Search

For ensuring a sustainable production of high-quality fruits, the fruit growing systems needs to adapt for climate change. Different irrigation strategies and their potential advantages for improving apple tree production of cultivar ‘Red Jonaprince’ in conditions of temperate climate were tested. The water regimes of the orchard were analysed through evapotranspiration balance and soil moisture. In apple trees, total growth, flowering intensity, total yield and yield in different fruit size classes including the crop load were evaluated. The results of the study confirm the need for supplementary irrigation of the trees, especially in years with high yields and evapotranspiration. Considering the lower susceptibility to drought of triploid apple tree cultivars, 50 % coverage of evapotranspiration helps to maintain their water needs throughout the vegetation season irrespectively to used irrigation strategy. However, the soil moisture should not fall under 50% of available soil water capacity. Optimal crop load of the cultivar should be with 3,2 m high crowns between 160–180 fruits per tree allowing with plant distance of 3,5×1,2 m production of 70–80 t/ha.

IntroductionCytomegaloviruses (CMVs) extensively reorganize the membrane system of the cell and establish a new structure as large as the cell nucleus called the assembly compartment (AC). Our previous studies on murine CMV (MCMV)-infected fibroblasts indicated that the inner part of the AC contains rearranged early endosomes, recycling endosomes, endosomal recycling compartments and trans-Golgi membrane structures that are extensively tubulated, including the expansion and retention of tubular Rab10 elements. An essential process that initiates Rab10-associated tubulation is cargo sorting and retrieval mediated by SNX27, Retromer, and ESCPE-1 (endosomal SNX-BAR sorting complex for promoting exit 1) complexes.ObjectiveThe aim of this study was to investigate the role of SNX27:Retromer:ESCPE-1 complexes in the biogenesis of pre-AC in MCMV-infected cells and subsequently their role in secondary envelopment and release of infectious virions.ResultsHere we show that SNX27:Retromer:ESCPE1-mediated tubulation is essential for the establishment of a Rab10-decorated subset of membranes within the pre-AC, a function that requires an intact F3 subdomain of the SNX27 FERM domain. Suppression of SNX27-mediated functions resulted in an almost tenfold decrease in the release of infectious virions. However, these effects cannot be directly linked to the contribution of SNX27:Retromer:ESCPE-1-dependent tubulation to the secondary envelopment, as suppression of these components, including the F3-FERM domain, led to a decrease in MCMV protein expression and inhibited the progression of the replication cycle.ConclusionThis study demonstrates a novel and important function of membrane tubulation within the pre-AC associated with the control of viral protein expression.

Understanding how corruption affects the relationship between natural resources and growth has profound implications for sustainable development goals in Sub-Saharan Africa (SSA). This study investigates the nexus between natural resources, economic growth, and corruption in SSA from 1985 to 2022, by employing a Panel Smooth Transition Regression model. Results indicate that corruption distorts natural resources and lowers economic growth. The magnitude of the effect of corruption depends on the type of corruption. The impact of political corruption is more relevant than the others form of corruption. In a low corruption regime, mineral, oil, and forest resources do not affect growth; (b) in a high corruption regime, oil and forest resources hinder growth whereas mineral resources lead to higher economic growth. The analysis highlights the need for policy measures that encourage sustainable economic development and promoting reinforcement of institutional quality.

Abstract Human cytomegalovirus (HCMV) is a widespread pathogen that in immunocompromised hosts can cause life-threatening disease. Studying HCMV-exposed monocyte-derived dendritic cells by single-cell RNA sequencing, we observe that most cells are entered by the virus, whereas less than 30% of them initiate viral gene expression. Increased viral gene expression is associated with activation of the stimulator of interferon genes (STING) that usually induces anti-viral interferon responses, and with the induction of several pro- (RHOB, HSP1A1, DNAJB1) and anti-viral (RNF213, TNFSF10, IFI16) genes. Upon progression of infection, interferon-beta but not interferon-lambda transcription is inhibited. Similarly, interferon-stimulated gene expression is initially induced and then shut off, thus further promoting productive infection. Monocyte-derived dendritic cells are composed of 3 subsets, with one being especially susceptible to HCMV. In conclusion, HCMV permissiveness of monocyte-derived dendritic cells depends on complex interactions between virus sensing, regulation of the interferon response, and viral gene expression.

One of the consequences of the globalisation of food markets is the effort enabling the control of food security and its origin. This might be traced by using different chemical composition analyses. However, for Central Europe, there is a lack of knowledge about the original reference values as well as their heterogeneity among the lands and countries. This study focused on characterizing the mineral profiles of apple tree fruits and comparing these profiles among different districts in Central Europe. The fruits of the apple cultivars ‘Gala’ and ‘Golden Delicious’ originated in the Czech Republic and Poland. The mineral and isotopic content of the apple fruit flesh was analysed using ICP-MS. The data were processed using the ANOVA test and compositely analysed using the PCA and LDA models. The results show relatively high variation in element distribution, particularly 87Sr/86Sr, Mn, Zn, Cu, Ca, P, and B, ranging between 20.6 and67.9% for both cultivars on average. However, their high variability within particular districts complicates the resolution of the LDA model. The reasons are linked to the geomorphological and pedological heterogeneity of the analysed districts as well as the particular sensitivity of the set of chosen primers to agronomic practices and tree performance. For this region, only partial separation among districts could be obtained by P, Ca, and Cu content, as well as the isotopic ratio of 10B/11B. However, the resolution of the geographical discrimination needs to be improved by an enhanced set of primers, the use of more precise analytical techniques for the Sr isotopic ratio, or by multiple chemical analyses. Furthermore, the heterogeneity of the analysed districts could be tackled by more detailed analyses at the level of micro-regions.

Plant-based indicators of water status, such as midday stem water potential (Ψstem) and leaf stomatal conductance (gs), are used to optimise irrigation scheduling in horticultural crops because they integrate the effect of soil and climatic conditions and the internal physiological constraints. The use of these indicators relies on experimentally acquired thresholds that relate the value of the indicator to negative effects on yield. In five irrigation treatments, we monitored yield, fruit size and the courses of Ψstem and gs throughout four consecutive growing seasons. We found that Ψstem was more sensitive to irrigation treatment than gs. Both indicators increased with available soil water content (ASWC) and decreased with evaporative demands of the atmosphere (ETC). On a seasonal basis, crop load had a stronger impact on gs than Ψstem. In summary, our study explored the effect of environmental conditions and crop load on plant-based indicators of tree water status and can be useful for establishing thresholds for irrigation scheduling in apple tree orchards.

Apples are an important source of polyphenolic substances that contribute to the antioxidant activity of the fruit. Presence of apples in the diet significantly contributes to the intake of polyphenols. Apple consumption is associated with prevention of degenerative diseases. Polyphenolic profiles of apples from different regions of the Czech Republic and Poland were analyzed using high-performance liquid chromatography with diode array detection. Four phenolic substances quercitric, epicatechin, phloridzin, and chlorogenic acid were determined in selected apples samples. No statistically significant differences between the regions of the given country were determined for the varieties ‘Golden Delicious’ and ‘Idared’. Chlorogenic acid was the major polyphenol in apples from Poland and its content tended to be higher being 45.4 ± 20.2 µg/g for the ‘Golden Delicious’ and 74.1 ± 2.6 µg/g for the ‘Idared’ varieties, while in the Czech Republic it was 34.2 ± 14.0, respectively 44.3 ± 21.3 µg/g. In contrast, Czech apples contained more epicatechin and quercitrin in these varieties.

