BackgroundThe identification and characterization of the transcriptional regulatory networks governing the physiology and adaptation of microbial cells is a key step in understanding their behaviour. One such wide-domain regulatory circuit, essential to all cells, is carbon catabolite repression (CCR): it allows the cell to prefer some carbon sources, whose assimilation is of high nutritional value, over less profitable ones. In lower multicellular fungi, the C2H2 zinc finger CreA/CRE1 protein has been shown to act as the transcriptional repressor in this process. However, the complete list of its gene targets is not known.ResultsHere, we deciphered the CRE1 regulatory range in the model cellulose and hemicellulose-degrading fungus Trichoderma reesei (anamorph of Hypocrea jecorina) by profiling transcription in a wild-type and a delta-cre1 mutant strain on glucose at constant growth rates known to repress and de-repress CCR-affected genes. Analysis of genome-wide microarrays reveals 2.8% of transcripts whose expression was regulated in at least one of the four experimental conditions: 47.3% of which were repressed by CRE1, whereas 29.0% were actually induced by CRE1, and 17.2% only affected by the growth rate but CRE1 independent. Among CRE1 repressed transcripts, genes encoding unknown proteins and transport proteins were overrepresented. In addition, we found CRE1-repression of nitrogenous substances uptake, components of chromatin remodeling and the transcriptional mediator complex, as well as developmental processes.ConclusionsOur study provides the first global insight into the molecular physiological response of a multicellular fungus to carbon catabolite regulation and identifies several not yet known targets in a growth-controlled environment.

Summary Trichoderma reesei is an industrial producer of enzymes that degrade lignocellulosic polysaccharides to soluble monomers, which can be fermented to biofuels. Here we show that the expression of genes for lignocellulose degradation are controlled by the orthologous T. reesei protein methyltransferase LAE1. In a lae1 deletion mutant we observed a complete loss of expression of all seven cellulases, auxiliary factors for cellulose degradation, β-glucosidases and xylanases were no longer expressed. Conversely, enhanced expression of lae1 resulted in significantly increased cellulase gene transcription. Lae1-modulated cellulase gene expression was dependent on the function of the general cellulase regulator XYR1, but also xyr1 expression was LAE1-dependent. LAE1 was also essential for conidiation of T. reesei. Chromatin immunoprecipitation followed by high-throughput sequencing (“ChIP-seq”) showed that lae1 expression was not obviously correlated with H3K4 di- or trimethylation (indicative of active transcription) or H3K9 trimethylation (typical for heterochromatin regions) in CAZyme coding regions, suggesting that LAE1 does not affect CAZyme gene expression by directly modulating H3K4 or H3K9 methylation. Our data demonstrate that the putative protein methyltransferase LAE1 is essential for cellulase gene expression in T. reesei through mechanisms that remain to be identified.

Due to its capacity to produce large amounts of cellulases, Trichoderma reesei is increasingly being investigated for second-generation biofuel production from lignocellulosic biomass. The induction mechanisms of T. reesei cellulases have been described recently, but the regulation of the genes involved in their transcription has not been studied thoroughly. Here we report the regulation of expression of the two activator genes xyr1 and ace2, and the corepressor gene ace1, during the induction of cellulase biosynthesis by the inducer lactose in T. reesei QM 9414, a strain producing low levels of cellulase (low producer). We show that all three genes are induced by lactose. xyr1 was also induced by D-galactose, but this induction was independent of D-galactose metabolism. Moreover, ace1 was carbon catabolite repressed, whereas full induction of xyr1 and ace2 in fact required CRE1. Significant differences in these regulatory patterns were observed in the high-producer strain RUT C30 and the hyperproducer strain T. reesei CL847. These observations suggest that a strongly elevated basal transcription level of xyr1 and reduced upregulation of ace1 by lactose may have been important for generating the hyperproducer strain and that thus, these genes are major control elements of cellulase production.

Numerous bacterial genetic markers are available for the molecular detection of human sources of fecal pollution in environmental waters. However, widespread application is hindered by a lack of knowledge regarding geographical stability, limiting implementation to a small number of well-characterized regions. This study investigates the geographic distribution of five human-associated genetic markers (HF183/BFDrev, HF183/BacR287, BacHum-UCD, BacH, and Lachno2) in municipal wastewaters (raw and treated) from 29 urban and rural wastewater treatment plants (750–4 400 000 population equivalents) from 13 countries spanning six continents. In addition, genetic markers were tested against 280 human and nonhuman fecal samples from domesticated, agricultural and wild animal sources. Findings revealed that all genetic markers are present in consistently high concentrations in raw (median log10 7.2–8.0 marker equivalents (ME) 100 mL–1) and biologically treated wastewater samples (median log10 4.6–6.0 ME 100 mL–1) regardless of location and population. The false positive rates of the various markers in nonhuman fecal samples ranged from 5% to 47%. Results suggest that several genetic markers have considerable potential for measuring human-associated contamination in polluted environmental waters. This will be helpful in water quality monitoring, pollution modeling and health risk assessment (as demonstrated by QMRAcatch) to guide target-oriented water safety management across the globe.

