Extreme overall mushroom genome expansion in Mycena s.s. irrespective of plant hosts or substrate specializations.

Mycena s.s. is a ubiquitous mushroom genus whose members degrade multiple dead plant substrates and opportunistically invade living plant roots. Having sequenced the nuclear genomes of 24 Mycena species, we find them to defy the expected patterns for fungi based on both their traditionally perceived saprotrophic ecology and substrate specializations. Mycena displayed massive genome expansions overall affecting all gene families, driven by novel gene family emergence, gene duplications, enlarged secretomes encoding polysaccharide degradation enzymes, transposable element (TE) proliferation, and horizontal gene transfers. Mainly due to TE proliferation, Arctic Mycena species display genomes of up to 502 Mbp (2-8× the temperate Mycena), the largest among mushroom-forming Agaricomycetes, indicating a possible evolutionary convergence to genomic expansions sometimes seen in Arctic plants. Overall, Mycena show highly unusual, varied mosaic-like genomic structures adaptable to multiple lifestyles, providing genomic illustration for the growing realization that fungal niche adaptations can be far more fluid than traditionally believed.

Mycena species can be opportunist-generalist plant root invaders.

Traditional strict separation of fungi into ecological niches as mutualist, parasite or saprotroph is increasingly called into question. Sequences of assumed saprotrophs have been amplified from plant root interiors, and several saprotrophic genera can invade and interact with host plants in laboratory growth experiments. However, it is uncertain if root invasion by saprotrophic fungi is a widespread phenomenon and if laboratory interactions mirror field conditions. Here, we focused on the widespread and speciose saprotrophic genus Mycena and performed (1) a systematic survey of their occurrences (in ITS1/ITS2 datasets) in mycorrhizal roots of 10 plant species, and (2) an analysis of natural abundances of 13 C/15 N stable isotope signatures of Mycena basidiocarps from five field locations to examine their trophic status. We found that Mycena was the only saprotrophic genus consistently found in 9 out of 10 plant host roots, with no indication that the host roots were senescent or otherwise vulnerable. Furthermore, Mycena basidiocarps displayed isotopic signatures consistent with published 13 C/15 N profiles of both saprotrophic and mutualistic lifestyles, supporting earlier laboratory-based studies. We argue that Mycena are widespread latent invaders of healthy plant roots and that Mycena species may form a spectrum of interactions besides saprotrophy also in the field.

In vitro evidence of root colonization suggests ecological versatility in the genus Mycena.

The root-associated habit has evolved on numerous occasions in different fungal lineages, suggesting a strong evolutionary pressure for saprotrophic fungi to switch to symbiotic associations with plants. Species within the ubiquitous, saprotrophic genus Mycena are frequently major components in molecular studies of root-associated fungal communities, suggesting that an evaluation of their trophic status is warranted. Here, we report on interactions between a range of Mycena species and the plant Betula pendula. In all, 17 Mycena species were inoculated onto B. pendula seedlings. Physical interactions between hyphae and fine roots were examined using differential staining and fluorescence microscopy. Physiological interactions were investigated using 14 C and 32 P to show potential transfer between symbionts. All Mycena species associated closely with fine roots, showing hyphal penetration into the roots, which in some cases were intracellular. Seven species formed mantle-like structures around root tips, but none formed a Hartig net. Mycena pura and Mycena galopus both enhanced seedling growth, with M. pura showing significant transfer of 32 P to the seedlings. Our results support the view that several Mycena species can associate closely with plant roots and some may potentially occupy a transitional state between saprotrophy and biotrophy.

The bacterial microbiota in first-void urine from men with and without idiopathic urethritis.

BACKGROUND: Non-gonococcal urethritis (NGU) is a common syndrome in men. NGU may have several causes, but many cases are caused by sexually transmitted infections that may also cause complications in their female partners. Chlamydia trachomatis and Mycoplasma genitalium are the most common causes of NGU, but in up to 35% of the cases, none of the known viral or bacterial causes are found. Traditionally, pathogens have been detected using various culture techniques that may not identify all species present in the urethra. To address this, we used culture-independent methods for analysis of the male urethral microbiota. METHODS: This case-control study analysed first void urine samples, collected at STD clinics in Stockholm, Sweden from men with idiopathic urethritis (IU), i.e. negative for Neisseria gonorrhoeae, Chlamydia trachomatis, Mycoplasma genitalium, Ureaplasma urealyticum, Trichomonas vaginalis, adenovirus, and herpes simplex virus type 1 and -2 together with samples from men without urethritis. Forty-six controls and 39 idiopathic urethritis patients were analysed. RESULTS: The microbiota was highly diverse: None of the 302 operational taxonomic units (OTUs) found in negative controls and IU patients were found in all of the samples or even in all of the samples in one group. More than 50% of the OTUs were only found in one or two of the total of 85 samples. Still the most dominant 1/6 of the genera constituted 79% of the sequences. Hierarchical clustering in a heatmap showed no specific clustering of patients or controls. A number of IU patient samples were dominated by a single genus previously related to urethritis (Gardnerella, Haemophilus, Ureaplasma). CONCLUSION: The male urethra contain a very diverse composition of bacteria, even in healthy controls. NGU may be caused by a number of different bacteria but more studies including a higher number of samples are needed for elucidation of the role of each species.

Towards diagnostic metagenomics of Campylobacter in fecal samples.

