Hydraulic characteristics and incubation methods for enhancing durability of Fungi-Mycelium treated silica sand using Rhizopus oligosporus and Rhizopus oryzae combination

Nowadays,the application of Fungi as a bio-mediated soil improvement technique is developing.The hydraulic properties of Rhizopus Fungi-Mycelium Treated Soil are unknown,and the treated sample tends to have low durability.This article presents experimental results on the hydraulic conductivity and shear strength of Fungi-mycelium-treated silica sand.The fungi used in the experiments are a combination ofRhizopus oligosporus andRhizopus oryzae,which are popular for making Tempeh,a local soybean cuisine from Indonesia.The samples were prepared by mixing the sand with Tempeh inoculum at various treatments and Tempe inoculum and rice flour dosages for enhancing the durability of the treated soil.The results showed that the saturated permeability of the treated soil could be reduced by about 10 times compared to the untreated soil.In addition,the Soil-Water Characteristic Curve of the treated soil also developed.The effect of the fungi appears to fill the void of soil and hence increases the Air Entry Value and residual suction of soil.The curing method outside the mold(O-method)with 10%Tempeh inoculum,and 5%Tempeh inoculum with 5%rice flour is proven can extend the durability of the treated sample,the undrained compressive strength is about 40 kPa on day 14.Scanning electron microscope was performed on the samples,which lasted for 4 months.The mycelium and hyphae are still clearly seen covering all sand particles with different percentages of Tempeh inoculum and rice flour.When the mycelium covered all the sand particles and filled the pores,the water flow was partially blocked.It might be attributed to the strong hydrophobicity of the fungi,which could prevent water from penetrating the soil.

Mixture of Bacillus Amyloliquefaciens and Bacillus Pumilus Modulates Community Structures of Rice Rhizosphere Soil to Suppress Rice Seedling Blight

Rice seedling blight,caused by various fungi,including Fusarium oxysporum,poses a severe threat to rice production.As awareness grows regarding the environmental and safety hazards associated with the application of fungicides for managing rice seedling blight,there has been a shift in focus towards biological control agents.In this study,we isolated biocontrol bacteria from paddy fields that significantly inhibited the growth of F.oxysporum in vitro and identified the strains as Bacillus amyloliquefaciens T40 and Bacillus pumilus T208.Additionally,our findings indicated that the combined application of these Bacillus strains in soil was more effective in reducing the incidence of rice seedling blight than their individual use.Analysis of 16S and internal transcribed spacer rRNA gene sequencing data revealed that the mixture of the T40 and T208 strains exhibited the lowest average clustering coefficients,which were negatively correlated with the biomass of F.oxysporum-inoculated rice seedlings.Furthermore,this mixture led to higher stochastic assembly(average|βNTI|<2)and reduced selection pressures on rice rhizosphere bacteria compared with individual strain applications.The mixture of the T40 and T208 strains also significantly increased the expression of defense-related genes.In conclusion,the mixture of the T40 and T208 strains effectively modulates microbial community structures,enhances microbial network stability,and boosts the resistance against rice seedling blight.Our study supports the development and utilization of biological resources for crop protection.

培养过程参数对霉菌Rhizopus oryzae IFO细胞催化植物油脂合成生物柴油的影响研究

采用脂肪酶催化可再生动植物油脂合成生物柴油已经成为目前研究的热点,其中利用全细胞催化剂是一个重要的研究方向。文中直接利用霉菌R.oryzaeIFO细胞催化植物油脂甲醇解反应合成生物柴油,系统研究了培养过程参数对细胞生长和该细胞催化剂催化甲醇解反应活性的影响。研究表明,细胞培养过程中所添加的油脂不同,细胞在后续反应中催化特定油脂进行生物柴油制备时所表现出的催化活性也会有所差别;由某种油脂作为碳源得到的细胞催化剂催化对应油脂与甲醇转酯化反应制备生物柴油时,表现出比催化其他油脂和甲醇反应制备生物柴油更高的催化活性。在优化的操作参数(大豆精制油20g/L,蛋白胨70g/L,NaNO31.2g/L,KH2PO41.2g/L,MgSO4·7H2O0.5g/L,培养温度35℃,摇床转速130r/min)下,培养得到的细胞催化剂能有效催化大豆油与甲醇三步转酯化反应,生物柴油(脂肪酸甲酯)最终得率可达到86%。

