Avaliação da interação entre leveduras isoladas de kombucha e bactérias probióticas / Evaluation of the interaction between isolated kombucha yeasts and probiotic bacteria

Um dos maiores desafios no desenvolvimento de produtos probióticos é entender como os microrganismos interagem entre si e com o hospedeiro. Quando falamos em alimentos fermentados tradicionais, este obstáculo aumenta porque a matriz alimentar já possui um microbioma intrínseco. No entanto, também é conhecido que muitos microrganismos podem interagir e cooperar para sobreviver quando condições de estresse são encontradas. Assim, o objetivo deste trabalho foi isolar leveduras de quatro diferentes kombuchas em distintos momentos fermentativos e verificar a influência que leveduras isoladas de kombucha têm na manutenção da viabilidade da bactéria probiótica Bifidobacterium animalis subsp. lactis HN019 em condições de aerobiose. Meyerozyma guilliermondii, Candida albicans, Rhodotorula mucilaginosa e Pichia membranifaciens foram leveduras encontradas nas kombuchas, das quais as duas últimas favoreceram a manutenção da alta viabilidade de HN019 em cocultura por 14 dias. Observou-se a viabilidade da bactéria acima de 9 log ao longo de todo o experimento, o que não foi observado em monocultura. Ademais, utilizou-se de análise de autoagregação, hidrofobicidade, atividade enzimática de proteases e fosfolipases das leveuras para analisar seu potencial patogênico. Observou-se que R. mucilaginosa demonstrou características semelhantes à Saccharomyces cerevisiae subsp. boulardii, e sua interação benéfica com HN019 reforça a possibilidade de que esta levedura seja uma chave para a inserção da bactéria em uma kombucha probiótica. Análises metabólicas foram realizadas e encontrou-se uma vasta diversidade de dipeptídeos, principalmente os compostos de prolina, durante a cocultura da bactéria com as leveduras. Tais dipeptídeos apresentam importantes mecanismos de ação no controle biológico e quorum sensing de bactérias e leveduras, e supostamente regulam a manutenção das relações mutualísticas entre ambos microrganismo

One of the biggest challenges in the development of probiotic products is to understand how microorganisms interact with each other and with the host. When we talk about traditional fermented foods, this obstacle increases because the food matrix already has an intrinsic microbiome. However, it is also known that many microorganisms can interact and cooperate to survive when stressful situations are encountered. Thus, the objective of this work was to isolate yeasts from four different kombuchas at different fermentation times and to verify the influence that yeasts isolated from kombucha have on maintaining the viability of the probiotic bacterium Bifidobacterium animalis subsp. lactis HN019 under aerobic conditions. Meyerozyma guilliermondii, Candida albicans, Rhodotorula mucilaginosa and Pichia membranifaciens were yeasts found in kombuchas, of which the last two favored the maintenance of HN019 high viability in co-culture for 14 days. Bacteria viability above 9 log was observed throughout the experiment, which was not observed in monoculture. In addition, analysis of autoaggregation, hydrophobicity, enzyme activity of proteases and phospholipases of yeasts was used to analyze their pathogenic potential. It was observed that R. mucilaginosa demonstrated characteristics similar to Saccharomyces cerevisiae subsp. boulardii, and its beneficial interaction with HN019 reinforces the possibility that this yeast is a key to the insertion of the bacterium in a probiotic kombucha. Metabolic analysis were performed and a wide diversity of dipeptides, mainly proline-based, was found during the co-culture of the bacteria with the yeasts. Such dipeptides have important mechanisms of action in the biological control and quorum sensing of bacteria and yeast, and supposedly regulate the maintenance of mutualistic relationships between both microorganism

Microbial Diversity and Characteristics of Kombucha as Revealed by Metagenomic and Physicochemical Analysis.

