Search

Background.One of the ways to use the genetic potential of bulbous barley, which is characterized by a number of valuable traits, is interspecific hybridization. In crosses ofH. vulgare(2x) ×H. bulbosum(2x) andH. vulgare(4x) ×H. bulbosum(4x) with a genome ratio of 1Hv: 1Hb in a hybrid embryo, elimination of bulbous barley chromosomes is observed in many cases, and the intensity of the process and the result of the crossing depend on the genotypes of the parental forms. This limits the possibility of including a significant variety of parental forms in crosses. Сrossing of diploid forms ofH. vulgarewith tetraploid accessions ofH. bulbosum(4x) results in the formation of triploid hybrids (HvHbHb) with stable chromosomal composition in pollen mother cells (PMCs) at metaphase I (MI) of meiosis. These triploid hybrids can serve as a basis for obtaining series of introgressive lines of cultivated barley. One of the tasks of this type of work is to estimate the involvement of various chromosomes and their arms in homoeologous associations. The aim of this work was to study the possibility of homoeologous pairing of chromosomes of parental species at MI of meiosis in triploid hybrids using GISH and FISH with chromosome-specific markers, as well as to register the participation of individual arms of the cultivated barley chromosomes in homoeologous associations with the chromosomes of bulbous barley in triploid hybrids (HvHbHb).Materials and methods.Seven triploid hybrids ofH. vulgare×H.bulbosum(HvHbHb) obtained in four combinations of crosses with the participation of three diploid cultivars of cultivated barley and two tetraploid accession of bulbous barley were used in this study. The features of homoeologous pairing of chromosomes at MI were studied using the method of fluorescentin situhybridization (GISH and FISH) with chromosome-specific markers.ResultsAll the studied hybrid plants are characterized by a stable chromosomal composition in PMCs at the MI stage of meiosis. Meiotic configurations formed by homoeologous chromosomes of the parental species, ranging from 0.87 to 1.40 on average per cell, were identified in all the studied plants. Among them, vbb trivalents prevailed. Analysis of chromosome pairing at MI in triploid hybrids revealed the participation of all chromosome arms ofH. vulgarein homoeologous Hv-Hb associations, except for the short arm of chromosome 1H. In all the studied triploid hybrids, there is a tendency for a higher frequency of involvement of the long arms of chromosomes in the formation of homoeologous associations; this feature is most clearly manifested in case of chromosome 5H.ConclusionsIntergenomic associations with the participation of all arms ofH. vulgarechromosomes, except for the short arm of chromosome 1H, were revealed at MI inH. vulgare×H. bulbosumtriploid hybrids (HvHbHb). Chromosome 5H, as well as any other cultivated barley chromosome, is characterized by a higher involvement of its long arm in homoeologous associations Hv-Hb, as compared to the short arm.

Lameness in pigs is one of the major reasons for culling and early losses in pigs. This can be linked to osteoporosis due to pathologic alterations in bone mineral density (BMD) or bone mineral content (BMC) and may also be linked to the sex. Dealing with the ban on piglet castration without anaesthesia in Germany 2021, we have three male “sex” types: entire boars (EB), immunocastrated boars (IB), and surgically castrated boars (SB). The hypothesis of the present study is that BMC or BMD varies between different male sex types. If sex has an effect on bone mineralization (BMC or BMD) and if this affects leg health, it could result in more lameness and problems during fattening in the negatively affected sex type. The present study evaluated bone mineralization (in terms of BMD and BMC) and body composition traits using dual-energy X-ray absorptiometry (DXA) three times during growth at 30, 50, and 90 kg live body weight. Nine body regions were analysed for bone mineral traits and compared for different male sex types and the fattening season. Significant differences were found regarding BMD (and BMC) among EB, IB, and SB for whole-body BMD (BMC). Additionally significant differences were found in the front and lower hind limbs, where SB showed a significantly higher BMD compared to EB, with IB in between. Additionally regional differences were detected among the groups. Further studies are needed to evaluate the effect of these differences in bone mineralization on leg health.

The objective of this study was to compare the reproductive performance, milk yield and composition, and udder health and conformation traits of Holstein (Ho), F1, and R1 crossbred Ho × Simmental (Sim) cows. Three commercial dairy farms in south Brazil were used as the research units. All farms held Ho, F1, and R1 crossbred Ho × Sim (¾ Ho × ¼ Sim and ¾ Sim × ¼ Ho) cows. The collection of milk samples and evaluation of udder conformation traits occurred during four visits to each farm. In addition to the actively collected data, retrospective reproduction records of the farms served as the basis for the statistical analysis using analysis of variance models using SAS. The F1 crossbred Ho × Sim cows and ¾ Sim (first rotational crossbreeding generation = R1 using Sim semen) cows had a shorter calving interval and calving to first service interval compared to the Ho cows (P

The advancement of new immunotherapies necessitates appropriate probes to monitor the presence and distribution of distinct immune cell populations. Considering the key role of CD4+T cells in regulating immunological processes, we generated novel single-domain antibodies (nanobodies, Nbs) that specifically recognize human CD4. After in depth analysis of their binding properties, recognized epitopes, and effects on T cell proliferation, activation and cytokine release, we selected CD4 Nbs that did not interfere with crucial T cell processesin vitroand converted them into immune tracers for non-invasive molecular imaging.By optical imaging, we demonstrate the ability of a high-affinity CD4-Nb to specifically visualize CD4+cellsin vivousing a xenograft model. Furthermore, time-resolved immune positron emission tomography (immunoPET) of a human CD4 knock-in mouse model showed rapid accumulation of64Cu-radiolabeled CD4-Nb in CD4+T cell-rich tissues. We propose that the CD4 Nbs presented here could serve as versatile probes for stratifying patients and monitoring individual immune responses during personalized immunotherapy in both cancer and inflammatory diseases.

Skeletal disorders, including fractures and osteoporosis, in laying hens cause major welfare and economic problems. Although genetics have been shown to play a key role in bone integrity, little is yet known about the underlying genetic architecture of the traits. This study aimed to identify genes associated with bone breaking strength and bone mineral density of the tibiotarsus and the humerus in laying hens. Potentially informative single nucleotide polymorphisms (SNP) were identified using Random Forests classification. We then searched for genes known to be related to bone stability in close proximity to the SNPs and identified 16 potential candidates. Some of them had human orthologues. Based on our findings, we can support the assumption that multiple genes determine bone strength, with each of them having a rather small effect, as illustrated by our SNP effect estimates. Furthermore, the enrichment analysis showed that some of these candidates are involved in metabolic pathways critical for bone integrity. In conclusion, the identified candidates represent genes that may play a role in the bone integrity of chickens. Although further studies are needed to determine causality, the genes reported here are promising in terms of alleviating bone disorders in laying hens.

Keel bone damage is an important animal welfare problem in laying hens. Two generations of four layer lines, differing in phylogenetic background and performance level and kept in single cages or floor pens were weighed and scored for keel bone deformities (KBD) during the laying period. KBD, keel bone fractures (KBF) and the bone mineral density (BMD) of the keels were assessed post mortem. For BMD, relationships to laying performance and body growth were estimated. Caged hens showed more deformities, but fewer fractures and a lower BMD of the keel bone than floor-housed hens. White-egg layers had a lower BMD (0.140–0.165 g/cm2) and more KBD than brown-egg layers (0.179–0.184 g/cm2). KBF occurred more often in the high-performing lines than the moderate-performing ones. However, in the high-performing lines, BMD was positively related to total egg number from 18 to 29 weeks of age. The adult body weight derived from fitted growth curves (Gompertz function) had a significant effect (p &lt, 0.001) on keels’ BMD. The study contributes to the understanding of predisposing factors for keel bone damage in laying hens. It showed that the growth rate has a rather subordinate effect on keels’ BMD, while the BMD itself greatly affects KBD.

