Analysis of the Bioactive Metabolites of the Endangered Mexican Lost Fungi Campanophyllum - A Report from India.

Meghalaya, (in India), in the region of the mega-biodiversity hotspots, is home to a plethora of wild mushrooms. The present study concerns the exploration of the order Agaricales, which includes rare gilled mushrooms considered endangered under IUCN A4c criteria, due to the declining habitat. Electron microscopy of the gill sections revealed an abundance of clamp connections, hyphal cell walls, cystidia, and basidia. This rare species which belongs to the family Cyphellaceae, exhibits morphological and molecular differences from the Cyphella spp. Phylogenetic analysis revealed that it formed a clade under the genus Campanophyllum of the order Agaricales, confirmed by both Neighbor Joining (NJ) and Bayesian phylogenetic analysis. Being nutritionally potent along with its efficient antioxidant value, the fungal extract shows significant rise of two-fold in the antimicrobial activity along with the commercial antibiotics. The compound, Phenol, 2, 4-bis (1, 1-Dimethylethyl) (2, 4-DTBP) showed in ample range in the fungal extract along with aliphatic hydrocarbons, terpene, alcohol and volatile organic compounds on further characterization in GCMS. The present study indicates the endangered Campanophyllum proboscideum could be a rich source of natural antioxidants and an effective pharmaceutical agent.

Lentinula edodes based GIS mapping, biometabolites and antiinflamatory activity of wild edible mushrooms from tropical 'sacred grove' forests of Meghalaya, India / Mapas basados en GIS deLentinula edodes, biometabolitos y actividad antiinflamatoria de los hongos silvestres comestibles de los bosques tropicales "bosque sagrado" de Meghalaya, India

AbstractThe biodiversity rich state of Meghalaya, India located in the realms of mega-biodiversity hotspots, is home to numerous species of wild edible macrofungi that are used extensively by the mycophillic ethnic population, as a part of their traditional cuisine and medicine systems. However, habitat loss, due to deforestation and climate change, is destroying the natural population of these mushrooms, depleting their availability to the local communities. In the present investigation, a GIS guided habitat search, using Lentinula edodes as a representative species, was used in mapping the habitats of wild edible macrofungi of the study region. Sampling of around 4 000 specimens per distinct morphological type available in the traditional markets and "sacred grove" forests indicated presence of ten common genera, belonging to nine different families of wild edible mushrooms. Nutritional profiling of the representative species Lentinula edodes was carried out by evaluation of its moisture, total fat, crude protein and carbohydrates contents by standard methods. Similarly, bioactive components determination was performed by estimation of total phenols, flavonoids, ascorbic acid, β-carotene and lycopenes. Bioactivity of the mushrooms extracts was studied using the DPPH radical scavenging and Human Red Blood Cell (HRBC) membrane stabilization assays. The present investigation successfully attempted to explore remote sensing technologies and GIS (Geographic Information System) based system to predict the natural habitats of wild edible mushrooms of Meghalaya, India which we believe will lead to the generation of a mushroom specific non-wood forest resource mapping system in the near future. Results of nutritional profiling and biological activity studies on the representative species of wild edible mushrooms from the studied region revealed that it is a rich source of essential nutrients and antioxidants.

