4 Material und Umwelt
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The termites are a derived eusocial lineage of otherwise non-social cockroaches. Understanding the proximal and ultimate drivers of this major evolutionary transition represents an important goal in biology. One outstanding question concerns the evolution of termite immunity, which is thought to have undergone broad-sweeping adaptations in order to enhance group-level immune protection. To understand the evolutionary origins of termite immunity, we conducted qualitative and quantitative transcriptome analyses along a gradient of sociality. Firstly, we aimed to identify large-scale genetic shifts in immune traits linked to eusociality by comparing immune gene repertoires in solitary and subsocial cockroaches and across a range of eusocial termite lineages. Secondly, we compared the responses of a solitary cockroach, a subsocial wood-roach and different castes of a lower termite species to a non-specific immune challenge, in order to understand how sociality may have influenced the evolution of immune gene regulation. Firstly, we found that termites have a broadly representative repertoire of canonical immune genes as compared to gregarious cockroaches and subsocial wood-roaches. Secondly, with respect to immune challenge, the solitary cockroach and the subsocial wood-roach displayed a similarly comprehensive induced response, while the termite response was considerably dampened by comparison and strongly influenced by caste; with reproductives displaying a generally higher constitutive level of immune-gene expression compared to sterile castes. In summary we did not find termite eusociality to be associated with significant changes in immune gene diversity, but rather to be linked with significant modifications to the regulation of immunity following the origin of division of labour.
A broad suite of immune adaptations have evolved in social insects which hold close parallels with the immune systems of multicellular individuals. However, comparatively little is known about the evolutionary origins of immunity in social insects. We tackle this by identifying immune genes from 18 cockroach and termite species, spanning a gradient of social lifestyles. Termites have undergone contractions of major immune gene families during the early origin of the group, particularly in antimicrobial effector and receptor proteins, followed by later re-expansions in some lineages. In a comparative gene expression analysis, we find that reproductive individuals of a termite invest more in innate immune regulation than other castes. When colonies encounter immune-challenged nestmates, gene expression responses are weak in reproductives but this pattern is reversed when colony members are immune-challenged individually, with reproductives eliciting a greater response to treatment than other castes. Finally, responses to immune challenge were more comprehensive in both subsocial and solitary cockroaches compared to termites, indicating a reduced overall ability to respond to infection in termites. Our study indicates that the emergence of termite sociality was associated with the evolution of a tapered yet caste-adapted immune system.
Fungi that share light-flooded habitats with phototrophs may profit from excess photosynthetic products. But to cope with sunlight-associated stresses it is important for fungi to accurately sense and respond to changes in light. To test the hypothesis that light is an environmental cue that Ascomycota use to coordinate growth, stress responses as well as to establish pathogenic or symbiotic relationships, the photoreceptor (PR) distribution in species from different ecological niches was analysed. The genomes of black fungi from phyllosphere and exposed solid surfaces contain multiple photoreceptors (PRs). The filamentous foliar plant pathogen Botrytis cinerea (Leotiomycetes) has a highly sophisticated photosensory and signalling system that helps to avoid light and to locate susceptible hosts. Rock-inhabiting Dothideo- and Eurotiomycetes including Knufia petricola possess equal numbers of PRs along with the same set of protective pigments. This similarity between black fungi from plant and rock surfaces suggests that photoperception and photoregulation are important for fungi that avoid loss of energy and nutrients through cooperation with phototrophs.
Fungi that share light-flooded habitats with phototrophs may profit from excess photosynthetic products. But to cope with sunlight-associated stresses it is important for fungi to accurately sense and respond to changes in light. To test the hypothesis that light is an environmental cue that Ascomycota use to coordinate growth, stress responses as well as to establish pathogenic or symbiotic relationships, the photoreceptor (PR) distribution in species from different ecological niches was analysed. The genomes of black fungi from phyllosphere and exposed solid surfaces contain multiple photoreceptors (PRs). The filamentous foliar plant pathogen Botrytis cinerea (Leotiomycetes) has a highly sophisticated photosensory and signalling system that helps to avoid light and to locate susceptible hosts. Rock-inhabiting Dothideo- and Eurotiomycetes including Knufia petricola possess equal numbers of PRs along with the same set of protective pigments. This similarity between black fungi from plant and rock surfaces suggests that photoperception and photoregulation are important for fungi that avoid loss of energy and nutrients through cooperation with phototrophs.
