源自不同寄主的灰葡萄孢生物学特性的比较研究

本研究以分离自番茄、辣椒、草莓、葡萄的灰葡萄孢Botrytis cinerea为供试菌株,从生长温度、pH适应性、碳源、氮源营养利用等方面对不同寄主来源的灰葡萄孢菌株的生物学性状进行了比较研究。结果表明,5个不同寄主来源的灰葡萄孢菌株的菌丝生长温度范围相同,均为0-35℃;但它们的最适生长温度和分生孢子致死温度存在差异,来自和县番茄菌株HX12最适生长温度为20℃,分生孢子致死温度为47℃ 10min,其余最适生长温度均为25℃,分生孢子致死温度均为48℃ 10min;不同菌株在相同温度下的生长速率有显著差异。pH对不同寄主来源灰葡萄孢菌株菌丝生长的影响存在差异,来自长丰辣椒的菌株LJ菌丝在pH2-9的范围内均能生长,以在pH3-6.5时生长较快,pH6时最快;其余4个菌株在pH2-12的范围内均能生长,以在pH3-9时生长较快,pH6左右最快。不同碳源、氮源营养对灰葡萄孢菌株菌丝生长和分生孢子产生均有显著影响,不同寄主来源的菌株间在碳源、氮源营养利用差异均极显著。在相同碳源、氮源营养条件下,不同寄主来源的菌株的线性生长、菌丝干重和分生孢子产量均有显著差异。     

蛹虫草子囊孢子萌发及其后代群体培养性状观察

从野生和人工栽培的蛹虫草子实体共分离了6个子囊孢子群体,对子囊孢子形态、萌发过程、培养性状及子实体产生等进行了观察。结果表明,蛹虫草子囊孢子为线形、多细胞,169.78~364.57×1.72~2.04μm;每个子囊孢子有56~114个细胞,每个细胞大小为1.77~4.53×1.72~2.04μm。子囊孢子弹射后13h开始萌发,30h后开始形成分生孢子。多数子囊孢子的每个细胞都能萌发,但少数子囊孢子的部分细胞不萌发。子囊孢子群体的培养性状表现多样性,以I型所占比列较大。野生群体的菌落颜色以杏橙色和淡柠檬黄色为主,而人工栽培的群体以橙铬色所占比例最大。不同个体的菌落生长速率有差异。野生群体产生扇形突变的比例高于人工栽培群体。在人工栽培的CM群体中出现了气生菌丝较多、白色、生长缓慢的个体。菌落为I型,橙铬色或杏橙色,生长速率正常,无突变的单子囊孢子菌株产生子实体的可能性较大,子实体产量较高,但大部分单子囊孢子菌株不能产生发育良好的子实体。     

番茄灰霉病拮抗内生放线菌的筛选、鉴定及其活性评价

对安徽省淮北市番茄植株根、茎、叶中内生放线菌进行了分离、筛选,并测定了其抑菌活性。结果表明:番茄根、茎和叶中的内生放线菌的数量分别为5.66×104、0.67×104和0.39×104CFU.g-1鲜重。根据分离部位和表型特征,从健康番茄植株体内分离到93株内生放线菌,通过对峙实验,筛选到7株对番茄灰霉病菌有拮抗作用的菌株,占所分离内生放线菌总数的7.5%。来自根组织中的菌株HNU-EA27的抑菌效果最佳,抑菌圈直径达28.3mm。根据形态特征、培养特征、生理生化特性、细胞壁组分和16SrDNA序列分析,将菌株HNU-EA27鉴定为毒三素链霉菌(Streptomyces toxytricini)。室内测定菌株HNU-EA27发酵滤液对灰霉病菌菌丝生长及分生孢子萌发的抑制作用,结果表明:菌株HNU-EA27发酵滤液可以抑制灰霉病菌菌丝生长和分生孢子萌发,且浓度越高,抑制能力越强;当发酵滤液浓度为30%时则完全抑制灰霉病菌菌丝生长和分生孢子萌发。盆栽防效试验结果表明:30%菌株HNU-EA27发酵滤液对番茄灰霉病的预防与治疗效果分别为80.6%和73.8%,均高于50%多菌灵可湿性粉剂600倍液。本研究表明,菌株HNU-EA27是防治番茄灰霉病潜在的优良生防菌株,具有良好的开发应用价值。     