ResumenLa rica biodiversidad del estado de Meghalaya, India situado en los reinos de los hotspots mega-biodiversidad es el hogar de numerosas especies de macro hongos silvestres comestibles que se utilizan ampliamente por la población étnica micofílica como parte de su alimentación y medicina tradicional. Sin embargo, la pérdida de hábitat debido a la deforestación y el cambio climático está destruyendo la población natural de estos hongos, que agotan su disponibilidad para las poblaciones locales. En la presente investigación, se utilizó el SIG para la búsqueda guiada de hábitat usando Lentinula edodes como especie representativa en la cartografía de los hábitats de macrohongos silvestres comestibles de la región de estudio. Alrededor de cuatro mil ejemplares de distinto tipo morfológico fueron muestreados según su disponibilidad en los mercados tradicionales y los "sacred grove" (bosques sagrados) lo que indicó la presencia de diez géneros comunes pertenecientes a nueve familias de setas silvestres comestibles. Perfiles nutricionales de las especies representativas Lentinula edodes se llevaron a cabo mediante la evaluación de su humedad, grasa total, proteína cruda y contenido de carbohidratos. Del mismo modo, la determinación de componentes bioactivos se realizó por estimación de fenoles totales, flavonoides, ácido ascórbico, β-caroteno y licopeno. La bioactividad de los extractos de setas se estudió mediante la captación de radicales DPPH y el ensayo de estabilización de la membrana de glóbulos rojos de la sangre humana (hRBC). Los resultados de estudios de perfiles nutricionales y actividad biológica en hongos comestibles revelan que es una fuente rica en nutrientes esenciales y antioxidantes. La presente investigación ha intentado con éxito explorar las tecnologías de teledetección y SIG (Sistema de Información Geográfica) para predecir el hábitat natural de los hongos silvestres comestibles de Meghalaya, India que creemos dará lugar a la generación de un mapeo de los recursos forestales no madereros, específicamente de hongos, en el futuro cercano.

Lentinula edodes based GIS mapping, biometabolites and antiinflamatory activity of wild edible mushrooms from tropical 'sacred grove' forests of Meghalaya, India.

The biodiversity rich state of Meghalaya, India located in the realms of mega-biodiversity hotspots, is home to numerous species of wild edible macrofungi that are used extensively by the mycophillic ethnic population, as a part of their traditional cuisine and medicine systems. However, habitat loss, due to deforestation and climate change, is destroying the natural population of these mushrooms, depleting their availability to the local communities. In the present investigation, a GIS guided habitat search, using Lentinula edodes as a representative species, was used in mapping the habitats of wild edible macrofungi of the study region. Sampling of around 4 000 specimens per distinct morphological type available in the traditional markets and "sacred grove" forests indicated presence of ten common genera, belonging to nine different families of wild edible mushrooms. Nutritional profiling of the representative species Lentinula edodes was carried out by evaluation of its moisture, total fat, crude protein and carbohydrates contents by standard methods. Similarly, bioactive components determination was performed by estimation of total phenols, flavonoids, ascorbic acid, ß-carotene and lycopenes. Bioactivity of the mushrooms extracts was studied using the DPPH radical scavenging and Human Red Blood Cell (HRBC) membrane stabilization assays. The present investigation successfully attempted to explore remote sensing technologies and GIS (Geographic Information System) based system to predict the natural habitats of wild edible mushrooms of Meghalaya, India which we believe will lead to the generation of a mushroom specific non-wood forest resource mapping system in the near future. Results of nutritional profiling and biological activity studies on the representative species of wild edible mushrooms from the studied region revealed that it is a rich source of essential nutrients and antioxidants.

Epiphytic and endophytic bacteria that promote growth of ethnomedicinal plants in the subtropical forests of Meghalaya, India / Bacterias epífitas y endófitas que promueven el crecimiento de plantas etnomedicinales en los bosques subtropicales de Meghalaya, India

The present study was aimed to investigate the endophytic and epiphytic bacteria associated with selected ethnomedicinal plants from the pristine subtropical forests of Meghalaya and analyse them for plant growth promotion and antagonistic ability. This study is an attempt to explore plant associated bacteria which are beneficial to host plants, and thus aid in the conservation of ethnomedicinal plants of the studied subtropical forests, which are dwindling due to exploitation. The plant growth promotion parameters like indole acetic acid (IAA) production, mineral phosphate solubilisation, acid phosphatase activity, presence of 1-aminocyclopropane-1-carboxylic acid deaminase (ACC) gene, nitrogen fixation, cellulose digestion, chitin and pectin degradation were screened among the isolates. The study revealed significant differences in bacterial population not only between the epiphytic and endophytic microhabitats, but also amongst the host plants. Out of the 70 isolated plant associated bacteria, Bacillus sp., Serratia sp., Pseudomonas sp., Pantoea sp., and Lysinibacillus sp. showed potent plant growth promotion properties. Bacillus siamensis C53 and B. subtilis cenB showed significant antagonistic activity against the tested pathogens. This study indicated the isolates inhabiting the plants prevalent in the subtropical sacred forests could be explored for use as plant growth promoters while practising the cultivation and conservation of ethnomedicinal plants. Rev. Biol. Trop. 62 (4): 1295-1308. Epub 2014 December 01.

