Occurrence and diversity of yeasts involved in fermentation of West African cocoa beans

Samples of cocoa beans were taken on two separate occasions during heap and tray fermentations in Ghana, West Africa. In total 496 yeast isolates were identified by conventional microbiological analyses and by amplification of their ITS1-5.8S rDNA-ITS2 regions. For important species the identifications were confirmed by sequencing of the D1/D2 domain of the 5' end of the large subunit (26S) rDNA. Assimilations of organic acids and other carbon compounds were conducted. For dominant yeasts intraspecies variations were examined by determination of chromosome length polymorphism (CLP) using pulsed-field gel electrophoresis. For the heap fermentations maximum yeast cell counts of 9.1 x 10(7) were reached, whereas maximum yeast counts of 6.0 x 10(6) were reached for the tray fermentations. Candida krusei was found to be the dominant species during heap fermentation, followed by P. membranifaciens, P. kluyveri, Hanseniaspora guilliermondii and Trichosporon asahii, whereas Saccharomyces cerevisiae and P. membranifaciens were found to be the dominant species during tray fermentation followed by low numbers of C. krusei, P. kluyveri, H. guilliermondii and some yeast species of minor importance. For isolates within all dominant species CLP was evident, indicating that several different strains are involved in the fermentations. Isolates of C. krusei, P. membranifaciens, H. guilliermondii, T. asahii and Rhodotorula glutinis could be found on the surface of the cocoa pods and in some cases on the production equipment, whereas the origin of e.g. S. cerevisiae was not indicated by the results obtained. In conclusion, the results obtained show that fermentation of cocoa beans is a very inhomogeneous process with great variations in both yeast counts and species composition. The variations seem to depend especially on the processing procedure, but also the season and the post-harvest storage are likely to influence the yeast counts and the species composition.     

Gas chromatographic determination and mechanism of formation of D-amino acids occurring in fermented and roasted cocoa beans, cocoa powder, chocolate and cocoa shell

Summary. Pseudomonas sp. strain phDV1, being a phenol degrading bacterium, has been found to utilize phenol as sole carbon source via the meta pathway. Blue native polyacrylamide gel electrophoresis (BN-PAGE) is widely used for the analysis of oligomeric state and molecular mass non-dissociated protein complexes. In this study, a number of proteomic techniques were used to investigate the oligomeric state enzymes involved in the aromatic degradation pathway. In particular, the Pseudomonas sp. strain phDV1 proteome was monitored under two different growth substrate conditions, using glucose or phenol as sole carbon source. The protein complexes map was compared by BN-PAGE after fractionation by sucrose density centrifugation of the cell extracts. Multiple differences were detected. Further, analysis and identification of the subunit composition of these complexes was carried out using MALDI-TOF MS, allowing the identification of 49 proteins. Additionally, functional information regarding protein–protein interactions was assembled, by coupling 2-D BN-PAGE with MALDI-TOF MS. Application of this functional proteomics method resulted in an higher number of the identified proteins.     

Bacterial diversity at different stages of the composting process

Background  

Composting is an aerobic microbiological process that is facilitated by bacteria and fungi. Composting is also a method to produce fertilizer or soil conditioner. Tightened EU legislation now requires treatment of the continuously growing quantities of organic municipal waste before final disposal. However, some full-scale composting plants experience difficulties with the efficiency of biowaste degradation and with the emission of noxious odours. In this study we examine the bacterial species richness and community structure of an optimally working pilot-scale compost plant, as well as a full-scale composting plant experiencing typical problems. Bacterial species composition was determined by isolating total DNA followed by amplifying and sequencing the gene encoding the 16S ribosomal RNA.     