In light of the COVID-19 pandemic, there is an ongoing need for diagnostic tools to monitor the immune status of large patient cohorts and the effectiveness of vaccination campaigns. Here, we present 11 unique nanobodies (Nbs) specific for the SARS-CoV-2 spike receptor-binding domain (RBD), of which 8 Nbs potently inhibit the interaction of RBD with angiotensin-converting enzyme 2 (ACE2) as the major viral docking site. Following detailed epitope mapping and structural analysis, we select two inhibitory Nbs, one of which binds an epitope inside and one of which binds an epitope outside the RBD:ACE2 interface. Based on these, we generate a biparatopic nanobody (bipNb) with viral neutralization efficacy in the picomolar range. Using bipNb as a surrogate, we establish a competitive multiplex binding assay ("NeutrobodyPlex") for detailed analysis of the presence and performance of neutralizing RBD-binding antibodies in serum of convalescent or vaccinated patients. We demonstrate that NeutrobodyPlex enables high-throughput screening and detailed analysis of neutralizing immune responses in infected or vaccinated individuals, to monitor immune status or to guide vaccine design.

Crossbreeding in dairy cattle has been used to improve functional traits, milk composition, and efficiency of Holstein herds. The objective of the study was to compare indicators of the metabolic energy balance, nonesterified fatty acids (NEFA), beta-hydroxybutyrate (BHBA), glucose, body condition score (BCS) back fat thickness (BFT), as well as milk yield and milk composition of Holstein and Simmental cows, and their crosses from the prepartum period until the 100th day of lactation at the Livestock Center of the Ludwig Maximilians University (Munich, Germany). In total, 164 cows formed five genetic groups according to their theoretic proportion of Holstein and Simmental genes as follows: Holstein (100% Holstein, n = 9), R1-Hol (51&ndash, 99% Holstein, n = 30), first generation (F1) crossbreds (50% Holstein, 50% Simmental, n = 17), R1-Sim (1&ndash, 49% Holstein, n = 81) and Simmental (100% Simmental, n = 27). The study took place between April 2018 and August 2019. BCS, BFT blood parameters, such as BHBA, glucose, and NEFA were recorded weekly. A mixed model analysis with fixed effects breed, week (relative to calving), the interaction of breed and week, parity, calving year, calving season, milking season, and the repeated measure effect of cow was used. BCS increased with the Simmental proportion. All genetic groups lost BCS and BFT after calving. Simmental cows showed lower NEFA values. BHBA and glucose did not differ among genetic groups, but they differed depending on the week relative to calving. Simmental and R1-Sim cows showed a smaller effect than the other genetic groups regarding changes in body weight, BCS, or back fat thickness after a period of a negative energy balance after calving. There was no significant difference for milk yield among genetic groups, although Simmental cows showed a lower milk yield after the third week after calving. Generally, Simmental and R1-Simmental cows seemed to deal better with a negative energy balance after calving than purebred Holstein and the other crossbred lines. Based on a positive heterosis effect of 10.06% for energy corrected milk (ECM), the F1, however, was the most efficient crossbred line.

Depending on the breed or crossbreed line, cows have to cope with a more or less severe negative energy balance during the period of high milk yields in early lactation, which can be detected by beta-hydroxybutyrate (BHBA) and non-esterified fatty acids (NEFAs) in blood. Preventing cows from undergoing a severe negative energy balance by breeding and/or feeding measures is likely to be supported by the public and may help to improve the sustainability of milk production. The aim was to compare BHBA and NEFA concentrations in the blood of Holstein and Simmental cows and their crosses during the prepartum period until the end of lactation. In total, 164 cows formed five genetic groups according to their theoretic proportion of Holstein and Simmental genes as follows: Holstein (100% Holstein; n = 9), R1-Hol (51–99% Holstein; n = 30), F1 crossbreds (50% Holstein, 50% Simmental; n = 17), R1-Sim (1–49% Holstein; n = 81) and Simmental (100% Simmental; n = 27). NEFA and BHBA were evaluated once a week between April 2018 and August 2019. A mixed model analysis with fixed effects breed, week (relative to calving), the interaction of breed and week, parity, calving year, calving season, milking season, and the repeated measure effect on cows was used. Holstein cows had higher NEFAs (0.196 ± 0.013 mmol/L), and Simmental cows had the lowest NEFA concentrations (0.147 ± 0.008 mmol/L, p = 0.03). R1-Sim, F1 and R1-Hol cows had intermediate values (0.166 ± 0.005, 0.165 ± 0.010, 0.162 ± 0.008 mmol/L; respectively). The highest NEFA value was found in the first week after calving (0.49 ± 0.013 mmol/L). BHBA did not differ among genetic groups (p = 0.1007). There was, however, an interaction between the genetic group and week (p = 0.03). While Simmental, R1-Sim and F1 cows had the highest BHBA value, the second week after calving (0.92 ± 0.07 and 1.05 ± 0.04, and 1.10 ± 0.10 mmol/L, respectively), R1-Hol and Holstein cows showed the BHBA peak at the fourth week after calving (1.16 ± 0.07 and 1.36 ± 0.12 mmol/L, respectively). Unexpectedly, Holstein cows had a high BHBA peak again at week 34 after calving (1.68 ± 0.21 mmol/L). The genetic composition of the cows affects NEFA and BHBA. Simmental and R1-Sim cows mobilize fewer body reserves after calving. Therefore, dairy cows with higher degrees of Simmental origin might be more sustainable in comparison with Holstein genetics in the present study.

Simple Summary Brittle or fractured bones due to continuous demineralisation cause major welfare and economic problems in laying hens. Bone weakness in laying hens is frequently attributed to long-term selection for increased egg production, but this is controversially discussed in the scientific literature. We aimed at characterizing factors influencing the bone breaking strength of laying hens, focusing mainly on the effect of eggshell production. By examining four different chicken layer lines separately, a genetically diverse spectrum of laying hen origins was included in our study. It was shown that bone strength is primarily influenced by bone mineral density. A strong association between bone strength and eggshell production was not observed within each of the lines studied. This applied to all layer lines. Our results suggest that a high egg number does not generally impair bone stability within layer lines. Findings from this study contribute to the discussion on the improvement of bone stability in poultry breeding programs and thus lead to increased animal welfare in egg production. Abstract Impaired animal welfare due to skeletal disorders is likely one of the greatest issues currently facing the egg production industry. Reduced bone stability in laying hens is frequently attributed to long-term selection for increased egg production. The present study sought to analyse the relationship between bone stability traits and egg production. The study comprised four purebred layer lines, differing in their phylogenetic origin and performance level, providing extended insight into the phenotypic variability in bone characteristics in laying hens. Data collection included basic production parameters, bone morphometry, bone mineral density (BMD) and bone breaking strength (BBS) of the tibiotarsus and humerus. Using a multifactorial model and regression analyses, BMD proved to be of outstanding importance for bone stability. Only for the tibiotarsus were morphometric parameters and the bone weight associated with BBS. Within the chicken lines, no effect of total eggshell production on BBS or BMD could be detected, suggesting that a high egg yield itself is not necessarily a risk for poor bone health. Considering the complexity of osteoporosis, the estimated genetic parameters confirmed the importance of genetics in addressing the challenge of improving bone strength in layers.

With the increasing anthropogenic impacts on fish habitats, it has become more important to understand which primary resources sustain fish populations. This resource utilization can differ between fish life stages, and individuals can migrate between habitats in search of resources. Such lifetime information is difficult to obtain due to the large spatial and temporal scales of fish behavior. The otolith organic matrix has the potential to indicate this resource utilization and migration with δ13C values of essential amino acids (EAAs), which are a direct indication of the primary producers. In a proof‐of‐concept study, we selected the Acoupa weakfish, Cynoscion acoupa, as a model fish species with distinct ontogenetic migration patterns. While it inhabits the Brazilian mangrove estuaries during juvenile stages, it moves to the coastal shelf as an adult. Thus, we expected that lifetime resource utilization and migration would be reflected in δ13CEAA patterns and baseline values in C. acoupa otoliths. By analyzing the C. acoupa otolith edges across a size range of 12–119 cm, we found that baseline δ13CEAA values increased with size, which indicated an estuarine to coastal shelf distribution. This trend is highly correlated with inorganic δ13C values. The δ13CEAA patterns showed that estuarine algae rather than mangrove‐derived resources supported the juvenile C. acoupa populations. Around the juvenile size of 40 cm, resource utilization overlapped with those of adults and mean baseline δ13CEAA values increased. This trend was confirmed by comparing otolith core and edges, although with some individuals potentially migrating over longer distances than others. Hence, δ13CEAA patterns and baseline values in otoliths have great potential to reconstruct ontogenetic shifts in resource use and habitats. The insight could aid in predictions on how environmental changes affect fish populations by identifying the controlling factors at the base of the food web.