Kombucha is a fermented tea made from a Symbiotic Culture of Bacteria and Yeast (SCOBY) with a long history of use as a health tonic. It is likely that most health benefits come from the tea and fermentation metabolites from specific microbial communities. Despite its growing importance as a functional health drink, the microbial ecosystem present in kombucha has not been fully documented. To characterize the microbial composition and biochemical properties of 'The Good Brew' original base kombucha, we used metagenomics amplicon (16S rRNA and ITS) sequencing to identify the microbial communities at the taxonomic level. We identified 34 genera with 200 microbial species yet described in kombucha. The dominance of organic acid producing microorganisms Acetobacter, Komagataeibacter and Starmerella are healthy for the human gut and their glucose metabolising activities have a putative role in preventing conditions such as diabetes and obesity. Kombucha contains high protein (3.31 µg/mL), high phenolic content (290.4 mg/100 mL) and low sugars (glucose: 1.87 g/L; sucrose 1.11 g/L; fructose: 0.05 g/L) as compared to green tea. The broad microbial diversity with proven health benefits for the human gut suggests kombucha is a powerful probiotic. These findings are important to improve the commercial value of kombucha and uncover the immense prospects for health benefits.

Patterns of yeast diversity distribution and its drivers in rhizosphere soil of Hami melon orchards in different regions of Xinjiang.

BACKGROUND: The unique climatic conditions of the Xinjiang region nurture rich melon and fruit resources, the melon and fruit sugar sources provide sufficient nutrients for the survival of yeast, and the diverse habitats accompanied by extreme climatic conditions promote the production of yeast diversity and strain resources. However, the relationship between yeast species and their relationship with environmental factors in the soil of Xinjiang specialty cash crop Hami melon is not clear. Here, we aimed to characterize the diversity, community structure, and relationship between yeast species and environmental factors in Hami melon orchards soils in different regions of Xinjiang, China. RESULTS: Based on Illumina MiSeq high-throughput sequencing analysis of the D1 domain of the LSU rRNA genes, the community richness of yeast in the soil of Northern Xinjiang was higher than in the Southern and Eastern Xinjiang, but the community diversity was significantly lower in the Northern Xinjiang than in the Southern and Eastern Xinjiang. A total of 86 OTUs were classified into 59 genera and 86 species. Most OTUs (90.4%) belonged to the Basidiomycota; only a few (9.6%) belonged to Ascomycota. The most dominant species in the Southern, Eastern and Northern Xinjiang were Filobasidium magnum (17.90%), Solicoccozyma aeria (35.83%) and Filobasidium magnum (75.36%), respectively. Principal coordinates analysis (PCoA) showed that the yeast community composition in the soils of the three regions were obviously different, with the Southern and Eastern Xinjiang having more similar yeast community. Redundancy analysis (RDA) showed that soil factors such as conductivity (CO), total phosphorus (TP) and Total potassium (TK) and climate factors such as average annual precipitation (PRCP), relative humidity (RH) and net solar radiation intensity (SWGNT) were significantly correlated with yeast communities (P < 0.05). CONCLUSION: There are abundant yeast resources in the rhizosphere soil of Hami melon orchard in Xinjiang, and there are obvious differences in the diversity and community structure of yeast in the three regions of Xinjiang. Differences in climatic factors related to precipitation, humidity and solar radiation intensity and soil factors related to conductivity, total phosphorus and total potassium are key factors driving yeast diversity and community structure.

Fermentation of Coffea canephora inoculated with yeasts: Microbiological, chemical, and sensory characteristics.

This work aimed to evaluate Coffea canephora's microbiological, chemical, and sensory characteristics at 300 and 600 m elevation plantations processed by the natural method inoculated with yeasts. The coffee was spread on suspended terraces and sprayed with approximately 107 cfu/mL of Meyerozyma caribbica CCMA 1738 or Pichia kluyveri CCMA 1743, separately. Cherries containing bark and parchment were collected during fermentation for microbial groups counting, qPCR, quantification of organic acids, and sugars (HPLC). Volatile compounds (GC-MS) and sensory analyses, cupping test with expert coffee tasters and triangular test with consumers, were performed on roasted coffee beans. The inoculated yeasts persisted during the entire fermentation process. M. caribbica reduced the filamentous fungal population by 63% and 90% in the 300- and 600-m coffees, respectively. The 300-m coffee fruits showed higher concentrations of organic acids in all fermentation times when compared to the 600-m reaching out to 8 times more. Twenty-four volatile compounds were identified in the roasted coffee beans, with the predominance of pyrazines. The 600-m coffee inoculated with M. caribbica showed an increase of more than one point in the score given by certified tasters. Consumers noticed the M. caribbica inoculation in the 300- and 600-m-elevation coffees. M. caribbica is a promising starter culture for Conilon coffee with the potential to increase the beverage quality.

