The single spores were isolated from chlamydospores of Ustilaginoidea virens with three different maturities by PSA. The isolated single spores were cul- tured on different media at different temperatures under natura...The single spores were isolated from chlamydospores of Ustilaginoidea virens with three different maturities by PSA. The isolated single spores were cul- tured on different media at different temperatures under natural light for inducing conidia to explore the optimum isolation technique of single spore and optimum cul- ture condition of conidia. The results showed that the successful isolating rate of single spore from yellow rice false smut balls reached 90.00%. The sporulatina quantities of isolated single spores cultured on PSD and PDB media at 22 -29 ~C （variable temperature under natural light） or 28 ℃ （constant temperature under dark condition） for 12 d were up to 6.3× 107 and 1.1× 106 spore/mL, respectively. PSA was the most effective method to isolate single spores from yellow rice false smut balls of U. virens. The optimum conidia culture condition included PSD or PDB medium, 22 -29℃ or 28℃, natural light and vibration culture. Key words Ustilaginoidea virens; Single spore isolation; Conidia; Culture condition展开更多
Streptomyces is a model bacterium to study multicellular differentiation and the major reservoir for antibiotics discovery.However,the cellular‐level lifecycle of Streptomyces has not been well studied due to its com...Streptomyces is a model bacterium to study multicellular differentiation and the major reservoir for antibiotics discovery.However,the cellular‐level lifecycle of Streptomyces has not been well studied due to its complexity and lack of research tools that can mimic their natural conditions.In this study,we developed a simple microfluidic chip for the cultivation and observation of the entire lifecycle of Streptomyces development from the single‐cell perspective.The chip consists of channels for loading samples and supplying nutrients,microwell arrays for the seeding and growth of single spores,and air chambers beside the microwells that facilitate the development of aerial hyphae and spores.A unique feature of this chip is that each microwell is surrounded by a 1.5µm nanogap connected to an air chamber,which provides a stabilized water–air interface.We used this chip to observe the lifecycle development of Streptomyces coelicolor and Streptomyces griseus germinated from single spores,which revealed differentiation of aerial hyphae with progeny spores at micron‐scale water–air interfaces and air chambers.Finally,we demonstrated the applicability of this chip in phenotypic assays by showing that the microbial hormone A‐Factor is involved in the regulatory pathways of aerial hyphae and spore formation.The microfluidic chip could become a robust tool for studying multicellular differentiation,single‐spore heterogeneity,and secondary metabolism of single‐spore germinated Streptomyces.展开更多
Perilla frutescens(L.)Britt.is widely cultivated in China for its medicinal and culinary uses.Rust disease is prevalant,and hyperparasitism was usually observed in Perilla producing regions annually.The urediniospores...Perilla frutescens(L.)Britt.is widely cultivated in China for its medicinal and culinary uses.Rust disease is prevalant,and hyperparasitism was usually observed in Perilla producing regions annually.The urediniospores of Perilla rust fungus are spherical,ovoid,or pyriform,and the conidia of its hyperparasite are oblong,elliptical,or short-cylindrical.A pure culture of the hyperparasite was isolated using the single spore isolation method.The 28S large ribosomal subunit rDNA gene(nrLSU)of rust fungus,and the internal transcribed spacer region(ITS-rDNA)of the hyperparasite were amplified.These two fungi shared 100%identity with Coleosporium plectranthi and Ramularia coleosporii.Phylogenetic trees showed that whole Perilla rust isolates clustered in a clade with C.plectranthi,while the hyperparasite fungus clustered in a clade with R.coleosporii,with a bootstrap support value of 93%and 98%.Combining micro-morphology and molecular sequence alignment,the rust fungus and its hyperparasite were identified as C.plectranthi and R.coleosporii.Urediniospore pustules of C.plectranthi gradually turned white after inoculation of R.coleosporii,while pustules inoculated with PDA plugs showed no change,which confirmed their hyperparasitic relationship.This study determined C.plectranthi as the causal agent of Perilla rust in China,and R.coleosporii as its hyperparasite.展开更多