Sunlight, the major source of energy, drives life but in excess it provokes UV-induced DNA damage, accumulation of reactive oxygen species, desiccation, osmotic stresses and so on. Biological strategies to survive excess light include protection, avoidance and active utilisation. Phototrophic organisms including cyanobacteria, green algae and plants use all three strategies. Fungi occupying phototrophic niches may profit from the surplus photosynthetic products which also obliges them to cope with the same light-induced stresses. One way of handling these stresses would be to use similar signalling pathways to sense the presence and to interact with their phototrophic partners. Obviously, the levels of adaptation and responses to light will depend on the environment as well as fungal genotype and phylogenetic position. Black fungi – a polyphyletic group – accumulate the dark pigment DHN melanin in their cell walls and often occupy light-flooded habitats from phyllosphere to rock surfaces. Here we compare two sequenced melanised fungi of different lifestyles in their response to light.
The Leotiomycete Botrytis cinerea is an aggressive pathogen that primarily infects the above-ground parts of plants. It possesses large numbers of photoreceptors that respond to a broad spectrum of light. As a consequence, light controls morphogenesis in which it induces conidiation for disease spreading and represses sclerotial development for survival and/or sexual recombination. Cellular components involved in photo-perception and regulation of morphogenesis, stress responses and virulence have been identified and appear to regulate propagation, survival and infection. These include phytochromes, a group of photoreceptors which are particularly enriched in the Leotiomycetes and that mediate coordinated responses to light and elevated temperatures. Assuming that photo-regulation may be equally important for fungi that live in mutualistic relationships with phototrophs either by forming composite organisms or biofilms, we investigate the role of light in the rock-inhabiting Eurotiomycete Knufia petricola. Like other black yeasts, K. petricola grows slowly, does not form specialised reproductive structures and constitutively produces DHN melanin as well as carotenoids. Combining K. petricola with the cyanobacterium Nostoc punctiforme, we developed a model system for studying biofilm formation and bio-weathering. A genetic toolbox to manipulate this model system is being developed. K. petricola strain A95 possesses ten putative photoreceptors, more than found in filamentous Eurotiomycetes suggesting that light plays an important role for abiotic and biotic interactions in extremo-tolerant and symbiosis-capable fungi.
Simple particulate matter sensors are gaining popularity due to their low price, easy handling and good temporal resolution. In this presentation, we report on the performance of a Sharp optical PM sensor GP2Y1010AU0F, which costs less than 15 €.
The sensor is built around an infrared emitting diode (ILED) and a phototransistor detecting the light scattered from the aerosol particle. An electronic circuit shapes the detected light in a pulsed signal. The manufacturer advises sampling the output signal 280 microseconds after the ILED pulse. The measured output voltage is an indicator of dust concentration.
We have built two identical simple PM monitoring devices using Raspberry Pi 3 computer interfacing the PM sensor with Microchip’s MCP3002 ADC via SPI. The ADC is capable of more than 100 ksamples/s at 10-bit resolution. The Rpi3 was pulsing the sensor at 10Hz, digitizing and saving the data and sending the results wirelessly.
Sensor’s output pulse shape was sampled with 10 microsecond time steps and saved, thus making offline analysis possible. A time jitter of output pulses can be observed and suggests a peak fitting as a better approach to the signal readout compared to the single sampling at a fixed time after pulse triggering We compared both methods.
In arid sun-exposed environments, rock-inhabiting fungi dominate microbial communities on rock surfaces. Fungi are known to enhance rock weathering, although the exact mechanisms are not completely understood. These natural processes can be simulated by reproducible experimental systems. Particularly attractive are genetically amenable rock-weathering fungi, where certain traits can be knocked-out to identify weathering-relevant functions. Here, we studied the effect of the rock-inhabiting fungus Knufia petricola A95 and its melanin-deficient mutant (ΔKppks) on the weathering of a Fe-bearing olivine. We examined the olivine dissolution kinetics and looked at the olivine-fungus interface. For the former we inoculated olivine powder with the fungus in batch and mixed flow reactors and analysed the medium solution by ICP-OES, for the latter biofilm-covered olivine sections from flow-through reactors were analysed by FIB-TEM.