Isolation of microorganisms from the feces of ring-tailed coati related to the production of "misha coffee" in the central forest of Peru and evaluation of some features of technological importance.

A diversity of yeasts and lactic bacteria were isolated from the feces of ring-tailed coaties bred in captivity and related to the production of "misha coffee". Isolation of yeasts was carried out using oxytetracycline-glucose-yeast extract agar containing 100 mg/L oxytetracycline and, lactic bacteria using de Man-Rogosa and Sharpe agar containing 20 mg/L of vancomicin. Then, isolates were biochemically analysed using API strips (ID 32C for yeasts and 50CHL for lactic bacteria) followed by 16S and 26S rRNA gene sequencing. Among the yeasts, Debaryomyces hansenii, Pichia kluyveri, Pichia kudriavzevii, and Candida sorboxilosa were the most frequent, whereas Weissella cibaria, Weissella paramesenteroides, Enterococcus thailandicus and Enterococcus faecalis were the most important lactic bacteria. Cultivation of the isolated yeasts under agitated conditions, showed that Pichia kluyveri LBFT.Lev3 (0.15 ± 0.01 h-1) and Pichia kudriavzevii LBTF.Lev7 (0.14 ± 0.01 h-1) had higher specific growth rates than Debaryomyces hansenii LBFT.Lev9 (0.09 ± 0.01 h-1), whereas cultivation of lactic bacteria under static fashion showed that Weisella paramesenteroides LBTF.Bal2 (0.16 ± 0.01 h-1) and Weisella cibaria LBTF.Bal3 (0.18 ± 0.01 h-1) had better growth than Enterococcus thailandicus LBTF.Bal1 (0.1 ± 0.015 h-1) and Enterococcus faecalis LBTF.Bal7 (0.14 ± 0.01 h-1). Additionally, evaluation of pectinolytic activity revealed that Pichia kudriavzevii LBTF.Lev7 and Debaryomyces hansenii LBFT.Lev9 were able to use pectin as carbon source for their growth. On the other hand, W. cibaria LBTF.Bal3, E. thailandicus LBTF.Bal1 and W. paramesenteroides LBTF.Bal2 showed inhibitory activity against S. mutans ATCC 35668, B. subtilis subsp. spizizenii ATCC 6633 and Staph. epidermidis ATCC 14990. Results of this study are useful for the search of potential application of the isolated yeasts and lactic bacteria in coffee and other food fermentations.

Metabolome-microbiome signatures in the fermented beverage, Kombucha.

Kombucha is a fermented tea. Here we investigate the fermentation kinetics, metabolite production, microbiome and potential health promoting properties of three different kombucha consortia. Shotgun metagenomic sequencing revealed several dominant bacterial genera such as Komagataeibacter, Gluconacetobacter and Gluconobacter. Brettanomyces and Schizosaccharomyces were the most dominant yeasts identified. Species distribution reflected different patterns of sugar consumption, with S. pombe being present in samples with the highest sugar conversion. Liquid-liquid extractions were performed with organic solvents in order to obtain dried extracts, which were later characterized. HPLC-DAD and GC-MS analysis revealed differences in the production of organic acids, sugars, alcohols and phenolic compounds, where the presence of caffeine, propanoic acid and 2,3 butanediol differ greatly across the three kombuchas. Metabolomic analysis exhibited a link between the microbiota and the production of bioactive compounds in kombucha fermentation. In vitro assays were carried out in order to evaluate potential health-promoting features of the fermented teas, with notable outcomes including antioxidant ability against DPPH radical and against the 15-lipoxygenase enzyme, indicating a potential anti-inflammatory activity. These investigations considerably enhance our understanding of the relationship between the microbiota and metabolites as well as health promoting potential of kombucha and have the potential for the development of future generations of kombucha products in which these relationships are optimized.

Valorisation of pectin-rich agro-industrial residues by yeasts: potential and challenges.