The aim of this study was to evaluate the possibility of predicting boar taint in live pigs non-invasively. For this, different magnetic resonance imaging based body composition traits (including testis volume) were compared with androstenone, skatole, and indole levels (ng/g fat) of the carcass` fat tissue after slaughtering. Additionally, in order to find traits which could help in predicting boar taint, the related sensory test results were included in the analysis. A number of 34 entire boars (EB) and 34 immunocastrated boars (IB; first injection at an age of 77 ± 1 days and second injection at an age of 137 ± 1 days) were scanned two times during growth (at 60 and 90 kg live body weight) using magnetic resonance imaging (MRI). Three body regions (shoulder, loin, and ham) were examined. Additionally the testes volume was calculated. After slaughtering, boar taint samples were taken for olfactory testing and stable isotope dilution assays. These data were compared with selected body composition traits in order to analyze the relationship between MRI traits and boar taint. The results showed that IB tend to have greater subcutaneous fat layers (belly fat, shoulder fat, back fat) than EB. Within EB, larger testis volumes and a higher amount of body fat (especially belly fat) are associated with a higher level of androstenone.

Objective Laron syndrome (LS) is a rare, autosomal recessive disorder in humans caused by loss-of-function mutations of the growth hormone receptor (GHR) gene. To establish a large animal model for LS, pigs with GHR knockout (KO) mutations were generated and characterized. Methods CRISPR/Cas9 technology was applied to mutate exon 3 of the GHR gene in porcine zygotes. Two heterozygous founder sows with a 1-bp or 7-bp insertion in GHR exon 3 were obtained, and their heterozygous F1 offspring were intercrossed to produce GHR-KO, heterozygous GHR mutant, and wild-type pigs. Since the latter two groups were not significantly different in any parameter investigated, they were pooled as the GHR expressing control group. The characterization program included body and organ growth, body composition, endocrine and clinical-chemical parameters, as well as signaling studies in liver tissue. Results GHR-KO pigs lacked GHR and had markedly reduced serum insulin-like growth factor 1 (IGF1) levels and reduced IGF-binding protein 3 (IGFBP3) activity but increased IGFBP2 levels. Serum GH concentrations were significantly elevated compared with control pigs. GHR-KO pigs had a normal birth weight. Growth retardation became significant at the age of five weeks. At the age of six months, the body weight of GHR-KO pigs was reduced by 60% compared with controls. Most organ weights of GHR-KO pigs were reduced proportionally to body weight. However, the weights of liver, kidneys, and heart were disproportionately reduced, while the relative brain weight was almost doubled. GHR-KO pigs had a markedly increased percentage of total body fat relative to body weight and displayed transient juvenile hypoglycemia along with decreased serum triglyceride and cholesterol levels. Analysis of insulin receptor related signaling in the liver of adult fasted pigs revealed increased phosphorylation of IRS1 and PI3K. In agreement with the loss of GHR, phosphorylation of STAT5 was significantly reduced. In contrast, phosphorylation of JAK2 was significantly increased, possibly due to the increased serum leptin levels and increased hepatic leptin receptor expression and activation in GHR-KO pigs. In addition, increased mTOR phosphorylation was observed in GHR-KO liver samples, and phosphorylation studies of downstream substrates suggested the activation of mainly mTOR complex 2. Conclusion GHR-KO pigs resemble the pathophysiology of LS and are an interesting model for mechanistic studies and treatment trials., Highlights • GHR-deficient pigs reveal postnatal growth retardation, disproportionate organ growth and an increased total body fat content. • GHR-deficient pigs show markedly reduced serum IGF1 and IGFBP3 levels, and transient juvenile hypoglycemia. • Increased expression and phosphorylation of IRS1 in liver of adult GHR-deficient pigs suggest increased insulin sensitivity. • Increased phosphorylation of JAK2 in liver of GHR-deficient pigs may be explained by higher serum leptin levels and activation of hepatic LEPR.

Interleukin (IL)-36 cytokines are important regulators of mucosal homeostasis and inflammation. We previously established that oral epithelial cells strongly upregulate IL-36γ expression in response to the bacterial pathogen Porphyromonas gingivalis. Here, we have established that IL-36γ stimulates the expression of the IL-12 cytokine family members, IL-23p19 and Epstein-Barr Virus-Induced Gene 3 (EBI3), by oral epithelial cells; their expression was also selectively stimulated by IL-36α. Notably, IL-23p19 and EBI3 expression was not stimulated by P. gingivalis, thus suggesting that their expression by the oral epithelium in response to P. gingivalis is likely to be mediated in an autocrine manner by IL-36γ. The IL-36γ-inducible expression of IL-23p19 and EBI3 was found to be diametrically regulated by the mitogen-activated protein kinase/extracellular signal regulated kinase (MEK)-extracellular signal-regulated kinase 1/2 (ERK1/2) pathway, whereby the activation of MEK-ERK signaling likely functions as a negative feedback mechanism to limit EBI3 expression. Furthermore, epidermal growth factor receptor (EGFR) signaling, which is important for mucosal homeostasis, was demonstrated to modulate, in a MEK-ERK-dependent manner, the stimulation of IL-23p19 and EBI3 expression by IL-36γ. IL-23p19 and EBI3 have recently been shown to heterodimerize to form the novel cytokine IL-39 and promote neutrophil expansion. EBI3 has been shown to also have IL-12 cytokine family independent functions (e.g. mediating IL-6 trans-signaling). Thus, this study not only advances our understanding of how IL-36 cytokines may control mucosal inflammation, but also establishes EGFR signaling as a potentially important modulator of IL-36 cytokine function.

In a previous study, genome-wide mapping of quantitative trait loci (QTL) for five body composition traits, three bone mineral traits and live weight was performed using whole-body dual-energy X-ray absorptiometry (DXA) data. Since QTL for bone mineral traits were rare, the current study aimed to clarify whether the mapping results were influenced by the analysed body regions. Thus, the same material (551 pigs) and methods as in the whole-body QTL mapping study were used. However, for evaluation of the DXA scans, we manually defined two body regions: (i) from the last ribs to the pelvis (A) and (ii) including the pelvis and the hind limbs (P). Since live weight was not affected by the regional analysis, it was omitted from the QTL mapping design. Our results show an overall high consistency of mapping results especially for body composition traits. Two thirds of the initial whole-body QTL are significant for both A and P. Possible causes for the still low number of bone mineral QTL and the lower consistency found for these traits are discussed. For body composition traits, the data presented here show high genome-wide Pearson correlations between mapping results that are based on DXA scans with the time-saving whole-body standard setting and mapping results for DXA data that were obtained by time-consuming manual definition of the regions of interest. However, our results also suggest that whole-body or regional DXA scans might generally be less suitable for mapping of bone mineral traits in pigs. An analysis of single reference bones could be more useful.

We aimed to determine the relationship of milk yield, milk composition, and somatic cell score (SCS) with functional traits (lameness score, udder cleanliness score, udder depth, and teat end hyperkeratosis score) and physiological status (parity and days in milk (DIM) of cows housed in compost bedded pack barn system (CBP) using multivariate analysis. Data were collected in September and October of 2017 and evaluated using factor and cluster analysis. Four factors were formed, with the first factor showing the traits responsible for altering the lactose content in milk, the second factor comprising the effect of DIM, the third factor representing the teat end hyperkeratosis score, and the fourth factor demonstrating the udder cleanliness. The cluster analysis formed three clusters. Clusters 2 and 3 differed for protein and lactose content, SCS, parity, DIM, lameness score, and udder depth. Cluster 1 differed from the others, especially due to the high milk yield. Cow conformation traits and physiological status affected milk yield and composition in CBP. There is an association between udder depth, parity, and SCS with negative impact on lactose content. Parity is also associated with higher milk yield and teat end hyperkeratosis score. Cows with lower lameness score presented dirtier udders due to increased movements on the feed parlor and water troughs areas. Special attention to the improvement of the udder conformation traits, as well as management practices that reduce hyperkeratosis, are necessary to keep high yielding, healthy, and cows with higher longevity. The cleanliness in the feed parlor and water troughs area is very important to preserve milk quality and healthy cows.

