Laboratory of Environmental genomics

We aim to understand how the absence of JH affects reproduction.

JH regulates nutritional homeostasis, and the absence of epoxidized JH leads to loss of reproductive capacity in female and male mosquitoes. Transcriptomic analysis of ovaries from adult females of our mutant line and wild-type mosquitoes are currently being used to identify a set of differentially expressed genes that will be functionally tested.

Trajectories of genome evolution in convergently evolving insect lineages

Organisms often evolve similar adaptations to similar environments in the process of convergent evolution, indicating that the trajectory of phenotypic evolution is amenable to predictions. Whether the genomic trajectories of convergent organisms are parallel - and thus predictable in principle - is unclear due to the scarcity of genomic evidence from convergent organismal groups. To answer these questions, we are using a model system consisting of rove beetles (Staphylinidae: Aleocharinae) and scuttle flies (Phoridae) that adapted more than twenty times repeatedly to live in symbiosis with termites. Using phylogenetics, comparative genomics, transcriptomics, and microtomographic reconstructions of phenotypes, we will infer the extent, the timescale, and the hierarchical level of genome sequence evolution at which parallel patterns of genome evolution accompany the evolution of convergent adaptations.
Research in collaboration with Tom Bourguignon (OIST, Japan) and Taisuke Kanao (Yamagata University, Japan).
Funding: 5-year research grant JUNIOR STAR GAČR from the Czech Science Foundation(25M CZK  ~ 1M EUR, 2023-2027).

Evolution of defense in termites

One of the likely key adaptations that led to the evolutionary success of termites is defense. While some termite workers evolved remarkable self-sacrificial chemical defensive strategies, the principal termite colony caste responsible for active defense are the soldiers. Termite soldiers typically combine their capacity to produce chemical secretions (Jirošová et al. 2017, Bourguignon et al. 2016) and mechanical weapons in the form of their mandibles (Beránková et al. 2022). One remarkable mandibular defense strategy - so called mandibular snapping - presumably evolved many times independently in termites. In this strategy, soldiers of some termite species evolved mandibles that lost the biting ability and instead adapted into springs that can deliver a lethal strike to insect competitors of termites. Remarkably, this highly derived mandibular strategy apparently evolved multiple times convergently. Currently, we are conducting a comparative evolutionary study across termite lineages to uncover the evolutionary trajectories that lead to mandibular snapping.
Research started in the labs of Tom Bourguignon (OIST, Japan) and Jan Šobotník (CULS, Czechia).

Funding: 2-year research grant KAKENHI from the Japan Society for the Promotion of Science (2023-2024).

TIMPAOMP project - The IMPortance of Anaerobic Oxidation of Methane in Peatlands

Financial suppport: Czech Science Foundation (GACR); Duration: 2023-2025

Principal Investigator: Vojtech Tlaskal

Team members: Renata Kruzikova

Ecosystem management of natural wetlands is an important tool of human land use practises that often have untargeted impacts on greenhouse gas (GHG) emissions such as methane. Systems such as northern peatlands are very sensitive to management practises and at the same time have the potential to release large amounts of GHGs due to their large storage of organic matter. As a result of practised clear-cutting of peatland forests the water table rises. At the same time, anoxic conditions in peat are expanding, creating a new niche for microbial greenhouse gas producers, but also for anaerobic methanotrophs that depend on substantial methane production and the presence of oxidised electron acceptors. There is a lack of knowledge about the proportion of anaerobically oxidised methane, the identity of the bacteria and archaea that carry out such process, their electron acceptors and nitrogen sources in peatlands. Filling these knowledge gaps, which are urgently needed to improve ecosystem models and reduce GHG emissions, is the subject of this project.

Side effects of transgenic plants

The aim of this project is to evaluate the effects of genetically modified maize on the biotic components of an agroecosystem. The study will be conducted with maize varietiety MON 88017, which tolerate the herbicide glyphosate and are resistant to rootworm infestation. This pest, which was introduced into the EU from the USA, is spreading rapidly and has taken over some of the country's maize-growing areas. Current chemical and biological control methods are unsatisfactory, so genetically modified maize is being considered as an alternative, provided no adverse effects on non-target organisms are identified. Herbicide-tolerant plants dominate the market for genetically modified plants, but their cultivation is not yet permitted in the EU.

PI: Oxana Skoková Habuštová

Integrated protection against vectors of viral diseases in seed potatoes and other crops

The project is carried out in cooperation with the Czech University of Agriculture in Prague, the Research Institute of Plant Production in Prague, the Potato Research Institute in Havlíčkův Brod and the Potato Farm in Havlíčkův Brod and is funded by the National Agency for Agricultural Research (NAZV).

The project deals with the regulation of potato virus vectors in the Czech Republic, such as peach aphids, hop aphids and buckthorn-potato aphid. The project offers a very comprehensive solution for the control of aphids. Due to the increasing resistance of aphids to the preparations previously used in Europe, a change of active ingredients is recommended.

The aim of the project led by Biology Centre Institute of Entomology is therefore to develop and test the efficacy of new biological preparations against viral vectors based on EPF such as Lecanicillium sp., Beauveria bassiana, Metarhizium anisopliae, Isaria sp. and Nomuraea rileyi, testing secondary metabolites from EPN, environmentally friendly botanical pesticides that also have anti-feeding and repellent effects. The project can thus support the selection and registration of new effective preparations.

PI: Oxana Skoková Habuštová

Endosymbionts of trypanosomatids and diplonemids

Although most trypanosomatid flagellates do not harbor endosymbiotic bacteria, members of the genera Angomonas, Strigomonas, Novymonas and Kentomonas do so. They acquired their endosymbionts by at least two independent events and seem to obtain from them various macromolecular precursors and vitamins. Diplonemids (for more information see other project abstract) contain numerous endosymbionts obtained by multiple independent (and probably repeated) acquisitions, but nothing is known about their metabolic interaction with their hosts. However, we predict that in both trypanosomatids and diplonemids, endosymbiotic bacteria have significant impact on their hosts, including their behavior, life cycle progression and ecological functions.

We are dissecting the relationship between the eukaryotic host and its bacterial symbiont using protein tagging and functional genomics. We have already identified in Novymonas a protein that is critical for the intracellular distribution of bacteria. In diplonemids, we succeeded to experimentally remove endosymbionts from the host cells and reinfect them. We believe that the protist-bacterium systems that are established in our laboratory will allow us fine dissection of their intricate relationships.

Population genomics of speciation and adaptation in parasites

We study genomic imprints of speciation and adaptation processes in two parasitic species, the tapeworm Ligula intestinalis and the bedbug Cimex lectularius. Despite differences in their ecology and evolution, both species show wide geographical range and many oportunities for emergence of specific adaptations (associated with host specificity in the tapeworm, or with insecticide resistance in the bedbug). Using SNP profiling, transcriptomis and WG sequencing we're evaluating the impact of geographic structuring and gene flow on adaptive processes in the two parasites (P.I.: J.Štefka).

Effect of benign helminth on intestinal inflammatory diseases

Helminths (or worms) are regarded as master manipulators of host immunity. Over the centuries of co-evolution with host organisms (including humans), they have developed a number of sophisticated mechanisms that divert or redirect the host's immune response, particularly to stop inflammatory mechanisms leading to their direct destruction. Despite the fact that many helminths are pathogenic to humans (under certain conditions), there is a new evidence that total loss of these organisms has detrimental consequences for human health. Epidemiological studies have shown that the absence of helminths in humans correlates with the worldwide increase in Chronic Inflammation-Associated Diseases (CIADs) in industrialized countries. CIADs are a group of diseases that have common, ostensibly unrelated, risk factors and occur predominantly in modern societies across the world (e.g., Crohn's disease, multiple sclerosis, various allergies, etc.). Recent intensive research on human gut microbiome has shown that reintroduction of helminths into the intestinal ecosystem appears to be a promising approach in the treatment of CIADs. The principle is to use controlled exposure to non-pathogenic or mildly pathogenic worms to activate anti-inflammatory immune responses. Our laboratory focuses on testing helminth, rat tapeworm (Hymenolepis diminuta), which host colonizations are mostly asymptomatic. We have recently demonstrated a beneficial effect of colonization of Hymenolepis on intestinal inflammation in an experimental model (Jirků-Pomajbíková et al., 2018, Parasitol). Despite testing in clinical trials and experimental systems, the study of helminth therapy reaches a milestone where it is necessary to proceed several other research directions that are necessary for its subsequent and successful development. One of these approaches is to investigate interactions in the ‘host-helminth complex’ together with defining conditions and factors affecting the potency of worm anti-inflammatory effect. The aim of this project is therefore to define the influence of dietary composition (and other specific factors as well as their combinations) on the interactions between the host and H. diminuta, which can influence its efficacy for therapy in selected inflammatory diseases. This tapeworm is at the forefront of helminth therapy research and is considered to be a very suitable therapeutic candidate. The proposed project is solved in close collaboration with William Parker leading the laboratory at the Duke University (NC, USA), which is focused on studying of the effect of H. diminuta on neuropsychiatric disorders (e.g., autism).

*Chronic inflammation-associated diseases (CIADs): refers to a broad class of diseases and conditions caused by chronic inflammation. Accompanied by immune dysregulation, this group of diseases and conditions includes allergies, a range of autoimmune conditions, some neuropsychiatric disorders, and some digestive diseases.

Systems biology approaches to better understand disease vector lifecycles

(a) I. ricinus transmits Borrelia burgdorferi (Lyme disease) and several other arboviral diseases via the salivary glands. We used massive de novo sequencing to characterize the transcriptional dynamics of the salivary and midgut tissues of nymphal and adult I. ricinus at various time points after attachment on the vertebrate host (Kotsyfakis et al., 2015, Sci Rep. 5, 9103). A number of gene family members showed stage- and time-specific expression, with 34 candidate histone-modifying proteins suggesting epigenetic control. Midgut transcriptome analysis revealed several enzymes associated with protein, carbohydrate, and lipid digestion, transporters and channels associated with nutrient uptake, and immune-related transcripts including AMPs.

