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Humboldt-Universität zu Berlin - Ecology and Evolution of molecular Parasite-Host Interactions

Ökologie und Evolution molekularer Parasit-Wirts Interaktion

         

Prof. Dr. Emanuel Heitlinger

emanuelheitlinger@gmail.com

W1-S Juniorprofessor in collaboration with
the Leibniz Institute for Zoo and Wildlife research
 

Main projects of my group

Eimeria and other Pathogens in a house mouse hybrid zone

My group's work combines evolutionary questions with mechanistic aspects of the interaction between parasites and their hosts. For this purpose we aim to establish a model system for coevolution and local adaptation approachable both in its original ecological settings and in laboratory experiments. 

The hybrid zone of the two subspecies of the house mouse Mus musculus musculus und Mus musculus domesticus often determines population structure of parasites and can thus be regarded as a natural laboratory for adaptations and coevolution. It is possible to identify parasite genotypes adapted to a subspecies on the background of the well investigated mouse genomes. We use the house mouse as an approachable model host in both the field and the laboratory and establish its parasites as novel parts of a model system for coevoltionary processes. 

We have made first progress establishing such a system using whole genome sequencing of the mouse parasite Eimeria falciformisEimeria, with over one thousand species, is the largest genus in the phylum Apicomlexa. These species are highly host-specific and tightly adapted, each to its own of a huge variety of vertebrates. Up zu 16 Eimeria species have been described in house mice and are found at high prevalence throughout their hosts range, including the hybrid zone.  

My group tests in how far the population structure of these Eimeria species coincides with the host hybrid zone. We analyze whether specific genetic elements in Eimeria strains and species correlate with host genotypes and whether this is driven by an „adaptive introgression“ via hybridization. 

To this end we analyze neutral marker genes as well as candidate genes for parasite-host interaction and additionally develop high throughput approaches („genome wide capture“) for unbiased genome scans with an ultra dense set of makers. 

Many pathogenicity factors, well investigated in the close relative Toxoplasma gondii, are found in the genomes of Eimeria spp. We test the hypothesis, that especially variation in these factors determines the specificity of parasite strains. We analyze in a comparative genomics approach whether identified candidate regions and genes for a reciprocal adaptation are hot spots of differentiation acting as speciation genes.

In future experiments we will compare naturally occurring compatible and incompatible combinations of host and parasite strains in cross-infections. We will analyze how the identified combinations of host and parasite genotypes influence parasitological, histological and immunity related phenotypes. We will especially compare gene expression data from field studies with these infection experiments and infections with immunized and immune deficient (knock-out) mice. This work has  the potential to link phenotypes of a productive infection with genotypes of both host and parasite and to provide an understanding of the underlying molecular mechanisms. 

 

Collaborating scientists:
  • Joelle Gouy de Bellocq, Stuart J.E. Baird and Jaroslav Pialek (Institute of Vertebrate Biology of the Academy of Sciences of the Czech Republic). 
  • Richard Lucius, Nishith Gupta and Simone Spork (Humboldt University Berlin)

 

 


 


Metabarcoding analyses of parasite community ecology

 

It is often necessary to go beyond a „one host one parasite“ system to infer reciprocal adaptation and to include the whole parasite community of a host species and its ecology in the analysis.  As a first goal we want to test whether the occurrence of a specific parasite correlates with ecological and immunological parameters of the host and therefore need a fine resolution in diagnosing parasite species or strains. Secondly we want to study interactions between components of the parasite community and the interactions of this community composition with the mentioned ecological and immunological parameters of host individuals. To achieve this we need both comprehensive diagnosis of infections and high sample sizes. 

To this end we develop DNA-sequencing based high throughput methods to investigate non-invasive samples (i.e. fecal samples) for the presence of eukaryote parasites and other pathogens based on marker genes. These methods use highly conserved regions to enrich crude DNA extractions for marker genes of parasites. The enriched DNA contains variable in addition to the conserved regions, which are sequenced on high throughput platforms (currently predominantly Illumina MiSeq). 

We develop universal and specific primers and probes, which are sensitive for a broad variety of pathogens, while avoiding binding to host genes. We work on solutions for bioinformatic data analysis to complement the wet-lab approaches and to make full use of the generated information. 