Mycobacterium avium subsp. paratuberculosis (MAP) is a pathogen causing paratuberculosis in cattle and small ruminants. During the long asymptomatic subclinical stage, high numbers of MAP are excreted and can be transmitted to food for human consumption, where they survive many of the standard techniques of food decontamination. Whether MAP is a human pathogen is currently under debate. The aim of this study was a better understanding of the host-pathogen response by analyzing the interaction of peripheral blood lymphocytes (PBL) from cattle with MAP in their exoproteomes/secretomes to gain more information about the pathogenic mechanisms of MAP. Because in other mycobacterial infections, the immune phenotype correlates with susceptibility, we additionally tested the interaction of MAP with recently detected cattle with a different immune capacity referred as immune deviant (ID) cows. In PBL, different biological pathways were enhanced in response to MAP dependent on the immune phenotype of the host. PBL of control cows activated members of cell activation and chemotaxis of leukocytes pathway as well as IL-12 mediated signaling. In contrast, in ID cows CNOT1 was detected as highly abundant protein, pointing to a different immune response, which could be favorable for MAP. Additionally, MAP exoproteomes differed in either GroEL1 or DnaK abundance, depending on the interacting host immune response. These finding point to an interdependent, tightly regulated response of the bovine immune system to MAP and vise versa.

Malaria claims about half a million lives each year. Plasmodium falciparum, the causative agent of the most severe form of the disease, uses proteins that are translocated to the surface of infected erythrocytes for immune evasion. To circumvent the detection of these gene products by the immune system, the parasite evolved a complex strategy that includes gene duplications and elaborate sequence polymorphism. STEVORs are one family of these variant surface antigens and are encoded by about 40 genes. Using deep RNA sequencing of blood-stage parasites, including free merozoites, we first established stevor expression of the cultured isolate and compared it with published transcriptomes. We reveal a biphasic expression of most stevor genes and confirm this for individual STEVORs at the protein level. The membrane topology of a rhoptry-associated variant was experimentally elucidated and linked to host cell invasion, underlining the importance of this multifunctional protein family for parasite proliferation., During its intraerythrocytic development, the malaria parasite Plasmodium falciparum exposes variant surface antigens (VSAs) on infected erythrocytes to establish and maintain an infection. One family of small VSAs is the polymorphic STEVOR proteins, which are marked for export to the host cell surface through their PEXEL signal peptide. Interestingly, some STEVORs have also been reported to localize to the parasite plasma membrane and apical organelles, pointing toward a putative function in host cell egress or invasion. Using deep RNA sequencing analysis, we characterized P. falciparum stevor gene expression across the intraerythrocytic development cycle, including free merozoites, in detail and used the resulting stevor expression profiles for hierarchical clustering. We found that most stevor genes show biphasic expression oscillation, with maximum expression during trophozoite stages and a second peak in late schizonts. We selected four STEVOR variants, confirmed the expected export of these proteins to the host cell membrane, and tracked them to a secondary location, either to the parasite plasma membrane or the secretory organelles of merozoites in late schizont stages. We investigated the function of a particular STEVOR that showed rhoptry localization and demonstrated its role at the parasite-host interface during host cell invasion by specific antisera and targeted gene disruption. Experimentally determined membrane topology of this STEVOR revealed a single transmembrane domain exposing the semiconserved as well as variable protein regions to the cell surface.

Mycobacterium avium subsp. paratuberculosis (MAP) is a pathogen causing paratuberculosis in cattle and small ruminants. During the long asymptomatic subclinical stage, high numbers of MAP are excreted and can be transmitted to food, where they survive many of the standard techniques of food decontamination. If these MAP are harmful to the consumers is currently under debate. In general, there is a lack of information regarding interaction of the hosts immune system with MAP.In this study, we tested the interaction of peripheral blood lymphocytes (PBL) from cattle with MAP in their exoproteomes/secretomes. Because in other mycobacterial infections, the immune phenotype correlates with susceptibility, we additionally tested the interaction of MAP with recently detected immune deviant cows.In PBL, different biological pathways were enhanced in response to MAP dependent on the immune phenotype of the host. PBL of control cows activated members of cell activation and chemotaxis of leukocytes pathway as well as IL-12 mediated signaling. In contrast, in ID cows CNOT1 was detected as highly abundant protein, pointing to a different immune response, which could be favorable for MAP. Additionally, MAP reacted different to the hosts. Their exoproteomes differed in either GroEL1 or DnaK abundance, depending on the interacting immune response.These findings point to an interdependent, tightly regulated response of MAP and the immune system.

Background:Activin A and follistatin belong to an anti-inflammatory auto-regulatory cycle. Patients with rheumatoid arthritis (RA) have increased activin A levels in the synovial fluid and tissue. During inflammation, activin A is released systemically, then inducing its antagonist follistatin. This negative feedback is active in different cell types but not RA synovial fibroblasts (SF). Fibroblasts interact with endothelial cells in inflamed tissues inducing angiogenesis.Objectives:Evaluation of the role of activin A and follistatin on RASF and endothelial cell interactions.Methods:RA synovium was used for RASF isolation, HUVEC were commercially obtained. RASF and HUVEC were stimulated in mono- and coculture. Direct: RASF together with HUVEC; indirect: inserts with HUVEC separated by a membrane from RASF. Stimuli: activin A 15ng/ml, follistatin 500ng/ml, IL-1β 1ng/ml. Proliferation was analyzed by BrdU assay. RASF were Calcein-AM stained. Cells were transferred to 24-well plates after 18h stimulation. After adhesion for 1h, non-adherent cells were removed by shaking 3x for 5 min. Afterwards, fluorescent cells were quantified. For the flow-adhesion assay, HUVEC were cultured on rattail collagen coated capillary slides. HUVEC and RASF were stimulated for 4h with TNFα, TNFα+activin A or TNFα+follistatin. After stimulation, 2x10^6 RASF were resuspended in 20ml medium and sent through the capillaries. Two 1min videos were recorded (18.4ml/h, 30.5ml/h). Settings: TNFα-stimulated RASF on HUVEC stimulated wit TNFα or activin A+TNFα or follistatin+ TNFα. For migration assays, 2% FCS medium with 1x10^5 cells were placed in inserts (8µm membrane) into wells with 10% FCS (control: 2%FCS vs. 2%FCS) and stimulated with, IL-1, IL-1+activinA and IL-1+follistatin. After 16h, migrated cells were quantified. For scratch-assay 4x10^5 cells were cultured overnight, then cells were scratched and stimulated, afterwards 3 pictures per scratch were taken at start, after 10h and every 1.5h. Cells migrating into the scratch area were quantified.Results:IL-1 induced activin A in RASF and HUVEC in all settings. IL-1-induced activin A release was reduced by follistatin in HUVEC monoculture and both cocultures compared to IL-1 alone but not in RASF monoculture. IL-1-induced IL-6 release was reduced by activin A in HUVEC and indirect coculture but not in RASF monoculture and direct coculture. Follistatin did not alter IL-6 responses. IL-1 induced VEGF in RASF but not in HUVEC and was not altered by activin A. Short-term adhesion showed no significant influence of activin A or follistatin (n=4). Flow adhesion assay showed reduced adherence/rolling of RASF on HUVEC stimulated with TNFα and activin A (n=4). Migration assays showed that IL-1 decreased migration but without significant difference between the induced effects mediated by IL-1+activinA and IL-1+follistatin (n=4). Scratch assay showed increased migration in dicrect coculture with greater difference between stimulated and unstimulated cells in RASF monoculture and indirect coculture (n=4). Proliferation was not altered by activin A or follistatin.Conclusion:In direct and indirect coculture of HUVEC with RASF the effect on HUVEC was dominant leading to reduced IL-1-induced activin A release. However, the IL-1-induced IL-6 release in RASF or HUVECs was decreased by activin A in HUVEC monoculture and indirect coculture but not during cell-contact between both cell types. The direct interaction of RASF with HUVEC seems to prevent the reducing activin A effect on IL-6 release in HUVECs. Activin A seems to not to have an impact on short-term cell adhesion but first observations show, that activin A alters selectin-mediated adhesion under flow conditions. The migration assay shows that IL-1-induced effects on cell migration were enhanced by activin A and follistatin. Migration assay shows that IL-1-induced decrease on migration more prominent in indirect coculture and RASF monoculture than in direct coculture although in gap migration in the scratch assay was highest in direct coculture.Disclosure of Interests:None declared

Anaerobic biodegradation of hydrocarbons mainly related to petroleum can be observed in numerous natural and anthropogenically overprinted environments. While microbiological and chemical tracers are used to study the development of such habitats, not much is known about the impact of particular bacteria on the system. Therefore, we used Aromatoleum sp. HxN1, an n-alkane-utilizing betaproteobacterium capable of degrading crude oil constituents under strictly anoxic conditions, as a model organism to examine its potential to release carboxylic acid metabolites into the surrounding environment. We conducted cultivation experiments with pure substrates, as well as with crude oil, and demonstrated that intermediates of catabolic pathways were highly abundant in the exometabolome. Strikingly, the metabolites found in highest concentrations in the cell-free culture supernatant of strain HxN1 grown with crude oil were associated with the transformation of potentially toxic aromatic hydrocarbons, which do not support growth of this particular bacterium. Additionally, a deficiency in coenzyme A caused by a highly productive substrate transformation may induce the excretion of a surplus of metabolites in order to regain coenzyme A. Under natural conditions, i.e., in the presence of mixed communities, such transformation products may be further degraded by other organisms, ultimately leading to mineralization.