(b) We identified hemocyte transcripts from semi-engorged female ticks by mass sequencing a hemocyte cDNA library and annotating immune-related transcripts based on their hemocyte abundance and their ubiquitous distribution (Kotsyfakis et al., 2015, PLoS Negl Trop Dis. 9, e0003754). 327 transcripts were significantly overexpressed in the hemocyte libraries, including those coding for scavenger receptors, AMPs, pathogen recognition proteins, proteases, protease inhibitors, lipid metabolism, and immune function. The coding sequences deposited to GenBank significantly increased the publicly available dataset now supporting biochemical, pathophysiological, and translational studies. These gene expression analyses (Chmelař et al., 2016a, Trends Parasitol. 32, 242–254) represent a milestone in the field, and the application of quantitative proteomics to ticks with unknown genomes has provided deeper insights into the molecular mechanisms underlying tick-host-pathogen interactions.

Tripartite interactions between the disease vector/vector-borne pathogens and the host

Noncoding RNAs (ncRNAs) are now known to be (I) transmitted by the vector to possibly modulate vertebrate host responses and (II) regulated in the host by parasites to favor parasite survival. We provided an overview of the involvement of ncRNAs in the parasite-vector-host triad and their effect on host homeostasis (Bensaoud et al., 2019, Trends Parasitol. 35, 715–724). We also reviewed the role that nanovesicles play during pathogen spread, host colonization, and disease pathogenesis (Chávez et al., 2019, J Cell Sci. 132, jcs224212), focusing on a potential role for arthropod exosomes in this tripartite interaction, especially as carriers of long non-coding (lnc)RNAs (Hackenberg and Kotsyfakis, 2018, Trends Parasitol. 34, 549-552).

Diverge or converge: do harsh abiotic condition restrict divergence of defences in alpine willows?

Plants employ a bewildering diversity of defences to withstand herbivory pressure and detrimental environmental conditions. Exploring the factors that drive the variability in plant defensive strategies is crucial for understanding plant evolution in face of herbivory and harsh environment. Within this project we aim at addressing the question: How the interplay between herbivory and environmental pressures form the evolution of plant chemical defences? To achieve this, we combine state of the art metabolomics and bioinformatics combined with field sampling of highland and lowland species of willows. The core of the project includes analysing macroevolutionary patterns in willow defences and comparing them between highland and lowland species. This project will show that changes along elevational gradients strongly affect the way how host-plant defences evolve, directing the prevalent macroevolutionary trends among related willow species (Figure 2).

Figure 2. Proposed elevational trends in plant defences. Higher biomass costs at high elevations drive an increase in plant defences and support defensive stragies and evolutionary trends alternative to the ones shown by lowland plants.

Multi-taxa responses of tropical Lepidoptera to climate changes and anomalies on Barro Colorado Island in Panama

Climate changes represent one of the major causes of global biodiversity decline with species attrition and plausible dramatic scenarios for terrestrial biodiversity concentrated in the tropics. Extreme droughts and disruption in rainfall have been affecting species composition and productivity among multiple organisms. However, we still have little understanding of the effects of climate changes on terrestrial organisms responsible for the maintenance of crucial ecosystem services, such as insect herbivores. Our main objective seeks to understand the multiple responses of tropical Lepidoptera to ongoing and predicted climate changes. We predict contrasting changes in ten phylogenetically and ecologically Lepidoptera taxa and study their past and future directional responses to extreme events of climate that occurred during 2009-2020. We will compare species responses using three different analytical approaches and models to explore the best choice of target model species for monitoring programs and studies of climate change. Our study system also includes reproducible field protocols, laboratory experiments (thermal and desiccation tolerance) and data modelling.

Duration: 01-01-2019 to 01.06.2022

Connections between maintenance of diversity and variation in nature

How do species coexist in diverse ecological networks despite competition, predation and parasitism? This is the most enigmatic question in community ecology, with an analogy at the population level: How is genetic variation maintained within species despite selection and drift? These twin questions are typically studied separately, yet diversity and variation are likely interdependent, with important consequences for basic understanding and biodiversity conservation.

ERC-CZ grant (2020 - 2025)

Innovation of integrated potato protection against the Colorado potato beetle based on new knowledge of genetic and biological characteristics

Colorado potato beetle (CPB) is our main pest in the above-ground mass of potatoes with the potential for crop losses up to 50%. However, protection against it is currently very difficult due to insecticide resistance. In addition to the use of chemical protection and controversial genetic modifications, the development and use of biological protective methods are becoming increasingly promising. Use of entomopathogenic fungi (EPF) and parasitic nematodes (EPN) for protection does not cause a pest resistance. A great advantage is the support in use of mycoinsecticides by sustainable and organic farming programs. Apart from conservation studies, molecular and genetics methods are used wordwide to address the invasive and genetic potential of CPB. Project NAZV ZEMĚ 3 č. QK1910270; PI: Oxana Skoková Habuštová.

Protective measures against the double spined bark beetle, Ips duplicatus

The aim of the project is to fill in gaps in the knowledge of biology and propose effective methods of management of a new pest of spruce monocultures in the Czech Republic, the double spined bark beetle. Field experiments will verify the effectiveness of new protective measures such as standing poisoned trap trees prepared by surface spraying and/or microinjection of insecticides as well as their impact on non-target arthropod species. Due to the unknown abundance of the double spined bark beetle, it is necessary to clarify the actual proportion of timber infested by this pest. Without such knowledge no precise calculation of the number of protective measures can be made. Project TAČR ZÉTA No. TJ02000025; PI: Markéta Davídková.

Selected aspects of bionomy of the large pine weevil and their importance in forest protection

The large pine weevil is a significant pest of coniferous plantations in the Czech Republic. Adults live up to five years, while life expectancy, fertility and rate of development depend on the species of host tree in which adults developed. The aim of the project is to provide the above-mentioned characteristics for the most important domestic species of conifers, as well as gain information on the mechanisms of overwintering and cold resistance of larvae and adults. Special attention will be given to the effectiveness of various mechanical methods of seedling protection available on the market. Based on the results, protective measures against this pest and the species composition of seedlings planted on clearcuts will be adjusted. Project supported by Grant Agency of the Forests of the Czech Republic, state enterprises, 08/2017; PI: Petr Doležal.

Endocrine control of stress situations

The goal of the project is to better understand the physiological and biochemical processes that insects' bodies go through in response to stress. These processes are triggered by peptides from the adipokinetic hormone (AKH) family. The study focuses on the function of AKH in the defence response to pathogens like entomopathogenic nematodes and fungi, as well as against naturally occurring toxins like the venom of the honey bee Apis mellifera and the parasitic wasp Habrobracon hebetor. Additionally, the project involves examining the mechanisms by which AKHs encourage pathogens to intensify their negative effects on the host organism. The emphasis is placed (1) on the study of anti-oxidative stress reactions, including their biochemical mechanism of action, (2) on the analysis of ultrastructural changes in insect tissues (muscles) after the application of toxins, and (3) on the function of enzymes, nutrients, and other metabolites in stressful situations that disrupt the insect organism's homeostasis.

PI: Dalibor Kodrík

Mutualistic dynamics

Many mutualisms rely on inter-specific resource-service exchanges (e.g., pollination, ant fungus gardens, cleaning mutualisms). Our goal is to study how resource-based mutualisms embed within food webs, and their effects on the dynamics and diversity of ecological communities. This is accomplished by modelling of the dynamics of resource production (e.g., nectar, pollen, honeydew) and the effects of their consumption on the dynamics of producers and consumers (the mutualists), as well as third parties (e.g., competitors, herbivores, predators). Adaptability (e.g., by behavior or evolution) in the provision of resources and services is also considered. This research can advance our understanding natural communities, and the consequences of species invasions and management.

Applications

Within this long-term project we study primarily the possibilities of the use of entomopathogenic and molluscoparasitic nematodes in biocontrol. Recently, we assessed the possibility of the use of entomopathogenic nematodes for the control of the bulb mite Rhizoglyphus robinii. This study has shown that EPNs can invade and kill bulb mite but the mortality is not sufficient for the use of EPNs against these pests. In similar way and with similar results we investigated whether the slug parasitic nematode Alloionema appendiculatum could be used in the control of noxious slugs Arion spp. Presently we investigate the possibility of EPN use against Colorado potato beetle and bark beetles.  Within this project we also identified the method of DNA extraction for taxonomic identification of early developmental stages of forensically important flies (Calliphoridae and Sarcophagidae). It was adopted for the routine work at the Department of forensics of the Institute of Criminalistics, Prague. Important publications: Nermuť et al, 2019a; Nermuť et al. 2019b; Olekšáková et al., 2018.

Impact of combined stressors on freshwater biota

We aim to unravel the combined impacts of anthropogenic chemical pollution and climate change on freshwater ecosystems, using a combination of laboratory and mesocosm experiments focusing on the biota of small standing waters dominated by freshwater invertebrates. We currently lack data and unified framework to predict responses of freshwater ecosystems to these combined stressors. To fill this knowledge gap, we will use laboratory and mesocosm experiments to understand how commonly found pharmaceutically active compounds (PhACs) and pesticides in combination with warming affect individuals, trophic interactions, whole ecosystems and energy flow in communities in small standing waters.

Research team: D. Boukal (PI), V. Kolář, A. Csercsa
External collaborators: C. Duchet (Faculty of Science, University of South Bohemia), T. Randák, K. Grabicová (both Faculty of Fisheries and Protection of Waters, University of South Bohemia), R. Stoks (KU Leuven)

Freshwater ectotherms under climate change: the role of phenotypic plasticity in life histories and trophic interactions

Understanding climate change impacts on ecosystems is one of the most important challenges to current ecology and conservation biology. While many freshwater ectotherms are particularly vulnerable to rising temperatures due to their limited dispersal ability, they may mitigate the impact of thermal stress by plastic responses. However, the role of phenotypic plasticity in the context of climate change is little understood. Using laboratory experiments and numerical simulations, we examine the role of individual thermal performance curves and thermally induced plasticity in life histories and species interactions of freshwater ectotherms under the anticipated climate change.