El presente estudio tuvo como objetivo investigar las bacterias endófitas y epífitas asociadas con plantas medicinales seleccionadas de los bosques subtropicales prístinos de Meghalaya, para estimular el crecimiento de plantas y la capacidad antagónica. Este estudio es un intento de explorar las plantas asociadas a bacterias que benefician a la planta hospedera, y así ayudar en la conservación de plantas medicinales de los bosques subtropicales estudiados, los cuales son cada vez más escasos debido a la explotación. Los parámetros de promoción de crecimiento de las plantas, tales como: índice de producción de ácido acético (IAA), solubilización de fosfato mineral, actividad de la fosfatasa ácida, presencia del gen 1-aminociclopropano-1-ácido carboxílico desaminasa (ACC), fijación de nitrógeno, digestión de celulosa, quitina y pectina fueron seleccionados entre los aislamientos. El estudio reveló diferencias significativas en la población bacteriana, no sólo entre los microhábitats epífitos y endófitos, sino también entre las plantas hospederas. De las 70 bacterias aisladas de plantas asociadas, Bacillus sp., Serratia sp., Pseudomonas sp., Pantoea sp. y Lysinibacillus sp. mostraron potentes propiedades estimulantes del crecimiento vegetal. Bacillus C53 siamensis y B. subtilis cenB mostraron actividad antagónica significativa contra los patógenos probados. Este estudio indicó que los aislamientos que habitan en las plantas predominantes en los bosques sagrados subtropicales podrían explorarse para su uso como promotores del crecimiento vegetal, mientras se practica el cultivo y conservación de plantas medicinales.

Epiphytic and endophytic bacteria that promote growth of ethnomedicinal plants in the subtropical forests of Meghalaya, India.

The present study was aimed to investigate the endophytic and epiphytic bacteria associated with selected ethnomedicinal plants from the pristine subtropical forests of Meghalaya and analyse them for plant growth promotion and antagonistic ability. This study is an attempt to explore plant associated bacteria which are beneficial to host plants, and thus aid in the conservation of ethnomedicinal plants of the studied subtropical forests, which are dwindling due to exploitation. The plant growth promotion parameters like indole acetic acid (IAA) production, mineral phosphate solubilisation, acid phosphatase activity, presence of 1-aminocyclopropane-1-carboxylic acid deaminase (ACC) gene, nitrogen fixation, cellulose digestion, chitin and pectin degrada- tion were screened among the isolates. The study revealed significant differences in bacterial population not only between the epiphytic and endophytic microhabitats, but also amongst the host plants. Out of the 70 isolated plant associated bacteria, Bacillus sp., Serratia sp., Pseudomonas sp., Pantoea sp., and Lysinibacillus sp. showed potent plant growth promotion properties. Bacillus siamensis C53 and B. subtilis cenB showed significant antagonistic activity against the tested pathogens. This study indicated the isolates inhabiting the plants prevalent in the subtropical sacred forests could be explored for use as plant growth promoters while practising the cultiva- tion and conservation of ethnomedicinal plants.

Cultivable bacterial diversity along the altitudinal zonation and vegetation range of tropical Eastern Himalaya.

