Structural characterization and protective effect on PC12 cells against H2O2-induced oxidative damage of a polysaccharide extracted from mycelia of Lactarius deliciosus Gray.

The crude polysaccharide LDP was extracted from mycelia of Lactarius deliciosus Gray and then purified by DEAE-52 cellulose and Sephadex G-200 to obtain a novel polysaccharide named LDP-CP. LDP-CP was mainly composed of mannose, glucose and galactose with an average molecular weight of 2.33 × 103 kDa. The structure of LDP-CP was determined by FT-IR, methylation and NMR analysis, and the results showed that the sugar linkage units of LDP-CP were composed of (1 â†’ 3)-linked ß-D-Manp, (1 â†’ 2,4)-linked α-D-Manp, (1→)-linked α-D-Manp, (1 â†’ 4)-linked ß-D-Glcp, (1 â†’ 2)-linked ß-D-Manp, (1 â†’ 4,6)-linked α-D-Manp, (1 â†’ 4)-linked α-D-Galp, (1 â†’ 2,3)-linked α-D-Glcp and (1→)-linked α-D-Glcf. The protective effects of LDP and LDP-CP on PC12 cells against H2O2-induced oxidative injury were exhibited by enhancing cell viability and morphological protection. The improvement to the level of LDH, SOD and GSH further indicated that LDP and LDP-CP had ability to alleviate H2O2-induced oxidative damage on PC12 cells. The polysaccharides in Lactarius deliciosus Gray mycelia exhibited the great advantages in the management of oxidative toxicity, which indicated that the polysaccharides can be further developed in application of natural functional food source.

Evaluación del crecimiento radial de Lactarius volemus en medio semisólido / Evaluation of radial growth of Lactarius volemus in semisolid medium

Objetivo: Evaluar el crecimiento radial de Lactarius volemus en cinco medios de cultivo semisólidos in vitro.Materiales y métodos: Cuerpos fructíferos de L. volemus provenientes de la Sierra Norte del estado de Oaxaca, México, se cultivaron en laboratorio en medios Agar Papa Dextrosa, Agar Czapek-Dox, Agar Extracto de Malta, Agar Papa Sacarosa y Agar Dextrosa Saboraud; mediante dos técnicas de sembrado. Se evaluaron las características morfológicas de colonias obtenidas de distintas muestras del cuerpo fructífero, así como el crecimiento radial de cada una.Resultados: El crecimiento colonial evaluado permitió seleccionar un medio que reúne las condiciones óptimas para el cultivo de Lactarius volemus in vitro. No todas las muestras utilizadas desarrollan un crecimiento abundante: la muestra proveniente del látex presenta un crecimiento escaso.Conclusiones: Con la evaluación del crecimiento radial de Lactarius volemus se obtiene una referencia directa del ciclo de crecimiento de esta especie; es posible identificar las fases exponencial y estacionaria pero las condiciones del medio no permiten evaluar la fase de muerte debido a la deshidratación y reducción del agar.(AU)

Objective: To evaluate the radial growth of Lactarius volemus in five semi-solid culture media in vitro.Materials and methods: Fruitful bodies of L. volemus from the Sierra Norte of the state of Oaxaca, Mexico, were cultured in the laboratory in Potato Dextrose Agar Papa, Czapek-Dox Agar, Malt Extract Agar, Potato Sucrose Agar and Dextrose Saboraud Agar; using two seeding techniques. The morphological characteristics of colonies obtained from different samples of the fruiting body were evaluated, as well as the radial growth of each one.Results: The evaluated colonial growth allowed to select a culture medium that meets the optimal conditions for the cultivation of Lactarius volemus in vitro. Not all samples used develop abundant growth: the sample from latex shows little growth.Conclusions: With the evaluation of the radial growth of Lactarius volemus a direct reference to the growth cycle of this species is obtained; it is possible to identify the exponential and stationary phases but the conditions of the medium do not allow evaluating the phase of death due to dehydration and reduction of the agar.(AU)

Successes and challenges in the sustainable cultivation of edible mycorrhizal fungi - furthering the dream.