Research team: D. Boukal (PI), A. Csercsa, S. Dijoux
External collaborators: J. Mocq, J. Okrouhlík (Faculty of Science, University of South Bohemia), L. Gvoždík (Institute of Vertebrate Biology CAS), A. Sentis (University of Aix-Marseille)

Larval specializations, bionomics, and habitat shifts in two selected orders of Insecta during the Mesozoic

Ontogenetic diet and habitat shifts from larvae to adults allow them to efficiently explore resources and minimize intraspecific competition. This evolutionary advantage helped insects become the most speciose group of organisms on Earth. However, larval adaptation of fossil insects are insufficiently explored. We will focus on new material of Mesozoic insects. This crucial period for insect evolution was characterized by accelerated diversification in some lineages connected with profound changes in the palaeoclimate and palaeoecosystems. As model taxa, we will use aquatic Ephemeroptera and terrestrial polyphagan Coleoptera that cover a wide range of newly established palaeohabitats. This approach will allow us to track habitat shifts and stases in their evolutionary history.

Research team: P. Sroka, R. Godunko
External collaborators: J. Prokop (PI), J. Batelka, K. Rosová (Charles University, Prague), A.H. Staniczek (Stuttgart State Museum of Natural History)

Biogeography and diversification of Neotropical biodiversity

The American tropics, or the Neotropics, is a biogeographic region home of unique animal and plant species. The rich diversity of life, forms, and genes found in this region goes hand in hand with the variety of climate, landscapes, and elevations. A long-standing question, since the time of early naturalists, is still unresolved: is the unique and rich biodiversity of the Neotropics the result of environmental complexity?

We use phylogenomic datasets to infer robust species trees, which are critical to resolve evolutionary relations, estimate divergence times, and study the tempo and mode of species diversification. Furthermore, the phylogenomic trees provide information on how regions become sources or sinks of diversity by means of dispersal.

Ecological and population genomics

Species and their genomes are under constant interaction with the environment. The responses of genomes, at local and regional scales, can be quantified using efficient tools such as high-throughput DNA sequencing. We aim to understand how past changes in the landscape and climates have shaped the diversity and distribution of populations.

We study local adaptation and signatures of selection in the genomes of butterflies. We estimate how the Pleistocene climatic fluctuations have modulated the structure and gene flow of populations across geographic regions. We also study genomic islands of divergence among sympatric species with high morphological resemblance that are either closely related (sister species) or evolutionarily distant.

Seasonal effects on telomerase activity and aging in honeybees

In honeybees, the rate of aging is modulated through social interactions and seasonal effects. Winter generation workers, which hatch at the end of summer, have remarkably extended lifespans as an adaptation to the cold season when the resources required for the growth and reproduction of colonies are limited and the bees need to maintain the colony until the next spring. In contrast, the summer bees only live for several weeks. To better understand the lifespan differences between summer and winter bees, we studied the fat bodies of honeybee workers and identified several parameters that fluctuate in a season-dependent manner. We reported that telomerase activity, DNA replication, fat body mass, and vitellogenin levels gradually increased as the winter season proceeded (Figure 2). Our study showed that although the fat bodies of winter bees differ significantly from those of summer bees, these differences are not a priori set when bees hatch at the end of summer or in early autumn but instead gradually evolve throughout the season, depending on environmental factors (Koubová et al. (2021) Scientific Reports 11:1-11).

Currently, we investigate effects of various environmental factors on honeybee aging in more detail.

Mapping of Butterflies of the Czech Republic

Butterfly fauna of the Czech Republic consists of 161 species. As many as 18 have already been lost, and about half is endangered. See lepidoptera.cz for National recording scheme, coordinated by our member Jiri Benes.

HYDROLYZE: Innovative utilization of hydrolysate from animal waste for improvement of agricultural soil quality

Technology Agency of the Czech Republic (TA ČR), principal investigator, 2023-2025

The project aim is to test a new type of bioadditive, hydrolysate from animal waste, in order to protect physical, chemical, and biological properties of agricultural soils and decrease greenhouse gases emissions. The main aim is to test the efficiency and longevity of hydrolysate phase and dose in improving the soil quality and health. The secondary aim is to verify the efficiency of hydrolysate in the stabilization of plant biomass from harvesting remnants in the soil and in decreasing greenhouse gases emissions from the soil. All this will be tested considering the type of soil and crop ensuring their sustainability, maximum water retention, plenty of nutrients and microbial communities.

under construction: the linden bug:

Pyrrhocoris apterus as a new model species

The linden, Pyrrhocoris apterus, served as an excellent model of classical insect endocrinology for more than 50 years. However, neither functional genetics, nor genomic tools were available. Our goal is to introduce these techniques to P. apterus and establish this insect species as a model organism with full palette of experimental tools, especially:

• Reverse genetics: Systemic RNA interference is routinely used for gene knock down in P. apterus (see Smykal et al. 2020, Bajgar et al., 2013a; Smykal et al., 2014; Buresova et al., 2016). To create either complete null mutants or specifically modify gene of interest, we established genome editing in P. apterus (Kotwica-Rolinska et al. 2019). Or first unique mutants are phenotypically characterized.

• Neuropeptide inventory: P. apterus served as a model for insect endocrinology, yet, its neuropeptide repertoire is only very poorly known. Therefore we prospected the genome and transcriptome for genes and transcripts coding for neuropeptides and their receptors. The physical existence of processed peptides is further confirmed by MALDI-TOF-MS/MS in collaboration with Dr. Christian Wegener (Wurzburg University), and anatomical localization is assessed with ICC experiments. See Aska's paper on a new insect neuropeptide (Kotwica-Rolinska et al.).

• Microsurgery: The size and anatomy of P. apterus allows for fine microsurgical operations, such as removing Corpus Allatum (P. apterus has only one gland), extirpation of pars intercerebralis region (Bajgar et al., 2013b), or eye removal.

Pathogesis and therapy of emerging viral diseases

(The National Institute for Virology nad Bacteriology, 2022-2025, coordinator for BC: Daniel Růžek)

MYXOZOA

Myxozoans are diverse group of cnidarian parasites, that have been morphologically highly reduced as an adaptation to parasitism and that have conquered a range of different aquatic habitats. Myxozoans have a complex life cycle involving a vertebrate (fish) and an invertebrate (bryozoan, polychaete or oligochaete) host and some species cause important losses in the aquaculture sector. We conduct a wide range of studies on myxozoans:

Functional Genomics of ticks

• RNA interference (RNAi): This well-established technique allows us to silence specific transcripts in ticks, revealing the critical roles their protein products in their biology.
• Ex vivo membrane feeding with protein inhibitors: This method enables us to deliver enzyme inhibitors and receptor blockers to ticks while they feed, providing insights into the functions of specific proteins and pathways within the context of blood feeding.
• Laboratory models for tick-borne diseases: We have set up in our laboratory, complete transmission models for Borrelia, Babesia, and Anaplasma infections. We use them mainly to test the involvement of tick candidate genes in tick-pathogen interactions using the RNAi method.

• Tick innate immunity: We explore how ticks defend themselves against pathogens and other threats.
• Haemoglobin and iron biology: We unravel the intricate processes by which ticks manage ingested blood components like haem or iron from host blood macromolecules.
• Proteolytic systems and protease inhibitors of ticks: We examine the enzymes involved in breaking down blood proteins in the tick's digestive system and enzymes or inhibitors secreted by the salivary glands to facilitate feeding.
• Chitin biology: We study chitin, a key component of the tick's exoskeleton, and its role in tick development and molting and transmission of pathogens.

Queuosine: The role of an essential tRNA modification in parasitic protist Trypanosoma brucei

Transfer RNAs are typical for the large number of posttranscriptional modifications. Most of the tRNA modifications are present in the anticodon loop, which have crucial role in proper translation of proteins. Queuosine is one of the most complex tRNA modifications. Despite its omnipresence among bacteria and eukaryotes, role of queuosine tRNA modification is not clear. The main aim of this project is to evaluate the function and subunit composition of the enzyme responsible for queuosine formation in T. brucei. Using the RNAi knock-down strategy we want to address the principal question regarding the role of queuosine tRNA modification with respect to biology and physiology of this protozoan parasite. 

MitoSignal: Determining signaling mechanisms that drive cellular differentiation of Trypanosoma brucei

Mitochondria perform three essential functions: ATP production, metabolite synthesis and cellular signaling. These signals, communicating the bioenergetic and biosynthetic fitness of the organelle to the nucleus, play a powerful role in determining cellular fate.

Our lab focuses on incorporation of mitochondrial reactive oxygen species (mROS) in cellular signaling. We utilize the unicellular parasites, Trypanosoma brucei and T. congolense, as simplified but elegant models to define mROS-driven cellular differentiation. As these protists undergo programmed development between several distinct life cycle forms, there are striking changes to the structure and physiology of their single mitochondrion that manifest in elevated ROS levels. Importantly, we demonstrated that these ROS molecules are essential for the developmental progression of the parasite. Employing these well-chosen models and combining next-generation biosensors, advanced bioenergetic methods, redox proteomics and a CRISPR/Cas9 genetic screen, we are answering the following fundamental questions: Does mROS drive Trypanosoma cellular differentiation? What molecular processes are responsible for the elevated mROS levels during differentiation? How is the redox signal propagated to the rest of the cell? Our research aspires to unravel the fundamental mechanisms underlying the intricate communication between mitochondria and the rest of the cell, featuring cellular hallmarks of cell fate decision.

AQUAPARA-OMICS: Aquatic parasitism meets biomics – addressing key biological questions using novel datasets and modern analytical tools.

