葡萄果粒表皮酵母菌多样性研究

从新疆、甘肃、陕西、宁夏、山东主要酿酒葡萄产区收集葡萄果粒并分离得到酵母258株,利用26S rDNA的D1/D2区序列分析并结合形态学、生理学特征对这些菌株进行了分类学研究,探讨了这些地区葡萄果粒表皮酵母的物种多样性及其分布.共鉴定出13属26种,其中优势属为汉逊酵母属Hanseniaspora(5种),假丝酵母属Candida(4种),毕赤酵母属Pichia(4种)和伊萨酵母属Issatchenkia(2种).对分离自不同地域的同种内不同菌株进行了D1/D2序列分析比较,以探讨不同地理起源地酵母种内序列稳定性及其变异.     

葡萄果粒表皮酵母菌多样性研究

从新疆、甘肃、陕西、宁夏、山东主要酿酒葡萄产区收集葡萄果粒并分离得到酵母258株, 利用26S rDNA的D1/D2区序列分析并结合形态学、生理学特征对这些菌株进行了分类学研究, 探讨了这些地区葡萄果粒表皮酵母的物种多样性及其分布。共鉴定出13属26种, 其中优势属为汉逊酵母属Hanseniaspora(5种), 假丝酵母属Candida(4种), 毕赤酵母属Pichia(4种)和伊萨酵母属Issatchenkia(2种)。对分离自不同地域的同种内不同菌株进行了D1/D2序列分析比较, 以探讨不同地理起源地酵母种内序列稳定性及其变异。     

红富士苹果园酵母多样性研究

从北京顺义和山东泰安红富士苹果园采集果实、叶片、树皮和土壤等不同基物,分离酵母菌,利用26S rDNA的D1/D2区域序列分析并结合形态学特征和SSCP分析对这些菌株进行了分类学研究,探讨了苹果园酵母的物种多样性及其分布。北京苹果园共分离酵母菌129株,鉴定为13属21种,优势属为Pichia（4个种）,Cryptococcus（3个种）,Pseudozyma（3个种）,子囊菌占较大优势,分布于8属12种,占总种数的57.1%。山东苹果园共分离酵母291株,鉴定为13属26种,优势属为假丝酵母Candida（6个种）,毕赤酵母Pichia（4个种）和隐球酵母Cryptococcus（3个种）,并且子囊菌占较大优势,分布于7属17种,占总种数的65.4%。     

假丝酵母属疑难菌株大亚基rDNA D1/D2区域序列分析及其分类学意义*

对根据常规形态和生理生化性状难以确定分类学地位的8株假丝酵母菌,进行了以大亚基(26S) rDNA中D1/D2区域(约500~600 bp)的碱基序列分析为依据的分子分类学研究。根据系统树上所显示的供试菌株与假丝酵母属及相关子囊菌酵母已知种的亲缘关系,以及与最近缘种模式菌株D1/D2区域序列的相似性比较,确定了各个菌株的归属。本研究也显示了DNA序列分析在假丝酵母菌快速鉴定中的优越性。     

假丝酵母属疑难菌株大亚基rDNA D1/D2区域序列分析及其分类学意义

对根据常规形态和生理生化性状难以确定分类学地位的8株假丝酵母菌,进行了以大亚基(26S) rDNA中D1/D2区域(约500～600 bp)的碱基序列分析为依据的分子分类学研究.根据系统树上所显示的供试菌株与假丝酵母属及相关子囊菌酵母已知种的亲缘关系,以及与最近缘种模式菌株D1/D2区域序列的相似性比较,确定了各个菌株的归属.本研究也显示了DNA序列分析在假丝酵母菌快速鉴定中的优越性.     

假丝酵母属疑难菌株大亚基rDNAD1／D2区域序列分析及其分类学意义

对根据常规形态和生理生化性状难以确定分类学地位的8株假丝酵母菌,进行了以大亚基（26S）rDNA中D1/D2区域（均500-600bp)的碱基序列分析为依据的分子分类学研究。根据系统树上所显示的供试菌株与假丝酵母属及相关子囊菌酵母已知种的亲缘关系,以及与最近缘种模式菌株D1/D2区域序列的相似性比较,确定了各个菌株的归属,本研究也显示了DNA序列分析在假丝酵母菌快速鉴定中的优越性。     