BACKGROUND: The development of diagnostic metagenomics is driven by the need for universal, culture-independent methods for detection and characterization of pathogens to substitute the time-consuming, organism-specific, and often culture-based laboratory procedures for epidemiological source-tracing. Some of the challenges in diagnostic metagenomics are, that it requires a great next-generation sequencing depth and unautomated data analysis. RESULTS: DNA from human fecal samples spiked with 7.75 × 101-7.75 × 107 colony forming unit (CFU)/ml Campylobacter jejuni and chicken fecal samples spiked with 1 × 102-1 × 106 CFU/g Campylobacter jejuni was sequenced and data analysis was done by the metagenomic tools Kraken and CLARK. More hits were obtained at higher spiking levels, however with no significant linear correlations (human samples p = 0.12, chicken samples p = 0.10). Therefore, no definite detection limit could be determined, but the lowest spiking levels found positive were 7.75 × 104 CFU/ml in human feces and 103 CFU/g in chicken feces. Eight human clinical fecal samples with estimated Campylobacter infection loads from 9.2 × 104-1.0 × 109 CFU/ml were analyzed using the same methods. It was possible to detect Campylobacter in all the clinical samples. CONCLUSIONS: Sensitivity in diagnostic metagenomics is improving and has reached a clinically relevant level. There are still challenges to overcome before real-time diagnostic metagenomics can replace quantitative polymerase chain reaction (qPCR) or culture-based surveillance and diagnostics, but it is a promising new technology.

Local diversity of heathland Cercozoa explored by in-depth sequencing.

Cercozoa are abundant free-living soil protozoa and quantitatively important in soil food webs; yet, targeted high-throughput sequencing (HTS) has not yet been applied to this group. Here we describe the development of a targeted assay to explore Cercozoa using HTS, and we apply this assay to measure Cercozoan community response to drought in a Danish climate manipulation experiment (two sites exposed to artificial drought, two unexposed). Based on a comparison of the hypervariable regions of the 18S ribosomal DNA of 193 named Cercozoa, we concluded that the V4 region is the most suitable for group-specific diversity analysis. We then designed a set of highly specific primers (encompassing ~270 bp) for 454 sequencing. The primers captured all major cercozoan groups; and >95% of the obtained sequences were from Cercozoa. From 443 350 high-quality short reads (>300 bp), we recovered 1585 operational taxonomic units defined by >95% V4 sequence similarity. Taxonomic annotation by phylogeny enabled us to assign >95% of our reads to order level and ~85% to genus level despite the presence of a large, hitherto unknown diversity. Over 40% of the annotated sequences were assigned to Glissomonad genera, whereas the most common individually named genus was the euglyphid Trinema. Cercozoan diversity was largely resilient to drought, although we observed a community composition shift towards fewer testate amoebae.

Aminobacter MSH1-Mineralisation of BAM in Sand-Filters Depends on Biological Diversity.

BAM (2,6-dichlorobenzamide) is a metabolite of the pesticide dichlobenil. Naturally occurring bacteria that can utilize BAM are rare. Often the compound cannot be degraded before it reaches the groundwater and therefore it poses a serious threat to drinking water supplies. The bacterial strain Aminobacter MSH1 is a BAM degrader and therefore a potential candidate to be amended to sand filters in waterworks to remediate BAM polluted drinking water. A common problem in bioremediation is that bacteria artificially introduced into new diverse environments often thrive poorly, which is even more unfortunate because biologically diverse environments may ensure a more complete decomposition. To test the bioaugmentative potential of MSH1, we used a serial dilution approach to construct microcosms with different biological diversity. Subsequently, we amended Aminobacter MSH1 to the microcosms in two final concentrations; i.e. 10(5) cells mL(-1) and 10(7) cells mL(-1). We anticipated that BAM degradation would be most efficient at "intermediate diversities" as low diversity would counteract decomposition because of incomplete decomposition of metabolites and high diversity would be detrimental because of eradication of Aminobacter MSH1. This hypothesis was only confirmed when Aminobacter MSH1 was amended in concentrations of 10(5) cells mL(-1). Our findings suggest that Aminobacter MSH1 is a very promising bioremediator at several diversity levels.

Ultrastructure and phylogenetic position of Regin rotiferus and Otto terricolus genera et species novae (Bicosoecida, Heterokonta/Stramenopiles).

We describe two novel flagellates isolated from soil, Regin rotiferus and Otto terricolus, genera et species novae, which we cultivated and characterized by light and transmission electron microscopy and by 18S rDNA sequence analysis. Both strains exhibit the key characteristic structural feature of Bicosoecida; i.e. the L-shaped cytostomal root system with an x-fiber, used for feeding. Otto terricolus displays unique novel morphological traits; thus, it has a basal swelling on each flagellum, a root 3/root 2 distribution of 10 + 1 microtubules, and an amoeboid stage in its life cycle. Regin rotiferus has flagella without swellings and a root 3/root 2 distribution of 7 + 3 microtubules, a pattern commonly observed in the Bicosoecida. We present an updated exhaustive maximum likelihood phylogeny of 48 cultured, complete or nearly complete (+1600 bp) 18S rDNA Bicosoecida sequences. Both new species fall into a well-supported freshwater Siluaniidae clade, without being particularly closely related. The morphology and phylogeny do not conclusively support Rictus as a member of Bicosoecida.