Summary: Murine cytomegalovirus (MCMV) infection of macrophages relies on MCMV-encoded chemokine 2 (MCK2), while infection of fibroblasts occurs independently of MCK2. Recently, MCMV infection of both cell types was found to be dependent on cell-expressed neuropilin 1. Using a CRISPR screen, we now identify that MCK2-dependent infection requires MHC class Ia/β-2-microglobulin (B2m) expression. Further analyses reveal that macrophages expressing MHC class Ia haplotypes H-2b and H-2d, but not H-2k, are susceptible to MCK2-dependent infection with MCMV. The importance of MHC class I expression for MCK2-dependent primary infection and viral dissemination is highlighted by experiments with B2m-deficient mice, which lack surface expression of MHC class I molecules. In those mice, intranasally administered MCK2-proficient MCMV mimics infection patterns of MCK2-deficient MCMV in wild-type mice: it does not infect alveolar macrophages and subsequently fails to disseminate into the salivary glands. Together, these data provide essential knowledge for understanding MCMV-induced pathogenesis, tissue targeting, and virus dissemination.

Myelin is formed of proteins of long half-lives. The mechanisms of renewal of such a stable structure are unclear. Here, the authors show that myelin integrity requires continuous myelin synthesis at the inner tongue, contributing to the maintenance of a functional axon-myelin unit.

The total aboveground biomass production, nutritional status, bearing and branching behaviors of the central leader and one year old shoots of young apple trees have been analyzed. The shoots were further characterized according to the length, shoot demography, and the production of terminal and lateral flowers. All the characteristics are described in connection with nitrogen supply and cultivar. Nitrogen represents one of the major macronutrients involved in the growth and development of the fruit trees. The understanding of the effect of nitrogen supply for flower bud formation can be further improved by detailed analyses of tree architecture. While the biomass production was cultivar specific, the trees within particular cultivar were characterized by almost similar growth with respect to the nitrogen supply. Cultivar ´Rubinola´ exhibited similar branching pattern, but higher vigor than ´Topaz´. As a result of higher apical dominance, ´Rubinola´ produced higher proportion of long shoots, but a lower quality of short shoots than ´Topaz´. Consequently, cultivar ´Rubinola´ produced only few terminal flowers on short shoots and lateral flowers dominantly in the distal zone, while ´Topaz´ was characterized by intensive terminal flowering, but the lateral flowers were more abundant in the median zone. Even a lower dose of spring nitrogen improved the flower bud formation on both terminal and lateral positions extending the flowering zone on one-year-old shoots. This further changed the branching and bearing behavior of the apple trees, which particularly allows to optimize their fertilization management. However, this effect appears to be further regulated by mechanism connected with apical dominance.

The presented work is focused on the creation and validation of original tools for the detection of apple fruits with false indication of geographical origin by analysing the mineral composition and isotope ratios using the method (ICP-MS) mass spectrometry with inductively coupled plasma. The obtained profiles of the element composition (P, K, Mg, Fe, Cu a Zn) and isotope ratios (86Sr/87Sr a 10B/11B) of individual samples were statistically processed using principal component analysis (PCA). The achieved results of the analysis of the mineral composition and isotope ratios of Polish and Czech apples show different trends, especially for the elements P, Cu and for the isotope ratio of 10B/11B. However, these trends alone do not yet allow for a sufficient distinction among the geographical origin of apples. These trends were not statistically detected for other analysed elements and for the isotope ratio 86Sr/87Sr. Therefore, in the future we plan to perform a combined multidimensional analysis with other methods, such as the evaluation of selected phenolic metabolites in the fruit flesh.

Replant disease represents an important obstacle in replantation of horticultural crops. There is a lack of knowledge about the influence of replant disease on nutritional status of fruit trees. This study was focused on comparison of the growth and production characteristics and nutritional status of apple trees ‘Gala Brookfield’ among different location according to replant problems and used irrigation strategy. In this trial, growth intensity, flowering intensity, yield, fruit quality, and leaf mineral content of N, P, K, Mg, and Ca were assessed. We found that the apple trees planted in replant soils grows 30 % less and bears 16 % less fruit than those in fresh soils. They are more susceptible to water deficit, documented by lower yield and mean fruit weight. Lower yield efficiency and fruit quality of the vigorously growing apple trees in fresh soil shows to the need for establishment of better light conditions in orchard. The nitrogen content was more related to the proportion between generative and vegetative growth, whereas the phosphorus, potassium, magnesium, and calcium were more site specific. Generally, apple trees are prone to accumulate more magnesium in leaves on fresh soils. The use of regulated deficit irrigation with intense fruit thinning may require corrections of the fertilization management with P, Mg, and Ca.

ABSTRACT Morphogenesis of herpesvirus particles is highly conserved; however, the capsid assembly and genome packaging of human cytomegalovirus (HCMV) exhibit unique features. Examples of these include the essential role of the small capsid protein (SCP) and the existence of the β-herpesvirus-specific capsid-associated protein pp150. SCP and pp150, as well as the UL77 and UL93 proteins, are important capsid constituents, yet their precise mechanism of action is elusive. Here, we analyzed how deletion of the open reading frames (ORFs) encoding pUL77, pUL93, pp150, or SCP affects the protein composition of nuclear capsids. This was achieved by generating HCMV genomes lacking the respective genes, combined with a highly efficient transfection technique that allowed us to directly analyze these mutants in transfected cells. While no obvious effects were observed when pUL77, pUL93, or pp150 was missing, the absence of SCP impeded capsid assembly due to strongly reduced amounts of major capsid protein (MCP). Vice versa, when MCP was lacking, SCP became undetectable, indicating a mutual dependence of SCP and MCP for establishing appropriate protein levels. The SCP domain mediating stable MCP levels could be narrowed down to a C-terminal helix known to convey MCP binding. Interestingly, an SCP-EGFP (enhanced green fluorescent protein) fusion protein which also impaired the production of infectious progeny acted in a different manner, as capsid assembly was not abolished; however, SCP-EGFP-harboring capsids were devoid of DNA and trapped in paracrystalline nuclear structures. These results indicate that SCP is essential in HCMV because of its impact on MCP levels and reveal SCP as a potential target for antiviral inhibitors. IMPORTANCE Human cytomegalovirus (HCMV) is a ubiquitous pathogen causing life-threatening disease in immunocompromised individuals. Virus-specific processes such as capsid assembly and genome packaging can be exploited to design new antiviral strategies. Here, we report on a novel function of the HCMV small capsid protein (SCP), namely, ensuring stable levels of major capsid protein (MCP), thereby governing capsid assembly. Furthermore, we discovered a mutual dependence of the small and major capsid proteins to guarantee appropriate levels of the other respective protein and were able to pin down the SCP domain responsible for this effect to a region previously shown to mediate binding to the major capsid protein. In summary, our data contribute to the understanding of how SCP plays an essential part in the HCMV infection cycle. Moreover, disrupting the SCP-MCP interface may provide a starting point for the development of novel antiviral drugs.