不同种质资源蛹虫草菌丝生长特性及其多糖产量的研究

研究本试验中6株不同来源的蛹虫草菌株(150707,150715,150725,150808,52244,51762)的菌丝生长特性及其多糖产量。测定这6株蛹虫草菌丝生长速度、分生孢子产量,利用水提醇沉法提取不同菌株粗多糖。研究发现在不同培养基上不同来源的这6株蛹虫草菌丝生长速度、产分生孢子量、多糖得率不同,在加富PDA培养基上生长速度较快,但是在血平板培养基上孢子产量及多糖得率较优。分析得出150707、52244两个菌株孢子产量和多糖产量相对较高,为优势菌株,血平板培养基为蛹虫草孢子及多糖产量较优培养基。     

培养基及培养条件对冬虫夏草菌固体发酵产分生孢子的影响

为获得冬虫夏草菌固体发酵产分生孢子的最优工艺,以野生分离的冬虫夏草菌为材料,对其固体发酵产分生孢子的培养基及培养条件进行了研究。试验结果表明：泥炭土为最佳基础培养基,该培养基中冬虫夏草菌气生菌丝生长一般,但产分生孢子最多,可达4.2×10³个/g;泥炭土培养基中添加0.1‰ IAA（吲哚乙酸）、0.1‰ IBA（吲哚丁酸）和0.1‰ NAA（萘乙酸）能促进冬虫夏草菌气生菌丝的生长和分生孢子的产生,其分生孢子达8.1×10³个/g;该基础培养基中,冬虫夏草菌于18℃培养30d后,在10℃、相对湿度45%、蓝光照射进行诱导,分生孢子可达1.0×10⁴个/g。本研究建立了一种大量获取冬虫夏草菌分生孢子的方法,为冬虫夏草繁育奠定了基础。     

从天然蛹虫草不同部位分离菌株特性比较

以采集自泰山地区的6株野生蛹虫草为研究对象,分别通过菌核、子座组织分离和子囊孢子分离法共获得18个菌株,经ITS鉴定均为蛹虫草菌株后,进一步对18个菌株进行了菌丝生长速度、菌落形态、出草试验、主要活性物质（虫草素、腺苷、虫草多糖、β-类胡萝卜素、叶黄素）和交配型的测定分析。结果表明：在菌丝长势、出草产量和整齐度方面,从子囊孢子分离得到的菌株普遍优于组织分离菌株;而在活性物质含量方面,从子囊孢子和子座分离得到的菌株普遍优于菌核分离菌株。通过交配型测定发现,从菌核、子座和子囊孢子分离得到的18个菌株均不存在交配型缺失现象。     

番茄灰霉病拮抗内生细菌的筛选、鉴定及其活性

对安徽省淮北市番茄植株根、茎、叶中内生细菌及其数量进行了调查和筛选,并测定了其抑菌活性。结果表明,番茄根、茎和叶中的内生细菌的数量分别为5.69×105、5.16×105和2.83×105CFU.g-1鲜重。根据分离部位和表型特征,从健康番茄植株体内分离到267株内生细菌,通过对峙实验,筛选到11株对番茄灰霉病菌有拮抗作用的菌株,占所分离内生细菌总数的4.12%。来自茎组织中的菌株XF136的抑菌效果最佳,抑菌带宽度达32.2mm。根据形态特征、生理生化特性、(G+C)mol%和16SrDNA序列分析,将菌株XF136鉴定为解淀粉芽孢杆菌(Bacillus amyloliquefaciens)。室内测定菌株XF136发酵滤液对灰霉病菌菌丝生长及分生孢子萌发的抑制作用,结果表明,菌株XF136发酵滤液可以抑制灰霉病菌菌丝生长和分生孢子萌发,且浓度越高,抑制能力越强;当发酵滤液浓度为20%时则完全抑制灰霉病菌菌丝生长和分生孢子萌发。盆栽防效试验结果表明,10%菌株XF136发酵滤液对番茄灰霉病防治效果与50%多菌灵600倍液相当,20%发酵滤液对番茄灰霉病的防治效果高于50%多菌灵600倍液。本研究表明,菌株XF136是防治番茄灰霉病潜在的优良生防菌株,具有良好的开发应用价值。     