Parasitism is one of the most successful lifestyles of living organisms. Despite the majority of parasites being aquatic, at least during part of their life cycle, comprehensive molecular datasets for them are still scarce. AQUAPARA-OMICS aims to explore the uncharted biodiversity of greatly undersampled groups of aquatic parasites such as the Myxozoa and Dicyemida, and monitor selected human and fish pathogens, by eDNA metabarcoding. Production of new genomic and transcriptomic datasets of basal neodermates, parasitic cnidarians and dicyemids will allow us to unveil their parasitic origin, evolutionary history and characterize unique and shared genomic traits. AQUAPARA-OMICS further aims to explore key genes and molecular mechanisms involved in parasite development, replication, survival strategies and host immune responses. This includes basic research related to the functional characterisation of candidate molecules for future studies on new therapies and vaccine developments, which are urgently required for emerging aquatic pathogens.

Czech Science Foundation – projects of excellence in basic research (EXPRO) (19-28399X; P.I.: T. Scholz; 2019–2023).

Chromerid algae

Chromerids Chromera velia and Vitrella brassicaformis, alveolate algae isolated from Australian stony corals, are the main objects of our scientific interest (Moore & Oborník et al., 2008; Oborník et al., 2012). These algae are believed to represent the closest known phototrophic relatives to apicomplexan parasites (Moore & Oborník et al., 2008; Oborník et al., 2009; Janouškovec et al., 2010; Woo et al, 2015). We have so far studied chromerid ultrastructure, morphology, life cycles (Oborník et al., 2011, 2012), plastid genomes of both algae (Janouškovec et al., 2010) as well as their mitochondrial genomes (Flegontov et al., 2015). We have participated on the sequencing of nuclear genomes of chromerids as well (Woo et al., 2015). We suppose to continue with this research in the future.

Targeted mutagenesis in insects

Targeted mutagenesis is one of the key methods for functional gene analysis. Two methods of gene targeting were established for Drosophila by Rong and Golic (2000) and Bibikova et al. (2002). The first one is based on a pair of site-specific DNA modifying enzymes from yeast, a recombinase and endonuclease that release a linear DNA fragment containing a modified sequence of the target gene in primordial germ cells. The other method is based on custom designed zinc finger nucleases (ZFNs), which are chimeric enzymes consisting of a zinc finger DNA recognition domain and the nonspecific nuclease domain of the Fok I restriction enzyme (Kimet al. 1996).

We pioneered the first method in Drosophila by knockouting the entire gene family using homologous recombination. We also evaluated a simplified variant of the second method and used direct microinjection of custom-designed Zinc Finger Nuclease (ZFN) mRNAs into silkworm embryos. We have a world priority of using ZFNs to generate germline mutations in an insect species other than Drosophila.

Stress physiology of honey bees

The project focuses on the physiological processes that occur in bees when defensive anti-stress responses are triggered following intoxication with synthetic or natural toxins. The project concentrates on the investigation of vibroacoustic signals, their artificial intelligence-based analysis, and the correlation between these signals and biochemical traits like the level of nutrients, the activity of digestive enzymes, the level of oxidative stress, the composition of bee venom components, the level of stress hormones, the amount of vitellogenin in the haemolymph, and the amount of biogenic amines in the body of bees. The project's goal is to understand the physiological and pathological phenomena that occur in the bee's body during stressful situations and, if necessary, to apply them in practical beekeeping.

New approaches to metabolite profiling in biological samples of animal origin

Multicomplex analysis of low molecular weight substances of diagnostic value has been a challenge in current metabolite research. This approach is promissing to getting an overview of the metabolic status and global biochemical events associated with a cellular or biological system. Our research aims are focused on high-end sample preparation and separaration procedures enabling comprehensive profiling of diagnostic compounds in complex biological, industrial and environmental matrices. Design and investigation of novel haloformates and their implementation in rapid, fast and reproducible analytical protocols have result in a comprehensive portfolio of unique analytical methods for a wide range of compound classes involving polyols, lipids, amines, organic acids, small peptides and steroids. The developed analytical protocols have been integrated with a house-made Metabolite Mapper tool enabling automated peak detection, metabolite library searching, component matrix formation and data exports for further data mining and statistical processing.

New alternative approaches in pest control of the horse-chestnut leafminer Cameraria ohridella

New data on applicability of entomopathogenic fungi against important insect pest of the horse-chestnut leafminer Cameraria ohridella will be obtained. It will be verified if the fungi have any negative effects on biodiversity of the leafminer parasitoids.

Phylogeny of butterflies as a tool for understanding of evolution of Ecological phenomena

Butterflies rank among best known groups of insects. Life history, habitat requirements,
seasonal occurrence and other details are known for most of European species, but many
phenomena are tractable only with understanding the evolutionary history of entire
monophyletic groups, typically genera. Our phylogeny analyses contributed to elucidating the
evolution of seasonal polyphenism of European map butterfly (Fric et al. 2004, J. Evol. Biol.),
the evolution of parasitic myrmecophily in Large blues (Pech et al. 2004, Cladistics; Fric et
al. 2007, Syst. Entomol.). Ongoing research targets the complex radiation patterns in Erebia
satyrines, dwellers of northern and mountain areas of Northern hemisphere.

SOMForClim: Soil organic matter fractions and soil carbon storage as affected by forest type and climate change

Czech Science Foundation (CSF), principal investigator, 2022-2024

Temperate forests hold a great potential for storing soil organic C (SOC). The bulk SOC, however, is partitioned into fractions of soil organic matter (SOM) that differ in terms of formation, persistence, and function. As a result, the fractions might be differently affected by carbon (C) input chemistry (quality) and climate change. Here, we compare the amounts of C in SOM fractions among deciduous, coniferous, and mixed forest soils along the soil profile. We also determine how the formation and persistence of SOM fractions are affected by C input quality (i.e., litter leachates vs. root exudates), increased temperature, and soil fauna and microorganisms. The proposed project consists of field samplings and observations, a slightly manipulated field experiment that explains a part of the process, and two heavily manipulated laboratory experiments that explain the process in detail. Finally, the acquired data will be compiled and used to improve and validate selected soil C models.

Insect seasonality:

Photoperiodic timer and mechanisms regulating insect Diapause

Our major interest is diapause and its regulation, particularly seasonal switch depending on day-length measurement called photoperiodic timer (clock). Our goal is identifying architecture of photoperiodic timer at the genetic, molecular and anatomical levels. We mostly use two insect species: Pyrrhocoris apterus (Heteroptera) and Chymomyza costata (Diptera).

Pyrrhocoris apterus (Heteroptera) undergoes adult reproductive diapause, which is characterized by absence of Juvenile Hormone synthesis.

• Using gene silencing in P. apterus, we confirmed the role of the circadian clock genes in photoperiodic timer (Kotwica-Rolinska et al., 2017).

• Using P. apterus, we described plasticity in Juvenile Hormone (JH) reception, where one universal receptor protein (MET) interacts with various partners in tissue specific manner (Smykal et al, 2014). One of responses requires interaction of MET with circadian proteins and output relies on novel, non-cyclical feedback between Par domain protein 1 and Cryptochrome (Bajgar et al., 2013a; Bajgar et al., 2013b).
• Surprisingly, JH not required for male mating behavior or fertility, whereas circadian genes are needed for the photoperiod-dependent switch from diapause to reproductive growth of MAGs and for mating (Urbanova et al., 2016)

Drosophilid fly Chymomyza costata undergoes photoperiodically regulated larval diapause.

• With robust photoperiodic diapause and adaptaion to temperate regions, Chymomyza costata is an interesting species to study circadian clock (Dolezel, 2019).
• Using RNAseq technology, we characterized transcription profiles associated with photoperiodic diapause induction. Short day photoperiod triggering diapause was associated to remarkable inhibition of 20-hydroxy ecdysone (20-HE) signalling during the photoperiod-sensitive stage of C. costata larval development (Poupardin et al., 2015).
• Using cell culture assays, we defined molecular mechanism of non-diapause (npd) mutant strain in C. costata. The 1855 bp deletion in npd strian removes crucial regulatory cis-elements as well as the minimal promoter, being subsequently responsible for the lack of tim mRNA expression (Kobelkova et al., 2010).

Representative publications:

• Smýkal V., Pivarči M., Provazník J., Bazalová O., Jedlička P., Lukšan O., Horák A., Vaněčková H., Beneš V., Fiala I., Hanus R., Doležel D. (2020) Complex evolution of insect insulin receptors and homologous decoy receptors, and functional significance of their multiplicity. Molecular Biology and Evolution 37 : 1775-1789. DOI: 10.1093/molbev/msaa048

• Kotwica-Rolinska J., Pivarčiová L., Vaněčková H., Doležel D. (2017) The role of circadian clock genes in the photoperiodic timer of the linden bug, Pyrrhocoris apterus, during the nymphal stage. Physiological Entomology 42 : 266-273.
  DOI: 10.1111/phen.12197

• Urbanová V., Bazalová O., Vaněčková H., Doležel D. (2016) Photoperiod regulates growth of male accessory glands through juvenile hormone signaling in the linden bug, Pyrrhocoris apterus. Insect Biochemistry and Molecular Biology 70: 184-190. DOI: 10.1016/j.ibmb.2016.01.003
• Doležel D. (2015) Photoperiodic time measurement in insects. Current Opinion in Insect Science 7: 98-103. DOI: 10.1016/j.cois.2014.12.002
• Poupardin R., Schöttner K., Korbelová J., Provazník J., Doležel D., Pavlinic D., Beneš V., Koštál V. (2015) Early transcriptional events linked to induction of diapause revealed by RNAseq in larvae of drosophilid fly, Chymomyza costata. BMC Genomics 16: 702. DOI: 10.1186/s12864-015-1907-4
• Smýkal V., Bajgar A., Provazník J., Fexová S., Buřičová M., Takaki K., Hodková M., Jindra M., Doležel D. (2014) Juvenile hormone signaling during reproduction and development of the linden bug, Pyrrhocoris apterus. Insect Biochemistry and Molecular Biology 45: 69-76.
• Bajgar A., Jindra M., Doležel D. (2013) Autonomous regulation of the insect gut by circadian genes acting downstream of juvenile hormone signaling. Proceedings of the National Academy of Sciences of the United States of America 110: 4416-21.