Pectin-rich agro-industrial residues are feedstocks with potential for sustainable biorefineries. They are generated in high amounts worldwide from the industrial processing of fruits and vegetables. The challenges posed to the industrial implementation of efficient bioprocesses are however manyfold and thoroughly discussed in this review paper, mainly at the biological level. The most important yeast cell factory platform for advanced biorefineries is currently Saccharomyces cerevisiae, but this yeast species cannot naturally catabolise the main sugars present in pectin-rich agro-industrial residues hydrolysates, in particular D-galacturonic acid and L-arabinose. However, there are non-Saccharomyces species (non-conventional yeasts) considered advantageous alternatives whenever they can express highly interesting metabolic pathways, natively assimilate a wider range of carbon sources or exhibit higher tolerance to relevant bioprocess-related stresses. For this reason, the interest in non-conventional yeasts for biomass-based biorefineries is gaining momentum. This review paper focuses on the valorisation of pectin-rich residues by exploring the potential of yeasts that exhibit vast metabolic versatility for the efficient use of the carbon substrates present in their hydrolysates and high robustness to cope with the multiple stresses encountered. The major challenges and the progresses made related with the isolation, selection, sugar catabolism, metabolic engineering and use of non-conventional yeasts and S. cerevisiae-derived strains for the bioconversion of pectin-rich residue hydrolysates are discussed. The reported examples of value-added products synthesised by different yeasts using pectin-rich residues are reviewed. Key Points • Review of the challenges and progresses made on the bioconversion of pectin-rich residues by yeasts. • Catabolic pathways for the main carbon sources present in pectin-rich residues hydrolysates. • Multiple stresses with potential to affect bioconversion productivity. • Yeast metabolic engineering to improve pectin-rich residues bioconversion. Graphical abstract.

Ecological diversity, evolution and metabolism of microbial communities in the wet fermentation of Australian coffee beans.

The microbial ecology in the fermentation of Australian coffee beans was investigated in this study. Pulped coffee beans were kept underwater for 36 h before air dried. Samples were collected periodically, and the microbial communities were analyzed by culture-dependent and independent methods. Changes in sugars, organic acids and microbial metabolites in the mucilage and endosperm of the coffee beans during fermentation were monitored by HPLC. Culture-dependent methods identified 6 yeast and 17 bacterial species, while the culture-independent methods, multiple-step total direct DNA extraction and high throughput sequencing, identified 212 fungal and 40 bacterial species. Most of the microbial species in the community have been reported for wet fermentation of coffee beans in other parts of the world, but the yeast Pichia kudriavzevii was isolated for the first time in wet coffee bean fermentation. The bacterial community was dominated by aerobic mesophilic bacteria (AMB) with Citrobacter being the predominant genus. Hanseniaspora uvarum and Pichia kudriavzevii were the predominant yeasts while Leuconostoc mesenteroides and Lactococcus lactis were the predominant LAB. The yeasts and bacteria grew significantly during fermentation, utilizing sugars in the mucilage and produced mannitol, glycerol, and lactic acid, leading to a significant decrease in pH. The results of this study provided a preliminary understanding of the microbial ecology of wet coffee fermentation under Australian conditions. Further studies are needed to explore the impact of microbial growth and metabolism on coffee quality, especially flavour.

Yeast diversity in Vitis non-vinifera ecosystems / Diversidad de levaduras en ecosistemas de Vitis no-vinifera

The surface of grapes lodges a complex community of yeast species responsible for spontaneous alcoholic fermentation. The study of indigenous Saccharomyces and "non-Saccharomyces" yeasts during grape must fermentation constitutes a major research area in microbial enology. Although there are detailed studies on the microbiota of Vitis vinifera grapes, little is known about the diversity of yeast communities present in non-vinifera Vitis ecosystems (i.e., grapes and spontaneously fermenting grape musts). Potentially scientific and/or enological valuable yeast strains from these non-vinifera Vitis ecosystems might never be isolated from V. vinifera L. In this updated review, we summarize relevant aspects of the microbial studies conducted on V. non-vinifera grapes and spontaneously fermenting grape musts.

La superficie de las uvas aloja una comunidad compleja de especies de levaduras responsables de la fermentación alcohólica espontánea. El estudio de estas levaduras Saccharomyces y «no-Saccharomyces¼ durante la fermentación del mosto de uvas constituye un área relevante de investigación microbiológica en enología. Si bien existen estudios detallados de la microbiota de uvas de Vitis vinifera L., poco se sabe sobre la diversidad de comunidades de levaduras presentes en ecosistemas de Vitis no-vinifera (i.e., uvas y mostos en fermentación espontánea). Cepas de levaduras presentes en ecosistemas de Vitis no-vinífera, con valor potencial científico y/o enológico, podrían no estar presentes en V. vinifera L. En esta revisión actualizada, resumimos los aspectos relevantes de los estudios microbiológicos efectuados en mostos en fermentación espontánea de uvas de V. no-vinifera.