The microbial faecal pollution of rivers has wide-ranging impacts on a variety of human activities that rely on appropriate river water quality. Thus, detailed knowledge of the extent and origin of microbial faecal pollution is crucial for watershed management activities to maintain safe water use. In this study, the microbial faecal pollution levels were monitored by standard faecal indicator bacteria (SFIB) along a 2580 km stretch of the Danube, the world's most international river, as well as the Danube's most important tributaries. To track the origin of faecal pollution, host-associated Bacteroidetes genetic faecal marker qPCR assays for different host groups were applied in concert with SFIB. The spatial resolution analysis was followed by a time resolution analysis of faecal pollution patterns over 1 year at three selected sites. In this way, a comprehensive faecal pollution map of the total length of the Danube was created, combining substantiated information on both the extent and origin of microbial faecal pollution. Within the environmental data matrix for the river, microbial faecal pollution constituted an independent component and did not cluster with any other measured environmental parameters. Generally, midstream samples representatively depicted the microbial pollution levels at the respective river sites. However, at a few, somewhat unexpected sites, high pollution levels occurred in the lateral zones of the river while the midstream zone had good water quality. Human faecal pollution was demonstrated as the primary pollution source along the whole river, while animal faecal pollution was of minor importance. This study demonstrates that the application of host-associated genetic microbial source tracking markers in concert with the traditional concept of microbial faecal pollution monitoring based on SFIB significantly enhances the knowledge of the extent and origin of microbial faecal pollution patterns in large rivers. It constitutes a powerful tool to guide target-oriented water quality management in large river basins.

Over the past 15 years, pioneering interdisciplinary research has been performed on the microbiology of hydrogeologically well‐defined alpine karst springs located in the Northern Calcareous Alps (NCA) of Austria. This article gives an overview on these activities and links them to other relevant research. Results from the NCA springs and comparable sites revealed that spring water harbors abundant natural microbial communities even in aquifers with high water residence times and the absence of immediate surface influence. Apparently, hydrogeology has a strong impact on the concentration and size of the observed microbes, and total cell counts (TCC) were suggested as a useful means for spring type classification. Measurement of microbial activities at the NCA springs revealed extremely low microbial growth rates in the base flow component of the studied spring waters and indicated the importance of biofilm‐associated microbial activities in sediments and on rock surfaces. Based on genetic analysis, the autochthonous microbial endokarst community (AMEC) versus transient microbial endokarst community (TMEC) concept was proposed for the NCA springs, and further details within this overview article are given to prompt its future evaluation. In this regard, it is well known that during high‐discharge situations, surface‐associated microbes and nutrients such as from soil habitats or human settlements—potentially containing fecal‐associated pathogens as the most critical water‐quality hazard—may be rapidly flushed into vulnerable karst aquifers. In this context, a framework for the comprehensive analysis of microbial pollution has been proposed for the NCA springs to support the sustainable management of drinking water safety in accordance with recent World Health Organization guidelines. Near‐real‐time online water quality monitoring, microbial source tracking (MST) and MST‐guided quantitative microbial‐risk assessment (QMRA) are examples of the proposed analytical tools. In this context, this overview article also provides a short introduction to recently emerging methodologies in microbiological diagnostics to support reading for the practitioner. Finally, the article highlights future research and development needs.This article is categorized under: Engineering Water > Water, Health, and SanitationScience of Water > Water ExtremesWater and Life > Nature of Freshwater Ecosystems

BackgroundFilamentous fungi are frequently used as production platforms in industrial biotechnology. Most of the strains involved were known as reproducing exclusively asexually thereby preventing the application of conventional strain breeding techniques. In the last decade, evidence was obtained that a number of these imperfect fungi possess a sexual life cycle, too. Trichoderma reesei, an industrial producer of enzymes for food, feed and biorefinery purposes, is heterothallic and takes a special position among industrially utilized species as all industrial strains are derived from the single MAT1-2 isolate QM6a. Consequently, strain improvement by crossing is not feasible within this strain line as this necessitates a MAT1-1 mating partner. Simply switching the mating type in one of the mating partners to MAT1-1, however, is not sufficient to produce a genotype capable of sexual reproduction with QM6a MAT1-2.ResultsWe have used a systems biology approach to identify genes restoring sexual reproduction in the QM6a strain line. To this end, T. reesei QM6a was crossed with the MAT1-1 wild-type strain CBS999.97. The descendants were backcrossed 8-times in two lineages with QM6a to obtain mating competent MAT1-1 strains with a minimal set of CBS999.97 specific genes. Comparative genome analysis identified a total of 73 genes of which two—encoding an unknown C2H2/ankyrin protein and a homolog of the WD-protein HAM5—were identified to be essential for fruiting body formation. The introduction of a functional ham5 allele in a mating type switched T. reesei QM6a allowed sexual crossing with the parental strain QM6a.ConclusionThe finding that Trichoderma reesei is generally capable of undergoing sexual reproduction even under laboratory conditions raised hope for the applicability of classical breeding techniques with this fungus as known for plants and certain yeasts. The discovery that the wild-type isolate QM6a was female sterile, however, precluded any progress along that line. With the discovery of the genetic cause of female sterility and the creation of an engineered fertile strain we now provide the basis to establish sexual crossing in this fungus and herald a new era of strain improvement in T. reesei.Electronic supplementary materialThe online version of this article (doi:10.1186/s13068-015-0311-2) contains supplementary material, which is available to authorized users.

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