不同演替阶段臭柏种群的遗传多样性

为了阐明分子变异与演替阶段或生态过渡带的联系,应用随机扩增多态性DNA (random amplified polymorphic DNA, RAPD) 标记方法对毛乌素沙地图克境内4个不同演替阶段的臭柏种群进行了分子生态学研究.用17个随机引物扩增出190条清晰谱带,其中173条为多态性谱带,并利用POPGENE 3.2 Version 1.31软件对数据进行处理.结果表明,臭柏种群的遗传多样性较丰富,各种群多态位点百分比在64.21%～74.63%,以演替早期的半固定沙地油蒿+臭柏种群最高(74.63%),演替亚顶极时期的固定沙地臭柏种群最低(64.21%);臭柏种群间分化较小,种群间遗传分化系数G_st= 0.1761, 82.39%的遗传变异存在于种群内;聚类分析显示,演替阶段相近的种群聚集到一起,反映了臭柏种群的遗传分化和演替阶段密切相关.利用Nei指数统计RAPD数据,各种群基因多样度在0.2163～0.2564之间,也证实了大部分的遗传变异存在于种群内.     

GC-MS detection of chiral markers in cocoa beans of different quality and geographic origin

Fermented cocoa beans (Theobroma cacao L., Sterculiaceae) from different countries of origin (Ecuador, Ghana, Trinidad) and cocoa beans roasted under defined conditions (industrial roasting; 150-220 degrees C for 20 min, dry roasting in conventional oven) were analyzed for their contents of certain chiral hydroxy acids, catechins, and amino acids. Cocoa beans are fermented, dried, and industrially transformed by roasting for the production of chocolate, cocoa powders, and other cocoa-related products. Fermentation and roasting conditions influence the contents of chiral compounds such as hydroxy acids, amino acids, and polyphenols, depending on technological procedures as well as some technical parameters. The aim of this work was to check if the content and nature of the named chiral compounds present both in fermented and roasted cocoa beans could be related to the traditional parameters used to classify the variety of seeds and the degree of fermentation. The extent of racemization of amino acids in fermented cocoa beans was low while it slowly increased during roasting, depending on the temperature applied. L-lactic acid was always higher than the D-form while citric acid was generally the most abundant hydroxy acid detected in beans. A correlation was found between polyphenol content and degree of fermentation, while epimerization of (-)-epicatechin to (+)-catechin was observed during roasting. On the whole, results showed that several chiral compounds could be considered as good quality markers for cocoa seeds and cocoa-related products of different quality and geographic origin.     

黔西北地区不同演替阶段植物群落结构与物种多样性特征

该文采用"空间代替时间"的方法,研究了贵州省威宁县喀斯特地区植被演替过程中的群落结构、物种组成、生活型谱和物种多样性的变化规律。结果表明:(1)该调查共记录到种子植物174种,隶属于52科117属,物种分布较多的有菊科、蔷薇科、禾本科、杜鹃花科、小檗科、唇形科、蓼科。(2)随着植被的正向演替,物种丰富度逐渐增加,群落结构趋于复杂,高位芽植物所占比例逐渐增大。(3)随着植被的恢复,群落层次分化逐渐明显,大径级植株所占比例呈现增加趋势。(4)随着植被的恢复,群落各层次的ShannonWiener多样性指数(H)、Simpson多样性指数(DS)、均匀度指数(J)和Margalef丰富度指数(DM)逐渐增加;不同演替阶段植物群落之间的Srensen相似系数呈现先上升后下降的趋势,Cody指数则表现为逐渐增加的趋势。黔西北地区不同演替阶段植物群落结构和物种多样性不同,建群种和关键种发生了明显变化,不同演替阶段植物群落结构和物种多样性的研究对喀斯特地区植被演替规律的认识和生态恢复具有重要意义。     

不同树龄杨树人工林的根系呼吸季节动态

根系呼吸是准确评估森林生态系统土壤碳收支的一个重要依据。基于LI-COR-6400-09土壤呼吸系统连续2a测定的3个生长阶段杨树人工林的根系呼吸数据,分析了根系呼吸的季节变化规律及树龄、土壤水热因子和细根生物量对它的影响。结果表明:3个不同树龄人工林的根系呼吸速率均呈明显的季节变化,最大值出现在夏初,最小值出现在秋末,基本上与表层土壤温度的季节变化相一致。根系呼吸的峰值早于土壤温度和细根生物量的峰值,说明林木根系的季节生长节律、地下碳分配模式都可能影响根系呼吸的季节变化。2年生人工林的根系呼吸速率最高,平均为3.78μmolCO2m-2s-1,并随树龄增长呈下降趋势。3个树龄人工林根系呼吸占土壤呼吸的比例介于38.6%-58.0%之间,且2年生人工林最大。不同林龄之间根系呼吸的差异主要与根系的生长周转速率及代谢活性随生长阶段的变化有关。总的来说,表层土壤温度和细根生物量的协同作用可解释根系呼吸速率变化的76%。此外在评估一个轮伐期内的根系呼吸强度时,应考虑不同生长阶段对它的影响。     