Actinomucor elegans、Aspegillus oryzae和Rhizopus oligosporus产蛋白酶条件及蛋白酶性质的比较

比较了腐乳生产菌株Actinomucor elegans、豆酱和酱油生产菌株Aspegillus oryzae以及天培生产菌株Rhizopus oligosporus产生蛋白酶的条件和所产蛋白酶的性质。结果表明,不同的菌株产酶条件及蛋白酶的性质有较大的差异:少孢根霉主要产生酸性蛋白酶,在pH2.5-4.0的酸性介质中、32℃条件下培养时产酶能力较强,所分泌的蛋白酶系在pH5.0时酶活力最高,在pH5.0附近最稳定;米曲霉可以产生酸性、中性及碱性蛋白酶,所产生的蛋白酶活力显著高于少孢根霉和毛霉,米曲霉在酸性条件下产酸性蛋白酶能力强,在中性条件下产中性蛋白酶能力强,在碱性条件下产碱性蛋白酶能力强,在28-32℃时产酶能力强,所分泌的蛋白酶系在pH5.0-9.0的广泛范围内有很强的活力,在pH6.0-8.0的范围内稳定性强;毛霉可以产生酸性、中性及碱性蛋白酶,但酶活力明显低于米曲霉,毛霉在中性偏酸性(pH5.5)的介质中产酸性蛋白酶的能力较强,但介质的酸碱度对毛霉产中性及碱性蛋白酶没有影响,在28℃时产酸性、中性和碱性蛋白酶的能力都比较强,毛霉所分泌的蛋白酶系在pH5.0-9.0的广泛pH范围内有活力,在pH5.0-6.0时酶活力最高,在pH5.0-7.0时稳定强。

Rhizopus oryzae产酸性蛋白酶条件及其酶学性质研究

本文以少孢根霉（Rhizopus oligosporus）为对照研究了豆豉中的一个分离菌株米根霉（Rhizopus oryzae）产生酸性蛋白酶的条件及所产蛋白酶的性质，结果表明这个茵株的产酶条件和蛋白酶的性质与少孢根霉相比有相似性但也存在一些差异：米根霉在水分含量57％-59％、pH2．5-3．0的酸性介质中、28-31℃下培养36h时产酸性蛋白酶能力最强，所分泌的蛋白酶系在pH4．0和pH6．0附近有最强的催化活性，在pH3．0-6.．0的范围内有较好的稳定性，催化反应的最适作用温度为50℃，它的温度稳定性很差，在50℃保温30min已完全失活；少孢根霉在水分含量52％-55％、pH2．5-3．0的酸性介质中、31℃下培养48h时产酸性蛋白酶能力最强，在35℃条件下培养36h也能产生较高的酶活力，少孢根霉分泌的蛋白酶系在pH3．0和pH6．0附近有最强的催化活性，在pH4．0—6．0范围内很稳定，催化反应的最适作用温度可达55～60℃，但它的温度稳定性较差，在50℃保温30min，酶活力损失达到90%，保温120min酶几乎完全失活。

Production of Ammonium Lactate by Fed-batch Fermentation of Rhizopus oryzaefrom Corncob Hydrolysate

L-(+)-Lactic acid production from corncob hydrolysate as a cheap carbohydrate source by fed-batch fermentation of Rhizopus oryzaeHZS6 was studied. After 96 h of fermentation in a 5 L fermentor, the final concentration of ammonium L-(+)-lactate, average productivity(based on initial xylose concentration) and maximum dry cell weight were 132.4 g/L, 1.38 g/(L·h), and 8.9 g/L respectively. The optical purity of L-(+)-lactate was 98.8%.

Rhizopus oryzae LS1利用丢糟水解液发酵生产乳酸

研究了一株诱变米根霉Rhizopus oryzae LS-1利用丢糟水解液发酵生产乳酸的可行性。首先考察R.oryzae LS-1利用葡萄糖和木糖的糖代谢差异特性,并通过正交实验优化丢糟水解液发酵生产乳酸的工艺参数。结果表明,R.oryzae LS-1能代谢利用葡萄糖和木糖,且二者存在协同互补作用,有利于乳酸生成和糖酸转化,可用于木质纤维原料的乳酸生产。丢糟水解液发酵生产乳酸的实验表明,氯化铵是R.oryzae LS-1适宜的氮源。在接种量为3.0%、p H为6.5、发酵时间为96h、Ca CO3添加量为80g/L的条件下,乳酸生成浓度为13.27g/L,糖利用率为79.61%。说明诱变菌株R.oryzae LS-1具备发酵丢糟水解液制备乳酸的潜力。