The cultivation of edible mycorrhizal fungi (EMF) has made great progress since the first cultivation of Tuber melanosporum in 1977 but remains in its infancy. Five cultivation steps are required: (1) mycorrhizal synthesis, (2) mycorrhiza development and acclimation, (3) out-planting of mycorrhizal seedlings, (4) onset of fructification, and (5) performing tree orchards. We provide examples of successes and challenges associated with each step, including fruiting of the prestigious chanterelles in Japan recently. We highlight the challenges in establishing performing tree orchards. We report on the monitoring of two orchards established between Lactarius deliciosus (saffron milk cap) and pines in New Zealand. Saffron milk caps yields reached 0.4 and 1100 kg/ha under Pinus radiata and P. sylvestris 6 and 9 y after planting, respectively. Canopy closure began under P. radiata 7 y after planting, followed by a drastic reduction of yields, while P. sylvestris yields still hovered at 690 to 780 kg/ha after 11 y, without canopy closure. The establishment of full-scale field trials to predict yields is crucial to making the cultivation of EMF a reality in tomorrow's cropping landscape. Sustainable EMF cultivation utilizing trees in non-forested land could contribute to carbon storage, while providing revenue and other ecosystem services.

Two new guaiane sesquiterpenes from the fruiting bodies of Lactarius Deliciosus.

Two new guaiane sesquiterpenes, 7-(1-hydroxyethyl)-4-methyl-1-azulenecarboxaldehyde (1) and 7-isopropenyl-4-methyl-1-azulenecarboxylic acid (2), together with 5 known sesquiterpenes, were isolated from the fruiting bodies of Lactarius deliciosus. All structures were elucidated based on extensive spectroscopic methods, including 1 D and 2 D-NMR spectroscopy and high resolution mass spectrometry.

Elemental bioaccumulation and nutritional value of five species of wild growing mushrooms from South Africa.

The collection of wild edible mushrooms for use is an ancient practice. In this study, the elemental concentrations in five mushrooms were compared as a function of species and geographical location. The accumulation of metals from the substrate was found to be species dependent. Mushrooms excluded As, Be, Ca, Cd, Co, Cr, Fe, Mn, Ni and Si, and accumulated elements in the following order: Amanita pantherina (K > Na > Zn > Cu > Mg); Boletus edulis (edible) (K > Cu > Zn > Se > Na > Mg); Boletus mirabilis (edible) (K > Cu > Zn > Na > Mg); Lactarius deliciosus (edible) (K > Zn > Mg); and Russula sardonia (K > Na > Zn > Cu > Mg). Statistical analysis showed soil parameters to affect elemental accumulation by edible mushrooms more than inedible ones, especially for uptake of Se by B. edulis, which was antagonistic with As and Pb in the soil. The results also showed the edible mushrooms to contain proteins (25 - 55%), carbohydrates (34 - 69%), ash (3-6.5%) and lipids (0.8-5.3%).

Chemical Composition, Antioxidant and Antihyperglycemic Activities of the Wild Lactarius deliciosus from China.