Lactic acid bacteria dynamics during spontaneous fermentation of cocoa beans verified by culture-independent denaturing gradient gel electrophoresis

Cocoa is naturally fermented in the field before the cocoa seeds are removed for processing. We assessed the dynamics of lactic acid bacteria during cocoa fermentation in Bahia, Brazil. During five days of fermentation, temperature and pH were measured and beans were collected for genomic DNA extraction every 12 h. The DNA was used as a template for amplification with Lac1-Lac2 and Lac3-Lac2 for denaturing gradient gel electrophoresis analyses. pH values ranged from 3.34 to 4.98, while the temperature varied from 23° to 50°C. Lac1-Lac2 primers permitted detection of 11 operational taxonomic units. Twenty-eight operational taxonomic units were obtained with the primer pair Lac3-Lac2. It was observed that there were variations between the numbers of operational taxonomic units throughout the process, probably because of changes in pH and temperature. The greatest similarity in amplified samples was obtained with the primers Lac3-Lac2.     

Diversity and dynamics of bacterial communities associated with Haematococcus pluvialis at different life stages

Journal of Applied Phycology - The physiological performance and productivity of microalgae are closely associated with their bacterial communities. Haematococcus pluvialis is an important...     

Recruitment of exogenous macrophages into metastases at different stages of tumor growth

Summary The endogenous tumor-associated macrophage content and recruitment of labeled peritoneal exudate cells into experimental murine B16 melanoma metastases has been examined at different stages in the progressive growth of metastatic lesions. The recruitment of thioglycollate-elicited peritoneal exudate cells and peritoneal exudate cells activated in vitro with muramyl dipeptide was studied. Tumor-associated macrophages and labeled peritoneal exudate cells were identified in paraffin sections by specific histochemical staining and their density in individual metastases measured morphometrically. The density of tumor-associated macrophages and exogenously recruited peritoneal exudate cells was high in very small lesions but decreased rapidly as a function of enlargement of metastases, MD:Aⁿ; where MD is macrophage density, A is the cross-sectional area of the lesion and n is a negative number. No significant difference was observed in the recruitment of activated and nonactivated peritoneal exudate cells. These results suggest that decreased recrutiment of macrophages from the circulation may explain the decrease in the density of tumor-associated macrophages as metastases grow and indicate that macrophage activation is not accompanied by enhanced localization and/or uptake of macrophages into metastases.     

Orientation sensitivity at different stages of object processing: evidence from repetition priming and naming

Background

An ongoing debate in the object recognition literature centers on whether the shape representations used in recognition are coded in an orientation-dependent or orientation-invariant manner. In this study, we asked whether the nature of the object representation (orientation-dependent vs orientation-invariant) depends on the information-processing stages tapped by the task.

Methodology/ Findings

We employed a repetition priming paradigm in which briefly presented masked objects (primes) were followed by an upright target object which had to be named as rapidly as possible. The primes were presented for variable durations (ranging from 16 to 350 ms) and in various image-plane orientations (from 0° to 180°, in 30° steps). Significant priming was obtained for prime durations above 70 ms, but not for prime durations of 16 ms and 47 ms, and did not vary as a function of prime orientation. In contrast, naming the same objects that served as primes resulted in orientation-dependent reaction time costs.

Conclusions/Significance

These results suggest that initial processing of object identity is mediated by orientation-independent information and that orientation costs in performance arise when objects are consolidated in visual short-term memory in order to be reported.     