In general olivine dissolution was lower when olivine was incubated without fungi: the abiotic olivine dissolution rates were lowest and the surface of the abiotically reacted olivine sections showed no etching. Various chemical analyses show Fe (oxyhydr)oxide precipitation on top of the abiotically reacted olivine, indicating its role in inhibiting olivine dissolution. Both the wild type (WT) and ΔKppks solubilised and bound significant amounts of Fe released by olivine dissolution. Fe (oxyhydr)oxide precipitation was thus reduced, explaining the enhanced olivine dissolution in their presence. Particularly efficient at preventing Fe precipitation were attached WT cells: the inhibition of olivine dissolution almost completely disappeared. The attachment capacity of the WT is most likely caused by the production of WT-specific extracellular polymeric substances. Our presented experimental systems allowed the precipitation of Fe (oxyhydr)oxides and included a rock-inhabiting fungus and thus simulated weathering processes relevant to natural ecosystems.
Rice is staple food of nearly half the world’s population. Rice yields must therefore increase to feed ever larger populations. By colonising rice and other plants, Herbaspirillum spp. stimulate plant growthand productivity. However the molecular factors involved are largely unknown. To further explore this interaction, the transcription profiles of Nipponbare rice roots inoculated with Herbaspirillum seropedicae were determined by RNA-seq. Mapping the 104 million reads against the Oryza sativa cv. Nipponbare genome produced 65 million unique mapped reads that represented 13,840 transcripts each with at least two-times coverage. About 7.4% (1,014) genes were differentially regulated and of these 255 changed expression levels more than two times. Several of the repressed genes encoded proteins related to plant defence (e.g. a putative probenazole inducible protein), plant disease resistance as well as enzymes involved in flavonoid and isoprenoid synthesis. Genes related to the synthesis and efflux of phytosiderophores (PS) and transport of PS-iron complexes were induced by the bacteria. These data suggest that the bacterium represses the rice defence system while concomitantly activating iron uptake. Transcripts of H. seropedicae were also detected amongst which transcripts of genes involved in nitrogen fixation, cell motility and cell wall synthesis were the most expressed.
In the modern world there is an increased understanding that design and performance monitoring of materials have to be tested in connection to chemical, physical and (micro)biological challenges. A systematic study on how biofilms interact with materials and what could be done to engineer biofilms and/or materials in order to maximize the resistance of the material (surface) or the resistance the biofilm-modified material (bulk) is in strong need. In the Department “Materials and the Environment” of the BAM new experimental platform is being developed. With the help of different type of device for high throughput and microbiologically-controlled environment simulation we establish a new approach to clarify the mechanisms of biofilm/material interactions. Despite the focus on fundamental research, the main results of this project proposal will be transferable into material technology and construction chemistry and will influence the development of standardization in this topic. As the interactions of biofilms and materials have implications for most constructions as well as climate change, the results of the research generates additional value.
Mechanisms of olivine dissolution by rock-inhabiting fungi explored using magnesium stable isotopes
(2019)
To unravel the dissolution mechanisms of olivine by a rock-inhabiting fungus we determined the stable isotope ratios of Mg on solutions released in a laboratory experiment. We found that in the presence of the fungus Knufia petricola the olivine dissolution rates were about seven-fold higher (1.04×10−15 mol cm−2 s−1) than those in the abiotic experiments (1.43×10−16 mol cm−2 s−1) conducted under the same experimental condition (pH 6, 25 °C, 94 days). Measured element concentrations and Mg isotope ratios in the supernatant solutions in both the biotic and the abiotic experiment followed a dissolution trend in the initial phase of the experiment, characterized by non-stoichiometric release of Mg and Si and preferential release of 24Mg over 26Mg. In a later phase, the data indicates stoichiometric release of Mg and Si, as well as isotopically congruent Mg release. We Attribute the initial non-stoichiometric phase to the rapid replacement of Mg2+ in the olivine with H+ along with simultaneous polymerization of Si tetrahedra, resulting in high dissolution rates, and the stoichiometric phase to be influenced by the accumulation of a Si-rich amorphous layer that slowed olivine dissolution. We attribute the accelerated dissolution of olivine during the biotic experiment to physical attachment of K. petricola to the Si-richamorphous layer of olivine which potentially results in ist direct exposure to protons released by the fungal cells. These additional protons can diffuse through the Si-rich amorphous layer into the crystalline olivine. Our results also indicate the ability of K. petricola to dissolve Fe precipitates in the Si-rich amorphous layer either by protonation, or by Fe(III) chelation with siderophores. Such dissolution of Fe precipitates increases the porosity of the Si-rich amorphous layer and hence enhances olivine dissolution. The acceleration of mineral dissolution in the presence of a rock-dissolving fungus further suggests that its presence in surficial CO2 sequestration plants may aid to accelerate CO2 binding.
