Giardia duodenalis in Wildlife: Exploring Genotype Diversity in Italy and across Europe

Fragmented data are so far available on genotype diversity of G. duodenalis in wildlife in different countries in Europe, in particular, in Italy. In the present study, G. duodenalis sequences obtained from different Italian wild animals [12 porcupines (Hystrix cristata), 4 wild boars (Sus scrofa), 1 wolf (Canis lupus italicus), 6 Alpine chamois (Rupicapra rupicapra rupicapra)] were compared with those available from wild host species in Europe to add new data on the geographic distribution of Giardia assemblages/sub-assemblages and their transmission patterns among natural hosts. Thirty-eight sequences were obtained by MLG analysis (SSU-rRNA, bg, gdh, and tpi genes) and subsequently compared by phylogenetic and network analyses with those from wild species monitored in the last decades in Europe. The results revealed the presence of potentially zoonotic (A-AI, A-AII from wild boar; B from porcupine) and host-adapted (D from wolf; E, A-AIII from chamois) assemblages and sub-assemblages and represent the first report for Italian wild boar. The analysis did not find any evidence of spatial or host segregation for specific genetic variants, mostly shared between different hosts from different European countries. However, conflicting evidence was found in genotypic assignment, advocating for data improvement and new genomic approaches.

Differential Network Testing Reveals Diverging Dynamics of Organ System Interactions for Survivors and Non-survivors in Intensive Care Medicine

Statistical network analyses have become popular in many scientific disciplines, where an important task is to test for differences between two networks. We describe an overall framework for differential network testing procedures that vary regarding (1) the network estimation method, typically based on specific concepts of association, and (2) the network characteristic employed to measure the difference. Using permutation-based tests, our approach is general and applicable to various overall, node-specific or edge-specific network difference characteristics. The methods are implemented in our freely available R software package DNT, along with an R Shiny application. In a study in intensive care medicine, we compare networks based on parameters representing main organ systems to evaluate the prognosis of critically ill patients in the intensive care unit (ICU), using data from the surgical ICU of the University Medical Centre Mannheim, Germany. We specifically consider both cross-sectional comparisons between a non-survivor and a survivor group and longitudinal comparisons at two clinically relevant time points during the ICU stay: first, at admission, and second, at an event stage prior to death in non-survivors or a matching time point in survivors. The non-survivor and the survivor networks do not significantly differ at the admission stage. However, the organ system interactions of the survivors then stabilize at the event stage, revealing significantly more network edges, whereas those of the non-survivors do not. In particular, the liver appears to play a central role for the observed increased connectivity in the survivor network at the event stage.

The build-up of dominance hierarchies in eusocial insects

Reproductive division of labour is a hallmark of eusocial insects. However, its stability can often be hampered by the potential for reproduction by otherwise sterile nest-mates. Dominance hierarchy has a crucial role in some species in regulating which individuals reproduce. Compared with those in vertebrates, the dominance hierarchies in eusocial insects tend to involve many more individuals, and should require additional selective forces unique to them. Here, we provide an overview of a series of studies on dominance hierarchies in eusocial insects. Although reported from diverse eusocial taxa, dominance hierarchies have been extensively studied in paper wasps and ponerine ants. Starting from molecular physiological attributes of individuals, we describe how the emergence of dominance hierarchies can be understood as a kind of self-organizing process through individual memory and local behavioural interactions. The resulting global structures can be captured by using network analyses. Lastly, we argue the adaptive significance of dominance hierarchies from the standpoint of sterile subordinates. Kin selection, underpinned by relatedness between nest-mates, is key to the subordinates' acceptance of their positions in the hierarchies. This article is part of the theme issue ‘The centennial of the pecking order: current state and future prospects for the study of dominance hierarchies’.

Evaluation of CRISPR gene-editing tools in zebrafish

Background Zebrafish have practical features that make them a useful model for higher-throughput tests of gene function using CRISPR/Cas9 editing to create ‘knockout’ models. In particular, the use of G0 mosaic mutants has potential to increase throughput of functional studies significantly but may suffer from transient effects of introducing Cas9 via microinjection. Further, a large number of computational and empirical tools exist to design CRISPR assays but often produce varied predictions across methods leaving uncertainty in choosing an optimal approach for zebrafish studies. Methods To systematically assess accuracy of tool predictions of on- and off-target gene editing, we subjected zebrafish embryos to CRISPR/Cas9 with 50 different guide RNAs (gRNAs) targeting 14 genes. We also investigate potential confounders of G0-based CRISPR screens by assaying control embryos for spurious mutations and altered gene expression. Results We compared our experimental in vivo editing efficiencies in mosaic G0 embryos with those predicted by eight commonly used gRNA design tools and found large discrepancies between methods. Assessing off-target mutations (predicted in silico and in vitro) found that the majority of tested loci had low in vivo frequencies (< 1%). To characterize if commonly used ‘mock’ CRISPR controls (larvae injected with Cas9 enzyme or mRNA with no gRNA) exhibited spurious molecular features that might exacerbate studies of G0 mosaic CRISPR knockout fish, we generated an RNA-seq dataset of various control larvae at 5 days post fertilization. While we found no evidence of spontaneous somatic mutations of injected larvae, we did identify several hundred differentially-expressed genes with high variability between injection types. Network analyses of shared differentially-expressed genes in the ‘mock’ injected larvae implicated a number of key regulators of common metabolic pathways, and gene-ontology analysis revealed connections with response to wounding and cytoskeleton organization, highlighting a potentially lasting effect from the microinjection process that requires further investigation. Conclusion Overall, our results provide a valuable resource for the zebrafish community for the design and execution of CRISPR/Cas9 experiments.