海南热带雨林腐木上两个无性型子囊菌酵母新种

用含木糖为唯一碳源和含葡萄糖及7.6%乙醇的两种富集培养基对采自海南热带雨林腐木样品中的酵母菌进行了分离培养。对分离出的酵母菌株进行的分子分类学研究表明,其中两株酵母菌X2WZ07-4和G2WZ06-1代表两个无性型子囊菌酵母新种。大亚基(26S)rRNA基因D1/D2域序列分析显示,与X2WZ07-4和G2WZ06-1亲缘关系最近的已知种分别为Candida cylindracea和C.llanquihuensis。在D1/D2域,X2WZ07-4与C.cylindracea模式菌株的碱基序列差异为2.5%;G2WZ06-1与C.llanquihuensis模式菌株的序列差异为3.9%,均远大于酵母菌种间在此区域的序列差异(～1%)。这两个新种分别被命名为拟柱形假丝酵母Candida pseudocylindracea sp.nov.(模式菌株:X2WZ07-4T=AS2.3788T=CBS10854T)和五指山假丝酵母Candida wuzhishanensis sp.nov.(模式菌株:G2WZ06-1T=AS2.3784T=CBS10850T)。     

石河子不同树龄蟠桃土壤中可培养酵母多样性分析及功能酵母的筛选

为了探究新疆本土蟠桃园可培养酵母菌多样性,并挖掘功能酵母资源,本研究以新疆石河子蟠桃园3年、8年、15年树龄的根际和非根际土壤以及桃树叶片为材料,经过传统的分离培养方法获得可培养的酵母菌菌株,并进行形态学、生理生化以及26S r DNA的D1/D2区序列分析,共获得可培养酵母菌129株,从属于12个属17个种,其中子囊菌酵母为优势菌群,占分离属的88%,分布于威克汉姆酵母属(Wickerhamomyces),Vanrija属,Barnettozyma属和有孢圆酵母属(Torulaspora)等11个属的15个种。担子菌占分离属的12%,分布于隐球酵母属(Cryptococcus)的2个种。其中优势属威克汉姆酵母属,包括异常威克汉姆酵母(W. anomalus)和W. pijperi两个种,占总比例的33%,优势种异常威克汉姆酵母所占总株数比例为17%。从可培养酵母中共筛选出23株功能酵母,其中富硒酵母21株,优势种为白地霉(Galactomyces candidum),产蛋白酶酵母2株均属于隐球酵母属的Cryptococcus albidus。结果表明,新疆桃园中蕴含丰富的酵母菌资源,非根际土壤中的酵母多样性大于根际及叶片酵母多样性,且分离得到富硒酵母及产蛋白酶酵母。本研究挖掘了新疆本土可培养酵母菌资源,同时也为功能酵母的开发和利用提供理论指导。     

青海东部土壤中酵母物种多样性研究

从青海的互助、民和、门源等10个州县收集土样分离得到98株酵母菌, 利用26S rDNA D1/D2区域序列分析并结合形态学和生理生化特性对这些菌株进行了分类学研究, 探讨了青海东部土壤中酵母的物种多样性及其分布。共鉴定出10属13种(其中有两个疑似新种), 其中 Galactomyces geotrichum和Rhodotorula mucilaginosa为该地的优势种。     

用传统分离培养法和高通量测序技术分析印度块菌子囊果内细菌的群落结构

块菌是重要的经济真菌, 在其生长发育过程中, 细菌扮演了重要角色。本文利用传统分离培养方法和高通量测序技术分析了印度块菌(Tuber indicum)子囊果内细菌的群落结构。共分离得到细菌532株, 根据物种累积曲线, 选取其中的112株细菌进行了16S rRNA基因序列分析, 共鉴定出4属40种, 其中假单胞菌属(Pseudomonas)菌株占所测菌株数的80%, 不动杆菌属(Acinetobacter)占12.5%, 链霉菌属(Streptomyces)占5%, 贪噬菌属(Variovorax)占2.5%。通过对印度块菌子囊果16S rRNA基因的V1-V3区高通量测序分析, 共获得细菌序列9,862条, 分属于7门43属220种, 其中变形菌门、拟杆菌门和放线菌门的物种占总物种数的99.7%, 是印度块菌子囊果内的优势细菌。黄杆菌属(Flavobacterium)、壤霉菌属(Agromyces)、微杆菌属(Microbacterium)、剑菌属(Ensifer)和寡养食单胞菌属(Stenotrophomonas)的物种数占总物种的86.3%, 是印度块菌子囊果内细菌的优势属。研究结果表明, 采用胰蛋白大豆培养基仅分离得到印度块菌子囊果内少数细菌物种, 而采用高通量测序技术分析发现, 印度块菌子囊果内细菌物种种类丰富, 群落结构复杂。     

Yeast biodiversity in hydromorphic soils with reference to grass-Sphagnum swamp in Western Siberia and the hammocky tundra region( Barrow, Alaska)]