Biology, phylogeny and evolution

For historical reasons, current myxozoan taxonomy is based predominantly on the morphology and structure of the spores produced in the vertebrate (fish) host. However, phylogenetic relationships among the myxozoans based on different gene regions disagree with the traditional taxonomic classification: a number of myxosporeans with very similar spore morphology and belonging to the same genus were found to be phylogenetically distantly related, rendering most genera non-monophyletic. But how important is spore morphotype for the evolution of these highly reduced parasites? One of our principal aims is to investigate differences with regard to parasite development as well as life cycles and biology, in order to be able to understand why they cluster in different branches of the phylogenetic tree. By reconstructing the evolutionary history of myxozoans and their hosts, using dated metazoan trees and cophylogeny methods, we have recently demonstrated that myxozoans emerged approx. 700 mya and that they first invaded invertebrate hosts before including vertebrate hosts into their life cycles (Holzer et al. 2018, Mol Ecol 27:1651-1666). Following vertebrate invasion, fish hosts were acquired multiple times, leading to parallel cospeciation patterns in all major phylogentic lineages. We identified the acquisition of vertebrate hsots that facilitate alternative transmission and dispersion strategies as reason for the distinct success of the Myxozoa, and identify massive host specification-linked parasite diversification events. We currently study the beginning of the heteroxenous life cycle in myxozoans in old vertebrate hosts (Chondrichthyes and Agnatha; Lisnerová et al. 2020, Biology 9:10; Kodádková et al. 2015, Int J Parasitol 45:269-276) and try to understand how the secondary host was integrated into the life cycle of these parasites. The identification of characters and key events in myxosporean evolution lead to a better understanding of the broad diversity of modern species.

Unveiling Tick Symbiosis: A Functional Dive into Tick-Bacteria Partnerships

• Developing "apo-symbiotic" ticks: We utilise ex vivo blood-feeding systems with antibiotic-treated blood meals to create ticks devoid of their bacterial symbionts. This allows us to compare "symbiont-free" and "symbiont-harboring" cohorts of ticks and understand the impact of these partnerships.
• Focus on key model systems: We primarily study the partnership between the bacterium Midichloria mitochondrii and the European tick Ixodes ricinus, but also explore symbiosis in Francisella and Ornithodoros ticks. This comparative approach provides valuable insights into function of diverse tick-symbiont interactions.

• How do symbionts influence tick development, reproduction, and feeding success?
• What role do symbionts play in tick immunity and defense against pathogens?
• Can understanding these partnerships pave the way for novel tick control strategies that target the tick-symbiont relationship?

Role of the only tRNA intron in trypanosomatids

In yeast Saccharomyces cerevisiae and other model organisms, 20% of all tRNAs contain introns. Their removal is an essential step in the maturation of tRNA precursors. In T. brucei, there is only one intron containing tRNA: tRNATyrGUA. Since this tRNA is responsible for decoding all tyrosine codons, intron removal is essential for viability. Using molecular and biochemical approaches, several non-canonical editing events were identified within the intron-containing tRNATyrGUA. The RNA editing involves guanosine-to-adenosine transitions (G to A) and an adenosine-to-uridine transversion (A to U), which are both necessary for proper processing of the intron. We have been obtaining tRNA intron sequences from our collection of newly identified trypanosomatid species. We hope this will help us to understand the process of RNA editing and ultimately identify biological function for the presence of the only intron containing tRNA in these organisms.

Microsporidia and Cryptosporidium of wild great apes

Projekt: Microsporidia and Cryptosporidium of wild great apes

Population genetics, demography and molecular evolution in rodents and their parasites

Diversity and biology of planktonic diplonemids

 .

Lyme disease transmission model

GACR 13-12816P (2013-2015)

Lyme borreliosis is an emerging vector-borne disease of temperate climates, with a current distribution spanning North America and Eurasia. It is caused by spirochetes belonging to the Borrelia burgdorferi sensu lato group that are transmitted through Ixodes ticks. Although Lyme borreliosis is one of the best studied tick-borne zoonoses, the annual incidence leads other vector-borne diseases and continues to increase. There is currently no vaccine available to prevent Lyme disease in humans. One of the promising strategies to break Borrelia transmission is to develop of a vaccine affecting basic physiological processes of the tick. Development of a promising vaccine against Lyme borreliosis would be greatly facilitated by a reproducible vector host transmission model. Main aim of this project is to implement such model and to find a molecule with proven anti-borrelial effect.

Interaction between bZIP proteins Jun and Atf3 in epithelial morphogenesis during metamorphosis

Movements, fusions and closures of epithelial tissues are important for animal development, e.g. during embryogenesis but also for healing of wounds. The fruit fly, Drosophila melanogaster, offers several model examples of such epithelial movements and fusions. An interesting case is the formation of the adult fly abdomen. During metamorphosis of a maggot into fly, large larval epidermal cells (LEC) must leave the epithelium to free space for new cells (histoblasts), which rapidly proliferate and migrate to occupy the entire surface of the abdomen, then produce the adult cuticle. Regulation of this complex exchange of cell populations is unclear. It has been known that the entire process, like metamorphosis of other tissues, is stimulated by the steroid hormone ecdysone, and that the extrusion of LEC requires cytoskeleton changes dependent on the Rho signaling pathway. Unexpectedly we have implicated in the abdominal morphogenesis Atf3, a transcription factor of the bZIP family. Activity of the atf3 gene must be restricted during the exchange of the two cell populations, as deregulated atf3 blocks removal of LEC from the epithelium by interfering with Rho signaling and increasing LEC adhesiveness. These obsolete larval cells consequently obstruct closure of the adult epithelium. To exert this effect, Atf3 requires its binding partner, the oncoprotein Jun. Although interaction between these two regulatory proteins has been well documented by in vitro studies on mammalian cell lines, our results for the first time demonstrate a developmental function of the Atf3-Jun complex in vivo.

Function of bee venom toxins

The project focuses on the individual components that make up bee venom, especially melittin and phospholipase A2, which are the most abundant elements of the venom and the main contributors to pain and allergic reactions brought on by bee stings. They have also demonstrated in vivo anti-bacterial, anti-tumour, and anti-inflammatory activity. The ratio of representation of individual components in the venom and environmental influences, such as exposure to toxic substances in the form of insecticides or stress brought on by the presence of predators or pathogens in the bee colony, are investigated using molecular methods (primarily RT-PCR). Additionally, it is being researched whether the bee organism can use the venom's components for its own defence during an infection immune response or as an anti-stress reaction when the organism's integrity is compromised.

PI: Markéta Hejníková

Dynamics of structured populations

Many practical and theoretical problems in ecology, epidemiology and evolutionary biology require consideration of a type of population structure. For example, analyzing impacts of efficiency of a variety of age-specific interventions or of vaccine prioritization in the ongoing COVID-19 epidemics, as well as exploring changes in dynamics of animal populations subject to sexually transmitted infections or sex-selective predators, we need to structure the respective populations by age and sex, and mirror this structure in the corresponding mathematical models. Our current research in this respect covers a range of topics, including dispersal-driven metapopulation dynamics, impacts of mating strategies on dynamics of populations subject to sexually transmitted infections, evolution of life history in age-structured or sex-structured populations, and actually exploration of impacts of non-pharmaceutical interventions and vaccination strategies on dynamics of the COVID-19 epidemic. Our results have ramifications for conservation strategies, biocontrol programmes, and generally management of populations.

Ecology and conservation of butterflies in traditional managed landscape

The EU-protected Marsh fritillary still persists in strong populations in westernmost areas of the Czech Republic. Following its populations from 2002 onwards resulted into detailed understanding of its habitat requirements (Konvicka et al. 2003, EJE) and management needs (Hula et al. 2004, Entomol. Fenica). Our active involvement in the species conservation included designing NATURA 2000 sites, drafting a species action plan, and advisory participation in habitat management. Several publications used this endangered species as a model for studies of dispersal (Fric & Konvicka 2007, Basic and Applied Ecology) and metapopulation structure (Fric et al. 2010, Ecol. Res.) of butterflies inhabiting seminatural grasslands.
 

Population ecology, dispersal, and phylogeography

Animal movement and demographic structure are some of the key parameters affecting the survival of local populations, and thus they have become of particular interest for conservation biology of the threatened species. Using relevant approaches (e.g. mark-release-recapture or radio-tracking) we study population structure and dispersal of endangered beetles, with the main focus on Rosalia alpina and Cerambyx cerdo. Further, to better understand the genetic population structure and the evolutionary history of these species, we also employ molecular methods commonly used in ecological studies. Our findings are important for designing more efficient conservation strategies.

Related publications:

Drag & Čížek 2018: Radio-Tracking Suggests High Dispersal Ability of the Great Capricorn Beetle (Cerambyx cerdo). Journal of Insect Behavior.

Drag et al. 2018: Phylogeography of the endangered saproxylic beetle Rosalia longicorn, Rosalia alpina (Coleoptera, Cerambycidae), corresponds with its main host, the European beech (Fagus sylvatica, Fagaceae). Journal of Biogeography.

Phylogeography

Publications:

• Ditrich T., Janda V., Vaněčková H., Doležel D. (2018) Climatic variation of supercooling point in the linden bug Pyrrhocoris apterus (Heteroptera: Pyrrhocoridae). Insects 9 : article number 144.
  DOI: 10.3390/insects9040144

• Pivarciova, Provaznik, Vaneckova, Pivarci, Peckova, Bazalova, Wu, Cada, Kment, Kotwica-Rolinska, Dolezel: Unexpected geographic variability of free running period in the linden bug, Pyrrhocoris apterus (Journal of Biological Rhythms, DOI: 10.1177/0748730416671213)

Publication connected to geographic variability, but on a different insect model:

• Paolucci S., Dalla Benetta E., Salis L., Doležel D., van de Zande L., Beukeboom L.W. (2019) Latitudinal variation in circadian rhythmicity in Nasonia vitripennis. Behavioral Sciences 9 : article number: 115. DOI: 10.3390/bs9110115

Geographic adaptations of the linden bug, Pyrrhocoris apterus, and its phylogeography

We use our favorite model organism, Linden bug (Pyrrhocoris apterus), to study diapause and particularly its photoperiodic regulation. Seasonal environmental changes are mostly influenced by latitude and altitude of particular locality. Indeed, we observe remarkable variability in photoperiodic and circadian clocks between geographical field-lines of P. apterus. Robust analysis of P. apterus phylogeography is needed to shed some light on origin of these adaptations. In collaboration with Prof. Roger Butlin (University of Sheffield) we are performing RAD sequencing and analysis of samples from approximately 150 localities.