The Northeastern part of India sprawls over an area of 262 379km2 in the Eastern Himalayan range. This constitutes a biodiversity hotspot with high levels of biodiversity and endemism; unfortunately, is also a poorly known area, especially on its microbial diversity. In this study, we assessed cultivable soil bacterial diversity and distribution from lowlands to highlands (34 to 3 990m.a.s.l.). Soil physico-chemical parameters and forest types across the different altitudes were characterized and correlated with bacterial distribution and diversity. Microbes from the soil samples were grown in Nutrient, Muller Hinton and Luria-Bertani agar plates and were initially characterized using biochemical methods. Parameters like dehydrogenase and urease activities, temperature, moisture content, pH, carbon content, bulk density of the sampled soil were measured for each site. Representative isolates were also subjected to 16S rDNA sequence analysis. A total of 155 cultivable bacterial isolates were characterized which were analyzed for richness, evenness and diversity indices. The tropical and sub-tropical forests supported higher bacterial diversity compared to temperate pine, temperate conifer, and sub-alpine rhododendron forests. The 16S rRNA phylogenetic analysis revealed that Firmicutes was the most common group followed by Proteobacreria and Bacteroidetes. Species belonging to the genera Bacillus and Pseudomonas were the most abundant. Bacterial CFU showed positive but insignificant correlation with soil parameters like pH (r=0.208), soil temperature (r=0.303), ambient temperature (r=0.443), soil carbon content (r=0.525), soil bulk density (r=0.268), soil urease (r=0.549) and soil dehydrogenase (r=0.492). Altitude (r=-0.561) and soil moisture content (r=-0.051) showed negative correlation. Altitudinal gradient along with the vegetation and soil physico-chemical parameters were found to influence bacterial diversity and distribution. This study points out that this is a biome with a vast reservoir of bacteria which decrease with increasing altitudes, and highlights the microbiological importance of the poorly studied Eastern Himalayan range, justifying efforts to explore the prevalence of novel species in the biome.

Cultivable bacterial diversity along the altitudinal zonation and vegetation range of tropical Eastern Himalaya

The Northeastern part of India sprawls over an area of 262 379km² in the Eastern Himalayan range. This constitutes a biodiversity hotspot with high levels of biodiversity and endemism; unfortunately, is also a poorly known area, especially on its microbial diversity. In this study, we assessed cultivable soil bacterial diversity and distribution from lowlands to highlands (34 to 3 990m.a.s.l.). Soil physico-chemical parameters and forest types across the different altitudes were characterized and correlated with bacterial distribution and diversity. Microbes from the soil samples were grown in Nutrient, Muller Hinton and Luria-Bertani agar plates and were initially characterized using biochemical methods. Parameters like dehydrogenase and urease activities, temperature, moisture content, pH, carbon content, bulk density of the sampled soil were measured for each site. Representative isolates were also subjected to 16S rDNA sequence analysis. A total of 155 cultivable bacterial isolates were characterized which were analyzed for richness, evenness and diversity indices. The tropical and sub-tropical forests supported higher bacterial diversity compared to temperate pine, temperate conifer, and sub-alpine rhododendron forests. The 16S rRNA phylogenetic analysis revealed that Firmicutes was the most common group followed by Proteobacteria and Bacteroidetes. Species belonging to the genera Bacillus and Pseudomonas were the most abundant. Bacterial CFU showed positive but insignificant correlation with soil parameters like pH (r=0.208), soil temperature (r=0.303), ambient temperature (r=0.443), soil carbon content (r=0.525), soil bulk density (r=0.268), soil urease (r=0.549) and soil dehydrogenase (r=0.492). Altitude (r=0.561) and soil moisture content (r=-0.051) showed negative correlation. Altitudinal gradient along with the vegetation and soil physico-chemical parameters were found to influence bacterial diversity and distribution. This study points out that this is a biome with a vast reservoir of bacteria which decrease with increasing altitudes, and highlights the microbiological importance of the poorly studied Eastern Himalayan range, justifying efforts to explore the prevalence of novel species in the biome.