Keratinocytes are central to the barrier functions of surface epithelia, such as the gingiva and epidermis. RIPK4 is a key regulator of keratinocyte differentiation; however, the signalling pathways in which it functions remain poorly defined. In this study, we identified a regulatory relationship between RIPK4 and ELF3, an ETS family transcription factor. RIPK4 was shown to be important for the upregulation of ELF3 gene expression by the PKC agonist PMA in both oral and epidermal keratinocytes. RIPK4 promotes keratinocyte differentiation in part by phosphorylating and thereby activating the IRF6 transcription factor. Significantly, silencing of IRF6 inhibited the PMA-inducible expression of ELF3. A role for the GRHL3 transcription factor, a downstream target gene of IRF6, in the regulation of ELF3 expression was similarly demonstrated. ELF3 has previously been shown to regulate the expression of SPPR1A and SPRR1B, small proline-rich proteins that contribute to the cornification of keratinocytes. Consistently, RIPK4 and IRF6 were important for the PMA-inducible expression of SPRR1A and SPRR1B. They were also important for the upregulation of TGM1, a transglutaminase that catalyses the cross-linking of proteins, including small proline-rich proteins, during keratinocyte cornification. RIPK4 was also shown to upregulate the expression of TGM2 independently of IRF6. Collectively, our findings position RIPK4 upstream of a hierarchal IRF6-GRHL3-ELF3 transcription factor pathway in keratinocytes, as well as provide insight into a potential role for RIPK4 in the regulation of keratinocyte cornification.

Keratinocytes of the oral mucosa and epidermis play key roles in host defense. In addition to functioning as a physical barrier, they also produce cytokines to elicit inflammation in response to infection or injury. We recently established that receptor-interacting protein kinase 4 (RIPK4) and interferon regulatory factor 6 (IRF6) function as a cell-intrinsic signaling axis to regulate keratinocyte differentiation. In this study, we have demonstrated a functional relationship between RIPK4 and IRF6 in the control of proinflammatory cytokine expression in keratinocytes. The overexpression of RIPK4 by oral keratinocytes induced the strong expression of CCL5 and CXCL11. In contrast, the expression of other cytokines (e.g. IL8 and TNF) was largely unaffected, thus demonstrating specificity in the induction of proinflammatory cytokine expression by RIPK4. CCL5 and CXCL11 expression were also induced in response to the activation of the PKC pathway, and gene silencing experiments indicated that their inducible expression was dependent on RIPK4 and IRF6. Moreover, gene reporter assays suggested that RIPK4 induces CCL5 and CXCL11 expression by stimulating the transactivation of their promoters by IRF6. Accordingly, our findings suggest that the RIPK4-IRF6 signaling axis plays a multifaceted role in barrier epithelial homeostasis through its regulation of both keratinocyte inflammation and differentiation.

Deformability of Plasmodium falciparum gametocyte-infected erythrocytes (GIEs) allows them to persist for several days in blood circulation and to ensure transmission to mosquitoes. Here, we investigate the mechanism by which the parasite proteins STEVOR (SubTElomeric Variable Open Reading frame) exert changes on GIE deformability. Using the microsphiltration method, immunoprecipitation, and mass spectrometry, we produce evidence that GIE stiffness is dependent on the cytoplasmic domain of STEVOR that interacts with ankyrin complex at the erythrocyte skeleton. Moreover, we show that GIE deformability is regulated by protein kinase A (PKA)–mediated phosphorylation of the STEVOR C-terminal domain at a specific serine residue (S324). Finally, we show that the increase of GIE stiffness induced by sildenafil (Viagra) is dependent on STEVOR phosphorylation status and on another independent mechanism. These data provide new insights into mechanisms by which phosphodiesterase inhibitors may block malaria parasite transmission.

Research has shown that digestion of A1 &beta, casein (&beta, CN) affects gastrointestinal motility and opioid activity through the release of the peptide &beta, casomorphin-7 (&beta, CM7). In the case of the A2 variant, the cleavage of &beta, CM7 does not occur or occurs at a very low rate. Therefore, the aim of the study was to compare the effects of milk containing either homozygote A1 or A2 &beta, CN on health and growth parameters of dairy calves. Forty-seven neonatal calves (24 females, 23 males) of the breeds German Holstein (GH, n = 9), German Simmental (GS, n = 33) and their crossing (GH &times, GS, n = 5) were used in a 21-day feeding study. Fecal score (FS), respiratory frequency (RF), and rectal body temperature (BT) were recorded daily, whereas body weight was measured at birth and at day 21 to estimate the average daily weight gain (ADG). Additionally, blood was collected from calves three times during the experimental period and, for the first time, the respective plasma samples were analyzed for intact &beta, CM7. Consumption of A2-milk led to a lower daily milk intake (dMI) (p &lt, 0.05). Furthermore, fecal consistency was softer for calves fed A2-milk (p &lt, 0.05). Although 44% of A2-calves had diarrhea or revealed a tendency towards it (FS &ge, 3), A1-calves had a prevalence of 21%. Calves with a FS of 4 were offered an electrolyte solution and received a dietary food supplement for the stabilization of the fluid and electrolyte balance. Nevertheless, similar ADG and end weights (EW) of calves fed A1- or A2-milk (p &gt, 0.05) indicate that A2-milk may compensate higher diarrhea rates and lower dMI due to the associated higher protein content. This is the first report of intact &beta, CM7 in plasma of calves fed milk of either A1 or A2 &beta, CN. Evidence from this study suggests that due to the change in the amino-acid sequence, A2-milk might be able to prevent or, at least, to minimize the cleavage of &beta, CM7 in calves.

The authors wish to make the following corrections to this paper [...]

IL-36 cytokines are critical regulators of mucosal inflammation and homeostasis. IL-36γ regulates the expression of inflammatory cytokines and antimicrobial proteins by gingival epithelial cells (e.g. TIGK cells). Here, we show that IL-36γ also regulates the expression of matrix metalloproteinase 9 (MMP9) and neutrophil gelatinase-associated lipocalin (NGAL), important mediators of antimicrobial immunity and tissue homeostasis in mucosal epithelia. MMP9 and NGAL were not similarly induced by IL-17 or IL-22, thus indicating the importance of IL-36γ in the regulation of MMP9 and NGAL. Mechanistically, MMP9 and NGAL expression was demonstrated to be induced in an IRAK1- and NF-κB-dependent manner. Furthermore, signaling by p38 MAP kinase may enable their expression to be independently regulated by IL-36γ. The stronger IL-36γ-inducible expression of MMP9 and NGAL in terminally differentiating TIGK cells suggests that control of their expression is associated with the maturation of the gingival epithelium. Although MMP9 and NGAL expression in epithelial cells can also be induced by bacteria, their expression in TIGK cells was not induced by the periodontal pathogen Porphyromonas gingivalis, most likely due to antagonism by the gingipain proteinase virulence factors. This study advances our understanding of how IL-36γ may promote oral mucosal immunity and tissue homeostasis, and how this may be dysregulated by bacterial pathogens.