喀斯特断陷盆地区不同恢复阶段群落物种组成与多样性特征

研究自然植被恢复过程中的物种组成、群落结构及生物多样性的变化,能够为人工促进植被恢复的树种选择与群落结构的优化配置提供重要依据。本研究以空间代替时间对喀斯特断陷盆地典型区云南省建水县不同天然植被(草丛、灌丛、乔木林)进行群落学调查,对不同恢复阶段的植物群落按乔木、灌木、草本进行分层,分析各恢复阶段植物群落的物种组成、水平和垂直结构、生物多样性。结果表明:在总面积为3200 m²的12个样地中,共记录43科72属94种维管束植物,优势种以壳斗科(Fagaceae)、鼠李科(Rhamnaceae)、紫金牛科(Myrsinaceae)、蔷薇科(Rosaceae)、木犀科(Oleaceae)等科的植物为主;在草丛→灌丛→乔木林的恢复过程中,群落物种组成中的科数、属数、种数逐渐增加,低矮和小径级植物个体数所占比例逐渐减少,但整体仍以低矮的小径级植物为主。草本植物的丰富度和Shannon指数在植被恢复的初期即草丛阶段最大,而均匀度指数则以灌丛阶段最大;木本植物的丰富度和Shannon指数随着植被的恢复逐渐增大,但均匀度指数随着植被的恢复逐渐下降;随着植被的恢复,草本层和乔木层...     

青海云杉不同演替阶段林下草本多样性特征及其环境解释

林下草本的演变与森林演替密切关联,为了解祁连山地区青海云杉林在不同演替阶段下的草本多样性特征及影响因素,本研究采用空间代替时间的方法,以阔叶纯林(Ⅰ)→阔叶混交林(Ⅱ)→针阔混交林(Ⅲ)→云杉纯林(Ⅳ)构成的演替序列为研究对象,计算分析不同演替阶段的草本多样性指标差异,探究生物因素和非生物因素对草本多样性的影响,旨在为青海云杉林林分结构优化与功能提升提供理论依据。结果表明：(1)不同演替阶段下,共发现草本31科64属81种,其中阔叶纯林(Ⅰ)草本植物共21科33属42种,阔叶混交林(Ⅱ)共22科40属50种,针阔混交林(Ⅲ)共22科42属51种,青海云杉纯林(Ⅳ)共15科21属23种。(2)随着演替的进行,物种丰富度指数、Simpson指数、Shannon-Wiener指数、Pielou指数均呈现先增大后减少的趋势,草本多样性指标均在针阔混交林(Ⅲ)阶段最大,在青海云杉纯林(Ⅳ)阶段最低。(3)生物因子中：LAI是影响草本多样性的主导因子,林分密度、胸径、树高和叶面积指数与草本多样性负相关。(4)非生物因子中,土壤有机质和速效钾是主导因子,土壤水分、速效磷、碱解氮与多样性指标正相关。综...     

Evaluation of the extent and type of bacterial contamination at different stages of processing of cooked ham

In an attempt to determine the composition and origin of the spoilage flora of refrigerated vacuum-packed cooked ham, the changes in microbial numbers and types were followed along the processing line. Results revealed Lactobacillus sake and Leuconostoc mesenteroides ssp. mesenteroides as the major causative agents of spoilage of sliced ham stored at 4 °C and 12 °C, due to recontamination in the cutting room. On the contrary, the progressive deterioration of whole ham under the same storage conditions was associated with a non-identifiable group of leuconostoc-like bacteria. Except for lactic acid bacteria, no other organism grew in vacuum packs of either sliced or whole ham. Although atypical leuconostocs could not be detected among isolates recovered from freshly produced whole ham, they appeared to survive cooking and proliferate during storage. Neither these organisms however, nor Lact. sake and Leuc. mesenteroides were important in curing and tumbling as carnobacteria, mainly Carnobacterium divergens, and Brochothrix thermosphacta dominated at this stage. A progressive inversion of the ham microflora from mostly Gram-negative at the beginning of processing to highly Gram-positive prior to cooking was noted. Listeria monocytogenes cross-contaminated ham during tumbling. However, the pathogen was always absent from the vacuum-packed product provided that heating to a core temperature of 70 °C occurred and recontamination during slicing and packing was prevented. The percentage distribution of different species of lactic acid bacteria as well as the uncommon phenotypic characteristics of some strains were discussed.     