La parte noreste de la India se extiende sobre una superficie de 262 379km² en la cordillera oriental del Himalaya. Es un punto de acceso con altos niveles de biodiversidad y endemismo; desafortunadamente, también es una zona poco conocida, sobre todo su diversidad microbiana. En este estudio se evaluó la diversidad de bacterias cultivables del suelo, su diversidad y distribución de las tierras bajas a las altas (34 a 3 990m.s.n.m). Se caracterizaron los parámetros físico-químicos del suelo y tipos de bosques a lo largo del gradiente altitudinal y se correlacionaron con la distribución y diversidad bacteriana. Los microbios del suelo se cultivaron en placas de agar enriquecido Muller Hinton y Luria-Bertani, e inicialmente se caracterizaron mediante métodos bioquímicos. Parámetros tales como actividad de la deshidrogenasa y ureasa, temperatura, contenido de humedad y de carbono, pH y densidad aparente del suelo se midieron en cada sitio. Aislamientos representativos también se sometieron al análisis secuencial de 16S rADN. Un total de 155 aislamientos bacterianos cultivables se caracterizaron para estimar los índices de riqueza, equidad y diversidad. Los bosques tropicales y subtropicales albergan una mayor diversidad bacteriana en comparación con los bosques templados de pino y coníferas, y los bosques subalpinos de rododendro. El análisis filogenético de 16S rARN reveló que Firmicutes fue el grupo más común, seguido de Proteobacteria y Bacteroidetes. Especies pertenecientes a los géneros Bacillus y Pseudomonas fueron las más abundantes. Las UFC bacterianas mostraron una positiva pero insignificante correlación con los parámetros del suelo, tales como pH (r=0.208), temperatura (r=0.303), temperatura ambiente (r=0.443), contenido de carbón (r=0.525), densidad aparente (r=0.268), ureasa (r=0.549) y deshidrogenasa (r=0.492). La altitud (r=-0.561) y el contenido de humedad del suelo (r=-0.051) mostraron una correlación negativa. Se encontró que el gradiente altitudinal, junto con la vegetación y los parámetros físico-químicos influyeron en la diversidad bacteriana y la distribución. Este estudio señala que este es un bioma con un vasto reservorio de bacterias que disminuyen con la altitud y pone en relieve la importancia microbiológica de la pobremente estudiada zona del este del Himalaya, lo que justifica los esfuerzos para explorar la prevalencia de nuevas especies en el bioma.

Influence of roadside pollution on the phylloplane microbial community of Alnus nepalensis (Betulaceae)

The North Eastern region of India is undergoing industrial development at a faster rate than expected. Roads form the main system of transportation and communication owing to the hilly topography of the region. Automobiles discharge a number of gaseous and trace metal contaminants. Human activities like stone grinding, road construction and sand milling also increase the atmospheric dust and heavy metal contaminant level. These contaminants get settled on leaf surfaces at roadsides and enter in contact with phylloplane microorganisms. This study compares microorganisms on leaf surfaces of alder (Alnus nepalensis (Betulaceae)) on roadside and non-roadside environments. Two sites dominated by alder were selected. One at a busy road intersection on the National Highway no. 44 in Shillong with high traffic density (8 000-9 000 heavy vehicles/day), taken as the polluted site and the other one in a forest approximately 500 m away from the roadside considered as the unpolluted site. Analysis of phylloplane microorganisms, lead, zinc, copper, cadmium and sulphur was carried out from leaves. The bacterial population was higher at the unpolluted site. Bacterial population showed a significant negative correlation with lead, zinc, copper, cadmium and sulphur. Similarly, fungal population was higher at the unpolluted site. A total of 29 fungal species were isolated from the phylloplane of A. nepalensis (polluted site 16 species; unpolluted site 28 species). Some fungal forms like Mortierella sp., Fusarium oxysporum and Aureobasidium pollulans were dominant in the polluted site. Numbers of phylloplane fungi and bacteria were significantly reduced in the polluted site. The correlation coefficient indicated a detrimental effect of metals like lead, zinc, copper, cadmium and sulphur on the microbial community of leaf surfaces. The specificity of certain fungi to the unpolluted site may be attributed to their sensitivity to pollution. The predominance of Aureobasidium pollulans, Fusarium oxysporum and Mortierella sp. in the polluted site may be due to their resistance to pollutants in roadside conditions. Rev. Biol. Trop. 56 (3): 1521-1529. Epub 2008 September 30.