The microbiological analysis of 78 samples taken from a boreal bog in Western Siberia and from a tundra wetland soil in Alaska showed the presence of 23 yeast species belonging to the genera Bullera, Candida, Cryptococcus, Debaryomyces, Hanseniaspora, Metschnikowia, Mrakia, Pichia, Rhodotorula, Saccharomyces, Sporobolomyces, Torulaspora, and Trichosporon. Peat samples from the boreal bog were dominated by eurytopic anamorphic basidiomycetous species, such as Rhodotorula mucilaginosa and Sporobolomyces roseus, and by the ascomycetous yeasts Candida spp. and Debaryomyces hansenii. These samples also contained two rare ascomycetous species (Candida paludigena and Schizoblastosporion starkeyi-henricii), which so far have been found only in taiga wetland soils. The wetland Alaskan soil was dominated by one yeast species (Cryptococcus gilvescens), which is a typical inhabitant of tundra soils. Therefore, geographic factors may serve for a more reliable prediction of yeast diversity in soils than the physicochemical or ecotopic parameters of these soils.     

Three new ascomycetous yeasts from insect-associated arboreal habitats

A new species of Pichia and two new species of Candida are described and were determined to be genetically isolated from all other currently recognized ascomycetous yeasts from their sequence divergence in the species-variable D1/D2 domain of large subunit (26S) ribosomal DNA. The three species were primarily isolated from the frass of wood-boring insects living in pine and spruce trees. The new species and their type strains are the following: Pichia ramenticola NRRL YB-1985 (CBS 8699), mating type alpha (NRRL YB-3835, CBS 8700, mating type a), Candida piceae NRRL YB-2107 (CBS 8701), and Candida wyomingensis NRRL YB-2152 (CBS 8703). Pichia ramenticola and C. piceae assimilate methanol as a carbon source; P. ramenticola is the first known heterothallic ascomycetous yeast to utilize this compound.     

田间施药对自然发酵葡萄酒酵母菌群落结构的影响

【背景】酵母菌是葡萄酒发酵过程中一类非常重要的微生物,其多样性及群体组成对葡萄酒的质量有重要贡献。影响葡萄酒中酵母菌组成的因素有很多,但目前尚未见葡萄园田管理对葡萄酒酵母菌群落结构影响方面的报道。【目的】探索田间施药对自然发酵葡萄酒酵母菌群落结构的影响。【方法】采用分离培养、常规分子生物学鉴定和Illumina MiSeq宏基因组测序结合的方法分析不同样品中的酵母菌群落结构情况。【结果】从不使用内吸收型化学农药的葡萄样品自然发酵液中分离鉴定出Pichia、Hanseniaspora、Schizosaccharomyces、Candida、Saccharomyces、Zygoascus、Issatchenkia等7个属8个种的酵母菌,宏基因组测序结果表明有Pichia(29.42%)、Saccharomyces(21.91%)、Issatchenkia(17.99%)、 Hanseniaspora(12.10%)、 Candida(7.47%)、 Zygosaccharomyces(5.32%)、Schizosaccharomyces (3.07%)、Aureobasidium (0.29%)等属的酵母菌参与发酵;使用常规化学农药的葡萄样品自然发酵液中分离鉴定出Pichia、Hanseniaspora、Schizosaccharomyces、Candida、Cryptococcus等5个属6个种的酵母菌,宏基因组测序结果表明有Pichia (41.66%)、Hanseniaspora (21.54%)、Candida(19.11%)、 Zygosaccharomyces(7.78%)、 Schizosaccharomyces(4.04%)、 Cryptococcus(3.21%)、Saccharomyces (1.12%)、Aureobasidium (0.49%)等属的酵母菌参与发酵。【结论】两样品中酵母菌比例有显著差异,表明在酿酒葡萄的园田管理中化学农药的使用对自然发酵葡萄酒的酵母菌群落结构有较大影响。     

西藏曲拉和云南乳饼中酵母菌的鉴定及其生物多样性

【目的】探讨西藏曲拉和云南乳饼中酵母菌的生物多样性及其分布特征,为我国传统乳制品中酵母菌资源的利用提供基础数据。【方法】从西藏和云南分别采集的5份曲拉样品和8份乳饼样品中分离出41株酵母菌,利用26SrDNAD1/D2区域序列分析对这些菌株进行了分类鉴定。【结果】曲拉和乳饼样品中酵母菌的总数分别在106-107cfu/g和102-106cfu/g之间,曲拉样品的酵母菌平均数比乳饼样品中的高34倍。共鉴定出10属12种,其中西藏曲拉的优势菌株为发酵毕赤氏酵母(Pichia fermentans)和酿酒酵母(Saccharomyces cerevisiae);云南乳饼的优势菌株为类筒假丝酵母(Candida zeylanoides)和喜仙人掌毕赤氏酵母(Pichia cactophila)。毕赤氏酵母属(Pichia)是曲拉和乳饼的共同优势属。【结论】西藏曲拉和云南乳饼中的酵母菌都具有丰富的生物多样性,但其差异性很大。     