Optimization of L-lactic Acid Production of Rhizopus Oryzae Mutant RLC41-6 by Ion Beam Implantation at Low-Energy

In order to obtain an industrial strain with a higher L（＋）-lactic acid yield, the strain Rhizopus oryzae RF3608 was mutated by means of nitrogen ion beam implantation and the mutant strain RLC41-6 was isolated. Under optimal conditions the yield of L（＋）-lactic acid produced in a shake-flask reached 133 g/L - 137 g/L after 36 h cultivation, indicating that the conversion rate based on glucose was as high as 88% - 91% and the productivity was 3.75 g/L.h. It was almost a 115% increase in lactic acid production compared with the original strain RF3608.

Optimization of L(+)-Lactic Acid Production from Xylose with Rhizopus Oryzae Mutant RLC41-6 Breeding by Low-Energy Ion Implantation

In order to obtain an industrial strain with a higher L（＋）-lactic acid yield, the strain Rhizopus oryzae PW352 was mutated by means of nitrogen ion beam implantation and the mutant strain Rhizopus oryzae RLC41-6 was obtained. An experimental finding was made in surprise that Rhizopus oryzae mutant RLC41-6 is not only an L（＋）-lactic acid producer from corn starch but also an efficient producer of L（＋）-lactic acid from xylose. Under optimal conditions, the production of L（＋）-lactic acid from 100 g/L xylose reached 77.39 g/L after 144 h fed-batch fermentation, A high mutation rate and a wide mutation spectrum of low-energy ion implantation were observed in the experiment.

Improvement of L(+)-Lactic Acid Production of Rhizopus Oryzae by Low-Energy Ions and Analysis of Its Mechanism

The wild type strain Rhizopus oryzae PW352 was mutated by means of nitrogen ion implantation （15 keV, 7.8×10^14 ～ 2.08 ×10^15 ions/cm^2） to find an industrial strain with a higher L（＋）-lactic acid yield, and two mutants RE3303 and RF9052 were isolated. In order to discuss the mechanism primarily, Lactate Dehydrogenase of Rhizopus oryzae was studied. While the two mutants produced L（＋）-lactic acid by 75% more than the wild strain did, their specific activity of Lactate Dehydrogenase was found to be higher than that in the wild strain. The optimum temperature of Lactate Dehydrogenase in Rhizopus oryzae RF9052 was higher. Compared to the wild strain, the Michaelis constant （Km） value of Lactate Dehydrogenase in the mutants was Changed. All these changes show that L（＋）-lactic acid production has a correlation with the specific activity of Lactate Dehydrogenase. The low-energy ions, implanted into the strain, may improve the specific activity of Lactate Dehydrogenase by influencing its gene structure and protein structure.

Repeated-Batch and Continuous Production of L-Lactic Acid by Rhizopus oryzae Immobilized in Calcium Alginat Beads:ReactorPerformance and Kinetic Model

Repeated-batch and continuous production of L-lactic acid by immobilized Rhizopusoryzae with calcium alginate entrapment method in a three-phase fluidized-bed bioreactor was stud-ied.The operation conditions were optimized.The productivity based on total reactor volume wasabout 3 times higher than that with free cells in a traditional stirred tank bioreactor.A mathemat-ical model was proposed and the model predictions were in good agreement with the experimentaldat.

烘烤期烟叶霉烂病病原菌Rhizopus oryzae的生态学研究

旨在探明米根霉(Rhizopus oryzae)与烘烤期烟叶霉烂病相关的生态学关联性,为制订该病害防控策略提供依据。通过组织保湿法研究了烟草叶柄、茎杆不同部位自然发病率。通过人工接种法研究了叶柄不同含水量与发病率关系。用稀释平板分离法测定了烟草地块及后茬作物根围土内的米根霉含量。结果表明烟草根围土中根霉菌主要为米根霉,并能引起烟草叶柄发病;离茎杆越近的叶柄部位发病率越高,连作烟株茎杆发病重于新种烟株,米根霉能进入茎杆内部,主要侵染植株中下部1~14节;烟草叶柄含水量低于40%时不能发病,含水量在70%~80%之间时发病最重;米根霉数量在连作田烟株根围土中大于新植田烟株根围土,在黑胫病病株根围土中大于健株根际土;米根霉在后茬作物西葫芦和花菜根围土中较多,在豌豆和蚕豆根围土中较少。本研究结果对制订烘烤期烟叶霉烂病防控策略有重要意义。

