嗜热真菌DSM10635生产耐热木聚糖酶的小试研究

应用嗜热真菌Thermomyces lanuginosus DSM10635,采用固体发酵的方法探索耐热木聚糖酶的优化生产条件。在研究玉米芯,玉米皮,玉米秆,麸皮,松树屑,桦树屑等不同底物,在不同温度、玉米芯颗粒大小以及料水比条件下培养比较酶产量后,发现该嗜热真菌产耐热木聚糖酶的最佳底物为玉米芯或玉米皮,最佳培养温度为50℃--55℃,在加水量为1份玉米芯：2．8份水,玉米芯的颗粒直径大约为1mm时产酶量最高。实验结果显示,嗜热真菌DSM10635在优化后的培养条件下木聚糖酶产量可达到12525．80IU／g玉米芯。     

四种嗜热真菌的分离与鉴定

目的:在对嗜热真菌的资源调查中,分离到嗜热真菌20株。方法:通过形态学比较研究并结合分子分析方法。结果:鉴定出嗜热真菌4种,即杜邦青霉Penicillium dupontii、疏绵状嗜热丝孢菌Thermomyceslanuginosus、嗜热子囊菌Thermoascus aurantiacus、嗜热革节孢Scytalidium thermophilum。此外,还分离到耐热真菌1种,鉴定为不规则头梗霉Cephaliophora irregularis,为中国新记录种。结论:这些研究结果新增了嗜热真菌在中国的分布记载,丰富了我国西南地区嗜热真菌的菌种资源库,另外对分离获得的嗜热真菌进行木聚糖酶活性测试,发现嗜热子囊菌为高产木聚糖酶活力的菌株。     

千层塔内生真菌的分离与鉴定

目的：为从千层塔中分离具有药用价值的内生真菌奠定基础。方法：新鲜千层塔茎段,经酒精和升汞消毒后,接种于PDA平板培养基上进行内生真菌的分离、纯化;根据菌落形态和孢子等形态特征,结合核糖体基因居间序列（ITS序列）进行菌株鉴定。结果：从千层塔的茎中分离出4株内生真菌。内生真菌I菌落形态和孢子特征与枝状枝孢霉属的特征相符合,ITS序列与GenBank中多条属于枝状枝孢霉的ITS序列相似,鉴定该菌株属于枝状枝孢霉;内生真菌II菌落形态和孢子特征与黄青霉的特征相符合,鉴定该菌株属于黄青霉;内生真菌III菌落形态和孢子特征与尖孢镰刀菌的特征相符合,ITS序列与GenBank中多条属于尖孢镰刀菌的ITS序列相似程度高,鉴定该菌株属于尖孢镰刀菌;内生真菌Ⅳ菌落形态与盾壳霉相似,ITS序列与GenBank中6条属于盾壳霉的ITS序列具有较高的相似性,鉴定该菌株属于盾壳霉。结论：从千层塔中分离和鉴定出4株内生真菌,分别属于枝状枝孢霉、黄青霉、尖孢镰刀茵和盾壳霉。     

中国南海部分深海沉积物真菌多样性及其抗菌活性

【目的】从南海4个站位的深海沉积物中分离真菌,揭示其多样性并测定抗菌活性。【方法】使用4种培养方法和8种培养基,从12个深海沉积物样本中分离培养真菌,通过菌落形态观察和ITS序列系统发育分析进行鉴定。采用滤纸片扩散法和生长速率法分别测试真菌小量发酵液粗浸膏的抗细菌和抗真菌活性。【结果】共分离到125株纯培养真菌,基于形态和ITS序列分析,排重后得到18个种类型,这些真菌可以划分到12个属,大多数属于子囊菌门(Ascomycota),只有2株属于担子菌门(Basidiomycota)。4个站位可培养真菌多样性具有差异性。抑菌活性筛选显示,大多数真菌具有较好的抑菌活性;链格孢属(Alternaria)、青霉属(Penicillium)、匐柄霉属(Stemphylium)这几个属的真菌表现出对多种指示细菌有抑制作用,尤其是Alternaria tenuissima DN09、Alternaria alternata DN14和Penicillium chrysogenum DN16对G~+和G~–细菌均表现出抑制作用。【结论】本研究揭示了南海深海沉积物可培养真菌多样性和抑菌活性,为进一步利用深海沉积物来源真菌奠定了基础。     