Diversity and evolution of non-Saccharomyces yeast populations during wine fermentation: effect of grape ripeness and cold maceration

We have evaluated the effect of grape maturity and cold maceration prior to fermentation on the yeast ecology during wine fermentation. Non-Saccharomyces strains were selectively isolated and identified using two rapid PCR techniques, namely enterobacterial repetitve intergenic consensus-PCR and PCR-intron splice sites, in various wine fermentation conditions. These identifications were further complemented and confirmed by restriction fragment length poymorphism and sequencing analysis of the 5.8S-ITS and D1/D2 ribosomal regions, respectively. Eleven species belonging to five genera were identified. Candida stellata, Hanseniaspora uvarum and Hanseniaspora osmophila were the dominant species, representing almost 90% of the isolates. Minor strains presented different species of the genera Candida, Issatchenkia, Zygoascus and Zygosaccharomyces. Selective isolation made it possible to isolate some species that were hardly related to the wine-making process, such as Issatchenkia hanoiensis, a new species that has only been described recently.     

Yeast species associated with orange juice: evaluation of different identification methods

Five different methods were used to identify yeast isolates from a variety of citrus juice sources. A total of 99 strains, including reference strains, were identified using a partial sequence of the 26S rRNA gene, restriction pattern analysis of the internal transcribed spacer region (5.8S-ITS), classical methodology, the RapID Yeast Plus system, and API 20C AUX. Twenty-three different species were identified representing 11 different genera. Distribution of the species was considerably different depending on the type of sample. Fourteen different species were identified from pasteurized single-strength orange juice that had been contaminated after pasteurization (PSOJ), while only six species were isolated from fresh-squeezed, unpasteurized orange juice (FSOJ). Among PSOJ isolates, Candida intermedia and Candida parapsilosis were the predominant species. Hanseniaspora occidentalis and Hanseniaspora uvarum represented up to 73% of total FSOJ isolates. Partial sequence of the 26S rRNA gene yielded the best results in terms of correct identification, followed by classical techniques and 5.8S-ITS analysis. The commercial identification kits RapID Yeast Plus system and API 20C AUX were able to correctly identify only 35 and 13% of the isolates, respectively. Six new 5.8S-ITS profiles were described, corresponding to Clavispora lusitaniae, Geotrichum citri-aurantii, H. occidentalis, H. vineae, Pichia fermentans, and Saccharomycopsis crataegensis. With the addition of these new profiles to the existing database, the use of 5.8S-ITS sequence became the best tool for rapid and accurate identification of yeast isolates from orange juice.     

Molecular assessment of indigenous yeast population from traditional balsamic vinegar

AIMS: To identify and describe the indigenous yeast population involved in traditional balsamic vinegar (TBV) fermentation. METHODS AND RESULTS: Using the restriction analysis of the ribosomal region 5.8S (5.8S rRNA) and the internal transcribed spacers 1 and 2 (5.8S-ITS region) we were able to group 133 strains isolated from 17 cooked grape must samples into 10 different yeast species, included into 4 genera. Moreover, we sequenced the D1/D2 domains of the 26S rRNA and confirmed the reliability of each identification at species level. Most strains belonged to the genus Zygosaccharomyces. In particular, Zygosaccharomyces bailii was found in 41% of the samples, followed by Saccharomyces cerevisiae, Zygosaccharomyces pseudorouxii and Candida stellata. Strains belonging respectively to Zygosaccharomyces mellis, Zygosaccharomyces bisporus, Zygosaccharomyces rouxii, Hanseniaspora valbyensis, Hanseniaspora osmophila and Candida lactis-condensi species were also detected. Despite the great number of species recovered, the mtDNA restriction profiles showed low variability at strain level. Saccharomyces cerevisiae isolates with an higher degree of intraspecific variance were considered an exception. CONCLUSIONS: Many different indigenous yeast species were recovered and TBV yeasts population seems to be far more complex than what was reported in previous literature. SIGNIFICANCE AND IMPACT OF THE STUDY: This study has allowed us to gain a better understanding of the indigenous yeast species of TBV cooked must.