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Ectomycorrhizal fungi in the genus Tuber form hypogeous fruiting bodies called truffles. Many Tuber species are highly prized due to their edible and aromatic ascomata. Historically, there has been attention on cultivating and selling European truffle species, but there is growing interest in cultivating, wild-harvesting, and selling species of truffles endemic to North America. North America has many endemic Tuber species that remain undescribed, including some that have favorable culinary qualities. Here, we describe two such Tuber species from eastern North America. Maximum likelihood and Bayesian phylogenetic analyses of ITS (internal transcribed spacer), tef1 (translation elongation factor 1-alpha), and rpb2 (second largest subunit of RNA polymerase II) sequences were used to place these species within a phylogenetic context. We coupled these data with morphological analyses and volatile analyses based on gas chromatography-mass spectrometry. Tuber cumberlandense , sp. nov. (previously referred to as Tuber sp. 66), is a member of the Rufum clade that has been opportunistically harvested for commercial sale from T. melanosporum orchards across eastern North America. Tuber canirevelatum , sp. nov. belongs in the Macrosporum clade and thus far is only known from eastern Tennessee, USA. Both new species were discovered with the assistance of trained truffle dogs. The volatile profiles of T. canirevelatum and T. cumberlandense were measured in order to characterize aromas based on the chemical compounds produced by these fungi. Ascomata from both species were enriched in acetone, dimethyl sulfide, 1-(methylthio)-1-propene, and 1-(methylthio)propane. In this work, we celebrate and encourage the use of trained truffle-hunting dogs for fungal biodiversity discovery and research.

Species of genus Morchella are high-value edible mushrooms. They are sought after by culinary experts due to their aroma, flavor, meaty texture, and health benefits. M. rufobrunnea, M. sextelata, and M. americana were chosen in this study and investigated for their medicinal quality by using in vitro anti-inflammatory and antioxidant assays. This sampling represents conditions by which morels are produced (cultivated indoors, cultivated outdoors, and collected from natural habitats, respectively) for commercial markets. Both aqueous and methanolic extracts of all three morel species showed identical chromatographic and bioassay profiles, independent of their phylogenetic position or production method. In an antioxidant assay, aqueous and methanolic extracts of these mushrooms at 100 μg/mL inhibited lipid peroxidation (LPO) by 59%-62% and 33%-36%, respectively. In an anti-inflammatory assay using cyclooxygenase enzymes (COX-1 and COX-2), aqueous and methanolic extracts at 100 μg/mL showed COX-1 enzyme inhibition by 53%-57% and 30%-32% and COX-2 enzyme inhibition by 38%-44% and 16%-17%, respectively. Chromatographic purification and spectroscopic characterization of M. rufobrunnea extracts afforded five sugars (compounds 1-5), seven organic acids (compounds 6-13), three flavonoids (compounds 14-16), triglycerides, free fatty acids, and three sterols (compounds 17-19). This is the first report of COX-1 and COX-2 enzymes and LPO inhibitory activities of pure isolates (S)-morelid (compound 6), glutamic acid (compound 9), and brassicasterol (compound 19). This study also showed inhibitions of COX-1 (by 84%, 33%, and 37%), COX-2 (by 47%, 11%, and 22%), and LPO (by 74%, 48%, and 35%), respectively, at 25 μg/mL.

Soil archives are an important resource in agronomic and ecosystem sciences. If microbial communities could be reconstructed from archived soil DNA, as prehistoric plant communities are reconstructed via pollen data, soil archive resources would assume even greater value for reconstructing land-use history, forensic science, and biosphere modelling. Yet, the effects of long-term soil archival on the preservation of microbial DNA is still largely unknown. To address this, we assessed the capacity of high-throughput sequencing (Illumina MiSeq) of ITS (internal transcribed spacer) and prokaryotic 16S rRNA genes for reconstructing soil microbial communities across a 20 years time-series. We studied air-dried soil archives and fresh soil samples taken from Populus bioenergy and deciduous forest research plots at the Kellogg Biological Station. Habitat and archival time explained significant amounts of variation in soil microbial α- and β-diversity both in fungal and prokaryotic communities. We found that microbial richness, diversity, and abundance generally decreased with storage time, but varied between habitat and taxonomic groups. The high relative abundance of ectomycorrhizal species including Hebeloma and Cortinarius detected in older soil archives raises questions regarding traits such as long-term persistence and viability of ectomycorrhizal propagules in soils, with relevance to forest health and ecosystem succession. Talaromyces, Paecilomyces and Epicoccum spp. were detected in fresh and across 20-year-old archived soils and were also cultured from these soils demonstrating their long-term spore viability. In summary, we found that microbial DNA in air-dried soils archived over the past 20 years degraded with time, in a manner that differed between soil types and phylogenetic groups of microbes., Competing Interests: The authors have declared that no competing interests exist.