温度和水活度对冠突散囊菌菌丝生长和产孢的影响

冠突散囊菌(Eurotium cristatum)是茯砖茶发酵过程中的优势菌。通过调节甘油含量制备具有不同水活度(aw)的培养基,研究培养基水活度、培养温度以及不同温度和水活度组合对冠突散囊菌菌丝生长、分生孢子产生、萌发和菌落形态的影响。供试的冠突散囊菌菌株FZ-2、FZ-3和FZ-4在水活度为0.77～0.99的培养基中均能生长,最适培养基水活度为0.90～0.95。培养基水活度在0.77～0.95范围内有利于菌株FZ-2产生分生孢子,最适水活度为0.83;培养基水活度于0.83～0.95范围内有利于菌株FZ-4产孢;而菌株FZ-3适合产孢的培养基水活度为0.95。菌株FZ-2分生孢子萌发的最适水活度为0.87～0.95。菌株FZ-2适合的生长温度为28～37℃,温度38℃时菌体生长速度下降,至40～45℃时菌丝停止生长;菌株FZ-3和FZ-4最适生长温度均为28℃。在20～37℃条件下菌株FZ-2均能产生分生孢子,最适产孢温度为37℃;菌株FZ-3在28～37℃范围内产生分生孢子;菌株FZ-4在20～37℃内产生分生孢子,最适产孢温度为37℃。菌株FZ-2分生孢子可以在较宽的温度范围内(20～40℃)萌发,最适温度为25～33℃。研究结果表明不同水活度-温度组合对菌株FZ-2菌丝生长和菌落形态产生显著的影响,与温度作用相比,水活度的影响更为明显。本研究证明了冠突散囊菌适合在干燥的茶叶中生长和繁殖,为解释其在茯砖茶加工过程中的优势地位提供理论依据,研究结果也将促进茶叶发酵菌剂的开发利用和砖茶的标准化生产。     

圆盘菌科无性型有性生殖能力评估

本文评估了分离自子囊孢子和分生孢子的圆盘菌科无性型有性生殖能力。结果表明,由子囊孢子和分生孢子萌发获得的菌株的有性生殖能力具有显著差异,这可能决定于不同的遗传特性。该试验支持了一种假说：仅能进行无性生殖的无性型菌株,很可能来源于逐渐丧失了有性生殖能力的全型菌株。     

围小丛壳Glomerella cingulata子囊孢子交配繁殖、生物学特性及致病性

炭疽叶枯病（Glomerella leaf spot）是我国苹果上新发现的一种病害。为了解围小丛壳Glomerella cingulata子囊孢子的交配方式、生物学特性和致病性,从安徽砀山、山东牟平等地采集病害样品,经分离培养和纯化获得单孢菌株。在适宜条件下单孢菌株可产生子囊和子囊孢子,经过毛细管破子囊壁后单孢分离,获得12个子囊,每个子囊有8个子囊孢子。其中10个子囊中有4个“正”孢子（+）和4个“负”孢子（-）,2个子囊中只有“负”孢子。子囊孢子单孢菌株培养72h,“正”菌株菌落白色,以营养生长为主;“负”菌株菌落灰白色,直径略小于正菌株,菌丝稀疏,边缘菌丝白色,中部有大量橙色的分生孢子堆。“正”、“负”菌株异宗配合后,可产生大量可育子囊壳;单独的“正”菌株有性生殖产生稀疏丛簇状的可育子囊壳;单个的“负”菌株只能产生分散且不育的子囊壳。“正”、“负”菌株菌落的生长速度没有差异,对温度、营养、光照和pH值的敏感性也没有差异,但“正”、“负”菌株的致病性存在差异。正菌株的有性生殖没有导致rDNA-ITS、β-tubulin基因碱基序列变异。     

两种不同来源的绿色木霉降解木质纤维素研究

利用纤维素酶高产菌绿色木霉Trichoderma viride降解木质纤维素是实现废料资源化的重要手段。本研究选取来自不同生境的两株T. viride,分别以玉米秸秆和甘草药渣为基质,测定两者滤纸纤维素酶（filter paper cellulase,FPase）活性和还原糖产量。从时间、温度、水分、pH 4个方面比较两株T. viride的环境适应性和不同基质的差异性。结果表明,以玉米秸秆为基质,T. viride XJ最适初始料液比为1:4-1:5.5,T. viride AG最适初始料液比为1:5-1:5.5。初始料液比1:5.5时,T. viride AG产FPase活性显著高于T. viride XJ。两株T. viride最适发酵温度均为28℃,各温度处理下不同菌株间无显著差异。两株T. viride均表现为还原糖消耗。以甘草药渣为基质,T. viride XJ最适初始料液比为1:2-1:2.5,T. viride AG最适初始料液比为1:3-1:3.5。料液比高于1:3,T. viride AG产FPase活性显著高于T. viride XJ。T. viride AG最适发酵温度为28℃,T. viride XJ最适发酵温度为23-28℃。温度低于28℃,T. viride XJ产FPase活性显著高于T. viride AG。两株T. viride均表现为还原糖积累。两株T. viride最适初始pH均为6-7,最适发酵时间均为3d。最优发酵条件下FPase活性：T. viride AG>T. viride XJ。对T. viride产FPase诱导能力：甘草药渣>玉米秸秆。变差分解表明两株T. viride产FPase活性差异主要源于菌株对生境的生态适应。比较分析菌种来源、基质类型、环境条件对T. viride发酵效果的影响,将有助于该菌大规模应用性研究。     