The wild mushroom Lactarius deliciosus from China was studied for the first time to obtain information about its chemical composition, antioxidant, and antihyperglycemic activities. Nutritional value, dietary fiber, fatty acids, metal elements, free sugars, free amino acids, organic acids, flavor 5'-nucleotides, and volatile aroma compounds were determined. Potential antioxidant and antihyperglycemic activities were also tested by investigating 1,1-diphenyl-2-picrylhydrazyl (DPPH) and 2,2'-Azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) radicals scavenging activities, ferric ion reducing activity, as well as α-amylase and α-glucosidase inhibitory activities using ethanol and aqueous extracts. The results showed that L. deliciosus was a good wild mushroom with high protein, carbohydrate, and dietary fiber contents, while low in fat and calorie, extensive unsaturated fatty acids contents, with negligible health risks about harmful metal elements. Twenty kinds of free amino acids were detected with a total content 3389.45 mg per 100 g dw. Flavor 5'-nucleotides including 5'-CMP, 5'-UMP, 5'-IMP, and 5'-AMP were 929.85, 45.21, 311.75, and 14.49 mg per 100 g dw, respectively. Mannitol (7825.00 mg per 100 g dw) was the main free sugar, and quininic acid (729.84 mg per 100 g dw) was the main organic acid. Twenty-five kinds of volatile aroma compounds were identified, acids (84.23%) were the most abundant compounds based on content, while aldehydes (15 of 25) were the most abundant compounds based on variety. In addition, both ethanol and aqueous extracts from L. deliciosus exhibited excellent antioxidant activity. While in antihyperglycemic activity tests, only ethanol extracts showed inhibitory effects on α-amylase and α-glucosidase.

Effect on macrophage proliferation of a novel polysaccharide from Lactarius deliciosus (L. ex Fr.) Gray.

The fundamental mechanisms underlying the preventional and therapeutic effects of polysaccharides from fungi, including the immunostimulatory, antiviral and antitumor effects, are considered to occur through the modulation and stimulation of the macrophage and complement system. LDG-A, a novel polysaccharide from Lactarius deliciosus (L. ex Fr.) Gray exhibits marked antitumor activities in vivo. However, the underlying molecular mechanism of the antitumor activities of LDG-A remains unclear. In the present study, cell cycle analysis was performed in macrophages and B cells, and the transcriptomes of macrophages in the control group and LDG-A group were sequenced using Illumina sequencing technology to analyze the differentially expressed genes (DEGs), and elucidate the molecular mechanisms underlying the immunomodulatory and antitumor activities of LDG-A. The cell cycle analysis results indicated that LDG-A was able to promote the proliferation of B cells by promoting cell cycle progression in S phase and G2/M phase and eliminating cell cycle arrest in G0/G1, and promote the proliferation of macrophages by promoting cell cycle progression in G0/G1 phase and eliminating cell cycle arrest in G2/M phase. Of the total number of genes (8,140), ~77.00% were expressed [reads per kilobase per million reads (RPKM) ≥1] and 1,352 genes were highly expressed (RPKM >60) in the LDG-A group. Of 775 unigenes which were identified as DEGs, 469 were downregulated and 306 genes were upregulated. A protein chip method was also used to determine the cytokines secreted by macrophages. Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis and GO enrichment analysis indicated that the Janus kinase/signal transducer and activator of transcription, mitogen-activated protein kinase, chemokine, vascular endothelial growth factor and transforming growth factor ß signaling pathways are markedly enriched for DEGs.

Immunomodulatory effect of a polysaccharide fraction on RAW 264.7 macrophages extracted from the wild Lactarius deliciosus.

The mushroom polysaccharides are important substances with variety of functions, especially to the human body's immunomodulation effects. In this work, a polysaccharide fraction (LDP-1) was extracted and purified from the fruiting bodies of a rare wild Lactarius deliciosus. LDP-1 with molecular weight of 9.8 × 105 Da showed an obvious immunological activity to the RAW 264.7 cells. It had no significant suppressive but promotive effects on proliferation of the macrophages. The production of nitric oxide (NO) presented a concentration-dependent manner after treated with the LDP-1, and the maximum yield of NO was 39.15 µM. LDP-1 could promote the phagocytic uptake ability of the RAW 264.7 cells significantly, and many of the antennas produced around the cells correspondingly. The cytokines of TNF-α, IL-1ß and IL-6 were secreted increasingly in a concentration-dependent manner, which were 4.83, 17.8 and 11 times than that of the control, respectively. Western blotting analysis confirmed that NF-κB levels in the nucleus were increased while cytoplasmic inhibitor of NF-κB (IκB-α) degraded after treated with the LDP-1, indicating the RAW 264.7 cells probably be stimulated by LDP-1 through activating the IκB-α-NF-κB pathway. These results demonstrated that LDP-1 could be used as a kind of immunomodulatory agent for healthcare potentially.