La región nororiental de la India está experimentando un rápido desarrollo industrial y debido a la topografía montañosa de la región, los automóviles descargan variedad de contaminantes. Actividades humanas como moler piedra y construir caminos también aumentan el polvo atmosférico y la contaminación con metales pesados. Este estudio compara microorganismos en superficies foliares del árbol Alnus nepalensis (Betulaceae). Se estudió uno sitio en una intersección de Shillong, con alta densidad de tráfico (8 000-9 000 vehículos pesados/día), tomada como el sitio contaminado, y otro en un bosque aproximadamente a 500 m de la carretera (sitio no contaminado). Se contabilizaron bacterias, hongos, plomo, zinc, cobre, cadmio y sulfuro. La población bacteriana era más alta en el sitio no contaminado. La población bacteriana mostró una correlación negativa significativa con el contenido de plomo, cinc, cobre, cadmio y sulfuro foliares en el sitio contaminado, mientras que en el sitio no contaminado la correlación fue insignificante. La población de hongos también fue más alta en el sitio no contaminado. Un total de 29 especies de hongos fueron aislados A. nepalensis: 16 en el sitio contaminado y 28 en el no contaminado. Algunas especies de hongos como Mortierella sp., Fusarium oxysporum y Aureobasidium pollulans fueron dominantes en el sitio contaminado. El número de hongos y bacterias fue significativamente menor en el sitio contaminado. El coeficiente de correlación indicó un efecto perjudicial de metales como el plomo, el zinc, el cobre, el cadmio y el sulfuro en esta comunidad que habitan superficies de hojas. La especificidad de ciertos hongos al sitio no contaminado se puede atribuir a su sensibilidad a la contaminación. El predominio de Aureobasidium pollulans, Fusarium oxysporum y Mortierella sp. en el sitio contaminado puede ser debido a su resistencia a los agentes contaminadores.

Influence of roadside pollution on the phylloplane microbial community of Alnus nepalensis (Betulaceae).

The North Eastern region of India is undergoing industrial development at a faster rate than expected. Roads form the main system of transportation and communication owing to the hilly topography of the region. Automobiles discharge a number of gaseous and trace metal contaminants. Human activities like stone grinding, road construction and sand milling also increase the atmospheric dust and heavy metal contaminant level. These contaminants get settled on leaf surfaces at roadsides and enter in contact with phylloplane microorganisms. This study compares microorganisms on leaf surfaces of alder (Alnus nepalensis (Betulaceae)) on roadside and non-roadside environments. Two sites dominated by alder were selected. One at a busy road intersection on the National Highway no. 44 in Shillong with high traffic density (8 000-9 000 heavy vehicles/day), taken as the polluted site and the other one in a forest approximately 500 m away from the roadside considered as the unpolluted site. Analysis of phylloplane microorganisms, lead, zinc, copper, cadmium and sulphur was carried out from leaves. The bacterial population was higher at the unpolluted site. Bacterial population showed a significant negative correlation with lead, zinc, copper, cadmium and sulphur. Similarly, fungal population was higher at the unpolluted site. A total of 29 fungal species were isolated from the phylloplane of A. nepalensis (polluted site 16 species; unpolluted site 28 species). Some fungal forms like Mortierella sp., Fusarium oxysporum and Aureobasidium pollulans were dominant in the polluted site. Numbers of phylloplane fungi and bacteria were significantly reduced in the polluted site. The correlation coefficient indicated a detrimental effect of metals like lead, zinc, copper, cadmium and sulphur on the microbial community of leaf surfaces. The specificity of certain fungi to the unpolluted site may be attributed to their sensitivity to pollution. The predominance of Aureobasidium pollulans, Fusarium oxysporum and Mortierella sp. in the polluted site may be due to their resistance to pollutants in roadside conditions.