1. [3H]-triethylcholine (TEC) was actively transported by the renal tubule of the chicken at a rate 85% that of simultaneously administered p-aminohippuric acid (PAH). 2. TEC was demonstrated to be transported by the organic cation transport system in the kidney through inhibition with quinine and the bio-cation choline. 3. When the infusion of TEC was increased to 2 times 10(-6) mol kg(-1) min(-1) reaching the infused kidney, the transport of [3H]-TEC was inhibited, suggesting that an excretory transport maximum for TEC in the renal tubules had been reached. 4. The excretion of both choline and acetylcholine was enhanced by TEC loads as low as 1 times 10(-18) mol kg(-1) min(-1). Enhancement continued as TEC infusion was increased up to approximately 1 times 10(-7) mol kg(-1) min(-1) at which point this enhancement was converted to inhibition. 5. Possible mechanisms for the biphasic effect of TEC on organic cation transport are discussed.

Endogenous concentrations of free choline in plasma were measured in azotemic subjects receiving repetitive hemodialysis and excretion of free choline into the dialysate was determined. Chemical choline in plasma and dialysate was measured by adding choline kinase and measuring the production of radiolabelled phosphorycholine in the presence of radiolabelled adenosine triphosphate (ATP). Mean free choline concentration in plasma of azotemic subjects receiving hemodialysis was found to be 37 muM, which is about twice that of normal persons. The total excretion of choline into the dialysate during 360 min averaged 730 mumoles +/- 69 (SEM). Levels of free choline in plasma fell during hemodialysis at two hours but recovered toward predialysis values at six hours. The return of plasma choline concentrations toward control values during dialysis suggests that a feedback mechanism exists which was activated rapidly to produce homeostasis of plasma choline concentrations. In these patients, the degree of peripheral neuropathy as judged by measurement of nerve conduction velocities showed a significant inverse correlation with levels of free choline in plasma.

In vitro and in vivo renal tubular transport of 2,4,5-trichlorophenoxyacetate (2,4,5-T), a hormone-type herbicide, was studied in the chicken. Renal cortical slices incubated with 14C-labeled 2,4,5-T demonstrated a slice-to-medium ratio of the 14C-label of 26 after 2 hr of incubation. This accumulation was inhibited significantly by probenecid, phenol red, dinitrophenol and iodoacetamide. In vitro metabolism of 2,4,5-T was apparent in the renal slice studies; at least two metabolites were found by electrophoretic analysis. Even though chicken liver slices were able to accumulate 2,4,5-T, the accumulated herbicide was not metabolized. In addition, chicken kidney slices metabolized 2,4,5-T to a much greater extent than did rat or rabbit kidney slices, which produced only negligible amounts of metabolites. When [14C]-2,4,5-T was infused into the renal portal system of the chicken, the 14C-label originally associated with 2,4,5-T was shown to be secreted by the renal tubule. Probenecid inhibited the transport of the 14C-label without affecting the simultaneous transport of tetra-ethylammonium, an organic cation transported by the renal proximal tubule. At the present infusion rate of 2,4,5-T (2.9 X 10(-10) mol/min), the excreted label was associated primarily with an acidic metabolite(s). It can be concluded that the chicken transports 2,4,5-T by a probenecid sensitive mechanism; however, this transport appears to be associated with the metabolism of the herbicide.

The Sperber technique of infusion into the renal portal circulation in chickens was used to investigate in vivo the renal tubular transport and renal metabolism of trimethylamine (TMA). When 14C-TMA was infused at a rate of 1 x 10(-9) mol/min the transport efficiency (TE), that is, the tubular excretion of the 14C-label relative to excretion of simultaneously infused paminohippuric acid, was 0.70. Progressive addition of unlabeled TMA up to infusion rates of 1 x 10(-5) mol/min produced a progressive fall in the TE of the 14C-label. Identification of the 14C-label excreted in the urine revealed that approximately 85% of the infused 14C-TMA was excreted by the infused kidney as a single metabolite over the entire range of infusions. By use of the techniques of low-voltage electrophoresis, high-voltage electrophoretic mobility-pH profile, and gas chromatography/mass spectrometry, the renal metabolite was found to be identical with standard 14C-trimethylamine oxide (TMAO). At a TMA infusion rate of 1.5 x 10(-6) mol/kg/min reaching the infused kidney, the rate at which TMAO was formed and excreted by the kidney was 0.12 x 10(-6) mol per g of kidney per min. When 14C-TMAO was infused into chickens its TE was 0.11, which was not evidence for active excretory transport. Infused TMA was almost entirely metabolized in vivo to its N-oxide, TMAO, which then entered the urine. The renal tubular excretion of 14C during infusion of 14C-TMA was inhibited by the cationic blocker of transport, quinine, and by the anionic blocker of transport, probenecid.

Plasma choline levels were measured in patients who received a kidney transplant, in donors who underwent nephrectomy and in nonrenal surgical patients. Choline was measured using a choline kinase assay. Choline levels in patients receiving a kidney fell from 29.8 +/- 1.86 microM before transplantation to 15.7 +/- 2.32 1 day later; this normal level was maintained for at least 7 months and in a single case for 2 years. Kidney donors and nonrenal surgery patients showed a significant decrease in plasma choline on the day following surgery but choline levels returned to normal by 3 days after surgery. Thus a transplanted functional kidney reduced the high plasma choline levels, associated with uremia, to normal and maintained these normal levels throughout the period of observation.