Plasmid contents of lactic acid bacteria isolated from different types of sour doughs

The plasmid contents of 13 lactic acid bacteria isolated from different types of sour doughs were examined and compared with the plasmid contents of 11 culture collection strains and one commercial pure starter culture for sour doughs. In addition, plasmid analysis was used as a tool to study the stability of a starter culture during sour dough fermentation in a bakery.The tested strains varied in plasmid content from no plasmid up to six plasmids, with molecular weights from 1.5 to 43 MDal. In most cases, the wild-type strains contained a higher number of plasmids than the culture collection strains. The ability of the strains to ferment different carbohydrates was also investigated, but no obvious correlations between the fermentation patterns and the plasmid patterns could be observed. During the fermentation of the bakery sour dough, strains other than the inoculated starter culture gradually became dominant in the microflora. These new strains contained 1–3 plasmids, contrary to the plasmidless starter culture, and they also fermented more carbohydrates than the starter culture.     

Tempe fermentation: some aspects of formation of gamma-linolenic acid, proteases and vitamins

During a tempe fermentation the concentrations of linoleic, and alpha-linolenic acids (ALA) decreased while the concentration of oleic acid increased. During fatty acid synthesis Rhizopus sp. produced only gamma-linolenic acid (GLA) instead of ALA. The amount of GLA in tempe were influenced by varying external parameters.The proteolytic capacity of 36 strains of the genus Rhizopus isolated from Indonesian tempe or tempe inocula was examined. There was a distinct increase in the amount of free amino acids during tempe fermentation. Fermentations with mixed populations of bacteria and Rhizopus yielded a lower level of free amino acids, but an increase in total amount of amino acids. In comparison to intracellular, and extracellular proteases the proteases of the cell wall fraction are most responsible for proteolytic capacity of the different Rhizopus strains.Two isolated strains of Citrobacter freundii were found to be the best vitamin B(12) producers during the soaking of soybeans. In the solid substrate fermentation the Rhizopus molds formed vitamin B(6), riboflavin, and nicotinic acid. The addition of bacteria to the solid substrate fermentation resulted in a strong increase of active vitamin B(12) in tempe. In the presence of the Rhizopus mold, the vitamin B(12) formation by C. freundii was three times higher than that of a fermentation without the mold.     

Multiple strains of Saccharomyces cerevisiae on a single grape vine

M. POLSINELLI, P. ROMANO, G. SUZZI AND R. MORTIMER. 1996. On the basis of the levels of secondary product formation four different phenotypes were represented among the 28 strains of Saccharomyces cerevisiae isolated during the spontaneous fermentation of grape juice. The genetic analysis indicated that four different strains, representing each phenotypic class, were derived, one from the other, by mutation. The spontaneous fermentation of a Malvasia must was dominated by different strains of Saccharomyces cerevisiae at different stages of fermentation.     

Ecology of lactic acid bacteria in Italian fermented sausages: isolation, identification and molecular characterization

In this study, the ecology of the lactic acid bacteria (LAB) of three naturally fermented sausages produced in the Friuli-Venezia-Giulia region, in the North East of Italy, was investigated. A total of 465 strains isolated from three fermentations were identified by molecular methods and 12 different species of LAB were detected. Lactobacillus curvatus and Lactobacillus sakei were the most numerous (67 and 353 strains isolated, respectively) and they were subjected to RAPD-PCR. Clusters containing strains isolated from different plants were observed, underlining a coherent population distribution in three different fermentations. However, we also observed clusters formed by strains isolated from a specific fermentation, only. This could be explained considering the different technologies and recipes used for the production in three plants. Ingredient composition, fermentation and maturation parameters could play an important role in the selection of specific populations adapted in a specific environment.     