13C discrimination (Δ13C) has been used in research as an indicator of water availability in crops; however, few data are available concerning fruit trees. The aim of this study was to examine the effect of irrigation on the Δ13C values of apple leaves. We assumed that Δ13C would increase with irrigation intensity. The Δ13C of apple trees (Malus domestica) cv. ‘Red Jonaprince’ was determined in the years 2019–2022. Leaf samples were collected in spring, in June, at the beginning of the irrigation campaign, and in autumn, in September, following the harvest. The irrigation doses were applied to replenish the water consumption, 0% (ET0), 50% (ET50), 75% (ET75), and 100% (ET100), of the calculated evapotranspiration (ET) levels. In November, the leaves collected from different positions on the shoots were sampled, assuming the Δ13C signature would reflect the changes occurring in the water supply during their growth. The irrigation rates had a significant effect on the Δ13C of the leaves when the data for the spring and summer months were pooled. On average, Δ13C increased from 20.77‰ and 20.73‰ for ET0 and ET50, respectively, to 20.80‰ and 20.95‰ for ET75 and ET100, respectively. When the data obtained for the spring and summer months were analysed separately, the effect of irrigation was weak (p < 0.043). The Δ13C value was always higher for treatment ET100 than treatment ET0, for individual experimental years and terms; however, the differences were minor and mostly insignificant. The leaf position had a strong significant effect on Δ13C; the values gradually decreased from the leaves growing from two-years-old branches (22.50‰) to the youngest leaves growing at the top of the current year’s shoots (21.07‰). This order was similar for all the experimental years. The results of the experiment suggest that 13C discrimination in apples is affected not only by water availability during growth, but also by the use of C absorbed in previous years.

Invaginations of the nuclear membrane occur in different shapes, sizes, and compositions. Part of these pleiomorphic invaginations make up the nucleoplasmic reticulum (NR), while others are merely nuclear folds. We define the NR as tubular invaginations consisting of either both the inner and outer nuclear membrane, or only the inner nuclear membrane. Specifically, invaginations of both the inner and outer nuclear membrane are also called type II NR, while those of only the inner nuclear membrane are defined as type I NR. The formation and structure of the NR is determined by proteins associated to the nuclear membrane, which induce a high membrane curvature leading to tubular invaginations. Here we review and discuss the current knowledge of nuclear invaginations and the NR in particular. An increase in tubular invaginations of the nuclear envelope is associated with several pathologies, such as laminopathies, cancer, (reversible) heart failure, and Alzheimer’s disease. Furthermore, viruses can induce both type I and II NR. In laminopathies, the amount of A-type lamins throughout the nucleus is generally decreased or the organization of lamins or lamin-associated proteins is disturbed. Also, lamin overexpression or modulation of lamin farnesylation status impacts NR formation, confirming the importance of lamin processing in NR formation. Virus infections reorganize the nuclear lamina via (de)phosphorylation of lamins, leading to an uneven thickness of the nuclear lamina and in turn lobulation of the nuclear membrane and the formation of invaginations of the inner nuclear membrane. Since most studies on the NR have been performed with cell cultures, we present additional proof for the existence of these structures in vivo, focusing on a variety of differentiated cardiovascular and hematopoietic cells. Furthermore, we substantiate the knowledge of the lamin composition of the NR by super-resolution images of the lamin A/C and B1 organization. Finally, we further highlight the essential role of lamins in NR formation by demonstrating that (over)expression of lamins can induce aberrant NR structures.

Oligodendrocytes myelinate and metabolically support axons. The role of myelination in information processing beyond regulation of conduction velocity is unclear. Here, the authors show that myelination contributes to sustained stimulus perception in the auditory cortex, shaping neuronal responses.

Chemical (benzyladenine - BA, gibberelins - GA4/7), mechanical (leaf pinching), and combined treatments were tested in 2015 and 2016 to increase the number and length of sylleptic side shoots (feathers) on apple (´Rubinola´, ´Topaz´/M9) and plum (´Elena´, ´Tophit´/St. J. A) trees. The numbers of feathers of three different lengths (1-10, 10-30, and >30 cm) were measured as the main indicator of quality along with other important morphological characteristics. The numbers of all feathers on both apple cultivars were the highest when BA was applied four times. However, mainly an increase in the numbers of shorter feathers was observed in both years. The efficiency of leaf pinching of apple trees was low. The numbers of all feathers and feathers 1-10 and 10-30 cm in length were the highest in both plum cultivars when BA was applied four times in both years. In contrast to apples, the induction of feathers longer than 30 cm improved in plum cultivars when pinching was used. Thus this approach can be also effective in organic production.

In this contribution we introduce new multilayer (bilayer and trilayer) resist systems for the generation of nanostructures with high aspect ratios of up to 14:1 on 4-in. full wafer scale. The bilayer stack consists of a bottom resist layer (lift off polymer LOR1A) and an UV-curable top resist layer (UV-NIL resist mr-NIL210 200 nm). The top resist is structured by UV-nanoimprint lithography with a soft polydimethysiloxane (PDMS) stamp (soft UV-NIL). After removal of the residual layer a wet chemical development is performed to achieve an isotropic undercut underneath the nanostructures in the top layer. This undercut is mandatory in order to perform a reliable and precise lift-off. The bilayer system is applicable on both silicon and fused silica. For a higher variety and combination of different resists, a trilayer system is investigated. A layer stack with new materials for bottom and top layer is presented. An intermediate layer made of silicon oxide by low temperature ICP-PECVD is added between a tailor-made top resist (mr-NIL213FC 200 nm) and an organic transfer layer (UL1). The intermediate layer serves as hard mask in order to etch the bottom layer isotropically utilizing a plasma etch process and thus replacing the wet-chemical development step. Subsequently, a thin metal layer is deposited onto the structured resist stack by electron beam evaporation. After lift-off, a nanostructured metal mask remains on the substrate providing a high selectivity during the following plasma etch step. A cryogenic ICPRIE etch process creates high aspect ratio nanostructures within the substrate. An aspect ratio of 14:1 was achieved.

Oligodendrocytes facilitate rapid impulse propagation along the axons they myelinate and support their long-term integrity. However, the functional relevance of many myelin proteins has remained unknown. Here, we find that expression of the tetraspan-transmembrane protein CMTM5 (chemokine-like factor-like MARVEL-transmembrane domain containing protein 5) is highly enriched in oligodendrocytes and central nervous system (CNS) myelin. Genetic disruption of the Cmtm5 gene in oligodendrocytes of mice does not impair the development or ultrastructure of CNS myelin. However, oligodendroglial Cmtm5 deficiency causes an early-onset progressive axonopathy, which we also observe in global and tamoxifen-induced oligodendroglial Cmtm5 mutants. Presence of the WldS mutation ameliorates the axonopathy, implying a Wallerian degeneration-like pathomechanism. These results indicate that CMTM5 is involved in the function of oligodendrocytes to maintain axonal integrity rather than myelin biogenesis.

In a three-year experiment (2019–2021), the roots of 7-year-old apple trees (Malus domestica cv. ‘Red Jonaprince’) grown under drip irrigation were studied. The aim of the study was to determine the effect of irrigation on root density at different depths and distances from the trunk. The working hypothesis assumed that irrigation significantly affects the total length of apple roots. The irrigation treatments corresponding to the calculated water evapotranspiration (ET100), 50% of the calculated ET (ET50), a control (ET0, no irrigation, under rainfed conditions), and a treatment using double-drip lines (2Drops) were monitored. Soil cores were collected in spring and autumn. The total length of the roots (TRLt) and the length of new vital roots (TRLv) to a depth of 80 cm were evaluated. The effects of treatments were mostly insignificant for the TRLt; only in the dry season in 2019 were the TRLt values of the irrigated treatments (ET50 and ET100) significantly higher, 18.67 km·m−2 and 17.45 km·m−2, in comparison to 11.16 km·m−2 for the ET0, at a 10 cm distance from the tree trunk. The irrigation treatments had a statistically significant effect on the TRLv values near the trunk in 2019 and 2020, while in autumn 2020 and 2021, irrigation significantly affected the TRLv at greater distances from the tree trunk. In summary, the irrigation treatments mostly had no significant effect on the total root length. However, an effect of irrigation on the root length of new vital roots was observed at certain sampling dates and distances from the trunk.