Trace element and toxic metal intake from the consumption of canned mushrooms marketed in Spain.

Mushrooms are a valuable foodstuff because they are an important source of fiber and essential amino acids, but they have a great capacity to capture metals from substrate. Due to this reason, the content of 14 metals (B, Ba, Cr, Cu, Fe, Li, Mo, Ni, Sr, V, Zn, Al, Cd, Pb) of 100 samples of canned mushrooms grown in Spain were analyzed by inductively coupled plasma atomic emission spectroscopy. Fe (9.55 mg/kg) was the most notable trace element, followed by Zn (2.39 mg/kg). The most abundant toxic metal was Al (17.80 mg/kg). Significant statistical differences between the absorption of metals have been observed when comparing species. Lentinula edodes and Lactarius deliciosus are the species which had the highest metal contents. Taking the average consumption 300 g per day for adults into account, the exposure to trace toxic metals from the consumption of the studied mushrooms does not pose a health risk. However, contribution percentages of Pb (87.6%) and Al (59.2%) found in Lentinula edodes and Pholiota nameko samples, respectively, are near the maximum intake.

New Azulene-Type Sesquiterpenoids from the Fruiting Bodies of Lactarius deliciosus.

In the 1H NMR-guided fractionation of extracts from the edible mushroom Lactarius deliciosus, two new azulene-type sesquiterpenoids, 7-isopropenyl-4-methyl-azulene-1-carboxylic acid (1) and 15-hydroxy-3,6-dihydrolactarazulene (2), together with seven known compounds were characterized. Their structures were determined on basis of spectroscopic evidence, as well as by comparing with literature data. Amongst the known metabolites, the 13C NMR assignment of 15-hydroxy-6,7-dihydrolactarazulene (3) is reported here for the first time. Moreover, 7-acetyl-4-methylazulene-1-carbaldehyde (5) displayed a moderate antibacterial activity against Staphylococcus aureus. *Digital image of L. deliciosus. Retrieved March 17, 2017 from https://upload.wikimedia.org/wikipedia/commons/e/e3/Lactarius_deliciosus_1_(1).jpg .

Mercury accumulation of three Lactarius mushroom species.

Accumulation, distribution and potential dietary intake of mercury accumulated by mushrooms of Lactarius species L. delicious, L. volemus and L. deterrimus were studied in the Pomerania region of Poland. In total, 212 fruiting bodies and 106 underlying topsoil samples were analyzed. Analysis indicated that the concentrations of Hg were at low levels both in mushrooms and forest topsoils for a majority of the locations investigated. L. volemus that grew in soils with only a slightly elevated contamination (0.11±0.07mgkg(-1) of dried soil), very efficiently accumulated Hg in fruiting bodies and concentration levels were at 3.7±1.3mgkg(-1) of dry biomass in caps and at 1.9±0.9mgkg(-1) of dry biomass in stipes. Consumption of mushrooms foraged from the Sobowidz forest, which is close to a foundry using ferrous and non-ferrous metals could result in a Hg intake that exceeds the current statutory limits.

Structure elucidation and anti-tumor activities of water-soluble oligosaccharides from Lactarius deliciosus (L. ex Fr.) Gray.

