为分析中国本土野生有孢汉逊酵母属Hanseniaspora酵母菌的种间差异和种内差异,构建分子指纹图谱,为优良有孢汉逊酵母资源的遗传多样性分析及开发利用提供科学依据.采用依赖于培养的经典方法包含WL营养培养基、26S rRNA基因D1/D2区域序列分析和5.8S-ITS-RFLP分析进行118株贵州紫云县刺葡萄自然发酵过程中分离的有孢汉逊酵母属酵母菌种水平的鉴定.采用Tandem repeat-tRNA (TRtRNA)指纹图谱法研究有孢汉逊酵母属酵母在两对微卫星引物扩增下的分子指纹图谱,并利用DPS软件分析不同分子指纹图谱类别之间的遗传发育关系.结果 表明,118株贵州紫云县刺葡萄自然发酵过程中分离的野生有孢汉逊酵母属酵母菌,经依赖于培养的经典方法鉴定为3个种,包括Hanseniaspora opuntiae、H.uvarum和H.occidentalis.TRtRNA指纹图谱法的引物对一TtRNASC和5CAG扩增出5类图谱,而引物对二TtRNASC和ISSR-MB扩增出10类图谱,可以表现出所测118株有孢汉逊酵母属的种间差异和种内差异.其中种内差异主要表现在H.uvarum中.总体而言,贵州紫云县刺葡萄自然发酵过程中分离的有孢汉逊酵母属主要为H.opuntiae、H.uvarum和H.occidentalis,其中H.opuntiae和H.uvarum内部表现了不同的TRtRNA分子指纹图谱,表征了种内遗传多样性.

The oriental fruit fly,Bactrocera dorsalis(Diptera:Tephritidae)is a destructive horticultural pest which causes considerable economic losses every year.A collection of microorganisms live within the B.dorsalis gut,and they are involved in its development,physiology,and behavior.However,knowledge regarding the composition and function of the gut mycobiota in B.dorsalis are still limited.Here,we comprehensively characterized the gut mycobiota in B.dorsalis across different developmental stages.High-throughput sequencing results showed a significant difference in fungal species abundance and diver-sity among different developmental stages of B.dorsalis.Quantitative polymerase chain reaction and culture-dependent methods showed that yeast species was the dominant group in the larval stage.We isolated 13 strains of yeast from the larval gut,and found that GF(germ-free)larvae mono-associated with strain Hanseniaspora uvarum developed faster than those mono-associated with other tested fungal strains.Supplementing the larval diet with H.uvarum fully rescued B.dorsalis development,shortened the larval developmental time,and increased adult wing lengths,as well as the body sizes and weights of both pupae and adults.Thus,our study highlights the close interactions between gut fungi,especially H.uvarum,and B.dorsalis.These findings can be applied to the sterile insect technique program to promote host development during mass insect rearing.

[背景]NX11424酵母是一株具有良好发酵特性且能赋予赤霞珠葡萄酒浓郁果香的宁夏本土酿酒酵母.[目的]解析NX11424酵母发酵的赤霞珠葡萄酒中的水果香气特征.[方法]以赤霞珠为材料,设置3个发酵处理:自然发酵、灭菌接种NX11424的发酵和直接接种NX11424的发酵,利用26SrDNAD1/D2区测序分析法鉴定发酵过程中酵母菌的种类,并通过顶空固相微萃取和气相色谱-质谱联用技术定量测定不同发酵处理下赤霞珠葡萄酒的香气成分及含量.[结果]三个发酵处理下赤霞珠葡萄酒各理化指标无显著性差异.所分离到的酵母菌鉴定为2属 3种:萄葡汁有孢汉逊酵母(Hanseniaspora uvarum)、酿酒酵母(Saccharomyces cerevisiae)和布拉迪酵母(Saccharomyces boulardii,S.boulardii);这2属3种均存在于自然发酵中,接种发酵中仅存在H.uvarum和S.cerevisiae两种酵母.三个发酵处理下的赤霞珠葡萄酒中香气物质种类无差异,均为69种;其中,酯类28种,醇类25种,有机酸5种,萜烯类2种和其他类化合物9种;但各发酵处理下香气物质的含量存在显著差异.聚类分析表明,69种香气成分被聚为3类.第1类香气物质包括香茅醇、丙醇等9种成分,其中7种香气物质的含量在灭菌接种发酵中较高;第2类香气物质包括己酸乙酯、棕榈酸乙酯等31种成分,其含量均在直接接种发酵中较高;第3类香气物质包括丁酸乙酯、乳酸乙酯等29种成分,其中27种香气物质的含量在自然发酵中较高.虽然直接接种发酵处理中酵母菌的多样性低于自然发酵处理,但是该处理的赤霞珠葡萄酒中酯类香气物质含量较多,水果香气浓郁,对赤霞珠葡萄酒香气的改善更明显.[结论]NX11424与发酵中的其他本土酵母间的相互作用可以改善葡萄酒的质量,为宁夏本土酵母NX11424在赤霞珠葡萄酒酿造过程中改善葡萄酒质量奠定了坚实的基础.