Avian haemosporidian parasites of accipitriform raptors

Background The order Accipitriformes comprises the largest group of birds of prey with 260 species in four families. So far, 21 haemosporidian parasite species have been described from or reported to occur in accipitriform birds. Only five of these parasite species have been characterized molecular genetically. The first part of this study involved molecular genetic screening of accipitriform raptors from Austria and Bosnia-Herzegovina and the first chromogenic in situ hybridization approach targeting parasites in this host group. The aim of the second part of this study was to summarize the CytB sequence data of haemosporidian parasites from accipitriform raptors and to visualize the geographic and host distribution of the lineages. Methods Blood and tissue samples of 183 accipitriform raptors from Austria and Bosnia-Herzegovina were screened for Plasmodium, Haemoproteus and Leucocytozoon parasites by nested PCR, and tissue samples of 23 PCR-positive birds were subjected to chromogenic in situ hybridization using genus-specific probes targeting the parasites’ 18S rRNAs. All published CytB sequence data from accipitriform raptors were analysed, phylogenetic trees were calculated, and DNA haplotype network analyses were performed with sequences from clades featuring multiple lineages detected in this host group. Results Of the 183 raptors from Austria and Bosnia-Herzegovina screened by PCR and sequencing, 80 individuals (44%) were infected with haemosporidian parasites. Among the 39 CytB lineages detected, 18 were found for the first time in the present study. The chromogenic in situ hybridization revealed exo-erythrocytic tissue stages of Leucocytozoon parasites belonging to the Leucocytozoon toddi species group in the kidneys of 14 infected birds. The total number of CytB lineages recorded in accipitriform birds worldwide was 57 for Leucocytozoon, 25 for Plasmodium, and 21 for Haemoproteus. Conclusion The analysis of the DNA haplotype networks allowed identifying numerous distinct groups of lineages, which have not yet been linked to morphospecies, and many of them likely belong to yet undescribed parasite species. Tissue stages of Leucocytozoon parasites developing in accipitriform raptors were discovered and described. The majority of Leucocytozoon and Haemoproteus lineages are specific to this host group, but most Plasmodium lineages were found in birds of other orders. This might indicate local transmission from birds kept at the same facilities (raptor rescue centres and zoos), likely resulting in abortive infections. To clarify the taxonomic and systematic problems, combined morphological and molecular genetic analyses on a wider range of accipitriform host species are needed.

Scheduling by NSGA-II: Review and Bibliometric Analysis

NSGA-II is an evolutionary multi-objective optimization algorithm that has been applied to a wide variety of search and optimization problems since its publication in 2000. This study presents a review and bibliometric analysis of numerous NSGA-II adaptations in addressing scheduling problems. This paper is divided into two parts. The first part discusses the main ideas of scheduling and different evolutionary computation methods for scheduling and provides a review of different scheduling problems, such as production and personnel scheduling. Moreover, a brief comparison of different evolutionary multi-objective optimization algorithms is provided, followed by a summary of state-of-the-art works on the application of NSGA-II in scheduling. The next part presents a detailed bibliometric analysis focusing on NSGA-II for scheduling applications obtained from the Scopus and Web of Science (WoS) databases based on keyword and network analyses that were conducted to identify the most interesting subject fields. Additionally, several criteria are recognized which may advise scholars to find key gaps in the field and develop new approaches in future works. The final sections present a summary and aims for future studies, along with conclusions and a discussion.