Interleukin-36 (IL-36) cytokines are important regulators of mucosal homeostasis and inflammation. We have previously established that oral epithelial cells upregulate IL-36γ expression in response to the bacterial pathogen Porphyromonas gingivalis. Here, we have established that IL-36γ can stimulate the gene expression of mechanistically distinct antimicrobial proteins, including the peptidoglycan amidase PGLYRP2, in oral epithelial cells (e.g., TIGK cells). PGLYRP2 gene expression was not stimulated by either IL-17 or IL-22, thus demonstrating selectivity in the regulation of PGLYRP2 by IL-36γ. The IL-36γ-inducible expression of PGLYRP2 was shown to be mediated by IRAK1- and p38 mitogen-activated protein (MAP) kinase-dependent signaling. Furthermore, our finding that IL-36γ-inducible PGLYRP2 expression was reduced in proliferating TIGK cells but increased in terminally differentiating cells suggests that control of PGLYRP2 expression is associated with the maturation of the oral epithelium. PGLYRP2 expression in TIGK cells can also be directly stimulated by oral bacteria. However, the extracellular gingipain proteases (Kgp and RgpA/B) produced by P. gingivalis, which are critical virulence factors, can antagonize PGLYRP2 expression. Thus, the expression of IL-36γ by oral epithelial cells in response to P. gingivalis might enable the subsequent autocrine stimulation of PGLYRP2 expression. In summary, our data identify how IL-36γ may promote oral mucosal homeostasis by regulating PGLYRP2 expression.

A BVD vaccine with novel production technology and adjuvant was introduced to improve immune responses against BVD. However, 5-10% of vaccinated cattle produced pathogenic antibodies, which transferred bovine neonatal pancytopenia (BNP) via colostrum to almost any calf given respective colostrum. Diseased calves had a very high lethality rate of 90%. Dysregulated immune response of dams to the novel vaccine is not fully understood to date. In this study, we clarified by in-depth characterization of immune capacities if the immune response was changed by this particular vaccine or if these cows were prone to a different type of immune response per se. There was a marked difference in the response after polyclonal in vitro stimulation of peripheral blood derived lymphocytes (PBL). BNP dams showed a highly significant hyperproliferation to Pokeweed Mitogen (PWM), Concanavalin A (ConA), Musa paradisiaca banana lectin (BanLec) and interleukin-2 (IL-2). Subsequent differential proteome analyses revealed substantial qualitative differences in immune capacities between both cow groups. PBL of BNP dams reacted with a STAT3 regulated response compared to PBL of controls, which used STAT1 pathway after ConA stimulation. We could detect this immune response pattern also in cows that were never vaccinated at all. Therefore, our data prove that respective BVD vaccination did not cause this deviant immune response, but that a deviant immune phenotype (ID) already existed in cattle and still exists. PBL of these ID cows prefer the IL-2/JAK2/STAT3 pathway for immune responses. Since ID cows additionally have a two-fold increased mastitis incidence, our findings have clinical relevance.

Interferon regulatory factor (IRF) family members impart cell-type specificity to toll-like receptor (TLR) signalling, and we recently identified a role for IRF6 in TLR2 signalling in epithelial cells. TLR3 has a well-characterized role in wound healing in the skin, and here, we examined TLR3-dependent IRF6 functions in human keratinocytes. Primary keratinocytes responded robustly to the TLR3 agonist poly(IC) with upregulation of mRNAs for interferon-β (IFN-β), the interleukin-12 (IL-12) family member IL-23p19 and the chemokines IL-8 and chemokine (C-C motif) ligand 5 (CCL5). Silencing of IRF6 expression enhanced poly(IC)-inducible IFN-β mRNA levels and inhibited poly(IC)-inducible IL-23p19 mRNA expression in primary keratinocytes. Consistent with these data, co-transfection of IRF6 increased poly(IC)-inducible IL-23p19 promoter activity, but inhibited poly(IC)-inducible IFN-β promoter activity in reporter assays. Surprisingly, poly(IC) did not regulate IL-12p40 expression in keratinocytes, suggesting that TLR3-inducible IL-23p19 may have an IL-23-independent function in these cells. The only other IL-12 family member that was strongly poly(IC) inducible was EBI3, which has not been shown to heterodimerize with IL-23p19. Both co-immunoprecipitation and proximity ligation assays revealed that IL-23p19 and EBI3 interact in cells. Co-expression of IL-23p19 and EBI3, as compared with IL-23p19 alone, resulted in increased levels of secreted IL-23p19, implying a functional role for this heterodimer. In summary, we report that IRF6 regulates a subset of TLR3 responses in human keratinocytes, including the production of a novel IL-12 family heterodimer (p19/EBI3). We propose that the TLR3-IRF6-p19/EBI3 axis may regulate keratinocyte and/or immune cell functions in the context of cell damage and wound healing in the skin.

Linear type evaluations of 214 horses of the Old Kladrub breed for 32 traits were used to analyse the effect of variety (gray and black), sex (stallions and mares) and age. The linear model included the fixed effects of variety, sex and age and interactions variety x sex, variety x age and sex x age. Significant differences were found between the varieties in almost two thirds of the traits. The topline was longer and weaker in the black variety than in the gray one. The Shoulder of the gray horses was longer and more sloped than of the black one. The animals of the gray variety had a wider and deeper ehest. The stance of forelimbs and hind limbs was more correct in the gray variety than in the black one. Significant differences between sexes were observed in a few traits. Only 12 of 32 traits showed significant changes with age. In 10 of 32 traits, at least one of the interactions was significant. The estimates of this study characterise the Old Kladrub breed and its varieties in its current state and allow for more accurate evaluation of the horses.

Title of the paper: Relationship between body composition measured by dual energy x-ray absorptiometry and reproduction performance in gilts This study was conducted to evaluate the relationship between body composition and reproduction traits in gilts. A number of 176 gilts of different genotypes were categorized in three different groups. Group 0 consists of gilts that were disqualified for breeding according performance test results or showed no occurrence of oestrus. Group 1 consists of all gilts, which farrowed after an artificially insemination. Group 2 was composed of gilts that were artificially inseminated but did not give birth to a litter. Body composition was determined in vivo using dual energy x-ray absorptiometry at a live weight of about 90 kg. Initial reproduction data from group 1 were also evaluated. Results of the performance test showed considerable variation among the different groups. Group 1 was significantly heavier and the daily gain was significantly higher than in group 0. However, results of breeding index and value were clearly below the results of the other two groups. The body fat content was significantly higher in gilts with a litter (>1.65 absolutely) than in gilts without a litter (group 2) and in disqualified gilts (group 0). Group 2 gilts showed the smallest body fat mass. However, the relation between DXA fat percentage and litter size for group 1 showed in tendency that the litter size might decrease with an increasing fat content of gilts. Therefore, an optimum body fat content for conventional breeding gilts of European or American origin needs to be determined for a maximum fertility.

Rubber flooring in dairy cows has been discussed controversially, because different studies found as well advantages as disadvantages regarding claw health. Until today little is known regarding the effect of rubber flooring on the reproductive performance of dairy cows. Therefore, the study aimed at comparing fertility and the milk yield of dairy cows on concrete or rubber flooring. Days to first breeding (DFB), days open (DO), services per conception (SC), heat detection rate (HDR), conception rate (first service; CR1st service) and the corresponding 100 days in milk (DIM) milk yield were evaluated for 139 lactation periods of 91 cows of the University Munich Livestock Center dairy herd. The cows entered the study as heifers and were housed on the same flooring system during their whole productive life. All cows underwent the same management, feeding, and climatic conditions. A mixed model procedure was used to analyse data. Cows on rubber (RSF) compared with cows on concrete flooring (CSF) showed less DFB (76.8/94.6; P

1. Bone stability of five purebred layer lines differing in performance efficiency and phylogenetic origin was investigated within two housing systems. 2. Hens were kept in single cages or floor pens until the end of the 74 th week of age. At that time, they were sacrificed and examined for humeral and tibial traits. 3. Within a phylogenetic group, hens of the low performing lines had higher bone breaking strengths compared to their high performing counterparts. The bone mineral density of the tibia was significantly lower in the high performing brown egg layers than in the low performing ones. 4. Within and among layer lines, birds in floor housing had significantly higher tibia and humerus breaking strengths as well as higher bone weights than caged hens. 5. The differences between the high and low performing layer lines suggest a negative association between egg laying performance and bone stability in laying hens, irrespective of their phylogenetic origin. Moreover, the results confirm the already known effect of the housing system on bone strength.