Gammaherpesviruses, such as Epstein-Barr virus and Kaposi’s sarcoma-associated herpesvirus, are important human pathogens involved in lymphoproliferative disorders and tumorigenesis. Herpesvirus infections are characterized by a biphasic cycle comprised of an acute phase with lytic replication and a latent state. Murine gammaherpesvirus 68 (MHV-68) is a well-established model for the study of lytic and latent life cycles in the mouse. We investigated the interplay between the type I interferon (IFN)-mediated innate immune response and MHV-68 latency using sensitive bioluminescent reporter mice. Adoptive transfer of latently infected splenocytes into type I IFN receptor-deficient mice led to a loss of latency control. This was revealed by robust viral propagation and dissemination of MHV-68, which coincided with type I IFN reporter induction. Despite MHV-68 latency control by IFN, the continuous low-level cell-to-cell transmission of MHV-68 was detected in the presence of IFN signaling, indicating that IFN cannot fully prevent viral dissemination during latency. Moreover, impaired type I IFN signaling in latently infected splenocytes increased the risk of virus reactivation, demonstrating that IFN directly controls MHV-68 latency in infected cells. Overall, our data show that locally constrained type I IFN responses control the cellular reservoir of latency, as well as the distribution of latent infection to potential new target cells.

Herpesviruses cause severe diseases particularly in immunocompromised patients. Both genome packaging and release from the capsid require a unique portal channel occupying one of the 12 capsid vertices. Here, we report the 2.6 Å crystal structure of the pentameric pORF19 of the γ-herpesvirus Kaposi's sarcoma-associated herpesvirus (KSHV) resembling the portal cap that seals this portal channel. We also present the structure of its β-herpesviral ortholog, revealing a striking structural similarity to its α- and γ-herpesviral counterparts despite apparent differences in capsid association. We demonstrate pORF19 pentamer formation in solution and provide insights into how pentamerization is triggered in infected cells. Mutagenesis in its lateral interfaces blocked pORF19 pentamerization and severely affected KSHV capsid assembly and production of infectious progeny. Our results pave the way to better understand the role of pORF19 in capsid assembly and identify a potential novel drug target for the treatment of herpesvirus-induced diseases.

The promising branching agent cyclanilide [1-(2,4-dichlorophenylaminocarbonyl)-cyclopropane carboxylic acid] is not permitted in the EU and the low vigour rootstocks preferred by growers make the branching of young sweet cherry (Prunus avium) trees difficult in the temperate zone production regions. To increase the number and length of new branches ("feathers") on such trees, chemical (6-benzyladenine [BA], gibberellic acid [GA4/7]), mechanical (leaf pinching, summer notching) and various combined treatments were tested from 2015-2018. The numbers of feathers divided into three different lengths (1-10 cm, 10-30 cm and > 30 cm) were assessed as the main indicator of quality, supplemented with other morphological characteristics. Summer notching induced an insufficient number of feathers and led to some negative effects on the trees. The number of feathers from BA or BA with GA4/7 treatment also was insufficient. The effect of pinching alone was variable. The number of all feathers generally was highest when pinching was combined with BA, regardless of the year or cultivar.

During 1999-2016, the performance of 'Red Bartlett', 'Conference' and 'Beurré Alexander Lucas' grafted on four pear (Pyrus) rootstocks OH × F 69, OH × F 87, OH × F 230, OH × F 333 and on quince (Cydonia) rootstock BA 29 was evaluated at the RBIP Holovousy Ltd. For each combination, trunk cross-section area, cumulative yield, yield efficiency and mean fruit weight were assessed. The growth vigour of the scion cultivars on OH × F rootstocks was similar or slightly higher comparing to the growth on BA 29. Throughout the years, the growth vigor and productivity of the rootstock combinations may change. Less vigorous combinations of rootstock/scion were linked with higher yields in young trees, but usually with just moderate yields in later seasons. By contrast, combinations with higher growth vigour had usually higher yields in the later seasons. This balance is genotype specific, where aside of rootstock an important role is played by the scion cultivar vigour. The bearing precocity may be influenced by the cultivar as well. The results can vary with different climatic conditions and orchard management.

ABSTRACT The plasma membrane (PM) must be overcome by viruses during entry and release. Furthermore, the PM represents the cellular communication compartment and the immune system interface. Hence, viruses have evolved sophisticated strategies to remodel the PM, for instance to avoid immune sensing and clearance of infected cells. We performed a comprehensive analysis of cell surface dysregulation by two human-pathogenic viruses, human cytomegalovirus (HCMV) and human immunodeficiency virus type 1 (HIV-1), in primary macrophages, which are classical antigen-presenting cells and orchestrators of the immune system. Scanning ion conductance microscopy revealed a loss of roughness and an overall smooth phenotype of HCMV-infected macrophages, in contrast to HIV-1 infection. This phenotype was also evident on the molecular level. When we screened for cell surface receptors modulated by HCMV, 42 of 332 receptors tested were up- or downregulated, whereas HIV-1 affected only 7 receptors. In particular CD164, CD84, and CD180 were targeted by HCMV. Mechanistically, HCMV induced transcriptional silencing of these receptors in an interferon (IFN)-independent manner, and expression was reduced not only by lab-adapted HCMV but also by clinical HCMV isolates. Altogether, our plasma membrane profiling of human macrophages provides clues to understand how viruses evade the immune system and identified novel cell surface receptors targeted by HCMV. IMPORTANCE The PM is a key component that viruses have to cope with. It is a barrier for infection and egress and is critically involved in antiviral immune signaling. We hence asked the question how two immunomodulatory viruses, HIV-1 and HCMV, dysregulate this compartment in infected macrophages, relevant in vivo targets of both viruses. We employed a contact-free microscopic technique to image the PM of infected cells and performed a phenotypic flow cytometry-based screen to identify receptor modulations on a molecular level. Our results show that HIV-1 and HCMV differentially manipulate the PM of macrophages. While HIV-1-mediated changes are relatively subtle, HCMV induces major alterations of the PM. We identify novel immune receptors manipulated by HCMV and define mechanisms of how HCMV interferes with receptor expression. Altogether, our study reveals differential strategies of how two human-pathogenic viruses manipulate infected cells and identifies potential novel pathways of HCMV immune evasion.

[This corrects the article DOI: 10.1371/journal.ppat.1008560.].