疏绵状嗜热丝孢菌的糖基化热稳定锰超氧化物歧化酶及真菌锰超氧化物歧化酶的系统学（英文）

疏绵状嗜热丝孢菌Thermomyces lanuginosus是一种广泛分布的嗜热真菌,产生的酶在高温条件下具有高活性和稳定性。从该嗜热真菌中克隆了一种锰超氧化物歧化酶(manganese superoxide dismutase,MnSOD)基因Tlmnsod1(EU364837),cDNA全长791bp,开放阅读框654bp,编码217个氨基酸,没有信号肽和前导肽序列。Tlmnsod1基因在酵母中高效表达,经纯化获得了电泳均一分子量约为27kDa的重组蛋白TlMnSOD1。表达的TlMnSOD1酶是糖基化蛋白,具高的热稳定性,最适反应pH和温度分别为8.0和55℃。通过对真菌的40种锰超氧化物歧化酶的细胞定位和系统学分析可知:真菌的锰超氧化物歧化酶可以分为胞内MnSOD、线粒体MnSOD和胞外MnSOD;锰超氧化物歧化酶可以作为分子标记来研究真菌的进化关系;3种嗜热真菌MnSOD分别与非嗜热真菌MnSOD聚类在3个不同的组中。本研究为MnSOD的糖基化和作为分子标记用于真菌种的鉴定提供了证据。     

天目山山胡椒不同部位内生真菌组成及多样性分析

采用表面消毒法,从浙江天目山野生山胡椒[ Lindera glauca( Sieb.et Zucc.) Blume]的茎、叶和树皮中分离出内生真菌,基于ITS序列分析进行分类鉴定;并以内生真菌的分离率、定殖率、分离频率、多样性指数(H')及相似性系数为指标,分析了山胡椒内生真菌的菌群组成及多样性.结果显示:在26株山胡椒样株的728块组织块中共分离得到328株内生真菌(茎、叶和树皮中分别有161、40和127株);共鉴定出44个分类单元(茎、叶和树皮中各有19、18和28个),其中25个分类单元鉴定到种、17个鉴定到属、2个鉴定到科,ITS序列的GenBank登录号从JF502420至JF502462.在44个分类单元中,有40个分类单元属于子囊菌(310株),存在于山胡椒的各个部位;仅有4个分类单元属于担子菌(18株),且仅存在于茎和树皮中.山胡椒茎、树皮和叶中内生真菌的定殖率分别为65％、60％和15％,分离率分别为0.77、0.61和0.19;叶和树皮中内生真菌的多样性指数均为2.63,远大于茎(H’=1.83).山胡椒内生真菌的优势属为Phomopsis、Paraconiothyrium、Phoma和Colletotrichum,大量存在于叶、茎和树皮中.山胡椒茎与树皮、茎与叶及叶与树皮间内生真菌的相似性系数分别为0.27、0.19和0.18,显示树皮和叶之间以及树皮和茎之间内生真菌的组成极不相似.研究结果表明:山胡椒体内存在大量的内生真菌,其茎、叶和树皮的内生真菌菌群组成具有一定程度的多样性和差异性,且内生真菌的分布具有组织特异性.     

三种红树林植物健康与非健康叶片中真菌的比较

红树林叶片感染真菌后可能会向非健康状态变化,为了比较三种红树林植物海漆、秋茄和桐花树健康与非健康叶片真菌类群,明确其中真菌类群的差异,该研究从广西茅尾海红树林自然保护区采集海漆、秋茄和桐花树健康与非健康叶片,对叶片中真菌分离纯化并进行形态学鉴定,提取真菌的DNA,采用RAPD多样性、ITS序列对真菌进行分子鉴定。经过初步分析,从海漆、秋茄和桐花树中共分离到157株真菌,经过形态学和RAPD分析,可能为19种不同的真菌。采用真菌ITS序列对19种真菌代表性菌株进行分析,结果表明:19种真菌都属于子囊菌门,有15株与已有的真菌ITS序列相似性在97%以上;有4株相似性低于95%,可能为新种。海漆健康和非健康叶片分别分离到真菌7种和5种,健康叶片含了非健康叶片中所有真菌;秋茄健康和非健康叶片分离到真菌1种和9种,且健康叶片中分离到的内生真菌Dothiorella aegiceri也包含于非健康叶片分离的真菌中;桐花树健康和非健康叶片分离到真菌1种和3种,且桐花树分离的内生真菌也是Dothiorella aegiceri,其包含于非健康叶片中分离到的真菌。该研究证明,海漆内生真菌同时也可能是植物病原菌,内生真菌与植物病原菌没有明显界限;另一方面,该研究发现秋茄和桐花树叶片的病变是由外部的病原真菌感染引起,感染的原因可能是昆虫叮咬红树林植物后留下了创口,这些创口有利于真菌的感染。     