Porphyromonas gingivalis is a keystone pathogen in chronic periodontitis. Its expression of gingipain proteases (Kgp and RgpA/B) is central to the stimulation of chronic inflammation. In this study, we investigated the inflammatory response of oral epithelial cells to P. gingivalis. The cells responded by upregulating the expression of the orphan chemokine CXCL14. The stimulation of CXCL14 expression was largely triggered by the gingipain proteases and was dependent on the host protease-activated receptor PAR-3. Significantly, CXCL14 expression was transcriptionally repressed in response to epidermal growth factor (EGF)-induced activation of the MEK-ERK1/2 pathway. P. gingivalis overcomes the repression of CXCL14 via the gingipain protease-mediated degradation of EGF. Therefore, P. gingivalis not only directly stimulates CXCL14 expression via PAR-3 but also promotes its expression by antagonising EGF signalling. In addition to chemotactic activity, some chemokines also have antimicrobial activities. CXCL14 was demonstrated to have bactericidal activity, against commensal oral streptococci associated with health. Notably though, P. gingivalis was not susceptible to killing by CXCL14, potentially because the gingipain proteases can degrade CXCL14. This suggests that the stimulation of dysregulated CXCL14 expression by P. gingivalis may help promote dysbiosis and the development of chronic periodontitis.

Plastic waste is continuously introduced not only into marine, but also freshwater environments, where it fragments into microplastics. Organisms may be affected by the particles themselves due to ingestion and indirectly via incorporated additives such as plasticizers, since these substances have the ability to leach out of the polymer matrix. Although it has been indicated that the likelihood of additives leaching out into the gut lumen of organisms exposed to microplastics is low, studies distinguishing between the effects of the synthetic polymer itself and incorporated additives of the same polymer are scarce. Since this is obligatory for risk assessment, we analyzed the chronic effects of flexible polyvinylchloride (PVC), a widely used polymer, containing the plasticizer diisononylphthalate (DiNP) on morphology and life history of the freshwater crustacean Daphnia magna and compared these effects with the effects of rigid PVC, lacking DiNP, as well as a glass bead control. After up to 31 days of exposure, rigid PVC and glass beads did not affect body length and relative tail spine length of D. magna, whereas flexible PVC led to an increased body length and a reduced number of offspring. None of the treatments increased the mortality significantly. We were able to show that 2.67μg/L DiNP leached out of the flexible PVC into the surrounding medium using GC-MS. Yet, we were not able to measure leachate inside the gut lumen of D. magna. The effects emerged towards the end of the experiment, due to the time dependent process of leaching. Therefore, the results highlight the relevance of long-term chronic exposure experiments, especially as leaching of additives takes time. Further, our study shows the importance to distinguish between microplastics containing leachable additives and the raw polymer in ecotoxicological testing.

A Hordeum bulbosum L. (Poaceae) clone A17 was identified, which showed complete resistance to Barley yellow dwarf virus (BYDV) and Cereal yellow dwarf virus (CYDV). It was not possible to infect plants of A17 with BYDV-PAV, -MAV, or with CYDV-RPV by the aphid vectors Rhopalosiphum padi (L.) or Sitobion avenae (Fabricius) (both Hemiptera: Aphididae). Plants of the A17 clone and of the BYDV-susceptible H. bulbosum clone A21 revealed some resistance to R. padi compared to the susceptible winter barley cultivar Rubina [Hordeum vulgare L. (Poaceae)]. The development time to the imago was longer and the number of nymphs was reduced on both clones compared with cv. Rubina. The probing and feeding behaviour of R. padi on plants of the H. bulbosum clones was studied over 12 h and compared with that on plants of the barley cv. Rubina. Principal component analysis of the results of the feeding behaviour revealed a clear separation of the H. bulbosum genotypes from Rubina. On H. bulbosum the number of penetrations was higher but total feeding time was shorter. Significant differences were mainly found in the phloem feeding parameters for plants of both clones in comparison to Rubina, with the virus resistant A17 clone having the strongest effect and the susceptible A21 clone being intermediate. Most significant differences were found in parameters of the phloem salivation phase. On A17, an average of less than one (0.9) E1 phase per plant was observed (3.3 on A21 and 5.7 on Rubina) and its duration was reduced to less than 1 min (0.9 min) in comparison to 2.4 min on A21 and 5.7 min on Rubina. Also, the phloem feeding (E2) phase was clearly reduced on A17 plants with 0.5 E2 phases per test and a mean duration of 1.1 min in contrast with 2.9 and 3.5 E2 phases per test and 34.1 and 421.3 min for A21 and Rubina, respectively. These results point towards a phloem-localized factor for aphid resistance in H. bulbosum, i.e., on A17 plants the phloem salivation time is too short for a successful infection by BYDV leading to vector resistance.

The distribution of free sterols in a sulfur-rich lacustrine sediment of Miocene age deposited in the ancient crater lake of the Nordlinger Ries (southern Germany) was investigated and compared with the corresponding distributions of esterified and kerogen-bound sterols. The three fractions exhibited the same suite of principal sterols, with 4α,24-dimethyl-5α- cholestan-3β-ol, dinosterol and dinostanol displaying the highest concentrations and relative abundances. The distributions of sterols, steroidal ketones, n-alkanols and isoprenoid alcohols suggest a prevalent deposition of autochthonous aquatic organic matter under saline conditions. In particular, the high abundance of 4α,24-dimethyl-5α-cholestan-3-one, dinosterone and dinostanone and the corresponding 4-methyl sterols indicates the importance of dinoflagellate productivity in this former crater lake and its significant contribution to the sedimentary lipids in the Nordlinger Ries sediment. The similarity in structures and relative abundance of the major 4-methyl sterols and the corresponding steroidal ketones suggest that they are biosynthetically linked.

In addition to their established roles in host defence, Toll-like Receptors (TLRs) have emerging roles in control of homeostasis, injury and wound repair. The dsRNA-sensing receptor, TLR3, has been particularly implicated in such processes in several different tissues including the skin, intestine and liver, as well as in the control of reparative mechanisms in the brain, heart and kidneys, following ischemia reperfusion injury. In this review, we provide an overview of TLR3 signalling and functions in inflammation, tissue damage and repair processes, as well as therapeutic opportunities that may arise in the future from knowledge of such pathways.

The purpose of this study was to use quantitative magnetic resonance (QMR) to measure changes in the body composition of piglets during growth from birth to 4 kg body weight. Using QMR, 60 pigs were scanned an average of 5 times starting at 2.7±1.3 days of age (1.95 kg) and finally at 13.1±4.3 days (4.14 kg). Regression analysis revealed that the rates of total body growth and fat and lean deposition were linear throughout this period. Subsequently, a second group of 235 pigs (109 males and 126 females) were scanned twice, first at 2.7±1.2 days of age and then at 13.4±3.1 days of age. The mean (±SD) rate of total body growth was 230±57 g/day. The rates of fat and lean deposition were 40±13 g/day and 191±52 g/day, respectively. The rates of both fat and lean deposition were highly correlated (P

The growth and composition of the neonatal pig is of interest because of potential impact on subsequent growth and finally, composition at market weight. The purpose of this study was to compare at weaning the growth and body composition of the largest and smallest pigs from each of 38 litters. At weaning (27±1.7 d) the largest (9.3±1.1 kg) and smallest (6.2±1.5 kg) pigs were selected for body composition measurement by dual energy X-ray absorptiometry (DXA). The body composition of the largest pigs consisted of 38 % more fat, 32 % more lean, and 29 % more bone mineral content (P0.05). A second study consisted of 12 pairs of pigs from 8 litters that were selected on the basis of having the same birth weight, but one pig out gaining the other by at least 50 g/day. At 21 days of age the selected pigs were scanned by DXA. For both groups combined, the correlation (r) between body weight and lean mass was 0.99, between body weight and fat mass it was 0.87, and between body weight at birth and body weight at weaning it was 0.56. The results of these studies revealed that, at weaning, the fastest and slowest growing pigs had similar proportions of fat, lean and bone mineral and, consistent with previous results, the rates of both fat and lean deposition were highly correlated (P

Colony stimulating factor-1 (CSF-1) mediates its pleiotropic effects on macrophages through the CSF-1 receptor (CSF-1R), a receptor tyrosine kinase. Current models of CSF-1 signalling imply that the CSF-1R activates signalling pathways exclusively at the plasma membrane and the subsequent internalisation of the CSF-1R simply facilitates its lysosomal degradation in order to prevent on-going signalling. Here, we sought to establish if the CSF-1R may in fact continue to signal following its internalisation. Erk1/2, Akt and Stat3 activation were abrogated when the internalisation of the CSF-1R was impaired, with the effects on Stat3 distinct from those for Erk1/2 and Akt. Pharmacologic inhibition of the CSF-1R following its internalisation resulted in less sustained Erk1/2 and Akt activity, whereas Stat3 activity was unaffected. Significantly, the suppressive effects of the CSF-1R inhibitor on the up-regulation of gene expression by CSF-1 (e.g. cyclin D1 and Bcl-xL gene expression) were comparable irrespective of whether the inhibitor was added prior to CSF-1 stimulation or following the internalisation of the CSF-1R. Similarly, pharmacologic inhibition of Erk1/2 (or Akt) activity either prior to CSF-1 stimulation or subsequent to CSF-1R internalisation had comparable effects on the regulation of gene expression by CSF-1. Together, our data argue that key signalling responses to CSF-1 depend on the ability of the CSF-1R to signal from endosomes following its internalisation, thus adding an important spatiotemporal aspect to CSF-1R signalling.