The story of how black fungi survive harsh conditions on sun-exposed desert rocks and material surfaces is fascinating. In the presentation examples of how knowledge of these organisms can be of practical value (e.g., in biodeterioration studies, such as on stone markers in cemeteries) would be given. Among other stories, the example of how roof tiles amended with a titanium oxide layers as a biocide actually selected for the black fungi will be told. A hypothesis about how these organisms would likely be found on solar panels and some early BAM work in that area will be presented. This then brings the connection to a study ripe for investigation in North Carolina. The talk presents the development of a study aiming at deciphering the influence of microbial biofilm formation on the energy conversion efficiency of solar photovoltaic panels or modules at two facilities (one facility under the impact of high intensity of animal agriculture and high deposition of ammonia from atmosphere and the other with low animal agriculture and lower atmospheric ammonia deposition) in North Carolina. The main hypothesis of the study is that microbial biofilm formation on solar photovoltaic panels will lead to significant decreases in energy conversion efficiency of solar photovoltaic modules and biofilm formation will also be accelerated by high ammonia concentration in the ambient atmosphere and high nitrogen deposition.
Interface between the atmosphere and mineral substrates is the oldest terrestrial habitat. Morphologically simple microbial biofilms were the first settlers on these inhospitable surfaces at times when the Earth was inhabited only by microorganisms and the solid substrates represented only by natural rock surfaces i.e. lithosphere. Miniature, self-sufficient microbial ecosystems continue to develop on subaerial (i.e. air-exposed) solid surfaces at all altitudes and latitudes where direct contact with the atmosphere and solar radiation occurs – on rocks, mountains, buildings, monuments, solar panels. All these sub-aerial biofilms develop under fluctuating and hostile conditions – and thus frequently harbour stress-tolerant black fungi inherently able to cope with the stresses of bright sunlight and constantly changing atmospheric conditions. Black fungi – a polyphyletic group of Ascomycetes– accumulate the dark pigment DHN melanin, diverse carotenoids and mycosporines in their cells and thus successfully colonise sunlight-flooded habitats from phyllosphere to rock surfaces. Various chemical and physical extremes and fluctuating environments belong to the challenges effectively mastered by black fungi. In our laboratory we isolate novel black fungi from man-made habitats like building materials and solar panels. Using Knufia petricola A95 as a model we conduct experiments to clarify interactions of black fungi with inorganic substrates. We use available mutants to determine the functional consequences of changes in the outer cell wall envelopes – from excreted EPS to layers of protective pigments. A genetic toolbox to manipulate this Chaetothyriales representative is in further development. Our long-term goal is to understand the fundamental mechanisms how black fungi are able (i) to adhere to dry atmosphere-exposed surfaces, (ii) to survive multiple stresses and (iii) to change the underlying substrates including rocks.
Social insects nesting in soil environments are in constant contact with entomopathogens and have evolved disease resistance mechanisms within a colony to prevent the occurrence and spread of infectious diseases. Among these mechanisms: mutual grooming reduces the cuticular load of pathogens, and burial of cadavers and cannibalism can prevent pathogens from replicating within the group. We explored how the rate and type of collective behavioural response is determined by stepwise infection dynamics operating at the level of the individual. Specifically, we infected the eastern subterranean termite Reticulitermes flavipes with different types of infectious particle and infection route of the entomopathogenic fungus Metarhizium anisopliae and recorded behavioural responses of nestmates to individuals at different times during the progression of infections. As expected, termites groomed conidia-exposed individuals significantly more than controls. Interestingly, grooming was significantly elevated after fungal germination than before, suggesting that pathogen growth cues act as strong stimulators of allogrooming. Conidia-exposed termites were cannibalized, but only after they became visibly ill. By contrast, termites did not groom blastospore-injected individuals more than controls at any time-point following infection. Instead, we found that blastospore-injected individuals were continually cannibalized at a low-level following injection with either viable or heat-killed blastospores, with a marked increase in cannibalism after termites injected with viable blastospores became visibly ill and were close to death. Together, these findings point to the importance of host condition as a cue for social hygienic behavior, and that the host itself appears to emit essential sickness cues that act as targets for its own sacrifice. This demonstrates that termites have independently evolved to both identify and destructively respond to sickness.