Occurrence of oleaginous yeast from mangrove forest in Thailand.

A total of 191 yeasts were isolated from 197 samples collected from eight estuarine mangrove forests along four different coastlines of Thailand (Andaman Sea and the East, North and West coasts of the Gulf of Thailand). Of these, 178 isolates were identified as 32 species in 16 genera of Ascomycota, 12 species in nine genera of Basidiomycota, and 13 isolates as potential new species, respectively. Mangroves located along the Andaman Sea coastline had a higher yeast diversity at the species and genera levels than those along the Gulf of Thailand. Kluyveromyces siamensis was the most frequently isolated species, whilst Candida tropicalis was the only species isolated at all eight sites. Screening isolated yeast strains belonging to genera previously reported as oleaginous yeast plus the 13 potential new species, revealed two oleaginous strains, Rhodotorula sphaerocarpa 11-14.4 and Saitozyma podzolica 11-11.3.1. Both of these strains were isolated from the same mangrove forest on the Andaman Sea coastline. They could accumulate lipid when suspended in glucose solution without any supplementation, while the fatty acid composition and oil profile of Rh. sphaerocarpa 11-14.4 and Sait. podzolica 11-11.3.1 were similar to vegetable oil and cocoa butter, respectively.

The utilisation of nitrogenous compounds by commercial non-Saccharomyces yeasts associated with wine.

Commercial wine fermentation is commonly conducted by inoculated strains of Saccharomyces cerevisiae. However, other non-Saccharomyces yeast species have recently become popular co-inoculants. Co-inoculated yeast species compete with each other for nutrients, and such competition may impact fermentation kinetics and aroma production. Understanding the specific nutrient requirements of non-Saccharomyces strains therefore is essential to better characterize the competitive potential of each strain, and to support rational decision making for nutrient supplementation during wine making. This study investigated the nitrogen source preference of commercial non-Saccharomyces yeasts by conducting pure culture and sequential culture fermentations in synthetic grape musts with adjusted nitrogen contents. Amino acid and ammonium uptake varied between yeast species. Lachancea thermotolerans and Torulaspora delbrueckii assimilated more nitrogen at a faster rate than Pichia kluyveri and Metschnikowia pulcherrima. Significant variation in amino acid preference between species was observed. Sequential fermentations confirmed the more competitive behaviour of L. thermotolerans and T. delbrueckii, with consequences for fermentation kinetics and aroma production. Furthermore, the data suggest that declining populations of non-Saccharomyces yeasts release nitrogen and supports the activity of S. cerevisiae. The data provide the most detailed assessment of nitrogen utilisation by the investigated yeast strains in a wine environment.

Yeast diversity in Vitis non-vinifera ecosystems.

The surface of grapes lodges a complex community of yeast species responsible for spontaneous alcoholic fermentation. The study of indigenous Saccharomyces and "non-Saccharomyces" yeasts during grape must fermentation constitutes a major research area in microbial enology. Although there are detailed studies on the microbiota of Vitis vinifera L. grapes, little is known about the diversity of yeast communities present in non-vinifera Vitis ecosystems (i.e., grapes and spontaneously fermenting grape musts). Potentially scientific and/or enological valuable yeast strains from these non-vinifera Vitis ecosystems might never be isolated from V. vinifera L. In this updated review, we summarize relevant aspects of the microbial studies conducted on V. non-vinifera grapes and spontaneously fermenting grape musts.

Microbial cell factories for the sustainable manufacturing of B vitamins.

Vitamins are essential compounds in human and animal diets. Their demand is increasing globally in food, feed, cosmetics, chemical and pharmaceutical industries. Most current production methods are unsustainable because they use non-renewable sources and often generate hazardous waste. Many microorganisms produce vitamins naturally, but their corresponding metabolic pathways are tightly regulated since vitamins are needed only in catalytic amounts. Metabolic engineering is accelerating the development of microbial cell factories for vitamins that could compete with chemical methods that have been optimized over decades, but scientific hurdles remain. Additional technological and regulatory issues need to be overcome for innovative bioprocesses to reach the market. Here, we review the current state of development and challenges for fermentative processes for the B vitamin group.