We amplify for example large proportions of the rDNA gene clusters for all bigger taxonomic groups of parasites using parallelized microfluidics PCRs (the Fuidigm Access Array), resulting in sufficient genetic resolution to allow differentiation of parasite species. Doing so we provide a universal high throughput assay for the diagnosis of eukaryote parasites and other pathogens from fecal samples to groups of the IZW. The obtained data has the quality to not only group parasites into described species but to also define taxonomic units for unsubscribed species. 

We want to expand the approach in future to allow quantitative assessment of parasite infections using correlations between DNA-content and classical infection parameters (i.e. egg counts). We will try to complement these classical microscopy methods for parasite eggs with particle differentiation and counting or flow cytometry to scale such approaches to the throughput of our sequencing based assay. 

 

We apply these metabarcoding methods to target host species relevant for ecology and conservation collaborating with groups within the IZW: 

 

  1. We analyze the relation between reproduction, disease, immune and stress status of hyenae with the occurrence of different parasites and parasite strains. In this host the social status of the animals within the pack is considered and another focus is on the complex influence of dry and wet seasons, the resulting migrations and changes in prey spectrum on the parasite fauna.
  2. We genotype the parasites of the wolf using intestinal contents of wolves found dead and wolf feces. We analyze whether wolves are infected by parasites of farm and companion animals and reciprocally whether new parasites are introduced by wolves, where they threaten humans and their animals. 
  3. We also asses the complete parasite community of the house mouse sampled in the project described above. This will allow us to include the effect of additional pathogens in our analysis of a correlation of host and parasite genotypes with immunological parameters and to control for them. Furthermore this provides an unbiased approach to screen for all parasites corresponding in their population structure to that of the house mouse hybrid zone.

 

Collaborating scientists:
  • Oliver Krone and Ines Lesniak (IZW, wolf project)
  • Marion East und Heribert Hofer (IZW, hyena project)

 

 


 

Ph.D. students

 

Alice Balard

 

alice.balard@fu-berlin.de

 

Co-evolution of house mouse and an intracellular parasite, Eimeria spp.

 

 

My PhD project focuses on population genomics of parasite-host interaction. I am using next-generation sequencing, bioinformatic tools and statistical genetics to investigate the genome evolution of Eimeria spp. in two closely related subspecies of the house mouse. We are establishing the European house mouse hybrid zone (HMHZ) East of Berlin as a system to study host-parasite interactions. The HMHZ is a meeting point for two subspecies of the house mouse (Mus musculus domesticus and Mus musculus musculus), where they can hybridize. Eimeria spp. occur at high prevalence (around 30%) and differ in specificity for the mouse subspecies. This system allows us to investigate the interplay of adaptation and parasite specialization vs. generalism. We aim to address the effect of recombination between parasite strains and host genotypes on these processes.

 


 

Enas Khalifeh

 

khalifeh_enas@yahoo.com

 

Immunology in natural host-parasite interactions: Mus musculusEimeria model

My current research on the immunology of parasitic infections integrates molecular parasitology with immunology and pathology to understand how obligate intracellular parasites interact with host cells. I work on the intracellular parasite Eimeria spp. and its mouse host. Due to its short and direct life cycle Eimeria is ideal model for understanding immune reactions against intracellular parasites. Multiple species of Eimeria have been described from house mice and some of them are using different niches in the intestine (Duszynski 2001). My immunological approach uses Eimeria spp. from house mice hybrid zone. Preliminary data indicates that each of the investigated parental mice taxa (Mus m. musculus and Mus m. domesticus) preferentially harbor two different Eimeria strains.

 

 

The project includes, on one hand, studies with field isolates of parasites and their natural mouse host, on the other hand, infection studies with sympatric vs. allopatric Eimeria spp. – mouse host pairs. We test the immune status of mice (using e.g. measurements on cytokines and immunoglobulins) and reproductive output of parasite. We aim to identify immune strategies that may have arisen through antagonistic coevolution with these specific natural parasites. The specific objectives of the project are:

 

  1. Test whether immune system parameters correlated with efficiency in parasite elimination change between mice subspecies (Mus m. musculus and Mus m. domesticus) in HMHZ.
  2. Compare the mechanisms of intestinal tissue responses between the two mice subspecies in the context of resistance and susceptibility to the adapted- none adapted natural parasite.