Viral infections are controlled, and very often cleared, by activated T lymphocytes. The inducible co-stimulator (ICOS) mediates its functions by binding to its ligand ICOSL, enhancing T-cell activation and optimal germinal center (GC) formation. Here, we show that ICOSL is heavily downmodulated during infection of antigen-presenting cells by different herpesviruses. We found that, in murine cytomegalovirus (MCMV), the immunoevasin m138/fcr-1 physically interacts with ICOSL, impeding its maturation and promoting its lysosomal degradation. This viral protein counteracts T-cell responses, in an ICOS-dependent manner, and limits virus control during the acute MCMV infection. Additionally, we report that blockade of ICOSL in MCMV-infected mice critically regulates the production of MCMV-specific antibodies due to a reduction of T follicular helper and GC B cells. Altogether, these findings reveal a novel mechanism evolved by MCMV to counteract adaptive immune surveillance, and demonstrates a role of the ICOS:ICOSL axis in the host defense against herpesviruses.

Human cytomegalovirus (HCMV) causes serious complications to immune compromised hosts. Dendritic cells (iDCgB) expressing granulocyte-macrophage colony-stimulating factor, interferon-alpha and HCMV-gB were developed to promote de novo antiviral adaptive responses. Mice reconstituted with a human immune system (HIS) were immunized with iDCgB and challenged with HCMV, resulting into 93% protection. Immunization stimulated the expansion of functional effector memory CD8+ and CD4+ T cells recognizing gB. Machine learning analyses confirmed bone marrow T/CD4+, liver B/IgA+ and spleen B/IgG+ cells as predictive biomarkers of immunization (≈87% accuracy). CD8+ and CD4+ T cell responses against gB were validated. Splenic gB-binding IgM-/IgG+ B cells were sorted and analyzed at a single cell level. iDCgB immunizations elicited human-like IgG responses with a broad usage of various IgG heavy chain V gene segments harboring variable levels of somatic hypermutation. From this search, two gB-binding human monoclonal IgGs were generated that neutralized HCMV infection in vitro. Passive immunization with these antibodies provided proof-of-concept evidence of protection against HCMV infection. This HIS/HCMV in vivo model system supported the validation of novel active and passive immune therapies for future clinical translation.

The research was conducted in the fruit nursery of the Research and Breeding Institute of Pomology (RBIP) Holovousy, Czech Republic in 2015, 2016 and 2017. The efficacy and potential phytotoxic effect of soil herbicides were assessed in spring‑planted and summer‑budded rootstocks M9, MA, Gisela 5 and St. Julien A in seven treatments with the following herbicides: linuron, metazachlor, metamitron, chlorotoluron, dimethenamid‑P, pendimethalin, and combination of pendimethalin and dimethenamid‑P in one product. The effects of the surfactant and simulated rainfall in combination with the pendimethalin + dimethenamid‑P treatment were also assessed. Herbicides were applied twice a season, i.e. in spring after planting and in autumn to control weeds emerging during the season. The efficacy of the herbicide combining both pendimethalin and dimethenamid‑P was the highest in all three seasons. Metazachlor, pendimethalin and dimethenamid‑P provided satisfactory efficacy with adequate soil moisture. Pre‑emergent efficacy of metamitron, linuron and chlorotoluron was not satisfactory, especially in dryer conditions. The synergistic effect of the surfactant and rainfall on the efficacy of pendimethalin in combination with dimethenamid‑P was proved in the dry year only. None of the tested herbicides had a negative effect either on the growth and health status of planted rootstocks or subsequently on budded plants in the following year.

In the present study we investigated the possibility to use air ozone in fruit tree storage. The variables were: rate of ozonization, length of exposure to ozonized air and storage temperature. We assessed the impact of ozone on the vitality of the trees or bench grafted rootstocks, any changes appearing after storage and the feasibility of this technique under real conditions. Supplementary research was done to test the impact of ozonization on the potential occurrence of microorganisms and pests on trees. Results showed that trees can withstand relatively high rates of ozonization except for peach which is the most sensitive fruit; all the ozonization treatments impaired peach vitality. Other species withstand three months of storage in 1 °C and one hour of exposure to ozone in fortnight intervals without a significant impact on their vitality. A higher (5 °C) storage temperature with or without ozonization decreased the vitality of trees. Although it has been proved that ozonization can suppress damage‑causing microorganisms (bacteria and fungi) on trees, the fortnight frequency of ozonization did not prevent mould to develop. Results also indicated the potential effect of ozonization on the disinfestation of plant material from overwintering pests. The main operation restrictions of ozone exposure is the ability of ozone to decompose some materials and that it is harmful to human health.

Abstract Human cytomegalovirus (HCMV) is a major human pathogen, causing serious diseases in immunocompromised populations and congenially infected neonates. One of the main immune cells acting against the virus are Natural Killer (NK) cells. Killing by NK cells is mediated by a small family of activating receptors such as NKp30 that interact with the cellular ligand B7-H6. The outcome of B7-H6-NKp30 interaction was, so far, mainly studied with regard to NK recognition and killing of tumors. Here, we demonstrated that the expression of B7-H6 is upregulated following HCMV infection and that HCMV uses two of its genes: US18 and US20, to interfere with B7-H6 surface expression, in a mechanism involving endosomal degradation, in order to evade NK cell recognition.

Quiescent long-term hematopoietic stem cells (LT-HSCs) are efficiently activated by type I interferon (IFN-I). However, this effect remains poorly investigated in the context of IFN-I-inducing virus infections. Here we report that both vesicular stomatitis virus (VSV) and murine cytomegalovirus (MCMV) infection induce LT-HSC activation that substantially differs from the effects triggered upon injection of synthetic IFN-I-inducing agents. In both infections, inflammatory responses had to exceed local thresholds within the bone marrow to confer LT-HSC cell cycle entry, and IFN-I receptor triggering was not critical for this activation. After resolution of acute MCMV infection, LT-HSCs returned to phenotypic quiescence. However, non-acute MCMV infection induced a sustained inflammatory milieu within the bone marrow that was associated with long-lasting impairment of LT-HSC function. In conclusion, our results show that systemic virus infections fundamentally affect LT-HSCs and that also non-acute inflammatory stimuli in bone marrow donors can affect the reconstitution potential of bone marrow transplants.

In most temperate fruit trees, fruits are located on one-year old shoots. In Prunus species, flowers and fruits are born in axillary position along those shoots. The axillary bud fate and branching patterns are thus key components of the cultivar potential fruit production. The objective of this study was to analyze the branching and bearing behaviors of 1-year-old shoots of apricot cultivars and clones genetically closely related. Shoot structures were analyzed in terms of axillary bud fates using hidden semi-Markov chains and compared depending on the genotype, year and shoot length. The shoots were composed of three successive zones containing latent buds (basal zone), central flower buds (median zone) and vegetative buds (distal zone), respectively. The last two zones contained few associated flower buds. The zones length (in number of metamers) and occurrence strongly depended on shoot development in the two successive years. With decrease in the number of metamers per shoot, the last two zones become shorter or may not develop. While the number of metamers of the basal and distal zones and the number of associated flower buds correlated to the number of metamers of the shoot, the number of metamers of the median zone and the transition probability from the median to the distal zone were cultivar specific.