Current collection of P. apterus (see the map) was only possible thanks to generous help of following colleagues: Adam Bajgar, Alejandro Cabezas-Cruz, Aleksandra Konjevic, Barbara Lis, Jerzy A. Lis, Carl-Cedric Coulianos, Rodolfe Costa, Dora Nagy, Eva Hola, Hanka Vaneckova, Iva Fukova, Jana Pavlová, Joanna Kotwica-Rolinska, Jula Lukes, Kai Schuette, Kajka Straznicka, Lucia Salis, Lukas Cizek, Lukas Drag, Manuel Baena, Marek Jindra, Marketa Ondrackova, Martin Kaltenpoth, Martin Vacha, Matilde Eizaguirre, Milan Stech, Milena Damulewicz, Olina Bazalova, Petr Kment, Petra Sekyrova, Plamen Kalushkov, Radka Zavodska, Ramon Albajes, Stanislav Rada, Teemu Rintala, Vlastimil Smykal, Xanti Pagola, Zejlko Popovic,

However, several geographical regions are either underrepresented or even absent completely (see red circles in the map below). Therefore, we would really welcome samples (either dead or alive) from following regions:

Europe – England, Sweden, Norway, Denmark, Mediterranean islands (Sicily, Corsica, Sardinia, islands in Aegean sea), Romania, Moldavia, Portugal, Belarus, Ukraine, Russia,

North Africa– Morocco, Tunisia, Algeria, Egypt

East – Turkey, Syria, Lebanon, Jordan, Iraq, Iran, Afghanistan, Pakistan, Oman, Saudi Arabia, Emirates, Georgia, Azerbaijan, Turkmenistan, Tajikistan, Uzbekistan, Kyrgyzstan, Kazakhstan, Russia, South West Mongolia, North West China

Sample collection: The optimal way for storing and transporting samples seems to be 96% ethanol (non-denatured). 2ml screw cup tubes can accommodate 5 - 8 bugs from the same locality. We are ready to send prepared tubes filled with EtOH, just send an e-mail request, please. Although we are happy even for one individual bug, 10-15 specimens from one locality are optimal. It is important to have the report of the locality (such as GPS position or locality name from the map, elevation is also very useful). Please, if possible, it would be also great to mention, if the locality was urbanized area (town, park and similar area modified by human) or if the locality is more similar to original nature-like type (forest, steppe, mountains). In ideal case we would like to have samples collected approximately 200 km apart in lowlands, while in mountains or complex terrain even higher density is helpful (down to tens of kilometers).

Living bugs – an alternative is to collect live specimens. Adults survive with wet piece of cotton or cellulose in small paper box for more than one week, as long as it is not too hot. We are really happy to obtain living specimens for establishing colonies, where circadian and photoperiodic phenotypes can be characterized.

Species identification - Pyrrhocoris apteruscan be quite easily identified thanks to its aposematic coloration. Both, larvae or adults can be collected and are welcome. We are also seeking for samples of closely related pyrrhocorids, such as P. sibiricus, P. marginalis, and genus Scantius (see the pictures below).

Contact:  David Dolezel,  david.dolezel@entu.cas.cz; Institute of Entomology; Biology center CAS; Branisovska 31; 370 05 Ceske Budejovice; Czech Republic

Tick-borne diseases in urban environments - where does the real danger lurk?

(Czech Health Research Council, 2023-2026, P.I.: Václav Hönig)

Research and development of vaccine prototypes against Lyme disease

• produce in an E. coli expression system and purify native his-tagged nanovaccines, immunise mice, and efficiently (100% effective) block transmission of B. afzelii (local strain) from laboratory-infected I. ricinus nymphs to naïve mice.
• describe mechanistic understanding of the mode-of-action of the anti-OspA antibodies inhibit the B. afzelii spirochetes in the tick Ixodes ricinus and whether tick physiology and tick (immune) proteins contribute to the elimination of antibody-bound Borrelia.

Non Stop Blastocrithidia: the trypanosomatids with all three stop codons reassigned

Genetic code previously thought to be universal for all the life forms but soon variations from canonical code were described in many organisms all across the tree of life, most of the deviation involves reassignment of one or two stop codon to sense codons leaving 1 or 2 stop codons to confer the termination of translation. Interestingly, in 2016 two groups of protist trypanosomatid Blastocrithidia and several ciliates were shown to reassign all the tree stop codons in their nuclear genome to code amino acids. Transcriptome and proteome analysis of Blastocrithidia revealed UGA has been reassigned to code tryptophan, while UAG and UAA are reassigned to code glutamate. This finding challenges our current understanding of very fundamental process of life “Translation”.

Blastocrithidia is an ideal organisms to study this phenomenon as it belong to well studied group of kinetoplastida where all the known kinetoplastids have canonical genetic code. We have successfully optimized the genetic manipulation of Blastocrithidia which will allow us to explore different aspect of translation.

Recent development in ribosome profiling which involves deep sequencing of ribosome protected mRNA fragment is a powerful tool for monitoring in vivo translation optimization of ribosome profiling and genetic manipulation of Blastocrithidia will allow us to address following questions: Qualitative and quantitative analysis of translation process, Biased in usage of canonical and non canonical synonymous codons, how does the termination of translation work without defined stop codon and evolution of genetic code.

The role of ATP synthase structure in the biogenesis and bioenergetics

Mitochondrial  cristae are inner membrane convolutions where protein factories responsible for bioenergy conversion reside. The cristae exhibit extremely large variability in their ultrastructure, except for one common attribute - the presence of ATP synthase dimer rows at the crista ridges. Little is known about the role of these arrays in cristae structure and mitochondrial bioenergetics. However, Trypanosoma brucei is an excellent model system as the singular mitochondrion of the digenetic parasite is drastically remodeled structurally and metabolically as it progresses through a complex life cycle. Notably, the highly branched, cristae-containing and ATP-producing mitochondrion transitions to a streamlined tubular, cristae-lacking and ATP-consuming organelle. Combining traditional biochemical methods with state-of-art structural approaches (single-particle cryo-EM), we solved the ATP synthase dimer structure, identified a dimer-specific subunit and explored its role in cristae shaping.

Now we aim to decipher the role of ATP synthase dimers in mitochondrial bioenergetics and biogenesis during the parasite differentiation.

Trematodes in sub-Arctic lake food webs: development of quantitative diversity baselines and a framework for community ecology research in the Arctic.

The project is focused on increased resolution in assessments of host-parasite interactions in sub-Arctic freshwater ecosystems by an integrated framework for the discovery of trematode diversity. We aim to provide the first contemporary quantitative baselines for abundance and community structure in model host-parasite systems and to validate an ecological framework for future comparative studies in lake ecosystems of the sub-Arctic. A principal goal is to provide comprehensive baselines for trematode diversity and abundance in key fish and snail species in three model lake ecosystems in Iceland. Having established the genetic species diversity, we shall identify the composition and structure of parasite communities in the model lake ecosystems addressing key issues related to (i) parasite-parasite interactions in local food webs; (ii) host-parasite associations and recognition of host-parasite interaction networks; and (iii) development of quantitative baselines and assessment of the predictability of parasite populations and communities over different temporal and spatial scales.

Czech Science Foundation (18-18597S; P.I: A. Faltýnková; 2018–2020).

Synthesis of tetrapyrroles

Tetrapyrroles heme and chlorophyll are essential for life on Earth. We are interested in evolution of the heme biosynthetic pathway, particularly in origins of involved enzymes and their intracellular localizations. We focus mainly on algae that have passed through a process of secondary endosymbiosis, such as chromerids, dinoflagellates, diatoms and cryptophytes, and at the end also apicomplexan parasites. We showed that phototrophic alga C. velia produces 5-aminolevulinate similarly to apicomplexan parasites from glycine and succinyl CoA in the mitochondrion. All other eukaryotic phototrophs synthesize this precursor from tRNA-GLU in the plastid, where the entire pathway is located. We study heme pathway also in other organisms such as Vitrella brassicaformis, Bigelowiella natans, Paulinella chromatophora, Glenodinium foliaceum and others.

Babesiosis transmission model

Babesiosis is a tick-borne malaria-like disease of mammals. Because of global changes and expansion of the tick range, importance of babesiosis as an emerging zoonosis is increasing. Interplay between the parasite, the tick and the vertebrate host represent a complex system of multiple molecular interactions. To date, only a limited number of molecules have been identified to play a role in this system. Our research is focused on the identification and characterization of molecular mechanisms of Babesia persistence within the tick vector and its transmission to the vertebrate host.  We are currently working on the setting of Babesia transmission model in our laboratory and use of this model for testing of the tick immune genes in Babesia infection by RNA interference.

Overwintering in insects of economic importance

We follow the course and the success of overwintering in insect pest species (or beneficial insects) in their field habitats. We assess the seasonal timings of diapause induction, initiation, termination and the onset of spring activity. We assess a whole set of physiological parameters, which correlate and/or are causally involved in cold tolerance and winter mortality. We use various techniques (supercooling point, vapour pressure and Clifton nanoliter osmometers, differential scanning calorimetry, gas exchange and respirometry analysis, high resolution mass spectrometry, etc.). Select target species: Ips typographus, Pityogenes chalcographus, Cydia pomonella, Ostrinia nubilalis, Culex pipines and others.