BACKGROUND: Oligosaccharides are composed of a variable number of monosaccharide units and very important in the biologically diverse of biological systems. MATERIALS AND METHODS: Crude water-soluble oligosaccharide was extracted from the fruiting bodies with water and then successively purified by DEAE-cellulose 52 and Sephadex G-100 column chromatography, yielding one major oligosaccharides fractions: LES-A. Structural features of Lactarius deliciosus (L. ex Fr.) Gray oligosaccharide (LDGO-A) were investigated by a combination of monosaccharide component analysis by thin layer chromatography, infrared spectra, nuclear magnetic resonance spectroscopy, scanning electron microscopy, and high-performance gel permeation chromatography analysis. RESULT: The results indicated that LDGO-A was composed of D-glucose and D-xylose, and the average molecular sizes was approximately 945 Da. The anti-tumor activity of LDGO-A was evaluated in vivo. The inhibitory rate in mice treated with 40 mg/kg LDGO-A can reach 40.02%, being the highest in the three doses, which may be comparable to mannatide. Histology of immune organs shows that the tissues arranged more regular and firmer, but the tumor tissue arranged looser in LDGO-A group than those in the control group. Meanwhile, there is no obvious damage to other organs, such as heart. The anti-tumor activity of the LDGO-A was usually believed to be a consequence of the stimulation of the cell-mediated immune response because it can significantly promote the lymphocyte and macrophage cells in the dose range of 100-400 µg/mL in vitro. LDGO-A also effected the expression of some housekeeping genes mRNA in S180 tumor. CONCLUSION: Accordingly, the LDGO-A might serve as an effective healthcare food and source of natural anti-tumor compounds.

Immunostimulant Activity of a Novel Polysaccharide Isolated from Lactarius deliciosus (L. ex Fr.) Gray.

More and more fungal polysaccharides have been reported to exhibit a variety of biological activities, including antitumor, antioxidant and immunostimulant activity. The non-starch polysaccharides have emerged as an important class of bioactive natural products. In this study, the immune activities of a novel polysaccharide (LDG-A) isolated from Lactarius deliciosus (L. ex Fr.) Gray were investigated at 20, 40 and 80 mg/kg dose levels. The inhibitory rate in mice treated with 80 mg/kg LDG-A can reach 68.422%, being the highest in the three doses, which may be comparable to mannatide. Histology of immune organs showed that the tissues were arranged in more regular and firm pattern, but the tumor tissue arranged looser in LDG-A group than those in control group. Meanwhile, there was no obvious damage to other organs, such as heart, lung, and kidney. The antitumor activity of the LDG-A was usually believed to be a consequence of the stimulation of the cell-mediated immune response because it can significantly promote the lymphocyte and macrophage cells in the dose range of 50-200 µg/ml and 100-400 µg/ml in vitro, respectively. The level of cytokines (IL-6, TNF-α, and NO) of macrophage cells induced by LDG-A treatment was similar to lipopolysaccharides at different concentrations. The expression of all these genes studied (TNF-α, IL-6, and iNOS mRNA) in the untreated macrophage was little, but increased dramatically in a dose-dependent manner in the LDG-A-treated cells. The results obtained in the present study indicated that the purified polysaccharide of L. deliciosus (L. ex Fr.) Gray is a potential source of natural immune-stimulating substances.

Molecular approach to characterize ectomycorrhizae fungi from Mediterranean pine stands in Portugal.

Stone pine (Pinus pinea L.), like other conifers, forms ectomycorrhizas (ECM), which have beneficial impact on plant growth in natural environments and forest ecosystems. An in vitro co-culture of stone pine microshoots with pure mycelia of isolated ECM sporocarps was used to overcome the root growth cessation not only in vitro but also to improve root development during acclimation phase. Pisolithus arhizus (Scop.) Rauschert and Lactarius deliciosus (L. ex Fr.) S.F. Gray fungi, were collected, pure cultured and used in in vitro co-culture with stone pine microshoots. Samples of P. arhizus and L. deliciosus for the in vitro co-cultures were collected from the pine stands southwest Portugal. The in situ characterization was based on their morphotypes. To confirm the identity of the collected material, ITS amplification was applied using the pure cultures derived from the sporocarps. Additionally, a molecular profile using PCR based genomic fingerprinting comparison was executed with other genera of Basidiomycetes and Ascomycetes. Our results showed the effectiveness of the techniques used to amplify DNA polymorphic sequences, which enhances the characterization of the genetic profile of ECM fungi and also provides an option to verify the fungus identity at any stage of plant mycorrhization.