Modulation of Host Immune Response during Leishmania infantum Natural Infection: A Whole-Transcriptome Analysis of the Popliteal Lymph Nodes in Dogs

Leishmania infantum, the etiological agent of canine leishmaniosis (CanL) in Europe, was responsible of the largest outbreak of human leishmaniosis in Spain. The parasite infects and survives within myeloid lineage cells, causing a potentially fatal disease if left untreated. The only treatment option relies on chemotherapy, although immunotherapy strategies are being considered as novel approaches to prevent progression of the disease. To this aim, a deeper characterization of the molecular mechanisms behind the immunopathogenesis of leishmaniosis is necessary. Thus, we evaluated, for the first time, the host immune response during L. infantum infection through transcriptome sequencing of the popliteal lymph nodes aspirates of dogs with CanL. Differential expression and weighted gene co-expression network analyses were performed, resulting in the identification of 5,461 differentially expressed genes (DEGs) and four key modules in sick dogs, compared to controls. As expected, defense response was the highest enriched biological process in the DEGs, with six genes related to immune response against pathogens (CHI3L1, SLPI, ACOD1, CCL5, MPO, BPI) included among the ten most expressed genes; and two of the key co-expression modules were associated with regulation of immune response, which also positively correlated with clinical stage and blood monocyte concentration. In particular, sick dogs displayed significant changes in the expression of Th1, Th2, Th17 and Tr1 cytokines (e. g. TNF-α, IFN-γ, IL-21, IL-17, IL-15), markers of T cell and NK cell exhaustion (e. g. LAG3, CD244, Blimp-1, JUN), and B cell, monocyte and macrophage disrupted functionality (e. g. CD40LG, MAPK4, IL-1R, NLRP3, BCMA). In addition, we found an overexpression of XBP1 and some other genes involved in endoplasmic reticulum stress and the IRE1 branch of the unfolded protein response, as well as one co-expression module associated with these processes, which could be induced by L. infantum to prevent host cell apoptosis and modulate inflammation-induced lymphangiogenesis at lymph nodes. Moreover, 21 lncRNAs were differentially expressed in sick dogs, and one key co-expression module was associated with chromatin organization, suggesting that epigenetic mechanisms could also contribute to dampening host immune response during natural L. infantum infection in the lymph nodes of dogs suffering from clinical leishmaniosis.

Expectation violations, expectation change, and expectation persistence: The scientific landscape as revealed by bibliometric network analyses

Expectation violations occur when there is a discrepancy between expected and perceived events or experiences. However, expectations often persist despite disconfirming evidence. Therefore, research on expectation violations, expectation change, and expectation persistence has been conducted in several fields of psychology with wide-ranging theoretical assumptions and empirical considerations. In the present review, we analyzed how these research fields relate to each other via bibliometric network analyses. For this purpose, we conducted a systematic literature search to identify scientific publications on expectation violations, expectation change, and expectation persistence. The literature corpus was then quantitatively analyzed using similarity measures that allow a data-driven classification of publications into groups, revealing their conceptual, theoretical, and empirical commonalities. Our results indicate that many influential publications have focused on finding reactivity measures (e.g., brain activation) to the discrepancy experienced between expectations and outcomes. Furthermore, these measures have been used to assess when and to which degree learning and behavioral adaptation (i.e., expectation change) takes place. We discuss the potential application of these measures for understanding expectation violations in more complex settings (e.g., social interaction) as well as phenomena such as expectation persistence. The goal of this review was to foster interdisciplinarity in psychology, enabling scientists and practitioners to identify new topics, promising empirical approaches and previously neglected variables.

Isoforms of the transcriptional cofactor SIN3 differentially regulate genes necessary for energy metabolism and cell survival

The SIN3 scaffolding protein is a conserved transcriptional regulator known to fine-tune gene expression. In Drosophila, there are two major isoforms of SIN3, SIN3 220 and SIN3 187, which each assemble into multi-subunit histone modifying complexes. The isoforms have distinct developmental expression patterns and non-redundant functions. Gene regulatory network analyses indicate that both isoforms affect genes encoding proteins in pathways such as the cell cycle and cell morphogenesis. Interestingly, the SIN3 187 isoform uniquely regulates a subset of pathways including post-embryonic development, phosphate metabolism and apoptosis. Target genes in the phosphate metabolism pathway include nuclear-encoded mitochondrial genes coding for proteins responsible for oxidative phosphorylation, important for energy metabolism. Here, we investigate the role of SIN3 isoforms in regulating energy metabolism and cell survival genes. We find that ectopic expression of SIN3 187 represses expression of several nuclear-encoded mitochondrial genes affecting production of ATP and generation of reactive oxygen species (ROS). Forced expression of SIN3 187 also activates several pro-apoptotic and represses a few anti-apoptotic genes. In the SIN3 187 expressing cells, these gene expression patterns are accompanied with an increased sensitivity to paraquat-mediated oxidative stress. These findings indicate that SIN3 187 influences the regulation of mitochondrial function, apoptosis and oxidative stress response in ways that are dissimilar from SIN3 220. The data suggest that the distinct SIN3 histone modifying complexes are deployed in different cellular contexts to maintain cellular homeostasis.