Thevetia cardiac glycosides can lead to intoxication, thus they are important indicators for forensic and pharmacologic surveys. Six thevetia cardiac glycosides, including two with unknown structures, were isolated from the seeds of the Yellow Oleander (Thevetia peruviana (Pers.) K. Shum., Apocynaceae). LC-ESI⁺-MS(/MS) analysis under high-resolution conditions used as a qualitative survey of the primary glycosides did not lead to fragmentation of the aglycones. Acid hydrolysis of the polar and non-volatile thevetia glycosides under severe conditions yielded the aglycones of the thevetia glycosides and made them amenable to GC-MS analysis. Comparison of mass spectral fragmentation patterns of the aglycones, as well as high-resolution mass spectrometric and NMR data of four of the primary thevetia glycosides including the two unknowns, revealed the structures of the complete set of six thevetia glycosides. The identified compounds are termed thevetin C and acetylthevetin C and differ by an 18,20-oxido-20,22-dihydro functionality from thevetin B and acetylthevetin B, respectively. The absence of an unsaturated lactone ring renders the glycosides cardio-inactive. The procedures developed in this study and the sets of analytical data obtained will be useful for screening and structure assessment of other, particularly polar, cardiac glycosides.

The improvement of carcass quality is one of the main breeding goals in pig production. To select appropriate breeding animals, it is of major concern to exactly and reliably analyze the body composition in vivo . Therefore, the objective of the study was to examine whether the combination of dual-energy X-ray absorptiometry (DXA) and magnetic resonance imaging (MRI) offers the opportunity to reliably analyze quantitative and qualitative body composition characteristics of different pig breeding groups in vivo . In this study, a total of 77 pigs were studied by DXA and MRI at an average age of 154 days. The pigs originated from different autochthonous or conventional breeds or crossbreeds and were grouped into six breed types: Cerdo Iberico (Ib); Duroc × Ib (Du_Ib); White Sow Lines (WSL, including German Landrace and German Large White); Hampshire/Pietrain (Pi_Ha, including Hampshire, Pietrain × Hampshire (PiHa) and Pietrain × PiHa); Pietrain/Duroc (Pi_Du, including Pietrain × Duroc (PiDu) and Pietrain × PiDu); crossbred WSL (PiDu_WSL, including Pietrain × WSL and PiDu × WSL). A whole-body scan was performed by DXA with a GE Lunar DPX-IQ in order to measure the amount and percentage of fat tissue (FM; %FM), lean tissue (LM; %LM) and bone mineral, whereas a Siemens Magnetom Open with a large body coil was used for MRI in the thorax region between 13th and 14th vertebrae in order to measure the area of the loin (LA) and the above back fat area (FA) of both body sides. A GLM procedure using SAS 9.2 was used to analyze the data. As expected, the native breed Ib followed by Du_Ib crossbreeds showed the highest %FM (27.2%, 25.0%) combined with the smallest LA (46.2 cm 2 , 73.6 cm 2 ), whereas Ib had the lowest BW at an average age of 154 days. Pigs with Pi_Ha origin presented the least %FM (12.4%) and largest LA (99.5 cm 2 ). The WSL and PiDu_WSL showed an intermediate body composition. Therefore, it could be concluded that DXA and MRI and especially their combination are very suitable methods to reliably identify differences in body composition and carcass traits among different pig lines in vivo .

Humic substances (HS) constitute 50–80% of total dissolved organic matter (DOM) in freshwaters but in the open ocean 60% of the HA were decomposed. Highest fractions were decomposed in treatments in which the bacterial communities were fed by HA media of a foreign salinity, e.g., of 14 in Exp1 and of 28 in Exp2. Some of the HA material was not decomposed but broken down to smaller moieties entering the fulvic acid (FA) fraction. The analysis of the HA media and their residuals after bacterial decomposition by pyrolysis GC/MS showed that individual organic compounds were decomposed. In Exp1, 32 aliphatic and aromatic compounds, including lignin biomarkers, were detected in the HA medium of which mainly aliphatic compounds were decomposed. In Exp2, 49 compounds were detected of which ~40–60% were not detected any more after bacterial decomposition in the HA fraction but still in the FA fraction. The results show that estuarine and marine bacterial communities can decompose large amounts of HA and that this process is important in reducing the amount of terrestrial HS and DOM entering the estuarine and coastal region.

Mangroves are the dominant type of vegetation along many tropical coasts. Organic matter (OM) derived from mangrove leaf litter and root material is stored in sediments and is a major contributor to the amount and chemical composition of sedimentary OM. A set of organic biomarkers in sediments was applied as a palaeo-indicator for the Holocene dynamics of a mangrove Estuary (Rio Caeté, Pará, Brazil). Six sediment cores were collected perpendicular to the present coast line and analysed for triterpenols and sitosterol. The influence of microbial biomarker degradation was implemented from a previous study. Biomarker profiles were validated with pollen data and multivariate statistics to test whether these compounds were suitable indicators for the palaeo-vegetation. Sediments deposited up to 2 Ma BP showed biomarker assemblages similar to those of recent surface sediment. In two cores, the biomarker composition revealed a transition from marsh to mangrove vegetation. Taraxerol, germanicol and b-amyrin provided the most significant chemotaxonomical information and, especially in combination, served as reliable proxies for OM from Rhizophora mangle in northern Brazil. The maximum age of the mangrove system ranged between 1000 and 5100 yr depending on the topographic elevation of the drilling location. Fil: Koch, Boris Peter. Alfred Wegener Institute for Polar and Marine Research; Alemania. University of Applied Sciences; Alemania Fil: Souza Filho, Pedro. Federal University of Pará; Brasil Fil: Behling, Hermann. University of Göttingen; Alemania Fil: Cohen, Marcelo C. L.. University of Pará; Brasil Fil: Kattner, Gerhard. Alfred Wegener Institute for Polar and Marine Research; Alemania Fil: Rullkötter, Jürgen. Carl von Ossietzky University of Oldenburg; Alemania Fil: Scholz Böttcher, Barbara. Carl von Ossietzky University of Oldenburg; Alemania Fil: Lara, Ruben Jose. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Bahía Blanca. Instituto Argentino de Oceanografía. Universidad Nacional del Sur. Instituto Argentino de Oceanografía; Argentina. Center for Tropical Marine Ecology; Alemania

Osteochondrosis (OC) or leg weakness is an economically important disease of young fast growing pigs and is a concern of animal welfare. The etiology and pathogenesis of osteochondrosis is not fully understood yet, but any abnormalities in the formation of hypertrophic chondrocytes and disrupted blood supply to the growth cartilage are very important predisposing factors. Matrix gla protein (MGP) as a potential calcification inhibitor of extracellular matrix might contribute to the development of OC. Molecular characterization, polymorphisms analysis, methylation at promoter region and expression of MGP gene and protein were performed in both healthy and OC cartilage collected from a Duroc x Pietrain resource population. The porcine MGP gene consists of 4 exons and 3 introns. The full-length MGP cDNA isolated from articular cartilage consists of 606 bp with a 69-bp 5' UTR, a 312-bp open reading frame with a start codon, a 225-bp 3' UTR. Three single-nucleotide polymorphisms (SNP) were detected in the intron 1 (A-115G, C-1073T and C-1135A) and one in the 3'UTR (C-3767T). The relative abundance of MGP mRNA was lower (P