The effects of probiotics in lactose intolerance: A systematic review.

Over 60 percent of the human population has a reduced ability to digest lactose due to low levels of lactase enzyme activity. Probiotics are live bacteria or yeast that supplements the gastrointestinal flora. Studies have shown that probiotics exhibit various health beneficial properties such as improvement of intestinal health, enhancement of the immune responses, and reduction of serum cholesterol. Accumulating evidence has shown that probiotic bacteria in fermented and unfermented milk products can be used to alleviate the clinical symptoms of lactose intolerance (LI). In this systematic review, the effectiveness of probiotics in the treatment of LI was evaluated using 15 randomized double-blind studies. Eight probiotic strains with the greatest number of proven benefits were studied. Results showed varying degrees of efficacy but an overall positive relationship between probiotics and lactose intolerance.

Isolation and molecular characterization of biosurfactant producing yeasts from the soil samples contaminated with petroleum derivatives.

In this study, 65 yeast strains were isolated from different environmental samples contaminated with various petroleum hydrocarbons such as activated sludges and soil samples from automobile workshops. The yeast isolates were tested for biosurfactant production using various screening methods such as parafilm M test, oil displacement assay, drop collapse assay, determination of surface tension reduction, and emulsification index. Nineteen of the isolates were found positive for biosurfactant production and their molecular characterizations were carried out by sequencing analysis of the ITS1-5.8S-ITS2 region and D1/D2 domain of 26S rDNA. The results indicated that these strains were from a wide range of yeast genera including Rhodotorula, Candida, Yarrowia, Geotrichum, Galactomyces, and Cystobasidium. The studies to determine the emulsification index revealed that the biosurfactants produced by Yarrowia lipolytica strains (TEMGS33, TEMOS12, and TEMOS14) and Apiotrichum loubieri strain (TEMOS16) were the most potent and capable of forming stable emulsions with emulsion index (E24 ) up to 68%. In addition, quantitative measurements of the surface tension reduction of the biosurfactants produced by these strains were carried out by Du Noüy ring method. Biosurfactants produced from Yarrowia lipolytica strain TEMGS33 and Apiotrichum loubieri strain TEMOS16 gave the best results reducing the surface tension to 34.7 ± 1.15 and 35.3 ± 0.55 mN m-1 , respectively. Based on these data, biosurfactants from Yarrawia lipolytica strains (TEMGS33, TEMOS12, and TEMOS14) and Apiotrichum loubieri strain (TEMOS16) showed promising results and might be implemented in numerous industrial fields such as bioremediation and food industry.

Understanding Kombucha Tea Fermentation: A Review.

Kombucha is a beverage of probable Manchurian origins obtained from fermented tea by a microbial consortium composed of several bacteria and yeasts. This mixed consortium forms a powerful symbiosis capable of inhibiting the growth of potentially contaminating bacteria. The fermentation process also leads to the formation of a polymeric cellulose pellicle due to the activity of certain strains of Acetobacter sp. The tea fermentation process by the microbial consortium was able to show an increase in certain biological activities which have been already studied; however, little information is available on the characterization of its active components and their evolution during fermentation. Studies have also reported that the use of infusions from other plants may be a promising alternative. PRACTICAL APPLICATION: Kombucha is a traditional fermented tea whose consumption has increased in the recent years due to its multiple functional properties such as anti-inflammatory potential and antioxidant activity. The microbiological composition of this beverage is quite complex and still more research is needed in order to fully understand its behavior. This study comprises the chemical and microbiological composition of the tea and the main factors that may affect its production.

Diversity of microbiota found in coffee processing wastewater treatment plant.