The rotation of the Earth causes a day/night cycle that is characterized by changing light and temperature conditions. Fungi actively sense the environmental light conditions to induce protective mechanisms against the light-associated stresses and to regulate development. Since fungi adapted to habitats with different light regimes, the complexities of their ‘visual’ systems and photoresponses may vary significantly. Fungi that are associated with plants experience a special light regime because the host seeks optimum light conditions for photosynthesis – and the fungus must cope with them. Moreover, fungi living under the canopy are confronted with an altered spectrum enriched for green and far-red light. Plants sense light to coordinate growth and morphogenesis: a low red:far-red ratio indicates the presence of competitors (shading by other plants) and triggers the shade avoidance response which is accompanied by increased susceptibility to nectrophic pathogens.
Botrytis cinerea, the causal agent of gray mold diseases on many plant species in moderate climate zones (high humidity, temperatures around 20°C, seasons), exhibits striking photoresponses accompanied by a broad action spectrum and a high number of photoreceptors. Light is the most important factor controlling morphogenesis: it induces conidiation for disease spreading (summer cycle) and represses sclerotial development for survival and/or sexual recombination (winter cycle). In the past years, we identified several cellular components involved in photoperception and regulation of photomorphogenesis and virulence indicating that the same signaling pathways are of relevance for both processes, propagation and infection. Notably, we recognized the role of the phytochromes (red/far-red light sensors) for coordinated responses to light and elevated temperatures (photo-/thermomorphogenesis) – similar to the function of Arabidopsis PhyB. BcPHY2-like thermosensors are likely restricted to the Leotiomycetes, which include several plant pathogens of the moderate climate zones.
Results suggest a reduction in immune gene repertoires in termites and possible complementary expression between termite castes. With comparative genomics we will investigate the evolution of gene families related to immunity, try to understand where reductions and expansions take place and relate these changes to shifts in sociality and ecology. The role of TEs in expansions and contractions of immune gene families will be investigated. For these analyses, we propose to generate high quality, highly contiguous genomes of species from different levels of sociality, covering all major termite families. With comparative transcriptomics we will investigate the expression of immune genes in different castes. Via network analyses we will identify pathways indicated in differential immunity between castes and between species of different sociality levels. We will investigate how these pathways have been rewired along the transitions to higher levels of sociality and how, intra-specifically, they change between castes.
Wild bees are important pollinators for agricultural crops and solitary species such as Osmia bicornis are particularly suitable for pollination management. Wild bees share floral resources with managed honey bees and may be exposed to emerging infectious diseases. Although studies have explored the prevalence of pathogens in solitary wild bee species, data regarding the impact of pathogens on solitary bee health are lacking.
We carried out experiments examining whether the solitary bee species O. bicornis is susceptible to infection with the emerging pathogen The results obtained indicate that N. ceranae may be able to infect O. bicornis but its impact on host fitness is negligible: survival rates did not differ between Control and inoculated bees, although male survival was marginally lower after infection. To explore the possible field-relevance of our findings, we collected wild bees near an infected and a non-infected hive and showed that N. ceranae was shared between managed and wild bees, although only the in presence of infected honey bees.
The findings of the present study show that O. bicornis is susceptible to pathogen spillover and could act as a potential reservoir host for N. ceranae in pollinator networks. Additional studies on this species incorporating sublethal effects, multiple infections and other interacting stressors are warranted.
Wild bees are important pollinators of wild plants and agricultural crops and they are threatened by several environmental stressors including emerging pathogens. Honey bees have been suggested as a potential source of pathogen spillover. One prevalent pathogen that has recently emerged as a honey bee disease is the microsporidian Nosema ceranae. While the impacts of N. ceranae in honey bees are well documented, virtually nothing is known about its effects in solitary wild bees.
The solitary mason bee Osmia bicornis is a common pollinator in orchards and amenable to Commercial management. Here, we experimentally exposed larvae of O. bicornis to food contaminated with N. ceranae and document spore presence during larval development. We measured mortality, growth parameters, and timing of pupation in a semi-field experiment. Hatched individuals were assessed for physiological state including fat body mass, wing muscle mass, and body size. We recorded higher mortality in the viable-spore-exposed group but could only detect a low number of Spores among the individuals of this treatment. Viable-spore-treated individuals with higher head capsule width had a delayed pupation start. No impact on the physiological status could be detected in hatched imagines. Although we did not find overt evidence of O. bicornis infection, our findings indicate that exposure of larvae to viable N. ceranae spores could affect bee development.