Linking functional traits of three organism levels as driving mechanisms of ecosystem functions in the Arctic

Czech Science Foundation (CSF), team member, 2017-2019

There is an increasing evidence that functional traits of biota may serve as important indicators of ecosystem services. Although it is known that different organism levels interact in providing ecosystem services, there are so far only few studies linking functional traits of several trophic levels together. Here we proposed to interconnect functional traits of three organism levels plants, soil fauna and soil microorganisms - to ecosystem functions underlying ecosystem services in the Arctic terrestrial ecosystem, representing a less complex, model system. Several ecosystem functions were approached (C sequestration potential and soil stability, fertility and water retention) in a set of manipulative experiments. We expected that joining relevant functional traits across organism levels into "multitrophic" trait clusters will allow us to identify key traits underlying the ecosystem functions.

Neuropathogenesis linked to NS1 protein of tick-borne encephalitis virus

(Czech Science Foundation, 2023-2025, P.I. Martin Palus)

Innovation of Integrated Potato Protection Against the Colorado Potato Beetle Based on New Knowledge of Genetic and Biological Characteristics

Colorado potato beetle (CPB) is our main pest in the above-ground mass of potatoes with the potential for crop losses up to 50%. However, protection against it is currently very difficult due to insecticide resistance. In addition to the use of chemical protection and controversial genetic modifications, the development and use of biological protective methods are becoming increasingly promising. Use of entomopathogenic fungi (EPF) and parasitic nematodes (EPN) for protection does not cause a pest resistance. A great advantage is the support in use of mycoinsecticides by sustainable and organic farming programs. Apart from conservation studies, molecular and genetics methods are used wordwide to address the invasive and genetic potential of CPB.

Funding: Ministry of Agriculture; Project NAZV ZEMĚ No. QK1910270 (PI: O. Skoková Habuštová).

Key outputs:

Zemek R., Konopická J., Jozová E., Skoková Habuštová O. (2021) Virulence of Beauveria bassiana strains isolated from cadavers of Colorado potato beetle, Leptinotarsa decemlineata. Insects 12: article number: 1077.

Evolution of trypanosomatids pathogenicity

Trypanosomatids undoubtedly belong to the most successful parasites on Earth with unique life style and ingenious adaptations to the hostile environment of the host. Because they cause devastating diseases and therefore represent major threat not only for developing countries, several dixenous (two-host) species from the genera Trypanosoma and Leishmania are subjects to thousands of studies aiming to help us understand various aspects of their molecular and cellular biology. Some of them had their genomes sequenced and now high quality reference genomes are available.

Nevertheless, it seems unlikely that genome analyses of just the dixenous species will reveal specific genetic elements responsible for successful parasite invasion. We would like to derive this information from genome sequencing and comparative analyses of monoxenous (single-host) species. For this purpose are sequencing, assembling and annotating genomes of selected monoxenous trypanosomatids belonging to phylogenetic clades, so far without a sequenced representative.

Acyclic nucleaoside phosphonates as potent nhibitors of purine salvage pathway enzymes

The purine salvage pathway (PSP) in unicellular parasites complies with key requirements for essentiality, potential druggability, availability of in vitro assays, and availability of structural information of target enzymes. In contrast to their mammalian hosts, trypanosomatids (as well as malaria parasites) lost their ability to synthesize nucleoside monophosphates de novo and they rely entirely on the acquisition of purines from the host environment. Recently several enzymes of the PSP (i.e., GMP synthase, hypoxanthine-guanine-(xanthine) phosphoribosyltransferase HG(X)PRT) have been experimentally validated as potential therapeutic targets. It has also been shown that selected prodrugs of potent acyclic nucleoside phosphonates (ANPs), inhibitors of HGXPRT and HGPRT, block the growth of above-mentioned parasites.

The current study focused on Trypanosoma brucei (Tbr) purine salvage pathway (PSP). To complement our preliminary studies on key PSP enzymes, 6-oxopurine PRTs and their inhibitors (ANPs), we decided to add adenine PRT (APRT) to the studied portfolio of enzymes and study APRT side by side to 6-oxopurine PRTs.

Epidemiology and immunology of microsporidiosis

The immune response mechanisms against the microsporidian Encephalitozoon cuniculi were studied. The role of specific anti-microsporidia antibodies in protection of infected mice was described based on results of experimental infections of immunocompetent mice compared with those of immunodeficient mice. Mice with disrupted IFN-? or IL-12 genes, and those with severe combined immunodeficiency, served as models of immunodeficient hosts. Adoptive transfer experiments of CD4+ and CD8+ T lymphocytes, in combination with antibodies applied in vivo, were used to examine the contribution of antibodies to protective immunity against microsporidians.

Free-living and captured mammals and birds are examined for microsporidia spores. New species of the genus Trachipleistophora has been described from muscles of Tenrec ecaudatus

Landscape ecology

To understand the drivers that affect insect communities at larger spatial scales and in long term, we study landscape changes using Geographic Information Systems (GIS). We characterize land cover types in past and in present, combine them with ecological background of studied organisms or mathematical models and relate these to current distribution of target species or target habitats. Our studies suggest that transition from traditional silvicultural practices to modern agriculture and forestry managements during last two centuries had an important negative impact on biodiversity of several groups of organisms.

Related maps:

Veteran trees and saproxylic insects in the floodplains of lower Morava and Dyje rivers: http://goo.gl/oeBgtn

Pollard willows in South Moravia: https://kfgg.maps.arcgis.com/apps/webappviewer/index.html?id=4d390794c9ec46b889269e744a8c5222

Related publications:

Miklin et al. 2018: Past levels of canopy closure affect the occurrence of veteran trees and flagship saproxylic beetles. Diversity and Distributions.

The impacts of tropical forest degradation and fragmentation on ant-plant mutualisms

Project investigates how the mutualism between early succession Macaranga trees and their ant symbionts changes with logging and fragmentation of tropical forest, conversion to oil palm and experimental forest regeneration in Borneo. We will quantify network structure, and assess benefits for ants (food and living space) and trees (protection from herbivores). We will use field experiments to assess critical degrees of isolation from ant colonists for Macaranga and the potential for ant-assisted Macaranga to facilitate other trees.

Funding: 2016-2018: 16-09427S Czech Science Foundation (PI: T. Fayle)

Key studies:

Houadria M.Y.I., Klimes P., Fayle T.M., Gullan P.J. Host-plant dissections reveal contrasting distributions of Crematogaster ants and their symbionts in two myrmecophytic Macaranga species. Ecological Entomology 43: 601–611

Houadria M.Y.I. Feldhaar H., Fiala B., Lestina D.,  Chung A., Salleh A., Justin H., Kokorova P., Fayle T.M. (2020) Reduced benefits of ant occupation for ant-trees in oil palm compared with heavily logged forest. Symbiosis 81: 79-91.

Organic matter decomposition and carbon sequestration on a natural gradient of labile carbon sources in coniferous temperate forest soils

Czech Science Foundation (CSF), principal investigator, 2017-2019 

This project dealt with the effect of labile carbon (C) from various sources (litter, rhizodeposition, honeydew) on soil organic matter (SOM) decomposition and C sequestration in temperate coniferous forest soils, which are of global importance in terms of organic C storage. The proposed project mainly dealt with: (i) the rate of SOM decomposition and C sequestration on a natural gradient of labile C sources; (ii) potential increase in SOM decomposition and C sequestration with increased labile C input; (iii) effect of microbial and faunal community on labile C flux; and (iv) the role of respective labile C sources in C fluxes through soil. Field samplings and observations were combined with field and laboratory manipulation experiments, both followed by laboratory analyses.

Gene knockdown and functional research of Myxozoa

Despite the era of next-generation sequencing of Myxozoa, reverse genetics techniques are limited in myxozoan research. A significant step forward to explore the biological aspects of unprecedented Myxozoa embodies RNA interference (RNAi)-triggered gene knockdown. Therefore, we optimized and experimentally demonstrated a workflow for gene knockdown of unusual actins (Kyslík et al. 2024) related to the unique motility of a model system of carp infecting Sphaerospora molnari established in our laboratory from in vivo infected common carp. In this direction, we can use this invaluable research tool to explore previously unknown genes restricted to Myxozoa. We continue to investigate myxozoa-specific genes of unknown function using RNAi, which considerably supports the unraveling of many aspects of this mysterious group of parasites.

The assembly of iron-sulfur clusters in Trypanosoma brucei

Trypanosoma brucei is characterized by a number of unique cellular features. Since methods of reverse genetics are available for this flagellate, it can now be considered a model protist. Iron-sulfur (Fe-S) clusters are ancient and ubiquitous cofactors of proteins that are involved in a variety of biological functions, including enzyme catalysis, electron transport and gene expression. Nevertheless, little is known about how Fe-S clusters are assembled in T. brucei. So far, by means of RNA interference, we have down-regulated several evolutionary highly conserved components of the pathway, such as cysteine desulfurase IscS, metallochaperone IscU, frataxin, ferredoxin, and IscA. With the exception of IscA, all are essential for the parasite and their down-regulation results in reduced activity of the marker Fe-S enzyme aconitase in both the mitochondrion and cytosol. Moreover, interfering with these genes also decreased the activity of succinate dehydrogenase and fumarase, affected membrane potential of the mitochondrion and general oxygen consumption. This supports the hypothesis that the mitochondrion plays a fundamental and evolutionary conserved role in cellular Fe-S cluster assembly throughout the eukaryotes. Interestingly, we have rescued the frataxin know-down in T. brucei with its homologue from the hydrogenosome of Trichomonas vaginalis containing the hydrogenosome-targeting signal peptide. Further analyses of this rescue and the various RNAi knock-downs are under way.

Immune response in gastric mucosa during Cryptosporidium muris infection

Although scarcely any T-cells are presented in gastric mucosa of healthy rodent hosts, significantly elevated migration of T-lymphocytes (more than 10,000 fold), especially CD8+ T-lymphocytes, to the stomach mucosa occurred during primary infection and persisted for more than two months after its resolution. Most immunocompetent hosts self-cured during 35 days post infection (DPI). The ex vivo cultures of splenocytes revealed very low levels of IFN-? production during the course of the primary infection. These infected animals acquire a protective immunity against re-infection with same or another gastric Cryptosporidium. No cross immunity between gastric Cryptosporidium species of mammals and C. parvumhas been detected. The hosts with CD8+ or CD4+ T-cells deficiency suffered from chronic cryptosporidiosis; nevertheless, they were able to self-cure during 100 or 150 DPI, respectively.