基金Science and Technology Project of Guizhou Province[QKH Major Project( 2012 ) 6012 ]Supported by Science and Technology Fund of Guizhou Province ( QKH J[2009]2103)+1 种基金Science and Technology Research Programof Guizhou Province ( QKH NY[2012]3031)Project of Guizhou Academy of Agricultural Sciences ( [2010]033 and QNKH[Major]07016)
文摘The single spores were isolated from chlamydospores of Ustilaginoidea virens with three different maturities by PSA. The isolated single spores were cul- tured on different media at different temperatures under natural light for inducing conidia to explore the optimum isolation technique of single spore and optimum cul- ture condition of conidia. The results showed that the successful isolating rate of single spore from yellow rice false smut balls reached 90.00%. The sporulatina quantities of isolated single spores cultured on PSD and PDB media at 22 -29 ~C （variable temperature under natural light） or 28 ℃ （constant temperature under dark condition） for 12 d were up to 6.3× 107 and 1.1× 106 spore/mL, respectively. PSA was the most effective method to isolate single spores from yellow rice false smut balls of U. virens. The optimum conidia culture condition included PSD or PDB medium, 22 -29℃ or 28℃, natural light and vibration culture. Key words Ustilaginoidea virens; Single spore isolation; Conidia; Culture condition
基金We would like to thank Dr.Liang Ma from Dawei Biotechnologies(Beijing,China)for his insightful suggestion.This study was supported by the National Key Research&Development Program of China(2021YFC2301000,2021YFA0717000,and 2021YFC2103300)the National Natural Science Foundation of China(31970091,91951103,and 21822408).
文摘Streptomyces is a model bacterium to study multicellular differentiation and the major reservoir for antibiotics discovery.However,the cellular‐level lifecycle of Streptomyces has not been well studied due to its complexity and lack of research tools that can mimic their natural conditions.In this study,we developed a simple microfluidic chip for the cultivation and observation of the entire lifecycle of Streptomyces development from the single‐cell perspective.The chip consists of channels for loading samples and supplying nutrients,microwell arrays for the seeding and growth of single spores,and air chambers beside the microwells that facilitate the development of aerial hyphae and spores.A unique feature of this chip is that each microwell is surrounded by a 1.5µm nanogap connected to an air chamber,which provides a stabilized water–air interface.We used this chip to observe the lifecycle development of Streptomyces coelicolor and Streptomyces griseus germinated from single spores,which revealed differentiation of aerial hyphae with progeny spores at micron‐scale water–air interfaces and air chambers.Finally,we demonstrated the applicability of this chip in phenotypic assays by showing that the microbial hormone A‐Factor is involved in the regulatory pathways of aerial hyphae and spore formation.The microfluidic chip could become a robust tool for studying multicellular differentiation,single‐spore heterogeneity,and secondary metabolism of single‐spore germinated Streptomyces.
基金supported by grants from the National Key R&D Program of China(2022YFC3501504)the CAMS Innovation Fund for Medical Sciences(2021-I2M-1-032).
文摘Perilla frutescens(L.)Britt.is widely cultivated in China for its medicinal and culinary uses.Rust disease is prevalant,and hyperparasitism was usually observed in Perilla producing regions annually.The urediniospores of Perilla rust fungus are spherical,ovoid,or pyriform,and the conidia of its hyperparasite are oblong,elliptical,or short-cylindrical.A pure culture of the hyperparasite was isolated using the single spore isolation method.The 28S large ribosomal subunit rDNA gene(nrLSU)of rust fungus,and the internal transcribed spacer region(ITS-rDNA)of the hyperparasite were amplified.These two fungi shared 100%identity with Coleosporium plectranthi and Ramularia coleosporii.Phylogenetic trees showed that whole Perilla rust isolates clustered in a clade with C.plectranthi,while the hyperparasite fungus clustered in a clade with R.coleosporii,with a bootstrap support value of 93%and 98%.Combining micro-morphology and molecular sequence alignment,the rust fungus and its hyperparasite were identified as C.plectranthi and R.coleosporii.Urediniospore pustules of C.plectranthi gradually turned white after inoculation of R.coleosporii,while pustules inoculated with PDA plugs showed no change,which confirmed their hyperparasitic relationship.This study determined C.plectranthi as the causal agent of Perilla rust in China,and R.coleosporii as its hyperparasite.