Diagnostic of the nutritional status in fruit trees can be estimated using different models. However, these models usually do not fully match the character of the perennial crops and their accuracy needs to be improved. This study aimed at deepening the understanding of the aspects controlling mineral content of macro- and micronutrients and fruit quality of the apple trees with respect to the crop load and vegetative shoot growth. The focus was given on description of the nutrient content variability in time and space, its relation to important physiological events, and conditions altering their gradient in leaf and fruit. The evaluation was carried out with six years old ’Golden Delicious’ apple trees on M9, planted in an experimental orchard. Flowering rate, total yield, shoot growth intensity, mean fruit weight and quality along with mineral content of leaves, and fruit in different development stages were analysed according to intensity of natural or regulated flowering and fruit set, respectively. We found that aside of macroelements, iron, and manganese depended on the crop load. For nitrogen, phosphorus, and potassium content, their gradient increased up to 10–25% within the sufficiency range as the trees tried to maintain certain level of these nutrients in leaves and fruit simultaneously during deficit. In contrary, the higher content of Mg, Fe, and Mn related to the avoidance of deficit through up to 20–30% higher uptake in trees with higher crop load. Phosphorus, potassium, and calcium content in short fruiting shoots was more stage dependent, showing relation with the timing of cessation of particular meristematic growth activity. If no intense deficit occurred, the trees kept unrestricted initial investment of nutrients in the fruit during cell division stage whatever the crop load was. Their content at the later development stages appeared to be more dependent upon the consumption by particular sink. These findings allowed to improve the interpretation of the fruit trees nutrition status due to adaptation of the reference nutrient content in low cropping trees by a reduction of 5–10% N and P, 10–15% Ca, as well as by an increase of 15–20% K content in leaves at the stage with fruit size of 20 mm, and without change in P content at the stage of about 70% of final fruit size.

The research was focused on the comparison of two pruning methods (winter pruning and winter + summer pruning) applied to apple trees trained to a standard and a modified slender spindle. The orchard of ‘Topaz’ trees, grafted on rootstock M 9, was planted in spring 2011. In the years 2013 – 2016, trunk cross sectional area (TCSA), crown volume, cumulative yields, yield efficiency, relative proportion of fruit size classes, number of cuts and dry matter of pruned wood were analyzed. The total growth intensity of the trees, measured by TCSA, was similar among the treatments. The trees of modified slender spindle had lower mean crown volume (2.751 – 2.765 m3) comparing to slender spindle with additional summer pruning (3.355 m3) and proved to better control the tree size. The modified slender spindle brought similar or slightly lower cumulative yields, but significantly higher proportion of good sized fruits (in categories above ř 70 mm) comparing to slender spindle. The pruning of modified spindle brings generally higher number of cuts removing a higher amount of woody biomass in comparison to slender spindle, regardless if combined with summer pruning. The additional summer pruning brought no beneficial effect in reduction of growth, fruit production and fruit size of the modified slender spindle.

The rapid activation of pattern recognition receptor (PRR)-mediated type I interferon (IFN) signaling is crucial for the host response to infection. In turn, human cytomegalovirus (HCMV) must evade this potent response to establish life-long infection. Here, we reveal that the HCMV tegument protein UL35 antagonizes the activation of type I IFN transcription downstream of the DNA and RNA sensors cGAS and RIG-I, respectively. We show that ectopic expression of UL35 diminishes the type I IFN response, while infection with a recombinant HCMV lacking UL35 induces an elevated type I IFN response compared to wildtype HCMV. With a series of luciferase reporter assays and the analysis of signaling kinetics upon HCMV infection, we observed that UL35 downmodulates PRR signaling at the level of the key signaling factor TANK-binding kinase 1 (TBK1). Finally, we demonstrate that UL35 and TBK1 co-immunoprecipitate when co-expressed in HEK293T cells. In addition, we show that a previously reported cellular binding partner of UL35, O-GlcNAc transferase (OGT), post-translationally GlcNAcylates UL35, but that this modification is not required for the antagonizing effect of UL35 on PRR signaling. In summary, we have identified UL35 as the first HCMV protein to antagonize the type I IFN response at the level of TBK1, thereby enriching our understanding of how this important herpesvirus escapes host immune responses.

Human cytomegalovirus (HCMV) latency is typically harmless but reactivation can be largely detrimental to immune compromised hosts. We modeled latency and reactivation using a traceable HCMV laboratory strain expressing the Gaussia luciferase reporter gene (HCMV/GLuc) in order to interrogate the viral modulatory effects on the human adaptive immunity. Humanized mice with long-term (more than 17 weeks) steady human T and B cell immune reconstitutions were infected with HCMV/GLuc and 7 weeks later were further treated with granulocyte-colony stimulating factor (G-CSF) to induce viral reactivations. Whole body bio-luminescence imaging analyses clearly differentiated mice with latent viral infections vs. reactivations. Foci of vigorous viral reactivations were detectable in liver, lymph nodes and salivary glands. The number of viral genome copies in various tissues increased upon reactivations and were detectable in sorted human CD14+, CD169+, and CD34+ cells. Compared with non-infected controls, mice after infections and reactivations showed higher thymopoiesis, systemic expansion of Th, CTL, Treg, and Tfh cells and functional antiviral T cell responses. Latent infections promoted vast development of memory CD4+ T cells while reactivations triggered a shift toward effector T cells expressing PD-1. Further, reactivations prompted a marked development of B cells, maturation of IgG+ plasma cells, and HCMV-specific antibody responses. Multivariate statistical methods were employed using T and B cell immune phenotypic profiles obtained with cells from several tissues of individual mice. The data was used to identify combinations of markers that could predict an HCMV infection vs. reactivation status. In spleen, but not in lymph nodes, higher frequencies of effector CD4+ T cells expressing PD-1 were among the factors most suited to distinguish HCMV reactivations from infections. These results suggest a shift from a T cell dominated immune response during latent infections toward an exhausted T cell phenotype and active humoral immune response upon reactivations. In sum, this novel in vivo humanized model combined with advanced analyses highlights a dynamic system clearly specifying the immunological spatial signatures of HCMV latency and reactivations. These signatures can be merged as predictive biomarker clusters that can be applied in the clinical translation of new therapies for the control of HCMV reactivation.

Human cytomegalovirus (CMV) and mouse cytomegalovirus (MCMV) infection share many characteristics. Therefore infection of mice with MCMV is an important tool to understand immune responses and to design vaccines and therapies for patients at the risk of severe CMV disease. In this study, we investigated the immune response in the lungs following acute infection with MCMV. We used multi-color fluorescence microscopy to visualize single infected and immune cells in nodular inflammatory foci (NIFs) that formed around infected cells in the lungs. These NIFs consisted mainly of myeloid cells, T cells, and some NK cells. We found that the formation of NIFs was essential to reduce the number of infected cells in the lung tissue, showing that NIFs were sites of infection as well as sites of immune response. Comparing mice deficient for several leukocyte subsets, we identified T cells to be of prime importance for restricting MCMV infection in the lung. Moreover, T cells had to be present in NIFs in high numbers, and CD4 as well as CD8 T cells supported each other to efficiently control virus spread. Additionally, we investigated the effects of perforin and interferon-gamma (IFNγ) on the virus infection and found important roles for both mechanisms. NK cells and T cells were the major source for IFNγ in the lung and in in vitro assays we found that IFNγ had the potential to reduce plaque growth on primary lung stromal cells. Notably, the T cell-mediated control was shown to be perforin-independent but IFNγ-dependent. In total, this study systematically identifies crucial antiviral factors present in lung NIFs for early containment of a local MCMV infection at the single cell level.