Impact of degradation of forests on ant/termite communities and their inter-species interactions

In this project we will assess the way that interactions between soil dwelling ants and termites change when tropical rainforest is logged, fragmented and converted into oil palm plantation. Interactions will be quantified using environmentally constrained null models of species cooccurrence, molecular identification of gut contents, and observations and experiments linking the soil biota to soil properties and nutrient redistribution rates. Using these data we will create whole-network models which will allow prediction of the stability of ecosystem diversity
and functioning under a range of species extinction scenarios. 

Funding: 2014-2016: How do changes in species interaction networks affect ecosystem function when tropical forests are degraded? Czech Science Foundation (PI: T. Fayle) 

Key studies:

Luke, S.H.; Fayle, T. M.; Eggleton, P.; Turner, E.C.; Davies, R.G. (2014) Biodiversity and Conservation 23 (11): 2817-2832.

Fayle T.M. et.al. (2015) PLoS ONE 10: e0122533.

Tuma J., Fleiss S., Eggleton P., Frouz J., Klimes P., Lewis O., Yusah K.M., Fayle T.M. 2019. Applied Soil Ecology 144: 123-133.

Host-parasite interaction

We use transcriptomics and proteomics in combination with in vitro and in vivo experiments to better understand how myxozoan proliferative stages in carp blood are able to multiply rapidly in the host and to use resources effectively, while avoiding host immune defense mechanisms. Our model organism, S. molnari uses highly derived cytoplasmic actin to generate a completely novel type of cellular motility, which is of great importance for evading the attachment and lysis by host immune cells (Hartigan et al. 2016, Sci Rep 6:39093). Apart from this motility, S. molnari uses a wide repertoire of proteolytic enzymes and their inhibitors to survive and propagate in the host blood, some of which are expressed only during proliferation and are of particular importance to the early propagation of the parasite infection in fish (Hartigan et al. 2020, BMC Genomics 21:404; Bartošová-Sojková et al. 2021, Biology 10:110; Esterbauer et al. 2020, Microorganisms 8:1502). We are currently describing and characterizing some proteases and specific protease inhibitors biochemically and structurally. The main source of nutrients for the parasite's blood stages are red blood cells (Korytar et al. 2020, Parasite Immunol, 42:e12683). Their immunological responses and energy transfer/nutrition roles are under further investigation. In parallel, we explore the immunological reaction of carp to S. molnari which exists of several phases, an initial covert infection with minor proinflammatory characteristics which is skewed to a strong anti-inflammatory type once proliferative blood stages occur (Korytar et al. 2019, Parasit Vect 12:208). Additionally, we expanded the available knowledge of the kinetics of leukocytes subpopulations during the model infection and evaluated their proliferative capacity in main lymphoid organs. The proliferation led to a substantial increase in the number of circulating B lymphocytes producing S. molnari specific antibodies. These antibodies, exerting potent killing capacity in vitro, do not prevent the proliferation of the parasite in vivo, indicating a possible antibody evasion strategy employed by the parasite. Current work focuses on the characterization of changes in the antigenic composition during the parasite's life cycle and its impact on B cell responses. 

Functional Analysis of Babesia Calcium-Dependent Protein Kinases

Project:GAČR: 21-11299S

EN: This research is aimed at tick-borne apicomplexan parasites of the genus Babesia, which infects a range of animals and humans. We focus on the parasite's ability to invade host erythrocytes, a trait it shares with its relative Plasmodium, the cause of malaria. Our team is employing cutting-edge transgenic techniques to conduct a thorough examination of previously unknown calcium-dependent protein kinases (CDPKs) in Babesia divergens. By exploring the role of these enzymes in key biological processes, we aim to understand their function and potential as drug targets. This includes using Bumped Kinase Inhibitors (BKIs) to selectively inhibit these enzymes and assessing their effectiveness in combating the parasite.

Researc Articles:

PMID: 36014069

PMID: 38156314

PMID: 38104025

PMID: 38287902

PARASITIC PROTISTS

Apart from our main focus on myxozoans, we investigate a wide diversity of protists in aquatic environments and hosts, studying their distribution, host-parasite interaction, pathology, ecology as well as functional and evolutionary aspects. Here’s some of our recent and ongoing research in our on lab and as collaborators:                                                                      

Master proteases driving the apical complex of Babesia parasites

GAČR: GA23-07850S

This project focuses on specific proteolytic enzymes appearing on the top of a proteolytic cascade directly driving the function of apical complex during host cell egress and invasion by apicomplexan parasites. An in silico comparative analysis of Babesia omics datasets utilizing Toxoplasma and Plasmodium queries determined candidate proteolytic enzymes in Babesia divergens that will be further functionally and biochemically characterized to confirm their essential role for the parasite and their potential druggability with low molecular weight inhibitors.  Main focus is dedicated to two B. divergens aspartyl protease homologues of Plasmodium falciparum plasmepsins IX/X  and Toxoplasma gondii TgASP3  tagged BdASP3a/b as potential master drivers of the apical complex and molecular events associated with egress and invasion of host red blood cells.  

Research Articles: PMID: 37271664

Research into free-living and amphizoic amoebae

A large-scale phylogenomic study focussed on free-living testate amoebe because they have, due to their shells, a fossile record and can provide invaluable information on early eukaryote evolution. Based on 324 genes and 19 testate amoebae it was shown that the major lineages of testate amoebae were already diversified before the Sturtian glaciation (720 mya), supporting the hypothesis that massive eukaryotic diversification took place in the early Proterozoic (Lahr et al. unpublished). Applied research encompases studies on the pathological agent involved in outbreaks of amoebic gill disease (AGD) in Atlantic salmon in Northern Europe (Paramoeba perurans) and we are testing a variety of substances for their efficacy against P. perurans and their suitability as in-feed treatments against AGD. Most of this research is sponsored by the R&D department of Skretting Aquaculture Research Centre. We also screen for of a variety of pathogenic amoebae in water sources impacted by human actions, using molecular methods, and we have a unique, comprehensive culture collection of amphizoic amoebae from different aquatic habitats and hosts, with hundreds of different strains.                                                                    

Functional analysis of RNA editing in Trypanosoma brucei

We have generated knock-down cell lines of T. brucei procyclics, in which RNA binding proteins MRP1 and MRP2 are down-regulated by RNA interference. We have shown that these proteins exist in a complex composed of two MRP1 and MRP2 molecules each, which is involved in RNA editing and stabilization of mitochondrial transcripts. In a collaborative effort, we would like not only to decipher the 3D structure of this complex, but also to understand how RNA molecules are bound to it. Moreover, we are using the newly developed TAP-purification method to detect binding partners of the MRP proteins and another RNA-binding protein, TbRGG1. Furthermore, we will dissect the function of individual MRP protein domains.

Running research projects

• Harnessing the Win-Win potential: Enhancing sustainability and disease control in aquaculture by using black soldier fly larvae fed on agricultural waste. Ministry of Education, Youth and Sport of the Czech Republic, program Inter-Excellence, subprogram Inter-Action. (LUAIZ24009, 2024-2027, PI: S. Nayak)
• Food or foe: Interaction of myxozoans with red blood cells. Czech Science Foundation and Austrian Science Fund. (24-13238L, PI: P. Sojková. 2024-2027).
• Unveiling the impact of human activities on the distribution of parasites in aquatic ecosystems using metagenomic analysis.  Ministry of Education, Youth and Sport of the Czech Republic, program Inter-Excellence, subprogram Inter-Action. (LUAUS24281, 2024-2027, PI: I. Fiala)
• Biodiversity Shifts in Myxozoan and Microsporidian Parasites: The Role of Human Activity and Sewage Treatment Plant in Živný Stream. Grant Agency of the University of South Bohemia. (PI: M. Bürgerová) 
• Living jewels under the water surface of Šumava. Interreg Bavorsko – Česko BYCZ01-020. https://www.fishjewels.cz/​  (Inst. of Parasitology Co-PI: I.Fiala)
• Myxozoan cystatins and fish host cathepsins: Revealing potential crosstalk during infection. Czech Science Foundation (21-16565S, PI: P. Sojková. 2021-2023).
• Complex characterisation of myxozoan mitochondria: Genome, proteome and ultrastructure approach. Czech Science Foundation. (21-29370S, PI: I. Fiala, 2021-2023)

Analysis of dyskinetoplastic trypanosomes

Trypanosoma brucei is a kinetoplastid flagellate, the agent of human sleeping sickness and ruminant nagana in Africa. Kinetoplastid flagellates contain their eponym kinetoplast DNA (kDNA), consisting of two types of interlocked circular DNA molecules: scores of maxicircles (each ~23 kb) and thousands of minicircles (~1.0 kb). Maxicircles have typical mt genes, most of which are translatable only after RNA editing. Minicircles encode guide (g) RNAs, required for decrypting the maxicircle transcripts. The life cycle of T. brucei involves a bloodstream stage (BS) in vertebrates and a procyclic stage (PS) in the tsetse fly vector. Partial (dyskinetoplastidy, Dk) or total loss (akinetoplastidy, Ak) of kDNA locks the trypanosome in the BS form. Transmission between vertebrates becomes mechanical without PS and tsetse mediation, allowing the parasite to spread outside the African tsetse belt. Trypanosoma equiperdum and Trypanosoma evansi are agents of dourine and surra, diseases of horses, camels, and water buffalos. We have characterized representative strains of T. equiperdum and T. evansi by numerous molecular and classical parasitological approaches. We show that both species are actually strains of T. brucei, which lost part (Dk) or all (Ak) of their kDNA. These trypanosomes are not monophyletic clades and do not qualify for species status. They should be considered two subspecies, respectively T. brucei equiperdum and T. brucei evansi, which spontaneously arose recently. Dk/Ak trypanosomes may potentially emerge repeatedly from T. brucei.