNA strands was almost identical, and &minus, RNA was absent during antiviral treatments. ViewRNA ISH with our custom probes also showed a good signal during acute and persistent enterovirus infections in cell and mouse models. Considering these results, along with the established bDNA FISH protocol modified by us, the effects of antiviral drugs and the emergence of enterovirus RNAs in general can be studied more effectively.

Multiple-strain pathogens often establish mixed infections inside the host that result in competition between strains. In vector-borne pathogens, the competitive ability of strains must be measured in both the vertebrate host and the arthropod vector to understand the outcome of competition. Such studies could reveal the existence of trade-offs in competitive ability between different host types. We used the tick-borne bacteriumBorrelia afzeliito test for competition between strains in the rodent host and the tick vector, and to test for a trade-off in competitive ability between these two host types. Mice were infected via tick bite with either one or two strains, and these mice were subsequently used to create ticks with single or mixed infections. Competition in the rodent host reduced strain-specific host-to-tick transmission and competition in the tick vector reduced the abundance of both strains. The strain that was competitively superior in host-to-tick transmission was competitively inferior with respect to bacterial abundance in the tick. This study suggests that in multiple-strain vector-borne pathogens there are trade-offs in competitive ability between the vertebrate host and the arthropod vector. Such trade-offs could play an important role in the coexistence of pathogen strains.