基于高通量测序的不同虫态洋虫内共生微生物群落分析

【目的】为分析不同虫态下洋虫内共生微生物菌群组成情况,以成虫和幼虫两种虫态为研究对象。【方法】通过对洋虫内共生微生物提取及高通量测序分析,获得不同样品中微生物组成结构及变化情况。【结果】不同虫态下,随着生长发育的进行,洋虫体内细菌在丰度和多样性两个方面都有很大变化,成虫中的菌群数量相对较少,多样性较幼虫也明显降低。在几种特定微生物组成上不同虫态也存在着较大的差异,成虫中,漫游球菌属和乳酸菌菌属为优势菌属;幼虫体内乳酸菌菌属为优势菌属。真菌变化不明显,孢囊线黑粉酵母菌属为其优势菌属。【结论】研究结果将为后继洋虫生物资源的研究提供相关信息,为洋虫内共生微生物资源开发与利用提供理论依据。     

Investigation on diversity and population succession dynamics of endophytic bacteria from seeds of maize (Zea mays L., Nongda108) at different growth stages

Plant seeds are carriers of both beneficial bacteria and pathogens. Using the 16S rRNA gene clone library technique, we conducted a preliminary study on the community diversity and population succession dynamics of endophytic bacteria in seeds of reciprocal cross hybrid maize at different seed developmental stages. In both hybrid lines (108A and 108B), more types of endophytic bacteria were found at the proembryo-forming stage than in the other two stages, including 29 and 23 bacterial operational taxonomic units (OTUs), respectively. Undibacterium (39.20 and 30.00 % in 108A and 108B, respectively) was the first dominant bacterium to appear. At the milky stage, fewer types of endophytic bacteria in 108A and 108B appeared, including 18 and 16 OTUs, respectively, and the abundance of the dominant genus Burkholderia in the two seed samples reached 73.38 and 80.43 %, respectively. Limnobacter appeared as the second and third endophytic dominant bacterium in 108A (4.55 %) and 108B (5.07 %), respectively, in both seed samples. At the dough stage, the abundance of the first dominant bacterium, Burkholderia, in 108A and 108B was 78.26 and 84.80 %, respectively. Pantoea appeared as the second endophytic dominant bacterium in the both seeds (9.42 and 4.80 % in 108A and 108B, respectively). This is the first study on endophytic bacteria present during several crucial stages of the dynamic grain growth process of plant seeds conducted using culture-independent methods.     

Circulation time and dynamics of 3H-thymidine incorporation into DNA at various stages of chick embryo development

The most adequate method for carrying out radioautographic investigation on proliferation in the chorioallantoic tissues and in the chick embryo proper is dropping 3H-thymidine on the membrane under the shell in the area where the chorioallantois grows under. Using the application method, the radioautographic analysis of proliferation could be performed by the saturation method.     

Time course of meiotic progression after transferring primary spermatocyte into ooplasm at different stages

This study attempted to investigate the time course of meiotic progression after transferring primary spermatocyte (PS) into ooplasm at different maturing stages. In present experiments, PSs were introduced into maturing ooplasts or oocytes by electrofusion. Higher fusion rate was obtained by phytohemagglutinin (PHA) agglutination than by perivitelline space (PVS) insertion. When the ooplasms prepared at 0, 2, 5, and 8.5 hr of in vitro maturation (IVM) were used as recipients and PSs were used as donors, the reconstructed cells extruded the first polar body (PB1) approximately 8.5, 7, 5.5, and 3 hr after electrofusion, respectively. Especially, when ooplasm cultured for 8.5 hr in vitro after GV removal was fused with PS, the PB1 was emitted 7-11 hr after electrofusion. Additionally, the PB1 extrusions of GV and pro-MI oocytes fertilized with PSs were 2.5 hr earlier than control oocytes. The results suggest that (1) PSs undergo the first meiosis in different time courses when introduced into ooplasm at different maturing stages; (2) GV material plays an important role in determining the timing of PB1 extrusion; and (3) first meiotic division of GV and pro-MI oocytes can be accelerated by introducing PS.