The receptor tyrosine phosphatase CD45 is expressed on the surface of almost all cells of hematopoietic origin. CD45 functions are central to the development of T cells and determine the threshold at which T and B lymphocytes can become activated. Given this pivotal role of CD45 in the immune system, it is probably not surprising that viruses interfere with the activity of CD45 in lymphocytes to dampen the immune response and that they also utilize this molecule to accomplish their replication cycle. Here we report what is known about the interaction of viral proteins with CD45. Moreover, we debate putative interactions of viruses with CD45 in myeloid cells and the resulting consequences—subjects that remain to be investigated. Finally, we summarize the evidence that pathogens were the driving force for the evolution of CD45.

Human cytomegalovirus (HCMV) has a large 240 kb genome that may encode more than 700 gene products with many of them remaining uncharacterized. Mutagenesis of bacterial artificial chromosome (BAC)-cloned CMV genomes has greatly facilitated the analysis of viral gene functions. However, the roles of essential proteins often remain particularly elusive because their investigation requires the cumbersome establishment of suitable complementation systems. Here, we show that HCMV genomes can be introduced into cells with unprecedented efficiency by applying a transfection protocol based on replication-defective, inactivated adenovirus particles (adenofection). Upon adenofection of several permissive cell types with HCMV genomes carrying mutations in essential genes, transfection rates of up to 60% were observed and viral proteins of all kinetic classes were found expressed. This enabled further analyses of the transfected cells by standard biochemical techniques. Remarkably, HCMV genomes lacking elements essential for viral DNA replication, such as the lytic origin of replication, still expressed several late proteins. In conclusion, adenofection allows the study of essential HCMV genes directly in BAC-transfected cells without the need for sophisticated complementation strategies.

Development of an effective vaccine against human cytomegalovirus (HCMV) is a need of utmost medical importance. Generally, it is believed that a live attenuated vaccine would best provide protective immunity against this tenacious pathogen. Here, we propose a strategy for an HCMV vaccine that aims at the simultaneous activation of innate and adaptive immune responses. An HCMV strain expressing the host ligand ULBP2 for the NKG2D receptor was found to be susceptible to control by natural killer (NK) cells, and preserved the ability to stimulate HCMV-specific T cells. Infection with the ULBP2-expressing HCMV strain caused diminished cell surface levels of MHC class I molecules. While expression of the NKG2D ligand increased the cytolytic activity of NK cells, NKG2D engagement in CD8+ T cells provided co-stimulation and compensated for lower MHC class I expression. Altogether, our data indicate that triggering of both arms of the immune system is a promising approach applicable to the generation of a live attenuated HCMV vaccine.

The receptor-like protein tyrosine phosphatase CD45 is expressed on the surface of cells of hematopoietic origin and has a pivotal role for the function of these cells in the immune response. Here we report that following infection of macrophages with mouse cytomegalovirus (MCMV) the cell surface expression of CD45 is drastically diminished. Screening of a set of MCMV deletion mutants allowed us to identify the viral gene m42 of being responsible for CD45 down-modulation. Moreover, expression of m42 independent of viral infection upon retroviral transduction of the RAW264.7 macrophage cell line led to comparable regulation of CD45 expression. In immunocompetent mice infected with an m42 deletion mutant lower viral titers were observed in all tissues examined when compared to wildtype MCMV, indicating an important role of m42 for viral replication in vivo. The m42 gene product was identified as an 18 kDa protein expressed with early kinetics and is predicted to be a tail-anchored membrane protein. Tracking of surface-resident CD45 molecules revealed that m42 induces internalization and degradation of CD45. The observation that the amounts of the E3 ubiquitin ligases Itch and Nedd4 were diminished in cells expressing m42 and that disruption of a PY motif in the N-terminal part of m42 resulted in loss of function, suggest that m42 acts as an activator or adaptor for these Nedd4-like ubiquitin ligases, which mark CD45 for lysosomal degradation. In conclusion, the down-modulation of CD45 expression in MCMV-infected myeloid cells represents a novel pathway of virus-host interaction.

Human cytomegalovirus (HCMV) infections of healthy individuals are mostly unnoticed and result in viral latency. However, HCMV can also cause devastating disease, e.g., upon reactivation in immunocompromised patients. Yet, little is known about human immune cell sensing of DNA-encoded HCMV. Recent studies indicated that during viral infection the cyclic GMP/AMP synthase (cGAS) senses cytosolic DNA and catalyzes formation of the cyclic di-nucleotide cGAMP, which triggers stimulator of interferon genes (STING) and thus induces antiviral type I interferon (IFN-I) responses. We found that plasmacytoid dendritic cells (pDC) as well as monocyte-derived DC and macrophages constitutively expressed cGAS and STING. HCMV infection further induced cGAS, whereas STING expression was only moderately affected. Although pDC expressed particularly high levels of cGAS, and the cGAS/STING axis was functional down-stream of STING, as indicated by IFN-I induction upon synthetic cGAMP treatment, pDC were not susceptible to HCMV infection and mounted IFN-I responses in a TLR9-dependent manner. Conversely, HCMV infected monocyte-derived cells synthesized abundant cGAMP levels that preceded IFN-I production and that correlated with the extent of infection. CRISPR/Cas9- or siRNA-mediated cGAS ablation in monocytic THP-1 cells and primary monocyte-derived cells, respectively, impeded induction of IFN-I responses following HCMV infection. Thus, cGAS is a key sensor of HCMV for IFN-I induction in primary human monocyte-derived DC and macrophages.

Human cytomegalovirus (HCMV) harmfully impacts survival after peripheral blood hematopoietic stem cell transplantation (PB-HSCT). Delayed immune reconstitution after cord blood (CB)-HSCT leads to even higher HCMV-related morbidity and mortality. Towards a feasible dendritic cell therapy to accelerate de novo immunity against HCMV, we validated a tricistronic integrase-defective lentiviral vector (coexpressing GM-CSF, IFN-α, and HCMV pp65 antigen) capable to directly induce self-differentiation of PB and CB monocytes into dendritic cells processing pp65 (âSmyleDCpp65â). In vitro, SmyleDCpp65 resisted HCMV infection, activated CD4+ and CD8+ T cells and expanded functional pp65-specific memory cytotoxic T lymphocytes (CTLs). CD34+ cells obtained from PB and CB were transplanted into irradiated NOD.Rag1â/â.IL2γcâ/â mice. Donor-derived SmyleDCpp65 administration after PB-HSCT stimulated peripheral immune effects: lymph node remodeling, expansion of polyclonal effector memory CD8+ T cells in blood, spleen and bone marrow, and pp65-reactive CTL and IgG responses. SmyleDCpp65 administration after CB-HSCT significantly stimulated thymopoiesis. Expanded frequencies of CD4+/CD8+ T cell precursors containing increased levels of T-cell receptor excision circles in thymus correlated with peripheral expansion of effector memory CTL responses against pp65. The comparative in vivo modeling for PB and CB-HSCT provided dynamic and spatial information regarding human T and B cell reconstitution. In vivo potency supports future clinical development of SmyleDCpp65.

Receptors of the signalling lymphocyte-activation molecules (SLAM) family are involved in the functional regulation of a variety of immune cells upon engagement through homotypic or heterotypic interactions amongst them. Here we show that murine cytomegalovirus (MCMV) dampens the surface expression of several SLAM receptors during the course of the infection of macrophages. By screening a panel of MCMV deletion mutants, we identified m154 as an immunoevasin that effectively reduces the cell-surface expression of the SLAM family member CD48, a high-affinity ligand for natural killer (NK) and cytotoxic T cell receptor CD244. m154 is a mucin-like protein, expressed with early kinetics, which can be found at the cell surface of the infected cell. During infection, m154 leads to proteolytic degradation of CD48. This viral protein interferes with the NK cell cytotoxicity triggered by MCMV-infected macrophages. In addition, we demonstrate that an MCMV mutant virus lacking m154 expression results in an attenuated phenotype in vivo, which can be substantially restored after NK cell depletion in mice. This is the first description of a viral gene capable of downregulating CD48. Our novel findings define m154 as an important player in MCMV innate immune regulation.