米根霉(Rhizopus oryzae)脂肪酶基因ProROL在毕赤酵母(Pichia pastoris)中的高效表达

米根霉(Rhizopus oryzae)脂肪酶在食品和油脂加工等方面具有广泛的应用前景。本研究通过PCR技术获得了米根霉脂肪酶前肽基因序列与成熟肽基因序列(ProROL),并将该基因克隆到表达载体pPIC9K上。线性化表达质粒后电转化感受态毕赤酵母菌株(Pichia pastoris)GS115,通过筛选得到了阳性重组菌。摇瓶及7 L罐发酵结果显示,以p-NPP为底物,上清胞外酶活分别为10.6 U/mL和126.8 U/mL。酶学性质研究发现,该酶的最适温度为40℃左右,在低于40℃条件下,保温30 min后,能够保存60%以上的酶活力;其最适pH为8.5左右,在pH 6.0-10.0范围内保持1 h后,均能保留60%以上的酶活力。本研究构建的基因重组菌株将为该酶的工业化生产与应用提供一定的理论基础。

Optimization of L(+)-Lactic Acid Fermentation Without Neutralisation of Rhizopus Oryzae Mutant RK02 by Low-Energy Ion Implantation

In order to get an industrial strain which can yield a high concentration of lactic acid for ISPR （in situ product removal）, the original strain Rhizopus oryzae RE3303 was mutated by low-energy ion beam implantation. A mutant RK02 was screened, and the factors such as the substrate concentration, nitrogen source concentration, inoculum size, seed age, aeration and temperature that affect the production of lactic acid were studied in detail. Under optimal con- ditions, the maximum concentration of L（＋）-lactic acid reached 34.85 g/L after 30 h shake-flask cultivation without adding any neutralisation （5% Glucose added）, which was a 146% increase in lactic acid production after ion implantation compared with the original strain. It was also shown that RK02 can be used in ISPR to reduce the number of times of separation.

米根霉Rhizopus oryzae LS-1对糠醛抑制物的耐受性

通过考察不同糠醛浓度对米根霉(Rhizopus oryzae LS-1)发酵产乳酸过程中糖酸转化率和糖利用率的影响,探索了解Rhizopus oryzae LS-1对木质纤维水解副产物糠醛抑制物的耐受性,寻找Rhizopus oryzae LS-1对糠醛抑制物的耐受浓度。结果表明:Rhizopus oryzae LS-1对糠醛的耐受浓度为12.5g/L,在0~12.5g/L可提高糖酸转化率和糖利用率。当糠醛浓度大于12.5g/L时,菌株生长完全受到抑制。研究结果对乳酸生产用木质纤维素水解液的脱毒处理工艺具有指导作用。

The response of roots and the rhizosphere environment to integrative cultivation practices in paddy rice

Integrative cultivation practices(ICPs)are essential for enhancing cereal yield and resource use efficiency.However,the effects of ICP on the rhizosphere environment and roots of paddy rice are still poorly understood.In this study,four rice varieties were produced in the field.Each variety was treated with six different cultivation techniques,including zero nitrogen application(0 N),local farmers’practice(LFP),nitrogen reduction(NR),and three progressive ICP techniques comprised of enhanced fertilizer N practice and increased plant density(ICP1),a treatment similar to ICP1 but with alternate wetting and moderate drying instead of continuous flooding(ICP2),and the same practices as ICP2 with the application of organic fertilizer(ICP3).The ICPs had greater grain production and nitrogen use efficiency than the other three methods.Root length,dry weight,root diameter,activity of root oxidation,root bleeding rate,zeatin and zeatin riboside compositions,and total organic acids in root exudates were elevated with the introduction of the successive cultivation practices.ICPs enhanced nitrate nitrogen,the activities of urease and invertase,and the diversity of microbes(bacteria)in rhizosphere and non-rhizosphere soil,while reducing the ammonium nitrogen content.The nutrient contents(ammonium nitrogen,total nitrogen,total potassium,total phosphorus,nitrate,and available phosphorus)and urease activity in rhizosphere soil were reduced in all treatments in comparison with the non-rhizosphere soil,but the invertase activity and bacterial diversity were greater.The main root morphology and physiology,and the ammonium nitrogen contents in rhizosphere soil at the primary stages were closely correlated with grain yield and internal nitrogen use efficiency.These findings suggest that the coordinated enhancement of the root system and the environment of the rhizosphere under integrative cultivation approaches may lead to higher rice production.