Black yeasts form a polyphyletic group of Ascomycota that colonize bare surfaces like rocks, facades of buildings, and solar panels. Their protective adaptations enable an adequate response to fluctuating and diverse temperature, water and UV radiation stresses. Together with bacteria and algae they form sub-aerial biofilms (SAB) this way discoloring and weathering the surfaces they grow on. Strain A95 of Knufia petricola (Eurotiomycetes, Chaetothyriales) displays both typical yeast-like cell growth and constitutive dihydroxynaphthalene (DHN) melanogenesis. Along with the cyanobacterium Nostoc punctiforme as photobiont, it is already used in a model system for studying SAB formation and bio-weathering. Applying the recently developed tools for the generation of deletion mutants will allow to define gene functions and to identify genes critical for abiotic and biotic interactions. We present a chromosome-level genome assembly and annotation for K. petricola A95. The genome was assembled with MaSuRCA using a hybrid assembly approach of Illumina MiSeq and PacBio SMRT sequencing data. The resulting assembly consists of 17 contigs including the complete mitochondrial genome and five complete chromosomes. It shows indication of repeat-induced point mutations (RIP). Supported by RNA sequencing data from eight different growth conditions, 10,994 genes were predicted with the BRAKER2 pipeline. Functional annotation of genes was obtained from general functional annotation databases and the fungal specific database FungiPath. Comparative analyses are in progress to identify genes specific to black yeasts, that may facilitate the survival on exposed surfaces. In sum, the genome sequence of K. petricola is a valuable resource to gain insight into the protein inventory and functional pathways of extremotolerant and symbiosis-capable fungi.
Termitidae comprises 80% of all termite species that play dominant decomposer roles in Tropical cosystems. Two major events during Termite evolution were the loss of cellulolytic gut protozoans in the ancestor of Termitidae and the subsequent gain in the termitid subfamily Macrotermitinae of fungal symbionts cultivated externally in ‘‘combs’’ constructed within the nest. How these symbiotic transitions occurred remains unresolved. Phylogenetic analyses of mitochondrial data previously suggested that Macrotermitinae is the earliest branching termitid lineage, followed soon after by Sphaerotermitinae, which cultivates bacterial symbionts on combs inside its nests. This has led to the hypothesis that comb building was an important evolutionary step in the loss of gut protozoa in ancestral termitids. We sequenced genomes and transcriptomes of 55 termite species and reconstructed phylogenetic trees from up to 4,065 orthologous genes of 68 species. We found strong support for a novel sister-group relationship between the bacterial comb-building Sphaerotermitinae and fungus comb-building Macrotermitinae.
This key finding indicates that comb building is a derived trait within Termitidae and that the creation of a comb-like ‘‘external rumen’’ involving bacteria or fungi may not have driven the loss of protozoa from ancestral termitids, as previously hypothesized.
Instead, associations with gut prokaryotic symbionts, combined with dietary shifts from wood to other plant-based substrates, may have played a more important role in this symbiotic transition. Our phylogenetic tree provides a platform for future studies of comparative termite evolution and the evolution of symbiosis in this taxon.
Black microcolonial fungi (MCF) are persistent inhabitants of rock surfaces in hostile desert environments. In these niches, MCF have evolved mineral-weathering and symbiotic capabilities as well as mechanisms to cope with multiple stresses such as solar irradiation, temperature extremes and low water activity. Due to their stress tolerance these ascomycetes are prominent in modern terrestrial ecosystems – like man-made material surfaces from roof to solar panels. MCF interactive capabilities support their facultative symbiotic relationships with cyanobacteria and ensure their rock-weathering geochemical activity.
Using the rock-inhabiting fungus K. petricola A95 (Chaetothyriales), we developed transformation protocols and deleted genes responsible for production of the protective pigments melanins and carotenoids. To confirm that the mutant phenotypes were not due to hidden mutations, melanin synthesis was restored by complementing the mutants with the respective wild type genes. Strains of K. petricola carrying gene variants for fluorescent proteins EGFP and DsRed are available. We successfully labelled the cytoplasm, nuclei, peroxisomes and mitochondria. Targeted and ectopic integrations result in stable transformants suitable for further phenotypical characterization. As K. petricola is a non-pathogenic fungus with all characteristic features of MCF, including meristematic growth, melanized cell-walls, extracellular polymeric substances and extensive pigment production, our results will shed light on protective role of pigments during cell wall maturation and oxidative stress defence in rock-inhabiting MCF. Genes involved in environmental sensing or substrate and phototroph interactions are currently targeted. With the help of a mutant collection and fluorescently labelled K. petricola we will be able to investigate interactions of MCF with environmental stressors, mineral substrates, soil matrices and phototrophic symbionts.