 


 

Totta Kasemo

Totta Kasemo

 

totta.kasemo@gmail.com

 

In my project I ask how hosts recognize intestinal, unicellular parasites. As parasite models I study the intracellular mouse parasite E. falciformis, and the extracellular multi-host parasite G. duodenalis.

 

G. duodenalis cause giardiasis - sometimes
G. duodenalis infects about 280 million humans every year but data is ambiguous on which factors that determine the outcome of an infection. Patients range from being asymptomatic, to having the disease giardiasis with 2­-3 weeks of diarrhea, nausea, abdominal pain. If infection becomes chronic, malabsorption and intestinal barrier 
dysfunction follow. What determines the outcome of an infection is not known.
 
What does the parasite do?
During an infection, G. duodenalis remains on the surface of the small intestinal wall, where it attaches to cells but do not penetrate them. Electron microscopy images show marks ­like footprints in the villi structure where parasites have been attached, but apart from this it is unclear how the host might recognize its presence

 

 

 

 

 

 

My G. duodenalis project
In one part of my project, I investigate the role of G. duodenalis caused amino acid depletion, with a special interest in arginine. Previous work (references in e.g. Bartelt and Sartor, 2015) has shown both that host arginine is utilized by the parasite, but also that the parasite releases an arginine degrading enzyme, arginine deiminase, ADI. I design experiments to study how an infected host responds to this particular type of arginine depletion, and ask which cellular pathways are involved. The e xperiments aim to elucidate whether there is a specific cellular response to parasite induced arginine depletion, which is distinct from other types of arginine decrease or constitutively low levels. (References in e.g. Ryan and Cacció, 2013, Klotz and Aebischer, 2015, Bartelt and Sartor, 2015)

 

Apicomplexan parasite life cycle as seen by its RNA­seq transcriptome
In a RNA­seq transcriptome study, I compare E. falciformis’ development in wild­type laboratory mice with immunocompromised mice, in both first infection and in challenge infections.

 

 

E. falciformis i s closely related to Toxoplasma gondii which causes toxoplasmosis in humans and animals, and to Plasmodium spp., which can cause malaria. There are also hundreds, if not thousands, of related Eimeria spp., which cause severe problems in livestock such as chicken (e.g. Eimeria tenella).
 
 
Therefore, the life­cycle transcriptome of E. falciformis in a well described and natural host, the mouse, with different immune competences, is informative. Among other things, our data confirms host immune activity late in infection, as well as parasite upregulation of specific metabolic and biosynthetic activities at different life­cycle stages.
 

 


 Victor Jarquín


 

 vhjarquind@gmail.com

 

Filling the gap between metabarcoding and genotyping - a case study in Eimeria  spp., an intracellular parasite of house mice (Mus musculus). 
 
My research focus is the establishment of methods for amplification, sequencing and analysis of marker genes for parasite detection and phylogenetics. I am using diverse sets of genetic loci ranging from a few conserved targets, which can be amplified in a large range of taxa and have good representation in sequence databases, to large genome-wide sets of markers. I test which of those marker sets are capable to resolve “species level” taxa.
 
In first instance, I apply this strategy to intracellular parasites of the genus Eimeria. In this model we have the opportunity to use genes from three different genomes (nuclear, mitochondrial and one from an organelle called apicoplast). I assess methods and markers to distinguish different species and strains of Eimeria detected in two sub-species of the European house mouse Mus musculus musculus (eastern) and Mus musculus domesticus (western). I will focus on a deep level of resolution between closely related taxa . I will screen for strains of potentially hybrid parasites and conduct an analysis of the co-phylogeny of house mice and these parasite strains. 
 
In a second project, I improve methods to detect and distinguish a broad taxonomic range of protist and metazoan parasites. I develop, use and analyse markers which can be amplified in this range of taxa. I will test in how far these universal markers are able to resolve between closely related taxa. To do so I compare this approach with a the Eimeria-targetd approach described above in the house mouse model. 
I will then apply the resulting generalized metabarcoding approach with a defined power to assess parasites in a diverse set of wildlife species in collaboration with other groups at the IZW.
 

 


 

Little helpers:

 

Deborah Dymke   -  student assistant in the lab

Marius Bäsler       -  student assistant managing the hard-, and software