Rhinocerebral mucormycosis caused by Rhizopus oryzae in a patient with acute myeloid leukemia: A case report

BACKGROUND Rhinocerebral mucormycosis(RCM)is a rare fatal fungal infection which is on the increase among immunocompromised hosts such as patients who have had hematological cancers,or have received immunosuppressive drugs,corticosteroids,or other T cell suppressing agents.CASE SUMMARY We report a case of RCM caused by Rhizopus oryzae,one of the most common opportunistic pathogens,in a patient suffering from a fourth relapse of acute myeloid leukemia.The patient developed RCM after he had received long-term antibiotic agents and corticosteroids.The pathogen was isolated three times from nasal secretions collected from the deep parts of the nasal cavity and was identified by morphology and internal transcribed spacer sequencing.Blood infection was excluded by droplet digital polymerase chain reaction and blood culture.The patient was empirically treated with caspofungin and voriconazole for several days while the lesions continued to progress.The patient was given amphotericin B in combination with caspofungin after RCM was suspected,and the lesions improved over the course of treatment,which lasted several days.However,the patient eventually died of the primary disease.CONCLUSION This case indicates that immunosuppressive drugs,including corticosteroids and antimetabolites in hematological tumor,do increase the risk of infections of this type.Early diagnosis,prompt and frequent surgical debridement,and treatment with amphotericin B without delay are all essential in combatting RCM.

The DNA damage repair complex MoMMS21-MoSMC5 is required for infection-related development and pathogenicity of Magnaporthe oryzae

The conserved DNA damage repair complex,MMS21-SMC5/6(Methyl methane sulfonate 21-Structural maintenance of chromosomes 5/6),has been extensively studied in yeast,animals,and plants.However,its role in phytopathogenic fungi,particularly in the highly destructive rice blast fungus Magnaporthe oryzae,remains unknown.In this study,we functionally characterized the homologues of this complex,MoMMS21 and MoSMC5,in M.oryzae.We first demonstrated the importance of DNA damage repair in M.oryzae by showing that the DNA damage inducer phleomycin inhibited vegetative growth,infection-related development and pathogenicity in this fungus.Additionally,we discovered that MoMMS21 and MoSMC5 interacted in the nuclei,suggesting that they also function as a complex in M.oryzae.Gene deletion experiments revealed that both MoMMS21 and MoSMC5 are required for infection-related development and pathogenicity in M.oryzae,while only MoMMS21 deletion affected growth and sensitivity to phleomycin,indicating its specific involvement in DNA damage repair.Overall,our results provide insights into the roles of MoMMS21 and MoSMC5 in M.oryzae,highlighting their functions beyond DNA damage repair.

立枯丝核菌Rhi-milR9829-5p与水稻和大豆互作的分子机理

立枯丝核菌可侵染水稻、大豆等260多种植物,是作物生产中的主要真菌性病害致病菌之一。microRNA(miRNA)是一类由20~25个核苷酸组成的非编码RNA分子,可以通过降解mRNA或抑制mRNA的翻译从而在转录或翻译水平上负调控靶基因的表达。已有研究表明,miRNA参与了真菌与植物互作等多种生物学过程。在前期研究中,课题组鉴定到一个能够负调控玉米抗病性的立枯丝核菌Rhi-milR9829-5p,为了进一步研究其在不同作物中的致病功能和机理,本研究获得了Rhi-milR9829-5p成熟体过量表达的水稻转基因植株,对阳性T1代植株接种立枯丝核菌后发现,Rhi-milR9829-5p过量表达的水稻植株对纹枯病的抗性显著降低。利用生物信息学方法分别预测了Rhi-milR9829-5p在水稻和大豆中的下游靶基因并进行功能注释分析,结果表明Rhi-milR9829-5p通过多种生物学途径调控水稻和大豆的抗病性,且在水稻和大豆中存在明显不同。研究结果初步揭示了立枯丝核菌Rhi-milR9829-5p调控水稻和大豆抗病性的分子机理,进一步验证了miRNA的跨物种调控功能,为作物抗病分子育种提供了基因资源。