Herpesviruses establish a lifelong latent infection posing the risk for virus reactivation and disease. In cytomegalovirus infection, expression of the major immediate early (IE) genes is a critical checkpoint, driving the lytic replication cycle upon primary infection or reactivation from latency. While it is known that type I interferon (IFN) limits lytic CMV replication, its role in latency and reactivation has not been explored. In the model of mouse CMV infection, we show here that IFNβ blocks mouse CMV replication at the level of IE transcription in IFN-responding endothelial cells and fibroblasts. The IFN-mediated inhibition of IE genes was entirely reversible, arguing that the IFN-effect may be consistent with viral latency. Importantly, the response to IFNβ is stochastic, and MCMV IE transcription and replication were repressed only in IFN-responsive cells, while the IFN-unresponsive cells remained permissive for lytic MCMV infection. IFN blocked the viral lytic replication cycle by upregulating the nuclear domain 10 (ND10) components, PML, Sp100 and Daxx, and their knockdown by shRNA rescued viral replication in the presence of IFNβ. Finally, IFNβ prevented MCMV reactivation from endothelial cells derived from latently infected mice, validating our results in a biologically relevant setting. Therefore, our data do not only define for the first time the molecular mechanism of IFN-mediated control of CMV infection, but also indicate that the reversible inhibition of the virus lytic cycle by IFNβ is consistent with the establishment of CMV latency.

The unfolded protein response (UPR) is an endoplasmic reticulum (ER)-to-nucleus signaling cascade induced in response to ER stress. The UPR aims at restoring homeostasis, but can also induce apoptosis if stress persists. Infection by human and murine cytomegaloviruses (CMVs) provokes ER stress and induces the UPR. However, both CMVs manipulate the UPR to promote its prosurvival activity and delay apoptosis. The underlying mechanisms remain largely unknown. Recently, we demonstrated that MCMV and HCMV encode a late protein to target IRE1 for degradation. However, the importance of its downstream effector, X Box binding protein 1 (XBP-1), has not been directly studied. Here we show that deletion of XBP-1 prior to or early after infection confers a transient delay in viral propagation in fibroblasts that can be overcome by increasing the viral dose. A similar phenotype was demonstrated in peritoneal macrophages. In vivo, acute infection by MCMV is reduced in the absence of XBP-1. Our data indicate that removal of XBP-1 confers a kinetic delay in early stages of MCMV infection and suggest that the late targeting of IRE1 is aimed at inhibiting activities other than the splicing of XBP-1 mRNA.

Las rocas metamórficas e ígneas del Paleozoico, que forman el basamento de la parte norte del terreno Mixteca en el sur de México, se han agrupado como el Complejo Acatlán. Seis fases de deformaciones distintas se han diferenciado por di' tintas facies metamórficas, grados de ductilidad y correlación temporal. La primera deformación (D) está caracterizada por metamorfismo de alta temperatura (M1), que se sobreimpuso a eclogitas y culminó en anatexis y la intrusión de granitos. Durante esta deformación crecieron grandes micas claras y megablastos de feldespato potásico en las rocas metasedimentarias. La segunda deformación (D2) causó retrometamorfismo (diaftóresis) llegando hasta un metamorfismo de bajo grado (esquisto ·Verde). Las micas de la M1 están conservadas como porfidoclastos dentro de la foliación S1• Durante esta defor- mación se han fracturado con una temperatura menor, mientras que en la foliación S2 están sustituidas por sericita. La defor- mación (D2) causó recristalización dinámica de cuarzo. La tercera deformación (D3) plegó el área en estructuras isoclinales y estrechas con vergencia hacia el poniente. Los sedimentos, depositados entre D2 y D3, sufrieron un metamorfismo de muy bajo grado, y tienen al igual que los granitos, intrusados durante la D3 a un nivel de la corteza superior, una foliación S3. La cuarta deformación (D4) está caracterizada por pliegues tipo “kink”, una zona empinada de cizalla cataclástica y planos de estrías. Los cálculos de tensores de paleoesfuerzo dan ejes subhorizontales con una dirección de NE-SW. La mayor parte de las poblaciones de fallas frágiles está relacionada con la quinta deformación (D5), con ejes y también subhorizontales, con una dirección de ENE-WSW. Fallas normales definen la sexta deformación (D6) con ejes subverticales y ejes subhorizontales con direcciones alrededor de N-S.

Paleozoic metamorphites and magmatites forming the base¬ment of the northern Mixteca terrane in southern Mexico define the Acatlán Complex. Six stages of deformation are defined by metamorphic facies, structural behaviour, and over¬printing relationships. The first deformation (D1) is accompanied by high temperature metamorphism (M1), which over¬prints pre-existing eclogites (D0), and culminates in intrusions of granites. During this deformation, large crystals of mica and megablasts of potassic feldspar grew. The second deformation (D2) is indicated by diaphtoresis up to greenschist fa¬cies. Mica crystals of the M1 are preserved as relictic porphyroclasts in the S1 foliation planes. On D2 planes they were broken and substituted by sericite. D2 caused elongation and recrystallization of quartz. During the third deformation (D3) the Acatlán Complex was folded into W-vergent, tight to isoclinal folds. Sediments deposited between D2 and D3 were metamorphosed by very low grade metamorphism during D3. As the granites intruded into an upper crustal level, they show S3 foliation. The fourth deformation phase is characterized by kinkfolds, cataclastic shear¬zones, and faults with slicken¬sides. Calculations of the paleostress tensors yielded subhorizontal σ1- and σ3-axis indicating NE-SW orientation of σ1. Most of the fault populations belong to the fifth deformation (D5) with subhorizontal σ1- and σ3-axis with ESE-WNW σ1 direction. Normal faults define the sixth deformation (D6) with subvertical σ1-axis and N-S trending σ3-axes.

Neonates, including mice and humans, are highly susceptible to cytomegalovirus (CMV) infection. However, many aspects of neonatal CMV infections such as viral cell tropism, spatio-temporal distribution of the pathogen as well as genesis of antiviral immunity are unknown. With the use of reporter mutants of the murine cytomegalovirus (MCMV) we identified the lung as a primary target of mucosal infection in neonatal mice. Comparative analysis of neonatal and adult mice revealed a delayed control of virus replication in the neonatal lung mucosa explaining the pronounced systemic infection and disease in neonates. This phenomenon was supplemented by a delayed expansion of CD8(+) T cell clones recognizing the viral protein M45 in neonates. We detected viral infection at the single-cell level and observed myeloid cells forming "nodular inflammatory foci" (NIF) in the neonatal lung. Co-localization of infected cells within NIFs was associated with their disruption and clearance of the infection. By 2-photon microscopy, we characterized how neonatal antigen-presenting cells (APC) interacted with T cells and induced mature adaptive immune responses within such NIFs. We thus define NIFs of the neonatal lung as niches for prolonged MCMV replication and T cell priming but also as sites of infection control.