Cultivable microbiota presents in a coffee semi-dry processing wastewater treatment plant (WTP) was identified. Thirty-two operational taxonomic units (OTUs) were detected, these being 16 bacteria, 11 yeasts and 4 filamentous fungi. Bacteria dominated the microbial population (11.61 log CFU mL- 1), and presented the highest total diversity index when observed in the WTP aerobic stage (Shannon = 1.94 and Simpson = 0.81). The most frequent bacterial species were Enterobacter asburiae, Sphingobacterium griseoflavum, Chryseobacterium bovis, Serratia marcescens, Corynebacterium flavescens, Acetobacter orientalis and Acetobacter indonesiensis; these showed the largest total bacteria populations in the WTP, with approximately 10 log CFU mL- 1. Yeasts were present at 7 log CFU mL- 1 of viable cells, with Hanseniaspora uvarum, Wickerhamomyces anomalus, Torulaspora delbrueckii, Saturnispora gosingensis, and Kazachstania gamospora being the prevalent species. Filamentous fungi were found at 6 log CFU mL- 1, with Fusarium oxysporum the most populous species. The identified species have the potential to act as a biological treatment in the WTP, and the application of them for this purpose must be better studied.

Specialist nectar-yeasts decline with urbanization in Berlin.

Nectar yeasts are common inhabitants of insect-pollinated flowers but factors determining their distribution are not well understood. We studied the influence of host identity, environmental factors related to pollution/urbanization, and the distance to a target beehive on local distribution of nectar yeasts within Robinia pseudoacacia L. and Tilia tomentosa Moench in Berlin, Germany. Nectar samples of six individuals per species were collected at seven sites in a 2 km radius from each target beehive and plated on YM-Agar to visualise the different morphotypes, which were then identified by sequencing a section of the 26S rDNA gene. Multivariate linear models were used to analyze the effects of all investigated factors on yeast occurrence per tree. Yeast distribution was mainly driven by host identity. The influence of the environmental factors (NO2, height of construction, soil sealing) strongly depended on the radius around the tree, similar to the distance of the sampled beehive. Incidence of specialist nectar-borne yeast species decreased with increasing pollution/urbanization index. Given that specialist yeast species gave way to generalist yeasts that have a reduced dependency on pollinators for between-flower dispersal, our results indicate that increased urbanization may restrict the movement of nectar-specialized yeasts, via limitations of pollinator foraging behavior.

Enumeration and rapid identification of yeasts during extraction processes of extra virgin olive oil in Tuscany.

The aim of this study was to evaluate the occurrence of yeast populations during different olive oil extraction processes, carried out in three consecutive years in Tuscany (Italy), by analysing crushed pastes, kneaded pastes, oil from decanter and pomaces. The results showed yeast concentrations ranging between 10(3) and 10(5) CFU/g or per mL. Seventeen dominant yeast species were identified by random amplified polymorphic DNA with primer M13 and their identification was confirmed by restriction fragments length polymorphism of ribosomal internal transcribed spacer and sequencing rRNA genes. The isolation frequencies of each species in the collected samples pointed out that the occurrence of the various yeast species in olive oil extraction process was dependent not only on the yeasts contaminating the olives but also on the yeasts colonizing the plant for oil extraction. In fact, eleven dominant yeast species were detected from the washed olives, but only three of them were also found in oil samples at significant isolation frequency. On the contrary, the most abundant species in oil samples, Yamadazyma terventina, did not occur in washed olive samples. These findings suggest a phenomenon of contamination of the plant for oil extraction that selects some yeast species that could affect the quality of olive oil.

Culture-Dependent and -Independent Methods to Investigate the Predominant Microorganisms Associated with Wet Processed Coffee.

The fermentation process of Yunnan arabica coffee is a typical wet fermentation. Its excellent quality is closely related to microbes in the process of fermentation. The purpose of this study was to isolate and identify the microorganisms in the wet method of coffee processing in Yunnan Province, China. Microbial community structure and dominant bacterial species were evaluated by traditional cultivated separation method and PCR-DGGE technology, and were further analyzed in combination with the changes of organic acid content, activity of pectinase, and physical parameters (pH and temperature). A large number of microorganisms which can produce pectinase were found. Among them, Enterobacter cowanii, Pantoea agglomerans, Enterobacteriaceae bacterium, and Rahnella aquatilis were the predominant gram-negative bacteria, Bacillus cereus was the predominant gram-positive bacterium, Pichia kluyveri, Hanseniaspora uvarum, and Pichia fermentans were the predominant yeasts, and all those are pectinase-producing microorganisms. As for the contents of organic acids, oxalic was the highest, followed by acetic and lactic acids. Butyrate and propionate, which were unfavorable during the fermentation period, were barely discovered.