产木聚糖酶嗜热真菌鉴定、酶基因克隆及生物信息学分析

木聚糖酶是一类木聚糖降解酶系,在食品、饲料领域应用广泛.本实验对实验室前期保存的一株产木聚糖酶真菌TL01进行形态学、18S rDNA及产酶鉴定,其结果表明TL01为梳棉状嗜热真菌(Thermomyces lanuginosus),以木糖为底物,测得粗酶液酶活为0.250±0.03 U/mL.对其两种主要的木聚糖酶基因(xyn11A和xyl43)进行克隆并对其编码蛋白(Xyn1 1A和Xy143)进行了生物信息学分析.Xyn11A预测蛋白分子量24.36 kD,等电点为4.77,含有19个氨基酸的信号肽序列,属于亲水性蛋白,无跨膜结构域,其二级结构主要是无规则卷曲(44.89％);Xyl43蛋白分子量38.24 kD,等电点为5.23,编码的胞内蛋白属于亲水性蛋白,无跨膜结构域,无规则卷曲(59.76％)是其主要的二级结构.本研究通过对Th.lanuginosus TL01的xyn11A和xyl43基因克隆及其编码蛋白的生物信息学分析,为后续进一步构建高效表达工程菌株奠定了基础.     

两种栽培型广藿香内生真菌群落组成变化

为探索内生真菌与广藿香互作间对宿主活性成分形成机制的影响,该研究以成分差异较大的牌香和湛香为对象,采用传统形态学方法对所获菌株归类,通过真菌通用引物ITS1/ITS4扩增菌株rDNA-ITS序列,鉴定其分类地位并研究其多样性。结果表明：(1)用PDA和LBA培养基对苗期、分枝期和成株期广藿香茎叶组织块进行内生真菌分离,共获得3 070株菌株,其中牌香(PX)分离出1 624株,鉴定出1 319株,分属于36属;湛香(ZX)分离出1 446株,鉴定出994株,分属于33属。牌香分离出7种特有内生真菌,分别为香柱菌(Epichloe typhina)、盘长孢状刺盘孢菌(Colletotrichum gloeosporioides)、座腔孢菌(Botryosphaeria sp.)、丝核菌(Rhizoctonia sp.)及截盘多毛孢菌(Truncatella sp.),并首次分离到疫霉菌(Phytophthora sp.)和指疫霉菌(Sclerophthora sp.),这2种菌属于卵菌门内生菌。湛香分离出拟青霉菌(Paecilomyces sp.)和尾孢菌(Cercospora sp.)...     

胃病患者胃黏膜真菌感染的检测

通过检测胃炎和胃溃疡、胃癌患者胃黏膜寄居的真菌,了解胃黏膜真菌的菌种多样性及其与胃溃疡的关系。采集消化科就诊患者胃镜钳取的胃黏膜标本63例,采用念珠菌显色培养基(CHROMagar)进行真菌分离培养鉴定。用玉米吐温80培养基进行真菌孢子形态学检查,用ITS(internal transcribed spacer region)序列限制性片段长度多态性(Restriction fragment length polymorphism,RFLP)检测分析真菌菌种多样性。分离培养真菌32(32/63,50.8%)株,经ITS序列RFLP法鉴定为白假丝酵母菌31株,光滑假丝酵母菌1株。真菌阳性率与病理诊断成正相关(r=0.263,P=0.027),与性别、年龄、吸烟、饮酒、学历的相关性均无统计学意义(P0.05)。结果表明,胃黏膜寄居的真菌存在多样性,且真菌阳性率与病理损害程度存在相关性。     

A Unique Protease Cleavage Site Predicted in the Spike Protein of the Novel Pneumonia Coronavirus (2019-nCoV) Potentially Related to Viral Transmissibility

正Dear Editor,In December 2019, a novel human coronavirus caused an epidemic of severe pneumonia(Coronavirus Disease 2019,COVID-19) in Wuhan, Hubei, China(Wu et al. 2020; Zhu et al. 2020). So far, this virus has spread to all areas of China and even to other countries. The epidemic has caused 67,102 confirmed infections with 1526 fatal cases     

Curcuminoids as inhibitors of thioredoxin reductase: A receptor based pharmacophore study with distance mapping of the active site

Curcumin is the yellow pigment of turmeric that interacts irreversibly forming an adduct with thioredoxin reductase (TrxR), an enzyme responsible for redox control of cell and defence against oxidative stress. Docking at both the active sites of TrxR was performed to compare the potency of three naturally occurring curcuminoids, namely curcumin, demethoxy curcumin and bis-demethoxy curcumin. Results show that active sites of TrxR occur at the junction of E and F chains. Volume and area of both cavities is predicted. It has been concluded by distance mapping of the most active conformations that Se atom of catalytic residue SeCYS498, is at a distance of 3.56 from C13 of demethoxy curcumin at the E chain active site, whereas C13 carbon atom forms adduct with Se atom of SeCys 498. We report that at least one methoxy group in curcuminoids is necessary for interation with catalytic residues of thioredoxin. Pharmacophore of both active sites of the TrxR receptor for curcumin and demethoxy curcumin molecules has been drawn and proposed for design and synthesis of most probable potent antiproliferative synthetic drugs.     