Development of a process for the production of the anticancer lead compound pleurotin by fermentation of Hohenbuehelia atrocaerulea

Pleurotin is a naphthoquinone antibiotic originally isolated from Pleurotus griseus. Two pleurotin producing strains of Hohenbuehelia atrocaerulea have been identified, which, on solid substrate fermentation for 2 months yield 1-2 mg/l of the antibiotic. Described here is the lengthy developmental process which resulted in a production protocol being developed which reliably yields pleurotin from liquid fermentation at >300 mg/l. Critical to obtaining this increase in titer was inclusion in the media of an aqueous extract of alder wood.     

鲍姆纤孔菌（桑黄）不同方式发酵的菌丝体生物活性比较

利用液体发酵、木屑固体发酵和米饭固体发酵3种方式培养鲍姆纤孔菌（桑黄）菌丝体,对菌丝体醇提物的体外抗氧化、抗肿瘤和抗衰老生物活性进行了研究。结果表明,木屑固体发酵、液体发酵和米饭固体发酵的菌丝体醇提物清除H2O2自由基的IC50值分别为78.28±0.32、27.73±0.57和7.84±0.37;米饭培养的桑黄菌丝体醇提物在低浓度500μg/mL下对超氧阴离子自由基清除作用到达80%,在相同的浓度下对DPPH自由基清除率也明显高于其他两种方法,表现出较高的抗氧化活性。木屑以及米饭培养方法得到的菌丝体对PC12神经细胞损伤修复均有较好的效果,液体培养的桑黄菌丝体表现的修复作用较低;液体发酵培养的菌丝体醇提物浓度在100μg/mL时,对肿瘤细胞HepG2的抑制率达70%,高于其他两种培养方法的抑制作用。     

Yeast communities and genetic polymorphism of Saccharomyces cerevisiae strains associated with artisanal fermentation in Brazil

Yeast communities and genetic polymorphism of prevalent Saccharomyces cerevisiae strains isolated from the spontaneous fermentation of the sugarcane juice during the production of aguardente in three distilleries in the state of Minas Gerais, Brazil, were studied. S. cerevisiae was the prevalent species during the process of aguardente production, but Schizosaccharomyces pombe was predominant in old fermentations in one distillery. Transient yeast species were found in a variable number, probably due to the daily addition of sugarcane juice, and they were different for each of the three distilleries studied. PFGE and PCR analysis of the predominant strains of S. cerevisiae isolated from the fermented must showed a high degree of genetic polymorphism among the three distilleries. A high molecular variability of S. cerevisae strains was also observed among strains isolated from the same vat at different fermentation ages. Our results showed that there was a succession of geneticly different strains of S. cerevisae during the process of aguardente production.     

Characterization of lactobacilli isolated from caper berry fermentations

AIMS: To determine the metabolic and functional properties of lactobacilli isolated from caper fermentation. METHODS AND RESULTS: A collection of 58 lactobacilli from fermentation of caper berries (including species of Lactobacillus plantarum, Lactobacillus paraplantarum, Lactobacillus pentosus, Lactobacillus brevis and Lactobacillus fermentum) was studied. Strains were classified in different clusters according to sugar fermentation patterns. Most strains of L. plantarum (the predominant species in the fermentation) clustered in a single group. Analysis of enzymatic activities revealed a high incidence of leucine aminopeptidase, acid phosphatase, beta-galactosidase and beta-glucosidase among the different strains of lactobacilli. A high number of strains were able to degrade raffinose and stachyose. Phytase activity and bile salt hydrolase activity were only detected in certain strains of L. plantarum. CONCLUSIONS: Lactobacilli from caper fermentation are metabolically diverse, and some strains display functional properties of interest. SIGNIFICANCE AND IMPACT OF THE STUDY: Strains of lactobacilli with selected functional properties could be good candidates for future development of commercial starters for industrial caper fermentation.     

Quantitative gas chromatography of Bacteroides species under different growth conditions

From 56 strains of strictly anaerobic gram-negative rods isolated from stool and purulent lesions the fermentation products in the presence and absence of hemin were determined by quantitative gas-solid chromatography, using a simple and more rapid chromatographic procedure. With hemin the fermentation products were propionic, acetic, lactic and succinic acid. Without hemin no or little succinic acid was formed and the main products were lactic and acetic acid. In both groups the distribution of subspecies was determined and the production of fatty acids measured quantitatively.Fourteen strains of the lesion group showed a higher metabolic activity, resulting in an increased total acid production caused by an excessive production of acetic and lactic acid. This characteristic is probably a virulence factor in these strains.All strains were protoporphyrin- and oxgall-dependent. It is postulated that these substances are used for the production of cytochromes which permits the formation of succinic acid by a fumarate reductase resulting in an increased growth rate and growth yield.