Influence of the Culinary Treatment on the Quality of Lactarius deliciosus.

The influence of culinary treatment on the nutritional value and quality of Lactarius deliciosus was established. Mushrooms: unblanched (I), blanched (II), and unblanched with onion and spices (III), were fried in oil for 10 min. Fried mushrooms were assessed before storage as well as after 48 h in storage at 20 °C, and after 48 and 96 h in storage at 4 °C. Frying increased the dry weight, protein, fat, ash, total carbohydrate, total polyphenol, and total flavonoid content, as well as the caloric value of the mushrooms. In addition, frying decreased the antioxidant activity, color parameters (a*, h*), and texture. The most significant changes due to culinary treatment and storage were observed in type II product. Microbiological analysis of the samples after a 48 h storage period at 20 °C revealed the total viable count over 106 and contamination with lactic acid bacteria. Fried mushrooms stored at 4 °C for 96 h were free from microorganisms.

Molecular approach to characterize ectomycorrhizae fungi from Mediterranean pine stands in Portugal

Stone pine (Pinus pinea L.), like other conifers, forms ectomycorrhizas (ECM), which have beneficial impact on plant growth in natural environments and forest ecosystems. An in vitro co-culture of stone pine microshoots with pure mycelia of isolated ECM sporocarps was used to overcome the root growth cessation not only in vitro but also to improve root development during acclimation phase. Pisolithus arhizus (Scop.) Rauschert and Lactarius deliciosus (L. ex Fr.) S.F. Gray fungi, were collected, pure cultured and used in in vitro co-culture with stone pine microshoots. Samples of P. arhizus and L. deliciosus for the in vitro co-cultures were collected from the pine stands southwest Portugal. The in situ characterization was based on their morphotypes. To confirm the identity of the collected material, ITS amplification was applied using the pure cultures derived from the sporocarps. Additionally, a molecular profile using PCR based genomic fingerprinting comparison was executed with other genera of Basidiomycetes and Ascomycetes. Our results showed the effectiveness of the techniques used to amplify DNA polymorphic sequences, which enhances the ­characte­rization of the genetic profile of ECM fungi and also provides an option to verify the fungus identity at any stage of plant mycorrhization.

Molecular approach to characterize ectomycorrhizae fungi from Mediterranean pine stands in Portugal

Stone pine (Pinus pinea L.), like other conifers, forms ectomycorrhizas (ECM), which have beneficial impact on plant growth in natural environments and forest ecosystems. An in vitro co-culture of stone pine microshoots with pure mycelia of isolated ECM sporocarps was used to overcome the root growth cessation not only in vitro but also to improve root development during acclimation phase. Pisolithus arhizus (Scop.) Rauschert and Lactarius deliciosus (L. ex Fr.) S.F. Gray fungi, were collected, pure cultured and used in in vitro co-culture with stone pine microshoots. Samples of P. arhizus and L. deliciosus for the in vitro co-cultures were collected from the pine stands southwest Portugal. The in situ characterization was based on their morphotypes. To confirm the identity of the collected material, ITS amplification was applied using the pure cultures derived from the sporocarps. Additionally, a molecular profile using PCR based genomic fingerprinting comparison was executed with other genera of Basidiomycetes and Ascomycetes. Our results showed the effectiveness of the techniques used to amplify DNA polymorphic sequences, which enhances the ­characte­rization of the genetic profile of ECM fungi and also provides an option to verify the fungus identity at any stage of plant mycorrhization.

Molecular Identification of Isolate of Lactarius deliciosus by ITS Analysis / 微生物学通报

In this study,the rDNA ITS sequence of isolate of Lactarius deliciosus(L.)Gray was sequenced.Molecular identification was accomplished using phylogenetic tree analysis of 22 Lactarius nrDNA-ITS nucleotide sequence including the 2 new and 20 GenBank sequence.