Comparative morphology of the carpel in the Liliaceae: Hewardieae, Petrosavieae, and Tricyrteae

The young pistils in the melanthioid tribes, Hewardieae, Petrosavieae and Tricyrteae, are uniformly tricarpellate and syncarpous. They lack raphide idioblasts. All are multiovulate, with bitegmic ovules. The Petrosavieae are marked by the presence of septal glands and incomplete syncarpy. Tepals and stamens adhere to the ovary in the Hewardieae and the Petrosavieae but not in the Tricyrteae. Two vascular bundles occur in the stamens of the Hewartlieae and Tricyrtis latifolia. Ventral bundles in the upper part of the ovary of the Hewardieae are continuous with compound septal bundles and placental bundles in the lower part. Putative ventral bundles occur in the alternate position in the Tricyrteae and putative placental bundles in the opposite. position in the Petrosavieae. The dichtomously branched stigma in each carpel of the Tricyrteae is supplied by a bifurcated dorsal bundle.     

Enterovirus 71 3C proteolytically processes the histone H3 N-terminal tail during infection

Highlights
1. The N-terminal tail of histone H3 is specifically cleaved during EV71 infection.
2. Viral protease 3C is identified as a protease responsible for proteolytically processing the N-terminal H3 tail.
3. Our finding reveals a new epigenetic regulatory mechanism for Enterovirus 71 in virus-host interactions.     

Detection of EBV and HHV6 in the Brain Tissue of Patients with Rasmussen’s Encephalitis

Rasmussen’s encephalitis (RE) is a rare pediatric neurological disorder, and the exact etiology is not clear. Viral infection may be involved in the pathogenesis of RE, but conflicting results have reported. In this study, we evaluated the expression of both Epstein-Barr virus (EBV) and human herpes virus (HHV) 6 antigens in brain sections from 30 patients with RE and 16 control individuals by immunohistochemistry. In the RE group, EBV and HHV6 antigens were detected in 56.7% (17/30) and 50% (15/30) of individuals, respectively. In contrast, no detectable EBV and HHV6 antigen expression was found in brain tissues of the control group. The co-expression of EBV and HHV6 was detected in 20.0% (6/30) of individuals. In particular, a 4-year-old boy had a typical clinical course, including a medical history of viral encephalitis, intractable epilepsy, and hemispheric atrophy. The co-expression of EBV and HHV6 was detected in neurons and astrocytes in the brain tissue, accompanied by a high frequency of CD8⁺ T cells. Our results suggest that EBV and HHV6 infection and the activation of CD8⁺ T cells are involved in the pathogenesis of RE.     

Perinatal Vertical Transmission of Chikungunya Virus in Ruili,a Town on the Border between China and Myanmar

正Dear Editor,Chikungunya virus (CHIKV), an arbovirus in the family of Togaviridae, genus Alphavirus, is transmitted by the A.aegyptii or A. albopictus mosquito, and causes disease in humans characterized by fever, rash, and arthralgia (Silva and Dermody 2017; Suhrbier 2019). It was first reported in 1953 in Tanzania, and caused only a few outbreaks and sporadic cases in Africa and Asia in last century. However, in the epidemic in 2004, CHIKV acquired mutations that conferred enhanced transmission by the A. albopictus mosquito(Schuffenecker et al. 2006). Since then, it has successively caused outbreaks in Africa, the Indian Ocean, South East Asia, the South America, and Europe (Zeller et al. 2016).     

Development of a Novel Reverse Transcription Loop-Mediated Isothermal Amplification Method for Rapid Detection of SARS-CoV-2

In conclusion, the novel visual RT-LAMP assay is a simple, rapid, and sensitive approach for detection of SARS-CoV-2, and it is ready for application in primary care and community hospitals or health care centers, and even patients' own houses in response to the current SARS-CoV-2 epidemic because the assay does not require sophisticated equipment and skilled personnel. Furthermore, it is also ready to be used in fields for screening samples from wild animals and environments to facilitate the identification of potential intermediate hosts that mediate the cross-species transmission of SARS-CoV-2 from bats to humans.